Nuclear Medicine Communications

Editorial

Radio-guided surgery of differentiated thyroid cancer using 131 I or 99mTc-Sestamibi Domenico Rubelloa, Massimo Salvatorib, Maria Rosa Pelizzoc, Lucia Rampina, Stefano Fantid, Michele Gregianine and Giuliano Marianif The classical therapeutic approach to patients with differentiated thyroid cancer (DTC) is based on total or near-total thyroidectomy, followed by 131I treatment and thyroid-stimulating hormone suppressive therapy. This approach allows complete cure in many patients, especially when the tumour is diagnosed at an early stage; it also allows long-term survival in patients with locoregional recurrences or distant metastases if they can be treated with 131I. In contrast, when metastatic DTC deposits lose their ability to trap 131I (non-functioning metastases), a worse prognosis is expected. Nevertheless, in patients with locoregional non-functioning recurrences, an early diagnosis and prompt surgical extirpation can lead to a favourable prognosis. In these cases, radical surgery is needed. This can be achieved with radio-guided surgery using a hand-held gamma probe and a tumour-seeking radiotracer to detect, intraoperatively, the smallest metastatic lesions. In this paper, we discuss the two principal techniques proposed in the literature for radio-guided

surgery of non-functioning DTC metastatic recurrences, the first using high doses of 131I and the second using low doses of 99mTc-Sestamibi. Nucl Med Commun 27:1–4

c 2006 Lippincott Williams & Wilkins.

Traditional treatment of differentiated thyroid cancer

of the lymph node dissection and methods of histological analysis [5]. Although, in some studies, lymph node involvement has been found to be associated with a higher risk of locoregional recurrences and tumour spread to distant sites, the impact of lymph node metastases on the survival of patients with DTC has not been elucidated completely [4,7]. In this regard, it has been reported in several series that, in spite of ‘radical’ initial treatment, 5–20% of patients develop locoregional recurrent disease and distant metastases [8].

Total thyroidectomy, followed by 131I treatment and thyroid-stimulating hormone (TSH) suppressive therapy, represents the standard of care for patients with differentiated thyroid cancer (DTC) [1]. The prognostic relevance of the extent of surgery is still controversial, although most authors currently recommend near-total or total thyroidectomy for all DTCs at any stage [2,3]. Indeed, total thyroidectomy is associated with a lower rate of local recurrences when compared with subtotal thyroidectomy or thyroid lobectomy. Moreover, the completeness of surgical excision has been proven to represent an independent prognostic variable of the overall survival and disease-free survival at multivariate statistical analysis [3,4]. In patients with DTC, first surgery is almost always followed by 131I treatment with the aim to ablate thyroid remnants and cure possible concomitant residual microscopic tumour foci. An additional advantage of this therapeutic approach is that it allows serum thyroglobulin and anti-thyroglobulin levels to be measured during follow-up [4–6]. Locoregional lymph node metastases at initial treatment are frequent, being present in approximately 50–60% of all patients with DTC. Their presence is related to the histological type of the tumour, size of the primary tumour, extension

Nuclear Medicine Communications 2006, 27:1–4 Keywords: differentiated thyroid cancer, 99m Tc-Sestamibi

131

I, radio-guided surgery,

a Nuclear Medicine Service – PET Unit, ‘S. Maria della Misericordia’ Hospital, Rovigo, bNuclear Medicine Service, Policlinico Gemelli, ‘Sacro Cuore’ Roma University, Rome, cDepartment of Surgery, Padova University, Padova, dNuclear Medicine Service – PET Unit, Policlinico S. Orsola-Malpighi, Bologna, eNuclear Medicine Service, PET Unit, ‘Umberto I’ Hospital, Mestre (VE) and fRegional Centre of Nuclear Medicine, Pisa University, Pisa, Italy.

Correspondence to Domenico Rubello MD, Nuclear Medicine Service – PET Unit, Istituto Oncologico Veneto (IOV), Viale Tre Martiri, 140, 45100 Rovigo, Italy. Tel: + + 39 (0425) 39 4427; fax: + + 39 (0425) 39 4434; e-mail: [email protected] Received 12 October 2005 Accepted 21 October 2005

Surgery is the principal treatment for locoregional DTC recurrences, and an accurate preoperative imaging workup is mandatory in order to localize precisely the site and extent of recurrence and to offer a better chance for complete surgical resection of neoplastic foci. Radiological imaging modalities, such as ultrasound, computed tomography (CT) scan and magnetic resonance imaging (MRI), are moderately sensitive (especially ultrasound), but unspecific; moreover, with these techniques, it is difficult to differentiate a postsurgical scar from viable tumour tissue [8,9]. Traditional scintigraphic imaging using single-photon emission tumour-seeking agents, such as 111Tl, 99mTc-Sestamibi (99mTc-MIBI) and 99m Tc-Tetrofosmin, has been proven to be able to overcome these problems, at least in part; however, the

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2 Nuclear Medicine Communications 2006, Vol 27 No 1

spatial resolution is no lower than 1–1.5 cm using singlephoton emission computed tomography (SPECT) [10]. More favourable results have been reported with positron emission tomography (PET) and PET/CT fusion imaging, which are characterized by a spatial resolution of 5 mm or lower. Unfortunately, some well-differentiated DTCs are characterized by a slow proliferation activity and thus may be PET negative [11]. To overcome these difficulties, several authors have proposed the use of an intraoperative gamma probe, following the administration of a tumour-seeking agent, to detect 131I-negative locoregional DTC recurrences [9,12,13]. In this paper, we discuss the principal intraoperative techniques proposed in the literature, their results, and the related advantages and limitations in patients with 131 I-negative DTC recurrent disease.

Radio-guided surgery with

131

I

Total or near-total thyroidectomy, followed by 131I ablation of thyroid remnants, is widely accepted as the most effective treatment modality in patients with DTC. After initial treatment, however, locoregional recurrences occur in 5–20% of patients and at twice the frequency of distant metastases [5]. Tumour recurrences, which are usually the consequence of incomplete initial treatment or an aggressive tumour, typically appear early during follow-up (during the first 5 years), but may occur 10–30 years later [5]. Lymph node metastases represent the most frequent type of locoregional recurrence: they can be associated with recurrent disease in the thyroid bed, soft tissue of the neck or invading the aerodigestive tract, strap muscles, recurrent laryngeal nerves and oesophagus [14]. Some patients are at higher risk of developing locoregional recurrences: young (less than 16 years of age) and elderly (more than 60 years of age) patients, patients with particular histological subtypes or with large tumours extending beyond the thyroid capsule, and patients with large, bilateral and multiple lymph node metastases at initial diagnosis [14]. Although locoregional recurrence has been reported to have controversial prognostic significance in the literature, and does not appear to be a clearly unfavourable independent prognostic factor in some studies, the tumour-specific mortality rate after lymph node recurrence is increased in most series, especially in patients over 45 years of age [14]. The therapeutic approaches to lymph node recurrences are 131I therapy, radical surgery and external radiotherapy. The efficacy of 131I therapy is essentially based on the size of the recurrence, radiotracer uptake and retention, and radiation dose delivered to recurrent disease. At the Memorial Sloan-Kettering Cancer Center in New York, Robbins and Schlumberger [15] found that a single dose of 131I can impair subsequent uptake of

131

I in approximately 65% of patients (stunning effect). Moreover, for macroscopic tumour deposits and radiation absorbed doses below 3500 cGy, cure could not be obtained [16]. The role of adjuvant external beam radiotherapy in lymph node recurrences remains controversial. It is not indicated in young patients (age < 45 years) presenting elevated 131I uptake, but some authors recommend external radiotherapy in patients over 45 years of age with extrathyroidal tumour invasion who fail to demonstrate significant 131I uptake [5]. Surgery represents the treatment of choice for I-negative locoregional recurrent disease in patients with DTC [12,13,15]. However, re-operation may be difficult, especially when the tumour foci are localized within extensive scarring related to previous surgery or in unusual sites behind vascular structures or in the mediastinum [9]. Moreover, neck dissection for locoregional recurrences should be performed in a single session, avoiding subsequent surgical procedures with a higher probability of intraoperative complications. To circumvent these difficulties, various authors have administered high 131I doses and have used an intraoperative gamma probe to detect and dissect locoregional tumour foci [9,17–19]. Because of the TSHdependent iodine uptake, withdrawal of thyroid hormone therapy is needed to achieve hypothyroidism; as an alternative, high serum TSH levels can be achieved by the administration of recombinant human TSH (rhTSH) [9,17].

131

The main inclusion criterion for radio-guided surgery with 131I in patients with DTC is the presence of persistent or recurrent radioiodine-positive locoregional disease after two or more 131I treatments [9,17]. The intraoperative protocol first proposed by Travagli et al. [9] takes place over 1 week. It begins on day 0 when the patient receives 100 mCi of 131I in a state of hypothyroidism (TSH > 30 mU/ml). 131I whole-body scan and spot neck images are obtained using a doubleheaded gamma camera equipped with a high-energy collimator on day 4. Five days after 131I administration and 1 day after scanning (day 5), a complete neck dissection of neoplastic areas is performed using an intraoperative hand-held gamma probe. After tumour extirpation, radioactivity is checked in the ‘empty’ operating basin to look for any residual radioactivity. The protocol finishes after 7 days (day 7) when a postoperative 131I whole-body scan is obtained to confirm the completeness of surgery. The radiation exposure dose to the surgeon using 131I as radiotracer for radio-guided surgery is approximately 30 mSv/h [17].

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Radio-guided surgery of thyroid cancer Rubello et al. 3

Radio-guided surgery with

99m

Tc-MIBI

131

I is the main radiotracer used in the evaluation and management of recurrent DTC. Although 131I is characterized by a very high specificity (99–100%), only a fraction of DTC recurrences are iodine avid (50–60% of papillary and 64–67% of follicular cancers) [14]. The loss of the ability to trap 131I by metastatic disease leads to limitations from both diagnostic and therapeutic viewpoints (iodine-negative or non-functioning metastases) [20]. Non-functioning metastases can be diagnosed at all ages, but their prevalence increases in the elderly, in poorly differentiated histotypes and in tumours revealing an aggressive pattern [20]. In these cases, the accurate localization of non-functioning metastases at an early stage, i.e. when they have spread to the neck and mediastinum only, is crucial because only a prompt radical re-operation can offer an increase in survival and the possibility of complete cure [13,21]. Radiological imaging modalities, such as CT scan and MRI, have shown a relatively low sensitivity in detecting locoregional metastases of DTC [20]. In contrast, high-resolution neck ultrasound and certain tumour-seeking scintigraphic methods, such as 99mTc-MIBI scan, have been proven to be highly sensitive in revealing cervical and mediastinal DTC recurrences [20–22]. On the basis of these considerations, Boz et al. [12] and Rubello et al. [13] proposed radio-guided surgery after 99mTc-MIBI injection for the intraoperative localization and resection of locoregional non-functioning DTC recurrences. This procedure was first described by Boz et al. [12], who reported a 30-year-old woman who had previously undergone two operations and 131I ablative treatment for a well-differentiated follicular thyroid carcinoma. Approximately 7 years after the initial treatment, a diagnosis of non-functioning, non-palpable neck recurrence was made on the basis of a progressive increase in serum thyroglobulin levels and a positive 99mTc-MIBI scan showing focal radiotracer uptake in the neck. In this patient, radio-guided surgery was performed 2 h after the intravenous injection of 20 mCi of 99mTc-MIBI. The tumour to background ratio in the tumour mass was high (4 : 1) and the recurrence embedded in the scar tissue was successfully removed. Complete tumour resection was confirmed by a subsequent 99mTc-MIBI scan control, and serum thyroglobulin levels decreased to undetectable values during subsequent follow-up. Rubello et al. [13] studied eight consecutive patients previously treated by total thyroidectomy and one or more 131I treatments who developed, during subsequent follow-up, locoregional non-functioning recurrences visualized at both 99mTc-MIBI scan and high-resolution neck ultrasound. At variance with Boz et al. [12], the radio-guided procedure used by Rubello et al. [13] consisted of the intravenous injection, in the operating theatre and a few minutes before the beginning of the

intervention, of a very low 99mTc-MIBI dose (1 mCi), followed by a flush of saline (30 ml) to avoid tracer stagnation. The authors chose to inject the radiotracer in the operating theatre a few minutes before the beginning of the intervention because it has been reported that DTC metastases can be characterized by a relatively rapid 99mTc-MIBI wash-out; this procedure was used to avoid possible false negative intraoperative results [20]. Moreover, the injection of the radiotracer just before the beginning of surgery allowed the authors to administer a very low 99mTc-MIBI activity, 20-fold lower than that used in the protocol of Boz et al. [12], thus minimizing the radiation exposure dose of the surgeon and operating theatre personnel. In this protocol, after radiopharmaceutical injection and under the guidance of the preoperative 99mTc-MIBI images, the patient’s neck was scanned with a hand-held 11-mm collimated gamma probe by the surgeon to individualize the cutaneous projection of the tumour focus/foci. Subsequently, during a traditional bilateral neck exploration, the surgeon used the gamma probe to detect the tumour focus (foci). Radioactivity was measured with the gamma probe on the tumour focus in vivo (T), on a background area (B) (apex of the lung contralateral to the injected arm), on the removed tumour mass ex vivo and on the tumour bed (T bed) after lesion extirpation. The last two measurements were performed to verify the completeness of surgery. The authors reported a lesion to background ratio higher than 2.0 in all cases (mean ± SD = 3.2 ± 0.8) at operation. Moreover, in seven patients, the radioactivity measured in the tumour bed after lesion extirpation fell to background activity values and, in six, stable normalization of serum thyroglobulin was achieved after reoperation, suggesting a complete cure. In the other patient, a persistent slight increase in serum thyroglobulin levels was observed after surgery despite the absence of pathological uptake in the neck and the absence of distant metastases evaluated using CT, MRI and 99mTcMIBI scans. This patient was judged to have biochemically persistent disease. In the eighth patient, the persistence of gamma probe-detected high radioactivity levels in the tumour bed after lesion removal correctly suggested disease persistence (deep tracheal infiltration), leading to the treatment of the patient with additional external radiotherapy. The authors did not find a significant correlation between tumour size and tumour 99m Tc-MIBI uptake; however, in most patients, a prompt fall in gamma probe-detected radioactivity in the neck after tumour removal greatly helped the surgeon to evaluate the completeness of extirpation [13]. Recently, the same authors have updated their experience on 38 patients using the same protocol, confirming the favourable preliminary results [21]. In this new larger study, seventy-nine metastatic lesions were removed (mean size, 18.7 ± 8.3 mm), 68 of which were 99mTcMIBI positive; the lesion to background ratio was

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4 Nuclear Medicine Communications 2006, Vol 27 No 1

2.44 ± 0.93 and the number of counts after removal was 0.9 ± 0.1. In this study, considering only the group of 25 patients with a minimum postoperative follow-up of 6 months, 18 (72%) were considered to be disease free, and seven had persistent metastatic disease [21]. The major limitation of radio-guided surgery with 99mTc-MIBI is that the metastatic deposits must be capable of trapping 99m Tc-MIBI; this condition is encountered in approximately 60–70% of cases [20]. The radiation exposure dose to the surgeon using this low-dose (1 mCi) 99mTc-MIBI approach for radio-guided surgery is 1.2 mSi/h, that is 25–30-fold less than the protocols using 131I [13,21].

Conclusion Radio-guided surgery for locoregional non-functioning DTC recurrences can be successfully performed using either a high-dose 131I protocol or a low-dose 99mTc-MIBI protocol. The 99mTc-MIBI protocol has the advantage of delivering a lower radiation dose to the surgeon and operating theatre personnel; however, metastatic lesions need to be capable of trapping 99mTc-MIBI. Conversely, the protocol based on high-dose 131I is potentially effective in all patients with DTC, but a higher radiation exposure dose is delivered to the surgeon and operating theatre personnel.

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3

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Shaha AP. Management of the neck in thyroid cancer. Otorhinolaryngol Clin North Am 1998; 31:823–883. Pelizzo MR, Boschin IM, Toniato A, Pagetta C, Piotto A, Rubello D. Natural history, diagnosis, treatment and outcome of papillary thyroid microcarcinoma (PTMC): a mono-institutional 12-year experience. Nucl Med Commun 2004; 25:547–552. Pelizzo MR, Toniato A, Boschin IM, Piotto A, Nibale O, Rubello D. Locally advanced differentiated thyroid carcinoma: a 35-year mono-institutional experience in 280 patients. Nucl Med Commun 2005; 26:965–968. Hay ID, Bergstrahl EJ, Goellner JR, Ebersold JR, Grant CS. Predicting outcome in papillary thyroid carcinoma: development of a reliable prognostic scoring system in a cohort of 1779 patients treated at one institution during 1940 through 1989. Surgery 1993; 114:1050–1058. Mazzaferri EL, Kloos RT. Current approaches to primary therapy for papillary and follicular thyroid cancer. J Clin Endocrinol Metab 2001; 86:1447–1463.

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Rubello D, Caasara D, Girelli ME, Piccolo M, Busnardo B. Clinical meaning of circulating antithyroglobulin antibodies in differentiated thyroid cancer: a prospective study. J Nucl Med 1992; 33:1478–1480. American Joint Committee on Cancer. Thyroid. In: American Joint Committee on Cancer. AJCC cancer staging handbook. 6th ed. New York: Springer; 2002. pp. 89–98. Tisell LE, Hansson G, Jansson S, Salander H. Reoperation in the treatment of asymptomatic metastasizing medullary thyroid carcinoma. Surgery 1986; 99:60–66. Travagli JP, Cailleux AF, Ricard M, Baudin E, Caillou B, Parmentier C, Schlumberger M. Combination of radioiodine (131I) and probe-guided surgery for persistent or recurrent thyroid carcinoma. J Clin Endocrinol Metab 1998; 83:2675–2860. Cherry SR, Sorenson JA, Phelps ME. The gamma camera: performance characteristics. In: Cherry SR, Sorenson JA, Phelps ME, editors. Physics in nuclear medicine. 3rd ed. Philadelphia, PA: Saunders; 2003. pp. 456–467. Schluter B, Bohuslavizki KH, Beyer W, Plotkin M, Buchert R, Clausen M. Impact of FDG PET on patients with differentiated thyroid cancer who present with elevated thyroglobulin and negative 131I scan. J Nucl Med 2001; 42:71–76. Boz A, Arici C, Gu¨ngo¨r F. Gamma probe-guided resection and scanning with Tc-99m MIBI of a local recurrence of follicular thyroid carcinoma. Clin Nucl Med 2001; 26:820–822. Rubello D, Piotto A, Pagetta C, Pelizzo MR, Casara D. 99mTc-MIBI radioguided surgery for recurrent thyroid carcinoma: technical feasibility and procedure, and preliminary clinical results. Eur J Nucl Med 2002; 29: 1201–1205. Voutilainen PE, Multanen MM, Leppaniemi AK. Prognosis after lymph node recurrence in papillary thyroid carcinoma depends on age. Thyroid 2001; 11:953–957. Robbins RJ, Schlumberger MJ. The evolving role of 131I for the treatment of differentiated thyroid carcinoma. J Nucl Med 2005; 46:28S–37S. Maxon HR, Englaro EE, Thomas SR. Radioiodine-131 therapy for well differentiated thyroid cancer. A quantitative radiation dosimetric approach: outcome and validation in 85 patients. J Nucl Med 1992; 33: 1132–1136. Salvatori M, Rufini V, Reale F, Samanes Gasate AM, Maussier ML, Revelli L, et al. Radio-guided surgery for lymph node recurrences of differentiated thyroid cancer. World J Surg 2003; 27:770–775. Gulec SA, Eckert M, Woltering EA. Gamma probe-guided node dissection (‘gamma picking’) in differentiated thyroid carcinoma. Clin Nucl Med 2002; 12:859–861. Gallowitsh HJ, Fellinger J, Mikosch P. Gamma probe-guided resection of a lymph node metastasis with I-123 in papillary thyroid carcinoma. Clin Nucl Med 1997; 22:591–592. Rubello D, Mazzarotto R, Casara D. The role of technetium-99m methoxyisobutylisonitrile scintigraphy in the planning of therapy and followup of patients with differentiated thyroid carcinoma after surgery. Eur J Nucl Med 1999; 27:431–440. Pelizzo MR, Pavan N, Piotto A, Rampin L, Nibale O, Rubello D. Sestamibi radioguided surgery in iodine-131 negative metastatic thyroid cancer. Eur J Nucl Med Mol Imaging 2005; 46:P607. Franceschi M, Kusic Z, Franceschi D, Luiniac L, Roncevic S. Thyroglobulin determination, neck ultrasonography and iodine-131 whole body scintigraphy in differentiated thyroid carcinoma. J Nucl Med 1996; 37: 446–451.

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Original article

Intra-individual comparison of sentinel lymph node scintigraphy on the day of injection and on the following day in breast cancer Fabrice Gutmana,b, Agathe Sansona,b, Jean-Michel Piquenotc, Anne Hitzela,b, Jean-Marie Ladonned, Phillipe Dessogned and Pierre Ve´raa,b Objectives To compare, intra-individually, the detection rates of sentinel node on lymphoscintigraphy performed on the day of injection (D0) and on the following day (D1) in breast carcinoma. We also compared 2-day and 1-day protocols in the two groups of patients. Methods The 2-day and 1-day protocols included 76 patients in group 1 and 23 patients in group 2. Patients from group 1 underwent lymphoscintigraphy twice – at 2 h (lymphoscintigraphy 1) and 18 h (lymphoscintigraphy 2) post-injection at four sites periareolar using 99mTc sulfur colloid. Patients from group 2 underwent lymphoscintigraphy only at 2 h post-injection. The detection rates and the number of sentinel nodes were compared in the two lymphoscintigraphy examinations for group 2. Results The detection rate on lymphoscintigraphy in group 1 was 92% at D0 and 96% at D1. The overall agreement between lymphoscintigraphy 1 and lymphoscintigraphy 2 was 69/76 (91%). In 2/76 women, the sentinel node disappeared at D1 on lymphoscintigraphy, but remained detectable during surgery, and in 5/76 women, the sentinel node appeared at D1 on lymphoscintigraphy. The mean number of sentinel nodes detected on lymphoscintigraphy was 2.05 ± 0.14 at D0 and 1.76 ± 0.11 at D1 (P = 0.004) in

Introduction Breast carcinoma is the most frequent cancer in women. The main prognostic factor is axillary node involvement for predicting its metastatic potential. Axillary clearance is an efficient procedure for both staging and preventing the local axillary relapse. However, more than 10% of women who undergo axillary dissection have post-operative complications in the arm (sensitivity dysfunction, physical limitations, lymphoedema), and psychological distress [1,2]. Small carcinomas of the breast (TNM stage T1 r 2 cm) have a low risk of lymph node involvement (less than 20%) and thus 80% of nodenegative patients with small breast carcinoma could avoid total lymph node dissection. This approach is gaining interest because of the increasing number of small breast carcinomas detected by screening programmes. Sentinel lymph node biopsy is a minimally invasive method for

group 2, the detection rate of the sentinel node was 20/23 (87%). Conclusion Our study demonstrated that for patients undergoing the 2-day protocol for sentinel node procedure in early stage breast cancer, the optimal imaging time would be to perform lymphoscintigraphy 1 h after injection, with the possibility of imaging patients the following day in cases where lymphoscintigraphy was negative. c 2006 Lippincott Williams & Nucl Med Commun 27:5–9 Wilkins. Nuclear Medicine Communications 2006, 27:5–9 Keywords: sentinel node imaging, breast cancer

a Department of Nuclear Medicine, Rouen University Hospital Charles-Nicolle and Henri Becquerel Center, Rouen, bLaboratoire universitaire QUANT.I.F., cService of Histopathology and dService of Surgery, Henri Becquerel Center, Rouen, France.

Correspondence to Dr Fabrice Gutman, De´partement de Me´decine Nucle´aire, Hoˆpital Tenon, 4 rue de la Chine, 75020 Paris, France. Tel: + 33 1 5601 6556; fax: 33 1 5601 6171; e-mail: [email protected]

Received 25 January 2005 Revised 27 July 2005 Accepted 17 August 2005

staging patients with early breast cancer. If the sentinel node, defined as the first regional node receiving lymphatic drainage from a cancer, is benign the patient may eventually be spared the morbidity of lymph node dissection. Numerous cases of sentinel node procedures for breast carcinomas have been published in the past few years, with a great heterogeneity of isotopic or blue dye techniques. Surprisingly, results using various techniques all seem to reliably identify the ‘true’ sentinel lymph node(s) in the axilla, providing very homogeneous results in terms of concordance rates with complete axilla dissection to predict the lymph node status (80–100%). The various sentinel node procedures differ in the radiopharmaceutical used [3], the activity administered [4], and the site of injection, either peritumoural,

c 2006 Lippincott Williams & Wilkins 0143-3636

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6 Nuclear Medicine Communications 2006, Vol 27 No 1

intra-tumoural or periareolar [5,6]. Radioisotope mapping and surgical excision usually involve one visit to the hospital but the operating room programme is often wasted in waiting for patients to arrive from lymphoscintigraphy. Thus, instead of performing a 1-day protocol, it would be better to perform the lymphoscintigraphy the day before surgery, using a 2-day protocol. To our knowledge, no previously published study has undertaken an intra-individual comparison of lymphoscintigraphy performed the day before surgery and the day of surgery. The aim of the present study was to compare, intraindividually, the images obtained on the day before surgery and, then, just before surgery.

Table 1

Characteristics of the two patient populations in this study

Characteristic Number of patients Mean age (years) Size of primary (mm) Histology Ductal, in situ Lobular, in situ Ductal Lobular Other malignant Benign

Group 1 (2-day protocol) Group 2 (1-day protocol) 76 57 ± 11 14 ± 9

23 61 ± 12 15 ± 11

4 2 43 13 10 6

2 1 12 5 3 0

Group 1 consisted of 76 patients who underwent the sentinel node procedure on two consecutive days, with lymphoscintigraphy on the day of injection and the day following injection. Group 2 consisted of 23 patients who underwent the sentinel node procedure within a single day.

Material and methods Patients

We performed a prospective study on 99 women who underwent the sentinel node procedure at the Henri Becquerel Center from January 2000 to January 2003. Patients were referred for a 99mTc labelled albumin nanocolloid study for lymphatic mapping after identification of breast tumours less than 15 mm and no evidence of palpable axillary lymph nodes. Patients with multifocal breast carcinomas and prior breast surgery were excluded from this study, which reflects our experience in the technique of locating sentinel nodes and is free of learning curve artifacts.

Group 1 included 76 patients who underwent the sentinel node procedure within two consecutive days. In this group, colloids were injected into the periareolar region the day before surgery, and lymphatic mapping was performed the day prior to surgery and the morning of surgery (1 h and 18 h post-injection). Group 2 included 23 patients who underwent the sentinel node procedure within a single day. In this group, colloids were injected into the periareolar region on the morning of surgery, and lymphatic mapping was performed 1 h after injection. Sentinel lymph node procedure

Study design

The 99 women were assigned to two groups (Fig. 1). Table 1 shows the characteristics of the two patient populations included in the study. Patients were not randomly assigned to either protocol but were allocated on the basis of the feasibility of scheduling the procedure.

Fig. 1

Group 1 (n = 76)

18 h

Sg

I

{

2 h LSG

LSG

D0

D1 Group 2 (n =23) Sg

{

I

2h

All patients received a four-site periareolar intra-dermal injection of 99mTc sulfur colloid (Nanocoll; Nycomed Amersham Sorin, Saluggia, Italy). Both groups received 74 MBq of the radiopharmaceutical. A brief massage was performed after injection. All patients underwent lymphoscintigraphy. Anterior and lateral projections were performed using a dual head camera with gantries at 901 (DST-XLi; General Electric Medical Systems) equipped with low energy, high resolution collimators. Static images were obtained with an acquisition time of 180 s in a 128  128 matrix. A cobalt flood source was used to silhouette the patient during image acquisition. The patient was positioned supine with the arm ipsilateral to the lesion in maximum abduction. A cutaneous projection of the sentinel node was marked from the anterior and lateral views to ease the per-operative detection. A gamma ray detection probe (Europrobe; Eurorad, France) was used to locate the lesion and to guide its surgical removal.

LSG

Evaluation D1 Design of the study: sentinel node procedure in the two groups. I: injection; D0: day before surgery; D1: day of surgery; Sg: surgery.

Two experienced observers together determined the number of visible sentinel nodes and the location of the first order sentinel node on all the images. The observers did not review images independently and a consensus was reached in all cases. In group 1, the success rate of lymphoscintigraphy and the number of visible sentinel

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Comparison of sentinel node scintigraphy on D0 and D1 Gutman et al. 7

nodes at D0 and D1 were assessed. Groups 1 and 2 were compared with regard to the detection rates of sentinel nodes on lymphoscintigraphy.

Fig. 2

D0

Anterior view

D1

Anterior view

Results Comparison of D0 and D1 lymphoscintigraphy in group 1 Visualization

Table 2 shows the results of D0 and D1 lymphoscintigraphy for the 76 patients. The success rate of lymphoscintigraphy in imaging the sentinel node was 70/76 (92%) on initial images (D0), and 73/76 (96%) on the delayed images (D1). In only one of the 76 cases, no sentinel node was visualized at either D0 or D1. The results of both lymphoscintigraphy examinations were in agreement in 69/76 cases with 68 positive detections and one non-visualization case. In seven cases, there was a disagreement between D0 and D1 lymphoscintigraphy with two cases where the sentinel node disappeared at D1. However, each time a sentinel node was visualized at D0 or D1 on lymphoscintigraphy, it was detected during the surgical procedure, including the two cases of disappearance of the sentinel node on the D1 images. Figure 2 shows the visualization of an axillary sentinel lymph node on delayed images.

Intramammary lymph pathway

Lateral view

Axillary SLN

Lateral view

Visualization of an axillary sentinel lymph node on delayed images. D0: day before surgery; D1: day of surgery.

and 22/23 in group 2 (96%) (surgery performed 2 h postinjection) (P = NS).

Discussion Number and location of sentinel nodes

The overall number of hot spots visualized on lymphoscintigraphy was 2.05 ± 0.14 at D0 and 1.76 ± 0.11 at D1 (P = 0.004). Hot spots represent both the ‘true’ sentinel node and higher-echelon nodes, which can perturb the surgical detection of the first order node. The same number of hot spots was detected on lymphoscintigraphy at D0 and D1 in 41/76 patients (54%). In 25/76 (33%), the number of hot spots visualized on lymphoscintigraphy was greater at D0, while in 10/76 (13%), the number of visualized hot spots was greater at D1. The breast lymphatic pathway was only in the axilla on lymphoscintigraphy in 74/76 patients at D0 and in 75/76 at D1, or both axilla and parasternal area in 2/76 at D0 and 1/76 at D1. When the sentinel node was visualized on lymphoscintigraphy at D0 and at D1 (in 68/76 patients), it was located in the same axillary area in 67/68 (99%). Comparison of groups 1 and 2

The surgical detection rate of sentinel node was 75/76 (99%) in group 1 (surgery performed 18 h post-injection) Results of lymphoscintigraphy on the day before surgery and on the day of surgery in group 1

Table 2

Day before surgery

Day of surgery

Total

Negative

Positive

Negative Positive

1 (1.3%) 2 (2.6%)

5 (6.6%) 68 (89.5%)

6 (7.9%) 70 (92.1%)

Total

3 (3.9%)

73 (96.1%)

76

We performed a prospective intra-individual study concerning the lymphoscintigraphic images performed on the day before and the day of surgery for sentinel node in early breast cancer. The aim of the study was to compare lymphoscintigraphic images performed on the day of injection and on the following day in order to decrease the waiting time for the surgical team. The rationale for periareolar injections is based on observations that suggest the lymphatic route to the axillary regional nodes may be the same for all breast sites. Even if this question remains debatable, several studies [7–10] showed similar results between sub-areoral or periareoral and peritumoural injections. However, one drawback of this technique could be the underestimation of an extraaxillay pathway, where therapeutic consequences are not evidence based [11]. Comparison of groups 1 and 2

The visualization rate of sentinel node on lymphoscintigraphy was higher in group 1 vs. group 2 (96% on late images vs. 87%), due to the possibility of delayed images in patients undergoing the 2-day protocol. Our study is in agreement with that of Veronesi et al. and Pijpers et al. [12,13] who showed the feasibility of injecting a radiopharmaceutical on the day before a successful surgical procedure in a large series of patients. Both sentinel node procedures in nuclear medicine and surgical departments can be performed within a single day. Although sentinel node localization based on this protocol is effective, operating room time is often wasted

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8 Nuclear Medicine Communications 2006, Vol 27 No 1

in waiting for patients to arrive from lymphoscintigraphy. The subsequent frustration of the surgeon and time pressure on the nuclear medicine physician may result in the omission of additional images that might reveal unexpected pathways of lymphatic drainage. These constraints have led surgeons to question the usefulness of lymphoscintigraphy. Moreover, Upponi et al. [14] showed that the surgical sentinel lymph node procedure in 62 patients, performed blindly to the images, found an axillary sentinel node each time the lymphoscintigraphy was positive. Also, McMasters et al. [15] showed that when the lymphoscintigraphy was negative, the sentinel node could be per-operatively detected in 78% in 126 patients. In contrast, lymphoscintigraphy with cutaneous marking guides the surgeon to the excision site and could also show an extra-axillary pathway. For these reasons, a 2day protocol seems to be more suitable because there is the possibility of performing lymphoscintigraphy with delayed images, without the constraints of a 1-day protocol.

(Fig. 3). The main drawback of this higher rate of ‘secondary lymph nodes’ is the risk of incorrect skin marking. Our study did not confirm the hypothesis of Morton et al. [17] and Tafra et al. [18] that the number of sentinel nodes increases with the time between injection and imaging. Yeung et al. [16] demonstrated that the same number of sentinel nodes was obtained at D0 and D1 in 666 patients (2.5 and 2.8, respectively). These contradictory results may probably be explained by the variations in the behaviour of radiopharmaceuticals with different particle sizes.

Conclusion Our study demonstrated that, in the 2-day protocol for the sentinel lymph node procedure in early stage breast cancer, the optimum time to perform lymphoscintigraphy would be 1 h after injection on the day before surgery. It is then possible to image patients on the following day if the lymphoscintigraphy is negative.

Acknowledgement Comparison of lymphoscintigraphy performed at D0 and D1 in group 1: the optimal time for imaging

The optimal time for imaging in the 2-day protocol remains controversial. So far, only inter-individual studies have been performed to evaluate the optimal time for imaging in the 2-day protocol. Yeung et al. [16] showed similar results for detection of sentinel nodes in 1-day and 2-day protocols in two groups of patients (the ratio of the success rates in the 2-day and 1-day protocols was 1.02). In our intra-individual study in 76 patients, the sentinel node visualization rate on lymphoscintigraphy at D0 and D1 was 92% and 96%, respectively. The sentinel node disappeared between D0 and D1 on lymphoscintigraphy only in two cases, but remained detectable during surgery. There was an increase in the ‘secondary’ sentinel nodes visualized at D0 compared to D1 on lymphoscintigraphy (2.05 ± 0.14 and 1.76 ± 0.11, respectively)

The authors thank R. Medeiros for his valuable advice in editing the manuscript.

References 1 2

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Fig. 3

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'Secondary' LN 9

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Injection site 11

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Visualization of ‘secondary’ lymph nodes on lymphoscintigraphy at D0, the day before surgery. D1: day of surgery.

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Ivens D, Hoe AL, Podd TJ, Hamilton CR, Taylor I, Royle GT. Assessment of morbidity from complete axillary dissection. Br J Cancer 1992; 66:136–138. Liljegren G, Holmberg L. Arm morbidity after sector resection and axillary dissection with or without postoperative radiotherapy in breast cancer stage I. Results from a randomised trial. Uppsala-Orebro Breast Cancer Study Group. Eur J Cancer 1997; 33:193–199. Goldfarb LR, Alazraki NP, Eshima D, Eshima LA, Herda SC, Halkar RK. Lymphoscintigraphic identification of sentinel lymph nodes: clinical evaluation of 0.22-micron filtration of Tc-99m sulfur colloid. Radiology 1998; 208:505–509. Valdes Olmos RA, Tanis PJ, Hoefnagel CA, Nieweg OE, Muller SH, Rutgers EJ, et al. Improved sentinel node visualization in breast cancer by optimizing the colloid particle concentration and tracer dosage. Nucl Med Commun 2001; 22:579–586. Krag D, Weaver D, Ashikaga T, Moffat F, Klimberg VS, Shriver C, et al. The sentinel node in breast cancer-a multicenter validation study. N Engl J Med 1998; 339:941–946. Smith LF, Cross MJ, Klimberg VS. Subareolar injection is a better technique for sentinel lymph node biopsy. Am J Surg 2000; 180:434–437; discussion 437–438. Zavagno G, Meggiolaro F, Rossi CR, Casara D, Pescarini L, Marchet A, et al. Subareolar injection for sentinel lymph node location in breast cancer. Eur J Surg Oncol 2002; 28:701–704. Shen P, Glass EC, DiFronzo LA, Giuliano AE. Dermal versus intraparenchymal lymphoscintigraphy of the breast. Ann Surg Oncol 2001; 8:241–248. Yoshida K, Yamamoto N, Imanaka N, Togawa T, Miyauchi M, Miyazaki M. Will subareolar injection be a standard technique for sentinel lymph node biopsy? Breast Cancer 2002; 9:319–322. Maza S, Valencia R, Geworski L, Zander A, Guski H, Winzer KJ, Munz OL. Peritumoural versus subareolar administration of technetium-99m nanocolloid for sentinel lymph node detection in breast cancer: preliminary results of a prospective intra-individual comparative study. Eur J Nucl Med Mol Imaging 2003; 30:651–656. Jansen L, Doting MH, Rutgers EJ, de Vries J, Olmos RA, Nieweg OE. Clinical relevance of sentinel lymph nodes outside the axilla in patients with breast cancer. Br J Surg 2000; 87:920–925. Veronesi U, Paganelli G, Galimberti V, Viale G, Zurrida S, Bedoni M, et al. Sentinel-node biopsy to avoid axillary dissection in breast cancer with clinically negative lymph-nodes. Lancet 1997; 349:1864–1867. Pijpers R, Meijer S, Hoekstra OS, Collet GJ, Comans EF, Boom RP, et al. Impact of lymphoscintigraphy on sentinel node identification with

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Comparison of sentinel node scintigraphy on D0 and D1 Gutman et al. 9

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technetium-99m-colloidal albumin in breast cancer. J Nucl Med 1997; 38:366–368. Upponi SS, McIntosh SA, Wishart GC, Balan KK, Purushotham AD. Sentinel lymph node biopsy in breast cancer–is lymphoscintigraphy really necessary? Eur J Surg Oncol 2002; 28:479–480. McMasters KM, Wong SL, Martin RC 2nd, Chao C, Tuttle TM, Noyes RD, et al. Dermal injection of radioactive colloid is superior to peritumoral injection for breast cancer sentinel lymph node biopsy: results of a multiinstitutional study. Ann Surg 2001; 233: 676–687.

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Yeung HW, Cody IH, Turlakow A, Riedel ER, Fey J, Gonen M, et al. Lymphoscintigraphy and sentinel node localization in breast cancer patients: a comparison between 1-day and 2-day protocols. J Nucl Med 2001; 42:420–423. 17 Morton DL, Bostick PJ. Will the true sentinel node please stand? Ann Surg Oncol 1999; 6:12–14. 18 Tafra L, Lannin DR, Swanson MS, Van Eyk JJ, Verbanac KM, Chua AN, et al. Multicenter trial of sentinel node biopsy for breast cancer using both technetium sulfur colloid and isosulfan blue dye. Ann Surg 2001; 233:51–59.

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Original article

Can [18F]fluorodeoxyglucose positron emission tomography imaging complement biopsy results from the iliac crest for the detection of bone marrow involvement in patients with malignant lymphoma? David Fustera, Stephen Chianga, Charalambos Andreadisb, Liang Guana, Hongming Zhuanga, Stephen Schusterb and Abass Alavia Objectives To assess the usefulness of [18F]fluorodeoxyglucose positron emission tomography in the detection of bone marrow involvement in malignant lymphoma, and its impact in clinical management. Methods One hundred and six consecutive patients with a confirmed diagnosis of lymphoma, referred for staging or restaging of Hodgkin’s lymphoma (n = 18) or non-Hodgkin’s lymphoma (n = 88), were reviewed retrospectively. A positron emission tomography scan and bone marrow biopsy of the iliac crest were performed in all patients. The assessment of bone marrow involvement by lymphoma was confirmed by histology and/or progression of bone marrow lesions in clinical follow-up. Results In 28 of 106 patients, bone marrow involvement was found. Positron emission tomography was more sensitive (86%) than bone marrow biopsy (57%). Positron emission tomography and bone marrow biopsy were concordant by positive correlation in 12 of 28 cases (43%) and by negative correlation in 77 of 78 cases (99%). Ten cases of non-Hodgkin’s lymphoma and two cases of Hodgkin’s lymphoma with positive positron emission tomography results and an initial negative bone marrow biopsy showed clinical progression of the bone marrow lesions and/or subsequent positive histology. These were considered as false-negative results for bone marrow biopsy. In seven of the 12 positive cases with negative

Introduction Bone marrow biopsy (BMb) is the gold standard method for determining the presence of bone marrow involvement in the initial staging of Hodgkin’s and nonHodgkin’s lymphoma (NHL). It is a painful and invasive procedure with a low yield in the detection of metastatic bone marrow disease [1–3]. [18F]Fluorodeoxyglucose positron emission tomography ([18F]FDG PET) is a non-invasive imaging technique that has been proven to be useful in the diagnosis and staging of several types of malignancy [4–8]. [18F]FDG PET imaging has also been used successfully in the detection of both primary and metastatic bone and bone marrow malignancies [5]. Several studies have indicated that PET can be very

bone marrow biopsy, positron emission tomography uptake distant from the site of the biopsy was seen. In four cases of follicular lymphoma, the bone marrow biopsy was positive and the positron emission tomography scan was normal. Conclusions Positron emission tomography and bone marrow biopsy are complementary in assessing the presence of bone marrow involvement in patients with malignant lymphoma. In our series, positron emission tomography was more sensitive than bone marrow biopsy in Hodgkin’s and non-Hodgkin’s lymphoma, except in follicular lymphoma. Nucl Med Commun 27:11–15

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:11–15 Keywords: bone marrow biopsy, diagnosis, [18F]fluorodeoxyglucose, lymphoma, positron emission tomography a Nuclear Medicine Division/Radiology Department and bHematology/Oncology Division/Department of Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, USA.

Correspondence to Dr David Fuster, Department of Nuclear Medicine, Hospital Clı´nic de Barcelona, Villarroel, 170, 08036 Barcelona, Spain. Tel: + 34-93-227-5516; fax: + 34-93-451-8137; e-mail: [email protected] Received 23 March 2005 Accepted 30 August 2005

useful in the staging and evaluation of recurrent disease and can change management in a substantial number of patients with lymphoma [9]. Studies comparing PETwith computed tomography (CT) for the staging of lymphoma have consistently shown that PET is highly successful in identifying nodal disease, and its greater accuracy aids the detection of small or borderline nodes and soft tissue involvement [9–13]. Magnetic resonance imaging (MRI) is a valuable tool and is probably the most sensitive procedure for the detection of local bone marrow involvement when a specific site is suspected of being affected by the disease. However, MRI lacks specificity and therefore cannot be relied upon for the accurate assessment of disease activity. Moreover, PET has the

c 2006 Lippincott Williams & Wilkins 0143-3636

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12 Nuclear Medicine Communications 2006, Vol 27 No 1

ability to screen the entire bone marrow, which is not practical using either MRI or CT. PET’s spatial and contrast resolution is superior to that of current existing nuclear medicine modalities for the assessment of bone marrow abnormalities, including conventional bone scans [14,15] and 67Ga imaging [16], which have been utilized to detect lymphomatous involvement of the marrow. It is imperative to detect and accurately determine the extent of bone marrow involvement in patients with lymphoma, especially in high-grade lymphomas where optimal staging is crucial for treatment planning [1–3]. The aims of this study were to assess the usefulness of PET in the detection of bone marrow involvement and its impact in the clinical management of lymphomas.

2.5D RAMLA iterative algorithm on the C-PET scanner and 3D RAMLA iterative algorithm on the Allegro scanner. All studies were interpreted to determine the presence of abnormal [18F]FDG uptake in the marrow of the bony structures included. Focal areas of increased uptake anywhere in the bone marrow were considered pathological and were recorded for comparison with BMb and patient follow-up results, as mentioned in the previous section. The results of the BMb were not known at the time of PET scan interpretation. Comparison of the PET results and unilateral iliac crest BMb results was performed in all patients. Biopsy samples were analysed following standard procedures.

Results Methods Patients

This study included 106 consecutive patients (54 women, 52 men) with a mean age of 53 ± 15 years and a confirmed diagnosis of lymphoid neoplasm by the World Health Organization criteria, who were referred for staging or restaging of known Hodgkin’s lymphoma (n = 18) or NHL (n = 88) by means of [18F]FDG PET imaging. The categories of NHL found in this series were diffuse large B-cell (n = 37), follicular (n = 21), peripheral T-cell (n = 7), mantle cell (n = 7), marginal zone (n = 5) and other cell types (n = 11). The time interval between BMb and [18F]FDG PET was 13 ± 12 days. A whole-body PET scan and unilateral BMb of the iliac crest were performed in all patients. Patients who received treatment between the two procedures were excluded from this study. Histology and/or clinical followup for at least 12 months following the PET scan was used for the final diagnosis of disease recurrence. The progression of bone marrow PET lesions during followup by PET and CT imaging and/or subsequent positive histology was considered as a true-positive finding. This retrospective review was approved by the Institutional Review Board of our institution. Method

The [18F]FDG PET protocol required the patients to fast for at least 4 h before the intravenous administration of 2.516 MBq
kg – 1 of [18F]FDG for C-PET studies and 5.18 MBq
kg – 1 for Allegro PET studies (dedicated PET, C-PET or Allegro; Philips-ADAC, Milpitas, California, USA). Sixty minutes later, whole-body PET images were acquired in three-dimensional mode on either the C-PET or Allegro scanner; the neck, chest, abdomen and pelvis were included. Attenuation correction was performed using a 137Cs transmission source employing a nonuniform background subtraction method with scatter correction. The images were reconstructed using the

Eighty-one patients were known to have active lymphoma, confirmed by histological findings, and 25 were considered in remission at the time of the study. Of the 81 patients with proven active disease, 28 were found to have marrow involvement by histological examination (16) or by progression of bone marrow lesions detected by [18F]FDG PET (12) (Table 1). PET showed pathological focal marrow uptake of [18F]FDG in 24 of the 28 cases (86%) with proven bone marrow disease (Fig. 1). BMb was less sensitive than [18F]FDG PET, showing positive results in only 16 of these 28 cases (57%). The sensitivity and specificity of PET for the detection of bone marrow involvement were 86% and 99%, respectively. PET and BMb were positively concordant in 12 of 28 (43%) of the positive cases and negatively concordant in 77 of 78 (99%) of the negative cases. Ten cases of NHL and two cases of Hodgkin’s lymphoma with positive PET uptake and an initially negative BMb showed progression of the disease in the marrow, demonstrated by subsequent positive histological findings when possible (five cases) and/or by PET and CT imaging follow-up (seven cases). These were considered as false-negative results for BMb. In seven of the 12 positive cases with negative BMb, PET uptake distant from the biopsied site was seen (Fig. 2). All the above patients presented with active lymphoma, but unsuspected bone marrow involvement, and PET findHistological type, disease activity and state of bone marrow involvement in all patients

Table 1

Histology Hodgkin’s lymphoma (18) Large B-cell lymphoma (37) Follicular lymphoma (21) Peripheral T-cell lymphoma (7) Mantle cell lymphoma (7) Marginal zone lymphoma (5) Other non-Hodgkin’s lymphoma (11) Total (106)

Presence of active disease

Bone marrow involvement

15/18 29/37 14/21 6/7 3/7 5/7 9/11 81/106

4/15 9/29 9/14 2/6 0/3 1/5 3/9 28/81

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PET and biopsy to detect lymphoma in bone marrow Fuster et al. 13

Fig. 1

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ANT LEFT

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Whole-body positron emission tomography scan in a patient with Hodgkin’s lymphoma and a negative initial bone marrow biopsy of the right iliac crest demonstrates pathological focal marrow uptake in the mid- and lower lumbar spine, a cluster of increased uptake in the sacrum and several foci of uptake in the posterior aspect of the left ischium (arrows).

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A whole-body positron emission tomography (PET) scan was performed in a patient with a grade 3 follicular lymphoma and a negative initial bone marrow biopsy of the iliac crest. Transaxial (a), sagittal (b) and coronal (c) PET slices show pathological marrow uptake in the sternum (small arrows). The image was highly suggestive of bone marrow disease, which was confirmed histologically. There is also lymph node involvement of the right hilum (large arrow).

ings led to an upgrade of the tumour stage. However, in four cases of follicular lymphoma (grade 1–2), the BMb was positive but PET images of the bone marrow were

normal; these were considered as false-negative results for PET (Table 2). There was only one false-positive result: PET imaging showed a mild, extensive uptake in

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14 Nuclear Medicine Communications 2006, Vol 27 No 1

Table 2 Comparison of the results obtained by positron emission tomography (PET) with the presence or absence of bone marrow involvement confirmed by histology and/or clinical follow-up Bone marrow involvement ( + )

Bone marrow involvement ( – )

24 4 28

1 77 78

PET ( + ) PET ( – )

25 81 106

the spine, but the lesion showed stability in the follow-up PET scan without further treatment. MRI of the spine demonstrated diffuse degenerative joint disease in the spine coinciding with PET findings, but no images suggesting malignant marrow disease were found.

Discussion BMb of the iliac crest is considered as the gold standard for the staging and restaging of patients with lymphoma. Unfortunately, this test is insensitive when the spread of cancer is focal and occurs at sites distant from the area in which the biopsy is taken. Therapy for lymphoma patients is related to staging and, as bone marrow involvement requires more aggressive treatment, a more accurate diagnosis of marrow disease is needed. The ability of PET imaging to screen the entire patient permits the detection of bone marrow involvement with a high degree of sensitivity anywhere in the body, as pointed out in the literature [17,18]. In our series, PET imaging showed bone marrow disease in 12 of 28 cases (43%) in which the initial BMb of the iliac crest was negative and subsequent histological examination or progression of the lesions detected by [18F]FDG PET demonstrated bone marrow involvement. In 25% of the cases with bone marrow involvement and negative BMb, PET showed focal disease distant from the iliac crest, which is very difficult or impossible to detect by BMb of the iliac crest. In these patients, bone marrow involvement was suspected only by the PET findings and led to a significant change in the initially planned chemotherapeutic strategy. PET was unable to detect marrow disease in four cases of follicular lymphoma (grades 1 and 2). It may not always be appropriate for low-grade NHL, and further studies are needed to evaluate the role of PET in these types of lymphoma [19]. The rate of false-positive findings with PET, taking into account all clinical variables, is quite low. In our series, there was only one false-positive case in a patient with Hodgkin’s lymphoma, in which PET showed a mild and extensive marrow uptake in the spine, misinterpreted as consistent with bone marrow involvement, which was in fact related to severe degenerative joint disease changes. A clinical situation which may stimulate bone marrow uptake is increased marrow

glucose metabolism as a result of previous systemic chemotherapy [20,21]. In the irradiated bone marrow, a transient increase in uptake may also be seen, which is followed by a rapid decline to baseline in a few days. These findings are easily classified because of the low intensity and diffuse pattern. In this setting, a significant increase in [18F]FDG uptake in the non-irradiated bone marrow in the rest of the body has been attributed to the release of cytokines during radiation therapy [22]. Our findings agree with those of other investigators in terms of the high overall accuracy (particularly the high sensitivity) of PET compared with BMb [17,18]. Moreover, there is also a consensus that false-negatives with PET tend to occur in cases of slow-growing lymphomas. Carr et al. [17] showed that, in two of three patients with histologically demonstrated marrow disease and no [18F]FDG uptake, there was no [18F]FDG uptake at the site of the primary lymph node disease. It was therefore unlikely that PET would be able to detect similar cells in the bone marrow. These results differ from ours in that we observed active nodal disease in three of the four false-negative cases. One possible explanation may be that there are factors, other than the degree of aggressiveness of the tumour, which may affect the sensitivity of PET in detecting marrow disease. In a study of 78 patients, Moog et al. [18] showed low sensitivity for [18F]FDG PET in detecting marrow involvement in lowgrade lymphomas. However, their series focused exclusively on the bone marrow and did not analyse other possible disease sites. Bone scanning has traditionally been used in the evaluation of marrow involvement by lymphoma, showing a high sensitivity (compared with conventional radiography) in the detection of active disease [23]. However, osteolytic lesions, which are commonly encountered in NHL, may not be identifiable by this scintigraphic technique [24]. Another limitation of bone scanning is the high incidence of false-positive results, which makes this method very non-specific. CT has been widely used to assess nodal and extranodal involvement of lymphoma, but it has now become clear that the accuracy of PET is substantially greater than this modality in most published series [9–13]. Bone marrow scanning with 99mTc-sulphur colloid, combined with MRI, has also been successfully used, and has been shown to provide better results than BMb, but there are serious shortcomings to this technique, making it impractical in most clinical settings [16]. In addition, MRI findings are non-specific, and disease activity cannot be determined in most instances. Although MRI has been found to be highly sensitive in detecting bone marrow involvement [25], it can only be used if the site of the metastatic focus is suspected in advance, or if there is evidence of diffuse marrow disease. Therefore, its use as a screening test for marrow metastasis is limited.

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PET and biopsy to detect lymphoma in bone marrow Fuster et al. 15

We conclude that PET and BMb are complementary techniques in assessing the presence of bone marrow involvement in patients with malignant lymphoma. In our series, PET was more sensitive than BMb in Hodgkin’s lymphoma and NHL, except in follicular lymphomas.

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References 1

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Original article

Impact of FDG PET for staging of Ewing sarcomas and primitive neuroectodermal tumours Tama´s Gyo¨rkea, Thomas Zajicb, Alexander Langeb, Oliver Scha¨ferc, Ernst Moserb, Erno˜ Mako´a and Ingo Brinkb Aim High-grade Ewing sarcomas and Primitive neuroectodermal tumours (PNET) make up the tumours of the Ewing family. Our purpose was to evaluate the value of [18F]fluorodeoxyglucose positron emission tomography (FDG PET) in patients with Ewing tumours. Patients and methods Twenty-four patients who had PET because of a suspected Ewing tumour during a 5-year period were included in this retrospective study. The images of 33 whole-body FDG PET investigations performed in primary or secondary diagnostics were analysed visually and semi-quantitatively by using standardized uptake values (SUVs). In 14 cases, PET was compared to bone scintigraphy regarding bone lesions. The final diagnosis was based on histology, imaging and follow-up. Results Histologically, the primary lesions were 10 Ewing sarcoma, 13 PNET and one osteomyelitis. The sensitivity and specificity of an examination-based analysis (presence of Ewing tumour and/or its metastases) were 96 and 78%, respectively. Altogether, 163 focal lesions were evaluated. Sensitivity and specificity regarding individual lesions were 73 and 78%. This lower sensitivity is mainly due to small lesions. In true-positive cases, the mean SUV was 4.54 ± 2.79, and the SUVs in two false-positive cases were 4.66 and 1.60. True-positive and false-positive cases could not be differentiated definitively based on SUVs because of

Introduction Positron emission tomography (PET) with 2-[18F]fluoro2-deoxy-D-glucose (FDG) has been widely used in the diagnostic evaluation of various malignancies. Although musculoskeletal sarcomas have also been investigated by PET to improve diagnosis of primary and recurrent disease, to discriminate benign from malignant lesions [1–10], to grade lesions non-invasively [7,9,11–20] and to detect response to therapy [9,21–24], the data about PET investigation of these tumour entities is insufficient, as stated in literature [23,25]. We retrospectively evaluated the whole-body FDG PET investigations performed at the Division of Nuclear Medicine of the Albert Ludwigs University in Freiburg

overlap and low values in true-positive lesions. In four cases, PET depicted 70 while bone scintigraphy depicted only eight bone metastases. Conclusion An FDG PET investigation is a valuable method in the case of Ewing tumours. PET is superior to bone scintigraphy in the detection of bone metastases of Ewing tumours. For the depiction of small lesions, mainly represented by pulmonary metastases, PET is less sensitive than helical computed tomography. Determination of the role of whole-body FDG PET in diagnostic algorithm needs further investigation. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:17–24 Nuclear Medicine Communications 2006, 27:17–24 Keywords: positron emission tomography, [18F]fluorodeoxyglucose, bone scintigraphy, Ewing sarcoma, primitive neuroectodermal tumour a Semmelweis University, Faculty of Medicine, Department of Diagnostic Radiology and Oncotherapy, Budapest, Hungary, and Divisions of, bNuclear Medicine and cDiagnostic Radiology, Albert Ludwigs University, College of Medicine, Department of Radiology, Freiburg, Germany.

Correspondence to Dr Tama´s Gyo¨rke, Semmelweis University, Department of Diagnostic Radiology and Oncotherapy, H-1082 Budapest, U¨llo˜i u´t 78/a., Hungary. Tel: + 0036 1 2100300; fax: + 0036 1 2100307; e-mail: [email protected] Received 25 April 2005 Revised 30 August 2005 Accepted 30 August 2005

between 1996 and 2002 on patients with histopathologically confirmed or suspected tumours of the Ewing family. Ewing tumours are the second most frequent malignant bone tumours in children and young persons. The group of Ewing tumours encompasses morphologically similar subtypes, like Ewing sarcoma, atypical Ewing sarcoma, and malignant peripheral neuroectodermal tumour (PNET). Histologically, they consist of small round cells. A rearrangement of chromosome 22 is characteristic. In cases of atypical Ewing sarcoma, large cells and/or the expression of a neuronal marker are detectable. In PNET, at least two neuronal markers are expressed, while Ewing sarcoma does not express any neuronal markers [26,27]. The aim of this study was to evaluate the diagnostic accuracy of FDG PET in Ewing tumours.

c 2006 Lippincott Williams & Wilkins 0143-3636

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18 Nuclear Medicine Communications 2006, Vol 27 No 1

Methods Patient population

Between January 1996 and June 2002, 47 whole-body FDG PET examinations were performed on 26 patients with a suspected or histologically confirmed Ewing tumour. Fourteen examinations could not be evaluated because of unknown reference state. We were able to include 24 patients (17 males, seven females; age range 6–62 years, mean 28.4 years) with 33 examinations (one in 17 patients, two in six patients and four in one patient) in this retrospective study. The histological diagnoses of primaries were as follows: 10 Ewing sarcoma of the bone, 13 PNET (six soft tissue, seven bone lesions) and one osteomyelitis. Sixteen examinations were performed at the first presentation with the primary lesion and 17 during the course of the disease.

photon attenuation. Coronal, sagittal and transaxial images were produced on the basis of an iterative reconstruction algorithm using ordered-subset expectation maximization and segmented attenuation correction after injection [29]. Whole-body bone scintigraphy was performed 3 h after the intravenous administration of 99mTc 2,3-dicarboxypropane-1,1-diphosphonic acid (99mTc-DPD) (Teceos; Schering, Germany) using a whole-body gamma camera (Siemens Bodyscan). For adults, the dose administered was about 700 MBq. For children, the dose was calculated according to the recommendations of the European Association of Nuclear Medicine [30]. All patients or parents gave their informed consent to all FDG PET and bone scan investigations. Image analysis

Accordingly, 16 examinations were performed without any kind of previous therapy, 17 examinations after chemotherapy (in three cases, completed within 2 months; and in 13 cases, completed between 2 months and 3 years before PET) and/or after radiotherapy (in one case, completed within 3 months; and in four cases, completed between 8 months and 3 years before PET) and/or after surgical intervention (two examinations, within 2 months; and six examinations, between 4 months and 3 years before PET).

The isotope and the radiopharmaceutical were produced and synthesized as reported previously [28]. After the patients had fasted for 12 h, 5 MBq FDG per kilogram of body weight were injected into a peripheral vein. All patients presented with normal blood sugar values at the time of FDG application. No patient had known diabetes mellitus. Patients rested during the 90 min uptake period.

Whole-body PET images were reviewed on hard copy and on a computer workstation (Sun SPARC 20; SUN Microsystems, Palo Alto, California, USA) linked to the data archiving and processing system commercially supplied by Siemens Medical Systems. The latter enabled the use of multiple operator-defined planes. The PET images were interpreted blindly to the results of other imaging studies and independently by two experienced investigators. The physicians interpreted any focally increased radiotracer uptake that exceeded the normal limits of regional FDG accumulation in the area as either benign or malignant. Interpretations were subsequently compared, and consensus was reached by discussion. For semi-quantitative evaluation of increased uptakes, observer-defined regions of interest (ROIs) were outlined around the entire area of suggestive pathological FDG accumulations in transaxial slices. The maximum activity in a selected ROI was chosen for calculation of standardized uptake values (SUVs) [31]. An SUV was compared with lesion size, when the exact size of a lesion at the time of PET investigation was known and available from morphological imaging (n = 33). Bone scans were analysed visually.

Imaging techniques

Reference methods

Static two-dimensional whole-body PET was performed with an ECAT-EXACT 922 tomograph (CTI Siemens, Knoxville, Tennessee). This device simultaneously records 47 planes, which encompass a 16.2 cm field of view. The spatial resolution is about 7.0 mm full width at half maximum. The scanner is calibrated for activity concentration (kBq
ml – 1). Beginning 90 min after tracer injection, an emission image and a transmission image (for subsequent photon attenuation correction using an external germanium source) were recorded at each bed position for 9 and 3 min, respectively. Data were corrected for the dead time of the scanner, decay and

The nature of findings in PET and/or other imaging methods was confirmed by histopathological analysis (in 22 lesions), results of imaging methods, i.e., plain radiography, ultrasonography, computed tomography (CT), magnetic resonance imaging (MRI) and in all but four cases a follow-up of at least 6 months (range 7–39 months, mean 20.8 ± 10.1 months), four patients died after 15–120 days.

The findings of 14 PET examinations in 14 patients were compared with whole-body bone scintigraphy performed 1–21 days (mean, 8.3 ± 7.9) before or after PET. Radiopharmaceutical

Statistical analysis

Diagnostic sensitivity, specificity and accuracy were calculated both on lesion-based analysis and on examination-based

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FDG PET in Ewing sarcomas and primitive neuroectodermal tumours Gyo¨rke et al. 19

analysis. The 95% confidence interval (CI) for each parameter is given. The statistical significance was calculated using the Mann–Whitney U test and a P-value of less than 0.05 was considered significant.

Fig. 1

Results PET investigations and/or reference methods revealed 174 lesions. Of these, 11 detected with PET in nine patients with multiple abnormalities could not be evaluated because of unknown reference state, i.e., no additional investigations were available and chemotherapeutic treatment was performed following PET. Analysing the remaining 163 lesions, 113 were categorized as true positives: 16 primaries (all of the primaries present at the time of PET investigation, Figs 1 and 2), five residual or recurrent lesions, 68 bone metastases, 13 pulmonary or pleural metastases (size, 8–30 mm), four liver metastases (size, 13–15 mm) and seven additional soft tissue metastases. The smallest true-positive lesion (among lesions with known size) was a lung metastasis 8 mm in length. In one case, PET detected more pulmonary or pleural metastases than did spiral CT because of the masking effect of pleural fluid collection at CT. In four cases, a magnetic resonance investigation detected a lesion that was suspected of being recurrent disease and PET classified these lesions as true negatives. Three further PET examinations showed no pathological uptake, considered also to be true negatives based on follow-up results. Two false-positive lesions were depicted in two patients: one osteomyelitis and in another patient a faintly increased uptake in the region of primary tumour without residual disease on follow-up. In seven patients, 41 metastatic or recurrent lesions were undetectable by using PET: 32 pulmonary metastases (smaller than 1 cm) (Fig. 3), six liver metastases (smaller than 1.3 cm), one bone metastasis (5 mm) and, following chemotherapy in one patient, two foci of recurrent Ewing sarcoma each with a diameter of 3 cm. FDG PET had a sensitivity of 73%, a specificity of 78% and an accuracy of 74% (Table 1) on lesion-based analysis. On examination-based analysis 23 examinations were true positives, seven true negatives, two false positives and one false negative, resulting in a sensitivity of 96%, a specificity of 78% and an accuracy of 91% (Table 1).

A Ewing sarcoma of the right humerus in a 23-year-old male patient. The standardized uptake value (SUV) was 5. (a) Coronal MRI, TIRM sequence and (b) coronal whole-body PET image.

The results of a separated analysis for examinations at primary diagnosis and in patients with recurrent tumours are also given in Table 1.

The standardized uptake values of true-positive lesions were between 1.15 and 18.07 with a mean value of 4.54 ± 2.79. The SUVs of lesions with a known size of

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20 Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 2

A primitive neuroectodermal tumour (PNET) of the left thigh in a 32-year-old female patient. The SUV was 18. (a) Transverse and (c) coronal contrast enhanced fat suppressed T1 weighted MRI, and corresponding (b) transverse and (d) coronal PET images.

15 mm, or smaller (n = 13) were between 1.15 and 4.25, mean 2.65 ± 1.12, for lesions larger than 15 mm (n = 20) between 2.46 and 18.07, mean 7.36 ± 4.87. The difference between the SUVs of these smaller and larger sized lesions was significant (P = 0.000081). The SUVs and lesion sizes in patients with the simultaneous manifestations of primary tumours and their metastases are listed in Table 2, demonstrating the heterogeneity of SUVs, even in the same patient at the same examination. Concerning the two false-positive lesions, SUV was 4.66 for osteomyelitis and 1.60 for the other indeterminate lesion. The available 14 whole-body bone scintigraphies detected all 11 primary bone lesions present at the time of investigation, but only eight of 70 bone metastases, revealed by PET and MRI. Sixty-two metastases of four patients remained undetected by bone scintigraphy (Fig. 4).

Discussion Several studies have been published about the FDG PET investigation of sarcomas including mixed histological types of soft tissue [5,7,9–11,15,17,18,20,22] or bone sarcomas [2–5,9,11,15,19–24]. There are publications about tumours of the same histology in case of lipomas and liposarcomas [12] and evaluating the response to neoadjuvant chemotherapy of osteosarcomas [23,24]. There is, however, a paucity of literature on PET scanning in patients with musculoskeletal sarcomas [23,25]. Ewing tumours are high-grade malignancies and, as PET is considered to be a very sensitive tool for detecting high-grade musculoskeletal tumours [8], we performed this retrospective study to evaluate the diagnostic accuracy of FDG PET in patients suffering from tumours of the Ewing family. Generally, according to our results, PET seems to be a valuable tool in the diagnostic work-up of Ewing tumours.

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FDG PET in Ewing sarcomas and primitive neuroectodermal tumours Gyo¨rke et al. 21

Table 2 Sizes (in millimetres) and standardized uptake values (SUVs) of lesions in three of four patients with simultaneous manifestation of primaries and metastases

Fig. 3

Lesion Primary Size SUV Metastasis Size SUV Metastasis Size SUV Metastasis Size SUV Metastasis Size SUV Metastasis Size SUV Metastasis Size SUV

PNET + lung metastases

ES + lung metastases

ES + bone metastases

70 18.1

70 6.29

55 6.30

20 15.1

15 1.22

10 2.10

16 8.86

10 1.15

15 1.45

14 3.84 11 3.05 11 3.90 8 3.73

PNET, primitive neuroectodermal tumour; ES, Ewing sarcoma.

Pulmonary metastases of a humerus Ewing sarcoma in a 9-year-old female patient. Concordant (a) transversal PET and (b) CT images. PET detected only larger metastasis (arrows); the smaller one was detected by CT (arrowhead) but not by PET.

Diagnostic capabilities of fluorodeoxyglucose positron emission tomography (FDG PET)

Table 1

Investigation and analysis

Sensitivity 95% CI

Specificity 95% CI

Accuracy 95% CI

0.73 0.67–0.80 0.96 0.89–1.00

0.78 0.71–0.84 0.78 0.64–0.91

0.74 0.66–0.80 0.91 0.76–0.98

For investigations at primary diagnosis Lesion based analysis 0.58 (n = 58) 0.45–0.71 Examination based analysis 1.00 (n = 16) –

– – – –

0.57 0.43–0.70 0.94 0.70–1.00

For investigations in recurrent disease Lesion based analysis 0.82 (n = 105) 0.75–0.89 Examination based analysis 0.89 (n = 17) 0.74–1.00

0.88 0.81–0.94 0.88 0.72–1.00

0.83 0.74–0.80 0.88 0.64–0.99

For all investigations Lesion based analysis (n = 163) Examination based analysis (n = 33)

Based on the mainly wider ranges of confidence intervals the separated analysis for primary diagnosis and for recurrent tumours is less reliable due to the smaller

number of cases and lesions. Furthermore, no specificities and confidence intervals could be calculated on primary diagnosis patients since these subjects were all positive (by definition) with reference methods for Ewing tumours (Table 1). The somewhat decreased sensitivity in lesion based analysis compared to examination based analysis can be explained mainly by small lesions undetectable for PET imaging. With one exception, the false-negative results were caused by the presence of small lesions (smaller than 13 mm). The smallest lesion detected with PET (among lesions with known size) was 8 mm in diameter. This is a particular problem in the case of small lung metastases, as reported in previous studies [2,3]. A recurrent tumour with two foci of 3 cm size was also not detected by PET. This patient had undergone chemotherapeutic treatment until 3 weeks before PET and subsequent histology confirmed only a small number of viable cells. Measuring the response of Ewing tumours to chemotherapy was beyond the scope of our investigation, but several studies have reported the successful use of FDG PET in measuring therapy response [21–24], so this result can be interpreted more likely as a sign of favourable treatment effect. According to the European guideline for FDG PET tumour imaging [32] radiotherapy can alter FDG uptake for up to 3 months after treatment, and surgery can cause FDG uptake due to healing processes for up to 2 months following intervention. However, we did not find any information concerning how long chemotherapy induced effects might alter FDG uptake into tumours. Based on own experiences and reports on post-therapy FDG PET evaluations of lymphomas [33,34] we presume that

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22 Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 4

interventions. Except for the previously mentioned two foci of recurrent tumour in one patient the false-negative results in the other six patients and the two false-positive lesions could not be explained by therapy induced changes, based on the previous time interval criteria. The most frequent metastatic sites for Ewing tumours are the lungs and bones and this was reflected in our study group as well. In the case of Ewing tumours, bone scintigraphy is the accepted screening method for bone metastases, which is the only whole-body examination in current staging procedures [35]. However, in agreement with the literature [4], PET detected bone metastases in our series with a high sensitivity while bone scintigraphy was rather insensitive. This is not very surprising because PET detects the primary metabolic changes in the bone marrow metastases themselves, while bone scintigraphy only detects the secondary changes in osteoblastic function. In our study the osseous dissemination of the disease was known prior to the PET investigation based on previous MRI findings. PET as a whole-body investigation detected more osseous metastases in further skeletal regions not evaluated by MRI, which, although a very sensitive method for diagnosing osseous metastases without radiation exposure, has a limited use as a method for screening osseous metastases because of the restricted volume imaged. Nevertheless, in the future, with the routine availability of whole-body MRI, it could be a real competitor in this field [3,36]. The specificity of tumour diagnostics with FDG PET is limited by the known accumulation of the radiopharmaceutical in inflammation [7,37]. In this study one of two false-positive findings was described in a patient with a radiologically and clinically suspected Ewing sarcoma of the clavicule. Histology revealed osteomyelitis confirming that the differentiation between osteomyelitis and Ewing sarcoma is also very difficult radiologically [38]. The other false-positive lesion was detected in a female patient with osseous PNET of the foot in the previous location of the primary tumour. The patient had completed high-dose chemotherapy and bone marrow transplantation 2 months before the PET investigation. During a follow-up of 2.8 years without any subsequent tumour therapy she was free of residual or recurrent malignancy, so the elevated FDG uptake was most likely due to inflammation.

Osseal PNET of the left sacro-iliac region in a 17-year-old female patient with multiple bone metastases throughout the skeleton detected by PET. (a) Coronal and sagittal whole-body PET slices and (b) anterior and posterior images of whole-body bone scintigraphy. Bone scintigraphy revealed only the primary at left ileosacral region and metastases only in the left iliac bone and the left femur.

chemotherapy can alter FDG uptake of tumours until up to 2 months. Altogether, we performed six investigations within such a short time period following therapeutic

The use of SUVs in grading musculoskeletal tumours has been investigated in previous studies. Folpe et al. reported heterogeneous SUVs in high-grade sarcomas, but no smaller SUV than 4.22 was found in untreated grade III tumours [15]. Nieweg et al. found a correlation between grade and glucose metabolic rate, but no correlation of grade and SUV [7]. Garcia et al. used a cut-off value of SUV = 3 to distinguish between active sarcomas and post-treatment changes [5], while Lucas et al. used a cut-off value of SUV = 2 to differentiate low-grade and

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FDG PET in Ewing sarcomas and primitive neuroectodermal tumours Gyo¨rke et al. 23

high-grade sarcomas [18]. In a recent meta-analysis Bastiaannet et al. found a significant difference between the SUVs of malignant sarcomas and benign tumours [9]. Lodge et al. found that the SUV and tumour metabolic rate measured at 4 h post-injection were more suitable parameters to help in grading soft tissue masses. Their kinetic analysis showed that the FDG uptake of certain high-grade malignant tumours does not reach maximum levels until approximately 4 h post-injection while benign and low-grade tumours either reached a plateau or falling phase within 30 min [17]. Hamberg et al. demonstrated in eight patients with non-small cell lung cancer that the FDG SUV (or dose-uptake ratio) plateau was not reached by 90 min in the majority of scans [39]. In contrast to other FDG PET tumour investigations performed routinely after an uptake time of 45–60 min [2,4,6–8, 15,18–21,32], we applied a prolonged uptake time of 90 min which may enable improved malignancy detection and more reliable distinction between malignant and benign lesions. Ewing tumours are high-grade malignancies, so elevated glucose metabolism and high SUVs should be expected. In contrast, only some of the SUVs of tumorous lesions in our study were high and they were heterogeneous, even in the same patient. In a previous publication, a theory was outlined about the different behaviour of primaries and their metastases [2] based on the results of another study carried out on lung cancers and their liver metastases [40]. According to this theory, the different behaviour should be a consequence of the different expression of glucose transporters in primary and metastatic sites. In our patient population, the simultaneous manifestation of a primary tumour and its metastases occurred in four cases (see Table 2). In these cases, the SUVs were lower in metastases than in the corresponding primary tumour. However, we also measured relatively low SUVs in several further lesions that were known to be less than 15 mm and we found a significant difference between the SUVs of lesions that were 15 mm or smaller and the SUVs of lesions that were larger. Consequently, the relatively low SUVs might be caused by the partial volume effect and the biological heterogeneity of primary tumours and metastases, as determined previously [41]. The partial volume effect might be further influenced and increased by movement blurring in thoracic and abdominal lesions [2].

was only quantitative, not qualitative, i.e., more pulmonary or bone metastases were depicted by PET in formerly known pulmonary or osseous metastatic states. Summarizing our results and the literature, the inherent advantage of a whole-body FDG PET investigation is the simultaneous investigation of soft tissues and bones. The results and figures of diagnostic accuracy are generally encouraging and PET is very sensitive in the detection of primary and recurrent lesions. In agreement with the literature, there is low sensitivity for small lesions, which is a disadvantage particularly in the screening for lung metastases, so spiral CT takes obvious priority in this regard [2,3,10]. PET remarkably exceeds whole-body bone scintigraphy in its ability to detect bone metastases [4], while, with a limited field of view, routine MRI was also very effective for this purpose [3,36]. PET cannot be used in the non-invasive differentiation between Ewing sarcoma and osteomyelitis, which, radiologically, are frequently indistinguishable [7,37]. SUV is of value in lesion characterization [5,9,15, 17,18,39] although in small lesions SUVs may be decreased due to technical reasons [41]. Evaluation of treatment effectiveness was not the object of this study, but based on literature data the possible role of PET in assessing therapy response is promising and for this purpose SUVs or other quantitative parameters may be essential [9,21–24,31]. In our patient population although PET proved to be very useful in ruling out viable tumour cells in suspected mass lesions and provided a higher degree of diagnostic safety, it did not lead to dramatic changes in patient management. In conclusion, an FDG PET investigation can be recommended whenever it is available, but the determination of its exact role and place in the diagnostic workup of Ewing tumours needs further investigation.

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Franzius C, Schulte M, Hillmann A, Winkelmann W, Jurgens H, Bockisch A, et al. Clinical value of positron emission tomography (PET) in the diagnosis of bone and soft tissue tumors. 3rd Interdisciplinary Consensus Conference ‘PET in Oncology’: results of the Bone and Soft Tissue Study Group. Chirurgie 2001; 72:1071–1077 [in German]. Delattre O, Zucman J, Melot T, Garau XS, Zucker JM, Lenoir GM, et al. The Ewing family of tumors – a subgroup of small-round-cell tumors defined by specific chimeric transcripts. N Engl J Med 1994; 331:294–299. Schmidt D, Herrmann C, Jurgens H, Harms D. Malignant peripheral neuroectodermal tumor and its necessary distinction from Ewing’s sarcoma. A report from the Kiel Pediatric Tumor Registry. Cancer 1991; 68:2251–2259. Hamacher K, Coenen HH, Sto¨cklin G. Efficient stereospecific synthesis of no-carrier-added 2-[18F]-fluoro-D-glucose using aminopolyether supported nucleophilic substitution. J Nucl Med 1986; 27:235–238. Mix M, Nitzsche EU. PISAC: a post-injection method for segmented attenuation correction in whole body PET. J Nucl Med 1999; 40(Suppl):297P. Piepsz A, Hahn K, Roca I, Ciofetta G, Toth G, Gordon I, et al. A radiopharmaceuticals schedule for imaging in paediatrics. Eur J Nucl Med 1990; 17:127–129. Young H, Baum R, Cremerius U, Herholz K, Hoekstra O, Lammertsma AA, et al. Measurement of clinical and subclinical tumour response using [18F]fluorodeoxyglucose and positron emission tomography: review and 1999 EORTC recommendations. European Organization for Research and Treatment of Cancer (EORTC) PET Study Group. Eur J Cancer 1999; 35:1773–1782. Bombardieri E, Aktolun C, Baum RP, Bishof-Delaloye A, Buscombe J, Chatal JF, et al. FDG-PET: procedure guidelines for tumour imaging. Eur J Nucl Med 2003; 30:BP115–BP124. Mikhaeel NG, Timothy AR, O’Doherty MJ, Hain S, Maisey MN. 18-FDG-PET as a prognostic indicator in the treatment of aggressive Non-Hodgkin’s lymphoma – comparison with CT. Leuk Lymphoma 2000; 39: 543–553. Spaepen K, Stroobants S, Dupont P, Thomas J, Vandenberghe P, Balzarini J, et al. Can positron emission tomography with [(18)F]-fluorodeoxyglucose after first-line treatment distinguish Hodgkin’s disease patients who need additional therapy from others in whom additional therapy would mean avoidable toxicity? Br J Haematol 2001; 115:272–278. Paulussen M, Ahrens S, Burdach S, Craft A, Dockhorn-Dworniczak B, Dunst J, et al. Primary metastatic (stage IV) Ewing tumor: survival analysis of 171 patients from the EICESS studies. European Intergroup Cooperative Ewing Sarcoma Studies. Ann Oncol 1998; 9:275–281. Daldrup-Link HE, Franzius C, Link TM, Laukamp D, Sciuk J, Jurgens H, et al. Whole-body MR imaging for detection of bone metastases in children and young adults: comparison with skeletal scintigraphy and FDG PET. Am J Roentgenol 2001; 177:229–236. Guhlmann A, Brecht-Krauss D, Suger G, Glatting G, Kotzerke J, Kinzl L, et al. Chronic osteomyelitis: detection with FDG PET and correlation with histopathologic findings. Radiology 1998; 206:749–754. Durbin M, Randall RL, James M, Sudilovsky D, Zoger S. Ewing’s sarcoma masquerading as osteomyelitis. Clin Orthop 1998; 357:176–185. Hamberg LM, Hunter GJ, Alpert NM, Choi NC, Babich JW, Fischman AJ. The dose uptake ratio as an index of glucose metabolism: useful parameter or oversimplification? J Nucl Med 1994; 35:1308–1312. Kurata T, Oguri T, Isobe T, Ishioka S, Yamakido M. Differential expression of facilitative glucose transporter (GLUT) genes in primary lung cancers and their liver metastases. Jpn J Cancer Res 1999; 90:1238–1243. Hoffman EJ, Huang SC, Phelps ME. Quantitation in positron emission computed tomography: 1. Effect of object size. J Comput Assist Tomogr 1979; 3:299–308.

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Original article 18

F-FEAU as a radiotracer for herpes simplex virus thymidine kinase gene expression: in-vitro comparison with other PET tracers

Anne Rixt Buursmaa, Vera Rutgersa, Geke A.P. Hospersb, Nanno H. Mulderb, Willem Vaalburga and Erik F.J. de Vriesa Objective The herpes simplex virus thymidine kinase (HSVtk) gene has frequently been applied as a reporter gene for monitoring transgene expression in animal models. In clinical gene therapy protocols, however, extremely low expression levels of the transferred gene are generally observed. Consequently, sensitive and selective radiotracers for imaging are required. This study describes the in-vitro evaluation of 20 -[18F]fluoro-5-ethyl-1-b-D -arabinofuranosyluracil (18F-FEAU) as a candidate tracer for HSVtk imaging with positron emission tomography (PET). Methods In cellular accumulation experiments, the potential of 18F-FEAU as a PET tracer for HSVtk was compared to the known acyclic guanosine derivatives 9-[(3-[18F]fluoro-1-hydroxy-2-propoxy)methyl]guanine (18F-FHPG) and 9-[4-[18F]fluoro-3-(hydroxymethyl)butyl]guanine (18F-FHBG), and the thymidine derivatives 30 deoxy-30 -[18F]fluorothymidine (18F-FLT), 20 -deoxy-20 [18F]fluoro-5-methyl-1-b-D-arabinofuranosyluracil (18F-FMAU) and 20 -deoxy-20 -[18F]fluoro-5-iodo-1-b-D -arabinofuranosyluracil (18F-FIAU). For this purpose, C6 control cells and HSVtk-expressing C6tk cells were incubated with the different tracers for various periods of time and cellular uptake and initial uptake rates were analysed. The initial rate of tracer uptake was determined from the slope of the plot of tracer uptake versus incubation time. Results After 2 h of tracer incubation, the C6tk/C6 accumulation ratio was 1.6 for 18F-FLT, 2.4 for 18F-FMAU, 5.5 for

Introduction The herpes simplex virus thymidine kinase (HSVtk) gene has been investigated as a reporter and/or suicide gene by several groups. Preclinical studies with HSVtk gene therapy appeared to be very promising. In clinical protocols, however, the success of gene therapy is limited by difficulties in accomplishing selective and efficient gene transfer to target cells. In order to assess the efficiency of HSVtk gene delivery protocols, it is essential to measure the extent of gene expression, the gene distribution and the HSVtk enzyme activity non-invasively. In this way, gene delivery techniques can be evaluated and optimized and the safety profile can be assessed. In recent years, much progress has been made

18

F-FHPG, 10.3 for 18F-FIAU, 40.8 for 18F-FHBG and 84.5 for 18F-FEAU. The initial tracer uptake rate in C6tk cells was in the order FLT > FMAU > FEAU > FIAU > FHBG > FHPG, whereas the initial tracer uptake rate in C6 control cells was FLT > FMAU > FIAU > FEAUDFHBGDFHPG. The highest HSVtk specific uptake was observed for FEAU. Conclusion This study indicates that the high uptake rate of FEAU together with its high selectivity make this tracer an excellent candidate as a PET tracer for HSVtk c 2006 gene expression. Nucl Med Commun 27:25–30
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:25–30 Keywords: herpes simplex virus thymidine kinase, gene therapy, imaging, radiotracers, 18F-FEAU Departments of aNuclear Medicine and Molecular Imaging and bMedical Oncology, University Medical Center Groningen, University of Groningen, The Netherlands. Sponsorship: This work was financially supported by a grant from the Dutch Cancer Foundation (RUG 2000-2310). Correspondence to Dr E.F.J. de Vries, Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, P.O. Box 30.001, 9700 RB Groningen, The Netherlands. Tel: + 0031 50 3613311; fax: + 0031 50 3611687; e-mail: [email protected]

Received 15 April 2005 Revised 9 August 2005 Accepted 30 August 2005

in the field of non-invasive nuclear imaging of HSVtk gene expression. The tracers that have been developed for imaging of HSVtk expression are phosphorylated by HSVtk in transfected cells only. After phosphorylation, the tracer is unable to cross the cell membrane and is trapped inside the HSVtk-expressing cell. As a result, the tracer accumulates in transfected tissue only. The accumulation of radioactivity can be imaged by positron emission tomography (PET) or single photon emission computed tomography (SPECT) and reflects the HSVtk enzyme activity. The optimal tracer for imaging of HSVtk should have a high affinity for the viral thymidine kinase and a very low

c 2006 Lippincott Williams & Wilkins 0143-3636


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26 Nuclear Medicine Communications 2006, Vol 27 No 1

affinity for host cell thymidine kinase. A pyrimidine nucleoside that has been shown to be highly active against herpes simplex viruses is 20 -fluoro-5-ethyl-1-b-D-arabinofuranosyluracil (FEAU). In a previous study, this compound exhibited similar anti-herpetic activity against herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) to that of 20 -deoxy-20 -fluoro-5-iodo-1-b-D-arabinofuranosyluracil (FIAU) and 20 -deoxy-20 -fluoro-5-methyl-1-b-D-arabinofuranosyluracil (FMAU) [1]. A major advantage of FEAU, however, seems to be a much lower toxicity towards host cells when compared with other pyrimidine nucleosides, because FEAU has been shown to exhibit a much higher affinity for viral thymidine kinases than for host cell enzymes [2,3]. In vivo, FEAU has been found to be metabolically stable [4]. From these studies, FEAU was proposed as a worthwhile candidate for further development as an anti-herpetic agent. Hitherto, the capacity of FEAU as a radiotracer for HSVtk imaging has not been described. However, based on the aforementioned information, we hypothesize that FEAU could also be a suitable tracer for HSVtk imaging. In order to investigate the potential of FEAU as a radiotracer for HSVtk imaging, we intended to compare FEAU with tracers that are known to have affinity for HSVtk and have been previously described in the literature. Such tracers can be divided in two classes: those derived from acyclic guanosine agents and those derived from thymidine, the natural substrate of HSVtk. Previously, we have shown that the acyclic guanosine derivative 9-[(3-[18F]fluoro-1-hydroxy-2-propoxy)methyl]guanine (18F-FHPG) selectively accumulates in HSVtk-expressing tumours [5]. Although FHPG proved to be a very selective tracer for HSVtk-expressing cells, its sensitivity might be insufficient, since the overall uptake of FHPG was rather low (a standardized uptake value (SUV) of r 1.2). In in-vitro experiments, the uptake of the thymidine derivative 20 -deoxy-20 -fluoro-5-methyl-1-bD-arabinofuranosyluracil (FMAU) was higher than the uptake of FHPG in HSVtk-expressing PA-317 and C6 cells [6]. In studies in mice bearing HSVtk-expressing fibrosarcoma and control tumours, accumulation of the thymidine derivative 20 -deoxy-20 -fluoro-5-iodo-1-b-D-arabinofuranosyluracil (FIAU) was up to 119-fold higher than FHPG uptake in the HSVtk transduced tumours [7]. Tjuvajev et al. compared FHPG, FIAU and the acycloguanosine agent 9-[4-fluoro-3-(hydroxymethyl)butyl]guanine (FHBG) as tracers for HSVtk expression. Paired comparisons (background corrected) revealed that FIAU uptake in HSVtk-expressing RG2 glioma xenografts was 18-fold higher than FHBG uptake and 185-fold higher than FHPG uptake, respectively [8]. In in-vitro studies in HT-29 colon cancer cells, FHBG proved to be a more sensitive tracer than FHPG, because the ratio of activity uptake between transduced and non-transduced cells was 2.1-fold higher with FHBG than with FHPG [9]. In a human breast cancer model, biodistribution studies and micro-PET images demonstrated higher total incorporation of FMAU in tumours as compared with FHPG and FHBG. However,

in terms of specificity, FHBG was superior to FMAU [10]. Recently, FHBG also has been shown to be very useful in combination with the HSVtk mutant HSV-sr39tk [11,12]. A mutual comparison of the above-mentioned tracers, however, is difficult on the basis of data reported in literature, because most of the tracers have been evaluated under different conditions and in different models, which can have a great influence on tracer characteristics. Such an effect was demonstrated in a study by Min et al., who reported differences in the relative accumulation of FHBG and FIAU between stably transfected cells and cells that were transiently transfected with adenovirus. The observed differences demonstrate that tracer accumulation depends on how the reporter gene is delivered to the cells [12]. In the present study, we investigated the potential of 20 deoxy-20 -[18F]fluoro-5-ethyl-1-b-D-arabinofuranosyluracil (18F-FEAU) as a radiotracer for HSVtk imaging. The novel thymidine derived tracer FEAU was compared with each of the above-mentioned tracers (the acycloguanosine derivatives FHPG and FHBG, and the thymidine derivatives FMAU and FIAU) under the same experimental conditions. In previous PET imaging studies, FMAU and FIAU have been labelled with 11C and 124I. Because of the possibility of de-iodination of 124I-FIAU and the short half-life of 11C, we labelled all tracers with 18 F in this study. In previous studies, the thymidine analogue 30 -deoxy-30 -fluorothymidine (FLT) has been efficiently phosphorylated by cytoplasmic thymidine kinase [13]. In order to investigate the capacity of HSVtk for the phosphorylation of FLT, we also included 30 deoxy-30 -[18F]fluorothymidine (18F-FLT) in our study. The chemical structures of the radiopharmaceuticals used in this work are shown in Fig. 1.

Methods and materials Dulbecco’s minimum essential medium (DMEM), geneticin (G418) and trypsin (2.5% (w/v)) were purchased from Invitrogen (Merelbeke, Belgium). Fetal calf serum (FCS) was obtained from PAA laboratories (Brunschwig, Amsterdam, The Netherlands). Phosphate buffered saline was purchased from the Department of Pharmacy (University Medical Center Groningen, Groningen, The Netherlands). 18

F-FHPG and 18F-FHBG were prepared according to the procedure described by Alauddin et al. [14]. 18F-FLT was synthesized according to the method of Machulla et al. [15]. 18 F-FMAU, 18F-FIAU and 18F-FEAU were prepared according to the procedure described by Mangner et al. [16]. Cell lines

C6 rat glioma cells obtained from the American Type Culture Collection were cultured in monolayers in

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In-vitro comparison of

Fig. 1

Thymidine derived tracers: O R3

HN HO

N O R2 O

FLT:

R1 = F; R2 = H; R3 =CH3

FMAU:

R1 = H; R2 = F; R3 =CH3

FIAU:

R1 = H; R2 = F; R3 =I

FEAU:

R1 = H; R2 = F; R3 =C2H5

R1 Acyclic guanosine derived tracers: O HN H2N

N N

FHPG:

X=O

FHBG:

X = CH2

F-FEAU with other PET tracers for HSVtk Buursma et al. 27

viable cells under a microscope. The radioactivity in the cell suspensions was measured in a gamma counter (LKB, Wallac, Turku, Finland) and normalized to the number of viable cells. Results are reported as percentage of activity accumulated per 106 cells: Acell 100% Atotal where Acell is the activity in 106 cells and Atotal is the total activity administered to the cell culture. For each tracer, three independent experiments were carried out on different days and each independent experiment was performed in duplicate. The results from the duplicate measurement were averaged and treated as independent variables for calculating the means and standard deviations as plotted in Figs 2 and 3. Statistical analysis

N

Differences in cellular uptake and differences in the initial uptake rate between the tracers were analysed using a two-sided unpaired Student’s t-test. P < 0.05 was considered statistically significant.

X

HO

18

F

The chemical structures of radiopharmaceuticals for nuclear imaging of HSVtk gene expression.

DMEM supplemented with 5% FCS in a humidified atmosphere with 5% CO2 at 371C. HSVtk positive C6 (C6tk) cells were obtained by transfection of the C6 cells with supernatant of PA-317tk packaging cells containing replication incompetent retroviruses carrying the HSVtk gene and the neomycin resistance (NeoR) gene. Subsequent G418 selection resulted in the C6tk cell line. Stable resistance in the C6tk cells was assured by culturing this cell line in the presence of G418 (0.5 mg  ml – 1). The packaging cell PA-317tk was a gift from Dr S.G. Marcus (Genetic Therapy Inc., Gaithersburg, Maryland, USA). Accumulation of tracers in C6 and C6tk cells

C6 cells and C6tk cells were seeded in 12-well culture plates with 1.5 ml of DMEM supplemented with 5% FCS. After 24 h at 371C, monolayers had grown. At time zero, 1.2 ± 0.7 MBq of tracer (specific activity, > 20 GBq  mmol – 1) was added to each well. After incubation times of 15, 30, 45, 60, 90 and 120 min, the culture medium was quickly removed and the monolayers were washed three times with 1 ml of cold phosphate buffered saline. The cells were harvested from the culture plates by treatment with 0.25 ml trypsin. When the cells were detached from the bottom of the well (after approximately 5 min), the cells were resuspended in 1 ml of culture medium to neutralize the trypsin. A 50 ml sample was taken and mixed with 50 ml trypan blue to count the number of

Results The uptake of the tracers in C6tk and control cells after 2 h is shown in Table 1. In C6tk cells, the uptake of the thymidine derived tracers is higher than the uptake of the acyclic guanosine agents. 18F-FLT and 18F-FMAU show the highest uptake in HSVtk-expressing cells. However, these tracers also show a high uptake in C6 control cells, whereas uptake of 18F-FEAU, 18F-FHBG and 18F-FHPG in control cells is negligible. In order to determine the selectivity of the tracers, the ratio between the uptake in C6tk and control cells was calculated as well. The uptake ratio between C6tk and C6 cells was 84.5 ± 5.6 for 18FFEAU, which was significantly higher than for any of the other tracers (P < 0.001). Figure 2 shows the cellular uptake of each tracer versus the incubation time in C6tk cells and in C6 cells. For tracers with high cellular uptake, the curves rapidly start to deviate from linearity. Especially for the thymidine derived tracers, > 10% of the tracer has accumulated in C6tk cells within 2 h. Consequently, pseudo-first-order kinetics do not apply to these tracers any more. As a result, the cellular uptake after 2 h of incubation underestimates the sensitivity and selectivity of the tracers. To correct for this effect, we calculated the initial rate of tracer uptake from the slope of each plot in Fig. 2. Curves following pseudo-first-order kinetics, were fitted according to the formula f ¼ y0 þ ax, where a is the slope, f is tracer uptake, y0 is the intercept of the plot and x is time. Exponential curves were fitted according to the formula f ¼ að1  ebx Þ, where a represents the maximum tracer uptake and where b is a constant (which depends on tracer uptake rate). The slope of the plot fitted at any given time x can be derived from the differential equation

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28 Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 2

Fig. 3

18 40

FLT FMAU FEAU FIAU FHBG FHPG

30

20

10

14 12 10 8 6 4 2 0

0 0

20

40

60 80 Time (min)

100

120

140

(b) 25 Accumulated activity per 106 cells (%)

FEAU FMAU FLT FIAU FHBG FHPG

16 Specific cellular uptake

Accumulated activity per 106 cells (%)

(a)

15

20

40

60 80 Time (min)

100

120

140

The specific uptake of 18F-FHPG, 18F-FHBG, 18F-FLT, 18F-FMAU, 18FFIAU and 18F-FEAU versus incubation time. The uptake in C6 control cells was subtracted from the uptake in C6tk cells. The results are reported as the percentage of activity accumulated per 106 cells. (Abbreviations are as given in the text).

FLT FMAU FIAU FEAU FHBG FHPG

20

0

Table 1 Cellular uptake after 2 h of tracer incubation expressed as percentage of activity accumulated per million cells (mean ± standard deviation)

10 5 0 0

20

40

60 80 Time (min)

100

120

140

The uptake of 18F-FHPG, 18F-FHBG, 18F-FLT, 18F-FMAU, 18F-FIAU and 18 F-FEAU versus incubation time in (a) C6tk cells and (b) C6 cells. The uptake was determined after 15, 30, 45, 60, 90 and 120 min incubation with the tracers at 371C. Results are reported as the percentage of activity accumulated per million cells, according to the equation
Acell=A total 100%. For each tracer, three independent experiments were performed in duplicate. Error bars represent the standard deviation. Curves following pseudo-first-order kinetics were fitted according to the equation f ¼ y0 þ ax. Exponential curves were fitted according to the equation f ¼ að1  ebx Þ. For all curves, r2 > 0.96 and P < 0.001. (Abbreviations and definitions are as given in the text).

Tracer

C6tk cells

C6 cells

Ratio C6tk/C6

FLT FMAU FEAU FIAU FHBG FHPG

30.3 ± 9.7 20.9 ± 4.8 15.9 ± 2.2 9.3 ± 1.4 5.9 ± 0.8 1.0 ± 0.4

18.4 ± 5.5 8.7 ± 1.6 0.19 ± 0.02 0.90 ± 0.17 0.14 ± 0.03 0.17 ± 0.06

1.6 2.4 84.5 10.3 40.8 5.5

Abbreviations as in the text.

The initial tracer uptake rate expressed as percentage of activity accumulated per million cells per min (mean ± standard deviation)

Table 2

Tracer

C6tk cells

C6 cells

Ratio C6tk/C6

FLT FMAU FEAU FIAU FHBG FHPG

0.40 ± 0.07 0.27 ± 0.07 0.21 ± 0.08 0.12 ± 0.03 0.0475 ± 0.0020 0.0068 ± 0.0035

0.25 ± 0.06 0.11 ± 0.02 0.0005 ± 0.0002 0.0065 ± 0.0012 0.0001 ± 0.0005 0.0001 ± 0.0004

1.6 2.5 410 18 475 68

Abbreviations as in the text.

of this formula: df ¼ ab expðbxÞ dx the slope at time zero (initial rate of tracer uptake) equals a  b. The initial uptake rate of the tracers in C6 and C6tk cells and the corresponding ratios are shown in Table 2. In the C6tk cells, the initial uptake rates of the thymidine derived tracer are significantly higher than those of the guanosine derivatives (P < 0.05). The order of initial uptake rates is FLT > FMAU > FEAU > FIAU >

FHBG > FHPG. For all tracers, the initial uptake rate in C6tk cells was significantly higher (P < 0.05) than in C6 control cells, except for 18F-FLT, where the difference in uptake rate between the cell lines approaches significance (P = 0.051). The initial uptake rate of 18FFHPG, 18F-FHBG and 18F-FEAU in C6 cells is negligible, whereas significantly higher uptake rates (P < 0.005) were observed for 18F-FIAU, 18F-FMAU and 18F-FLT. In C6 cells, the order in uptake rate is FLT > FMAU > FIAU > FEAUDFHBGDFHPG.

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In-vitro comparison of

To calculate the specific cellular uptake, the uptake in the control cells was subtracted from the uptake in the C6tk cells. Figure 3 shows the specific tracer accumulation versus incubation time. After 2 h incubation, the specific cellular uptake of 18F-FEAU (15.7 ± 1.7% accumulated activity per 106 cells) was significantly higher than that of 18F-FIAU (8.4 ± 1.3% accumulated activity per 106 cells, P = 0.004), 18F-FHBG (5.8 ± 0.3% accumulated activity per 106 cells, P = 0.0006) and 18F-FHPG (0.8 ± 0.4% accumulated activity per 106 cells, P = 0.0001), which are the most frequently used tracers for HSVtk imaging.

Discussion In this study, we investigated the potential of the thymidine derivative FEAU as a novel PET tracer for HSVtk imaging. The uptake of 18F-FEAU was compared to the known acyclic guanosine derivatives 18F-FHPG and 18 F-FHBG and the thymidine derivatives 18F-FLT, 18FFMAU and 18F-FIAU. Our results show that, in general, the thymidine derived tracers exhibit much higher uptake rates in HSVtk-expressing cells than do the acyclic guanosine derivatives FHPG and FHBG. The uptake rate of the tracers in C6tk cells, i.e., the sensitivity, was in the order FLT > FMAU > FEAU > FIAU > FHBG > FHPG. Differences in sensitivity of the tracers for HSVtkexpressing cells will considerably depend on the affinity of the tracers for the viral thymidine kinase. In a study where antiviral activities of several nucleosides were compared, the affinity of FMAU to HSV-1 was somewhat higher than that of FIAU, but in the same order of magnitude (Ki of 0.59 vs. 0.68) [17]. In a study where FMAU, FEAU and FIAU were compared, the antiherpetic activities against HSV-1 were similar [1]. These findings are in agreement with our results. However, in another study, it was shown that FHPG could efficiently be phosphorylated by a purified HSV-1 thymidine kinase, with a phosphorylation rate of 67% relative to thymidine. [18]. In contrast, our results show that the average initial uptake rate of the thymidine derivatives in HSVtkexpressing cells is more than 17 times higher than that of FHPG. However, differences in cellular uptake between the tracers not only depend on the affinity for HSVtk, but could also be due to differences in cellular accessibility. It was suggested by several groups that tracer transport across cell membranes affects accumulation of radioactivity [19,20]. Several nucleoside transporters are expressed in different cell lines and tissues and the expression of nucleoside transporters can be altered during cell growth and differentiation. Modulation of these nucleoside carriers could influence the cellular accessibility of the tracers [21]. It was also suggested that the transport limitations may affect the acyclic guanosine

18

F-FEAU with other PET tracers for HSVtk Buursma et al. 29

derivatives more than the thymidine derived tracers [8]. In inhibition/competition uptake experiments, we found that FHPG, like acyclovir, is transported by the purine nucleobase carrier [22]. This might be explained by the resemblance in the chemical structures of FHPG and acyclovir. The structure of FHBG is also very similar to acyclovir and FHPG, and therefore it is likely that FHBG also uses a nucleobase transporter to cross the cell membrane. Thus, the acycloguanosine derivatives probably use another transporter to enter the cell than the thymidine derived tracers, which might result in differences in cellular accessibility of the tracers and thus in differences in accumulation in HSVtk-expressing cells. When comparing tracers for HSVtk imaging, not only the affinity and cellular accessibility, but also the contrast between uptake in HSVtk transfected and untransfected cells, the selectivity, is an important factor. The ratios of the tracer’s initial uptake rate in C6tk and C6 cells were in the order FHBGDFEAU > FHPG > FIAU > FMAU > FLT. So, although FLT showed the highest accumulation in C6tk cells, it is an unsuitable tracer for HSVtk because its selectivity is poor. The selectivity of FMAU, which also demonstrated a very high uptake in C6tk cells, is only moderate. Our results show that FLT and FMAU both have a high affinity for kinases in host cells. This characteristic has been exploited for tumour detection. Thymidine kinase activity correlates with the degree of cellular proliferation. Because FLT and FMAU are phosphorylated by the host thymidine kinase, both tracers could be used to image tumour proliferation [23]. Based on our results, FLT could well be a better proliferation marker, because this tracer shows the highest uptake in C6 control cells. However, work has recently been published which indicates that 18F-FLT is not incorporated into DNA and therefore cannot directly measure proliferation, but could be used as an indirect indicator of proliferation potential [24]. FLT is currently under investigation as a new marker for monitoring tumour response to antiproliferative therapy [23]. In our experiments the affinity of FIAU was relatively high, but FIAU showed appreciable phosphorylation by mammalian cellular thymidine kinase as well. This is in agreement with previous reports in the literature [17]. This affinity of FIAU for host thymidine kinase could be a problem in vivo, in the detection of low HSVtk transfection levels in highly proliferating tumours. Still, FIAU is a well-known tracer for HSVtk imaging and has already been applied in humans [25,26]. The selectivity of FHBG was very high in our experiments. Although the affinity was much lower than that of the thymidine derivatives examined, its uptake in C6tk cells was still seven times higher than that of FHPG.

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Nuclear Medicine Communications 2006, Vol 27 No 1

Previously, FHBG has shown to be highly sensitive in combination with the HSVtk mutant HSV-sr39tk [12]. Together with its high selectivity, this makes FHBG a worthy probe for HSVtk imaging. A pharmacokinetic and dosimetry study of 18F-FHBG has already been performed in human volunteers and the results show that 18 F-FHBG is a safe tracer with a desirable pharmacokinetic profile [27]. For FEAU, a high affinity for HSVtk was found in our experiments, comparable to that of FMAU. However, the selectivity was much higher and comparable to that of FHBG. In early literature, biochemical and biological data demonstrate the high selectivity of FEAU against HSVtk. This could be due to the 5-substituent in the structure of the thymidine analogues (R3 in Fig. 1). In FEAU, R3 is an ethyl group. It already has been reported that the size of the 5-substituent is very important for the antiviral activity [1]. Several groups of workers have proposed that FEAU is a worthy candidate for further development as an anti-herpetic agent [1–4]. Based on the information provided in the literature, we hypothesized that 18FFEAU could be a suitable tracer for HSVtk imaging. Our results confirm this hypothesis. FEAU shares the high affinity for HSVtk and cellular accessibility that were found for all thymidine derivatives examined, but demonstrates a superior selectivity. For FEAU, the highest specific cellular uptake was found. This study shows the potential of FEAU as a novel radiotracer for HSVtk imaging and warrants the additional preclinical and clinical testing that is still required before FEAU can be applied in human gene therapy.

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Original article

Discordant localization of 2-[18F]-fluoro-2-deoxy-D-glucose in 6-[18F]-fluorodopamine- and [123I]-metaiodobenzylguanidinenegative metastatic pheochromocytoma sites Marcelo Mamedea, Jorge A. Carrasquilloa, Clara C. Chena, Pedro Del Corralb, Millie Whatleya, Ioannis Iliasb, Alejandro Ayalab and Karel Pacakb Background Although the majority of pheochromocytomas (PHEO) are benign, a subset is malignant. Computed tomography (CT) and magnetic resonance imaging (MRI) localize PHEO with high sensitivity but, because of limited specificity, [131I]- or [123I]-metaiodobenzylguanidine ([131I]or [123I]-MIBG) is often used as a complementary agent. 6-[18F]-fluorodopamine ([18F]-DA) has been developed as a radiopharmaceutical for the targeting of noradrenergic pathways, and has been shown to result in a better detection rate of PHEO sites than MIBG; however, [18F]-DA has shown a lack of accumulation in some patients with metastatic PHEO. Methods Five patients with widespread metastatic PHEO who had CT and MRI evidence of metastatic disease (one man and four women; age range, 25–64 years), and who underwent imaging with [123I]-MIBG, [18F]-DA and 2-[18F]fluoro-2-deoxy-D-glucose ([18F]-FDG), were evaluated retrospectively. Tomographic imaging was performed and positron emission tomography (PET) images were inspected visually and quantitatively. Results All five patients had [123I]-MIBG scans that grossly underestimated the extent of disease when compared with conventional CT and MRI. All lesions seen on [123I]-MIBG scans were detected on [18F]-DA scans, which also

Introduction Molecular imaging, employing nuclear medicine techniques, has been used successfully to localize neuroendocrine tumors [1–7]. In particular, with the development of new positron-labeled radiopharmaceuticals, advances in tumor localization have occurred [8–13]. In contrast to anatomic imaging studies, such as computed tomography (CT) and magnetic resonance imaging (MRI), which rely on size measurements, tissue density and vascular enhancement patterns, these radiopharmaceuticals rely on the targeting of specific receptors, transporters or pathways important in the disease process. In addition to anatomic imaging studies, the diagnostic imaging work-up of pheochromocytoma (PHEO) often involves the utilization of functional imaging studies due

detected additional lesions. Nonetheless, [18F]-DA also failed to detect numerous lesions seen on CT and MRI. In all of these cases, [18F]-FDG PET showed lesions that were not detected on either [123I]-MIBG or [18F]-DA scans. Conclusions When [123I]-MIBG or [18F]-DA fails to localize lesions seen on conventional imaging studies, [18F]-FDG may be recommended as an ancillary test for the diagnosis and localization of metastatic PHEO. This is particularly important in patients with aggressive PHEO. c 2006 Lippincott Williams & Nucl Med Commun 27:31–36 Wilkins. Nuclear Medicine Communications 2006, 27:31–36 Keywords: fluorodeoxyglucose, fluorodopamine, pheochromocytoma, positron emission tomography a Nuclear Medicine Department, Clinical Center and bReproductive Biology and Medicine Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA.

Correspondence to Karel Pacak MD, PhD, DSc, Reproductive Biology and Medicine Branch, National Institute of Child Health and Human Development, CRC, Room 1E-3140, 10 Center Drive, MSC-1109, National Institutes of Health, Bethesda, MD 20892-1109, USA. Tel: + 301 402-4594; fax: + 301 402-4712; e-mail: [email protected] Received 26 July 2005 Accepted 30 September 2005

to the limited specificity of both CT and MRI [14–16]. Thus the work-up of PHEO often includes the use of [131I]- or, more recently, [123I]-metaiodobenzylguanidine ([131I]- or [123I]-MIBG), both of which have high sensitivity and specificity for detecting PHEO [17,18]. Nonetheless, [131I]- and [123I]-MIBG both fail to detect approximately 10–20% of PHEO [6,7]. 6-[18F]-fluorodopamine ([18F]-DA) is a catecholamine precursor that has been developed at the National Institutes of Health (NIH) as a probe for sympathoneuronal innervation [19]. [18F]-DA binds to the cell membrane norepinephrine transporter and is internalized into intracytoplasmic vesicles through the vesicular monoamine transporter. As an imaging agent, [18F]-DA has a high sensitivity for the detection of PHEO and is superior to [131I]-MIBG in the localization of metastatic

c 2006 Lippincott Williams & Wilkins 0143-3636

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32 Nuclear Medicine Communications 2006, Vol 27 No 1

PHEO (16 of 16 versus 9 of 16 patients, respectively), as reported previously [8,9].

repeated. Patients were not on any medication known to block [123I]-MIBG uptake.

However, we have recently observed several patients in whom known metastatic sites of PHEO were negative with [18F]-DA as well as with MIBG. Thus, we considered that 2-[18F]-fluoro-2-deoxy-D-glucose positron emission tomography ([18F]-FDG PET), a functional imaging modality based on glucose metabolism, may be useful to detect metastatic lesions that are not localized by other imaging modalities.

For PET scanning, the patients fasted for at least 4 h before the intravenous injection of [18F]-DA (38.3 ± 1.3 MBq) or [18F]-FDG (458.4 ± 114.6 MBq) and were asked to avoid caffeine, tobacco and alcohol for at least 12 h before the scan. A lower dose of [18F]-DA was used than [18F]-FDG as previous studies showed adequate tumor imaging with this amount. [18F]-DA scanning was performed using an Advance scanner (General Electric Medical Systems) with a 15-cm axial field of view. Eight-minute emission images were obtained in two-dimensional mode from the neck to the pelvis starting approximately 3 min after tracer injection. Three-minute transmission scans were obtained for attenuation correction. The PET images were reconstructed on a 128  128 matrix using an iterative algorithm provided by the manufacturer. [18F]-FDG scanning was performed using a Discovery ST PET/CT scanner (General Electric Medical Systems) with a 15-cm axial field of view. CT studies for attenuation correction and anatomic co-registration were performed without oral or intravenous contrast, and with the following imaging parameters: 140 kVp, 90 mA, 0.8 s per CT rotation and a slice thickness of 3.25 mm. Emission images were also obtained in two-dimensional mode with a 5 min acquisition at each level starting 60 min after injection. The PET images were reconstructed in a 128  128 matrix using an iterative reconstruction algorithm provided by the manufacturer. [18F]-FDG and [18F]-DA scans were performed on different scanners because the [18F]-FDG scans were performed as clinical scans in the Nuclear Medicine Department which uses PET/CT and the [18F]-DA scans were performed in a research study in the PET Department which does not use PET/CT.

Methods We selected five patients (one man and four women; age range, 25–64 years) from a group of patients who had given written informed consent to participate in an NIH Institutional Review Board-approved study evaluating the diagnosis, pathophysiology and molecular biology of PHEO. These patients were selected retrospectively from a group of PHEO patients who had undergone CT, MRI, [123I]-MIBG, [18F]-DA and [18F]-FDG scanning. The selected patients were chosen because they had significantly more lesions noted on their [18F]-FDG than [18F]-DA scans. In a retrospective analysis of their medical records, all patients had undergone previous surgery for PHEO and had aggressive metastatic disease. At NIH, they underwent extensive biochemical testing as described previously [20,21]. In addition to CT and MRI, the five patients underwent [123I]-MIBG scintigraphy, [18F]-DA PET and [18F]-FDG scanning. All scintigraphic studies were performed within 7 days of each other, and CT and MRI scans were performed within 8 days of the scintigraphic studies. CT was performed from the neck to the pelvis with a Hi Speed Advantage scanner (General Electric Medical Systems, Milwaukee, Wisconsin, USA) as described previously, using oral and intravenous contrast [8]. In one patient, an outside CT was reviewed and not repeated at NIH. MRI from the neck to the pelvis was obtained with a 0.5 T scanner (Picker, Highland Heights, Ohio, USA) using multiple sequence and intravenous contrast with gadolinium diethylenetriaminepentaacetate, as described previously [8]. For [123I]-MIBG scanning, patients were imaged at approximately 24 h (and in two cases also at 48 h) following intravenous administration of approximately 10 mCi (370 MBq) of [123I]-MIBG. Both planar and single photon emission computed tomography (SPECT) imaging were performed on a dual-headed gamma camera (ADAC Laboratories, Milpitas, California, USA) or tripleheaded camera (Trionix Corp., Twinsburg, Ohio, USA) equipped with low-energy, high-resolution collimators. One patient had a good quality [123I]-MIBG scan recently performed at an outside facility and the scan was not

Anatomic imaging studies were interpreted by radiologists who were blind to the results of the scintigraphic studies. [123I]-MIBG, [18F]-DA and [18F]-FDG PET studies were each read by two nuclear medicine physicians (J.A.C. and M.M.) during separate reading sessions. These readers were blind to the results of all other imaging modalities. Any tracer accumulation above background that could not be explained by physiologic distribution was considered a possible lesion. Lesions were graded on a scale of 1–5 [1, negative; 2, probably negative; 3, equivocal; 4, probably positive; 5, definitively positive (for tumor)]. Lesions with scores of 4 and 5 were counted as positive findings. Discrepancies were resolved by consensus review of the modality in question. All scintigraphic studies were then reviewed together to correlate the different modalities, and PET/CT was used to determine the anatomical location when lesions were seen on [18F]-FDG scan. The standardized uptake values corrected for the lean body mass (SUVlbm) were calculated (SUVlbm = nCi/g per nCi injected  lean body

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Functional imaging of pheochromocytoma Mamede et al. 33

mass). The maximum SUVlbm value for each lesion was taken and then averaged for each patient.

Results Table 1 summarizes the lesions detected by [18F]-DA and [18F]-FDG PET in each patient. The data for [123I]MIBG are not shown as [18F]-DA detected all foci seen with [123I]-MIBG plus some additional lesions. Figures 1(a) and 1(b) show the relationship between the SUVlbm values calculated in lesions that were seen with both [18F]-DA and [18F]-FDG. In order to determine whether soft tissue and bone lesions behaved differently, we determined the SUVlbm values separately for lymph nodes and bony lesions. No apparent correlation could be observed between the SUVs calculated for lymph nodes and bony lesions with [18F]-DA and [18F]-FDG. Case 1

A 34-year-old man was initially diagnosed in June 2004 with a large heterogeneous mass at the level of the aortic bifurcation (7.7  4.4 cm), which was eroding through the sacrum and was associated with retroperitoneal lymphadenopathy. A CT-guided fine needle aspiration biopsy of the pelvic mass documented a neuroendocrine tumor. One month later, at NIH, a CT scan showed enlargement of the mass anterior to the sacrum (8.2  8 cm) and retroperitoneal lymphadenopathy. In addition, he was found to have a right retrocrural mass (2.3 cm) and a mass in the posterior mediastinum. Biochemical test showed high levels of plasma norepinephrine (2784 pg/ml; normal range, 80–498 pg/ml). Imaging with [18F]-DA did not localize in three lymph node regions detected with [18F]FDG (3.6–6.8 cm in size) and considerably underestimated the extent of bone and soft tissue involvement of the sacral mass when compared with [18F]-FDG (Table 1, Fig. 2). Case 2

A 46-year-old woman had a large PHEO (10  5 cm) resected from the left adrenal in 1990. She did well until

Table 1

Case 3

A 64-year-old woman with a diagnosis of long-standing hypertension perhaps suggestive of catecholamine excess underwent CT in October 2004. This revealed normal adrenal glands and a possible mass near the aortic bifurcation. Two months later, a CT scan showed multiple, new, enlarged lymph nodes in the neck, left axillary and subpectoral regions, the upper abdomen and proximal left common iliac region (2.2 cm). There was a large, heterogeneous, enhancing mass in the pelvis (7.4 cm) and an enhancing lesion near the upper pole of the left kidney. MRI of the spine showed multiple lumbar vertebral body lesions. Biochemical tests showed very high levels of plasma norepinephrine (22 587 pg/ml). [18F]-DA did not localize in the extensive adenopathy in the left axilla, subpectoral area, upper abdomen or pelvis, whereas these areas were well seen on [18F]-FDG

Summary of patients’ 2-[18F]-fluoro-2-deoxy-D -glucose ([18F]-FDG) and 6-[18F]-fluorodopamine ([18F]-DA) imaging results Case 1 [18F]-FDG

Site of lesion Lymph node

Mass Bone Liver SUVlbm

2002, when she developed severe hypertension and MRI showed recurrent tumors in the left suprarenal area (5.4 cm), liver metastases (largest lesion measuring 7 cm) and multiple bony lesions in the lumbar vertebra. [131I]MIBG scan showed increased uptake in the left suprarenal lesion, which was surgically removed. Two months later, she was found to have multiple new soft tissue lesions, and underwent chemotherapy in combination with external beam radiotherapy. Two years later, in 2004, the patient was referred to NIH. A repeat CT scan showed several new liver metastases and a new, complex infrasplenic mass, although the biochemical work-up showed a normal level of plasma norepinephrine (266 pg/ml). Imaging results are shown in Table 1. [18F]-DA scanning did not show uptake in two liver lesions, the right rib lesions or a T-spine lesion, whereas these were all positive with [18F]-FDG. In addition, the infrasplenic lesion was more prominent on [18F]-FDG scan. In retrospect, bilateral cervical nodes that were enlarged on CT were also shown to correspond to hypermetabolic [18F]-FDG nodes on PET/CT, but were negative on [18F]-DA scan.

Neck/supr/axil Mediastinum Abdomen Pelvis

Mean SD

– 2 5 – 1 1 – 18.1 5.4

Case 2

Case 3

Case 4

Case 5

[18F]-DA

[18F]-FDG

[18F]-DA

[18F]-FDG

[18F]-DA

[18F]-FDG

[18F]-DA

– 1 – – 1 – – 17.1 18.4

a

– – – – 1 1 – 5.7 1.1

7 1 5 – 1 5 – 4.9 3.2

1 – – – 1 5 (2b) – 20.0 16.6

2 4 – – – Mult – 8.9 3.8

1 1 – – – Het – 3.2 0.5

1

– – – 1 3 2 3.7 1.5

[18F]-FDG 3 11 6 4 2 – 5.5 3.8

[18F]-DA 2 8 6 (3b) 2 3 (2b) – 9.7 6.9

CT, computed tomography; Het, heterogeneous distribution of the radiotracer; Mass, soft tissue mass visualized in the abdomen or pelvis; MRI, magnetic resonance imaging; Mult, multiple lesions; Neck/supr/axil, lymph nodes in cervical, supraclavicular and axillary regions; PET, positron emission tomography; SD, standard deviation; SUVlbm, maximum standardized uptake value corrected for lean body mass of all lesions seen on both scans. a Lesion recognized on [18F]-FDG PET but not confirmed on CT/MRI. b Lesions recognized on [18F]-DA PET but not confirmed on [18F]-FDG PET or CT/MRI.

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34

Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 1

(a)

Fig. 2

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SUVs of [18F]-FDG (b)

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Single photon emission computed tomography (SPECT) and positron emission tomography (PET) reprojected images from case 1. (a) The [123I]-metaiodobenzylguanidine ([123I]-MIBG) scan shows an abnormal focus of high uptake in the large heterogeneous mass at the level of the aortic bifurcation (arrow head). (b) The 6-[18F]-fluorodopamine ([18F]DA) scan shows the same abnormal high uptake in the soft tissue mass in the abdomen (arrow head). (c) The 2-[18F]-fluoro-2-deoxy-D-glucose ([18F]-FDG) scan shows a larger soft tissue mass which is eroding the sacrum (arrow). Additional foci of activity (not observed with [123I]MIBG or [18F]-DA) are seen in a mass in the posterior mediastinum and in multiple lymph nodes in the abdomen (arrow head).

20 Fig. 3

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SUVs of [18F]-FDG (a) Correlation between standardized uptake values (SUVs) calculated in lesions that were seen with both 6-[18F]-fluorodopamine ([18F]-DA) and 2-[18F]-fluoro-2-deoxy-D-glucose ([18F]-FDG) in lymph nodes (r = 0.145, P = 0.59). (b) Correlation between SUVs calculated in lesions that were seen with both [18F]-DA and [18F]-FDG in bone metastases (r = 0.21, P = 0.97).

scan (Fig. 3); however, [18F]-DA did localize two bony lumbar lesions that were not detected by [18F]-FDG. Case 4

A 25-year-old woman had been diagnosed with PHEO at the age of 6 years with a primary resected at that time. She did well until 2002, when a chest wall paraganglioma was diagnosed and resected. In February 2004, a CT scan showed lesions in the right clavicle, right hilum, right pleura and left hemithorax, and bilateral patchy pulmonary infiltrates. Needle aspiration of the right clavicular lesion was performed and the pathology was consistent with metastatic PHEO. In July 2004, a subsequent CT scan showed progressive disease in the chest with extensive new bony lesions throughout the skeleton and new soft tissue masses in the right iliac fossa. Her workup showed an extremely high level of plasma norepinephrine (141 299 pg/ml). Imaging results are shown in

(a) The [123I]-metaiodobenzylguanidine ([123I]-MIBG) planar torso view shows abnormal uptake in a large heterogeneous mass in the pelvis posterior to the urinary bladder (black lower arrow) and two lesions in the lumbar spine (black middle arrows). (b) The reprojected 6-[18F]fluorodopamine ([18F]-DA) image shows the same abnormal uptake in the pelvis posterior to the urinary bladder (black lower arrow) and the two lesions in the lumbar spine (black middle arrows). In addition, faint uptake in a lymph node in the supraclavicular region (black upper arrow) is observed. In the abdomen and inguinal region, two faint areas of uptake were seen but were negative on 2-[18F]-fluoro-2-deoxy-Dglucose ([18F]-FDG) scan and computed tomography/magnetic resonance imaging (CT/MRI) (b Table 1). (c) The reprojected [18F]-FDG image shows similar increased uptake in the pelvic mass and in the lumbar spine (arrows). In addition, areas that were not visualized with [18F]-DA but were seen with [18F]-FDG include multiple lymph nodes in the neck, left axillary and subpectoral regions, which were confirmed as pheochromocytoma by fine needle aspiration (black arrow heads), and in the upper abdomen and iliac region (white arrow heads).

Table 1. [18F]-FDG showed more extensive disease with better contrast in all sites than did [18F]-DA. The diffuse bone involvement was much better recognized with [18F]-FDG, whereas, with [18F]-DA, some sites, such as

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Functional imaging of pheochromocytoma Mamede et al. 35

in the ribs, were not detected at all. [18F]-DA also failed to detect a lung lesion and several mediastinal lesions, as well as the pelvic extension of bony disease that was visualized with [18F]-FDG. On [18F]-FDG scan, there was also extensive uptake in the abdomen that overlapped mesenteric fat; the etiology of this uptake could not be determined. The patient underwent external beam radiation therapy of the upper body, but died in December 2004, 2 months after admission. Case 5

A 54-year-old woman had a history of PHEO diagnosed in 2002 in her right adrenal gland (5.5  5  2.2 cm), which was surgically removed. Two years later, she was found to have metastatic lesions in the left adrenal gland as well as bony lesions. She received a therapeutic dose of [131I]MIBG outside of NIH in 2004. However, after the therapeutic dose, a new CT scan showed multiple, new, mediastinal, retrocrural and para-aortic lymphadenopathies. Biochemical tests showed high levels of plasma norepinephrine (888 pg/ml). Compared with [18F]-DA, [18F]-FDG visualized more lymph node metastases. On the other hand, [18F]-DA showed better contrast than [18F]-FDG in bone lesions, and localized a few additional sites in bone that were not originally identified on [18F]FDG. Retrospective review of these sites on [18F]-FDG PET/CT showed that they were mildly hypermetabolic.

Discussion In this case series, we describe five patients selected to illustrate an imaging pattern of increased uptake of [18F]FDG in sites that were negative on [18F]-DA and [123I]MIBG scans. Clinically, these patients had aggressive metastatic PHEO, often with involvement of soft tissues and bones. The incidence reported for metastatic PHEO ranges (depending on the genetic background and tumor location) from 3 to 36% [22–26]. The survival rate for metastatic PHEO depends on the location of metastatic lesions. Short-term survivors (less than 2 years) tend to be those patients with metastatic lesions in the liver and lungs, whereas long-term survivors (more than 20 years) are those with metastatic lesions in bone. The overall 5-year survival rate varies between 34 and 60% [3,23]. This poor prognosis emphasizes the need to adequately identify those with metastatic disease. Several functional imaging methods using PET agents are available for the detection of PHEO [8–13]. Previous studies from this institution of [18F]-DA PET scanning in patients with known or suspected PHEO have shown the high sensitivity and specificity of this agent for both benign and metastatic disease [8,9]. Amongst the advantages of [18F]-DA for the imaging of PHEO is that it appears to be a better substrate for the cell membrane norepinephrine transporter than other compounds [27]. Despite its high sensitivity, however, we have recently

observed several cases in which [18F]-DA failed to localize in known metastatic sites of PHEO. The uptake of [18F]-FDG by cancer cells is based on an increased metabolic rate of glucose [28]. Several case reports and articles have shown that [18F]-FDG successfully localizes PHEO with a sensitivity in the region of 72% [11,29–31]. Shulkin et al. [11] demonstrated that it has a higher sensitivity for metastatic PHEO (82%) than for benign PHEO (58%). As in all tumors, however, [18F]FDG is non-specific. Shulkin et al. [11] also showed that, in benign PHEO, although [18F]-FDG localized 58% of cases, MIBG localized 83%; in contrast, in malignant PHEO, [18F]-FDG localized 82%, whereas MIBG localized 88%. In the series of Shulkin et al. [11] described above, there were four cases in which [18F]-FDG localized PHEO, which did not accumulate MIBG. Similarly, in this report, we describe five cases of metastatic PHEO that demonstrated discordant imaging results between [123I]-MIBG and [18F]-DA versus [18F]-FDG. One explanation for the imaging pattern of increased [18F]-FDG uptake in sites that are negative on [123I]-MIBG and [18F]-DA could be dedifferentiation of the tumor, with the loss of specific cellular characteristics, such as the cell membrane norepinephrine and vesicular monoamine transporter systems responsible for the uptake of [123I]-MIBG and [18F]-DA. Such dedifferentiation may be especially prevalent in aggressive cases of malignant PHEO, and, indeed, the five patients described in this series had aggressive metastatic PHEO, often with widespread involvement of soft tissues and bones. Although we have yet to confirm the possibility of dedifferentiation histologically, we hypothesize that this is what may have occurred in our patients. In patients who lose the ability to accumulate [18F]-DA and MIBG, [18F]-FDG could become the preferred functional imaging method to detect these lesions. On the other hand, we also observed a few lesions in these patients that accumulated [18F]-DA and not [18F]-FDG (Table 1), suggesting that individual lesions in the same patient can behave differently from each other; however, most of our patients’ positive [18F]-DA lesions also avidly accumulated [18F]-FDG. This is in accordance with studies that show that many MIBG-positive PHEO will also accumulate [18F]-FDG [11]. Although we used two different scanners (PET/transmission pin and PET/CT), the resulting [18F]-FDG attenuation-corrected images were of high quality and the differences observed were not subtle and could not be attributed to technical scanner differences. These cases illustrate a potential role for [18F]-FDG in a subset of patients with metastatic PHEO in whom highly

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specific functional imaging agents, such as [18F]-DA and MIBG, failed to localize sites of disease, a potentially significant problem when assessing a patient’s extent of disease or response to therapy. Our findings suggest that, in MIBG-negative patients, [18F]-DA and/or [18F]-FDG may be useful as a complementary modality. Issues raised in this article that require addressing in the future include the histologic confirmation of our hypotheses and the determination of whether these findings are seen with benign PHEO. Because we retrospectively selected patients with discordant imaging results and did not obtain [18F]-FDG scans in all our metastatic PHEO patients, there may be a selection bias, and thus the true incidence and nature of these discordant findings will require prospective evaluation of a larger series of consecutive patients.

Conclusion In conclusion, this case series shows that some cases of aggressive PHEO can be both [123I]-MIBG and [18F]-DA negative, and that, in these cases, [18F]-FDG may be useful in the diagnostic localization of metastatic PHEO. These findings may further contribute to a better diagnostic imaging algorithm in patients with PHEO.

10

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16 17

18

19

20

Acknowledgements The authors would like to thank Miss Karen T. Adams for her assistance with medical records and care of the patients, Dr Roberto M. Maas for his assistance with the technical aspects of the scans, and the PET and nuclear medicine radiopharmacists and technologists for technical assistance in performing patient imaging.

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Original article

A region-of-interest template for three-dimensional stereotactic surface projection images: Initial application to the analysis of Alzheimer’s disease and mild cognitive impairment Takao Kubota, Yo Ushijima, Chio Okuyama and Tsunehiko Nishimura Objective To construct a region-of-interest (ROI) template for Z-score images of three-dimensional stereotactic surface projections (3-D SSP) and to assess whether the ROI template can be a useful tool for evaluation of brain perfusion abnormalities of neurological disorders. Materials and methods We constructed the ROI template for Z-score images of 3-D SSP based on the standardized magnetic resonance imaging data of 10 healthy volunteers. We assigned a total of 26 ROIs to Z-score images and superimposed it on Z-score images constructed from the brain perfusion SPECT data of 15 patients with Alzheimer’s disease and 10 patients with mild cognitive impairment (MCI) who developed Alzheimer’s disease within the following 2 years. We then obtained the mean Z-scores of each ROI and examined them to determine whether the hypoperfusion typical of Alzheimer’s disease had been demonstrated quantitatively. We also visually inspected the Z-score image of each patient in both groups to determine whether the areas with the highest Z-scores were demonstrated within the ROIs of regions typical of Alzheimer’s disease. Results In the patients with Alzheimer’s disease, our ROI template quantitatively demonstrated hypoperfusion in regions typical of the disease and the Z-scores were very

Introduction Region-of-interest (ROI) analysis has been used to assess brain-function images, such as those obtained during brain perfusion studies using single photon emission computed tomography (SPECT) and in glucose metabolism studies using positron emission tomography (PET), but it has several disadvantages. ROI analysis does not usually include the entire brain, and since the data sampling is regionally biased, a priori hypothesis-based investigations are unavoidable, and unexpected results are unlikely to be revealed. In addition, reproducibility and consistency among investigators are limited due to its observer dependency. A three-dimensional stereotactic surface projection (3-D SSP) program was developed by Minoshima et al. to solve the problems associated with conventional ROI analysis and the 3-D SSP technique was described in detail [1]. Briefly, it involves three major steps. First, there is stereotactic re-orientation of each

high. In the MCI patients, the mean Z-scores of the ROI in the posterior cingulated gyrus were the highest among all regions. Visual inspection of the Z-score images of each patient in both groups confirmed that the areas with the highest Z-scores were demonstrated within the ROIs in regions typical of Alzheimer’s disease in all cases. Conclusion Use of 3-D SSP methods and our ROI template enables automated quantitative evaluation of brain function images over the entire brain surface. In addition, the ROI template may facilitate visual interpretation of functional images of individual patients with neurological c 2006 disorders. Nucl Med Commun 27:37–44 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:37–44 Keywords: ROI template, 3-D SSP, SPECT, Alzheimer disease Department of Radiology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Japan. Correspondence to Dr Takao Kubota, Department of Radiology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, 465 Kajii-cho, Kawaramachi-Hirokoji Kamigyo-ku Kyoto 602-8566, Japan. Tel: + 0081 75 251 5620; fax: + 0081 75 251 5840; e-mail: [email protected] Received 1 December 2004 Accepted 21 June 2005

brain into a standard shape by non-linear warping. Second, a method of data extraction is performed in which cortical activity is projected onto the brain surface. Finally, the cortical projections obtained are compared with a normal database on a pixel-by-pixel basis, and Z-score images are obtained. A Z-score is calculated for each surface pixel: Z-score = {(normal mean) – (individual value)/(normal standard deviation)}, and regions of hypoperfusion or hypometabolism are demonstrated as high Z-score areas on Z-score images. 3-D SSP programs allow analysis of functional images on a pixel-by-pixel basis over the entire brain. Because processing with 3-D SSP is fully automated and the results of the analysis do not depend on observers, 3-D SSP provides completely objective data. These advantages have recently led to 3-D SSP becoming a useful method for both clinical and research purposes [1–7].

c 2006 Lippincott Williams & Wilkins 0143-3636

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Reduced metabolism in the posterior cingulate gyri may be visualized on Z-score images of 3-D SSP in a very early stage of Alzheimer’s disease, even before a clinical diagnosis of probable Alzheimer’s disease is possible [4], and excellent diagnostic performance by this technique in visual image interpretation in individual cases has been shown already in previous reports [1,2]. Excellent diagnostic performance in individual cases has not been obtained with other analytical methods, such as statistical parametric mapping (SPM), which was originally developed for activation studies and data comparison between groups. 3-D SSP, however, has its own shortcomings. Since there are no anatomical landmarks in the current 3-D SSP image displays, it is difficult for researchers to analyse regional Z-scores quantitatively. The lack of anatomical landmarks for Z-score images has disadvantages not only in quantitative analysis but in diagnosis based on visual evaluation. When Z-score images in an individual case visualize areas of abnormal perfusion or metabolism, it is difficult for novice interpreters to identify the precise functional areas of the brain to which they correspond. To solve these problems with current 3-D SSP programs, in this study we constructed a ROI template for Z-score images of 3-D SSP based on the standardized brain atlas of Talairach and Tournoux, and investigated whether the ROI template can be a useful tool to assess brain perfusion abnormalities of neurological disorders.

Materials and methods Standardized surface brain magnetic resonance images of healthy volunteers

Magnetic resonance images and N-isopropyl-p-[123I] iodoamphetamine (123I-IMP) brain perfusion SPECT images of 10 healthy volunteers were used to obtain standardized stereotactic surface magnetic resonance images of the brain. The 10 volunteers consisted of five young adults (three men and two women; age, 20–31 years) and five elderly individuals (two men and three women; age, 66–72 years). None of them had a history of neurological or psychiatric disorders or major medical illness, and the results of a neurological examination on the day of the magnetic resonance and SPECT imaging were normal. The magnetic resonance images were obtained using a 1.5 T imager (Gyroscan Intera, Philips Medical Systems) with 3-D T1 weighted fast-field-echo sequence (TR/TE: 25/4.6, flip angle: 30, field of view: 230  184 mm, matrix: 256  205, pixel size: 0.9  0.9 mm, section thickness: 2 mm). Brain perfusion SPECT was performed by intravenously injecting 185 MBq of 123I-IMP (Nihon Mediphysics, Hyogo, Japan) with the subjects seated at rest with eyes open. SPECT imaging was commenced 22 min after the injection and continued for 16 min. A triple-head gamma camera (PRISM IRIX, Picker International, Cleveland, USA) and low energy, high resolution, parallel collimator were used. Projection data from each camera were obtained in a

128 128 format for 40 angles of 1201 at 8 s per angle (voxel size: 2 mm  2 mm  2 mm). Scatter correction was performed by the triple energy window method. The transaxial images were reconstructed by using the ordered subsets expectation maximization algorithm with an iteration number of 8 and subset of 10, and a post-reconstruction Butterworth filter was used. Attenuation correction was performed by Chang’s method. The magnetic resonance and SPECT images of each of the 10 volunteers were read by two experienced neuroradiologists (O.K. and K.Y.), and no abnormal findings were detected. We obtained standardized stereotactic brain surface magnetic resonance images of the volunteers by the method reported by Minoshima et al. [1]. Briefly, structures such as the scalp and bone marrow on the magnetic resonance imaging set were removed by applying a threshold suitable for delineating cortical margins, and residual extra-cerebral pixels on the magnetic resonance imaging set were removed manually. The right hemispheres of the brain were removed, and the left hemispheres were transposed to the right side to create symmetrical brain images. The magnetic resonance imaging sets were then transformed to binary image sets by assigning a value of 1 to pixels with intensity higher than that of cerebrospinal fluid and 0 to the rest of the pixels. Anatomical standardization techniques used in the 3-D SSP program (NEUROSTAT) [8–10] were applied to the SPECT images to create transformation parameters. These parameters were then used to transform the magnetic resonance image sets to the standard stereotactic atlas coordinates. As a result we obtained a standardized binary magnetic resonance image set for each healthy volunteer. The NEUROSTAT program was also used to transform the magnetic resonance image data. Construction of the ROI template for the 3-D SSP Z-score images

By viewing the standardized binary magnetic resonance image set in three dimensions, we obtained the standardized stereotactic brain surface images of each healthy volunteer (Fig. 1), and these surface magnetic resonance images were used to construct a ROI template for the Z-score images of 3-D SSP. We planned to assign a total of 26 ROIs (14 ROIs for lateral aspects and 12 ROIs for medial aspects) (Table 1) with the objective of reflecting the functional areas that had been reported as being characteristically affected in various neurological disorders, such as Alzheimer’s disease [4,11,12], dementia with Lewy bodies [13], Parkinson’s disease [14], frontotemporal dementia [15], corticobasal degeneration [16], progressive supranuclear palsy [16], and other psychoneurological diseases [17]. By referring to the standardized brain atlas of Talairach and Tournoux [18], we used many sulci identified on

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Applications of an ROI template for 3-D SSP images Kubota et al. 39

Fig. 1

(a)

Lateral

Superior

Medial

(b) 4 2

6 3

1 5

9

10 11

3

4

7

7

8 12

13

14

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8 11 12

6

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Medial

Lateral

Medial

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(a) The standardized surface brain magnetic resonance images of a healthy volunteer are viewed in three dimensions from the lateral, superior and medial aspects. (b) The ROI template we constructed from magnetic resonance data sets of 10 healthy volunteers. A total of 26 ROIs (14 ROIs for the lateral aspect and 12 ROIs for the medial aspect) was assigned. The number of each ROI corresponds to the ROI number in Table 1. (c) The ROI template superimposed on standardized magnetic resonance surface images of a healthy volunteer is shown. Note that the boundaries of each ROI properly fit the sulci.

standardized surface magnetic resonance images as the boundaries of ROIs, including the pre-central sulcus, the post-central sulcus, Sylvian fissure, the intraparietal sulcus, the cingulate sulcus, the superior frontal sulcus, the inferior frontal sulcus, the superior temporal sulcus, and the parieto-occipital sulcus. We referred to several previous articles [19–21] to accurately identify these sulci on each surface magnetic resonance images. These sulci were identified on each standardized surface magnetic resonance image of 10 volunteers and transferred to a coordinate plane. The mean position of each sulcus of the 10 volunteers on the coordinate plane were determined as ROI boundaries, and the construction of the ROI template were completed (Fig. 1). By using the NEUROSTAT program, the ROI template we constructed can be automatically superimposed on Z-score images of 3-D SSP. Assessment of the usefulness of the ROI template

To determine whether the ROI template for 3-D SSP image sets is useful for assessing brain perfusion

abnormalities in neurological diseases, we applied it to 3-D SSP image sets constructed from the brain perfusion SPECT data of 15 patients with Alzheimer’s disease (six men and nine women; mean age, 73.1 ± 6.9; Mini-Mental State Examination score, 18.9 ± 4.1) and 10 patients with mild cognitive impairment (MCI) (four men and six women; mean age, 75.2 ± 4.9; Mini-Mental State Examination score, 27.1 ± 1.5) who had developed Alzheimer’s disease during the following 2 years, and investigated whether our ROI template would demonstrate the characteristic abnormal pattern of Alzheimer’s disease. All 15 patients with Alzheimer’s disease included in our study satisfied the criteria for probable disease of the National Institute of Neurological and Communicative Disorders and Stroke (NINCDS) and the Alzheimer’s Disease and Related Disorders Association (ADRDA). Referring to previous reports describing the abnormal perfusion pattern in Alzheimer’s disease [4,11,12], we defined the typical perfusion pattern of the disease as bilateral parietotemporal lobe and/or posterior cingulate gyrus hypoperfusion with relative sparing of

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Table 1 Regions of interest for Z-score images of three-dimensional stereotactic surface projections Surface Lateral surface

Region of interest

Brain area

Brodmann area

1 2

9, 10 8, 9, 46

12 13 14

Prefrontal cortex Superior and medial frontal gyrus Broca’s area Premotor area Orbitofrontal cortex Primary sensorimotor cortex Superior parietal lobule Inferior parietal lobule Anterior temporal area Superior temporal association area Inferior temporal association area Posterior temporal lobe Occipital lobe Cerebellum

1 2 3 4 5 6 7 8 9 10 11 12

Inferior prefrontal cortex Superior prefrontal cortex Supplementary area* Primary sensorimotor cortex Orbitofrontal cortex Medial temporal area Precuneus Occipital lobe Anterior cingurate gyrus Posterior cingulate gyrus Brain stem Cerebellum

3 4 5 6 7 8 9 10 11

Medial surface

44, 45 6 11, 47 1, 2, 3, 4 5, 7 39, 40 38 22, 41, 42 20, 21 37 19

9, 10 8, 9 6, 8 1, 2, 3, 4 11, 12 38 7 17, 18, 19 24, 32, 33 23, 29, 30, 31

we used Z-score images with count normalization to the global brain area. A normal database for 3-D SSP analysis of patient data was constructed from the brain perfusion SPECT data of 20 healthy subjects (10 men and 10 women; mean age, 72.1 ± 6.2; Mini-Mental State Examination score, 29–30). None of these healthy subjects had a history of neurological or psychiatric disorders or major medical illness, and the results of their neurological examinations were normal. None of the healthy subjects had abnormal findings on brain perfusion SPECT and magnetic resonance images, including T1 weighted and T2 weighted magnetic resonance images.

Results The results of the quantitative analysis of the Z-score images with our ROI template are shown in Figs 2 and 3. Typical hypoperfusion was demonstrated quantitatively in Alzheimer’s disease. The mean Z-scores of the posterior cingulate gyrus and parietotemporal regions were higher than in the other regions, and the mean Zscores of the primary sensorimotor cortex, occipital lobe and cerebellum were relatively low. The mean Z-scores of the posterior cingulate gyrus, superior parietal lobule, inferior parietal lobule and inferior temporal association area were significantly higher than in the primary sensorimotor cortex.

*

A part of frontal eye field is included in ROI 3 (medial surface).

the primary sensorimotor cortex, occipital lobe, and cerebellum. All 10 MCI patients included in our study satisfied the commonly accepted diagnostic criteria for MCI defined by Petersen et al. [22] at the time of brain perfusion SPECT and met the NINCDS and ADRDA criteria for probable Alzheimer’s disease after clinical follow-up for 1 to 2 years. 123I-IMP brain perfusion SPECT data of Alzheimer’s disease and MCI patients were acquired by the same imaging protocol used for SPECT imaging of the healthy volunteers whose magnetic resonance data were used for construction of the ROI template. Each Zscore image of all the patients was automatically obtained by analysing the acquired SPECT data with the 3-D SSP program (NEUROSTAT). We again used NEUROSTAT to automatically superimpose our ROI template on their Z-score images, obtained the mean Z-scores in each ROI, and assessed whether the typical hypoperfusion pattern of Alzheimer’s disease was quantitatively demonstrated. We compared the mean Z-scores of each ROI with those of the primary sensorimotor cortex by Student’s paired two-tailed t-test, and P < 0.05 was considered statistically significant. We also visually assessed whether the areas with the highest Z-scores in each patient were demonstrated within the ROIs of typical hypoperfusion regions of Alzheimer’s disease (parietotemporal regions and posterior cingulate gyrus). In all of these evaluations,

The mean Z-scores in MCI were highest in the posterior cingulate gyrus, and they were significantly higher than in the primary sensorimotor cortex. Visual inspection of the Z-score images of each patient confirmed that the areas with the highest Z-scores were within the ROI of the typical hypoperfusion regions of Alzheimer’s disease (parietotemporal regions and posterior cingulate gyrus). The Z-score images of an Alzheimer’s disease patient and an MCI patient with our ROI template are shown as examples in Figs 4 and 5.

Discussion The 3-D SSP program allows analysis of functional images, including perfusion SPECT and glucose metabolism PET images, on a pixel-by-pixel basis over the entire brain. Since the process is fully automated and statistical, the 3-D SSP program enables completely objective evaluation of functional images, which is the most outstanding advantage of these programs. However, current Z-score image displays of 3-D SSP contain no landmarks that would indicate the corresponding location of each brain-function unit, and this has been a drawback of 3-D SSP and made it difficult for researchers to quantitatively analyse Z-score images of 3-D SSP by each functional unit of the brain. The ROI template we constructed in this study automates quantitative analysis of Z-score images of 3-D SSP by each functional unit. Use of the 3-D SSP technique and our ROI template for

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Applications of an ROI template for 3-D SSP images Kubota et al. 41

Fig. 2

ROl 12

ROl 11

ROl 9

ROl 8

ROl 7

ROl 6

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ROl 5

ROl 4

ROl 3

ROl 2

ROl 1

∗

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2 1.8 1.6 1.4 1.2 1 0.8 0.6 0.4 0.2 0

∗

ROl 2

∗

ROl 1

2 1.8 1.6 1.4 1.2 1 0.8 0.6 0.4 0.2 0

ROl 10

∗

(b)

(a)

The mean Z-scores of each ROI in 15 AD patients with Alzheimer’s disease. (a) Lateral aspects; (b) medial aspects. The ROI numbers correspond to the ROI numbers in Table 1. *Significantly higher than the sensory motor cortex.

Fig. 3

1.4

ROl 12

ROl 11

ROl 10

ROl 9

ROl 8

ROl 7

ROl 6

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∗

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1

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1.2

ROl 2

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ROl 2

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(a)

The mean Z-scores of each ROI in 10 patients with mild cognitive impairment. (a) Lateral aspects; (b) medial aspects. The ROI numbers correspond to those in Table 1. *Significantly higher than the sensory motor cortex.

analysis of SPECT or PET images automates quantitative evaluation not only of the regional abnormalities of neurological diseases but of longitudinal changes in individual cases, including disease progression and results of treatment. 3-D SSP image sets are known to be useful for diagnosis in individual cases [1,2], and this is an outstanding characteristic of the 3-D SSP technique. Usefulness for diagnosis of individual cases has never been reported for other image analysis methods, such as SPM. Our ROI template for 3-D SSP image sets can be used not only for automated quantitative analysis but for visual image interpretation in individual patients, and that is another

important advantage of our ROI template. Many neurological disorders, including Alzheimer’s disease, dementia with Lewy bodies, frontotemporal dementia, corticobasal degeneration, progressive supranuclear palsy, and other neurological and psychoneurological diseases, exhibit characteristic abnormal findings on brain functional images [4,11–17]. These characteristic perfusion or metabolic changes are often very helpful in making the diagnosis, and the precise localization of abnormalities is important in image interpretation. Superimposing our ROI template on the patients’ Z-score images facilitates precise identification of the functional parts of the brain to which abnormal areas correspond. In the present study, we visually confirmed that the areas with the highest

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42 Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 4

(a)

Right lateral

Left lateral

Left medial

Right medial

Z 1

(b)

Right lateral

2

3

Left lateral

4 Left medial

5 Right medial

Z 1

2

3

4

5

(a) Z-score images of a patient with Alzheimer’s disease (71-year-old male; MMSE score, 19). (b) The same Z-score images with our ROI template. The areas with high Z-scores are within the ROIs of the posterior cingulate gyrus, parietal lobe, and temporal lobe, and there is no area with high Z-scores within ROIs of the primary sensorimotor cortex, occipital lobe, or cerebellum, indicating that the boundaries of the ROIs have been properly assigned.

Z-scores in 3-D SSP images were within the ROI of regions typical of Alzheimer’s disease in all patients with the disease or with MCI. This indicates that superimposing our ROI template greatly facilitates visual interpretation of 3-D SSP images, at least in Alzheimer’s disease and MCI. There are several problems when an ROI template is constructed on standardized brain maps. First, it is impossible to avoid the effect of a margin of error in the process of image standardization. A previous study revealed that while the effect of a margin of error is smaller with the standardization technique of 3-D SSP methods and SPM than with other standardization techniques, such as linear transformation and the human brain atlas standardization technique, standardization is imperfect even with 3-D SSP methods [23]. Since the effect of a margin of error in the process of image standardization cannot be avoided, it is very important to be careful not to construct ROIs that are too small. If such ROIs are used in a quantitative analysis, the results are likely to be greatly affected by the margin of error resulting from the process of standardization. However, a large ROI that includes several different functional parts of the brain may not satisfy the demands of

neurological researchers and clinicians. In the present study, taking care not to construct a ROI that includes several different functional parts, we constructed relatively large ROIs. The second problem involved in constructing an ROI template is the individual difference in the anatomy of brain structures, including the sulci and gyri. Since our ROI template was constructed from the magnetic resonance data set of volunteers, the boundaries of each ROI may have been affected by the specificity of the individual’s brain structure. However, unlike researchers who constructed ROI templates for brain images from the magnetic resonance data set of a single healthy volunteer [24,25], we constructed the ROI template from the magnetic resonance data set of 10 healthy volunteers, including young adults and elderly individuals, and thus, our ROI template is unlikely to be markedly affected by the specificity of a single volunteer’s brain structure. Moreover, we also assessed the reliability of our ROI template by using data from Alzheimer’s disease patients and MCI patients who developed the disease within the following 2 years. We quantitatively and visually confirmed that the characteristic perfusion changes of the disease were detectable with our ROI template. We

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Applications of an ROI template for 3-D SSP images Kubota et al. 43

Fig. 5

(a)

Right lateral

Left lateral

Left medial

Right medial

Z 1

(b)

Right lateral

2

3

4 Left medial

Left lateral

5 Right medial

Z 1

2

3

4

5

(a) Z-score images of a patient with mild cognitive impairment (a 69-year-old male; MMSE score, 28). (b) The same Z-score images with our ROI template. By superimposing our ROI template on Z-score images, we can judge with a great certainty that the areas with the highest Z-scores exist within the posterior cingulate gyri.

confirmed that the hypoperfusion typical of Alzheimer’s disease was also detectable with our ROI template in MCI cases, suggesting that using the 3-D SSP technique and our ROI template enables reliable estimation of very slight perfusion changes in degenerative disorders. We believe that this method will be helpful in assessing various other neurological diseases in their early stages.

Conclusion Our ROI template for Z-score images of 3-D SSP enables automated quantitative evaluation of brain-function images. In addition, the ROI template may facilitate visual interpretation of 3-D SSP images of individual patients with neurological disorders.

Acknowledgements The authors thank Dr Satoshi Minoshima, Department of Radiology and Bioengineering, University of Washington, for his valuable suggestions. We also thank Kiyotaka Watanabe, Nihon Mediphysics Co. Ltd (Tokyo, Japan), for his technical support.

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Original article

Simultaneous surface registration of ictal and interictal SPECT and magnetic resonance images for epilepsy studies Erkan U¨. Mumcuog˘lua, Fatih Nara, Yasemin Yardımcıa, Umut Koc¸aka, ¨ mer Ug˘urb and Belkıs Erbas¸ b Eser Lay Ergu¨nb, Bilge Volkan Salancıb, O Background Subtraction of ictal and interictal single photon emission computed tomography (SPECT) images is known to be successful in localizing the seizure focus in the pre-surgical evaluation of patients with partial epilepsy. A computer-aided methods for producing subtraction ictal SPECT co-registered to the magnetic resonance image (MRI) (the SISCOM method) is commonly used. The two registrations involved in SISCOM are (1) between the ictal–interictal SPECT images, which was shown to be the more critical, and (2) between the ictal image and MRI. Objective To improve the accuracy of ictal–interictal registration in SISCOM by registering all three images (ictal, interictal SPECT, MRI) simultaneously. Methods The registration problem is formulated as the minimization of a cost function between three surfaces. Then, to achieve a global minimum of this cost function, the Powell algorithm with randomly distributed initial configurations is used. This technique is tested by a realistic simulation study, a phantom study and a patient study. Results The results of the simulation study demonstrate that, in surface-based registration, the triple-registration method results in a smaller ictal–interictal SPECT registration error than the pair-wise registration method (P < 0.05) for a range of values of the cost-function parameter. However, the improved registration error is still larger than that obtained by the normalized mutual

Introduction The SISCOM method has been shown to improve the sensitivity and specificity of single photon emission computed tomography (SPECT) in identifying the seizure focus in the pre-surgical evaluation of patients with partial epilepsy [1,2]. It has advantages over side-byside visual interpretation and manual registration approaches. Recent studies suggest that the sensitivity and specificity of SISCOM may surpass those of magnetic resonance imaging (MRI), positron emission tomography (PET), scalp-recorded electroencephalogram (EEG), interictal SPECT, and visual analysis of ictal SPECT [1,2]. Computer-aided methods for SISCOM are com-

information method (P < 0.001), which is a voxel-based registration algorithm. The phantom and patient studies reveal no observable difference between registration results. Conclusions Although the improved accuracy of triple registration is slightly worse than voxel-based registration, it will soon be possible to apply the results of this study in research utilizing the triple-registration principle to improving voxel-based results of ictal–interictal registrac 2006 Lippincott tion. Nucl Med Commun 27:45–55
Williams & Wilkins. Nuclear Medicine Communications 2006, 27:45–55 Keywords: SISCOM, registration, SPECT, MRI a Informatics Institute, Middle East Technical University and bDepartment of Nuclear Medicine, Hacettepe University, Faculty of Medicine, Ankara, Turkey.

Correspondence to Dr Erkan U¨. Mumcuog˘lu, Informatics Institute, Middle East Technical University, Ino¨nu¨ Bulvari, 06531, Ankara, Turkey. Tel: + 0090 312 210 3753; fax: + 0090 312 210 3745; e-mail: [email protected]

Sponsorship: Grant support for this study was supplied by the Middle East Technical University Grant BAP-2002-07-04-03, ‘3D Brain Image Analysis’; and the Turkish Government Planning Agency, DPT-YUUP Grant, ‘Visual Archival System for E-Government’.

Received 31 March 2005 Revised 18 July 2005 Accepted 21 September 2005

monly used and the SISCOM algorithm consists of the following steps [3]: 1. Image masking. By thresholding and morphological image processing, masks of cerebral volumes of ictal and interictal SPECT images are obtained. 2. Ictal and interictal image registration. Brain surface contours from mask images are used in surface-based registering. A voxel-based algorithm can be used as well. 3. Normalization. Mean brain pixel intensities of the ictal and interictal SPECT brain images are normalized to a constant value (to account for the differences in tracer uptake and decay).

c 2006 Lippincott Williams & Wilkins 0143-3636


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4. Subtraction. The normalized interictal image is subtracted from the normalized ictal image. Two standard deviations (SDs) and above are chosen only as significant activation pixels. 5. MRI co-registration. The extra-cerebral pixels on MRI are removed and the ictal image is co-registered to the extracted MRI brain by using surface matching. A fused display of colour-mapped SPECT activation areas are shown on the extracted brain MRI.

Of the two registrations involved in SISCOM, the registration error of ictal–interictal image registration is the more crucial, and is one of the major contributors to noise in the SISCOM method [4]. Registration errors (even sub-voxel) at this stage produce false positive activation areas and obscured true positive activation areas [4]. Therefore, the recommended approach is to register the two SPECT images rather than registering each to MRI [3]. The reported median surface registration error is 2.6 mm for ictal–interictal SPECT registration [1] and 4.4 mm for SPECT–MRI registration [5]. The reported best voxel-based registration error is between 0.5 and 2.0 mm for ictal–interictal SPECT registration [4], surpassing the surface-based techniques.

Fig. 1

(a)

Ictal SPECT (y)

Interictal SPECT (x) T

(b)

MRI (x)

T1

T2

Surface-based registration

In SISCOM, brain surface registration techniques [6,7] have traditionally been used to produce subtraction SPECT images [8,9]. Thresholding and morphological operations are generally used to extract the threedimensional brain surface. Surface registration consistently matches SPECT images with an accuracy better than 1 voxel [8]. An interpolated closest point transform which has been introduced recently for medical images claims that it can lead to a registration accuracy comparable to that of voxel-based methods in computed tomography (CT)–MRI registration [10]. An iterative closest point algorithm [11] is widely used in arbitrary surface matching problems. Jiang et al. proposed [7] a multi-resolution gradient descent approach to minimize the chamfer distance between two surfaces. Audette et al. have presented a recent survey paper [12] that overviews surface registration methods. Surface registration is achieved as the minimization of a squared distance function below with respect to a rigid transformation function T (it has three rotation and three translation parameters) between two SPECT images: 1 X D2pair ðT ðY Þ; XÞ ¼ ð1Þ k T ð yi Þ  x i k 2 i Ny in which, (yi) for i = 1,y, Ny is a set of points on the ictal surface Y, and xi = C(T(yi), X) is a point on the interictal surface X closest to T(yi), as shown in Fig. 1(a). (Note that T can be defined in any order, by changing ‘to’ and ‘from’ image volumes).

Ictal SPECT (y)

Interictal SPECT (z)

(a) A rigid transformation (T) defined betweeen ictal and interictal surfaces. (b) Two rigid transformations (T1 and T2) defined between ictal, interictal SPECT and MRI surfaces.

Voxel-based registration

Several voxel-based registration algorithms have been developed that were shown to provide increased registration accuracy in many cases [13–15]. A survey of other algorithms for co-registering SPECTor PET images is listed by Brinkmann et al. [3]. A few studies that have specifically addressed ictal–interictal SPECT registration accuracy [4] suggest that voxel-based registration is more accurate than surface matching, and the AIR algorithm [16] may be more robust to ictal–interictal blood flow changes than (normalized) mutual-information-based algorithms [17–20]. Using additional information in registration

To improve the registration accuracy of SPECT registration, use of additional information such as a simultaneously acquired transmission data [21,22], injection of a second radionuclide [23,24] or use of scatter window data [25] have been proposed. Pluim et al. [26] made a comprehensive survey of various mutual-informationbased registration techniques of more than two images,

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Simultaneous registration of SPECT images in epilepsy Mumcuog˘lu et al. 47

where multiple images are simultaneously registered. Among the papers surveyed by Pluim et al. [26] are those by Studholme et al. [27] and Andersson and Thurfjell [28]. Studholme et al. [27] incorporate segmented MRI set as third image in mutual information (voxel-based) registration of pelvis PET and MRI sets. The results demonstrate that additional information increases the overall information and produces distinct optimum close to true optimum. Andersson and Thurfjell [28] introduce a multivariate cost function based on information theory to register sets of emission tomography images obtained with different tracers (image volumes within each set are correctly aligned, but the transformation that maps one set to the other is sought). Another example of a multiple-image registration study is by Acton et al. [29], where a dynamic sequence of SPECT study frames are registered to each other (due to patient motion and patient repositioning) by using principal component analysis. It has been demonstrated by simulation and patient studies that simultaneous registration of image frames in a dynamic image study results in better performance than successive pair-wise registrations. Without acquiring additional data (such as transmission, second radionuclide or scatter window data), the MRI set is already available as additional information in SISCOM. The study presented investigates whether using MRI as additional information can improve the accuracy of ictal– interictal SPECT registration accuracy. The problem is formulated as simultaneous three-image-registration (ictal, interictal SPECT and MRI). Our initial investigation results have already been presented [30]. This article explains our methodology, and presents the results of simulation, phantom and patient studies.

Methods Proposed surface registration method

In our investigations for simultaneous three-image registration, a more straightforward surface-based approach is taken [30], because (1) formulation of the simultaneous voxel-based three-image registration (SPECT–SPECT– MRI) is not trivial; and (2) computational complexity would hinder a voxel-based investigation (see Discussion). However, the approach taken in this section can be extended to a voxel-based cost function. The registration problem is defined as simultaneous registration of ictal, interictal SPECT and MRI brain image surfaces. It is achieved as the minimization of the squared distance function below with respect to two rigid transformation functions T1 and T2 (each has three rotation and three translation parameters; a total of 12

parameters) between two SPECT and one MRI image sets, as shown in Fig. 1(b): D2triple ðT1 ðY Þ; T2 ðZÞ; XÞ ¼

1 X kT1 ðyi1 Þ  xi1 k2 Ny i1 1 X þ kT2 ðzi2 Þ  xi2 k2 Nz i2 1 X þa kT1 ðyi1 Þ  zi1 k2 Ny i1 ð2Þ

in which, (yi1) for i1 = 1,y, Ny is a set of points on the ictal brain surface Y; xi1 = C(T1(yi1), X) is a point on the MRI brain surface X closest to T1(yi1); (zi2) for i2 = 1,y, Nz is a set of points on the interictal brain surface Z; xi2 = C(T2(zi2), X) is a point on the MRI surface X closest to T2(zi2); and zi1 = C(T1(yi1), T2(Z)) is a point on the transformed interictal surface Z closest to T1(yi1), i.e., transformed ictal surface Y. In our implementation, zi1 = C(T2– 1T1(yi1), Z), whose result is identical to the previous formula. Note that in simultaneous three-image registration, there are only two degrees of freedom (i.e., two transformations, T1 and T2) to be determined. We use MRI as ‘to’ image volume in registration for both T1 and T2. Either transformation can be replaced by a transformation from ictal to interictal volumes (let us call it T3), but this will not affect the methodology because this transformation is a function of others: T3 = T2– 1T1. The first term in Equation 2 is the average squared distance between the transformed (T1) ictal surface and the MRI surface. The second term in Equation 2 is the average squared distance between the transformed (T2) interictal surface and the MRI surface. The third term in Equation 2 is the average squared distance between transformed (T1) ictal and transformed (T2) interictal surfaces. a is a constant to adjust the proportional weight of the squared distance between SPECT–SPECT surfaces versus that between SPECT–MRI surfaces. Such a term is considered appropriate in order to accommodate intramodality and inter-modality distance terms together in the same cost function (Equation 2). The optimum value of a is determined by realistic simulations: the value that results in the minimum registration error between ictal– interictal SPECT images is chosen (see the section ‘Results of the simulation study’). The cost function (Equation 2) is written as a function of surface points in MRI and SPECT image sets, and hence, is not free of error due to imaging data generation, reconstruction and surface extraction processes. Therefore, in comparing the registration accuracy of the three-image registration algorithm with that of others, the true registration error has to be used. (We used an average deviation of six points on the

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brain surface of the simulated image, as explained in detail at the end of the section ‘Simulations’.) Therefore, a is optimized according to this metric by realistic simulations. In order to evaluate the cost function in Equation 2, we need to compute the distance of a surface point in one data set to the other surface. To do this, the simplest approach is to determine the closest point among the points of the surface. In our implementation, interpolation [10] is not used, since the distance function is observed to be very smooth with respect to translation and rotation parameters (see Fig. 2). Searching for the closest point is computationally very expensive. The K-d tree technique is used [31,32] in our implementation for fast computation. An alternative technique would be to use chamfer distance-based fast computation [7]. To determine the optimum transformation parameters, the cost function (Equation 2) has to be minimized with Fig. 2

(a)

Cost function value

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7

−7 − 6 − 5 − 4 − 3 − 2 −1 0 1 2 3 4 5 Rotation around x-axis (degree)

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8

Our investigations have revealed that the cost function (Equation 2) has many local minima around the global minimum but that it behaves as a quadratic away from the global minimum. Therefore, after the first Powell run, it converges to a local minimum within very close proximity of the global minimum. Figure 2 shows the cost function (Equation 2) plotted as a function of ictal SPECT rotation. Note the smoothness of the cost function and the local and global minima on different scales. Figure 2(a) shows the quadratic nature of the cost function away from the global optimum, while Fig. 2(b) shows the local minima around the global optimum. Because of the above behaviour, we decided to use the randomly distributed initial configurations technique [17]. We initialized the Powell algorithm with 16 uniformly distributed random point sets (within ± 2 mm and ± 2 degrees of the first solution to the Powell run), and then chose the solution that achieves the minimum cost function. A genetic algorithm [34], simulated annealing [33] and a multi-resolution approach [17,35] are some of the other possible techniques to achieve a global minimum. The Powell algorithm for three-image registration takes approx. 2 min to run on a PC with a Pentium 4, 3.0 GHz processor. Since we used 16 random initializations after the first run (hence 1 + 16 runs), the total run time is (1 + 16)  2 min = 34 min for triple registration. The memory requirement of three-image registration is not significantly more than that of two-image registration algorithm. Simulations

(b)

Cost function value

respect to 12 parameters. We chose the Powell algorithm [33] to perform this optimization. The parameter order used is (t1x, t1y, t1z, r1x, r1y, r1z, t2x, t2y, t2z, r2x, r2y, r2z). The fractional tolerance parameter in Powell is set to 10 – 4.

Validation of this study is difficult due to lack of appropriate (ictal, interictal SPECT and MRI) data sets with external markers. Only SPECT–MRI data exist [15] with external markers, and we were not able to perform patient studies with external markers. For these reasons, a realistic simulation study was conducted to compare the accuracy of ictal–interictal registration in, firstly, the simultaneous ictal–interictal-MRI registration method (called ‘triple registration’) and, secondly, ictal–interictal registration (called ‘pair-wise registration’).

8

7

−1

0 Rotation around y-axis (degree)

The cost function (Equation 2) plotted as a function of ictal SPECT rotation (degrees) around the x-axis. (a) The quadratic nature of the cost function away from the global optimum, and (b) the local minima around the global optimum.

The Monte Carlo code developed by Singh and Mumcuog˘lu [36] was adapted for brain SPECT geometry. Photoelectric attenuation within the head and detectors are modelled. The system geometry was as follows: parallel hole collimator with aperture size 1.8 mm, aperture length 4.0 cm, septal thickness 0.3 mm. The detector was a 60  60 array with 0.35  0.35  0.35 cm

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Simultaneous registration of SPECT images in epilepsy Mumcuog˘lu et al. 49

elements (a detector total imaging area of 21  21 cm). The detector and collimator rotate around a diameter of 30 cm. A high resolution patient brain MRI volume (T1) is used (Fig. 3(a)) with 170  218  149 voxels of size 1  1  1 mm. Cerebral and extra-cerebral regions are first segmented using the ‘Brain Extraction Tool’ [37]. The cerebral region is next segmented using probabilistic SPM2 (2003) [38] software. Voting is then applied to identify non-overlapping grey matter, white matter, cerebrospinal fluid (CSF) and extra-cerebral regions (ECR) where the class having maximum probability by SPM segmentation is chosen (Fig. 3b). The image intensities in each region are modified so that radioactivity ratios of 24 : 10 : 1 : 1 are assigned to grey matter, white matter, CSF and ECR, respectively, to form an interictal SPECT template, as shown in Fig. 3(c). (Simulated radioactivity ratios are based on the study published by Yokoi et al. [39].) The ECR activity level is chosen to be high to simulate increased scalp activity in interictal SPECT studies. This interictal template is in perfect registration with MRI. In clinical practice, interictal acquisition is performed 15–30 min after radiopharmaceutical administration. This results in relatively high scalp activity. In order to create the ictal template, the interictal template is modified to represent contrast changes

between ictal–interictal images. Firstly, radioactivity ratios of 24 : 10 : 1 : 0.25 are assigned to grey matter, white matter, CSF and ECR, respectively. (In clinical practice, for ictal studies, patient stabilization takes some time and the patients’ acquisitions are performed with a delay of at least 30–90 min. Therefore, significantly diminished scalp activity is noted for ictal studies.) Secondly, volumetric Gaussian activity spots of varied size (SD = 10–20 mm) and amplitude (25–75% of the grey matter activity level) are added to form ictal SPECT templates (Fig. 3(d)). Activation spots are placed in cortical regions (to have an effect on the brain surface): right middle temporal gyrus, superior occipital gyrus, bilateral precuneus, and right superior frontal gyrus. Next, the Monte Carlo simulator is run using the ictal and interictal templates to generate projection data from a total of 120 angular views around 3601 to generate five pairs of ictal–interictal sinograms. The count rate is adjusted to obtain approximately 4.6 million counts for ictal and 7.0 million counts for interictal sinogram data. In our nuclear medicine department, 740–925 MBq 99mTcECD is prepared for the ictal study. For most patients, it is noted that seizures occur after prolonged periods without sleep. Therefore the patients are not allowed to sleep at night and have seizures in a predicted time interval (1–2 h). Nevertheless, the activity given to the patients for ictal studies is usually lower than that in interictal studies and it is difficult to establish the same

Fig. 3

A slice from the volume of (a) MRI, (b) segmented MRI into grey matter, white matter, CSF and extra-cerebral regions, (c) interictal template, (d) ictal template by contrast altering interictal template, (e) filtered back-projection (FBP) reconstructed interictal image, and (f) FBP reconstructed ictal image.

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amount of radioactivity. This fact is taken into account when the simulation parameters were established and total counts were lower in ictal study simulations when compared with interictal simulations. The sinogram data of opposing angles are geometrically summed and two-dimensional smoothing is applied by convolving with a 9  9 Gaussian filter of SD = 0.6  detector size. Then, planes of images are reconstructed by using a filtered back-projection (FBP) algorithm at a voxel size of 3.5 mm, as shown in Fig. 3(e and f) (the volume size is 100  100  60 in x, y and z, respectively). Voxel values are stored as 16-bit short integer maximum set to 30 000. Reconstructed image resolution is 0.9 cm in the centre of the field of view, which is about the same resolution as our clinical patient images. An attenuation correction is implemented in a post-processing step using a correction matrix based on Chang’s method (m = 0.12 cm – 1) [40]. Next, 15 sets of Gaussian distributed random translation and rotation parameters are generated (SD = 101 for rotation and SD = 10 mm for translation parameters). Using these parameters, five pairs of reconstructed ictal and interictal images are translated and rotated to obtain 15 image sets (Table 1) in random orientation for the registration algorithm in order to represent clinically unknown translation and rotation situations. Relative transformations between ictal and interictal SPECT is within ± 35.85 mm and ± 33.641. The MRI volume is kept at a fixed (initial) position for all data sets. Then, reconstructed SPECT images are thresholded (at an appropriate fixed threshold for all sets) to segment brain and background regions. Since the voxels in the CSF region can be lower than the threshold value, a fill algorithm is used to segment all the voxels within the brain. Next, simple 4-nearest-neighbour pixel checking logic is used to obtain the brain boundary voxels for each image set. There are approximately 50 000 brain boundary

voxels in MRI and 3750 in SPECT images to represent the brain surfaces. Next, pair-wise and triple registrations are performed by running the Powell algorithm with randomly distributed initial configurations (to reach a global minimum) on the cost function to compute registration parameters, using the calculated MRI, ictal and interictal SPECT boundaries. Finally, the registration accuracy of each method is measured by computing the average Euclidean distance of six predetermined points on the brain surface between ictal–interictal SPECT images (after estimated registration transformations are applied to misregistered images). Six points are on brain surface in orthogonal directions away from the centre of mass of the brain (Fig. 4). We call this average distance of six points between ictal–interictal images recovered by the registration method the registration error throughout the paper. Statistical analysis of simulation data

For the statistical analysis of the simulation study results, the normal distribution assumption is found to be valid, and each method has a homogenous distribution. Therefore, the paired t-test can be used in the analysis. (No correction is made for multiple comparisons). The level of statistical significance is set at P < 0.05 for all tests. The statistical analysis of the simulation study results was performed by using SPSS statistical software. Brain phantom study

The interictal phantom study was performed using a Hoffman brain phantom filled with 148 MBq [99mTc]pertechnetate. SPECT acquisition was performed using a double head gamma camera (Siemens ECAM, USA) and the parameters were set as follows: matrix: 128 128, 128 frame, with count rate of 60 kcounts/s. For the ictal study, a fish oil capsule that contained 0.9 MBq [99mTc]pertechnetate was placed in the third

Table 1 Misregistration translation (mm) and rotation (degrees) parameters (in the x, y, z axes) of interictal and ictal images with respect to the MRI set Image set

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Interictal image transformation parameters

Ictal image transformation parameter

tx (mm)

ty (mm)

tz (mm)

rx (deg)

ry (deg)

rz (deg)

tx (mm)

ty (mm)

tz (mm)

rx (deg)

ry (deg)

rz (deg)

18.27 6.54 – 15.45 – 3.75 2.08 – 7.66 – 1.06 3.39 10.34 – 14.05 – 10.31 – 6.43 1.71 13.45 19.36

7.41 8.12 – 1.43 – 1.00 – 8.00 4.93 12.38 12.96 – 2.78 2.17 6.31 – 5.49 2.30 3.55 5.21

– 6.16 13.46 9.75 – 23.78 – 10.92 – 3.26 – 20.12 15.68 2.33 6.46 – 11.29 1.97 16.97 7.26 7.93

– 6.73 – 9.02 – 1.55 9.47 15.50 4.29 – 5.61 1.79 – 7.72 – 9.43 – 14.08 – 19.06 – 0.65 6.72 2.06

– 0.08 0.20 – 5.58 18.86 – 2.20 – 14.14 – 3.03 – 5.70 – 1.21 – 3.90 – 8.44 – 17.38 – 4.50 – 15.48 – 0.96

9.08 23.70 5.20 4.11 10.53 4.29 12.95 – 1.86 1.31 – 6.58 – 7.59 – 5.95 8.12 0.70 – 18.34

– 4.11 5.11 – 11.99 – 0.96 4.46 – 2.96 – 1.68 1.80 4.21 16.78 19.97 6.97 – 13.66 3.63 – 5.67

– 10.44 6.97 4.84 – 1.94 – 3.78 – 8.86 – 18.40 – 16.28 – 11.74 – 4.15 1.75 2.29 – 12.41 7.00 4.27

14.55 – 5.10 – 0.07 – 5.26 7.18 10.88 5.01 27.72 – 1.60 4.29 – 19.67 – 5.46 – 18.88 – 1.08 – 13.16

1.98 14.69 3.66 – 4.43 – 0.49 0.78 19.58 – 0.73 9.39 – 0.80 – 8.01 3.09 10.52 – 16.64 – 10.91

– 1.92 4.63 – 9.24 – 6.50 6.23 – 13.35 10.48 8.63 – 6.42 6.60 12.94 3.15 8.60 1.29 0.17

– 0.73 – 9.94 – 7.47 – 0.31 9.88 – 5.99 14.77 – 8.14 6.45 – 13.10 – 8.67 – 4.74 2.22 18.71 1.10

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Simultaneous registration of SPECT images in epilepsy Mumcuog˘lu et al. 51

Fig. 4

performed and slices were set parallel to the orbito-meatal line. Attenuation correction was performed using the Chang algorithm (m = 0.12 cm – 1). Interictal SPECT study

Interictal SPECT was performed after a seizure-free day within the following week. Each patient received 740 MBq of 99mTc-ECD by i.v. injection in a dimly lit, silent room. Acquisition was performed after 15–30 min of radiopharmaceutical administration. The same acquisition and reconstruction parameters were used as in the ictal study. MRI study

A high resolution MRI volume was acquired using a 3 T system (Siemens) (T1), the voxel size was 1.0  1.0  1.0 mm, and the duration of data acquisition was 9 min.

Three rendered views of the brain and six points on the brain surface (in orthogonal directions away from the centre of mass of the brain) which are used to measure the registration accuracy.

ventricule adjacent to the posterior wall of the Hoffman brain phantom. The ictal SPECT acquisition was performed as described in the interictal study. The brain phantom containing the fish oil capsule was imaged by using 3 T MRI system (Siemens) (T1), the voxel size was 1.0  1.0  1.0 mm, and the duration of data acquisition was 9 min. Patient study Ictal SPECT study

Tc ethylenecysteinate dimer (99mTc-ECD), (Neurolite; Bristol Myers Squibb, Medical Imaging) was labelled according to the instructions on the package insert. Each patient was followed in an EEG monitoring room for 1 day. An intravenous line was set for each patient and 740– 925 MBq radioactivity was passed through the line. As soon as a clinical seizure started and had been confirmed by the EEG, the radioactivity prepared was injected by a trained nurse. After the radiopharmaceutical administration and the patient stabilized (in a time range of 30–120 min), the patient was brought to the nuclear medicine department for imaging. SPECT acquisition was performed using a dual-headed gamma camera (Siemens, ECAM, USA) equipped with high resolution, parallel hole collimator. During a 3601 rotation in 128  128 matrices, 128 frames (20 s/frame) were obtained. Filtered back-reconstruction with a Butterworth filter (cut-off 0.4, order 7) was

Since the true registration parameters of phantom and patient images could not be known, an absolute measure of registration accuracy could not be obtained. Brinkmann et al. [4] proposed the SD of non-zero (cerebral) pixels in their resultant subtracted images as a surrogate measure of accuracy. This was based on a correlation between the SD and registration error observed. Before incorporating this approach, we tried to validate this correlation by creating a plot of SD versus registration error using different registration method results on our simulation data. Unfortunately, we did not observe this correlation. Therefore, phantom and patient image registration tests were done by the visual comparison method.

Results Results of the simulation study

The results of simulation data registration are shown in Tables 2 and 3 for 15 image sets as the registration error between ictal–interictal SPECT images using different methods.

99m

Table 2 displays the registration error of the pair-wise registration method and the triple-registration method (for nine values of a). Also shown are the average and SD of error computed among 15 image sets for each method. The triple-registration method gives a smaller (P < 0.05) registration error than the pair-wise registration method only for a values 0.3, 0.5 and 0.7. However, when a = 0.0 or Z 1.0 a statistically significant reduction in error is not obtained. If the cost-function (Equation 2) is examined carefully, the result of the optimization for a = 0.0 corresponds to individual SPECT–MRI registrations; on the other hand, for high values of a, the solution converges to pair-wise (ictal–interictal) registration. Therefore, both limiting cases of a are not expected to be the optimum working

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52 Nuclear Medicine Communications 2006, Vol 27 No 1

Table 2 Ictal–interictal registration error (mm) of 15 image sets using surface-based pair-wise and triple-registration methods. The last row displays the average and standard-deviation (SD) of the error for each method Surface-based triple-registration: values of a

Image set

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Average (SD)

0.0

0.3

0.5

0.7

1.0

1.2

1.5

2.0

2.5

0.9073 1.8959 0.7287 1.5513 0.9623 0.6536 1.4430 1.0818 1.1212 0.9611 1.2002 1.7258 1.3942 2.0151 1.2270 1.26 (0.41)

0.7936 1.7508 0.8334 1.6350 0.7305 1.2757 1.4282 1.5487 0.6995 1.1129 1.1468 1.5812 1.3282 1.9521 1.2930 1.27 (0.39)

0.7692 1.7214 0.8485 1.6885 0.7505 1.3336 1.5065 1.5985 0.6796 1.1865 1.1729 1.5629 1.3162 1.8839 1.3152 1.29 (0.39)

0.7338 1.7736 0.8553 1.7024 0.7770 1.3902 1.4644 1.6496 0.7030 1.2219 1.1838 1.5479 1.3098 1.8650 1.3158 1.30 (0.39)

0.7158 1.7981 0.9014 1.7325 0.7894 1.4202 1.5519 1.6650 0.7254 1.2562 1.1937 1.5583 1.3382 1.8624 1.3369 1.32 (0.39)

0.7137 1.8500 0.9538 1.7499 0.8001 1.4336 1.5737 1.6745 0.7713 1.2992 1.2095 1.5406 1.3027 1.8850 1.3198 1.34 (0.39)

0.7139 1.8592 0.9665 1.8134 0.8148 1.4861 1.5776 1.6833 0.7885 1.3044 1.2035 1.5314 1.3081 1.8727 1.3439 1.35 (0.39)

0.7106 1.8555 1.1419 1.8404 0.8298 1.5243 1.6052 1.6730 0.8802 1.3204 1.2143 1.5385 1.3508 1.8538 1.2653 1.37 (0.37)

0.7056 1.8606 1.1604 1.8542 0.8223 1.5304 1.6084 1.7079 0.8612 1.3348 1.2324 1.5430 1.3357 1.8364 1.2807 1.38 (0.38)

Table 3 Ictal–interictal registration error (mm) of 15 image sets using the voxel-based pair-wise normalized mutual information (NMI) method. The last row displays the average and standard deviation (SD) of the error Image set 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Average (SD)

Surface-based pair-wise registration 0.6964 1.8811 1.6921 1.8860 0.8552 1.5842 1.6610 1.7208 0.8878 1.4162 1.2858 1.5435 1.2637 1.8826 1.1546 1.43 (0.39)

4. Analyze software (SISCOM: normalized mutual information registration). 5. Analyze software (SISCOM: surface-based registration).

Voxel-based (NMI) pair-wise registration 0.2599 0.3295 0.3814 0.4599 0.3744 0.5017 0.5552 0.5513 0.9375 0.2972 0.6746 0.5361 0.9793 0.5333 0.7552 0.54 (0.22)

point of triple registration, justifying the above statistical analysis results. The registration error of pair-wise ictal–interictal SPECT registration using the normalized mutual information (NMI) method is displayed in Table 3. By comparing this with Table 2, it can be seen that the NMI error is smaller than both the triple registration (all a) and pair-wise registration errors (P < 0.001).

Note that SISCOM (NMI registration; surface registration) is an integral part of Analyze software. All the views (transaxial/coronal/sagittal) of the registered images – MRI and ictal and interictal SPECT – are made available to physicians. Coloured activation images are overlayed on MRI (Fig. 5), and fused SPECT images (ictal, red; interictal, green) are overlayed on MRI. Phantom images were interpreted independently by four nuclear medicine physicians and the activation simulation area was identified at the posterior wall of the third ventricule in the ictal image set. Patients’ images were interpreted visually without the details of the clinical data being made available. Afterwards, image registration algorithms were performed and the resulting images were re-evaluated by the observers. Observer study results of phantom and patient data did not reveal any difference between the results of the five registration methods.

Discussion

In the evaluation of phantom and patient images, results for the following methods are compared:

We should point out that for the best performance of the method introduced in this paper, geometric and scale distortion in MRI should be corrected prior to application of the method [41]. In our phantom and patient studies, we were not able to do these corrections on MRI data, similar to many studies in the literature.

1. Our surface-based registration (pair-wise). 2. Our surface-based registration (triple, a = 0). 3. Our surface-based registration (triple, a = 1).

The purpose of our investigation was to explore whether incorporating the already available MRI into ictal–interictal SPECT registration can bring any benefit.

Results of the phantom and patient studies

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Simultaneous registration of SPECT images in epilepsy Mumcuog˘lu et al. 53

Fig. 5

deviation) is not obvious. Similarly, in the case of mutual information, the definition of the higher dimensional mutual information for three images is not well defined [26]. One definition [42] of higher dimensional mutual information for three images is I ðA; B; C Þ ¼H ð AÞ þ H ðBÞ þ H ðC Þ  H ðA; BÞ  H ðA; C Þ  H ðB; C Þ þ H ðA; B; C Þ where H denotes entropy. A property of this first definition is that it is not necessarily non-negative [26]. Another definition [27] of higher dimensional mutual information for three images is I ðA; B; C Þ ¼ H ð AÞ þ H ðBÞ þ H ðC Þ  H ðA; B; C Þ which can also be written as I ðA; B; C Þ ¼

X a;b;c

pða; b; c Þ log

pða; b; c Þ : pð aÞpðbÞpðc Þ

This second definition is not negative but it does not define the mutual information of three images as one would expect, namely as the information shared between all three images [26]. An alternative to a single three-image registration metric is to create an approximate cost function as a summation of three two-image co-registration metrics, similar to the surface-based metric we used (Equation 2). However, in this case, two voxel-based metrics (SPECT–SPECT and SPECT–MRI) would appear together, in a hybrid way, in the same cost function. (In the surface-based case, this is not a problem since only surface points are involved.) Combining a SPECT–SPECT and SPECT–MRI metric in the same equation would probably require more than adjustment of a coefficient, since SPECT–SPECT and SPECT–MRI voxel-based metrics can have significantly different characteristics (e.g., how fast they deviate from their optimum as registration parameters are changed). Activation images overlayed on brain MRI of a 52-year-old female patient with a resistant partial epilepsy. (a) Activation spot overlayed on MRI; and (b) activation spot only. In each panel, coronal (upper left), sagittal (upper right) and transaxial (lower left) slices through the activation area, as well as the rendered volume (lower right) are shown. The patient’s EEG revealed possible epileptiform activity of the parietal lobe. On visual examination of the brain perfusion images, cerebral hyperperfusion is identified in the posterior parietal lobe, which is supported by the SISCOM algorithm.

Although it is known that a voxel-based co-registration system is more accurate than the surface-based coregistration, there are two reasons why we chose to work on a surface-based framework. Firstly, in the case of AIR, the derivation of the threeimage registration metric (the normalized standard

Secondly, even if the above challenges of the voxel-based three-image registration method are dealt with, computational challenges exist, since voxel-based registration is known to be computationally heavier than surface-based registration (unless multi-resolution optimization and other acceleration methods are incorporated), and simultaneous three-image registration would require even more processing time. Hence, a very long investigation/ code debugging time would be necessary. For the above reasons, in order to show a proof of the principle, a straightforward and computationally feasible approach is taken. We hope that this can serve as a starting point for further exploration by other groups to investigate if a similar approach is beneficial.

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54

Nuclear Medicine Communications 2006, Vol 27 No 1

In the Analyze software package, voxel-based ictal– interictal registration, followed by surface-based ictal SPECT–MRI registration is recommended for best accuracy. Therefore, in triple-registration, a hybrid cost function can be formed in principle (the SPECT–SPECT error is in terms of the voxel-based measure, and the SPECT–MRI error is in terms of the boundary-based measure). Doing this in a robust way is, however, a subject of further investigation. The computation time of our current implementation of the triple-registration algorithm can be significantly reduced in the future by multi-resolution techniques.

6

7

8

9

10

11 12

A generalization of the technique introduced here to N data volumes (N > 3), can be considered. In such case, there will be N – 1 degrees of freedom (N – 1 independent transformations to be optimized simultaneously). The cost function will have as many error terms as all the possible combinations of image pairs. Hence, as N increases, the computation load will increase significantly, but for a few image volumes, simultaneous registration can be feasible and beneficial.

13 14

15

16

17

Conclusions The results of the simulation study have demonstrated that, in surface-based registration, the triple-registration method results in a smaller ictal–interictal SPECT registration error than the pair-wise registration method (P < 0.05) for a range of cost-function parameter values. However, the improved registration error is still higher than the NMI error (P < 0.001), which is a voxel-based registration algorithm. Nevertheless, the results of this study can be used in future research to apply the tripleregistration principle to improving the voxel-based registration results of ictal–interictal registration.

18

19 20

21

22

23

Acknowledgements The authors thank Serap Saygı, Ay ¸senur Cila, Dog˘an Bor and Osman Saka for their assistance, helpful discussions, and for providing the MRI set.

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Original article

QGS ejection fraction reproducibility in gated SPECT comparing pre-filtered and post-filtered reconstruction Janelle M. Wheat and Geoffrey M. Currie Introduction The aim of this investigation was to compare the QGS determined functional parameters using prefiltering to that using post-filtering in the gated myocardial perfusion single photon emission computed tomography (SPECT) reconstruction process. Methodology A total of 25 patient files were examined, each with both a gated rest and gated stress study, and were reconstructed using two strategies. The first employed pre-filtering with a Butterworth low pass filter (order 4.0 and cut-off 0.21) and the second employed postfiltering with a Butterworth low pass filter (order 5.0 and cut-off 0.21). Following reconstruction and reorientation, gated short axis slices were evaluated with QGS software.

cally significant difference between matched pairs (P < 0.0001) and a statistically significant difference was shown between the means (P = 0.005). Conclusion The impact of performing pre-filtering on data in the reconstruction process is significant with a 5.3% increase in the calculated ejection fraction over post-filtering. Clearly, this has the potential to undermine diagnostic and prognostic roles of functional parameters. Nucl Med Commun 27:57–59

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:57–59 Keywords: gated myocardial perfusion SPECT, pre-filter, post-filter, ejection fraction

Results The mean ejection fraction for the post-filtered data was 49.5% (95% CI, 45.8–53.1%) and for the prefiltered data was 54.8% (95% CI, 51.4–58.1%). Excellent correlation was demonstrated between the pre- and postfiltered ejection fractions with a correlation coefficient of 0.964. The mean difference between matched pairs of preand post-filtered ejection fraction data was 5.3% (95% CI, 4.3–6.3%). The match pair t-test demonstrated a statisti-

Introduction Gated myocardial perfusion single photon emission computed tomography (SPECT) was developed in the late 1980s with the initial role of differentiating artefact from pathology [1]. The high count density of 99mTc based myocardial perfusion studies allows SPECT images to be synchronized to the patient’s electrocardiogram (gated) because they maintain adequate count density in individual cardiac frames (intervals). This feature, in conjunction with acceptable spatial and contrast resolution, allows simultaneous assessment of myocardial perfusion and ventricular function [2]. Despite limitations associated with filtering gated SPECT data, it is universally recommended that default filter parameters are adhered to due to the danger of introducing false positive or false negative results following filter customization [3]. Over-filtering myocardial perfusion SPECT data is known to cause false negative results and under-filtering causes false positive results [3]. Since the quantitative gated SPECT (QGS) software (Cedars Sinai Medical Centre, Los Angeles, California) determines functional parameters utilizing

School of Clinical Sciences, Charles Sturt University, Wagga Wagga, Australia. Correspondence to Janelle Wheat, School of Clinical Sciences, Locked Bag 588, Charles Sturt University, Wagga Wagga 2678, Australia. Tel: + 0061 02 693 32750; fax: + 0061 02 693 32866; e-mail: [email protected] Received 31 March 2005 Accepted 2 September 2005

edge detection, filtering errors will also cause inaccuracies in these calculations. Kubo et al. [4] indicated that the ideal gated SPECT prefilter for ventricular volume determination was a Butterworth with a critical frequency of 0.54 cycle/cm but this filter caused an underestimation of left ventricular volumes by 17% for end diastolic and 8% for end systolic. Kubo et al. [4] conclude that pre-filters have little impact on the ejection fraction because of the simultaneous blur effect on end diastolic volume (EDV) and end systolic volume (ESV), effectively cancelling each other out. Fredericks et al. [5] found that QGS was robust to both variations in filter cut-off and orientation of the short axis. The authors of QGS recommend a Butterworth pre-filter of 0.3 cycle/pixel at a pixel size of 0.64 cm [6] which translates to a recommended critical frequency of 0.47 cycle/cm. This recommendation is discordant with the default filter prescribed by the QGS software manual [7] which requires a three-dimensional low pass postfilter with an order of 5.0 and a cut-off of 0.21. Not surprisingly then, 60.4% (32/53) of departments using QGS also employ a pre-filter [8]. This may be attributed

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in part to the requirement of pre-filtering in ungated quantitative software (e.g., CEqual), which was used widely prior to the widespread use of gating, and to the discordant recommendations in the literature. The aim of this investigation was to determine the impact on the QGS determined functional parameters by comparing pre-filtering to post-filtering in the gated myocardial perfusion SPECT reconstruction process.

Methodology This clinical investigation employed a retrospective repeat-measures design. This study design allowed assessment of the dependant variables (ejection fraction, EDV, ESV) by manipulating the independent variable (reconstruction strategy) for a single data set. Using this approach meant that a single clinical data set acted as both the control group (post-filtering) and the experimental group (pre-filtering). All patients in the study population followed one of two myocardial perfusion SPECT protocols: 2 day rest–stress or 2 day stress–rest. All myocardial perfusion SPECT studies employed a 740 MBq (20 mCi) dose of 99mTc tetrofosmin (Nycomed-Amersham, Amsterdam). A triple detector gantry was used to acquire all patient data. All data acquisitions employed low energy, high resolution collimation with step and shoot mode, elliptical orbits and a 64 matrix. The zoom was 1.23 and projections were acquired at 31 intervals for 20 s per projection to provide a total acquisition time of 15 min. All patients were positioned supine with their feet into the gantry for an eight interval gated SPECT acquisition. A total of 25 patient files were examined, each with both a gated rest and gated stress study and, thus, a total of 50 studies were produced for quantitative analysis with QGS.

independent means and proportions was calculated with a 95% confidence interval (CI). Correlation was evaluated with chi-squared analysis and reliability measured using Cohen’s kappa coefficient. Bland–Altman analysis [9] and the matched pairs t-test were used to assess agreement between paired data. Approval for this study was granted by the Charles Sturt University Ethics in Human Research Committee for the retrospective manipulation of the de-identified patient data.

Results All 25 clinical studies had both stress and rest data quantified with QGS software following reconstruction by both a pre-filtering algorithm and a post-filtering algorithm by two experienced observers. Inter-operator reproducibility was excellent with a correlation coefficient of 0.994 for post-filtered data and 0.979 for prefiltered data (R2 of 0.988 and 0.958, respectively). No statistically significant difference was noted between matched pairs comparing operators 1 and 2 for either the pre-filtered data (P = 1.0) or the post-filtered data (P = 0.06). Consequently, the following analysis was undertaken using the mean data of matched pairs between operators. The mean ejection fraction for the post-filtered data was 49.5% (95% CI, 45.8–53.1%) and for the pre-filtered data was 54.8% (95% CI, 51.4–58.1%). Excellent correlation was demonstrated between the pre- and post-filtered ejection fractions with a correlation coefficient of 0.964 and R2 of 0.929 (Fig. 1). While this supports a strong relationship between the pre- and post-filtered ejection

Fig. 1

80 70 60 Pre-filtered EF

The gated SPECT data for all clinical studies were reconstructed using two strategies. The first employed pre-filtering with a Butterworth low pass filter (order 4.0 and cut-off 0.21) followed by reconstruction and reorientation of gated short axis slices for QGS analysis. The second strategy employed post-filtering with a Butterworth low pass filter (order 5.0 and cut-off 0.21) during reconstruction followed by re-orientation of gated short axis slices for QGS analysis. All data was reconstructed using a 1801 filtered back-projection algorithm.

50 40 30 20 10

Pre-filter EF = 11.1 + 0.9 Post-filter EF

0

The statistical significance was calculated using the Student’s t-test for continuous data. A P value less than 0.05 was considered significant. Normality of distribution was determined using the Shapiro–Wilk W test with a P value less than 0.05 indicating that the data varies significantly from normality. The differences between

0

10

20

30 40 50 Post-filtered EF

60

70

80

Bivariate analysis of the ejection fraction (EF) calculated following pre-filtered reconstruction versus post-filtered reconstruction demonstrating excellent correlation (0.964).

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EF reproducibility between pre- and post-filtering in QGS Wheat and Currie 59

Discussion and conclusion

Fig. 2

It is worth noting that using an eight-bin gated acquisition underestimates the ejection fraction by 3.7% [10] requiring interpreting physicians to add 4% to the calculated ejection fraction [7]. An eight-bin gated SPECT acquisition processed using a pre-filter would not require this correction because it overestimates the ejection fraction by 5.3% compared to the prescribed post-filter. The uncorrected result, therefore, would leave the ejection fraction with a 1.6% overestimation of the ejection fraction. It is clear that accurate interpretation and reporting of functional parameters on gated myocardial perfusion SPECT using QGS requires either adherence to the prescribed acquisition and processing parameters or a thorough understanding of the implications of variations to these prescribed parameters.

15

Difference: Pre EF − Post EF

10

5 0

−5 −10 −15 10

20

30 40 50 60 Mean: (Pre EF + Post EF)/2

70

80

Bland–Altman analysis of the pre-filtered ejection fraction and the postfiltered ejection fraction demonstrating a mean difference of 5.3% (solid horizontal line).

fractions, it does not provide an indication of the strength of agreement between data. Consequently, Bland–Altman analysis was performed (Fig. 2) which indicated that the 95% limits of agreement included 96% of data points. The difference between matched pairs of pre-filtered and post-filtered ejection fractions demonstrated a normal distribution (P = 0.19). The mean difference between matched pairs of pre- and post-filtered ejection fraction data was 5.3% (95% CI, 4.3–6.3%) where a positive difference indicates that the pre-filtered ejection fraction is higher than that of the post-filtered. Despite the overlap of 95% CIs, the match pair t-test demonstrated a statistically significant difference between matched pairs (P < 0.0001) and a statistically significant difference was shown between the means (P = 0.005). The pre-filtered mean EDV was 106.6 ml and the postfiltered was 103.2 ml. Despite excellent correlation (correlation coefficient = 0.967), a statistically significant difference was noted between matched pairs (P < 0.0001). The mean difference between pre- and post-filtered EDV was – 3.38 ml indicating that the prefilter EDV was lower than the post-filtered. Similarly, the pre-filtered mean ESV was 57.7 ml and the post-filtered was 50.8. Despite a correlation coefficient of 0.974, a statistically significant difference was noted between matched pairs (P < 0.0001). The mean difference between pre- and post-filtered ESV was – 6.95 ml. The greater impact of filtering on ESV than EDV (absolute and relative) is responsible for the ejection fraction differences described above.

In general, pre-filtering in SPECT reconstruction with a ramp filter is thought to be superior to post-filtering. Armed with this knowledge and combined with variations in recommendations offered in the literature, discordance between clinical practice and the recommendations of the software distributor is not uncommon. Nonetheless, the default filtering requirements for QGS software includes post-filtering of reconstructed data. The impact of performing pre-filtering on data in the reconstruction process is significant with a 5.3% increase in the calculated ejection fraction over post-filtering. Clearly, this has the potential to undermine diagnostic and prognostic roles of functional parameters. Furthermore, precision and reliability of functional parameters are undermined, especially between centres employing different filtering strategies.

References 1

Go V, Bhatt M, Hendel R. The diagnostic and prognostic value of ECGgated SPECT myocardial perfusion imaging. J Nucl Med 2004; 45: 912–921. 2 Zaret B, Bellar G. Nuclear Cardiology. State of the Art and Future Directions, second edition. New York: Mosby; 1999. 3 DePuey E. Updated imaging guidelines for nuclear cardiology procedures. J Nucl Cardiol 2001; 8:G1–G58. 4 Kubo N, Mabuchi M, Katoh C, Morita K, Tsukamoto E, Morita Y, Tamaki N. Accuracy and reproducibility of left ventricular function from quantitative, gated, single photon emission computed tomography using dynamic myocardial phantoms: effect of pre-reconstruction filters. Nucl Med Commun 2002; 23:529–536. 5 Fredericks N, Baxter P, McKay E, Smart R. An assessment of the sensitivity of the Cedars-Sinai quantitative gated SPECT software to changes in the reconstruction of the short-axis slices. J Nucl Med Technol 1999; 27: 123–126. 6 Germano G, Berman D. Clinical Gated Cardiac SPECT. New York: Futura Publishing; 1999. 7 Picker International. Quantitative Gated SPECT (QGS) Operators Guide. Ohio: Picker International; 1997. 8 Wheat J, Currie G, Adams B. Myocardial perfusion SPECT in Australia: processing parameters. ANZ Nucl Med J 2005; 36:63–66. 9 Bland J, Altman D. Comparing two methods of clinical measurement: a personal history. Int J Epidemiol 1995; 24(Suppl 1):S7–S14. 10 Germano G, Kiat H, Kavanagh PB, Moriel M, Mazzanti M, Su HT, et al. Automatic quantification of ejection fraction from gated myocardial perfusion SPECT. J Nucl Med 1995; 36:2138–2147.

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Original article

Value of 99mTc-methoxyisobutylisonitrile (99mTc-MIBI) gated SPECT for the detection of silent myocardial ischemia in hemodialysis patients: clinical variables associated with abnormal test results Meltem Caglara, Babak Mahmoudiana, Kudret Aytemirb, Serkan Kahramanc, Mustafa Arıcıc, Giray Kabakcıb and Erdem Karabulutd Background Although coronary artery disease is a major cause of mortality and morbidity in patients undergoing hemodialysis, there is no accurate diagnostic strategy for these patients. Aim To assess the value of 99mTc-methoxyisobutylisonitrile (99mTc-MIBI) gated single-photon emission computed tomography for the detection of silent myocardial ischemia in patients undergoing hemodialysis and to evaluate the clinical variables associated with abnormal test results. Methods Thirty-one asymptomatic patients undergoing hemodialysis (20 men, 11 women), with a mean age of 45 years (range, 25–75 years), were included in the study. Serum electrolytes, creatinine, homocysteine and adhesion molecules were measured prior to dialysis. Ambulatory blood pressure, carotid intima–media thickness measurements, echocardiography and stress 99m Tc-MIBI imaging were performed in all patients, whereas coronary angiography was performed only in patients with abnormal myocardial perfusion scintigraphy and/or echocardiography. Results Gated myocardial perfusion scintigraphy results were abnormal in nine patients (29%) and coronary angiography was abnormal in six patients. After a median follow-up of 20 months (range, 14–28 months), nine patients experienced a cardiac event. Seven of the nine patients (78%) with abnormal myocardial perfusion scintigraphy suffered a cardiac event, compared with only

Introduction Cardiovascular disease (CVD) has been reported to account for more than one-half of all deaths in patients on hemodialysis [1–3]. The high prevalence of diabetes and hypertension contributes to the accelerated rate of CVD, in addition to uremia-related risk factors which play a role in the development of atherosclerosis. The mechanism of atherogenesis is complex and includes hypertension, smoking, hyperlipidemia, thrombogenesis and the production of vasoactive substances, growth factors, mediators of inflammation, soluble adhesion molecules, hyperhomocysteinemia and abnormal mineral metabolism [4–6].

two of 22 patients (9%) with normal myocardial perfusion scintigraphy (P < 0.0001). Patients with abnormal perfusion scintigraphy had higher serum C-reactive protein, homocysteine and adhesion molecule levels and the duration of hemodialysis was significantly longer. Conclusion In asymptomatic hemodialysis patients, gated myocardial perfusion scintigraphy is a safe and non-invasive screening technique for the detection of coronary artery disease and for predicting future cardiac events. The presence of ischemia correlates significantly with markers of inflammation. The discordant results (abnormal myocardial perfusion scintigraphy/normal coronary angiography) can be attributed to angiographically unrecognized occult atherosclerotic changes and abnormal vasodilatation capacity of the coronary circulation. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:61–69
Nuclear Medicine Communications 2006, 27:61–69 Keywords: coronary artery disease, hemodialysis, myocardial perfusion scintigraphy, silent ischemia, 99mTc-MIBI Departments of aNuclear Medicine, bCardiology, cInternal Medicine Division of Nephrology and dBiostatistics, Hacettepe University Medical Faculty, Ankara, Turkey. Correspondence to Professor Meltem Caglar MD, Department of Nuclear Medicine, Hacettepe University Medical Faculty, Sıhhıye, Ankara 06100, Turkey. Tel: 90 312 448 0810; fax: 90 312 309 3508; e-mail: [email protected] Received 14 July 2005 Accepted 5 October 2005

The diagnosis of coronary artery disease (CAD) by clinical symptoms is unreliable as a considerable number of patients are asymptomatic. Silent myocardial ischemia has been reported in 25% of chronic hemodialysis patients, defined as the presence of ST segment depression without associated chest pain [7]. The diagnostic value of exercise tests is limited, because the interpretation of electrocardiogram (ECG) findings is hampered in the presence of left ventricular hypertrophy, electrolyte disorders and medications. Furthermore, patients on hemodialysis frequently are unable to complete a maximal test due to anemia, debility, peripheral vascular disease or poor physical condition.

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62 Nuclear Medicine Communications 2006, Vol 27 No 1

Thus, CAD can be misdiagnosed when relying on clinical symptoms and ECG findings alone. According to current opinion, the demonstration of silent myocardial ischemia requires the detection of CAD, preferably by myocardial perfusion scintigraphy (MPS) or stress echocardiography before the patient is referred to angiography [8–11]. The use of pharmacological stress is an alternative to exercise, especially in patients with reduced exercise capacity. The purpose of this study was to assess the diagnostic accuracy of 99mTc-methoxyisobutylisonitrile (99mTcMIBI) scintigraphy for the detection of silent myocardial ischemia in patients undergoing hemodialysis and to investigate the clinical variables associated with abnormal test results.

Patients and methods Patient characteristics

Thirty-one patients with end-stage renal disease, continuously maintained on chronic hemodialysis treatment, were enrolled in the study on the basis of the following criteria: maintained on hemodialysis for more than 1 year; stable hemodynamic status; and absence of overt CAD, angina, myocardial infarction or congestive heart failure. Written informed consent was obtained from each patient and the study was approved by the local ethics committee (No: HEK 02/19-10). Twenty patients were male, and the mean age was 45 ± 11.8 years (range, 25–75 years). Regular hemodialysis was performed three times per week for 4 h per session. Body weight, blood pressure and heart rate were measured before hemodialysis, as were the serum urea nitrogen, creatinine, uric acid, glucose, total protein, albumin, electrolyte, total cholesterol, high and low density lipoprotein, homocysteine and hematocrit levels. The detailed features of the study group are described in Table 1. Serum levels of adhesion molecules were assayed in predialysis venous blood. Soluble adhesion molecules have been postulated to be risk predictors of cardiovascular events and their focal expression has been observed in atherosclerotic plaques. Serum concentrations of soluble E-selectin (sE-selectin), soluble P-selectin (sPselectin), soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1) were determined by enzyme-linked immunoabsorbent assay (ELISA) using standard kits (Bender MedSystems, Vienna, Austria). Serum concentrations of these soluble adhesion molecules were calculated by reference to standard curves obtained with the corresponding recombinant molecules. For all assays, the intraand inter-assay coefficients of variation were less than 5%

Table 1

Detailed patient characteristics

Male/female Age (years) Hemodialysis duration (months) Systolic/diastolic blood pressure (mmHg) Hematocrit (%) Cholesterol (normal, < 200 mg/dl) Blood urea nitrogen (normal, 4.6–23 mg/dl) Creatinine (normal, 0.6–1.2 mg/dl) Protein (normal, 6–8.7 g/dl) Albumin (normal, 3.2–4.8 g/dl) C-reactive protein (normal, 0.0–0.8 mg/dl) Homocysteine (normal, 5–15 mmol/l) sP-selectin (normal, 111–266 ng/ml) sE-selectin (normal, 23.0–79.2 ng/ml) sVCAM (normal, 675–1693 ng/ml) sICAM (normal, 129.9–297.4 ng/ml)

20/11 45.5 ± 11.7 77.7 ± 74.7 120 ± 24/78 ± 16 31 ± 5 157 ± 48 77 ± 24 11 ± 4 7 ± 0.7 4 ± 0.4 0.55 ± 0.37 23.4 ± 8.2 245 ± 113 129 ± 124 1053 ± 422 1065 ± 324

sE-selectin, soluble E-selectin; sICAM, soluble intercellular adhesion molecule; sP-selectin, soluble P-selectin; sVCAM, soluble vascular cell adhesion molecule.

and 10%, respectively. All results from ELISAs represent means from duplicated measurements and are expressed in nanograms per milliliter. Carotid intima–media thickness

This was measured using an ultrasound machine, generating a wide-band ultrasonic pulse with a middle frequency of 7.5 MHz (Toshiba SSA-270 A, Tokyo, Japan). A single observer, blind to the patients’ demographic data and cardiovascular risk, measured the combined thickness of the intima and media of the far wall of both common carotid arteries. Images were recorded from an anterolateral longitudinal view. The mean intima–media thickness was computed and right and left measurements were averaged for the statistical analysis. Blood pressure

Blood pressure profiles were obtained by ambulatory monitoring. Measurements were obtained every 15 min during the day and every 30 min during the night. Patients were asked to record their periods of activity and sleep. The following parameters were analyzed: mean 24-h systolic and diastolic blood pressure; mean systolic and diastolic blood pressure during the day; and mean systolic and diastolic blood pressure at night. Imaging protocol

This consisted of rest echocardiography and stress/rest gated MPS. Patients with abnormal MPS and/or echocardiography underwent cardiac angiography within 2 weeks. The interval between investigations and hemodialysis was 24 h in all patients in order to standardize the blood volume and preload, which affect the left ventricular ejection fraction (LVEF). Stress test and MPS

Myocardial perfusion single-photon emission computed tomography (SPECT) was performed with a 2-day stress/ rest protocol. All b-blockers and calcium channel blockers were discontinued 48 h before the study and nitrate

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Silent ischemia in hemodialysis patients Caglar et al. 63

each for 25 s, were acquired in a 64  64 matrix over a non-circular 1801 orbit, starting at the 451 right anterior oblique to the 451 left posterior oblique position. Rest images were acquired with the same protocol following 60 min after the injection of 740 MBq of 99mTc-MIBI.

compounds were withheld at least 24 h before the study. Patients underwent a graded treadmill test according to the Bruce protocol [12]. The standard 12-lead ECG and blood pressure were monitored during exercise. Patients with limited exercise capacity and those who failed to achieve at least 85% of the predicted heart rate underwent dobutamine–atropine stress test. None of the patients had severe hypertension, which precludes the administration of dobutamine. Dobutamine was infused through an antecubital vein starting at a dose of 10 mg/kg/ min for 3 min, increasing by 10 mg/kg/min every 3 min up to 40 mg/kg/min. Patients who were unable to achieve 85% of their predicted maximal heart rate, and without symptoms or signs of ischemia, were given atropine in addition to the maximal dose of dobutamine, starting with 0.5 mg intravenously and repeated up to a maximum of 2.0 mg. Test endpoints were the achievement of 85% of the maximum age-predicted heart rate, horizontal or downsloping ST segment depression of more than 2 mm, ST segment elevation of more than 1 mm in patients without previous myocardial infarction, severe angina, systolic blood pressure fall of more than 40 mmHg, blood pressure above 240/120 mmHg or significant arrhythmia. b-Blockers were available to reverse the side-effects of dobutamine and/or atropine. The exercise test was considered to be adequate if the patients reached 85% or more of their maximum predicted heart rate and/or when an ischemic endpoint (angina, ST segment depression) was reached.

After exclusion of the non-myocardial activity, slices were reoriented along short, vertical long and horizontal long axes. Data were filtered by backprojection using a Butterworth filter (order of 5; cut-off frequency, 0.5 cycles per pixel) and reconstructed. No scatter or attenuation correction was applied. Visual and semiquantitative interpretations were performed for the presence, location and severity of the defects by two independent observers who were unaware of the patients’ clinical data. The left ventricle was divided into 20 regions for each study (Fig. 1). The left anterior descending artery territory included the anterior and anterolateral walls, the septum and the apical wall (segments 1–3, 7–8, 13, 14, 19, 20). The inferior wall (segments 4, 9, 10, 15, 16) was assigned to the right coronary artery and the lateral wall (segments 5, 6, 11, 12, 17, 18) to the left circumflex artery (Fig. 1). Each ventricular segment was scored by the consensus of two observers using a five-point scoring system: 0, normal uptake; 1, slightly reduced uptake; 2, moderately reduced uptake; 3, severely reduced uptake; 4, absent uptake. A segment was considered to be abnormal if its score was two or more. Apparent perfusion defects, probably caused by soft tissue attenuation, were given a score of one. Scans that had perfusion defects on both stress and rest images were considered to have a fixed defect. When a perfusion defect on the stress image resolved completely or in part (score 0–1) on the rest images, a reversible defect was assigned.

Approximately 1 min before the termination of the stress test, an intravenous dose of 700–925 MBq of 99mTc-MIBI was injected intravenously, with dose variation based on the patient’s weight. Eight-frame per cardiac cycle gated SPECT imaging was initiated 15–45 min after the stress test using a double-headed rotating gamma camera (depending on the type of stress) (Siemens E-Cam, Erlangen, Germany) equipped with a low-energy, highresolution collimator. Energy discrimination was centered at 140 keV with a 20% window. Sixty-four projections,

A scintigraphic extension score, defined as the number of abnormal segments, was also obtained for each patient. The perfusion defect score was derived by the summation of the score of each myocardial segment at stress

Fig. 1

Short axis Apical

Vertical long axis

Mid

Basal 13

7 1 2

6 5

3 4

8

12

9

11 10

14

18

15

17

19 20

16

Assignment of myocardial regions used for regional wall motion and perfusion analysis (20-segment model).

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(SSS) and at rest (SRS). The difference was expressed as the summed difference score (SDS). SSS < 4 was considered to be normal, SSS = 4–8 mildly abnormal, SSS = 9–12 moderately abnormal and SSS Z 13 severely abnormal. The gated study was analyzed using the Quantitative Gated SPECT (QGS) program described by Germano et al. [13]. Echocardiography

Echocardiography was performed with a two-dimensional M-mode echo using a 2.5–3.5 MHz probe (System five, cardiac scanner, GE Vingmed Ultrasound, Horten, Norway). Images were acquired at rest in the parasternal long and short axes and apical four- and two-chamber views. Doppler echocardiography was performed to evaluate left ventricular regional wall motion, dimensions and ejection fraction. The left ventricular mass was calculated using the following ‘Devereux’ formula: LVMðgÞ ¼1:04½ðLVID þ VST þ PWTÞ3  ðLVIDÞ3   13:6 where LVM is the left ventricular mass, LVID is the left ventricle end-diastolic internal dimension, VST is the ventricular septal thickness and PWT is the posterior wall thickness. Left ventricular hypertrophy was defined when the mass was greater than 125 g/m2 in men and 110 g/m2 in women. Left ventricular wall motion was scored semiquantitatively as follows: 1, normal; 2, mild-to-moderate hypokinesia; 3, severe hypokinesia or akinesia; 4, dyskinesia; 5, aneurysm.

Angina pectoris was diagnosed by the presence of typical symptoms or reversible ischemic changes on ECG. Myocardial infarction was defined as prolonged chest pain accompanied by pathological Q waves or persistent ST segment modification, or high serum creatinine phosphokinase levels. Cardiac death was defined as a witnessed death that occurred within 1 h after the onset of symptoms, with no history of violence or accident having a role in the fatal outcome. Myocardial revascularization was performed after identification of severe coronary artery lesions on coronary angiography. Statistical analysis

This was performed using the SPSS statistical software package (SPSS 11.0 for Windows, SPSS Inc., Chicago, Illinois, USA). Comparison between groups was performed by Student’s t-test. Data are given as the mean ± standard deviation (SD). A P value of 0.05 or less was considered to be significant. The association between the risk factors and the presence of CAD was assessed using Spearman’s correlation coefficient (r). Stepwise logistic regression was used to determine the ability of patients’ clinical characteristics and MPS to predict future cardiac events. In the analysis, the following variables were introduced: age, sex, duration of hemodialysis, serum adhesion molecules, gated MPS and echocardiography results.

Results Patient characteristics

The 31 patients enrolled in the study had been treated by hemodialysis for 12–228 months (median, 36 months). The most common cause of renal failure was glomerulonephritis (n = 7, 23%). Only one patient had diabetes.

Coronary angiography

Coronary angiography was performed in patients with an SSS of more than four and/or in those who had at least two segments with a wall motion score of greater than three (severe hypokinesia or worse) on echocardiography. Coronary angiography was performed using the Seldinger technique, with multiple left and right anterior oblique projections and craniocaudal angulations (Siemens, Bicol, Germany). The presence of significant CAD was defined as a reduction in luminal diameter of more than 50%.

Stress test results and SPECT MIBI imaging

Baseline ECG abnormalities were present in 15 patients, which included ST segment changes in 11 and left ventricular hypertrophy in nine. Five patients managed to complete an adequate exercise stress test, whereas 26 individuals were given pharmacological stress. Atropine was administered to five patients who failed to reach the maximum predicted heart rate with dobutamine infusion, all of whom finally achieved the target heart rate.

Follow-up

Patients were followed up from the time of MPS or cardiac catheterization until June 2004. The median follow-up interval was 20 months (range, 14–28 months). The outcome events studied were cardiac morbidity and all-cause mortality. A cardiac event in this study was defined as cardiac death, myocardial infarction, angina pectoris or coronary revascularization, which were assessed by patient interviews and verified by hospital records.

The heart rate and systolic blood pressure increased from rest to peak stress (78 to 127 beats/min and 120 to 151 mmHg, respectively; P < 0.01), whereas the diastolic blood pressure did not change significantly (P > 0.05). ST segment depression occurred in four patients and one experienced chest pain. Minor symptoms, including dizziness, headache and nausea, were reported by six patients, but no severe episodes of ischemia or lifethreatening complications developed.

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Silent ischemia in hemodialysis patients Caglar et al. 65

Table 2

Semiquantitative analysis of stress (myocardial perfusion scintigraphy, MPS)

Stress MPS Scintigraphic extension score (range) Summed stress score (range)

Abnormal MPS (n = 9)

Normal MPS (n = 22)

P value

3.5 ± 1.6 (2–7) 7.7 ± 2.2 (4–10)

0.7 ± 0.8 (0–2) 0.9 ± 1 (1–3)

< 0.01 < 0.01

Table 3 Gated single-photon emission computed tomography (SPECT) results in patients with normal and abnormal myocardial perfusion scintigraphy (MPS) Measured parameter Left ventricular ejection fraction (%) End-systolic volume (ml) End-diastolic volume (ml)

All patients (n = 31)

Abnormal MPS (n = 9)

Normal MPS (n = 22)

P value

51 ± 11 56 ± 32 110 ± 46

44 ± 10 73 ± 45 129 ± 56

54 ± 10 50 ± 25 104 ± 42

< 0.05 < 0.05 > 0.05

Gated myocardial perfusion was normal in 22 patients (71%) with a mean SSS of 0.9 ± 1 (Table 2). Fixed perfusion defect was seen in one patient, involving the anterior wall, and reversible defects were seen in eight patients, with reduced myocardial uptake in the anterior, septal or apical zones in three studies and in the inferior/posterobasal wall in six (Fig. 2). One patient with normal perfusion had global hypokinesia involving the left ventricle on the post-stress study (Fig. 3). Gated SPECT results in patients with normal and abnormal MPS are summarized in Table 3. No significant difference was observed between the end-diastolic volumes in patients with normal and abnormal MPS, whereas LVEF was significantly lower and the endsystolic volume was higher in patients with abnormal MPS. Coronary angiography was performed in all eight patients with perfusion defects and one patient considered to be at high risk for CAD due to reduced LVEF on gated MPS following stress despite normal perfusion. Coronary angiography results were abnormal in six of the nine patients evaluated (19% of the total study population). Four patients presented with one-vessel stenosis, involving the right coronary artery in three and the left anterior descending artery in one. One patient had twovessel disease involving the circumflex and right coronary arteries and one patient had triple-vessel disease. The findings of MPS typically corresponded to the coronary artery lesions identified (Table 4). Using a 50% obstruction in at least one vessel to denote significant disease, three of the nine patients who were referred for coronary angiography had normal coronary arteries (Fig. 4). All of these patients had left ventricular hypertrophy (mean left ventricular mass, 171 g/m2). Coronary angiography was not performed in patients with normal MPS for ethical reasons, and it is known that a negative MPS result rules out CAD with a high probability [14]. Sixteen studies performed between 1994 and 2001, which reported 20 983 patients with normal MPS and a mean follow-up

of 28 months, showed a rate of cardiac death or myocardial infarction of 0.7% per year [15]. Similar findings have been reported in a multicenter registry of 4728 patients [16]. Echocardiography results were abnormal in nine patients (29%). These abnormalities included mild septal hypokinesia, pericardial effusion, left ventricular hypertrophy and diastolic dysfunction in three, three, six and one patient, respectively. The mean values of the indexed left ventricular mass and ejection fraction were 149 ± 51 g/m2 and 70 ± 7%, respectively. No significant difference was found in LVEF, end-systolic and end-diastolic volumes and left ventricular mass between patients with normal and abnormal MPS. Variables associated with abnormal test results

There were significant differences between patients with perfusion defects and those with normal MPS regarding the intima–media thickness of the common carotid artery and serum levels of C-reactive protein, homocysteine, sVCAM, sICAM, sP-selectin and sE-selectin. All of these variables were higher and the duration of hemodialysis was significantly longer in patients with abnormal MPS than in those with normal scans (132 ± 89 months versus 59 ± 60 months; P < 0.05) (Table 5). Abnormal stress test results were not significantly associated with systolic/ diastolic blood pressure, heart rate, age, sex, 24-h ambulatory blood pressure readings, serum lipids, apolipoproteins and albumin. Follow-up results

Follow-up lasted for 20 months (range, 14–28 months). During this period, one patient died from sepsis and two patients were admitted to hospital for angina pectoris in the group with normal MPS. In the group with abnormal MPS, seven patients sustained cardiac events. The prevalence of cardiac events was significantly different in the two groups (13% in the normal MPS group and 75% in the abnormal MPS group; P < 0.01). The prognostic positive predictive value of MPS was 77% and its negative predictive value was 91%. The echocardiographic findings

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Fig. 2

(a) Reversible perfusion defect is seen in the inferior and septal walls on 99m Tc-methoxyisobutylisonitrile (99mTc-MIBI) scan. (b) Quantitative analysis confirms the presence of inferoseptal wall ischemia. (c) Coronary angiography demonstrates severe narrowing in the right coronary artery.

were not significantly different between patients who suffered or did not suffer a cardiac event. The time between myocardial SPECT and cardiac event varied

Fig. 3

Myocardial perfusion scintigraphy with 99mTc-methoxyisobutylisonitrile (99mTc-MIBI) reveals normal tracer distribution on stress and rest images (a) and on quantitative analysis (b). (c) Quantitative gated single-photon emission computed tomography (SPECT) analysis shows a reduced post-stress left ventricular ejection fraction (39%) which is due to myocardial stunning. (d) Severe occlusion in the right coronary artery is seen on coronary angiography.

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Silent ischemia in hemodialysis patients Caglar et al. 67

Table 4

Comparison between scintigraphy and coronary angiography in patients with abnormal myocardial perfusion scintigraphy (MPS)

Patient

MPS

Stress gated

Summed stress score

LVH

Coronary angiography

1 2 3 4 5

Inferior and septal ischemia Apico-anteroseptal ischemia Basal inferior ischemia Septal ischemia Inferior ischemia

Inferior hypokinesia Apical hypokinesia Inferior hypokinesia Normal Inferior hypokinesia

10 4 6 5 9

– + + – +

6 7

Inferior ischemia Anterior myocardial infarction Inferior ischemia

Septal hypokinesia Global hypokinesia

4 6

+ +

8 9

Inferoseptal ischemia Normal

N Global hypokinesia

5 2

– +

RCA 50% Normal Normal LAD 50% RCA 70% CX 60% Normal RCA 60% CX 70% LAD 90% RCA 50% RCA 85%

CX, left circumflex artery; LAD, left anterior descending artery; LVH, left ventricular hypertrophy; RCA, right coronary artery. Table 5

Comparison between risk factors and myocardial perfusion scintigraphy (MPS)

Risk factor

Abnormal MPS (n = 9)

Normal MPS (n = 22)

P value

0.77 ± 0.39 29.6 ± 10.2 330 ± 111 201 ± 136 1345 ± 362 1257 ± 283 132 ± 89 0.87 ± 0.24

0.48 ± 0.35 22.99 ± 10.93 209 ± 95 98 ± 108 907 ± 379 969 ± 307 59 ± 60 0.59 ± 0.13

< 0.05 < 0.05 < 0.01 < 0.01 < 0.01 < 0.01 < 0.05 < 0.01

C-reactive protein (mg/dl) Homocysteine (mmol/l) sP-selectin (ng/ml) sE-selectin (ng/ml) sVCAM (ng/ml) sICAM (ng/ml) Hemodialysis duration (months) Intima–media thickness of common carotid artery (mm)

sE-selectin, soluble E-selectin; sICAM, soluble intercellular adhesion molecule; sP-selectin, soluble P-selectin; sVCAM, soluble vascular cell adhesion molecule.

Table 6

Long-term follow-up results of patients who sustained cardiac events Myocardial perfusion scintigraphy

Coronary angiography

Cardiac event

Time (months)a

1 2 3 4

Inferior/septal ischemia Apico-anteroseptal ischemia Basal inferior ischemia Inferior ischemia

AP AP AP PTCA

9 3 3 1

5 6

Inferior ischemia Anterior MI Inferior ischemia

MI PTCA

8 1

7

Normal perfusion Global hypokinesia Normal Normal

RCA 50% Normal Normal RCA 70% CX 60% Normal RCA 60% CX 70% LAD 90% RCA 85%

PTCA

1

AP AP

11 13

Patient

8 9, hemodialysis

b

Normal Normalb

AP, angina pectoris; CX, left circumflex artery; LAD, left anterior descending artery; MI, myocardial infarction; PTCA, percutaneous transluminal angioplasty; RCA, right coronary artery. a Time between MPS and cardiac event. b Coronary angiography was performed within 24 h of the cardiac event.

from 1 to 13 months. Logistic regression analysis showed that only SSS was significantly related to cardiac events (odds ratio, 20; P = 0.003; 95% confidence interval, 2.68–149.0). The pertinent data in patients with events are summarized in Table 6.

Discussion In this study, we found that nine (29%) of our hemodialysis patients, all asymptomatic, had perfusion and/or contraction abnormalities on gated MPS. Six of these nine patients were found to have significant CAD on angiography, yielding a positive predictive value of

67%; however, all three patients with normal coronary angiography had cardiac events in the follow-up period. Perfusion defects in a given vascular territory may result from an obstruction to flow at the site of narrowing or a more diffuse atherosclerosis with a progressive accumulation of resistance to blood flow. Formerly often claimed to be ‘false positive’, recent studies suggest that discordant MPS and coronary angiography results in patients with chronic renal failure may be due to endothelial dysfunction, abnormal coronary reserve and left ventricular hypertrophy [17–19]. Left ventricular hypertrophy, which is accompanied by coronary microcirculatory

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Fig. 4

dysfunction in the subendocardium, causes impaired vasodilator reserve and hypoperfusion [20,21]. Therefore, our ‘low’ positive predictive value may relate to the fact that MPS traces perfusion abnormalities, not necessarily caused by epicardial CAD, but possibly due to microvascular disease [22]. This underlines the important role of the microcirculation in the distribution of radiotracers. Verna et al. [23] have reported that angiographically unrecognized occult atherosclerotic defects and abnormal vasodilatation capacity of the coronary circulation can be seen on SPECT images as reversible perfusion defects, based on intravascular sonography studies. The diagnostic efficacy of MPS has been clearly shown previously for the dialysis population. In a study by Derfler et al. [24], in which 36 patients on chronic hemodialysis were evaluated, thallium MPS was found to be the only predictor for identifying hemodialysis patients at increased risk for cardiovascular events. Likewise, Feola et al. [25], who studied 61 pretransplant renal patients, found a significantly higher incidence of cardiac events in patients with reversible defects compared with those with a normal perfusion pattern on thallium scintigraphy. Our second goal was to identify predictors of test abnormalities. Our results indicate that circulating biomarkers of inflammation are related to myocardial ischemia. A large body of evidence points to the involvement of adhesion molecules in the pathophysiology of myocardial ischemia [26]. Focal expression of adhesion molecules has been observed in atherosclerotic plaques and their serum levels have been postulated to be useful risk predictors of cardiovascular events. When stimulated, leukocytes and endothelium express adhesion molecules, which play a pivotal role in ischemic myocardial damage through microvascular obstruction by releasing inflammatory cytokines and oxygen free radicals. Ridker et al. [27] reported that C-reactive protein correlated with CAD in a group of low-risk men, and high levels of C-reactive protein were found in subjects later developing atherosclerotic disease [28].

(a) Reduced uptake is seen in the inferior wall on myocardial perfusion scintigraphy (MPS). (b) Quantitative analysis confirms mild inferior wall ischemia (summed stress score = 4). (c) Coronary angiography was performed on this patient due to inferior wall ischemia and septal hypokinesia. Although coronary angiography was normal, this patient had myocardial infarction involving the inferior wall at 8 months of follow-up.

This study also confirms the relationship between serum homocysteine concentrations and myocardial perfusion defects. Significant homocysteinemia is present from the early stage of chronic renal failure and may constitute a risk factor for premature atherosclerosis [29]. The mechanism by which homocysteine enhances atherosclerosis is uncertain, although the postulated mechanisms include direct endothelial cytotoxicity [30,31] and the encouragement of smooth muscle growth [32]. Hyperhomocysteinemia is associated with altered endothelial-dependent vasodilatation, suggesting interference with nitric oxide [33]. In the present study, patients with abnormal MPS had a longer duration of hemodialysis. The time during which

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Silent ischemia in hemodialysis patients Caglar et al. 69

patients are uremic and may suffer hypertension has a major impact on the frequency and severity of CVD [34]. The increased risk is associated with a higher incidence of atherosclerotic disease, left ventricular hypertrophy and congestive heart failure.

16

17

18

In conclusion, the current study shows that gated SPECT MPS provides important diagnostic and prognostic information in patients undergoing hemodialysis; SSS was the single most important predictor of subsequent cardiac events. Many risk factors, including the duration of hemodialysis, elevated levels of serum adhesion molecules and homocysteine, account for the increased incidence of CAD.

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Shaw LJ, Hendel R, Borges-Neto S, Lauer MS, Alazraki N, Burnette J, et al. Prognostic value of normal exercise and adenosine Tc-99m-tetrofosmin SPECT imaging: results from the multicenter registry of 4,728 patients. J Nucl Med 2003; 44:134–139. Annuk M, Zilmer M, Fellstrom B. Endothelium-dependent vasodilatation and oxidative stress in chronic renal failure: impact on cardiovascular disease. Kidney Int Suppl 2003; 84:S50–S53. Narita M, Kuhira T. Scintigraphic assessment of patients with electrocardiographic left ventricular hypertrophy with ST-T changes without apparent cause. Clin Nucl Med 2002; 27:641–647. Wieneke H, Zander C, Eising EG, Haude M, Bockisch A, Erbel R. Non-invasive characterisation of cardiac microvascular disease by nuclear medicine single photon emission tomography. Herz 1999; 24: 515–521. Iriarte M, Caso R, Murga N. Microvascular angina pectoris in hypertensive patients with left ventricular hypertrophy and diagnostic value of Thallium201 scintigraphy. Am J Cardiol 1995; 75:335–339. Rajappan K, Rimoldi OE, Dutka DP, Ariff B, Pennell DJ, Sheridan DJ, et al. Mechanisms of coronary microcirculatory dysfunction in patients with aortic stenosis and angiographically normal coronary arteries. Circulation 2002; 105:470–476. Fragasso G. Diagnosing coronary artery disease in patients with hypertension: a resolved dilemma? Ital Heart J 2000; 1:726–731. Verna E, Ceriani L, Giovanella L, Binaghi G, Garancini S. ‘False-positive’ myocardial perfusion scintigraphy findings in patients with angiographically normal coronary arteries: insights from intravascular sonographic studies. J Nucl Med 2000; 41:1935–1940. Derfler K, Kletter K, Balcke P, Heinz G, Dudczak R. Predictive value of thallium-201-dipyridamole myocardial stress scintigraphy in chronic hemodialysis patients and transplant recipients. Clin Nephrol 1991; 36:192–202. Feola M, Biggi A, Ribichini F, Rovere A, Vado A, Camuzzini G, et al. Predicting cardiac events with Tl201 dipyridamole myocardial scintigraphy in renal transplant recipients. J Nephrol 2002; 15:48–53. Miyao Y, Miyazaki S, Goto Y, Itoh A, Daikoku S, Morii I, et al. Role of cytokines and adhesion molecules in ischemia and reperfusion in patients with acute myocardial infarction. Jpn Circ J 1999; 63: 362–366. Ridker PM, Cushman M, Stampfer MJ, Tracy RP, Hennekens CH. Inflammation, aspirin and the risk of cardiovascular disease in apparently healthy men. N Engl J Med 1997; 336:973–979. Lind L. Circulating markers of inflammation and atherosclerosis. Atherosclerosis 2003; 169:203–214. Chauveau P, Chadefaux B, Coude M, Aupetit J, Hannedouche T, Kamoun P, et al. Hyperhomocysteinemia, a risk factor for atherosclerosis in chronic uremic patients. Kidney Int Suppl 1993; 41:S72–S77. Starkebaum G, Harlan JM. Endothelial cell injury due to copper-catalyzed hydrogen peroxide generation from homocysteine. Clin Invest 1986; 77:1370–1376. Loscalzo J. The oxidant stress of hyperhomocysteinemia. J Clin Invest 1996; 98:5–7. Tsai JC, Perrella MA, Yoshizumi M, Hsieh CM, Haber E, Schlegel R, et al. Promotion of vascular smooth muscle cell growth by homocysteine: a link to atherosclerosis. Proc Natl Acad Sci USA 1994; 91: 6369–6373. Tawakol A, Omland T, Gerhard M, Wu JT, Creager MA. Hyperhomocyst(e)inemia is associated with impaired endotheliumdependent vasodilation in humans. Circulation 1997; 95:1119–1121. Denker BM, Chertow GM, Owen WF. Hemodialysis. In: Brenner BM (editor): The kidney. 6th ed. Pennsylvania: W.B. Saunders; 2000, pp. 2373–2453.

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Original article

Characterization of 133Xe gas washout in pulmonary emphysema with dynamic 133Xe SPECT functional images Kazuyoshi Suga, Yasuhiko Kawakami, Tomio Yamashita, Mohammed Zaki and Naofumi Matsunaga Purpose To characterize regional ventilation impairment of pulmonary emphysema using dynamic 133Xe single photon emission computed tomography (SPECT) functional images, compared with other forms of chronic obstructive pulmonary disease (COPD). Methods Dynamic 133Xe SPECT was performed in 34 patients with emphysema and 15 patients with other forms of COPD. Three-dimensional voxel-based functional images of the half-clearance time (T1/2) mainly reflecting the initial rapid washout of 133Xe gas from the large airways, and of the mean transit time (MTT) reflecting 133Xe gas washout from the entire lungs, including the small airways and alveoli, were created based on an area-over-height method. T1/2 and MTT values were compared with the regional extent of low attenuation areas (%LAA) on density-mask computed tomography images and the diffusing capacity of the lungs for carbon monoxide (DLCO). Results The MTT/T1/2 ratio in each lung in emphysema was significantly higher than that in other forms of COPD (1.60 ± 0.74 vs. 1.21 ± 0.26; P < 0.01). In the selected unilateral lungs with similar T1/2 values, MTT values were also significantly higher in emphysema. MTT values in each lung showed a significantly closer correlation with the corresponding %LAA values compared with T1/2 values in emphysema (R = 0.698, P < 0.0001 vs. R = 0.338, P < 0.01; P < 0.05); while only the T1/2 values showed a significant correlation in other forms of COPD (P < 0.0001). In correlation with DLCO, MTT values showed a significantly

Introduction Dynamic 133Xe gas single photon emission computed tomography (SPECT) is a useful tool in quantification of regional ventilation on cross-sectional lung planes in chronic obstructive pulmonary diseases (COPDs) [1–10]. The process of 133Xe gas washout can be divided into the initial, rapid, washout phase, which mainly reflects rapid emptying of 133Xe gas from the relatively large airways, and the late, slow, washout phase, which reflects the slow elimination of 133Xe gas via the smaller airways at the level of the terminal bronchioles and alveolar air spaces [3–5,10,11]. The late, slow, washout phase may be particularly affected in pulmonary emphysema with significant alveolar destruction, and the process of 133Xe gas washout in this disease may be different from that

closer correlation compared with T1/2 values in emphysema (R = 0.909, P < 0.0001 vs. R = 0.555, P < 0.001; P < 0.05); while either value did not show a significant correlation in other forms of COPD. Conclusion MTT values are more critically affected in emphysema compared with other forms of COPD without significant alveolar destruction, and MTT and T1/2 values appear to be differently correlated with the regional extent of LAA between these two disorders. Direct comparison of regional T1/2 and MTT values on functional images may contribute to the demarcation of lung pathology of these c 2006 two disorders. Nucl Med Commun 27:71–80 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:71–80 Keywords: single photon emission computed tomography (SPECT), 133Xe gas, lung ventilation, pulmonary emphysema, chronic obstructive pulmonary disease (COPD) Department of Radiology, Yamaguchi University School of Medicine, Japan. Correspondence to Dr Kazuyoshi Suga, Department of Radiology, Yamaguchi University School of Medicine, 1-1-1 Minamikogushi, Ube, Yamaguchi 755-8505, Japan. Tel: + 0081 836 22 2383; fax: + 0081 836 22 2285; e-mail: [email protected] Sponsorship: This study was supported, in part, by a Research Grant for Scientific Research (08671033) from the Japanese Ministry of Education. Received 18 January 2005 Accepted 15 July 2005

in other forms of COPD without significant alveolar destruction [9–11], although, morphologically, these two disorders show similar appearance of low attenuation areas (LAAs) on computed tomography (CT) images [12– 17]. The aim of the present study was to assess the difference in the pulmonary washout of 133Xe gas between emphysema and other forms of COPD, by using recently developed three-dimensional (3-D) volume-rendering functional images of dynamic 133Xe SPECT, which provide the voxel-based ventilatory parameters of the half-clearance time T1/2 mainly reflecting the initial rapid 133 Xe gas washout phase and of the mean transit time (MTT) reflecting both the initial rapid and late slow

c 2006 Lippincott Williams & Wilkins 0143-3636

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72 Nuclear Medicine Communications 2006, Vol 27 No 1

washout phases. For this assessment, we also compared these two ventilation parameters with regional extent of low attenuation areas (%LAA) on density-mask CT images, and the diffusing capacity of the lungs for carbon monoxide (DLCO) as an indicator of alveolar–capillary destruction.

negative for a pulmonary embolic event. The SPECT procedure was approved by the Institutional Review Board of the Yamaguchi University School of Medicine, and informed consent was obtained from all subjects after the nature of the procedure had been fully explained. Dynamic

Materials and methods Patient population

The subjects were 34 smoker patients (33 men and one woman; age, 54–83 years, mean 65 ± 8 years) with emphysema and 15 non-smoker patients (nine men and six women; age, 29–78 years, mean 59 ± 13 years) with other forms of COPD (six patients with obstructive bronchiolitis, five with chronic bronchitis, three with diffuse panbronchiolitis and one patient with bronchial asthma) underwent dynamic 133Xe gas SPECT between July 2001 and June 2004. All subjects underwent high resolution CT scanning within 2 weeks of dynamic 133Xe SPECT. The diagnosis of emphysema was made according to CT image findings, the smoking history and the criteria of the American Thoracic Society [18]. All 34 patients with emphysema had a long-term cigarette smoking history, with the mean Brinkman index (cigarettes consumption per day  years; 467 ± 52). Twenty-six of these patients had heterogeneously focal low attenuation areas (LAA) on high resolution CT images, and the remaining eight had diffuse LAA throughout the lungs, with or without bullae. The diagnosis of other forms of COPD was made according to clinical history and feature and typical findings on high resolution CT images. All 15 patients with this disorder had no history of cigarette smoking, and had focal LAA on CT images, with bronchial wall thickening, bronchiectasis or small abnormal centrilobular opacities indicative of bronchiolar inflammatory changes. Of these patients, six with obstructive bronchiolitis had undergone open lung biopsy within 6 months before 133Xe SPECT, which had shown no significant alveolar destruction. All patients had dyspnoea at rest or during mild exercise (grade 1 in three patients, grade 4 in 11 patients, grade 4 in 22 patients, grade 5 in 13 patients, according to Fletcher’s dyspnoea scale [19]), and the average values of predicted forced expiratory volume in 1 s (%FEV1) and vital capacity (%VC) were 48.3% ± 12.8 (range, 23.3–68.6%) and 82.1% ± 17.5 (range, 76.4– 108.7%), respectively. For reference, 133Xe SPECT findings of six non-smoker patients with normal high resolution CT findings and %FEV1 and %VC values (six women; age, 42–51 years, mean 51 ± 6 years) who underwent combined 133Xe gas ventilation and 99mTc macroaggregated albumin perfusion SPECT examinations for suspected pulmonary embolism because of the presence of deep venous thrombosis during the same study period. These patients showed normal ventilation and perfusion, being diagnosed as

133

Xe gas SPECT

Dynamic 133Xe gas SPECT was performed using a continuous repetitive rotating acquisition mode with a triple-detector SPECT system (GCA 9300 A/HG, Toshiba Medical System, Tokyo, Japan) and 133Xe gas control system (AZ-702, Anzai Medical, Osaka, Japan) [6–9]. On the image data processor of GMS-5500U, SPECT data were reconstructed into equilibrium and washout images in 51–64 one-pixel (3.2 mm) thick transaxial planes covering the entire lungs, using a Butterworth prefilter (order no. 8, cut-off frequency 0.13 cycle/cm) and ramp back-projection filter. The lung area was segmented with a threshold level of 25% of the maximal voxel 133Xe activity in the equilibrium data. Then, the real halfclearance time (T1/2 = 133Xe clearance time required to reach the 50% level of the equilibrium count rate during washout, in seconds) and the mean transit time (MTT) were calculated by a computer for every voxel. MTT values were estimated based on an area-over-height method (a modified Stewart–Hamilton method) according to the theory of MTT described by Zierler [20] and Sacker-Walker et al. [2]; MTT = V/F = ADT/H = the total counts collected during washout/the counts at equilibrium, where V is the volume of distribution, F is the air flow (assumed constant), H is the counts during an equilibrium interval of DT, and A is the collected counts during washout. T1/2 and MTT values were mapped in a voxel base on the underlying lung contour image derived from the equilibrium data. The colour scale of each voxel was made proportional to the magnitude of each parameter. These transaxial T1/2 and MTT images were also reconstructed into 3-D images, using a volumerendering and maximum intensity projection technique, with a 64  64  64 voxel. These 3-D images could be freely rotated at any angle of view. When the user interactively selected the lung section level, the corresponding orthogonal (transaxial, coronal and sagittal) section images were simultaneously displayed. All these images were created using the software (NSIS-034A, Lung function SPECT analysis program, Toshiba Medical System, Tokyo, Japan) equipped in the image data processor. Density-mask computed tomography images

CT images were obtained using the multidetector row CT scanner with four rows of a 0.5 mm detector (Siemens Volume Zoom, Siemens-Asahi Medical Ltd., Tokyo, Japan). High resolution CT images, 3 mm thick, were obtained at rest inspiratory breath-hold, using 3 mm collimation, scan time 1.0 s, 120 kVp, and 230 mA. To objectively assess the extent and location of LAA, a

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Dynamic

density-mask CT image of each lung section was automatically created by the supplemental software in the data processor. The density-mask software program highlighted each slice voxel with CT attenuation values less than – 960 Hounsfield units [8,12,13]. The densitymask program also automatically gave the area of the highlighted voxels of LAA for each slice, and the per cent lung involvement of LAA (%LAA) was calculated at any selected lung levels and in each lung. Data analysis

Two observers (T.Y. and Y.K.) compared the distributions of T1/2, MTT and LAA in each transaxial lung section. In this procedure, the series of SPECT and density-mask CT images were simultaneously displayed side-by-side on the image viewer (Yokogawa-GE Medical, Tokyo, Japan) connected to the CT and SPECT systems, and the matched CT and SPECT slices were selected at the same

133

Xe SPECT functional images Suga et al. 73

cranio-caudal level. An adequate correspondence of CT and SPECT slices was confirmed by comparing the slice geometry from the lung apex and diaphragmatic contours on CT and SPECT scout view images, and by correlating with the contours of the mediastinal and hilum structures. Although there might be slight mismatching between the selected slices, this effect seemed inconsequential, because each of T1/2, MTT and LAA almost similarly appeared in at least two or three adjacent sections above and/or below. If an apparent dissociation among those regional distributions was seen, the locations were reported by consensus of these two observers. For quantitative assessment, the mean T1/2 and MTT values in each unilateral lung were estimated in all subjects, and the values were compared with the corresponding %LAA values. T1/2 and MTT values were also estimated in the entire lungs in the patients,

Fig. 1

R

13

21

13

50

Transaxial

50

Transaxial

Sagittal T1/2

14

Sagittal MTT

0

0 Coronal

3D (Ant)

Coronal

3D (Ant)

A 42-year-old female normal subject without a history of smoking. Three-dimensional 133Xe gas SPECT functional images and their orthogonal section images show nearly uniform distribution of T1/2 and MTT values (the mean T1/2 and MTT values in the entire lungs; 35.5 s and 38.4 s, respectively), but with higher values in the lower ventral lungs, indicating a physiological gravitational effect on pulmonary ventilation (arrows).

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74 Nuclear Medicine Communications 2006, Vol 27 No 1

which were compared with DLCO as an indicator of alveolar–capillary destruction, using a linear regression analysis. DLCO was measured by the single-breath method (Pulmorecorder, model R1551S; Anima, Tokyo, Japan), according to the recommendation of the American Thoracic Society [21–23]. Statistical analysis

The values were expressed as mean ± standard deviation (SD), and a paired or unpaired Student’s t-test was applied to compare the differences. A P-value less than 0.05 was regarded as significant. A linear regression analysis to evaluate the correlation of T1/2 and MTT values with %LAA values was performed using the commercially available software (StatView 4.02 SE + Graphics; Abacus Concepts, Berkeley, California, USA), and a P-value less than 0.05 was considered significant

for each correlation coefficient (R). The difference of R values was evaluated by the Neyaman–Pearson test with Fischer’s Z transformation scores.

Results In the normal, control subjects, T1/2 and MTT images showed almost uniform distribution of these values, but with the higher MTT and T1/2 values in the lower ventral lungs, indicating a physiological gravitational effect on pulmonary ventilation (Fig. 1). In contrast, in patients with COPD, heterogeneously prolonged T1/2 and MTT values were seen with loss of physiological gravitational effect even in cases with symmetrical LAA and in the normally appearing areas on density-mask CT images (Figs 2–4). Apparent dissociation in the location of prolonged T1/2 and MTT values was found in 23 (76.4%) of patients with emphysema and in five

Fig. 2

R

400

400

Transaxial

Transaxial

Sagittal

MTT

T1/2

Coronal

Sagittal

3D (Ant)

0

Coronal

3D (Ant)

0

A 67-year-old man with pulmonary emphysema. CT and density-mask CT images (top) show low attenuation areas (LAA) in both lungs (the mean %LAA in the entire lung was 53.4%). Three-dimensional functional images and their orthogonal section images including the transaxial images at the same level of the CT plane show heterogeneously prolonged T1/2 and MTT values in both lungs, regardless of diffuse LAA (the mean T1/2 and MTT values in the entire lungs were 215.2 s and 315.3 s, respectively) (bottom). Apparent dissociations between T1/2 and MTT distributions are seen (arrows). DLCO was markedly decreased to 3.8 ml
min – 1 per mmHg.

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Dynamic

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Xe SPECT functional images Suga et al. 75

Fig. 3

19

R 400

400

Sagittal

Transaxial 10

Transaxial 9

T1/2

Coronal

Sagittal

MTT

3D (Ant)

0

Coronal

3D (Ant)

0

A 68-year-old man with emphysema. CT and density-mask CT images (top) show low attenuation areas (LAA) in both lungs (the mean %LAA in the entire lung was 62.5%). Three-dimensional functional images and their orthogonal section images including the transaxial images at the same level of the CT plane show heterogeneously prolonged T1/2 and MTT values in both lungs (the mean T1/2 and MTT values in the entire lungs were 225.4 s and 350.3 s, respectively) (bottom). MTT values are more prolonged than T1/2 values. DLCO was markedly decreased to 3.9 ml
min – 1 per mmHg.

(33.3%) patients with other forms of COPD, by consensus of the two observers (Fig. 2). All the T1/2, MTT and %LAA values in each lung in patients with emphysema and other forms of COPD were significantly higher compared with the corresponding values in the normal control subjects (P < 0.0001) (Table 1). Regardless of no significant difference in the %LAA values between these patients, the T1/2, MTT values were significantly higher in patients with emphysema (P < 0.005 and P < 0.0001, respectively). The MTT/T1/2 ratio in each lung in patients with emphysema was significantly higher than that in patients with other forms of COPD (1.60 ± 0.74 vs. 1.21 ± 0.26; P < 0.01) (Fig. 5). When comparing the mean MTT values in the 10 selected unilateral lungs of 10 patients with almost the same T1/2 values (144.7 s ± 64.4 vs.

145.2 s ± 66.1; NS), these values were significantly higher in emphysema (208.4 s ± 72.9 vs. 171.3 s ± 53.6; P < 0.01). The T1/2 and MTT values in each lung showed a significant correlation with the corresponding %LAA values (P < 0.01 and P < 0.0001, respectively), where the MTT values showed a significantly closer correlation compared with the T1/2 values in emphysema (R = 0.698, P < 0.0001 vs. R = 0.338, P < 0.01; P < 0.05) (Fig. 6(a)), while in other forms of COPD, only the T1/2 values were significantly correlated with %LAA values (P < 0.0001) (Fig. 6(b)). DLCO values of 7.5 ml
min–1 per mmHg ± 4.1 (range, 2.1–16.5) in patients with emphysema were significantly lower compared with those of 18.3 ml
min–1 per mmHg ± 2.1 (range, 15.3–22.1) in patients with other forms of COPD (P < 0.0001). The mean MTT values in

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76 Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 4

20

R

200

200

Sagittal

Transaxial

Transaxial

T1/2

14

Sagittal MTT

0

0 Coronal

˚)

3D (RAO 45

Coronal

3D (RAO

45˚)

A 29-year-old man with another form of COPD (obstructive bronchiolitis). CT and density-mask CT images (top) show LAA in both lungs (the mean %LAA in the entire lung was 75.9%). On three-dimensional functional images and their orthogonal section images including the transaxial images at the same level of the CT plane heterogeneously prolonged T1/2 and MTT values in both lungs (the mean T1/2 and MTT values in the entire lungs were 155.2 s and 164.6 s, respectively) (bottom). T1/2 and MTT distributions are almost consistent. DLCO was relatively preserved with 15.6 ml
min – 1 per mmHg, regardless of extended LAA.

Table 1 Comparison of half-clearance time (T1/2), mean transit time (MTT) and regional extent of low attenuation areas (%LAA) in each unilateral lung Subjects Normal control subjects (n = 12 unilateral lungs) Patients with pulmonary emphysema (n = 68 unilateral lungs) Patients with other forms of COPD (n = 30 unilateral lungs)

T1/2 (s)

MTT (s)

%LAA

34.5 ± 4.2 158.7 ± 61.0 (P < 0.005) 120.2 ± 25.8

35.2 ± 4.6 213.3 ± 78.6 (P < 0.0001) 142.6 ± 28.3

1.2 ± 0.9 31.6 ± 17.9 (NS) 39.7 ± 18.2

COPD, chronic obstructive pulmonary disease. All the T1/2, MTT and %LAA values in patients with pulmonary emphysema and other forms of COPD are significantly higher compared with those in the normal control subjects (P < 0.0001). Superscript: comparison between patients with pulmonary emphysema and other forms of COPD.

the entire lungs showed a significantly closer correlation with DLCO compared with T1/2 values in emphysema (R = 0.909, P < 0.0001 vs. R = 0.555, P < 0.001; P < 0.05) (Fig. 7(a)). In contrast, these two values were not significantly correlated with DLCO in other forms of COPD (Fig. 7(b)).

Discussion The present functional images showed certain differences in pulmonary washout of 133Xe gas between emphysema and other forms of COPD. Regional MTT values appear to be more critically affected in emphysema, as the MTT/T1/2 ratio in each lung was significantly

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Dynamic

Fig. 5

P < 0.01

MTT/T½ ratios in each unilateral lung

n = 68 unilateral lungs 5.5 5.0

1.60 ± 0.74

4.0

n = 30 unilateral lungs 3.0

1.21 ± 0.26

2.0 1.0 0 Pulmonary emphysema

Other forms of COPD

Comparison of MTT/T1/2 ratios in each unilateral lung between 34 patients with emphysema and 15 patients with other forms of COPD. Each box represents mean ± standard deviation (SD). MTT/T1/2 ratios are significantly higher in patients with emphysema than in patients with other forms of COPD (1.60 ± 0.74 vs. 1.21 ± 0.26; P < 0.01).

higher in this disorder compared with other forms of COPD. In the selected unilateral lungs with almost the same T1/2 values, MTT values were also significantly higher in this disorder. In a correlation of MTT and T1/2 values with the regional extent of LAA and DLCO, there were also certain differences between emphysema and other forms of COPD. Although the mean MTT and T1/2 values in each lung showed a significant correlation with the corresponding %LAA in emphysema, the T1/2 values alone showed a significant correlation in other forms of COPD. Although the mean MTT and T1/2 values in the entire lungs showed a significant correlation with DLCO in emphysema, both values did not show a significant correlation with DLCO. Airflow through the lung is principally divided into two different components: bulk and relatively rapid gas emptying from the large conducting airways (trachea down to the terminal bronchiole) and the subsequent slow gas elimination from the more distal small airways including the alveoli [3–5,10,11]. The entire 133Xe gas washout curves on dynamic SPECT can include these two different components of airflow; T1/2 values obtained from the initial rapid phase, and MTT values reflect the entire washout process including the later slow phase [3,4,10]. These two 133Xe gas washout processes are considered to be differently affected between emphysema and other forms of COPD. As seen in the significantly higher MTT/T1/2 ratios in patients with emphysema, regional MTT values are more critically affected in emphysema. Regionally destroyed alveolar space with

133

Xe SPECT functional images Suga et al. 77

reduced alveolar compliance and a loss of elastic lung coil strongly interrupt local air volume change and airflow rate in this disorder [10,24–27]. Rapid 133Xe gas emptying from the relatively large airways and the later, slow, washout process can be differently affected in regional lungs of emphysema, as an apparent dissociation in the location of prolonged T1/2 and MTT values was frequently seen. As indicated by the closer correlation of MTT values and DLCO compared with the T1/2 values in patients with emphysema, MTT values appear to more accurately reflect the severity of decreased integrity of the alveolar vasculature in this disorder. In contrast, in other forms of COPD without significant alveolar destruction, 133Xe gas washout from the relatively small airways and alveoli seems to be relatively preserved, regardless of prolonged T1/2 values. This may be attributed to less dissociation in the location of prolonged T1/2 and MTT values in patients with this disorder. Air trapping caused by alveolar air space destruction in emphysema and hyper-inflation caused by the relatively large airways obstruction in other forms of COPD similarly show LAA on CT images [11,14–16,28–37]. However, the present functional images revealed that regional ventilation were differently impaired between these two disorders. Regardless of no significant difference in the %LAA values in each lung, the T1/2 and MTT values were significantly higher in patients with emphysema. As MTT values in each lung showed a significantly closer correlation with %LAA compared with T1/2 values, LAA in emphysema appears to largely attribute to focal air trapping caused by alveolar air space destruction. In contrast, in other forms of COPD, T1/2 values in each lung alone showed a significant correlation with %LAA. LAA in other forms of COPD appears to be largely attributed to focal lung hyper-inflation caused by the obstruction of relatively large airways. Integrity of the alveolar vasculature seems to be relatively preserved regardless of the presence of LAA in this disorder, as no significant correlation was seen between T1/2 values and DLCO. The present functional images showed heterogeneously impaired ventilation even in the lung areas with diffuse LAA and in the areas that appear normal on density-mask CT images in patients with emphysema and other forms of COPD. Regardless of similar LAA appearance, regional ventilation should be heterogeneously affected according to the degree of airway obstruction, alveolar destruction, loss of lung elastic recoil and regional mechanical impairment of respiratory muscles [10,11,14,31]. Localized mild alveolar destruction in emphysema can be missed on high resolution CT images due to the limitation of spatial resolution [17,28–30,34]. Previous studies have shown several cases of emphysema which showed a relatively low prevalence of LAA on CT images,

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78 Nuclear Medicine Communications 2006, Vol 27 No 1

Fig. 6

Pulmonary emphysema

(a) n = 68 unilateral lungs y = 122.196 + 1.156x R = 0.338 P < 0.01

500

400

400

300

300

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T½(s)

500

200 100

200 100

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(b)

90 100

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30 40 50 % LAA

60

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60

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90 100

Other forms of COPD n = 30 unilateral lungs y = 84.397 + 0.922x R = 0.674, P < 0.0001

300

n = 30 unilateral lungs

300

250

250

200

200 MTT (s)

T½(s)

n = 68 unilateral lungs y = 116.348 + 3.07x R = 0.698 P < 0.0001

150

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50 % LAA

Correlation of T1/2 and MTT values with %LAA in each unilateral lung in 34 patients with emphysema (a) and 15 patients with other forms of COPD (b). In patients with emphysema (a), T1/2 and MTT values are significantly correlated with %LAA values (P < 0.01 and P < 0.0001, respectively), where the correlation coefficient (R) is significantly higher in MTT values (R = 0.698 vs. R = 0.338, Z transformation score = 2.916, P < 0.05) (a). However, in patients with other forms of COPD (b), T1/2 values alone are significantly correlated with %LAA values (P < 0.0001) (b).

regardless of severe airflow limitation in pulmonary function test [16,17,28–30]. The present functional images can display regional voxelbased ventilation parameters of T1/2 and MTT in a useful topographic form. Direct comparison of regional T1/2 and MTT values on these images is useful for clarifying the contribution of alveolar destruction and relatively large airways in ventilation impairment of COPD, as demonstrated in the present study. An inter-study comparison of these ventilation parameters would also be useful for an objective assessment of the treatment effects of bronchodilator or lung volume reduction surgery [11,34]. The drawbacks of the present study include the lack of histopathology in the majority of patients with other forms of COPD and the small study population of this

disease, although DLCO values indicated the relatively preserved integrity of the alveolar vasculature in these patients. A larger number of histologically proven cases and a study population that includes patients with bronchial asthma is warranted in a further study. More accurate measurement of %LAA on spirometrically controlled CT scanning is also desirable, because CT values of the lung are largely influenced by the depth of inspiration of the individual patient [6].

Conclusion The present volume-rendering functional images showed certain differences in the washout of 133Xe gas in patients with emphysema and other forms of COPD. MTT values appear to be well correlated with the severity of

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Dynamic

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Xe SPECT functional images Suga et al. 79

Fig. 7

(a)

Pulmonary emphysema 18

n = 34 patients y = 13.509 0.044x R = 0.555, P < 0.001

16 14

DLCO (ml·min−1 per mmHg)

DLCO (ml·min−1 per mmHg)

18

12 10 8 6 4 2 0

n = 34 patients y = 16.469 0.043x R = 0.909, P < 0.0001

16 14 12 10 8 6 4 2 0

50

100

150

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300

50

T½(s) Other forms of COPD

(b) n = 15 patients

25 DLCO (ml·min−1 per mmHg)

25 DLCO (ml·min−1 per mmHg)

100 150 200 250 300 350 400 450 MTT (s)

20 15 10 5 0 90

n = 15 patients

20 15 10 5 0 90 100 110 120 130 140 150 160 170 180 MTT (s)

100 110 120 130 140 150 160 170 180 T½(s)

Correlation of MTT and T1/2 values in whole lungs and DLCO in 34 patients with emphysema (a) and 15 patients with other forms of COPD (b). In patients with emphysema (a), T1/2 and MTT values are significantly correlated with DLCO (P < 0.001 and P < 0.0001, respectively), where the correlation coefficient is significantly higher in the MTT values (Z transformation score = 3.528, P < 0.05). However, in patients with other forms of COPD (b), these two values do not show a significant correlation with DLCO.

decreased integrity of the alveolar vasculature and to be more critically affected in emphysema compared with other forms of COPD without significant alveolar destruction. Regional MTT and T1/2 values also seem to be differently correlated with regional extent of LAA between these two disorders. Although further validation is required, direct comparison of regional T1/2 and MTT values on the functional images may be useful for demarcation of the lung pathology of these two disorders.

References 1 2

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Kreck TC, Krueger MA, Altemeier WA, Sinclair SE, Robertson HT, Shade ED, et al. Determination of regional ventilation and perfusion in the lung using xenon and computed tomography. J Appl Physiol 2001; 91: 1741–1749. 6 Suga K, Nishigauchi K, Kume N, Takano K, Koike S, Shimizu K, Matsunaga N. Ventilation abnormalities in obstructive airways disorder: detection with pulmonary dynamic densitometry by means of spiral CT versus dynamic Xe-133 SPECT. Radiology 1997; 202:855–862. 7 Suga K, Nishigauchi K, Kume N, et al. Dynamic pulmonary SPECT of xenon133 gas washout. J Nucl Med 1996; 37:807–814. 8 Suga K, Kume N, Matsunaga N, Ogasawara N, Motoyama K, Hara A, Matsumoto T. Relative preservation of peripheral lung function in smokingrelated pulmonary emphysema: assessment with 99mTc-MAA perfusion and dynamic 133Xe SPET. Eur J Nucl Med 2000; 27:800–806. 9 Suga K, Kume N, Nishigauchi K, Kawakami Y, Kawamura T, Matsumoto T, Matsunaga N. Three-dimensional surface display of dynamic pulmonary xenon-133 SPECT in patients with obstructive lung disease. J Nucl Med 1998; 39:889–893. 10 Travaline JM, Maurer AH, Charkes D, Urbain JL, Furukawa S, Criner G. Quantitation of regional ventilation during the washout phase of lung scintigraphy. Measurement in patients with severe COPD before and after bilateral lung volume reduction surgery. Chest 2000; 118:721–727. 11 Gelb AF, Zamel N. Unsuspected pseudophysiologic emphysema in chronic persistent asthma. Am J Respir Crit Care Med 2000; 162: 1778–1782.

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12 Sakai N, Mishima M, Nishimura K, Itoh H, Kuno K. An automated method to assess the distribution of low attenuation areas on chest CT scans in chronic pulmonary emphysema patients. Chest 1994; 106:1319–1325. 13 Kinsella M, Muller N, Abboud RT, Morrison NJ, dyBuncio A. Quantitation of emphysema by computed tomography using a ‘density mask’ program and correlation with pulmonary function tests. Chest 1990; 97:315–321. 14 Sandek K, Bratel T, Lagerstrand L, Rosell H. Relationship between lung function, ventilation–perfusion inequality and extent of emphysema as assessed by high-resolution computed tomography. Respir Med 2002; 96:934–943. 15 Newman KB, Lynch DA, Newman LS, Ellegood D, Newell Jr JD. Quantitative computed tomography detects air trapping due to asthma. Chest 1994; 106:105–109. 16 Park KJ, Bergin CJ, Clausen JL. Quantitative of emphysema with threedimensional CT densitometry: comparison with two-dimensional analysis, visual emphysema scores, and pulmonary function test results. Radiology 1999; 211:541–547. 17 Gevenois PA, De Vuyst P, de Maertelaer V, Zanen J, Jacobovitz D, Cosio MG, Yernault JC. Comparison of computed density and microscopic morphometry in pulmonary emphysema. Am J Respir Crit Care Med 1996; 154:187–192. 18 Thurlbeck WM, Muller NL. Emphysema: definition, imaging, and quantification. Am J Roentgenol 1994; 163:1017–1025. 19 Fletcher CM. The clinical diagnosis of pulmonary emphysema: an experimental study. Proc R Soc Med 1952; 45:577–584. 20 Zierler KL. Equations for measuring blood flow by external monitoring of radioisotopes. Circ Res 1965; 16:309–321. 21 American Thoracic Society. Single breath carbon monoxide diffusing capacity (transfer factor). Am Crit Care Med 1995; 152:2185–2198. 22 Cotton DJ, Soparkar GR. Diffusing capacity in the clinical assessment of chronic airflow limitation. Med Clin North Am 1996; 80:549–564. 23 Graham BL, Mink JT, Cotton DJ. Effects of increasing carboxyhemoglobin on the single breath carbon monoxide diffusing capacity. Am J Respir Crit Care Med 2002; 165:1504–1510. 24 Alpert NM, McKusick KA, Correia JA, Shea W, Brownell GL, Potsaid MS. Initial assessment of a simple functional image of ventilation. J Nucl Med 1975; 17:89–92. 25 Bunow B, Line BR, Horton MR, Weiss GH. Regional ventilatory clearance by xenon scintigraphy: A critical evaluation of two estimation procedures. J Nucl Med 1979; 20:703–710.

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Original article

In-vivo visualization of radiation-induced apoptosis using 125 I-annexin V Hiroshige Watanabea, Yuji Murataa, Masahiko Miuraa, Masatoshi Hasegawab, Tadafumi Kawamotoc and Hitoshi Shibuyaa Background As apoptosis occurs in tumors within a short time after irradiation, the detection of the frequency of apoptosis may be useful as an indicator of the effect of treatment. For the evaluation of apoptosis under these conditions, tissue extraction from patients is indispensable. Aim To develop a noninvasive imaging technique to measure and monitor apoptosis in tumor cells caused by X-irradiation using 125I-radiolabeled annexin V. Methods The tumors used were human ependymoblastomas, which were transplanted into nude mice. The tumors were irradiated at 2, 5 or 10 Gy. 125I-annexin V was administered intravenously 6 h after irradiation. In the 5 Gy irradiation group, the isotope was injected at various time intervals (3, 6 and 12 h) after irradiation. Three hours after the injection, the mice were sacrificed, the tumors were quickly removed and frozen sections were prepared at 6 and 40 lm thickness using a cryomicrotome. In autoradiographic imaging, the tumor-to-muscle ratios were compared in the respective irradiated groups. In addition, apoptosis detection by the in-situ end-labeling (Klenow) assay was conducted on the same sections. The number of Klenow-positive cells was counted in 100 ¾ fields for each section.

the control group (P < 0.05). Although immunohistochemical staining revealed a peak apoptosis frequency in the 5 Gy irradiated group, autoradiography revealed a peak in the group receiving a lower dose than 5 Gy. When the time from irradiation to annexin injection was varied, both imaging methods showed a peak apoptosis frequency in the group receiving the injection 6 h after irradiation. Conclusion It is possible to predict the effect of treatment in cancer in a noninvasive manner by apoptosis imaging in vivo after radiotherapy. Nucl Med Commun 27:81–89

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:81–89 Keywords: human tumor, apoptosis, radiotherapy

125

I-annexin V, in vivo, radiation-induced

a Department of Radiology, Faculty of Medicine, Tokyo Medical and Dental University, Tokyo, bDepartment of Radiology and Radiation Oncology, Gunma University, Gunma and cRadioisotope Research Institute, Tsurumi University School of Dental Medicine, Yokohama, Japan.

Correspondence to Hiroshige Watanabe MD, Department of Radiology, Faculty of Medicine, Tokyo Medical and Dental University, 5–45, 1-chome, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan. Tel: + 81-3-5803-5311; fax: + 81-3-5803-0147; e-mail: [email protected]

Results Both autoradiography and immunohistochemical staining showed a significantly higher frequency of apoptosis in the neoplasms in all irradiated groups than in

Sponsorship: This research was supported by a grant (14570840) from the Japan Society for the Promotion of Science.

Introduction

clinical information for the management of such neoplasms.

Apoptosis, as distinct from necrosis, is an active suicide process of cells. It is triggered by various stimuli [1–5], including radiotherapy for cancer [6–9]. It is believed that the curative effect of cancer therapy can be enhanced in the presence of enhanced apoptosis. This assumption, however, is not always valid, especially in the case of solid tumors, in which the frequency of apoptosis is very low. On the other hand, in neoplasms with a high apoptotic activity, such as malignant lymphomas, undifferentiated tumors or tumors with a high nuclear/ cytoplasmic (N/C) ratio, apoptosis is enhanced within a short time after irradiation, and the frequency of apoptosis may be a good indicator of the effect of treatment. The extent and time at which apoptosis occurs may be important for obtaining potentially useful

Received 1 June 2005 Accepted 4 October 2005

For the evaluation of apoptosis under these conditions, the extraction of tissue samples from patients is indispensable. Biopsy is highly invasive, however, and is often associated with sampling errors. Therefore, various imaging techniques have been employed as noninvasive strategies to detect apoptosis. One of these, namely radionuclide imaging using radiolabeled annexin V, appears to be promising [10–13]. One of the earliest events in programmed cell death is the externalization of phosphatidylserine, a membrane phospholipid that is normally restricted to the inner leaflet of the lipid bilayer. Annexin V, an endogenous human protein

c 2006 Lippincott Williams & Wilkins 0143-3636

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82 Nuclear Medicine Communications 2006, Vol 27 No 1

with a high affinity for membrane-bound phosphatidylserine [14–16], can be used both in vitro and in vivo to detect apoptosis even before the appearance of the other welldescribed morphologic and nuclear changes associated with programmed cell death. Therefore, radiolabeled annexin V may be useful for evaluating the efficacy of therapy and disease progression or regression.

with sterile food and sterile water. The protocol for this animal experiment was approved by the Institutional Animal Care and Use Committee of Tokyo Medical and Dental University, and the experiment was carried out in accordance with the Guidelines for Animal Experimentation of the Tokyo Medical and Dental University. Tumors

The purpose of this study was to develop a noninvasive imaging technique for clinical application to measure and monitor programmed cell death in tumor cells induced by tumor irradiation, using 125I-radiolabeled annexin V. As far as we know, no other reports on imaging after the irradiation of neoplasms have been described. The tumor examined in this study was ependymoblastoma, which is composed of undifferentiated cells and is known to be highly radiosensitive.

The tumor examined in this study was a human ependymoblastoma which is transplantable into nude mice [17]. The tumor specimen was broken up into 2–3 mm pieces, and a few of these pieces were subcutaneously transplanted to the right thigh of nude mice. The tumor diameter was measured every 3 days after transplantation and, when the calculated tumor volume had reached 200– 250 mm3, the experiment was begun. The tumor volume (V) was calculated using the following formula: V = 1/2a2 b (a is the short diameter and b is the long diameter).

Materials and methods Animals

125

Nude mice (BALB/cA Jcl-nu), 7–10 weeks of age, were used. The animals were housed in autoclaved cages, in groups of three to five animals per cage, and provided

125

I-annexin V

I-annexin V was synthesized by the Iodo-Gen method (Daiichi-Radioisotope Laboratory, Tokyo, Japan), as reported by Lahorte et al. [18]. The radiochemical purity

Fig. 1

Annexin V imaging of ependymoblastomas (arrows) at different doses of irradiation (2, 5 and 10 Gy). Significantly greater accumulation was observed in all of the irradiated groups (2, 5 and 10 Gy) than in the control group. Regions of interest were set in the neoplasm and the surrounding thigh muscle, and the tumor-to-muscle ratios (TMRs) were calculated in the respective groups.

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In-vivo visualization of apoptosis Watanabe et al. 83

of the synthesized 125I-annexin V, as measured by high performance liquid chromatography, was better than 95%.

Fig. 2

Uptake of 125I-annexin V 4.00

Irradiation

Hasegawa et al. [17], who used the same ependymoblastoma tumor cell line in their study, reported that radiation-induced apoptosis increased with increasing radiation dose until a plateau was reached. Therefore, the radiation doses selected in the present study were 2, 5 and 10 Gy.

3.50 ∗ 3.00 Tum or-to-muscle ratios

The X-irradiation to neoplasms was performed using a Stabliipan 2 (Siemens, Munich, Germany): 250 kV; 1 mm Cu filter; 15 mA; 0.83 Gy/min; distance, 50 cm. The unanesthetized mice were restrained, and only the tumors in the thighs were irradiated; the other body regions were shielded with a lead plate. The radiation was administered in single doses of 2, 5 or 10 Gy.

∗ ∗∗

2.50

2.00 2.84

2.37

1.50

1.00

1.42

0.50

0.00 125

2.61

Irradiation dose 2 Gy 5 Gy

I-annexin V (10–15 mCi, 50–100 mg/kg protein per animal) was administered intravenously into the tail vein 6 h after irradiation. To confirm the peak time of occurrence of apoptosis after irradiation, the isotope was injected at various time intervals (3, 6 and 12 h) after irradiation in the 5 Gy irradiated group. Three mice were prepared for each group. Three mice were also injected with 125I-annexin V and not irradiated; these were used as the control group.

Tumor-to-muscle ratios (TMRs) determined in annexin V images of the ependymoblastomas at various irradiation doses (2, 5 and 10 Gy). The TMRs were significantly higher in all of the irradiated groups than in the control group. Although the differences were not significant, the TMRs were greater at lower doses of irradiation. *P < 0.05 in comparison with the results in the control. **P < 0.01 in comparison with the results in the control.

Frozen sections

chemical staining and at 40 mm thickness for autoradiography.

Three hours after intravenous injection of radiolabeled annexin V, the mice were sacrificed and the tumors and surrounding tissues were quickly removed. All efforts were made to minimize the animals’ suffering. Kawamoto’s tape method [19–21] was used to prepare the frozen sections. The sections were prepared from unfixed and undecalcified frozen specimens using an adhesion film as support material. The same sections were used for autoradiography and immunohistochemical staining, and, when needed, the adhesion film sections were preserved by transcribing them on to glass slides. Sections were cut along the maximum diameter of the neoplasm, and were prepared for autoradiographic and immunohistochemical analyses one by one. The tumor samples were embedded in a 5% carboxymethyl cellulose (CMC) solution, frozen in hexane/dry ice and stored at – 801C. The CMC block was placed on the sample stage of a microtome (LKB PMV 2258, Bromma, Sweden) in a cryostat kept at – 201C and trimmed. The sectioning surface was covered with a sheet of Saran Wrap coated with a synthetic rubber adhesive (Azia Genshi, Gifuken, Japan), and frozen sections were cut at 6 mm thickness for immunohisto-

Control

10 Gy

Radionuclide imaging

For macroscopic autoradiography, in order to prevent chemographic artifacts, the freeze-dried sections, 40 mm thick, were covered with a 4 mm sheet of plastic film (Diafoil, Mitsubishi Plastic Co., Tokyo, Japan) and then pressed on to a Fuji imaging plate. The imaging plate is superior to film for quantitative evaluation. Densitometric analysis was performed using a BAS2000 system (Fuji Photo Co., Tokyo, Japan). In the annexin V imaging plate images, regions of interest (ROIs) were set up in the neoplasm and the surrounding thigh muscle. The tumor-to-muscle ratios (TMRs) for apoptosis were then calculated and compared. To measure the average intratumoral accumulation, ROIs were set by the method that did not contain the domains exhibiting a high luminosity, which are considered to originate from blood vessels within the tumor. Immunohistochemistry

The Klenow assay [22,23] was performed according to the manufacturer’s instructions (Trevigen, Gaithersburg, Maryland, USA). This is an in-situ end-labeling (ISEL)

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technique, the protocol of which is similar to that of the terminal deoxynucleotidyl transferase-mediated UTP nick end-labeling (TUNEL) assay, with only minor differences. The sections were fixed in 4% paraformaldehyde solution. After protease treatment, endogenous peroxidase was inhibited by incubation with hydrogen peroxide (2% in water, 5 min). Following 5 min of incubation with a labeling solution, the sections were incubated with a reaction mixture (containing the Klenow enzyme and biotinylated dNTP) for 1 h at 371C. The biotinylated nucleotides were detected using a streptavidin conjugate. The addition of insoluble colorless substrates (Trevigen Apoptotic Cell System Blue Label, TBL) to the treated sample resulted in the formation of an insoluble colored substrate in the regions of DNA fragmentation. The tissue sections were then counterstained to aid in morphologic characterization. Negative and positive controls were used instead of a reference area for the determination of the number of Klenow-positive cells. The number of Klenow-positive cells was counted by light microscopy in five randomly selected 100  fields, and the mean values were calculated.

Statistical analysis

Statistical analysis was performed for comparison of the TMR values in annexin V images between the irradiation groups and nonirradiated control group, using the unpaired Student’s t-test; P < 0.05 was considered to denote a significant difference. Similarly, statistical analysis was performed for comparison of the number of Klenow-positive cells per high-power field (100  ) in the immunohistochemically stained sections between the irradiation groups and the nonirradiated control group, using the unpaired Student’s t-test; P < 0.05 was considered to denote a significant difference.

Results Groups in which the radiation dose was varied (the time from irradiation to intravenous injection of annexin V was fixed at 6 h)

In annexin V imaging of ependymoblastomas, significant accumulation of the isotope was observed in the tumors in all irradiated groups (2, 5 and 10 Gy) when compared with that in the control group (Fig. 1). The TMR was significantly higher in all irradiated groups than in the control group (Fig. 2). Although no significant differences in the TMR values were observed between the three

Fig. 3

Klenow staining (200  ) of the same sections as in Figs. 1 and 2. The nuclei carrying fragmented DNA are darkly stained. Klenow-positive cells are hardly seen in the nonirradiated control group, whereas numerous Klenow-positive cells are observed in the irradiated groups. Scale bar, 100 mm.

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In-vivo visualization of apoptosis Watanabe et al. 85

irradiated groups, the TMR values were greater at lower doses of irradiation.

Fig. 4

Klenow-positive cells

Mean no of k lenow-positive cells HPF

100

∗

80

∗

Klenow staining was performed on the same specimens (Fig. 3). The number of Klenow-positive cells per highpower field (100  ) was higher in all irradiated groups than in the control group (P < 0.05) (Fig. 4); however, unlike annexin V imaging, the number of Klenow-positive cells increased with increasing radiation dose up to 5 Gy.

60 71.0 40

59.7

∗

20 27.5 2.75 0 Irradiation Dose Control

2 Gy

5 Gy

Groups in which the time from irradiation to intravenous injection of annexin V was varied (the radiation dose was fixed at 5 Gy)

In annexin V imaging of the neoplasms, the accumulation of the isotope was recognized as soon as 3 h after irradiation (Fig. 5). In the groups injected with the isotope 3 and 6 h after irradiation, the TMR value was significantly higher than that in the control group (P < 0.05) (Fig. 6). The TMR was highest in the group that received the injection 6 h after irradiation.

10 Gy

Mean number of Klenow-positive cells per high-power field (HPF) (100  ). The number of Klenow-positive cells at 9 h after irradiation increased with increasing radiation dose up to 5 Gy. *P < 0.05 in comparison with the results in the control.

Immunohistochemical staining of the same specimens also revealed that the number of Klenow-positive cells per high-power field was significantly higher in the irradiated groups than in the control group (P < 0.05) (Figs. 7 and 8). The number of Klenow-positive cells was

Fig. 5

Annexin V imaging of the ependymoblastomas (arrows) when the time from irradiation to intravenous annexin V injection was varied (3, 6 and 12 h). The accumulation of the isotope was recognized as soon as 3 h after irradiation.

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radiotherapy in the living body, using 125I-annexin V imaging. Ex-vivo imaging with a gamma camera or by positron emission tomography following intravenous injection of 124I/18F/99mTc-labeled annexin V might allow the detection and serial imaging of apoptosis occurring during cancer treatment.

Fig. 6

Uptake of 125I-annexin V 4.00

3.50

Tum or-to-muscle ratios

3.00

∗

2.50 2.00

∗

2.61

2.32

1.50

1.82 1.00

1.42 0.50

0.00 Time after X-ray irradiation Control

3h

6h

12 h

Tumor-to-muscle ratios (TMRs) in annexin V images of ependymoblastomas when the time from irradiation to intravenous annexin V injection was varied (3, 6 and 12 h). The TMRs were significantly higher in the groups receiving the isotope injection 3 and 6 h after irradiation compared with the values in the control group. TMRs were highest in the group receiving the isotope injection 6 h after irradiation. *P < 0.05 in comparison with the results in the control.

highest in the group that received the isotope injection 6 h after irradiation, similar to the findings in annexin V imaging. It is noteworthy that, for the same time from irradiation to intravenous injection, the immunohistochemical staining findings reflected the result of staining at the time of freezing of the specimen 3 h after intravenous injection, whereas the findings on annexin V imaging were a result of the accumulation of the isotope over 3 h after the injection. Therefore, the Klenowpositive cell number was thought to peak after the peak in isotope accumulation. An additional case

One neoplasm was only partially irradiated because the shield did not function properly at the time of the 2 Gy irradiation. Annexin V imaging of the neoplasm in which the irradiated regions could be differentiated from the nonirradiated regions revealed a clear difference in the incidence of apoptosis between the irradiated upper half and the nonirradiated lower half of the tumor (Fig. 9). These data have not been included in this report.

It is believed that the therapeutic efficacy of cancer treatment is significantly improved even with a slight enhancement in apoptosis; however, no clear conclusion has emerged regarding the correlation between the incidence of apoptosis after irradiation and the radiosensitivity of tumors. Some in-vitro studies have indicated that the incidence of apoptosis after irradiation is often correlated with the radiosensitivity of the tumor [24,25]. On the other hand, a negative correlation [26] and even the absence of correlation [27] between the two have also been reported. The correlation between the incidence of apoptosis and the treatment outcome is also not clear. This correlation may hold true for tumors originating from cells of the lymphoid system or the hematopoietic system, in which cell death often occurs by apoptosis. On the other hand, it may not be valid for solid tumors which show a very low incidence of apoptosis. In the case of solid neoplasms, it is thought that necrosis finally determines healing; therefore, to demonstrate that the incidence of apoptosis is correlated with the treatment outcome, it would be necessary to demonstrate that the frequency of apoptosis determines the frequency of necrosis in these tumors. In this study, a difference in the peak frequency of apoptosis was recognized between annexin V imaging and immunohistochemical staining when the radiation dose to the tumor was varied. For immunostaining, the frequency of apoptosis reached a peak at 5 Gy of irradiation. For lowdose irradiation, the dose–effect relationship was consistent with linearity. However, in the case of highly radiosensitive tumors, a plateau is reached immediately. The ependymoblastoma used for this study is a highly radiosensitive tumor. Moreover, in the experiment conducted by Hasegawa et al. [17] using the same tumor cell line, the frequency of apoptosis reached a plateau at 5 Gy, and no significant increase in the frequency of apoptosis was recognized at 10 Gy or higher doses of irradiation. In contrast, for annexin V imaging, the peak apoptosis frequency was observed at a radiation dose of less than 5 Gy. Unlike the Klenow assay, in which stained sections are examined, as annexin V is injected intravenously in vivo, it is necessary to also consider the hemodynamic influence.

Discussion This study demonstrates the possibility of detection of the frequency of apoptosis in a neoplasm following

Necrosis occurring as a result of the physical effects of irradiation is associated with cytolysis. The cellular

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In-vivo visualization of apoptosis Watanabe et al. 87

Fig. 7

Klenow staining (200  ) of the same sections as shown in Figs. 5 and 6. Numerous Klenow-positive cells are already seen in cases receiving the isotope injection 3 h after irradiation. Note that the neoplasm specimens were frozen 3 h after the injection. Scale bar, 100 mm.

contents begin to flow, triggering an inflammatory reaction in the surrounding surviving cells. On the other hand, cells undergoing apoptosis show nuclear condensation, and the whole cell, including the nucleus, undergoes fragmentation to become the so-called apoptotic body. As continuity of the cell membrane is maintained in the case of apoptosis, an apoptotic body is wrapped in a membrane and is therefore removed by macrophages, eliciting little inflammatory reaction in the surrounding tissues. As mentioned above, although blood flow reduction around cells is hardly observed in apoptosis, it is a common feature in necrosis. Although annexin V binds to both necrotic and apoptotic cells, intravenously injected annexin V cannot bind to necrotic cells because of the reduced blood supply around them. This may be the reason for the decrease in the accumulation of intravenously injected annexin V with increasing radiation dose. In addition, the time of detection of apoptosis was different between annexin V imaging and the ISEL assay. In the early stage of apoptosis, phosphatidylserine is

translocated from the inner side of the plasma membrane to the outer layer. As this change takes place at an earlier stage than DNA fragmentation, imaging using annexin V, which binds to phosphatidylserine, may allow apoptosis to be detected at an earlier stage than with the ISEL assay. In this study, the highest frequency of apoptosis was detected, by both annexin V imaging and immunohistochemical staining, in the group that received the isotope injection 6 h after irradiation (Figs. 6 and 8). It must be remembered, however, that immunohistochemical staining detected the apoptosis that had occurred at the time of freezing of the specimens 3 h after intravenous injection. In general, the majority of radiopharmaceuticals are taken up first-pass after intravenous injection, so that there may be a difference of 2 h or more in the time of apoptosis detection between annexin V imaging and immunohistochemical staining. Similarly, in the groups that received different radiation doses, the time from irradiation to apoptosis detection was different between annexin V imaging and ISEL assay, even in the same specimens. This may have also affected the difference in the peak frequency of apoptosis when the radiation dose was varied.

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Fig. 8 Klenow-positive cells 100

∗

Mean no of klenow-positive cells HPF

80

∗

60

There are some subjects for future research in this field that may be useful for clinical application. First, the feasibility and usefulness of apoptosis imaging must be evaluated in a variety of neoplasms. It has been suggested that ependymoblastomas, used in this study, show a higher frequency of apoptosis following irradiation than many other neoplasms [17,28–31]. Therefore, the detection of apoptosis by imaging must be confirmed in other neoplasms.

∗ 71.0

40 56.3 47.3

20

Second, the possibility of synergistic effects of other modalities of therapy, including chemotherapy, endocrine therapy and gene therapy, combined with radiotherapy must be considered. Radiotherapy is rarely used independently in clinical practice; therefore, evaluating the apoptosis frequency after radiation monotherapy may give an incorrect indication of the treatment outcome and subsequent treatment planning for combined therapy.

2.8

0 Time after X-ray irradiation Control

3h

6h

12 h

Mean number of Klenow-positive cells per high-power field (HPF) (100  ). The number of Klenow-positive cells was highest in the group receiving the isotope injection 6 h after irradiation. As apoptosis that had occurred at the time of specimen freezing, 3 h after injection, was assessed, the number of Klenow-positive cells was expected to peak after the peak in isotope accumulation. *P < 0.05 in comparison with the results in the control.

Fig. 9

Annexin V imaging of a neoplasm that was only partially irradiated because the shield did not function properly at the time of 2 Gy of irradiation. In the tumor (arrowheads), the boundary between the irradiated area and the nonirradiated area is represented by a line joining the two arrows together. A difference in the extent of accumulation between the irradiated upper half and the nonirradiated lower half is apparent. The data from this neoplasm are not included in this report.

Third, there is the possibility to predict the efficacy of therapy by the evaluation of spontaneous apoptosis before medical treatment; however, as the frequency of apoptosis before treatment is much lower than that after treatment, its evaluation may prove to be difficult. Fourth, in this study, after intravenous injection of the tracer, the neoplasm was enucleated and autoradiographic imaging was conducted. However, when ex-vivo imaging is conducted by positron emission tomography or gamma camera imaging following injection of 99mTc/18F/124Ilabeled annexin V in the clinical setting, a deterioration in image quality might be expected, because of the influence of absorption and scattering by the surrounding tissues and the use of a collimator. In addition, discrimination from physiological annexin V accumulation may become a problem. Therefore, it is expected that the visual confirmation of a change in the degree of accumulation in the tumor by this technique might be more difficult than by autoradiography. On the other hand, semiquantitative analyses, such as measurement of the TMR or the standardized uptake value (SUV), might allow even minute changes to be recognized. In conclusion, we imaged apoptosis induction after low-dose irradiation in human ependymoblastomas transplanted into nude mice by autoradiography using 125 I-radiolabeled annexin V. Both annexin V imaging and immunohistochemical staining revealed a higher apoptosis rate in neoplasms following irradiation at all of the doses examined in this study compared with that in the control group. It was shown that it is possible to provide noninvasive and continuous visualization of apoptosis by scintigraphy in a neoplasm after irradiation. This technique might become useful in the future for evaluating the therapeutic effect of not only radiotherapy, but also chemotherapy, endocrine therapy and gene therapy.

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In-vivo visualization of apoptosis Watanabe et al. 89

Acknowledgement

16

We wish to express our gratitude to Winn Aung (National Institute of Radiological Sciences, Japan) for his valuable contribution to this study.

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Original article

Review of monitoring free muscle flap transfers in reconstructive surgery: role of 99mTc sestamibi scintigraphy Husamettin Topa, Ali Sarikayab, A. Cemal Aygita, Erol Benliera and Medeni Kiyaka Background Free tissue transfer is a method of moving any tissue from a donor area of the body to a recipient site and re-attaching the arteries and veins to the blood vessels at the recipient site by microvascular surgery. Improved microsurgical techniques have resulted in a high percentage of successful free tissue transfers. Post-operative monitoring of tissue viability can detect early problems in free tissue transfer which may allow early intervention and salvage. Although many flap monitoring methods have been described, there is still no consensus on which of these techniques will become the standard accepted method for monitoring free muscle flaps. Objective In present study, we investigated the use of 99m Tc sestamibi scintigraphy in determining free muscle flap viability and complications, and also in directing treatment. Methods Thirteen patients were examined prospectively during the post-operative period after free tissue transfer for foot defects. The cause of the defect was diabetic foot ulcer in 10 patients, dermatofibrosarcoma in one patient, squamous cell carcinoma in one patient and gunshot wound in one patient. Foot defect covering was carried out with a free latissimus dorsi muscle flap and skin graft (n = 12) and a free gracilis muscle flap (n = 1). All patients were examined with a monitoring system that consisted of visual inspection, hand-held Doppler ultrasonography and scintigraphic examinations. Scintigraphic imaging of all cases was performed routinely within the first 48 h postoperatively, and also on days 10 and 91 in two patients. Results There were four flap failures during the study. One of these patients had viable findings upon visual inspection and no evidence of vascular compromise on Doppler at the first examination. In the other patient, visual inspection of

Introduction Free tissue transfer is a method of transferring any tissue from a donor site to a recipient site with re-attaching arteries and veins to the blood vessels at the recipient site by microvascular surgery. Over the past three decades, free tissue transfers have been used successfully for reconstruction of complex tissue defects after trauma or oncological ablative surgery. Success rates have improved

the flap showed that it was ischaemic in one region, but there was no vascular compromise on Doppler examination. Scintigraphic images of each of these patients showed a partial hypoperfused area in the flap region. Later, these two flaps showed positive clinical indications of hypoperfusion (colour of muscle and appearance of skin graft) and Doppler abnormalities. The remaining two patients had non-viable scintigraphic images as well as positive clinical indicators of hypoperfusion and evidence of vascular compromise on Doppler. Nine patients each had a viable flap. In these patients, all three examination tools demonstrated that the flaps were totally viable and there were no vascular complications. Conclusion According to the results of this study, 99mTc sestamibi scintigraphy appears to be a feasible and promising method in the evaluation of free muscle flap viability and complications. On the other hand, to demonstrate any impact on management or patient outcome, further evaluation of 99mTc sestamibi imaging, including comparative studies with different established methods in a larger patient population, is highly recommended. Nucl Med Commun 27:91–98

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:91–98 Keywords: free muscle flap, viability, scintigraphy Departments of aPlastic, Reconstructive and Aesthetic Surgery and bNuclear Medicine, Trakya University, Edirne, Turkey. Correspondence to Dr Husamettin Top, 1. Murat Mah., 5. Sok., Celik20 Apt. D:6, 22030, Edirne, Turkey. Tel: + 0090 284 21 31278; e-mail: [email protected] Received 10 May 2005 Revised 2 August 2005 Accepted 30 August 2005

steadily to 95% or more in most series [1–4]. Flap failures are caused by thrombosis of the anastomosed vessels or by disturbances in the microcirculation of the transferred tissue [5]. These vascular complications usually occur during the first post-operative days [6]. Through early intervention, the flap can be salvaged in 33–57% of cases [7]. Here, the time interval between detecting vascular complications or disturbances in the microcirculation of

c 2006 Lippincott Williams & Wilkins 0143-3636

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92 Nuclear Medicine Communications 2006, Vol 27 No 1

the transferred tissue and re-establishing vascular patency or microvascular perfusion is of particular importance for flap viability. Moreover, late complications, such as vascular and infectious, may lead to total or partial flap necrosis. Post-operative monitoring methods are therefore required for the early detection of complications in free flaps. For post-operative monitoring of free flaps, various strategies have been developed to address the issue of detecting post-operative flap complications in an effort to permit intervention and flap salvage [8]. The most widely used clinical monitoring method of tissue perfusion is visual inspection [9]. In the vast majority of cases an experienced clinician is able to ascertain the status of the circulation of the flap simply by its clinical characteristics. But this method is not reliable in buried and muscle flaps. Concomitantly, more objective flap monitoring methods have developed. These include Doppler ultrasonography [10–13], electrical impedance plethysmography [14], intravenous fluorescein [15], laser Doppler [16], photoplethysmography [17,18], pulse oximetry [19], temperature monitoring [20], tissue pH [21], transcutaneous oxygen monitoring [22,23], implantable Doppler monitoring [4], and muscle contractility testing [24]. These methods range in complexity, invasiveness and efficacy. But there is still no consensus on which of abovementioned techniques will become the standard accepted method for monitoring free muscle flaps [3,4]. Another choice for monitoring tissue viability is scintigraphy. 99mTc sestamibi has been employed in the evaluation of skeletal muscle perfusion in patients with periphery artery disease and other muscle disease [25– 28]. In previous preliminary studies with a few cases, we demonstrated that 99mTc sestamibi muscle scintigraphy is appropriate for monitoring free muscle flaps [29,30]. In this prospective study, we evaluated the usefulness of a 99mTc sestamibi scan in detecting free muscle flap viability and complications and also in directing treatment.

Patients and methods Thirteen patients (11 male, two female) were examined prospectively during the post-operative period after free tissue transfer for foot defects (Table 1). The patients ranged in age from 22 to 73 years, with an average age of 59 years. The cause of the defect was diabetic foot ulcer in 10 patients, dermatofibrosarcoma in one patient, squamous cell carcinoma in one patient, and gunshot wound in one patient. Vascular reconstruction was performed in eight patients. Covering of the foot defect was carried out with a free latissimus dorsi muscle flap and skin graft (n = 12) and a free gracilis muscle flap (n = 1). After free flap transfer, all patients were administered 2500 IU heparin four times daily, intrave-

nously, for 1 week. Post-operative evaluation consisted of visual inspection, hand-held Doppler ultrasonography and scintigraphic examinations. Visual inspections were conducted every 2 h for the first 24 h, every 4 h on the second and third days, and twice daily until discharge. Visual inspection consisted of reporting the colour of the muscle and the appearance of the skin graft. Doppler examination was performed routinely on the first three postoperative days. The findings of the visual inspection did not influence the timing of the scintigraphic imaging, which was performed routinely as early as possible in the first 48 h. The scintigraphic examination was performed in one patient on post-operative days 1 and 10 (patient 5) and in one patient on post-operative days 1 and 91 (patient 2). Scintigraphic images were obtained within 60 min after intravenous injection of 555 MBq 99mTc sestamibi in different positions. All studies were performed using a large-field-of-view gamma camera equipped with a low energy, parallel hole, high resolution collimator, and a 20% energy window centred at 140 keV. Images were acquired on a dedicated computer system in a 128  128 pixel matrix until 500 000 counts/view were obtained or for 5 min. Images were interpreted visually by two observers. Flap tissue was considered viable by visual analysis if it showed good uptake in the flap region compared with activity of adjacent soft tissue. Scintigraphic images were interpreted as having either a total or partial hypoperfused area in the flap tissue if there was decreased uptake in the flap region compared with the activity of adjacent soft tissue or the viable areas of flap tissue. Mean follow-up time was 26.07 months (range, 1–55 months).

Results Outcome analysis of the 13 patients is shown in Table 1. There were four flap failures (30.7%) during the study. One of these patients (patient 4) had viable findings upon visual inspection and no evidence of vascular compromise on Doppler in first examination (postoperative day 1). In the other patient (patient 2), visual inspection of the flap demonstrated that it was ischaemic in one region, but there was no vascular compromise on Doppler examination (post-operative day 90). Scintigraphic scan images of these patients showed partial hypoperfused area in the flap region. Later, these two flaps showed positive clinical indicators of hypoperfusion (colour of the muscle and appearance of the skin graft) and Doppler abnormalities on post-operative day 91 in patient 2, and post-operative day 2 in patient 4. The remaining two patients had non-viable scintigraphic images as well as positive clinical indicators of hypoperfusion and evidence of vascular compromise on Doppler. Eight patients each had a viable flap. In these patients, all three examination tools demonstrated that the flaps were totally viable and there was no vascular complication. Two of these eight patients died due to myocardial infarctus

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Table 1

Summary of patients

Patient Age (years) number sex

Cause

Lesion site

Flap

Imaging time (days)

99m

Tc sestamibi scan

Flap viability

Muscle flap colour normal (reddish-pink) Skin graft adherence good Muscle flap colour normal Skin graft adherence good (day 1 to discharge) Muscle flap colour bluishpurple (day 90) Muscle flap colour pale Skin graft adherence bad

+

1

Exitus due to MI at the end of post-operative first month

–

5

Near total necrosis at third month/below knee amputation

–

5

Muscle flap colour normal Skin graft adherence good (day 1)

–

7

Total flap necrosis, tumour recurrence/below knee amputation Total flap necrosis/reconstructed with cross-leg flap, healing

+

9

+

19

+

22

+

32

+

38

Distal flap necrosis/secondary healing Tumour recurrence, pulmonary metastasis/exitus No/Healing

+

42

No/Healing

+

40

No/Healing

+

45

No/Healing

–

53

Total flap necrosis/Secondary healing

1

64 F

DF

L foot

LD

1

Good uptake in all areas of flap region

–

2

56 M

DF

R foot

LD

1 and 90

Good uptake in all areas of flap region (day 1)

91

3

72 F

DFS

L ankle

LD

2

4

60 M

DF

R heel

LD

1

Decreased uptake area in the flap region (day 90) Decreased uptake in all areas of the flap region Decreased uptake area in the flap region

1

2

5

66 M

DF

R foot

LD

1 and 10

Good uptake in all areas of flap region (day 1 and 10)

–

6

58 M

DF

L foot

G

2

–

7

62 M

DF

L foot

LD

1

8

40 M

SCC

L heel

LD

1

9

64 M

DF

L foot

LD

2

10

73 M

DF

R foot

LD

1

11

71 M

DF

R foot

LD

2

12

60 F

DA

L foot

LD

2

13

22 M

G

L foot

LD

1

Good uptake in all areas of flap region Decreased uptake area in the flap region Good uptake in all areas of flap region Good uptake in all areas of flap region Good uptake in all areas of flap region Good uptake in all areas of flap region Good uptake in all areas of flap region Decreased uptake in all areas of the flap region

– – – – – – 1

Muscle flap colour pale Skin graft adherence bad (day 2) Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour normal Skin graft adherence good Muscle flap colour pale Skin graft adherence bad

Follow-up (months)

Complication/ conclusion

Lack of adaptation to recipient area and total graft necrosis due to infection/Healing No/Healing

DF, diabetic foot; L, left; R, right; LD, latissimus dorsi muscle free flap; DFS, dermatofibrosarcoma; MI, myocardial infarction; G, gracilis muscle free flap; SCC, squamous cell carcinoma; G, gunshot wound.

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Tc sestamibi scintigraphy in monitoring free muscle flap Top et al. 93

Visual inspection

99m

Doppler abnormality detected time (days)

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and metastasis. There was one distal flap necrosis, and healed flap with secondary intention (patient 7). Distal flap necrosis was detected initially by scintigraphic examination, and then visual findings were noted. There was no evidence of vascular compromise on Doppler examination in this patient.

Fig. 1

Three sample cases representing a non-viable free flap transfer (patient 4), a viable free flap transfer in a patient who had flap adaptation problem due to ongoing necrosis and infection (patient 5), and a viable free flap transfer (patient 9) are presented below. Patient 4

A 60-year-old diabetic male patient was referred to our clinic with an unstable scar with multiple fistula openings on his right heel. On physical examination, there was an 8 cm depressed scar on inferior surface and fistula openings on the medial and lateral surfaces of the right heel. The popliteal, posterior tibial and dorsalis pedis pulses were non-palpable bilaterally. After debridement, a free latissimus muscle flap was transferred and followed by meshed split-thickness skin grafting (Fig. 1). 99mTc sestamibi scintigraphy was performed on the first postoperative day. Scintigraphic images showed a decreased uptake area in muscle flap region (Fig. 2) while other examination methods demonstrated normal findings. Because these results suggested that there was hypoperfused area in the flap tissue, streptokinase therapy was started. The following day the Doppler tone was changed and the colour of muscle tissue and the appearance of the skin graft were worse. Revision showed total thrombosis of both vessels, which was removed completely before reanastomosis was performed. But this procedure was a failure. Necrotic tissues were removed and the tissue defect was reconstructed with a cross-leg flap.

Free latissimus muscle flap transfer was performed and covered with split-thickness graft.

Fig. 2

Patient 5

A 66-year-old diabetic male patient whose third and fourth right toes had been amputated 1 year before was referred to our clinic with necrotic tissues on his right foot. On physical examination, there were necrotic tissues on the anterior metatarsal area, the second and fifth toes and the plantar area of his right foot. The popliteal, posterior tibial and dorsalis pedis pulses were palpable bilaterally. Transmetatarsal amputation and debridement of necrotic tissues were performed. To cover the remaining defect, we decided on a free latissimus dorsi muscle flap plus meshed split-thickness skin graft. Scintigraphic images, Doppler examination and visual inspection findings demonstrated that flap tissue was totally viable on the first post-operative day. However, the flap did not adhere to the underlying recipient area in the following days. Tissue necrosis and infection were evident below the flap tissue and skin graft became totally necrotic. Ten days after surgery, as flap failure was indicated, we performed a scintigraphic examination. The

99m Tc sestamibi scan images of patient 4, obtained at the first postoperative day, showing focally decreased uptake (white arrowheads) at the flap region (black arrowheads). C: cranial.

scintigraphic images showed the the flap tissue was totally perfused (Fig. 3). After intensive wound care with dressings and antibiotic treatment, necrotic tissues were removed. The flap was re-adapted to the recipient area and resurfaced with a meshed split-thickness skin graft. Follow-up to 9 months after surgery showed healing without a problem (Fig. 4). Patient 9

A 64-year-old diabetic male patient was referred to our clinic with necrotic tissues on his left foot. On physical examination, the fifth toe was totally necrotic. There

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99m

Fig. 3

Tc sestamibi scintigraphy in monitoring free muscle flap Top et al. 95

Fig. 5

99m Tc sestamibi scan images of patient 5 obtained 10 days after surgery show good uptake in all the areas of the flap region (arrowheads). C: cranial.

Fig. 4

99m Tc sestamibi scan images of patient 9, obtained at the first postoperative day, show good uptake in all the areas of the flap region (arrowheads). C: cranial.

were necrotic tissues on the anterior metatarsal area and the lateral plantar area of his left foot. The popliteal pulses were palpable, but posterior tibial and dorsalis pedis pulses were not palpable bilaterally. After debridement, a free latissimus muscle flap was transferred and followed by meshed split-thickness skin grafting. 99mTc sestamibi scintigraphy was performed on the first postoperative day. Scintigraphic images showed an area in muscle flap region with good uptake (Fig. 5). Doppler ultrasonography indicated that both anastomoses were patent and visual inspection showed that the flap tissue was viable. Follow-up to 38 months after surgery showed healing without a problem (Fig. 6).

Discussion

Patient 5 could walk with the salvaged foot.

The first three decades of microsurgery have shown a steady increase in flap success rate. Enhanced flap survival was initially attributed to the accumulation of surgical experience [31,32]. Later, the development of more reliable flaps further increased flap survival. These flaps have larger donor sites and reliable vascular pedicles that facilitate the technical aspects of the microvascular anastomosis [33]. However, regardless of the experience

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Fig. 6

Final result. The patient could walk with the salvaged foot, but did not want a debulking procedure.

of the surgeon or the reliability of the donor site, primary and secondary thrombosis is an unavoidable potential complication. The first step in the salvage of a failing free flap is the rapid identification of microvascular problems. This rapidity is accomplished by various monitoring methods. These methods can be divided into two groups: those for direct imaging of blood vessels and those for direct and indirect imaging of tissue perfusion [8]. Clinical examination is still the most reliable method for monitoring free flaps for most microsurgeons rather than technical monitoring systems [34,35]. The experienced observer can evaluate the status of the circulation of a free flap by its clinical characteristics, such as skin colour, temperature and capillary refill, and by pinprick testing. But these clinical examination methods are not reliable in free muscle flaps. Although evaluation of muscle colour is a valuable monitoring choice, it is subjective. For this reason clinical examination should not be used as a single method for monitoring free muscle flaps. Electrical impedance plethysmography, laser Doppler, photoplethysmography, pulse oximetry, temperature monitoring, tissue pH, and transcutaneous oxygen monitoring require a portion of the flap to be exposed and/or provide

a measure of flap perfusion indirectly by assessing skin temperature or perfusion [4]. Muscle contractility is not a reliable test for showing impaired flap perfusion [24]. Doppler monitoring is a useful method for evaluating the patency of arterial and venous flow but it is generally unreliable in detecting any alteration in the microcirculation [36]. Therefore, it should not be used as a single tool for evaluation of free flap viability. Implantable Doppler is an invasive technique, requiring meticulous positioning of the probe, with the potential disadvantage that any change in probe position may lead to loss of Doppler signal and the requirement for a false-positive reexploration [34]. Recently, a few radionuclide imaging techniques have been used to assess post-operative flap viability [29,30,37]. Monitoring of free flap viability has been performed by Smith et al. using 2-[18F]fluoro-2-deoxy-Dglucose (FDG) positron emission tomography (PET) scanning [37]. Although FDG PET is unique in that it assesses viability non-invasively, it is expensive and has only limited availability. Recently, 99mTc sestamibi has been found to be an appropriate radionuclide agent for non-invasive imaging of perfusion of the extremities. In our previous studies, 99mTc sestamibi scintigraphy was used for both assessing extremity perfusion/viability and evaluating the response to treatment in many different diseases [26–28]. Our group was the first to describe 99m Tc sestamibi imaging as a promising technique in monitoring free muscle flap in a few cases [25,29]. An ideal monitoring method can evaluate both microvascular anastomoses and tissue perfusion. Because there is no single method that monitors the patency of microvascular anastomoses and evaluates the perfusion and microcirculation of flap tissue, the best monitoring system should include a combination of different monitoring techniques at the same time [38]. For this reason, we used Doppler monitoring for direct imaging of blood vessels, and clinical observation and scintigraphy for direct and indirect imaging of tissue perfusion. There are several advantages of scintigraphy in our monitoring system. First, the above-mentioned techniques are not reliable for monitoring free muscle flaps. Although Doppler monitoring and clinical examination (examination of how the skin graft has taken or the colour of the muscle) are useful methods for evaluating the viability of free muscle flaps, both are unreliable in monitoring the perfusion of a whole flap. In addition, evaluation of the colour of muscle is subjective and skin graft take may be affected by various factors such as infection and immobilization. Second, free tissue failure is not an all-or-none phenomenon. In other words, there is an instantaneous cessation of blood flow to a flap primarily due to thrombosis at the arterial or venous anastomotic site (primary thrombosis)

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99m

with complete flap loss as a result. However, free tissue transfers occasionally ‘die’ a slow, progressive and partial death. This most likely is due to gradual ‘shutting down’ of the microcirculation by showering microemboli downstream from the arterial anastomosis (secondary thrombosis). Secondary thrombosis is not detectable without using sophisticated instrumentation to assess muscle flap viability. It is usually noticed clinically late in course, several days after surgery, as a slow, progressive flap death without a dramatic change in Doppler tone [36]. 99mTc sestamibi scintigraphy can show microcirculation problems such as distal ischaemia or a partial hypoperfused area in muscle tissue. In our series, there were normal Doppler findings in three cases (patients 2, 4 and 7) while, as the 99mTc sestamibi scan showed, there was a partial hypoperfused area in two patients (numbers 2 and 4) and distal ischaemia in one patient (number 7). Additionally, the short half-life of 99mTc (6 h) permits two scintigraphic examinations to be performed within 24 h. Third, secondary thrombosis may result in partial or total flap necrosis. In cases of partial muscle flap necrosis, 99m Tc sestamibi scintigraphy can show borders of necrosis and this knowledge is useful in surgical debridement.

Tc sestamibi scintigraphy in monitoring free muscle flap Top et al. 97

References 1

2

3

4

5

6 7

8

9

10

11

Fourth, some risk factors such as infection, hypercoagulable state, arteriosclerosis and the use of a vein graft predispose to secondary thrombosis. 99mTc sestamibi scintigraphy should be performed in these cases for the evaluation of perfusion of free muscle flaps. In our study, there was atherosclerosis in all cases except two (patients 3 and 13) and use of a vein graft in eight patients.

12

Fifth, a free muscle flap can not adhere in the recipient area because of ongoing necrosis or infection. When the free flap does not adhere in the recipient area and infectious and pseudo-membranous tissues cover the muscle flap, clinicians suspect partial or total flap necrosis. In these situations, 99mTc sestamibi scintigraphy is valuable in the evaluation of free muscle viability. In our study, there was a lack of adaptation to the recipient area and total skin graft necrosis in one case (patient 5). However, the flap tissue was well perfused completely visible in scintigraphic images.

16

13 14

15

17

18 19 20 21

From our data we conclude that 99mTc sestamibi scintigraphy is feasible and promising in the evaluation of free muscle flap viability and complications. However, further work, including comparative studies with other established techniques, is required to determine any impact on management or patient outcome in extensive patient series.

22

Acknowledgements

26

We would like to thank Serhan and Erhan Cildavil (nuclear medicine technicians) for their skilful technical assistance.

23

24

25

27

Schusterman MA, Miller MJ, Reece GP, Kroll SS, Marchi M, Goepfert H. A single center’s experience with 308 free flaps for repair of head and neck cancer defects. Plast Reconstr Surg 1994; 93:472–478. Kroll SS, Schusterman MA, Reece GP, Miller MJ, Evans GR, Robb GL, Baldwin BJ. Choice of flap and incidence of free flap success. Plast Reconstr Surg 1996; 98:459–463. Hidalgo DA, Disa JJ, Cordeiro PG, Hu QY. A review of 716 consecutive free flaps for oncologic surgical defects: refinement in donor-site selection and technique. Plast Reconstr Surg 1998; 102:722–732. Kind GM, Buntic RF, Buncke GM, Cooper TM, Siko PP, Bunck HJ Jr. The effect of an implantable Doppler probe on the salvage of microvascular tissue transplants. Plast Reconstr Surg 1998; 101:1268–1273. Kroll SS, Schusterman MA, Reece GP, Miller MJ, Evans GR, Robb GL, Baldwin BJ. Timing of pedicle thrombosis and flap loss after free-tissue transfer. Plast Reconstr Surg 1996; 98:1230–1233. Hirigoyen MB, Urken ML, Weinberg H. Free flap monitoring: a review of current practice. Microsurgery 1995; 16:723–726. Thorniley MS, Sinclair JS, Barnett NJ, Shurey CB, Green CJ. The use of near-infrared spectroscopy for assessing flap viability during reconstructive surgery. Br J Plast Surg 1998; 51:218–226. Machens HG, Mailaender P, Rieck B, Berger A. Techniques of postoperative blood flow monitoring after free tissue transfer: an overview. Microsurgery 1994; 15:778–786. Walkinshaw M, Holloway A, Bulkley A, Engrav LH. Clinical evaluation of laser Doppler blood flow measurements in free flaps. Ann Plast Surg 1987; 18:212–217. Solomon GA, Yaremchuk MJ, Manson PN. Doppler ultrasound surface monitoring of both arterial and venous flow in clinical free tissue transfers. J Reconstr Microsurg 1986; 3:39–41. Jones BM, Greenhalgh RM. The use of the ultrasound Doppler flowmeter in reconstructive microvascular surgery. Br J Plast Surg 1983; 36: 245–253. Tsuzuki K, Yanai A, Bandoh Y. A contrivance for monitoring skin flaps with a Doppler flowmeter. J Reconstr Microsurg 1990; 6:363–366. Karkowski J, Buncke HJ. A simplified technique for free transfer of groin flaps, by use of a Doppler probe. Plast Reconstr Surg 1975; 55:682–686. Concannon MJ, Stewart DH, Welsh CF, Puckett CL. Impedance plethysmography: a new method for continuous muscle perfusion monitoring. Plast Reconstr Surg 1991; 88:292–298. Graham BH, Gordon L, Alpert BS, Walton R, Buncke HJ, Leitner DW. Serial quantitative skin surface fluorescence: a new method for postoperative monitoring of vascular perfusion in revascularized digits. Am J Hand Surg 1985; 10:226–230. Jenkins SD, Sepka RS, Barwick WJ, Serafin D, Klitzman B. Routine clinical use of laser Doppler flowmeter to monitor free tissue transfer: preliminary results. J Reconstr Microsurg 1987; 3:281–283. Scheker LR, Slattery PG, Firrell JC, Lister GD. The value of the photoplethysmograph in monitoring skin closure in microsurgery. J Reconstr Microsurg 1985; 2:1–5. Doyle DJ, Eng P. A microminiature photoplethysmograph probe for microvascular surgery. Microsurgery 1984; 5:105–106. Graham B, Paulus DA, Caffee HH. Pulse oximetry for vascular monitoring in upper extremity replantation surgery. Am J Hand Surg 1986; 11:687–692. Sudarsky LA, Salomon J. Liquid crystal temperature monitoring for microsurgery. J Reconstr Microsurg 1991; 7:13–14. Raskin DJ, Erk Y, Spira M, Melissinos EG. Tissue pH monitoring in microsurgery: a preliminary evaluation of continuous tissue pH monitoring as an indicator of perfusion disturbances in microvascular free flaps. Ann Plast Surg 1983; 11:331–339. Serafin D, Lesesne CB, Mullen RY, Georgiade NG. Transcutaneous PO2 monitoring for assessing viability and predicting survival of skin flaps: experimental and clinical correlations. J Microsurg 1981; 2:165–178. Smith AR, Sonneveld GJ, Kort WJ, van der Meulen JC. Clinical application of transcutaneous oxygen measurements in replantation surgery and free tissue transfer. Am J Hand Surg 1983; 8:139–145. Davison PM, Batchelor AG, Wilson GR, Sully L. The muscle twitch monitor: a rapid unequivocal method of monitoring free muscle transfers. Br J Plast Surg 1986; 39:356–360. Sayman HB, Urgancioglu I. Muscle perfusion with technetium-MIBI in lower extremity peripheral arterial diseases. J Nucl Med 1991; 32:1700–1703. Sarikaya A, Sarikaya I, Pekindil G, Firat MF, Pekindil Y. Technetium-99m sestamibi limb scintigraphy in post-traumatic reflex sympathetic dystrophy: preliminary results. Eur J Nucl Med 2001; 8:1517–1522. Aygit AC, Sarikaya A. Imaging of frostbite injury by technetium-99msestamibi scintigraphy: a case report. Foot Ankle Int 2002; 23:56–59.

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28 Sarikaya A, Ege T, Firat MF, Enver D. Assessment of digital ischemia and evaluation of response to therapy by technetium-99m sestamibi limb scintigraphy after local cooling of hand in patients with vasospastic Raynaud’s syndrome. Nucl Med Commun 2004; 25:207–211. 29 Aygit AC, Sarikaya A. Technetium 99m sestamibi scintigraphy for noninvasive assessment of muscle flap viability. Ann Plast Surg 1999; 43:338–340. 30 Sarikaya A, Aygit AC. Combined 99mTc MDP bone SPECT and 99mTc sestamibi muscle SPECT for assessment of bone regrowth and free muscle flap viability in an electrical burn of scalp. Burns 2003; 29: 385–388. 31 Harashina T. Analysis of 200 free flaps. Br J Plast Surg 1988; 41:33–36. 32 Irons GB, Wood MB, Schmitt 3rd EH. Experience with one hundred consecutive free flaps. Ann Plast Surg 1987; 18:17–23.

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Disa JJ, Cordeiro PG, Hidalgo DA. Efficacy of conventional monitoring techniques in free tissue transfer: an 11-year experience in 750 consecutive cases. Plast Reconstr Surg 1999; 104:97–101. Jones NF. Intraoperative and postoperative monitoring of microsurgical free tissue transfers. Clin Plat Surg 1992; 19:783–797. Neligan PC. Monitoring techniques for the detection of flow failure in the post-operative period. Microsurgery 1993; 14:162–164. Weinzweig N, Gonzalez M. Free tissue failure is not an all-or-none phenomenon. Plast Reconstr Surg 1995; 96:648–660. Smith GT, Wilson TS, Hunter K, Besozzi MC, Hubner KF, Reath DB, et al. Assessment of skeletal muscle viability by PET. J Nucl Med 1995; 36:1408–1414. Gross JE, Friedman JD. Soft tissue reconstruction. Monitoring. Orthop Clin North Am 1993; 24:531–536.

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NEWS AND VIEWS January 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: + 44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: + 44 (0)113 206 6930, E-mail: [email protected] Nuclear Medicine Communications, 2006, 27:99–100

The Radiology Integrated Training Initiative (RITI)

The Royal College of Radiologists, in collaboration with the Department of Health (DoH), has started work on an ambitious project aimed at increasing the throughput of trainees in radiology across the UK. The main trigger for this development was the embarrassing shortage of applicants for consultant radiology posts over the last few years, together with a belated realization by DoH that many of the new healthcare initiatives they have introduced recently – for example, the earlier diagnosis of common cancers and rapid access arrangements for patients with heart disease – have imposed a substantial additional burden on imaging facilities and staff, on top of the continuing growth in imaging arising from improvements in technology, in new applications, and the massive increase in monitoring new treatments, particularly in oncology. At the same time, post-graduate medical education is being completely overhauled, with a politically driven intention to substantially shorten the duration of higher specialist training. How could we produce more trained specialists in a shorter time, with no increase in the number of training centres and trainers? The approach which has been adopted is ambitious and, if successful, will have implications for training in other specialties.

The Radiology Integrated Training Initiative (RITI) project has three main strands: radiology academies, an electronic learning database and a validated case archive. The ‘radiology academy’ is a rather loose descriptive term for the physical environment in which an increased number of trainees will be accommodated. In some cases it could be a new building, most often it will be an enhanced or expanded area within existing facilities. I believe it is envisaged that all major training centres will eventually develop radiology academies, but as a first step three pilot sites in Yorkshire, East Anglia, and the south-west are now ‘operational’. The electronic learning database (ELD) represents an attempt to translate the whole of the radiology training curriculum up to FRCR2 level into a computer-based interactive learning format. It is estimated that about 1200 electronic tutorial sessions, each of about 20 min duration, will be needed. Production of these tutorial sessions is already well under way and it is hoped to complete them by early spring 2006. The third element in the new programme, the validated case archive (VCA), is envisaged to be a national repository of teaching cases, each of which has been thoroughly worked up and peer reviewed. The initial proposal is to aim for 10 000 validated cases covering all aspects of

imaging, which will be made available electronically to radiology trainees throughout the country. How will all this increase the throughput of trainees? Currently, radiology training is mainly an apprenticeship, in which sporadic episodes of formal teaching punctuate daily (and nightly!) experience in the radiology department. Under the new system, trainees will spend a substantial proportion of their time away from the ‘shop floor’, using the electronic learning facilities. In parallel with this, it is hoped that the use of web-cam technology and the like will allow individual trainers to interact simultaneously with a larger number of trainees than is possible in the conventional physical environment where students are clustered around viewing screens together with the teacher. Can all this work? Even if the plan is implemented as proposed, will it actually deliver a larger number of trained specialists without diminishing the quality of the training? If it does work, what would be the implications for training in other specialities? The answers to all these questions should become clear within the next 2–3 years. Currently, the tasks which loom large are creating the ELD, and populating the VCA. Professor Isky Gordon has bravely taken on the role of coordinating and editing the radionuclide imaging

c 2006 Lippincott Williams & Wilkins 0143-3636

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100 Nuclear Medicine Communications 2006, Vol 27 No 1

elements of the ELD. At present we envisage creating about 100 electronic sessions to deliver learning/ teaching in radionuclide imaging up to FRCR2 level. A team of nuclear medicine physicians and radionuclide radiologists has been identified to create these teaching units, but the time and effort needed is not inconsequential. My own experience is that when creating sessions from teaching material which I already had available electronically in the form of text, images, graphics, AVIs etc, it took 5–6 h to prepare the content of a 20 min session in a suitable form for the project’s software engineers to translate into an appropriate electronic learning framework. However, once the work is completed, it should offer a most useful resource for trainees, not only in other medical specialties but also in other professions. The detailed description of RITI which has recently appeared on the website of the Royal College of Radiologists gives the impression that the project is well-advanced and that the facilities envisaged in it are already available. This is sort of true, up to a point. The structure of the new learning environment is in place, the content has been described, the authors have been identified, and hopefully are hard at work, but as usual, implementation of the detail is taking much longer than was anticipated by the strategists who developed the plans. Even so, we may look with cautious optimism at the development of a valuable new training

initiative which may eventually have a much wider benefit than its initial target. Professor Philip Robinson, Isotope Department, St James’s Hospital, Leeds, UK Meeting Announcements

UK Radiopharmacy Group Workshop: Radiopharmacy Practice and Quality Control Update Date: 13 January 2006 Venue: Beeches Management Centre, Bournville, Birmingham, UK Website: www.ukrg.org.uk Applications of Radiotracers in Chemical, Environmental and Biological Sciences (ARCEBS 06) Dates: 23–27 January 2006 Venue: Saha Institute of Nuclear Physics, Kolkata, India Website: www.saha.ernet.in/arcebs Communications: Professor Susanta Lahiri [email protected] Annual Clinical PET CT Course – a user’s guide Dates: 8–10 March 2006 Overview of basic science, PET CT imaging and interactive teaching. Contact for information and registration: [email protected] BNMS Spring Meeting Dates: 27–29 March 2006 Venue: Manchester, UK Website: www.bnms.org 2nd European IRPA Congress on Radiation Protection Dates: 15–19 May 2006

Venue: Paris, France Website: www.irpa2006europe.com BNMS Autumn Meeting Dates: 4–5 September 2006 Venue: Cambridge, UK Website: www.bnms.org.uk EANM Annual Meeting Dates: 29 September to 3 October 2006 (provisional) Venue: Athens, Greece Website: www.eanm.org 9th World Congress of Nuclear Medicine and Biology Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/congress2006/ index02.htm Education and Training

EANM Learning Courses Dates: Weekend courses throughout 2005 including: 12–13 November, EANM Therapy–Dosimetry Course 19–20 November, EANM Neuroimaging Course 17–18 December, EANM learning course on PET/CT in oncology Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on Tel: + 43 1 212 8030; fax: + 43 1 212 80309 Website: www.eanm.org/education/ esnm/esnm_intro.php E-mail: [email protected]

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Editorial

PET and TNM: evolution or revolution? Heok K. Cheow, N. George Mikhaeel and Michael J. O’Doherty Nuclear Medicine Communications 2006, 27:101–103

Received 18 October 2005 Accepted 6 December 2005

Correspondence: Dr Heok Cheow, Clinical PET Centre, St Thomas’ Hospital, London SE1 7EH, UK. Tel: + 44 1223 217147; e-mail: [email protected]

The TNM staging system in the diagnosis and treatment of cancer was first proposed by Denoix in 1944 [1] and the concept has subsequently been adopted by the Union Internationale Contre le Cancer (UICC, also known as International Union Against Cancer) with the first edition of TNM Classification of Malignant Tumours published in 1968 as the ‘livre de poche’. The American Joint Committee on Cancer (AJCC) also introduced a TNM staging system. However, the UICC and AJCC TNM classifications developed separately with different definitions of the tumour (T), node (N) and metastases (M) categories. It was not until 1987 that the two organizations unified the definitions of TNM [2]. Systems used by international speciality oncology societies were blended into TNM if it was feasible or, if not, alternatives were adopted instead of TNM (e.g. FIGO classification for gynaecological tumours and Ann Arbor classification of lymphomas). This staging system has provided a benchmark for reporting the extent of disease, enabling the planning of treatment, providing an indication about prognosis, allowing the identification of similar patient groups to assess therapy outcome and evaluate different treatment methods between oncology centres and provide a mechanism for identifying patients for clinical trials. With the use of TNM for more than 50 years and with sequential editions, the classification has evolved to accommodate new knowledge. Clinical staging beyond physical examination has been widely conditioned by imaging techniques (e.g. computed tomography (CT) and magnetic resonance imaging (MRI)). The accepted minimal staging work-up for each tumour site has changed over the years. Yet for some tumour sites, surgical pathological staging is the mainstay of the classification (e.g. colorectal T staging). Several other factors beyond the anatomical extent of cancer (as expressed by stage) have an impact on the natural history and prognosis. These vary from simple laboratory tests, e.g. prostate specific antigen in prostrate cancer, to the novel ‘gene expression profiling’, e.g. in lymphomas. As our ability to diagnose cancer early, to

characterize tumours, to assess disease extent and to treat cancer improves each year, there is a need to keep pace with progress. From the clinicians’ point of view, a more adaptable classification that is relevant to contemporary clinical practice and which makes use of the available technology is preferable. Other uses of TNM, on the other hand, would benefit more from a more stable classification with less frequent changes. An example of the latter, is the use of TNM by cancer registries. Therefore, a balance needs to be struck. Positron emission tomography (PET) imaging as a novel functional imaging technique, complementing the largely ‘anatomical’ cross-sectional imaging modalities can offer a variety of molecular insights but can it fit neatly into a TNM classification or does it need to exist as a parallel stream? Over the last few years, there have been numerous observational clinical reports on the use of PET or PET/CT imaging in the primary staging of different cancers, but a number of questions are still to be answered. The choice of tracer may depend on the specific cancer in question. 18F labelled 2-fluorodeoxy-D-glucose (FDG) is, by far, the molecule most commonly used for imaging cancer but there are a host of other molecules that can complement FDG imaging, and in certain situations or cancer types, define the extent of disease more appropriately. 11C-methionine imaging in brain tumours may define the extent of the disease more accurately and 11Ccholine may define sites of metastatic prostate cancer more specifically than FDG. Here lies a major question of using PET for TNM staging: which tracer for which disease? The definition of the T stage varies in different tumour sites. It is based on size criteria for some tumours (e.g. lung and breast), local extent and invasion of surrounding structures (e.g. uterine cervix, head and neck, and lung), depth of wall infiltration (e.g. colorectal and bladder). The latter is unlikely to be easily evaluated by PET, even with the advent of PET/CT, and will probably remain a pathological system. The difficulty with edge detection, and issues related to motion with PET, make it unlikely

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102 Nuclear Medicine Communications 2006, Vol 27 No 2

that the technique will accurately define the edge of a tumour. The N stage also relies on the degree of nodal metabolic activity being sufficient to identify disease. This may be problematic in some cancers. In lung cancer, inflammation can cause increased uptake in nodes leading to false positive results [3] and in melanoma there is a threshold amount of disease required to detect nodal involvement resulting in false negative results [4,5]. Determination of the M stage is likely to be the most affected by adding PET to routine staging work-up, but it is not without limitations. Different tumours have extremely variable uptake with low-grade uptake seen in bronchoalveolar carcinoma [6], carcinoid [7] and mucinous adenocarcinomas of the colon [8] as well as a variety of other tumour types [9]. With these tumour types the identification of early metastatic disease would be difficult if not impossible using FDG. Even with highgrade tumours, pulmonary metastases may not be visualized with FDG, for an as yet undefined reason which is not just related to size of the metastasis [10–12], and the addition of PET/CT offers a method of ensuring that these metastases can be identified on the CT component. The timing of PET scanning in relation to FDG administration may have an effect on the accuracy of PET in staging. There is evidence for a number of tumour types that the peak uptake does not appear in a tumour until 2–5 h after the FDG injection. This has been apparent in lung cancers [13,14], sarcomas [15] and breast cancer [16]. Recent studies have suggested that by delaying whole-body imaging until 2 or 3 h after FDG injection, more metastases are demonstrated [17,18]. These areas of research raise an important question as to when is the optimum scanning time, since one only has to look at the published data to show that clinical scanning times vary from 30 min to 2 h post-injection [19]. There is patchy data on the value of FDG PET in the primary staging of patients with cancer. The available data varies from very good in the case of lung cancer [20] and lymphoma [21] to less satisfactory in melanoma [22] and head and neck cancer [23]. Despite this the overall sensitivity and specificity is higher than most conventional imaging studies. Even in lung cancer, where the strongest case for PET staging has been made, the available data is only on patients selected by other imaging for consideration of surgery or radical radiotherapy. The addition of PET/CT has improved the situation [24] but there are still false positives and negatives. Difficulties with respiratory motion and movement of abdominal structures may result in inaccurate localization of abnormal uptake. The use of oral contrast

on the CT and the effect of this on the attenuation corrected PET image quantification are still being debated. It is expected that the role of PET would not just be a road map, but with the biochemical information it provides and with the correct range of tracers for each disease process, management decisions can potentially be enhanced. Even with FDG alone in lung cancer, there is a suggestion that the level of uptake can indicate prognosis [25–27] that could help refine treatment decisions, but this work needs to be confirmed on a large scale before applying to clinical practice. This role of PET could add to the prognostic value of TNM stage in the future. Application to other tumour sites needs to be explored. It is clear that a great deal of work needs to be done on the evaluation of PET in the primary staging of different tumour sites (with sensitivity and specificity data) and also on the standardization of technique and choice of tracers. This work is essential towards the evidence-based integration of PET/CT in the routine staging of cancer. At present, trying to introduce PET/CT to a TNM stage prematurely without the necessary evidence to support this will only result in confusion for clinicians and possibly bring the technique into disrepute. A parallel system is likely to be required for a while. The introduction of PET to TNM staging certainly will not be a revolution but is more likely to be an evolution towards the establishment of PET in the primary staging of all cancers.

References 1

Denoix PF. Tumor, node, and metastasis (TNM). Bull Inst Nat Hyg 1944; 1:1–69. 2 Hermanek P, Sobin LH (editors). UICC TNM Classification of Malignant Tumours, 4th edition. Berlin: Springer-Verlag, 1992. 3 Takamochi K, Yoshida J, Murakami K, Niho S, Ishii G, Nishimura M, et al. Pitfalls in lymph node staging with positron emission tomography in non-small cell lung cancer patients. Lung Cancer 2005; 47:235–242. 4 Acland KM, Healy C, Calonje E, O’Doherty M, Nunan T, Page C, et al. Comparison of positron emission tomography scanning and sentinel node biopsy in the detection of micrometastases of primary cutaneous malignant melanoma. J Clin Oncol 2001; 19:2674–2678. 5 Wagner JD, Schauwecker D, Davidson D, Coleman 3rd JJ, Saxman S, Hutchins G, et al. Prospective study of fluorodeoxyglucose-positron emission tomography imaging of lymph node basins in melanoma patients undergoing sentinel node biopsy. J Clin Oncol 1999; 17:1508–1515. 6 Higashi K, Ueda Y, Seki H, Yuasa K, Oguchi M, Noguchi T, et al. Fluorine-18FDG PET imaging is negative in bronchioloalveolar lung carcinoma. J Nucl Med 1998; 39:1016–1020. 7 Erasmus JJ, McAdams HP, Patz Jr EF, Coleman RE, Ahuja V, Goodman PC. Evaluation of primary pulmonary carcinoid tumors using FDG PET. Am J Roentgenol 1998; 170:1369–1373. 8 Berger KL, Nicholson SA, Dehdashti F, Siegel BA. FDG PET evaluation of mucinous neoplasm, Correlation of FDG uptake with histopathologic features. Am J Roentgenol 2000; 174:1005–1008. 9 Lee FY, Yu J, Chang SS, Fawwaz R, Parisien MV. Diagnostic value and limitations of fluorine-18 fluorodeoxyglucose positron emission tomography for cartilaginous tumors of bone. J Bone Joint Surg Am 2004; 86-A: 2677–2685. 10 Mijnhout GS, Hoekstra OS, van Tulder MW, Teule GJ, Deville WL. Systematic review of the diagnostic accuracy of (18)F-fluorodeoxyglucose

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positron emission tomography in melanoma patients. Cancer 2001; 91:1530–1542. Fuster D, Chiang S, Johnson G, Schuchter LM, Zhuang H, Alavi A. Is 18F-FDG PET more accurate than standard diagnostic procedures in the detection of suspected recurrent melanoma? J Nucl Med 2004; 45:1323–1327. Lucas JD, O’Doherty MJ, Wong JC, Bingham JB, McKee PH, Fletcher CD, et al. Evaluation of fluorodeoxyglucose positron emission tomography in the management of soft-tissue sarcomas. J Bone Joint Surg B 1998; 80:441–447. Matthies A, Hickeson M, Cuchiara A, Alavi A. Dual time point 18F-FDG PET for the evaluation of pulmonary nodules. J Nucl Med 2002; 43:871–875. Hamberg LM, Hunter GJ, Alpert NM, Choi NC, Babich JW, Fischman AJ. The dose uptake ratio as an index of glucose metabolism: useful parameter or oversimplification? J Nucl Med 1994; 35:1308–1312. Lodge MA, Lucas JD, Marsden PK, Cronin BF, O’Doherty MJ, Smith MA. A PET study of 18FDG uptake in soft tissue masses. Eur J Nucl Med 1999; 26:22–30. Boerner AR, Weckesser M, Herzog H, Schmitz T, Audretsch W, Nitz U, et al. Optimal scan time for fluorine-18 fluorodeoxyglucose positron emission tomography in breast cancer. Eur J Nucl Med 1999; 26:226–230. Ma SY, See LC, Lai CH, Chou HH, Tsai CS, Ng KK, et al. Delayed (18)F-FDG PET for detection of paraaortic lymph node metastases in cervical cancer patients. J Nucl Med 2003; 44:1775–1783. Yen TC, Ng KK, Ma SY, Chou HH, Tsai CS, Hsueh S, et al. Value of dual-phase 2-fluoro-2-deoxy-D-glucose positron emission tomography in cervical cancer. J Clin Oncol 2003; 21:3651–3658. Lowe VJ, DeLong DM, Hoffman JM, Coleman RE. Optimum scanning protocol for FDG-PET evaluation of pulmonary malignancy. J Nucl Med 1995; 36:883–887.

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Shim SS, Lee KS, Kim BT, Chung MJ, Lee EJ, Han J, et al. Non-small cell lung cancer: prospective comparison of integrated FDG PET/CT and CT alone for preoperative staging. Radiology 2005; 236: 1011–1019. Isasi CR, Lu P, Blaufox MD. A metaanalysis of (18)F-2-deoxy-2-fluoro-Dglucose positron emission tomography in the staging and restaging of patients with lymphoma. Cancer 2005; 104:1066–1074. Kumar R, Mavi A, Bural G, Alavi A. Fluorodeoxyglucose-PET in the management of malignant melanoma. Radiol Clin N Am 2005; 43: 23–33. Stoeckli SJ, Steinert H, Pfaltz M, Schmid S. Is there a role for positron emission tomography with 18F-fluorodeoxyglucose in the initial staging of nodal negative oral and oropharyngeal squamous cell carcinoma. Head Neck 2002; 24:345–349. Cerfolio RJ, Ojha B, Bryant AS, Raghuveer V, Mountz JM, Bartolucci AA. The accuracy of integrated PET-CT compared with dedicated PET alone for the staging of patients with non small cell lung cancer. Ann Thorac Surg 2004; 78:1017–1023. Ahuja V, Coleman RE, Herndon J, Patz Jr EF. The prognostic significance of fluorodeoxyglucose positron emission tomography imaging for patients with nonsmall cell lung carcinoma. Cancer 1998; 83:918–924. Dhital K, Saunders CA, Seed PT, O’Doherty MJ, Dussek J. [(18)F]Fluorodeoxyglucose positron emission tomography and its prognostic value in lung cancer. Eur J Cardiothorac Surg 2000; 18:425–428. Vansteenkiste JF, Stroobants SG, Dupont PJ, De Leyn PR, Verbeken EK, Deneffe GJ, et al. Prognostic importance of the standardized uptake value on (18)F-fluoro-2-deoxy-glucose-positron emission tomography scan in non-small-cell lung cancer: An analysis of 125 cases. Leuven Lung Cancer Group. J Clin Oncol 1999; 17:3201–3206.

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Review paper

Ventricular mechanical dyssynchrony and resynchronization therapy in heart failure: a new indication for Fourier analysis of gated blood-pool radionuclide ventriculography G. Aernout Somsena,w, Hein J. Verberneb,w, Haran Burric, Osman Ratibc and Alberto Righettic In patients with decreased left ventricular ejection fraction and conduction disease, ventricular mechanical dyssynchrony has been demonstrated. To date, resynchronization by biventricular pacing is increasingly used since it improves ventricular function and exercise capacity in patients with heart failure. To optimize and evaluate the effect of resynchronization therapy and to identify patients who may benefit from biventricular pacing the assessment of left ventricular synchronicity is essential. Therefore, a non-invasive and reproducible technique to obtain information on ventricular synchrony is clinically valuable. In this review, the technical background and the role of phase analysis of gated blood-pool nuclear ventriculography in the assessment of ventricular mechanical synchrony, especially in heart failure patients subjected to biventricular pacing, will be discussed. Nucl

Introduction Ventricular systolic performance can be described as a function of force, velocity and synchrony of myocyte contraction. The value of the contractile force and velocity is widely recognized as a clinical indicator of cardiac function and prognosis in patients with heart failure. However, the clinical importance of synchronous segmental myocardial contraction was not recognized until the early 1980s when regional asynchronous activation was demonstrated in heart failure patients by temporal Fourier analysis of nuclear ventriculography data. This technique was predominantly used to quantify wall motion [1–4] and conduction abnormalities [5–7]. For several years, the assessment of ventricular synchrony has been gaining interest because resynchronization of ventricular contraction by pacing has been demonstrated to improve ventricular performance in patients with heart failure and conduction abnormalities. This illustrates that synchrony of ventricular contraction is not only interesting from a pathophysiological point of view but that it may also serve as a target for therapeutic intervention in patients with heart failure. This review addresses the current knowledge of ventricular mechanical synchrony in both the normal and the failing heart. In addition, the technical background and w

G. Aernout Somsen and Hein J. Verberne contributed equally to this manuscript.

Med Commun 27:105–112 Wilkins.


c

2006 Lippincott Williams &

Nuclear Medicine Communications 2006, 27:105–112 Keywords: gated blood-pool, heart failure, resynchronization therapy, ventriculography. a

Department of Cardiology, Onze Lieve Vrouwe Gasthuis, Amsterdam, Department of Nuclear Medicine, Academic Medical Center, University of Amsterdam, Netherlands and cDepartment of Cardiology and Radiology, University Hospital Geneva, Switzerland. b

Correspondence to Dr G.A. Somsen, Department of Cardiology, Onze Lieve Vrouwe Gasthuis, P.O. Box 95500, 1090 HM Amsterdam, The Netherlands. Tel: + 31 20 599 3032; fax: + 31 20 599 3997; e-mail: [email protected] Received 1 September 2005 Accepted 15 November 2005

clinical value of phase analysis of blood-pool nuclear ventriculography to assess ventricular mechanical synchrony, especially in heart failure patients subjected to biventricular pacing, will be discussed.

Ventricular mechanical synchrony The way mechanical activation spreads over the ventricles is determined by the propagation of ventricular depolarization and the excitation–contraction coupling. In the normal heart, duration of ventricular electrical activation (ranging from approximately 80 to 100 ms) and mechanical activation (ranging from approximately 150 to 200 ms) are closely related, although an excitation– contraction delay of approximately 20 ms exists. In the structurally normal heart, without conduction disease, mechanical activation spreads homogeneously. Activation usually commences in the basal septum. The right ventricle is activated slightly earlier than the left ventricle [8]. Subsequently, the activation spreads towards the lateral wall of the left ventricle (Fig. 1). The propagation of mechanical activation over each ventricle and over both ventricles is called intraventricular and inter-ventricular synchrony, respectively.

Ventricular dyssynchrony In patients with dilated cardiomyopathy, delayed electrical activation and impaired excitation–contraction coupling [9] result in a dispersion of regional mechanical

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Fig. 1

LV

LV RV

RV

RVEF = 50% std = 8.9 Mean = 256 Mode = 260 Skewn = −0.18

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LVEF = 65% std = 5.8 Mean = 269 Mode = 270 Skewn = −0.11

360/0 90

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std = 7.8 Mean = 245 Mode = 245 Skewn = −0.7 IVD = 13 ms

270 360/0 90

180 270 360 Degrees

Phase histogram of a normal mechanical activation of the right, left and both ventricles. Each colour in the histogram corresponds to the localization of pixels having the same phase angle. Clinically used parameters to express ventricular synchrony are shown. RVEF = right ventricular ejection fraction, LVEF = left ventricular ejection fraction, std = standard deviation of the phase, skewn = skewness, IVD = inter-ventricular delay. Data are expressed in degrees or milliseconds.

activation known as ventricular dyssynchrony and is associated with increased pre-systolic mitral valve regurgitation, decreased left ventricular diastolic filling time, reduced left ventricular contractility and occasionally with septal wall dyskinesia. Since approximately 35% of heart failure patients have a prolonged QRS duration ( > 120 ms), the number of patients with ventricular dyssynchrony is substantial. Moreover, the prevalence of dyssynchrony in these patients may be underestimated since recent reports have identified ventricular dyssynchrony even in the presence of normal QRS duration [10,11]. Since right and left ventricular function are interdependent, inter-ventricular synchrony contributes to left ventricular performance. The importance of inter-ventricular synchrony is illustrated by the observation that right ventricular apical pacing results in decreased systolic left ventricular function [12]. However, the prognostic importance of inter-ventricular synchrony is still debated. Recently, intra-ventricular but not inter-ventricular synchrony was an independent prognostic indicator in heart failure patients [13].

Resynchronization therapy in heart failure Restoration of ventricular synchrony, or resynchronization, can be accomplished by biventricular pacing. This therapy is characterized by standard placement of an atrial lead, and a ventricular lead which is positioned in the apex or inter-ventricular septum of the right

ventricle. An additional third pacemaker lead is ideally positioned in a lateral cardiac vein to stimulate the left ventricle. Following a sensed atrial electrical activation, both ventricles are stimulated to contract either simultaneously or with pre-excitation of one of the ventricles. This results in resynchronization of ventricular contraction which may improve left ventricular systolic function, decrease left ventricular end systolic and end diastolic diameters, reduce mitral regurgitation and optimize left ventricular filling, thereby reducing symptoms and increasing exercise capacity. According to the American Heart Association Guidelines, biventricular pacing is indicated in medically refractory, symptomatic (New York Heart Association class III or IV) patients with dilated or ischemic cardiomyopathy, prolonged QRS interval (greater or equal to 130 ms), left ventricular end diastolic diameter greater or equal to 55 mm, and ejection fraction less than or equal to 35% [14]. A recently published meta-analysis, pooling data from four randomized trials, demonstrated a 51% reduction in mortality rate due to progressive heart failure and a 29% reduction in hospitalization rate for heart failure, during a follow-up of 3–6 months, in patients subjected to biventricular pacing compared to control patients [15]. Moreover, a trend towards reduced all-cause mortality was observed in this analysis. The Comparison of Medical Therapy, Pacing, and Defibrillation in Heart Failure (COMPANION) trial showed that

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Ventricular dyssynchrony and gated blood-pool ventriculography Somsen et al. 107

cardiac-resynchronization therapy (CRT) alone or combined with an implantable defibrillator reduced the composite end point of death from any cause or hospitalization during a mean follow-up of 16 months. The decrease in the risk of death, however, was not significant with cardiac resynchronization therapy alone (P = 0.06) [16]. In addition the CARE–HF study showed that in patients with class III to IV heart failure, left ventricular ejection fraction (LVEF) = 35% and ventricular dyssynchrony (QRS > 150 ms or QRS > 120 ms and with echocardiographic mechanical dyssynchrony), CRT resulted in reduction of mortality, improvement of symptoms and quality of life [17]. In patients with heart failure treated with biventricular pacing, improvement of ventricular function is best predicted by the severity of left ventricular dyssynchrony at baseline, as was assessed by echocardiography [18,19]. Remarkably, the alteration in QRS duration was a less valuable predictor of a beneficial response. This may suggest that a reduction in electrical activation time is not essential to ameliorate mechanical activation, indicating that electrical and mechanical activation are not strictly correlated in these patients. In heart failure patients, the optimal stimulation site of the left ventricular pacing lead is still debated. In the acute phase, the optimal haemodynamic response is obtained by stimulating the lateral left ventricular wall [20,21]. However, recent data have shown that, during long-term follow-up, clinical outcome is independent of the stimulation site [22]. It can be hypothesized that the optimal pacing site may be the site with the most delayed activation and may therefore vary among these patients. These observations emphasize the clinical importance of left ventricular synchronicity assessment to optimize and evaluate the effect of resynchronization therapy and to identify patients who may benefit from biventricular pacing. Therefore, a non-invasive and reproducible technique to obtain information on ventricular synchrony is essential and clinically useful in patients with heart failure.

Gated blood-pool ventriculography and Fourier analysis

sults with respect to sensitivity and specificity of phase analysis in detecting abnormally contracting myocardial segments [4,24]. However, phase analysis of blood-pool nuclear ventriculography can be used to quantify the temporal sequence of systolic ventricular wall motion in a non-invasive and reproducible way [25]. Technical aspects

For equilibrium studies, a tracer is required which remains within the vascular compartment at a relative stable concentration during data collection. Although there a variety of tracers which fulfil this requirement, 99m Tc labelling of erythrocytes is almost universally employed. There are several methods for labelling erythrocytes with 99mTc. The principle behind the technique is that 99mTc, in its normal oxidized form, diffuses freely into and out from erythrocytes. In its reduced form, however, obtained by using stannous chloride, 99mTc binds tightly to the beta chain of haemoglobin resulting in effective labelling of erythrocytes. After labelling erythrocytes with 99mTc, triggered acquisitions are performed using continuous ECG monitoring to ensure R-wave gating of the QRS complex. The RR interval is divided into frames typically ranging from 16 to 64 frames. A higher temporal resolution (i.e., number of frames per cardiac cycle) allows for a more accurate determination of left ventricular function. Increasing temporal resolution, however, leads to a decrease in count density. To compensate for this decrease the image acquisition time can be prolonged. Since no commercial software is currently available for applying phase analysis to SPECT images, only the acquisition and analysis of planar images will be further discussed. Planar images should be acquired in left anterior oblique projection to allow for optimal separation of both ventricles and identification of the inter-ventricular septum. Regions of interest can be automatically or manually drawn around both ventricles, or any other region of interest, in end diastolic and end systolic frames. Background correction can be performed by using an extra-cardiac region of interest. Subsequently, a time–activity curve is constructed and ventricular mechanical activation in time can be assessed. The time–activity curve of the right and left ventricle as obtained from nuclear ventriculography resembles a cosine function.

Background of the technique

The recognition that timing of ventricular contraction is an important influence on ventricular contractile function has resulted in increasing interest in techniques to assess ventricular synchrony. Currently, a revival of a technique which has been described a few decades ago takes place [23]. The initial goal of applying Fourier analysis to ventriculographic images was to detect areas of abnormal contraction. Studies have demonstrated conflicting re-

To reduce random variation due to the Poisson nature of radioactive decay, data from blood-pool nuclear ventriculography can be smoothed using spatial and temporal filtering [26]. Temporal Fourier transformation using the heart rate as the fundamental frequency is performed for each pixel while using the R-wave on the ECG as a reference point. The time–activity curve for each pixel is fitted with a single cosine whose period equals the RR

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Fig. 2

LV

LV

RV

RV

RVEF = 35% std = 15.8 Mean = 236 Mode = 225 Skewn = 1.1

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LVEF = 17% std = 71 Mean = 283 Mode = 285 Skewn = 3.1

360/0

90

std = 62.7 Mean = 259 Mode = 273 Skewn = 4.2 IVD = 47 ms

180 270 360/0 Degrees

90

180 270 Degrees

360

Phase histogram of the right, left and both ventricles in a patient with dilated cardiomyopathy and left bundle branch block. Clinically used parameters to express ventricular synchrony are shown. (Abbreviations as in the legend to Fig. 1.)

interval. The single-cycle cosine fit for each pixel can be characterized by amplitude and phase, representing the maximum value and the shift in time of a given pixel’s cosine, respectively. Phase data are either expressed in degrees between 01 and 3601 or in milliseconds. Both units are interchangeable by using the algorithm phase angle=360 RR interval ðmsÞ: Phase data can be displayed using a colour-coded histogram in which the number of pixels within the ventricle having any phase angle are plotted on the Y axis and the phase angle is plotted on the X axis (Fig. 1). The phase angle corresponds to the relative sequence and pattern of ventricular contraction during the cardiac cycle. Each colour in the histogram corresponds with the localization of pixels having the same phase angle. Regional Fourier analysis of the ventriculographic data can also be performed (i.e., left or right ventricle, septum etc.) to determine local mechanical activation. A distribution curve (i.e., phase–amplitude curve) can be described using various mathematical parameters each reflecting different curve characteristics. In both, the normal and the failing heart, (regional) phase can be expressed in terms of the mean, median, standard deviation, mode and skewness (Figs 1 and 2).

The mean phase angle of the histogram represents the average of all phase angles within the distribution curve. The mean phase angle can be used to determine mechanical activation in different regions of interest, and can be used to evaluate (inter-) regional synchrony (Fig. 3). Inter-ventricular synchrony is expressed as the difference between the mean phase angle of the right and the left ventricle. In patients with dilated cardiomyopathy interventricular dyssynchrony may be present (Fig. 2). The inter-ventricular delay was positively correlated to QRS duration [13]. The degree of inter-ventricular dyssynchrony has been shown to correlate with LV ejection fraction [27]. In a study using radionuclide ventriculography, inter-ventricular asynchrony was a predictor for the magnitude of improvement in synchrony and left ventricular ejection fraction in heart failure patients subjected to biventricular pacing [27]. Currently, no data are available on the optimal inter-ventricular delay during biventricular pacing. Figure 4 shows data from a patient with dilated cardiomyopathy and left bundle branch block subjected to biventricular pacing with varying inter-ventricular delay. Both ventricular synchrony and left ventricular ejection function progressively improve by pre-excitation of the left ventricle.

Quantification of ventricular synchrony

Commonly used parameters to quantify the degree of mechanical synchrony assessed by nuclear ventriculography are: mean phase, median, mode and standard deviation of the phase histogram.

The median of the phase histogram is the midpoint of all the individual measures and has been related to the number of pixels. The median is more informative than the mean phase in case of a skewed distribution.

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Ventricular dyssynchrony and gated blood-pool ventriculography Somsen et al. 109

Fig. 3

(a)

(b) S S

LV

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EF = 23% std = 54.4 Mean = 249 Mode = 260 Skewn = −1.7

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270

360

Phase histogram of both ventricles in a patient with dilated cardiomyopathy and left bundle branch block. Panel A: Native ventricular conduction during atrial pacing (80 beats/min). Intra-ventricular dyssynchrony is shown. Panel B: Biventricular pacing (80 beats/min). Arrow indicates the preexcitation of the lateral left ventricular wall ( = left ventricular lead position). Improved intra-ventricular synchrony and systolic performance of the left ventricle during biventricular pacing is demonstrated as compared to native conduction. RV = right ventricle, LV = left ventricle, S = septum. (Other abbreviations as in the legend to Fig. 1.)

The standard deviation of the phase angle reflects the width of the phase histogram and serves as an index of total magnitude of intraventricular synchrony and is considered to be sensitive to segmental wall motion abnormalities [3]. Small standard deviations indicate synchronous contraction, whereas large values indicate intraventricular dyssynchrony. The standard deviation of the phase was an independent predictor of cardiac events such as death, worsening of heart failure and cardiac transplantation in patients with idiopathic dilated cardiomyopathy [13]. In patients with heart failure and intraventricular conduction disease, the standard deviation of the phase was a powerful predictor of beneficial response to biventricular pacing [18,21]. These data indicate that standard deviation of the phase is an important clinical parameter which can be used as a prognostic marker, and also for selecting patients who may benefit from resynchronization therapy. In addition, this parameter may allow for evaluation of the success of biventricular pacing and potentially guides optimization of lead position.

The skewness of the phase histogram reflects the symmetry or obliquity of the phase histogram which corresponds with the time at which most of the (a)synchrony occurs. A negatively skewed distribution is one whose elongated tail extends to the left end of the range, while a positively skewed distribution is one whose elongated tail extends to the right end of the range. An unskewed distribution, on the other hand, is one that is completely symmetrical. A negative skewness indicates early asynchrony (Fig. 3), whereas a positive skewness represents late dyssynchrony. Clinical studies failed to demonstrate a correlation between skewness and wall motion abnormalities [4]. Therefore, the clinical usefulness of this parameter remains to be determined.

Limitations and potential pitfalls of Fourier analysis of gated blood-pool nuclear ventriculography Temporal resolution is limited because, for example, 32 frames provide a resolution of 31.25 ms (for a heart rate of

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110 Nuclear Medicine Communications 2006, Vol 27 No 2

Fig. 4

AAI

LVEF = 16% std = 63 Mean = 259 IVD = 35 ms

0

LV (−40 ms)

LV = RV

90

180 270 360 Degrees

LVEF = 22% std = 37 Mean = 267 IVD = 15 ms

0

90

LVEF = 28% std = 28 Mean = 270 IVD = 10 ms

180 270 360 Degrees

0

90

180 270 Degrees

360

Phase histogram of both ventricles in a patient with dilated cardiomyopathy and left bundle branch block, during various modes of stimulation (with identical heart rate: 80 beats/min). Left panels: atrial stimulation (AAI). Middle panels: Stimulation of left and right ventricle at the same time (LV = RV). Right panels: Stimulation of left ventricle 40 ms before stimulation of the right ventricle (LV ( – 40 ms)). (Abbreviations as in the legend to Fig. 1.)

60 beats/min). Although other techniques such as magnetic resonance imaging (MRI) and echocardiography have a higher temporal resolution, these techniques have important other disadvantages. Due to the nature of MRI, patients with pacemakers can not be studied. Recently tissue synchronization imaging (TSI) with echocardiography has been described and validated. TSI portrays regional asynchrony on two-dimensional echocardiography by transforming the timing of regional peak velocity into colour codes, which allow immediate visual identification regional delay in systole by comparing the colour mapping of orthogonal walls [28]. However, echocardiography is highly user-dependent and therefore hampers the widespread use in a clinical setting. Attention should be paid to triggering of the acquisitions. Especially during pacing, the R-wave gating of the QRS complex may vary compared to gating during native cardiac rhythm. This may lead to alterations in triggering and subsequent timing of the phase curve. Although clinical value of phase analysis has been shown in patients with ischaemic cardiomyopathy, regional wall motion abnormalities, which are characterized by low amplitude, may lead to misinterpretation of the phase. Since three-dimensional phase analysis is currently not commercially available, only planar images can be used to assess ventricular synchrony. In planar images, the left anterior oblique projection causes over-projection of different areas with various contraction characteristics leading to underestimation of dyssynchrony of myocardial contraction. The use of left anterior oblique projection in

planar imaging implies that only mechanical activation of the septal, apical, lateral and inferior wall can be determined. The development of a technique that allows for three-dimensional phase analysis may overcome these drawbacks. Ventricular synchrony, expressed as the standard deviation of the phase histogram, has been related to heart rate [4]. An increase in pacing rate in a heart failure patient subjected to biventricular pacing diminishes diastolic filling time but may also cause a decrease in intravenricular and inter-ventricular synchrony which may be accompanied by a decrease in left ventricular ejection fraction (Fig. 5). This advocates the use of identical pacing rates during follow-up studies in patients with heart failure in whom the effect of biventricular pacing on ventricular synchrony is evaluated.

Conclusion and future perspectives Identification of responders and non-responders to biventricular pacing in heart failure is challenging. Up to 30% of the patients subjected to biventricular pacing may not benefit from this treatment. The QRS duration on the ECG at baseline is not a potent predictor of clinical outcome and has therefore limited value for the selection of patients. However, the degree of ventricular asynchrony as assessed by Fourier analysis of nuclear ventriculography may be more powerful to identify responders to resynchronization therapy. Since gated blood-pool planar nuclear ventriculography is non-invasive, highly reproducible and relatively easy to perform, it can be considered to be one of the most promising techniques to identify responders to resynchronization

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Ventricular dyssynchrony and gated blood-pool ventriculography Somsen et al. 111

Fig. 5

RR = 750 ms LVEF = 52% std = 24 IVD = 23 ms

0

90

180 270 Degrees

RR = 600 ms LVEF = 43% std = 26 IVD = 30 ms

360 0

90

RR = 500 ms LVEF = 36% std = 27 IVD = 45 ms

180 270 Degrees

360 0

90

180 270 360 Degrees

The effect of various RR intervals (ms) on ventricular synchrony in a patient with heart failure and left bundle branch block who is subjected to biventricular pacing. Inter-ventricular delay (dyssynchrony) increases with decreasing RR interval. RR = RR interval (Other abbreviations as in the legend to Fig. 1.)

therapy. This technique also enables the evaluation of the effect of biventricular pacing in patients with heart failure, which should be studied in clinical trials. In addition, optimization of resynchronization therapy, especially with respect to inter-ventricular delay and the position of the left ventricular pacemaker lead, will be possible using gated blood-pool ventriculography.

References 1

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Botvinick E, Dunn R, Frais M, O’Connell W, Shosa D, Herfkens R, et al. The phase image: its relationship to patterns of contraction and conduction. Circulation 1982; 65:551–560. Brateman L, Buckley K, Keim SG, Wargovich TJ, Williams CM. Left ventricular regional wall motion assessment by radionuclide ventriculography: a comparison of cine display with Fourier imaging. J Nucl Med 1991; 32:777–782. Vos PH, Vossepoel AM, Pauwels EK. Quantitative assessment of wall motion in multiple-gated studies using temporal Fourier analysis. J Nucl Med 1983; 24:388–396. Mancini GB, Peck WW, Slutsky RA. Analysis of phase-angle histograms from equilibrium radionuclide studies: correlation with semiquantitative grading of wall motion. Am J Cardiol 1985; 55:535–540. Botvinick EH, Frais MA, Shosa DW, O’Connell JW, Pacheco-Alvarez JA, Scheinman M, et al. An accurate means of detecting and characterizing abnormal patterns of ventricular activation by phase image analysis. Am J Cardiol 1982; 50:289–298. Links JM, Raichlen JS, Wagner Jr HN, Reid PR. Assessment of the site of ventricular activation by Fourier analysis of gated blood-pool studies. J Nucl Med 1985; 26:27–32. Raichlen JS, Links JM, Reid PR. Effect of electrical activation site on left ventricular performance in ventricular tachycardia patients with coronary heart disease. Am J Cardiol 1985; 55:84–88. Wendt III RE, Murphy PH, Clark Jr JW, Burdine JA. Interpretation of multigated Fourier functional images. J Nucl Med 1982; 23:715–724.

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Pieske B, Kretschmann B, Meyer M, Holubarsch C, Weirich J, Posival H, et al. Alterations in intracellular calcium handling associated with the inverse force–frequency relation in human dilated cardiomyopathy. Circulation 1995; 92:1169–1178. Fauchier L, Marie O, Casset-Senon D, Babuty D, Cosnay P, Fauchier JP. Reliability of QRS duration and morphology on surface electrocardiogram to identify ventricular dyssynchrony in patients with idiopathic dilated cardiomyopathy. Am J Cardiol 2003; 92:341–344. Yu CM, Lin H, Zhang Q, Sanderson JE. High prevalence of left ventricular systolic and diastolic asynchrony in patients with congestive heart failure and normal QRS duration. Heart 2003; 89:54–60. Tse HF, Lau CP. Long-term effect of right ventricular pacing on myocardial perfusion and function. J Am Coll Cardiol 1997; 29:744–749. Fauchier L, Marie O, Casset-Senon D, Babuty D, Cosnay P, Fauchier JP. Interventricular and intraventricular dyssynchrony in idiopathic dilated cardiomyopathy: a prognostic study with fourier phase analysis of radionuclide angioscintigraphy. J Am Coll Cardiol 2002; 40:2022–2030. Gregoratos G, Abrams J, Epstein AE, Freedman RA, Hayes DL, Hlatky MA, et al. ACC/AHA/NASPE 2002 Guideline Update for Implantation of Cardiac Pacemakers and Antiarrhythmia Devices-summary article: a report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines (ACC/AHA/NASPE Committee to update the 1998 Pacemaker Guidelines). J Am Coll Cardiol 2002; 40:1703–1719. Bradley DJ, Bradley EA, Baughman KL, Berger RD, Calkins H, Goodman SN, et al. Cardiac resynchronization and death from progressive heart failure: a meta-analysis of randomized controlled trials. JAMA 2003; 289:730–740. Bristow MR, Saxon LA, Boehmer J, Krueger S, Kass DA, De MT, et al. Cardiac-resynchronization therapy with or without an implantable defibrillator in advanced chronic heart failure. N Engl J Med 2004; 350:2140–2150. Cleland JG, Daubert JC, Erdmann E, Freemantle N, Gras D, Kappenberger L, et al. The effect of cardiac resynchronization on morbidity and mortality in heart failure. N Engl J Med 2005; 352:1539–1549. Breithardt OA, Stellbrink C, Kramer AP, Sinha AM, Franke A, Salo R, et al. Echocardiographic quantification of left ventricular asynchrony predicts an acute hemodynamic benefit of cardiac resynchronization therapy. J Am Coll Cardiol 2002; 40:536–545.

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112 Nuclear Medicine Communications 2006, Vol 27 No 2

19 Yu CM, Fung WH, Lin H, Zhang Q, Sanderson JE, Lau CP. Predictors of left ventricular reverse remodeling after cardiac resynchronization therapy for heart failure secondary to idiopathic dilated or ischemic cardiomyopathy. Am J Cardiol 2003; 91:684–688. 20 Auricchio A, Stellbrink C, Block M, Sack S, Vogt J, Bakker P, et al. for the Pacing Therapies for Congestive Heart Failure Study Group Effect of pacing chamber and atrioventricular delay on acute systolic function of paced patients with congestive heart failure. Circulation 1999; 99:2993–3001. 21 Nelson GS, Curry CW, Wyman BT, Kramer A, Declerck J, Talbot M, et al. Predictors of systolic augmentation from left ventricular preexcitation in patients with dilated cardiomyopathy and intraventricular conduction delay. Circulation 2000; 101:2703–2709. 22 Gasparini M, Mantica M, Galimberti P, Bocciolone M, Genovese L, Mangiavacchi M, et al. Is the left ventricular lateral wall the best lead implantation site for cardiac resynchronization therapy? Pacing Clin Electrophysiol 2003; 26:162–168. 23 Links JM, Douglass KH, Wagner Jr HN. Patterns of ventricular emptying by Fourier analysis of gated blood-pool studies. J Nucl Med 1980; 21:978–982.

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Original article

Is stress-only imaging practical when a 1-day stress–rest 99m Tc-tetrofosmin protocol is used? Andrew M. Cheetham, Victoria Naylor, Jean McGhie, Fabrice Ghiotto, M. Bashar Al-Housni and Andrew D. Kelion Objectives To investigate whether a significant number of normal 1-day stress–rest 99mTc-tetrofosmin myocardial perfusion scintigraphy (MPS) studies can be identified from the low-dose stress acquisition alone, and whether technical staff can find such studies reliably. Methods The supervising consultant and four technologists independently graded the stress acquisitions from 200 consecutive MPS studies using a five-point scale. Studies were classified as normal or abnormal according to the final clinical report based on the completed stress–rest protocol.

Conclusions Technologists could make the decision to avoid a resting study in up to 30% of patients following a normal low-dose stress acquisition. The supervising nuclear cardiologist would disagree in perhaps one in five cases, even when there was consensus between two or more technologists. These patients would suffer minor inconvenience by being recalled for a rest acquisition on a second day, but there would be important savings in time and radiation exposure for the majority. Nucl Med Commun 27:113–117

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:113–117

Results Between 31 and 62 studies (16–31%) were classified as definitely normal from the stress acquisition alone, of which 0–4 (0–9%) proved abnormal on the final report. Of stress studies graded definitely normal by each technologist, the consultant disagreed in 13–34% of cases. Of 78 stress studies graded definitely normal by at least one technologist, 6% turned out to be abnormal and the consultant disagreed in 33%. When there was agreement between at least two technologists (57 studies), the rates fell to 4% and 21% respectively.

Introduction Protocols for myocardial perfusion scintigraphy (MPS) using a 99mTc-labelled agent (sestamibi or tetrofosmin) involve separate stress and rest studies and are timeconsuming for both patients and nuclear medicine departments. Ideally stress and rest studies are performed on separate days (2-day protocol) so that the two studies are truly independent of each other, similar high doses of radiopharmaceutical can be used on each day to ensure optimal image quality and comparability, and there is ample opportunity to avoid an unnecessary rest study if the stress study is normal. Despite the advantages of a 2day protocol, many departments employ a 1-day protocol for convenience. A relatively small dose of radiopharmaceutical is used for the initial study, followed by a larger dose for the second study later the same day to swamp residual myocardial activity from the first injection. In the United Kingdom, 80% of 1-day 99mTc MPS studies employ a stress–rest order [1]. This offers the possibility of avoiding an unnecessary rest study if the stress study is normal, thereby saving a few hours of the patient’s time, reducing radiation exposure by a factor of 4, and freeing up

Keywords: stress-only, 1-day stress–rest protocol, tetrofosmin, myocardial perfusion scintigraphy, SPECT, technologist Department of Nuclear Medicine, Harefield Hospital, UK. Correspondence to Dr A.D. Kelion, Nuclear Medicine Department, Harefield Hospital, Hill End Road, Harefield, Middlesex UB9 6JH, UK. Tel: + 0044 (0)1895 826565; fax: + 0044 (0)1895 828880; e-mail: [email protected] Received 7 September 2005 Accepted 11 October 2005

gamma camera time for additional patients to be studied. However, in a busy department it is usually impossible for every stress acquisition to be reviewed by the reporting physician prior to the rest injection, and the decision to send a patient home may rest with technical staff. This decision may be particularly difficult because image quality for a low-dose stress study may be suboptimal and gating is impractical. Unless a significant number of unnecessary rest studies can be avoided using a stress–rest protocol, a rest–stress order would actually be more efficient because less delay is required between injections. We investigated whether a significant number of normal 1-day stress–rest 99mTc-tetrofosmin MPS studies can be identified from the low-dose stress acquisition alone, and whether technical staff are able to find such studies reliably.

Methods Patients and study design

The study population consisted of 200 consecutive patients referred for MPS in a busy nuclear medicine department (more than 2000 MPS studies per year). All

c 2006 Lippincott Williams & Wilkins 0143-3636

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114 Nuclear Medicine Communications 2006, Vol 27 No 2

patients were studied using a 1-day stress–rest 99mTctetrofosmin protocol. Clinical reports were written by a single experienced nuclear cardiologist (A.D.K.), based on clinical data, the stress test, and both the stress and gated rest SPECT images. Based on these reports, studies were classified as normal (with or without artefact) or abnormal. Six months later the nuclear cardiologist and four nuclear medicine technologists with varying levels of experience independently reviewed the 200 stress acquisitions. Readers were told the clinical indication for MPS and the results of the stress test, but did not have access to the resting study. Studies were graded on a five-point scale: 1, completely normal; 2, normal with minor artefact, re-injection not required; 3, probably normal with artefact, re-injection required; 4, probably abnormal; and 5, definitely abnormal. Protocol for myocardial perfusion scintigraphy

Patients were stressed using treadmill exercise or adenosine 140 mg
kg – 1
min – 1 over 6 min, as appropriate. 99m Tc-tetrofosmin (250 MBq) was administered intravenously 1 min prior to peak exercise or 3 min into the adenosine infusion. An ungated single photon emission computed tomography (SPECT) acquisition was obtained 45–60 min after tracer injection. At least 3 h after the stress injection, a further injection of Tc-tetrofosmin was given at the higher dose of 750 MBq. This was preceded by sublingual glyceryl trinitrate if the patient was known to have coronary artery disease or had a significant perfusion defect on the stress acquisition. A gated SPECT acquisition was obtained 2 h after tracer injection.

Table 1

Clinical details of the 200 patients

Characteristic

Number

Age (years) Sex Male

62 ± 11 120 (60%)

Indication for MPS Diagnostic Post-acute coronary syndrome Post-angiography Post-revascularization

85 23 30 62

(43%) (12%) (15%) (31%)

Stress for MPS Treadmill exercise Limiting chest pain Ischaemic ECG changes Adenosine

144 7 13 56

(72%) (5%) (9%) (28%)

MPS result Normal Fixed defect Reversible defect Mixed defect(s)

111 55 16 18

(56%) (28%) (8%) (9%)

MPS, myocardial perfusion scintigraphy; ECG, electrocardiogram.

MPS for diagnostic purposes, whilst 115 (58%) were already known to have coronary artery disease. One hundred and eleven (56%) MPS studies were reported as normal, with or without artefact, based on the complete stress–rest study, whilst 89 (45%) showed a perfusion defect.

99m

SPECT acquisitions

All studies were acquired on a Philips CardioMD gamma camera equipped with low energy high resolution collimators. A contoured 1801 orbit with 32 steps (45 s per step for ungated stress acquisition, 60 s per step for gated rest acquisition) was used, with a 64  64 matrix and an energy window of 140 keV ± 10%. Rest acquisitions were gated wherever possible, using 16 frames and an R–R window of ± 20%. Attenuation correction was not used. Stress and gated rest acquisitions were processed and displayed on a Philips Pegasys workstation using CedarsSinai AutoSPECT and AutoQuant software. Iterative reconstruction was performed, with a Butterworth filter (stress: cut-off 0.45, order 6.0; rest: cut-off 0.55, order 5.0). Acquisitions were reoriented to the standard orthogonal planes of the heart.

Results Clinical details of the 200 study patients are summarized in Table 1. Eighty-five (42%) patients were referred for

The five readers each identified between 31 and 62 studies (16–31%) as being definitely normal (grade 1 or 2, no resting injection required) from the stress acquisition alone (Table 2). For the two most experienced readers (consultant and technologist 1), none of these studies proved to be abnormal based on the final report; for the other readers, between 1 and 4 (3–9%) were abnormal. An important proportion of patients graded as 3 (probably normal with artefact, re-injection required) proved to have abnormal studies (10–29%). The majority of patients graded as 4 or 5 (probably or definitely abnormal) had abnormal studies (61–90%). In clinical practice, the reporting consultant would be the final arbiter of the requirement for a resting study. Of patients graded 1–2 by a technologist who could have been sent home without a resting study, 13–34% were graded 3–5 by the consultant and might have had to be recalled for a resting study on a second day (Table 3). The risk of misclassification of abnormal studies, or those requiring re-injection according to the consultant, decreased as consensus between the technologists increased (Table 4). Of 78 studies graded 1–2 by at least one technologist, 6% turned out to be abnormal and 33% would have required re-injection. The rates fell to 4% and 21%, respectively, for 57 studies graded 1–2 by two or more technologists.

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Stress-only imaging using a 1-day stress–rest

Table 2

99m

Tc protocol Cheetham et al. 115

Number of patients with an abnormal study in each stress-only category for each of the five readers Number of patients*

Stress-only category

Consultant

Technologist

Grade 1–2 3 4–5

Normal No rest study Probably normal Rest study Abnormal Rest study

n Abnormal n Abnormal n Abnormal

59 0 (0%) 57 13 (23%) 84 76 (90%)

1

2

3

4

57 0 (0%) 31 3 (10%) 112 86 (77%)

62 3 (5%) 52 15 (29%) 86 71 (83%)

31 1 (3%) 33 5 (15%) 136 83 (61%)

47 4 (9%) 50 14 (28%) 103 71 (69%)

*

n is the total number of patients in the group, and ‘Abnormal’ refers to the number of patients in that group who had an abnormal final study result.

Number of patients requiring a rest study (graded 3–5 by the consultant) in each stress-only category for each of the four technologists

Table 3

Number of patients*

Stress-only category

Technologist

Grade 1–2 3 4–5

Normal No rest study Probably normal Rest study Abnormal Rest study

n Rest n Rest n Rest

1

2

3

4

57 10 (18%) 31 20 (65%) 112 111 (99%)

62 15 (24%) 52 42 (81%) 86 83 (97%)

31 4 (13%) 33 16 (48%) 136 121 (89%)

47 16 (34%) 50 31 (62%) 103 71 (69%)

*

n is the total number of patients in the group, and ‘Rest’ refers to the number of patients in that group who required a rest study according to the consultant.

Table 4 Influence of increasing consensus between technologists on the likelihood of a study graded 1 or 2 being abnormal or requiring a rest study (graded 3–5 by the consultant) Number of technologists grading study 1 or 2 None One or more Two or more Three or more All four

Number of patients 122 78 57 44 18

Number abnormal (%) 84 5 2 1 0

(69%) (6%) (4%) (2%) (0%)

Number requiring rest study (%) 115 26 12 7 0

(94%) (33%) (21%) (16%) (0%)

Discussion In this study, technologists identified up to 30% of 1-day stress–rest 99mTc-tetrofosmin MPS studies as definitely normal, and not requiring re-injection at rest, from the low-dose stress study alone. Few such patients turned out to have an abnormal result based on the completed study, although as many as one in three were thought to need re-injection by the consultant. This could be improved to one in five if there was agreement between two or more technologists. Previous studies of the practicality and safety of stress-only MPS have relied on a high-dose stress study, usually with gating and/or attenuation correction, with interpretation performed by expert readers [2–4]. To our knowledge, this is the first study to investigate the everyday value of a low-dose stress-only acquisition in the hands of nonmedical readers of variable experience.

percent of these involve a 1-day protocol, with a stress–rest order in 80%. There are a number of technical problems associated with a 1-day stress–rest study. Severe inducible perfusion abnormalities are occasionally very prolonged following stress and can persist until the resting injection. Absolute myocardial perfusion is higher during stress than at rest, leading to a higher proportion of the injected tracer dose being taken up by the myocardium. The second high-dose injection is therefore less effective at swamping residual activity when a stress–rest order is used compared with a rest–stress order, which may lead to underestimation of defect reversibility. For these reasons, a significant delay is required between injections when a stress–rest order is used. A stress–rest order in a 1-day protocol has the one important advantage of allowing the resting study to be avoided if the stress study is normal. This saves time for both the patient and the department, whilst minimizing radiation exposure. A normal stress-only MPS study is as reassuring prognostically as a normal complete study [3]. In a study by Gibson and colleagues, 729 patients with a normal high-dose 99mTc-sestamibi stress-only attenuation corrected SPECT study were followed up. The hard cardiac event rate was 0.6% over 2 years, no higher than the rates quoted for a normal complete stress and rest study.

Advantages and disadvantages of a 1-day stress–rest protocol

Proportion of patients with a definitely normal stress-only study

In the UK, the majority of MPS studies are now performed using a 99mTc-based tracer [1]. Twenty-five

For the 1-day stress–rest order to be worthwhile, a significant proportion of normal studies would need to be

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116 Nuclear Medicine Communications 2006, Vol 27 No 2

identifiable from the stress study alone. Heller and colleagues [4] found that of 90 high-dose (1110 MBq) stress studies, 55% were normal when attenuation correction was used, similar to the 56% of studies which were normal in our study based on the final report. However, these workers found that only 20% of stress studies were definitely normal on uncorrected images, a lower figure than the 29% in our study if two technologists were in agreement. This may reflect the access to the clinical indication and stress data given to the readers in our study, reproducing the situation in everyday clinical practice. Worsley and colleagues [2] found a somewhat higher proportion of normal stress images (41%) amongst 148 patients undergoing a 2-day 99m Tc MPS study, but this probably reflects the exclusion of patients with a history of myocardial infarction. Reliability of reporting stress-only studies

In clinical practice, technical staff would have to be able to identify normal stress acquisitions reliably as a nuclear cardiologist is unlikely to be available throughout the day. In Worsley’s study, the resting acquisition did not alter the interpretation of any MPS study thought to have a normal stress-only acquisition. Similarly, in our study no patient with a stress acquisition graded as definitely normal (1–2) by the two most experienced readers turned out to have an abnormal complete study. Whilst the less experienced readers did make occasional errors of judgement (3–9%), this could be reduced by consensus (4% if two or more technologists agreed). Although they were unlikely to have an abnormal study, one in five patients with a definitely normal stress acquisition identified by at least two technologists could have expected to be recalled for a resting study after consultant review. This reflects the subjectivity involved in classifying studies, and in particular the fine line between grades 2 and 3. The failure to proceed to a resting study which is subsequently considered necessary by the reporting nuclear cardiologist is of little clinical consequence, as the patient can be recalled on a different day, but there may be significant inconvenience to patients and the department if this occurs frequently. Value of low-dose stress-only study in clinical practice

We have modelled the implications of our findings for own clinical service. We currently perform approximately 2000 1-day stress–rest 99mTc-tetrofosmin MPS studies each year. Five-hundred and eighty (29%) of the stress acquisitions would be normal based on agreement between two technologists, and the patients could be sent away without a rest study. The consultant would recall up to 122 (21%) for a rest study on a separate day, but this would still allow an additional 458 image acquisitions each year, or 229 whole patient studies.

In terms of radiation exposure, avoiding the resting study in a 1-day stress–rest protocol is even more beneficial than in a 2-day protocol. UK diagnostic reference levels for a 1-day 99mTc-based MPS study are 250 MBq followed by 750 MBq [5]. Avoidance of an unnecessary resting injection reduces a patient’s effective dose equivalent from approximately 10 mSv to only 2.5 mSv. This is particularly relevant given the high likelihood of longterm survival associated with a normal MPS study. Benefit of gating or attenuation correction

In our study it would have been impractical to gate the stress acquisitions because of the low tracer dose involved, and attenuation correction was not applied. In Heller’s study [4], the high-dose stress acquisition was gated and attenuation corrected, and images were read by 10 expert nuclear cardiologists. Attenuation correction, but not gating, increased the percentage of definitely normal stress-only studies substantially, from 20% to 55%. Whilst helpful to the experienced reader, attenuation correction makes additional demands and might be less suitable for technologist-only review of images. Limitations of the study

The nonmedical readers in this study did not routinely report MPS and received no specific training, and the definition of the thresholds between grades was left to an individual’s discretion. Moreover, the consultant made his assessment blind to that of the technologists, in contrast to the hierarchical sequence that would occur in clinical practice. These factors probably contributed to the variation in the proportion of patients graded 1–2 by different readers, as well as to the seemingly high proportions appearing to require recall by the consultant. It is likely that as a routine policy of avoiding unnecessary rest studies was implemented in clinical practice, feedback from the supervising consultant would lead to more uniformity between technologists and a lower rate of patient recall for a rest acquisition than our study predicts.

Conclusion We conclude that technologists can reasonably make the decision to avoid a resting study in up to 30% of patients following a normal low-dose stress acquisition. Accuracy can be improved if members of staff seek agreement from colleagues. Inevitably, a few patients would have to be recalled for a resting study on a second day, but this small inconvenience would be more than compensated by the savings in time and radiation exposure for the majority.

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methoxyisobutylisonitrile myocardial perfusion imaging. Eur J Nucl Med 1992; 19:441–444. Gibson PB, Demus D, Noto R, Hudson W, Johnson LL. Low event rate for stress-only perfusion imaging in patients evaluated for chest pain. J Am Coll Cardiol 2002; 39:999–1004.

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Tc protocol Cheetham et al. 117

Heller GV, Bateman TM, Johnson LL, Cullom SJ, Case JA, Galt JR, et al. Clinical value of attenuation correction in stress-only Tc-99m sestamibi SPECT imaging. J Nucl Cardiol 2004; 11:273–281. Administration of Radioactive Substances Advisory Committee (ARSAC). Diagnostic procedures – adult patients. 2002; 37–39.

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Original article

Early post-stress pulmonary uptake of 99mTc tetrofosmin during exercise (SPECT) myocardial perfusion imaging: Correlation with haemodynamic, perfusion and function parameters Panagiotis Georgouliasa, Nikolaos Demakopoulosb, Angelos Kontosb, Petros Xaplanterisc, Kostis Xydisa and Ioannis Fezoylidisa Objective To test the association of early post-stress lung/heart ratio (LHR) of 99mTc tetrofosmin radioactivity with gated-SPECT findings and angiographic results. Methods We studied 158 consecutive patients, with stress/rest 99mTc tetrofosmin myocardial SPECT and coronary angiography. Rest scans were obtained as gated SPECT and the left ventricular ejection fraction and end diastolic volume were calculated. To evaluate myocardial ischaemia, we calculated the summed stress score, summed rest score and summed difference score indices. For the LHR calculation, we acquired an anterior image, 4–6 min after radiotracer injection at stress; LHR was defined as mean counts/pixel in the lung region of interest divided by the mean counts/pixel in the myocardial region of interest. Results An early post-stress LHR value of 0.500 was defined as the upper normal limit. The most significant correlation (P < 0.001) was observed among early poststress LHR, summed stress score and the number of stenosed vessels. The incidence of multi-vessel coronary artery disease in the subgroup of patients with increased values of early post-stress LHR, was significantly higher than in the normal group (81% vs. 42%, P < 0.001).

Introduction A significant number of studies have elaborated on the clinical importance of increased pulmonary uptake of 201 Tl during myocardial perfusion imaging [1–7]. The quantification of lung uptake is usually estimated using the lung/heart ratio (LHR) of radioactivity (counts/pixel) in two regions of interest (ROIs): one in the lung (usually the left) and another over the left ventricle myocardium, delineated in the anterior images [1–4]. With 201Tl, the classic radionuclide used for this purpose, pulmonary uptake is assessed as an ancillary aspect of interpretation of myocardial scintigraphies. Increased post-stress lung concentration of 201Tl has been shown to be indicative of left ventricular dysfunction and to correlate with a larger number of abnormal scintigraphic segments; it is also linked to multi-vessel or severe coronary artery disease (CAD) [1–7]. The ratio of pulmonary to myocardial 201Tl

There was a significant difference (P < 0.001) of the early post-stress LHR value between patients with normal coronary arteries or one-vessel disease and patients with multi-vessel disease. Early post-stress LHR was an independent predictor of multi-vessel coronary artery disease (coefficient 1.85, SD 0.16, P < 0.001), with an incremental value for its identification. Conclusions Our results suggest that early post-stress Tc tetrofosmin LHR appears to be a useful index of extensive myocardial ischaemia dysfunction and multi-vessel coronary artery disease. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:119–126 99m

Nuclear Medicine Communications 2006, 27:119–126 Keywords: lung/heart ratio, technetium-99m tetrofosmin a

Department of Nuclear Medicine, University Hospital of Larissa, Departments of Nuclear Medicine and cCardiology, NIMTS Hospital, Athens, Greece.

b

Correspondence to Dr Panagiotis Georgoulias, 15 Kritis St. 19009 Rafina, Greece. Tel: + 0030 2294 026201 or + 0030 2410 682052; fax: + 0030 2294 078400 or + 0030 2410 670117; e-mail: [email protected] or [email protected] Received 5 September 2005 Accepted 15 November 2005

activity during myocardial perfusion scintigram, in combination with either treadmill exercise testing or pharmacological stress methods, is also considered a valuable prognostic index [3–7]. Although technetium-based radiopharmaceuticals have been widely used for several years, their limited use in lung uptake assessment has been regarded as a potential drawback. Moreover, previous studies have published controversial results regarding the clinical value of pulmonary uptake measurements with 99mTc sestamibi or 99mTc tetrofosmin and also about the method of calculation [8–21]. 99m

Tc tetrofosmin is an interesting alternative radiotracer for myocardial perfusion scanning, as it combines the exceptional physical properties of 99mTc with easy and fast preparation [22–25].

c 2006 Lippincott Williams & Wilkins 0143-3636

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120 Nuclear Medicine Communications 2006, Vol 27 No 2

The diagnostic and prognostic value of myocardial single photon emission computed tomography (SPECT) using this agent has already been well established, although a few studies have utilized the usefulness of lung uptake as an ancillary sign [18–27]. The aim of this study was to determine the clinical significance of early, post-stress, pulmonary/heart ratio of 99m Tc tetrofosmin activity during exercise/rest SPECT (gated tomo) myocardial perfusion scintigraphy.

Materials and methods Population

The study population consisted of 158 consecutive patients (111 males and 47 females), ranging in age from 33 to 84 years (mean ± standard deviation (SD): 58.3 ± 9.8), who were properly referred (based on their data), between April 2002 and December 2004, for a symptom-limited exercise testing combined with a SPECT myocardial perfusion imaging. One hundred and forty-six patients (92.4%) had undergone a recent coronary angiography within a 3 month period or underwent an angiography within 3 months. Of these, 19 patients (13%) had normal angiography, 45 (30.8%) had single-vessel disease, 43 (29.5%) had twovessel disease and 39 (26.7%) had three-vessel disease. In our study, we excluded patients whose tracer pulmonary uptake or myocardial perfusion imaging and function might have been affected by factors other than myocardial ischemia. We therefore excluded patients with left bundle branch block, a history of myocardial infarction or an irreversible defect on their scintigram, congenital or valvular heart disease, cardiomyopathy, pulmonary disease and those patients with an implanted pacemaker. We also excluded patients taking digoxin (due to its prolonged effect) and those with a contraindication or inability to perform treadmill testing or achieve a satisfactory exercise level because of a non-cardiac condition (e.g., sciatica, neuropathy, disability). In addition, patients were excluded if gated-SPECT data could not be obtained or if the images had excessive abdominal activity and/or motion artifacts interfering with assessment of the perfusion data and radiotracer lung uptake [28,29]. Concomitant medications that could possibly influence patient performance in exercise testing and related variables were temporarily discontinued; beta blockers and calcium channel antagonists were discontinued 48 h prior to study initiation as well as for the duration of the study. Similarly, nitrates were discontinued 24 h before and during the study. Any patient whose medication was not properly intermitted was excluded from the study.

Before testing, patients were asked to give informed consent according to the hospital ethics committee guidelines. Then, a structured interview was conducted from which data were collected on symptoms, medications, previous cardiac events, lung disease, coronary risk factors and cardiac/non-cardiac diagnoses. Hypertension was defined as a systolic blood pressure of 140 mmHg or greater at rest and/or a diastolic blood pressure of 90 mmHg or greater at rest, or treatment with antihypertensive medication. Diagnoses of diabetes mellitus and lipid disorder were derived from the interviews with the patients and the use of corresponding medications. Obesity was confirmed as a body mass index value of 30.0 or greater (calculated as weight in kilograms divided by height in metres squared). Prior to the study, patients were also given written directions on radioprotection. To determine the normal value of early post-stress LHR we considered a separate reference (normal) group of patients without angina or other typical symptoms of cardiovascular disease, normal exercise testing, normal myocardial perfusion and function (left ventricular ejection fraction function = 55%, end diastolic volume = 180 ml), and no evidence of coronary artery disease (referring to those who had undergone coronary angiography) or use of cardioactive medication. The reference group consisted of 40 patients (16 men and 24 women) ranging in age from 31 to 60 years (mean age 40.17 ± 12.21 years). Exercise testing

After discontinuing cardioactive medication and undertaking a fast for 6–12 h, patients underwent symptomlimited treadmill exercise testing (Bruce protocol). Data on symptoms and estimated workload in metabolic equivalents (METs) were obtained; functional capacity in METs was estimated using standard tables [30]. As a criterion of ischaemic ST-segment response, we considered greater than 1 mm horizontal or down-sloping ST-segment depression, 80 ms after the J point or more than 1 mm of additional ST-segment rise in leads without pathological Q waves. Chronotropic response was defined as the percentage of maximal age-predicted heart rate achieved at peak exercise (peak heart rate/(220 – age)); a value of 85% or greater was considered to be normal. SPECT myocardial perfusion imaging

Myocardial perfusion studies were performed with 99mTc tetrofosmin (Myoview; Amersham, UK). Scintigraphies were obtained after the intravenous injection of the radiotracer (dose, 185–296 MBq; based on weight), 1– 2 min before the cessation of exercise. Twenty minutes after administration of radiopharmaceutical, 250 ml of

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Early post-stress pulmonary uptake of

milk (3.5% fat) was ingested for optimum excretion of the tracer by the gallbladder and acquisition started 35– 45 min after the administration. Four hours after the fist injection, resting 99mTc tetrofosmin SPECTwas obtained, using a 555–888 MBq dose (based on weight). Images were acquired from 451 right anterior oblique to the 451 left posterior oblique position in step-and-shoot mode (32 projections, matrix 64 64, pixel size 6.8 mm, 30 s per projection for stress and 50 s per projection for rest scintigrams). The high-count rest scans were acquired as gated-SPECT studies (eight frames per cardiac cycle) and the left ventricular ejection fraction (LVEF) and end diastolic volume (EDV) were calculated using Sopha Medical Vision software. The acquisitions were obtained with a two-headed Sophy camera equipped with lowenergy, parallel-hole, high-resolution collimators. The energy window was set at ± 10% symmetrically along the 140 keV photopeak and data processing was accomplished with a Butterworth filter (cut-off 0.35, order 4.0). We performed polar and three-dimensional mapping (transient ischaemic dilation index (TID) was also calculated) in all studies. For interpretation of the SPECT scans, the myocardium of the left ventricle was divided into 17 segments according to previous reports [31,32]. Two independent experienced observers blindly evaluated the reconstructed images, the polar maps and the three-dimensional images of both stress and rest studies; this was done by scoring the uptake in each of the 17 regions, using a five-point scoring system (0, normal uptake; 1, mildly reduced uptake; 2, moderately reduced uptake; 3, severely reduced uptake; and 4, no uptake) [31,32]. If counts were reduced in a segment, and this was judged to be the result of attenuation artefact, the score was 0 [33]. Ischaemia was considered in every region with an uptake higher than 0 at stress imaging and a reduction of the score by at least one unit at rest. Finally, the ‘summed stress score’ (SSS) and ‘summed rest score’ (SRS) were obtained by adding the scores of the segments at stress and rest studies. In case of discordance between the two observers, the view of a third observer was requested and the disagreement was resolved by consensus [31–33]. Subsequently, a ‘summed difference score’ (SDS) was estimated by subtracting the SRS from the SSS to assess defect reversibility [31–33]. Studies with an SSS equal or lower than 2 were considered normal.

99m

Tc tetrofosmin Georgoulias et al. 121

Fig. 1

(a) Normal early post-stress lung/heart ratio (0.391) in a 52-year-old woman without coronary artery disease. (b) Elevated early post-stress lung/heart ratio (0.684) in a 64-year-old man with three-vessel disease (RCA 99%, LCX 95%, LAD 70%).

the myocardium (at a distance of at least 3 pixels above the anterolateral wall – ‘lung ROI’) and the other, manually drown irregular ROI, including the whole myocardial activity of the left ventricle (‘myocardial ROI’) [12,13] (Fig. 1). The pulmonary/heart ratio was determined as the mean counts/pixel in the lung ROI divided by the mean counts/ pixel in the myocardial ROI [1,4–9,12,13,18,19]. Each LHR value was derived as the average value of the calculations of the two observers. Cardiac catheterization

Quantification of pulmonary uptake

We acquired early anterior planar images (1000 kcounts, matrix 256  256), 4–6 min after radiotracer injection at stress. We then delineated two ROIs: one square, 15  15 pixels, placed over the left mid-lung area at the vicinity of

Cardiologists, based on patient data, requested cardiac catheterization studies. One experienced observer ‘blindly’ interpreted all the coronary arteriographies. A stenosis of the vessel lumen greater than 50% was considered haemodynamically significant, while the

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122 Nuclear Medicine Communications 2006, Vol 27 No 2

presence of a stenosis in the left mainstem was considered equivalent to a two-vessel disease. Statistical analysis

Continuous variables are presented as mean ± SD. Differences between groups were estimated by using the chi-squared test (Yates’ correction), ANOVA, Student’s t-test and the Wilcoxon rank sum test (when groups were not normally distributed). To determine the correlation between two variables, we used the Pearson correlation coefficient or the Spearman rank correlation coefficient (for groups not normally distributed). In addition, after adjusting for any confounding factors, we fitted a multivariate regression model to assess the relationship between the value of early post-stress LHR (response) and myocardial perfusion SSS, SRS and SDS, the presence of multi-vessel CAD and LVEF. We also fitted a binary logistic regression model to estimate the effect of the value of the other variables on the probability of normal early post-stress LHR and normal SSS, and a number of variables on the prediction of multivessel CAD. We calculated the normal value of early post-stress LHR by using the formula ‘mean value ± 2SD’, taking into consideration the values of the normal group [34]. All statistical analyses were performed using Minitab (Release 13) statistical software (http://www.minitab.com); the significance level was set at 0.05 (P < 0.05), for all analyses.

Results There were 158 patients who constituted the study population. There was a very good inter-observer agreement in the evaluation of myocardial scintigrams (r = 0.86, P < 0.0001) and the calculation of LHR (r = 0.82, P < 0.0001). Combining all the data (clinical, exercise testing, gated SPECT) apart from early post-stress LHR value, multivessel (two-vessel or three-vessel) CAD was detected with a sensitivity of 87% and a specificity of 78%, while the positive and negative predictive values were 84% and 82%, respectively. The mean value of early post-stress LHR was 0.519 ± 0.041, while the mean value of early post-stress LHR in the normal group was 0.426 ± 0.036. Thus, we considered 0.500 as the upper normal value. For simplicity, this was derived as 0.426 + (2  0.036) = 0.498, or approximately 0.500. There was a statistically significant (P < 0.001) difference in the early post-stress LHR value between the study population and the normal group.

Seventy-eight patients (49%) had an abnormal value of early post-stress LHR. Patients with an abnormal value of early post-stress LHR were older and more likely to be obese, with hypertension, diabetes mellitus or hyperlipidaemia. In addition, these patients were more likely to be males, smokers, and taking cardioactive medications (beta blockers, calcium channel antagonists, nitrates). The main characteristics of the patients according to the value of their early post-stress LHR (normal or abnormal value) are presented in Table 1. The mean value of early post-stress LHR was 0.423 ± 0.038 in the subgroup with normal angiography (19 patients), 0.432 ± 0.029 in patients with one-vessel disease (45 patients), 0.538 ± 0.026 in patients with twovessel disease (43 patients) and 0.552 ± 0.034 in patients with three-vessel disease (39 patients, Fig. 2). There was a significant difference (P < 0.001) between the mean value of early post-stress LHR in the subgroup with normal angiography or patients with one-vessel disease and patients with multi-vessel disease (82 patients in total). On the other hand, the difference of early poststress LHR between normal population (referred to the coronary arteriography) and patients with one-vessel disease, as well as between those with two-vessel and three-vessel disease, was not statistically significant. A statistically significant positive correlation was found between the values of early post-stress LHR and SSS (r = 0.60, P < 0.001) and also between early post-stress LHR and the number of stenosed coronary vessels (r = 0.56, P < 0.001), while a significant negative correlation was found between early post-stress LHR and LVEF (r = – 0.57, P < 0.001). Moreover, a considerable positive correlation was found between early post-stress LHR and SDS (r = 0.53, P < 0.001). In addition, a weak positive correlation was reckoned between the early post-stress LHR and SRS (r = 0.12), and TID and early post-stress LHR (r = 0.14), but the results were not significant (P > 0.05). Table 1 Characteristics of the study group according to early poststress lung/heart ratio (LHR) Characteristic Number of patients Age (years) Sex (male), no. (%) Obesity, no. (%) Smoking, no. (%) Hypertension, no. (%) Diabetes, no. (%) Lipid disorder, no. (%) Use of beta blockers, no. (%) Use of calcium channel antagonists, no. (%) Use of nitrates, no. (%)

Normal (LHR r 0.500)

Abnormal (LHR > 0.500)

80 50 ± 14 51 (64%) 27 (34%) 18 (23%) 30 (38%) 14 (18%) 38 (48%) 16 (20%) 17 (21%) 13 (16%)

78 62 ± 15*** 66 (85%)* 42 (54%)* 41(53%)*** 47(60%)* 31 (40%)** 61 (78%)*** 36 (46%)** 33 (42%)* 32 (41%)**

*

P < 0.05, P < 0.01, P < 0.001.

**

***

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Early post-stress pulmonary uptake of

Fig. 2

Tc tetrofosmin Georgoulias et al. 123

Table 2 Logistic regression model: multi-vessel coronary artery disease with summed stress score (SSS), lung/heart ratio (LHR), left ventricular ejection fraction (LVEF), transient ischaemic dilation index (TID) and outcome of exercise testing

0.6 Early post-stress LHR

99m

0.5 0.4 0.3 0.2

Predictor

Coefficient

SE Coefficient

P-value

Constant SSS LHR LVEF TID Exercise

– 18.031 0.6272 27.60 0.02923 0.423 – 0.7123

9.171 0.1692 13.73 0.07023 4.478 0.8322

0.049 0.000 0.044 0.677 0.925 0.392

0.1 Table 3

0 Normal angiography

1-vessel disease

2-vessel disease

3-vessel disease

Mean value ( ± SD) of early post-stress lung/heart ratio, in patients with normal angiography, one-vessel disease, two-vessel disease and threevessel disease.

After adjustments were made for age, gender, smoking, the presence or absence of hypertension, diabetes, obesity, lipid disorder; the use or non-use of cardioactive medications (beta blockers, calcium channel antagonists, nitrates), resting heart rate, exercise duration, maximal systolic blood pressure, METs, double product, angina and abnormal ST response during exercise testing and performing a multivariate regression analysis, the strongest correlation was between the values of early poststress LHR and SSS (coefficient, 7.19; SE, 1.62; P < 0.001) and also between early post-stress LHR and the number of diseased vessels (coefficient, 4.08; SE, 1.46; P < 0.001). The associations between the values of early post-stress LHR and SDS, LVEF were weaker (P = 0.031, P = 0.017). We fitted a binary logistic regression model with multivessel CAD as the outcome variable and SSS, LHR, LVEF, TID and exercise testing outcome as the predictive variables (Table 2). From the results, we deduced that only SSS and LHR are statistically significant in predicting multi-vessel CAD. After fitting binary logistic regression analysis, where the response was the probability of normal early post-stress LHR, the results confirmed that the SSS is the most significant predictive variable of this probability (coefficient, 3.92; SE, 0.56; P < 0.001) while LVEF was also an independent predictive variable (coefficient, – 1.97; SE, 0.24; P < 0.01). Considering the probability of normal perfusion imaging (SSS r 2) as the response, the early post-stress LHR was an independent predictor (coefficient, 0.82; SE, 0.04; P < 0.01). In addition, considering the probability of multi-vessel disease as the response, early post-stress LHR value was an independent predictor (coefficient, 1.85; SE, 0.16; P < 0.001). Moreover, the early post-stress LHR added incremental value to clinical, exercise testing, and myocardial perfusion and

Exercise performance and scintigraphic data

Data Resting heart-rate (beats/min) Exercise duration (min) Maximal heart-rate (beats/min) Maximal systolic blood pressure (mm Hg) Metabolic equivalents (METs) Double product (  100) Angina during exercise testing, no. (%) Abnormal ST-segment response, no. (%) Abnormal chronotropic response, no. (%) Myocardial perfusion SSS Myocardial perfusion SRS Myocardial perfusion SDS TID index LVEF (%) EDV (%)

Normal (LHR r 0.500)

Abnormal (LHR > 0.500)

69 ± 12 11.6 ± 3.2 156 ± 14 202 ± 18 11.3 ± 2.2 312 ± 34 7 (9%) 13 (16%) 17 (21%) 5.3 ± 1.8 2.0 ± 1.2 4.6 ± 0.9 1.04 ± 0.19 56 ± 6 153 ± 14

76 ± 13** 8.4 ± 2.8*** 108 ± 19*** 161 ± 22*** 8.9 ± 2.6*** 237 ± 42*** 28 (36%)*** 32 (41%)** 40 (51%)** 15.8 ± 2.7*** 2.2 ± 1.3 13.1 ± 1.7*** 1.09 ± 0.17 45 ± 7*** 182 ± 16***

*

P < 0.05, P < 0.01, *** P < 0.001. Abbreviations as in Tables 1 and 2. **

function data, for the identification of patients with multi-vessel disease. Specifically, if the cut-off point of early post-stress LHR value was set at 0.500, the sensitivity, positive and negative predictive value to detect multi-vessel CAD improved from 87%, 84% and 82% to 94%, 85% and 91%, respectively, while the specificity did not change considerably (78%). Patients with a normal value of early post-stress LHR had a significantly better performance during exercise testing and better gated-SPECT results, as compared to those who had an abnormal value (Table 3). Furthermore the incidence of multi-vessel disease (regarding the patients who underwent a coronary angiography) in the subgroup with increased values of early post-stress LHR was significantly higher (81% vs. 42%, P < 0.001).

Discussion Many studies have utilized the calculation and clinical evaluation of the pulmonary uptake of radiotracers during stress/rest myocardial perfusion imaging, using either 201 Tl or 99mTc sestamibi [1–17]. On the other hand, only a few reports have investigated whether the lung-tomyocardial ratio using 99mTc tetrofosmin also provides similar diagnostic and prognostic information. The calculation of lung uptake on myocardial perfusion scintigrams with 201Tl provides important additional

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124 Nuclear Medicine Communications 2006, Vol 27 No 2

diagnostic and prognostic data. Increased pulmonary concentration of 201Tl during stress myocardial imaging has been shown to be an index of left ventricular dysfunction and/or severe CAD. The results of assessing 99mTc sestamibi pulmonary-toheart ratio of activity on conventional images obtained 30–60 min after stress are ambiguous, while a few studies have suggested that measuring lung uptake of 99mTc sestamibi on immediate post-stress images may be more valuable [8–17]. In addition, the methodology for calculating LHR among investigators varies; for example, a large ROI that enclosed the entire left ventricle or most of the left ventricle respectively and a fixed-size ROI in the left lung [12,13]. Other widely used methods are the following: a transmural segment of the myocardium is outlined containing the area of peak counts and a crescenting ROI placed over the left lung or alternatively, a fixed small rectangular ROI placed over maximal myocardial and left lung activity [8,11,14,16,17]. Our method for calculating 99mTc tetrofosmin LHR is similar to that used by Choy et al. [12] and Patel et al. [13]. This approach may underestimate the counts from the heart, if severe defects are present, thus potentially overestimating the LHR value. On the other hand, this approach avoids problems related to hot spots or other artefacts that could have erroneous effects on LHR. We believe that our method is more representative of pulmonary and heart radioactivity, especially in cases with regional myocardial ischaemia. Other studies report comparable results with ours, using Tc sestamibi as the agent for myocardial imaging [24,26,35]. They have found that increased pulmonary to myocardial ratio calculated on images obtained almost immediately after stress (exercise or vasodilation) was associated with severe scintigraphic abnormalities and angiographic findings (mainly three-vessel disease or stenoses in the left mainstem), in concordance to 201Tl post-stress lung uptake [8,11,14,16]. On the other hand, conflicting results have been reported about the relation between the values of LHR measured in the delayed images and the presence of extensive myocardial ischemia or severe CAD [10,12,13,15].

99m

99m

Tc tetrofosmin The correlation we found between pulmonary uptake and left ventricular function variables is comparable to other published results [8,9,12–14]. The strong correlation we found between the early post stress LHR and the haemodynamic and myocardial perfusion variables is in general agreement with other published data [8,12–16]. On the contrary, Saha et al. [10] mention that increased lung uptake of MIBI is not related to indexes of exercise testing, left ventricular dysfunction or perfusion abnormalities.

In our study, patients with a normal early post-stress LHR value generally had a better performance in the treadmill testing than those with an abnormal value (higher peak heart rate, better chronotropic response, maximal systolic blood pressure, double product, and METs). A lower percentage of patients reported angina or had an abnormal ST-segment response during exercise testing, compared to the subgroup with an abnormal value. In addition, patients with a normal value of early post-stress LHR had significantly better myocardial perfusion (as it is estimated via the myocardial perfusion SSS and SDS) and function (higher mean value of LVEF, lower mean value of EDV) and better angiographic results (lower incidence of multi-vessel disease). The above-mentioned data are most probably related to the influence of ischaemia in the early post-stress LHR value and are generally analogous with other published data, while other investigators did not find similar results using 99mTc sestamibi as the radiotracer [8,10,11]. The higher value of early post-stress LHR in patients with multi-vessel disease as well as the higher frequency of multi-vessel disease among patients with abnormal LHR values, and the inverse correlation between 99mTc tetrofosmin pulmonary uptake and LVEF are generally in agreement with other published results, while Tsou et al. [18], Tanigaki et al. [19] and Okajima et al. [20] have also reported the incremental value of 99mTc tetrofosmin LHR compared to conventional myocardial perfusion imaging for the detection of multi-vessel disease. In general, we believe that the variability of specific values reported in various studies could be presumably attributed to technical differences such as time of imaging, stress method and method of delineating ROIs. 99m

Tc tetrofosmin is cleared rapidly from the lungs, so we calculated the LHR values in early post-stress images instead of delayed images and in our opinion this could be a reason which explains the significant correlation we found between the early post-stress LHR and the myocardial perfusion and function parameters as well as between the early post-stress LHR and the presence of multi-vessel CAD. The mechanism of lung uptake with 99mTc labelled myocardial perfusion tracers has not been directly established, but it is reasonable to consider that it shares similarities with 201Tl pulmonary uptake. Our findings that CAD severity, scintigraphic ischaemic abnormalities and clinical-exercise data of myocardial ischaemia have a considerable correlation with stress LHR, support the role of ischaemic left ventricular dysfunction with subsequent pulmonary congestion-increased pulmonary vascular transit time as previously suggested for thallium (although additional factors maybe also important) [12].

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Early post-stress pulmonary uptake of

Our study has some potential limitations. First, we did not perform a comparison between different methods for measuring LHR values using various ROIs (large ROIs that enclose the entire left ventricle, small ROIs over the myocardial area of peak counts, fixed-size ROIs placed over maximal myocardial and left lung activity, crescentic ROIs over the left lung, for example), although we did not find such a comparison in other studies too. Second, we did not calculate LHR values in delayed stress or postrest images, considering that early post-stress LHR measurements are most representative. Third, the early post-stress acquisition might need a modification of the daily schedule, especially in departments where many examinations are carried out. An additional short period under the camera would be required, thus leading to time implications in a very busy centre. However, this extra acquisition can also be obtained with a planar camera.

Conclusions: clinical implications In conclusion, our study suggests the clinical value of routine pulmonary uptake measurements during 99mTc tetrofosmin myocardial scintigram, in immediately poststress images. This ancillary scintigraphic sign appears to be associated with clinical and exercise data of myocardial ischaemia and CAD. In an era where continuous effort is made to derive as many elements as possible from the examinations, the calculation of early post-stress LHR during myocardial perfusion imaging could provide additional information and it may contribute to the selection of patients who are in need of coronary angiography. It seems that early poststress LHR can add incremental value in detecting multivessel CAD, although its value, as an additional criterion required to improve the decision making process, needs further investigation.

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Our conclusion that LHR has incremental predictive value could be strengthened by the prospective application of our findings to a large cohort of patients, thus justifying the routine measurement of LHR. Finally, the prognostic implication of increased 99mTc tetrofosmin lung uptake and the comparison with other prognostic factors need to be evaluated and compared to those of 201Tl in a large cohort of patients.

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20

Acknowledgements The authors thank Dr Alexia Zaganides for dealing with the medical statistics; Sokratis Kapikos, Stauroula Giannakou and Tatiana Raco for technical assistance; and Rena Roboti for secretarial assistance.

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Kurata C, Tawarahara K, Taguchi T, Sakata K, Yamazaki N, Naitoh Y. Lung thallium-201 uptake during exercise emission computed tomography. J Nucl Med 1991; 32:417–423.

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Original article

A model that accounts for the interdependence of extent and severity in the automatic evaluation of myocardial defects Henrik Hussein El-Alia, John Palmera, Lars Edenbrandtb,c and Michael Ljungberga Background The extent and severity are two important parameters when describing a regional defect in myocardial single-photon emission computed tomography (SPECT) perfusion imaging. Intuitively, these two parameters should be independent of each other, but we have shown in a previous study that there is an interdependence. This interdependence has been investigated in two commercially available analysis programs (AutoQUANT and 4D-MSPECT) using Monte Carlo-simulated SPECT data. Methods An anthropomorphic digital computer phantom and a Monte Carlo program were used to generate SPECT data. Several defects of different volumes and lesion activity uptake reductions were simulated and evaluated. Comparison of the measures of extent and severity with their corresponding known values led to the development of a correction model based on least-squares parameter estimation. This model was then applied to a test group consisting of 10 different computer-simulated patients. Results Our results showed that the interdependence was reduced significantly for most of our test cases after applying the correction method. The application of the correction model to the test group demonstrated that the model was robust with respect to different patient

Introduction The investigation of regional myocardial perfusion with single-photon emission computed tomography (SPECT) is a widely used method to diagnose ischaemic heart diseases [1–5]. The common procedure is to compare stress and rest images to determine regional reductions in blood flow. Commercially available automatic quantification programs are an important aid in the interpretation of the images [6], and AutoQUANT and 4D-MSPECT are two examples of such programs [7,8]. On the basis of input from reconstructed short-axis images, these programs process and display results as a combination of stress and rest perfusion images by comparing the regional count variation in a polar image of the heart with a corresponding sex-specific database of normal perfusion data. A clinical report from a myocardial SPECT investigation may require the determination of the extent of the defect, the severity of the defect and its location. The extent in this context is a measure of the

geometries. A further test with projections that simulated a perfect SPECT system revealed that the interdependence between the extent and severity was not caused by the limited spatial resolution of the SPECT system, but rather the inherent design of the algorithms of the analysis programs. Conclusions A model has been developed to take into account the interdependence of the extent and severity. c 2006 Lippincott Williams Nucl Med Commun 27:127–135
& Wilkins. Nuclear Medicine Communications 2006, 27:127–135 Keywords: correction model, extent, lesion, Monte Carlo, myocardial perfusion, sestamibi, severity. a Department of Medical Radiation Physics, Clinical Sciences, Lund, Lund University, bClinical Sciences, Malmoe, Lund University and cDepartment of Clinical Physiology, Gothenburg University Hospital, Gothenburg, Sweden.

Correspondence to H. Hussein El-Ali, PhD, Department of Medical Radiation Physics, Clinical Sciences, Lund, Lund University Hospital, SE-221 85 Lund, Sweden Tel: + 46 46 178543; fax: + 46 46 178540; e-mail: [email protected] Sponsorship: This work was supported by the Medical Faculty of Lund University, Lund, Sweden. Received 11 August 2005 Accepted 22 November 2005

volume of the defect, and is often expressed as a percentage of the whole left wall volume. The severity of the defect is a measure of the decrease in blood perfusion [9], and is often expressed as the number of standard deviations relative to the corresponding region for the heart data in the normal database. In order to understand how AutoQUANT and 4DMSPECT calculate the severity and extent, we can consider the following definitions. The severity at location i in the polar map is equal to (mi – Ci)/si, where mi and si are the mean number of counts and standard deviation at location i in the sex-specific normal database, respectively, and Ci is the count level in the patient at location i. The extent of a defect is defined as the ‘area’ of myocardial surface, where, as a result of the defect, the severity falls below a predetermined threshold. Pixel severities may be averaged within a given sector to produce a severity value for the sector. If the extent of a

c 2006 Lippincott Williams & Wilkins 0143-3636


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128 Nuclear Medicine Communications 2006, Vol 27 No 2

defect is zero, the severity can also (formally) be assigned a value of zero. Both the extent and severity measurements thus depend on the predetermined threshold, which may be region-specific, as well as being differently defined by the two analysis programs considered here. We have shown in a previous study [10], and also here (Figs. 1 and 2), that the two concepts of extent and severity, as obtained from SPECT images, are dependent on each other. The reason for the interdependence was not explicitly investigated in our previous study. The aims of this study were therefore three-fold: firstly, to investigate how the calculated severity could be influ-

enced by the true defect size; secondly, to determine whether we could develop a method to correct for the interdependence between the extent and severity of the defect; thirdly, to validate the correction method for different NCAT [11] patient configurations to investigate the robustness of the method.

Materials and methods We based our investigation on Monte Carlo simulations as this methodology allows for direct comparison between the calculated results of the severity and extent and the initial values known a priori. Realistic SPECT images were obtained by the combination of a realistic digital

Fig. 1 100LAUR

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Top: extent and severity given by AutoQUANT as a function of the true extent for different locations (LAD, left anterior descending coronary artery; RCA, right coronary artery; LCx, left circumflex coronary artery) and values of the lesion activity uptake reduction (LAUR). Bottom: corresponding values given by 4D-MSPECT.

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An extent–severity interdependence correction model El-Ali et al. 129

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Extent and severity for different defect sizes versus lesion activity uptake reduction (LAUR) values. The graphs in the top row show the values given by AutoQUANT and those in the bottom row the values given by 4D-MSPECT. LAD, left anterior descending coronary artery; RCA, right coronary artery; LCx, left circumflex coronary artery.

computer phantom, which allows for accurate heart motions and precise definitions of different defects, and an accurate SPECT camera simulation program, which provides realistic simulation of the photon trajectories towards the scintillation camera. The simulated SPECT projections were evaluated in exactly the same manner as for a real patient, using the same clinical software and instrument settings. The advantage of this methodology is that the definition of a defect in terms of the true extent and lesion activity uptake reduction (LAUR) is very precise. Hence, measured values of the extent and

severity (the latter being the counterpart of LAUR) can be directly compared and the accuracy established. The computer phantom

We used the realistic NCAT computer phantom developed by Segars et al. [11]. This phantom has been developed from accurate computed tomography and magnetic resonance images of humans, and is therefore anatomically accurate. It also includes realistic heart motions and many of the major organs in the body. In this study, we simulated a male patient with a heart size that

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was ‘normal’ according to the NCAT definitions, which means a heart with a long axis of 9.49 cm, a short-axis radius of 2.9 cm, leading to a left ventricle wall volume of 174 ml. We included image blurring due to heart motion by averaging phantom images created over eight segments of the cardiac cycle. In order to mimic a clinically realistic study, the activity concentrations of the other organs in the phantom were set to match a 3 h postadministration 99mTc-sestamibi distribution using previously published data [12] and by visually comparing simulated projections with several clinical patient cases. Defects were defined in the three regions of the heart commonly assigned to the left anterior descending coronary artery (LAD), the right coronary artery (RCA) and the left circumflex coronary artery (LCx). In each of these three regions, we defined defects with true defect volumes ranging from 8 to 30 ml and, for each of these defect sizes, we used LAUR values ranging from 20 to 100% in steps of 20% (where 100% denotes the absence of activity uptake). From the general definition of the extent (i.e. the fraction of defect volume of the left ventricle wall volume), the defect volume was expressed as a percentage ranging from 5 to 17% in the three heart regions. These defect volumes are denoted here as the true extent, which was compared with the corresponding measured extent given by AutoQUANT and 4DMSPECT. A heart simulation study without a defined defect was used as the normal rest image. SPECT simulations

Projection images were generated using the Monte Carlo program SIMIND [13]. In this generation, we assumed a SPECT system equipped with a low-energy, high-resolution collimator and a 20% energy window centred over the 140 keV photopeak. Sixty-four SPECT projections with a matrix size of 64  64 pixels and a pixel size of 0.63  0.63 cm2 were simulated using an elliptical 1801 orbit starting from right anterior oblique (RAO) 451 to left posterior oblique (LPO) 451. Physical effects, such as photon attenuation, contribution from photons scattered in the patient and distance-dependent collimator resolution blurring, were included in the simulated SPECT projections. The number of simulated photon histories per projection was sufficient to obtain good image quality [14]. Each of the projections was scaled in counts to match a 1 day rest : stress 99mTc-sestamibi protocol (300 : 900 MBq with a separation time of 3 h). In addition to Monte Carlo-simulated SPECT projections, we also created ideal SPECT projections by calculating projections from line integrals along the different projection angles from the three-dimensional NCAT transverse images of the activity distribution. These projections thus represent scatter- and attenuation-free data recorded by an imaging system with perfect

spatial resolution. These projections were reconstructed and used in the determination of the cause of the interdependence. Image reconstruction

The Monte Carlo-simulated SPECT projections and the ideal projections were reconstructed to give short-axis images [15,16] using commercial clinical processing software (Philips AutoSPECT, ADAC Laboratories, Milpitas, California, USA) that employs an MLEM iterative reconstruction method based on 12 iterations. The projections were post-filtered by a fifth-order Butterworth filter with a cut-off frequency of 0.66 Nyquist for the stress studies and 0.55 Nyquist for the rest study. Perfusion image evaluation

Reconstructed short-axis images were analysed with the AutoQUANT and 4D-MSPECT automatic quantification programs. From the short-axis data, the programs construct polar maps and compare the regional count levels with a program-specific normal database for males in order to calculate the extent and severity. Using these results, we then compared the measured extent and severity of the defects with the known defect volume (true extent) and LAUR values to determine the accuracy and consistency of the extent and severity calculated by these programs. The correction model

In a previous study, we showed that the measured severity was influenced by the true extent of the defect, and vice versa. To address this problem, we have developed a correction model which models the dependence of the measured extent and measured severity on the known lesion volume (expressed as a fraction of the left ventricle wall volume), that is the true extent, and LAUR. Ideally, the measured extent is proportional to the true extent and the measured severity is proportional to LAUR, with proportionality factors that contain the crossdependence of the measured extent on LAUR and the measured severity on the true extent. We found that such simple relationships, although approximately correct, could be improved by including adjustable offset terms f, g, h and i, thus: ME0 ¼ME  f ; TE0 ¼ TE  g; MS0 ¼MS  h; LAUR0 ¼ LAUR  i

ð1Þ

where ME denotes the measured extent, MS the measured severity and TE the true extent. The new primed quantities do not have any new physiological meaning; they merely facilitate the parameterization of the observations. By trial and error, equations of the following type were found to provide a useful

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An extent–severity interdependence correction model El-Ali et al. 131

Predicted values of the primed quantities of the lesion activity uptake reduction (LAUR0 ) and true extent (TE0 ), given as solutions to the expression [ – B ± O(B2 – 4AC)]/(2A) Table 1

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approximation of the observed data:   b 0 ME ¼ a  TE0 LAUR0  c MS0 ¼ ðd TE0 þ eÞLAUR0

ð2Þ ð3Þ

where a, b, c, d and e are coefficients. In order to predict the true extent and LAUR from the measured extent and measured severity, these equations must be invertible, which proves to be the case: it is possible to express TE0 and LAUR0 as roots of seconddegree equations with coefficients that depend on a, b, c, d and e only (Table 1). The coefficients a to i were obtained by non-linear least-squares fitting of Equations (2) and (3) to the data set obtained by simulation with a true extent in the range 5–17% and LAUR in the range 40–100%. The offset terms f, g, h and i occur in both Equations (2) and (3), and thus the least-squares fitting must be performed on both equations simultaneously. Robustness of the model

In order to study the robustness of the proposed model when applied to a patient population, 10 male patients with different heart sizes and different body sizes were simulated. The first four patients had the same body size as used to develop the model, but different heart sizes obtained by scaling the normal NCAT heart size by factors of 0.8, 0.9, 1.1 and 1.2. The remaining six patients had the same heart size as used to develop the model, but the body size was increased in steps of 10% of the NCAT normal body size, increasing photon attenuation and scatter effects. The defects were all located in the LCx region, and a single LAUR value of 60% was used for all 10 patient configurations.

Results The measured extent and severity are presented as a function of the true extent in Fig. 1 and LAUR in Fig. 2. These figures show that the measures of extent and severity given by the two quantification programs are essentially proportional to the true extent and LAUR, respectively. They also show that the measured severity is highly dependent on the true extent, and the measured extent on LAUR. The latter dependence is obviously

non-linear and is better described by the term [a – b/ (LAUR0 – c)] of Equation (2) than would have been possible with a linear expression. The two analysis programs, AutoQUANT and 4D-MSPECT, show some differences. Although 4D-MSPECT appears to treat defects in LCx fairly similarly to AutoQUANT, defects in LAD and RCA are generally assigned smaller values of extent and severity. Moreover, defects with low LAUR (20 and 40%) in the LAD and RCA regions are not reported by 4D-MSPECT. The values of the extent and severity given by the two programs are thus different, and this is especially apparent in the LAD and RCA regions. The values of extent given by AutoQUANT do not show a dependence on the location of the defect, in contrast with the results given by 4D-MSPECT. When compared with the true extent, both programs overestimate the extent in all three myocardial regions, but by different amounts ranging from 50 to 100% for small defect volumes. The interdependence of the extent and severity shown in Fig. 3 is still present despite the fact that we used images obtained from SPECT projections with perfect resolution and no motion. The pattern of interdependence is almost the same for lesions with different LAURs in Fig. 1 as in Fig. 3. The graphs in Fig. 4 show that the interdependence between the extent and severity can almost be cancelled out by implementing our correction method. The top panels in the figure present the corrected extent and corrected severity for AutoQUANT, and the bottom panels present the corresponding values for 4DMSPECT. The adjustable coefficients used in Equations (1)–(3) for solving defects in the LCx region are presented in Table 2. The robustness of the correction model was tested on a male patient group and the results are shown in Fig. 5. The ratios between the corrected extent and true extent for the values of extent given by AutoQUANT were slightly closer to unity than those given by 4D-MSPECT. In contrast, the ratios between the corrected LAUR and true LAUR for the values of severity given by 4DMSPECT were closer to unity than those given by AutoQUANT. Both the corrected extent/true extent and corrected LAUR/true LAUR ratios for the measures of extent and severity given by AutoQUANT and 4DMSPECT varied by approximately 10% at most. This indicates that the correction method is at least suitable for this group of simulated male patients.

Discussion The extent and severity are important parameters used to determine the degree of disease in myocardial SPECT

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132 Nuclear Medicine Communications 2006, Vol 27 No 2

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imaging, and should ideally be independent of each other. This study and our previous investigation showed that this was not the case, and that the calculation of the extent could vary substantially with LAUR and with the location of the defect. This implies that the extent and severity are, to some degree, connected with each other. The reason was found to be the quantification algorithms, as shown in Fig. 3.

The relation between the severity and extent was linear for almost all of the defined defect sizes (see Fig. 1), but the results given by AutoQUANT had slopes of different steepness. Defects with low values of LAUR analysed by 4D-MSPECT did not show this linearity. 4D-MSPECT did not detect defects with a LAUR of 20% when located in the LAD and RCA regions, probably due to the high threshold for abnormality (2.5s). Figure 2 shows the

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An extent–severity interdependence correction model El-Ali et al. 133

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2

2 0

4 6 8 10 12 14 16 18

120

LAD

4 6 8 10 12 14 1618

True extent (%)

120

RCA

120

100

100

100

80

80

80

60

60

60

40

40

40

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0

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0 4 6 8 10 12 14 1618

LCx

Corrected LAUR (%)

Corrected extent (%)

22

0

0 4 6 8 10 12 14 1618

LCx

4 6 8 10 12 14 16 18

4 6 8 10 12 14 1618

True extent (%)

Top: corrected extent and severity values in the different myocardial regions (LAD, left anterior descending coronary artery; RCA, right coronary artery; LCx, left circumflex coronary artery) given by AutoQUANT. Bottom: corrected values obtained using 4D-MSPECT. The reference lines (black lines) are given to aid comparison of the corrected value with the actual value. ^, lesion activity uptake reduction (LAUR) of 100%; !, LAUR of 80%; ~, LAUR of 60%; K, LAUR of 40%.

measured severities versus LAUR for different true extents. The results given by AutoQUANT followed the same pattern for all three regions, whereas those given by 4D-MSPECT showed a different pattern for each of the three regions. The measured severity for a particular defect in a particular region was, in general, not the same for the two programs; exceptions were the measured severities of defects with high LAUR values in the LCx region, which were reasonably similar. However, the magnitudes of severity given by both programs started to vary at low LAUR values.

The interdependence (Figs. 1 and 2) was reduced when the least-squares method was used to minimize the least-square error of the two relationships [Equations (2) and (3)] which give the best fit to the measured extent and severity data in Figs. 1 and 2, respectively. Once the coefficients (Table 2) in Equations (2) and (3) had been determined, a second-degree equation combining Equations (2) and (3) could be obtained. The corrected extent, which should not be influenced by LAUR (severity), and the corrected LAUR, that is severity, which is not influenced by the

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Table 2 Coefficients used to estimate the lesion activity uptake reduction and true extent from the measured severity and measured extent of defects located in the left circumflex coronary artery region Coefficient a b c d e f g h I

AutoQUANT

4D-MSPECT

1.38 35.74 – 34.0 0.003 – 0.07 – 27.64 – 24.32 1.09 – 4.45

2.58 152.4 – 89.4 0.004 – 0.011 – 5.61 – 2.37 4.36 30.21

Fig. 5

1.5

Auto QUANT

4D-MSPECT

CE / TE

1.0

0.5

0.0 #1

#2

#3

#4

#5 #6 #7 Patient no.

#8

#9 #10

1.5

1.0

Our aim was not to make an accurate calculation of the real extent and real severity, but rather to show that the interdependence between the extent and severity could be eliminated. Equations (2) and (3) were intended to provide the best fit to the extent and severity in the LCx region. The same equations were used for the values in the LAD and RCA regions, without taking into consideration the influence of location on the extent and severity. Further work should focus on finding a regiondependent set of equations. The correction of the extent and severity values given by 4D-MSPECT was found to be less accurate as our model did not take into consideration the location dependence, which is especially important for 4D-MSPECT. The calculation of the extent and severity by AutoQUANT is not so strongly dependent on the location of the defect. Our intention was, however, not to include a location parameter in our correction model, but rather to see if we could use the same equation for all defects regardless of size and location. Our results show that this type of correction model performs better for values given by AutoQUANT than for those given by 4D-MSPECT, as the defect threshold used in 4D-MSPECT is not dependent on location. The proposed correction model has only been tested on a small group of simulated male patients, but the results indicate good robustness. This correction model may also be suitable and useful for extent and severity corrections in different types of patients. Further studies must, however, be made in order to test the robustness in female patients, where larger variations in photon attenuation can be expected.

Conclusions

CL / TL

We conclude that there is an interdependence between the measured extent and severity calculated from myocardial perfusion SPECT images using two commonly employed quantification programs. This interdependence is at variance with the basic intuitive definitions of extent and severity. The interdependence can be reduced significantly by implementing the correction model proposed in this paper.

0.5

0.0 #1

#2

#3

#4

#5 #6 #7 Patient no.

#8

#9 #10

Top: ratio between the true extent and corrected measured extent (TE/ CE). The values, obtained from AutoQUANT and 4D-MSPECT programs, were corrected using our correction model. Bottom: corrected lesion activity uptake reduction (LAUR)/true LAUR ratios (CL/TL). The reference lines at the value 1.0 are to guide the eye.

References 1

2

3

extent, could be determined by solving this seconddegree equation (Table 1). The results are presented in Fig. 4.

Ladenheim ML, Pollock BH, Rozanski A, Berman DS, Staniloff HM, Forrester JS, et al. Extent and severity of myocardial hypoperfusion as predictors of prognosis in patients with suspected coronary artery disease. J Am Coll Cardiol 1986; 7:464–471. Gibbons RJ, Miller TD, Christian TF. Infarct size measured by single photon emission computed tomographic imaging with (99m)Tc-sestamibi: a measure of the efficacy of therapy in acute myocardial infarction. Circulation 2000; 101:101–108. Thomas GS, Miyamoto MI, Morello AP III, Majmundar H, Thomas JJ, Sampson CH, et al. Technetium 99m sestamibi myocardial perfusion imaging predicts clinical outcome in the community outpatient setting. The Nuclear Utility in the Community (NUC) Study. J Am Coll Cardiol 2004; 43:213–223.

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An extent–severity interdependence correction model El-Ali et al. 135

4

Sabharwal NK, Lahiri A. Role of myocardial perfusion imaging for risk stratification in suspected or known coronary artery disease. Heart 2003; 89:1291–1297. 5 Berman DS, Germano G, Shaw LJ. The role of nuclear cardiology in clinical decision making. Semin Nucl Med 1999; 29:280–297. 6 Germano G, Kavanagh PB, Berman DS. An automatic approach to the analysis, quantitation and review of perfusion and function from myocardial perfusion SPECT images. Int J Cardiol Imaging 1997; 13:337–346. 7 ADAC Laboratories. AutoQUANTt user’s manual 9201-0225B-ENG, Rev A. Milpitas, CA: ADAC Laboratories; 1999. 8 The user’s manual for 4D-MSPECT. Ann Arbor, MI: The University of Michigan Medical Center; 2003. 9 Sciagra R, Imperiale A, Antoniucci D, Migliorini A, Parodi G, Comis G, et al. Relationship of infarct size and severity versus left ventricular ejection fraction and volumes obtained from 99mTc-sestamibi gated single-photon emission computed tomography in patients treated with primary percutaneous coronary intervention. Eur J Nucl Med Mol Imaging 2004; 31:969–974. 10 El-Ali HH, Palmer J, Carlsson M, Edenbrandt L, Ljungberg M. Interdependence between measures of extent and severity of myocardial

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16

perfusion defects provided by automatic quantification programs. Nucl Med Commun 2005; 26:1125–1130. Segars WP, Lalush DS, Tsui BMW. A realistic spline-based dynamic heart phantom. IEEE Trans Nucl Sci 1999; 46:503–506. Wackers FJ, Berman DS, Maddahi J, Watson DD, Beller GA, Strauss HW, et al. Technetium-99m hexakis 2-methoxyisobutyl isonitrile: human biodistribution, dosimetry, safety, and preliminary comparison to thallium-201 for myocardial perfusion imaging. J Nucl Med 1989; 30:301–311. Ljungberg M, Strand S-E. A Monte Carlo program simulating scintillation camera imaging. Comp Meth Progr Biomed 1989; 29:257–272. El-Ali HH, Palmer J, Carlsson M, Edenbrandt L, Ljungberg M. Comparison of one- and two-day protocols for myocardial SPECT: a Monte Carlo study. Clin Physiol Funct Imaging 2005; 25:189–195. Germano G, Kavanagh PB, Chen J, Waechter P, Su HT, Kiat H, et al. Operator-less processing of myocardial perfusion SPECT studies. J Nucl Med 1995; 36:2127–2132. Germano G, Kavanagh PB, Su HT, Mazzanti M, Kiat H, Hachamovitch R, et al. Automatic reorientation of three-dimensional, transaxial myocardial perfusion SPECT images. J Nucl Med 1995; 36:1107–1114.

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Original article

Correlation of preoperative thallium SPECT with histological grading and overall survival in adult gliomas Fre´de´ric Comtea, Luc Bauchetb, Vale´rie Rigauc, Jean Robert Haueta, Michel Fabbrod, Philippe Coubesb, Jeanine Chevaliera, Denis Mariano-Goularta, Michel Rossia and Michel Zancaa Background The management and prognosis of a glioma depend on the tumour’s histological grade. Thus, preoperative prediction of the grade is routinely needed to indicate whether surgery or biopsies are required. It has been proposed that thallium single photon emission computed tomography (SPECT), in a relative short series, will aid this prediction.

optimal cut-off value of the thallium index for the detection of high grade glioma was determined. By using 2.2 as the value for the threshold thallium index, the sensitivity and specificity were 93% and 72%, respectively. Kaplan–Meier estimates of the overall survival curves, as a function of the thallium index, indicated that it was correlated with the overall survival (P < 0.001).

Aim To confirm the correlation between the results of preoperative thallium SPECT and grade of tumour as well as patient survival, and to define the cut-off value of the optimal thallium index for the detection of high grade gliomas in a large series of patients.

Conclusion Thallium SPECT provides useful information about the histological grade of the tumour and overall patient survival. Additionally, in spite of its relatively weak resolution, it appears to be a powerful routine clinical tool for the management of gliomas. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:137–142

Methods One hundred and eighteen patients treated for glioma were retrospectively included in this study. All patients underwent preoperative 201Tl SPECT upon initial presentation and were referred for neurosurgery. Initial scintigraphic findings were correlated with the histological grade of the tumour and overall patient survival. Results Thallium uptake was highly correlated with histological grade; the mean thallium indices for low grade and high grade gliomas were 1.8 and 4.9, respectively. On the basis of receiver operating characteristic analysis, the

Introduction Gliomas are primary brain tumours that are histologically classified as low grade or high grade malignant tumours. The management of gliomas depends on their histological characteristics and grade. Specific noninvasive explorations are necessary to choose the appropriate surgical strategy, and to assess the therapeutic response or the histological conversion of a low grade to a high grade lesion. 201Tl single photon emission computed tomography (thallium SPECT) has been proposed as an indicator of both the histological grade of the tumour and prognosis of brain lesions. Mountz et al. [1], in a microautoradiographic study, described the preferential uptake of thallium by tumour cells compared with non-involved brain tissue. Most recently, Gungor et al. [2] showed that thallium uptake can predict proliferative activity of a glioma as defined by the proliferating cell nuclear antigen labelling index. Several clinical studies have shown that thallium uptake is correlated with histological grade [3–9]

Nuclear Medicine Communications 2006, 27:137–142 Keywords: thallium, SPECT, glioma, survival, prognosis a Service de Me´decine Nucle´aire, bService de Neurochirurgie, cLaboratoire d’Anatomopathologie, CHU Montpellier and dService d’Oncologie, CRLC Val d’Aurelle, Montpellier, France.

Correspondence to Dr Fre´de´ric Comte, Service Me´decine Nucle´aire, CHU Gui de Chauliac, 80, avenue Augustin Fliche, 34295 Montpellier Cedex 5, France. Tel: + 0033 467 337 287; fax: + 0033 467 336 922; e-mail: [email protected] Received 19 September 2005 Accepted 11 October 2005

and survival after treatment [5,7,8,10–15]. Stereotaxic biopsies can also be successfully guided by thallium uptake [16,17]. Other authors [18,19] did not confirm this correlation. In addition, most of the aforementioned studies were performed on a small number of patients. The aims of this study were (1) to confirm the correlation between thallium uptake and histological grade in a large population of gliomas; (2) to evaluate the prognostic value of thallium SPECT performed before treatment; and (3) to define the optimal value of the thallium index in terms of sensitivity and specificity.

Methods Patients

One hundred and eighteen patients (47 females and 71 males; mean age 51 years) treated for glioma between January 1998 and December 2002 were included retrospectively in this study. Tumour histologies included glioblastomas (n = 42), anaplastic astrocytomas (n = 11),

c 2006 Lippincott Williams & Wilkins 0143-3636

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astrocytomas (n = 9), pilocytic astrocytomas (n = 4), oligodendrogliomas (n = 15), anaplastic oligodendrogliomas (n = 30), anaplastic oligoastrocytomas (n = 5), subependymoma (n = 1), and anaplastic ependymoma (n = 1). All patients underwent preoperative thallium SPECT upon initial presentation and were referred for neurosurgery. Optimal treatment was total surgery and radiotherapy (56–60 Gy). Chemotherapy (cisplatin–carmustine or temozolomide–carmustine) was indicated when partial surgery or biopsy alone was performed and in cases of recurrence. Sixty-six patients (56%) underwent total surgery, 15 patients (13%) partial surgery, and 37 patients (31%) stereotaxic biopsy.

Fig. 1

Thallium SPECT

Mean value of the thallium index (Tl index) for low (grades 1 and 2) and high (grades 3 and 4) grade tumours. The Tl index is significantly different (P < 0.0001) in low grade gliomas (Tl index = 1.8, standard deviation = 1.4) compared with high grade gliomas (Tl index = 4.95, standard deviation = 2.5).

All patients received an intravenous dose of 201Tl chloride (185 MBq, 5 mCi,). A triple-head gamma camera (Prism 3000 Philips) with fan-beam collimators (LEUHR fan) was used. Projection data were acquired 10 min after intravenous injection, in a 128  128 pixel matrix in 73 keV (15% window) and 170 keV (10% window) photopeaks with 30 steps over 1201 at 60 s per view. Images were reconstructed with a ramp filter and Wiener post-filter (cut-off frequency = 1.15) without attenuation correction. Tumour activity was evaluated on a 6.5 mm thick transaxial slice. A region of interest (ROI) was manually drawn around the hottest part of the lesion and duplicated on the opposite side in the normal brain. The thallium index was defined as the ratio of the mean activity in the tumour ROI to the mean activity in the contralateral ROI. Statistical analysis

The values of the thallium indices for low grade and high grade gliomas were compared. To define the thallium indices that discriminated between low grade and high grade gliomas, a receiver operating characteristic (ROC) curve analysis was performed; sensitivity, specificity and predictive values were computed with 95% confidence intervals. The prognostic value of the thallium indices was assessed by a survival analysis. The study population was divided into four groups as a function of the thallium indices. Survival rates of all groups were determined by using a log-rank test; multiple comparisons with Bonferroni’s correction were performed with a 95% confidence interval using a univariate Cox model.

Results Correlation of thallium uptake with histological grade

Twenty-nine tumours were low grade gliomas (24.6%), and 89 were high grade gliomas (75.4%). The mean thallium indices measured in low grade and high grade gliomas were significantly different (P < 0.0001), with values of 1.8 and 4.9 for low grade and high grade gliomas,

8 7 Thallium index

6 5 4 3 2 1 0 Low grade

High grade

respectively (Fig. 1). No significant difference was observed in histological sub-groups of high grade gliomas (WHO II and IV). In low grade gliomas, the mean thallium index for the pilocytic astrocytoma group (n = 4) was higher (3.0) than the indices for astrocytomas and oligodendrogliomas (1.9 and 1.6, respectively). These differences do not appear to be significant (P > 0.01) (Fig. 2). Accuracy of thallium SPECT for the detection of high grade lesions

For a thallium index of 2.7 the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV), obtained from the ROC curves (Fig. 3), were 89%, 79%, 93% and 70%, respectively. For a thallium index of 2.2, the sensitivity, specificity, PPV and NPV were 93%, 72%, 91% and 78%, respectively. Correlation of the thallium indices with overall survival

Kaplan–Meier estimates of the overall survival curves as a function of the thallium indices show a significant survival difference between groups with thallium indices smaller and greater than 2.2, whatever the histological grade (P < 0.0001). Beyond this threshold, no difference in survival could be observed, whatever the value of the index (Fig. 4). With a thallium index greater than 2.2, the death hazard ratio was 4.08 (95% confidence interval = 2.16–7.71). We observed six cases of low grade gliomas with high thallium indices, evoking false positive SPECT results; two cases corresponded to pilocytic astrocytoma (with short duration of survival) and four to oligodendroglioma or astrocytoma (with long duration of survival in three cases). Three patients (two with pilocytic astrocytoma and one with astrocytoma) showed a poor outcome and

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Thallium SPECT in adult gliomas Comte et al. 139

Fig. 2

9 8 7

Thallium index

6 5 4 3 2 1 0 Pilocytic astro

Oligodendro

Astro

Anaplastic oligoastro

Anaplastic astro

Anaplastic oligoastro

Glioblastoma

Mean value of the thallium index and standard deviation for pilocytic astrocytomas (3.0–2.5), oligodendrogliomas (1.6–1.2), astrocytomas (1.9–1.5), anaplastic oligoastrocytomas (3.4–1.3), anaplastic astrocytomas (3.7–1.7), anaplastic oligodendrogliomas (5.1–2.5) and glioblastomas (5.3–2.7).

Fig. 3

1.0

Fig. 4

1.00

0.9 0.8

Survival distribution function

0.7

Sensitivity

0.6 0.5 0.4 0.3

0.75

0.50 A

0.25 B

0.2

C

0.1 D

0.00 0.0

0 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1 − Specificity

Receiver operating characteristic curve analysis to determine the cut-off value of the thallium index that differentiates between low and high grade gliomas. The area under the curve, which represents the overall accuracy of the test, is 0.89 (95% confidence interval = 82.8–94.6%).

1

2

3

4

5

6

7

Overall survival (years) Survival curves as a function of the thallium index (Tl index). Tl index < 2.2 (group A); 2.2 < Tl index < 3 (group B); 3 < Tl index < 4.5 (group C), and Tl index > 4.5 (group D). The survival rate was significantly lower in patients with a Tl index > 2.2, compared with those with a Tl index < 2.2 (P < 0.0001).

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Table 1 Cases that showed discordance between the histological and scintigraphic findings Histological data Patient number 1 2 3 4 5 6 7 8 9 10 11 12

Type

Pilocytic astrocytoma Pilocytic astrocytoma Astrocytoma Astrocytoma Oligodendroglioma Oligodendroglioma Anaplastic astrocytoma Anaplastic oligodendroglioma Anaplastic oligodendroglioma Anaplastic oligodendroglioma Anaplastic oligodendroglioma Multiforme glioblastoma

Grade (WHO)

Thallium index

Overall survival (months)

I I II II II II III III III III III IV

3.7 5.0 2.7 5.6 5.4 3.0 1.6 1.0 1.0 1.5 1.7 2.2

5 0* 11 32 > 43 60 15 44 > 29 > 29 6 9

*

Postoperative death.

died within 11 months. For six high grade gliomas, the thallium indices were lower than 2.2, leading to false negative SPECT results. Three of these patients were still alive 29 months after the SPECT; one patient had a 15-month survival and the two others had poor outcomes and died within 9 months. Table 1 summarizes the cases where there was discordance between the scintigraphic, histological and clinical results.

Discussion The initial noninvasive evaluation of histological grade is a major factor in the management of gliomas. The monitoring of treated lesions and the early detection of transformation is important. Functional imaging with thallium or sestamibi can be used in these areas. Several studies have shown the ability of thallium SPECT to differentiate between low grade and high grade gliomas [3–5,9,20,21]. In these studies, thallium uptake was compared with histological findings in 22–29 presurgical patients with low grade and high grade gliomas. The thallium uptake was greater in high grade than in low grade lesions. Other authors have studied the performance of thallium in the assessment of recurrence. In 29–47 patients with previously treated high grade gliomas with suspected recurrence, the thallium indices not only correlated with the histopathological findings noted upon reoperation [6–8] but also with survival [7,8]. Some authors did not confirm the accuracy of thallium in the prediction of histological grade and showed the superiority of a computed tomography (CT) scan or magnetic resonance imaging. Kallen et al. [22] studied 37 patients with low and high grade gliomas by using CT scans and thallium SPECT and reported a lower accuracy for thallium (78%) compared to a CT scan (84%) in identifying high grade gliomas. Lam et al. [19], in a study that included 19 patients, concluded that thallium was not useful for the preoperative grading of gliomas. In fact, their study confirmed that no statistical difference could

be observed in thallium uptake between grades I and II (i.e., low grade gliomas) and grades III and IV (i.e., high grade gliomas). In a subgroup of only six patients (two grade II and four grade III), no significant difference was observed in thallium uptake between the grade II (low grade) and the grade III (high grade) gliomas. However, the inaccuracy of the thallium SPECT could not be established in such a small series. The results of our study, performed on a large group, showed that low grade gliomas could effectively be distinguished from high grade by preoperative thallium SPECT. The mean index was significantly different in these two groups of gliomas. The mean indices of the low and high grade gliomas, evaluated on 29 and 89 patients, were 1.8 and 4.9, respectively. However, no distinction could be made between the different high grade histological groups; glioblastomas, anaplastic astrocytomas, oligodendrogkiomas or oligoastrocytomas showed similar mean thallium indices. As reported by others [23], we also observed that the uptake of thallium was variable in pilocytic astrocytoma, which is a low grade glioma (WHO I): two patients among the four cases of pilocytic astrocytomas showed important uptake of thallium. However, these two patients had very poor outcomes and died within 5 months following surgery. The two other patients with pilocytic astrocytomas presented low thallium uptake and were still alive as of June 2005. The prognosis of pilocytic astrocytoma seems to be better for tumours with low thallium uptake compared with those of high thallium uptake. No conclusion can be drawn from this observation on only four cases, and additional studies on pilocytic astrocytomas are needed to evaluate the significance of thallium uptake in this group of low grade gliomas. The thallium uptake of gliomas clearly discriminated high grade from low grade gliomas. The mean uptake values were significantly different in the two groups. However, the definition of a discriminant threshold value remains difficult. Previous studies [6,7,24] reported a cut-off value of around 2 (1.5–2.5). The modality of the digital image treatment, particularly the choice of the algorithm of reconstruction and the correction or not of attenuation phenomena, could influence the absolute value of the thallium index. We used filtered back-projection without attenuation correction. Under these conditions, using a threshold index of 2.2, ROC analysis revealed a sensitivity and specificity of 93%, and 73%, respectively. The specificity could reach 79%, by using 2.7 as the threshold index, but in this case the sensitivity decreased to 89%. To preserve maximal sensitivity, we used 2.2 as the cutoff value for the prediction of high grade gliomas. The outcome and the prognosis of gliomas depend on histological type and tumour grade. The prognostic value of thallium SPECT in suspected recurrence of gliomas

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Thallium SPECT in adult gliomas Comte et al. 141

has been reported in previous studies [7,8,11,12] which showed that, in 22 to 57 patients with recurrent gliomas, thallium is a good predictor of outcome. However, in one study, survival was reported to be dependent on the thallium index, but only in grade III gliomas and not in grade IV [11]. Datta et al. [25] studied the postoperative prognostic value of thallium SPECT in 33 patients with high grade gliomas, before receiving radiotherapy, and showed that the Karnofsky performance status was the only significant predictor for progression-free survival. In the present study we showed that thallium SPECT, performed preoperatively, gave indications about the outcome, whatever the histological grade. Discordance between histological findings and scintigraphic findings was observed in 12 cases. Six low grade gliomas showed moderate or highly elevated thallium uptake. Three of them showed poor outcomes: two died within 5 months and another within 11 months. On the other hand, six patients with high grade gliomas showed no or low thallium uptake: three of them showed prolonged survival and the three others died within 15 months (Table 1). The Kaplan–Meier survival curve indicated that gliomas with low thallium uptake ( < 2.2) were associated with a longer survival than those with high thallium uptake, confirming the prognostic value of thallium SPECT in gliomas. However, the overall survival was not linearly correlated with maximal tumour intensity, as reported in previous studies [7,12], but we observed a threshold value (2.2) beyond which poor outcome was observed, whatever the value of the thallium index. Preoperative thallium SPECT thus provides valuable prognostic information as a complement to histological grade. The emergence of positon emission tomography (PET) imaging in routine clinical practice allows the use of specific tracers such as radiolabelled amino acids. PET imaging appears to be a more resolutive and more sensitive modality. [11C-methyl]methionine has been shown to be efficient in brain tumour imaging [9,27,28]. However, the short half-life of 11C is a limiting factor for its routine clinical use. Fluoro tracers such as [18F]fluoroethyl-L-tyrosine have been proposed in routine clinical practice for the exploration of gliomas [28–31]. Given this emergent modality, the place of thallium SPECT could be questionable. The performance, cost and availability of both modalities must be taken into account to define the accurate strategy for management of gliomas. Our study confirms on a relatively large scale the value of thallium SPECT in the detection of high grade gliomas and its prognostic value when performed preoperatively. To date, thallium SPECT appears to be a powerful, and available, tool for routine clinical use.

Conclusion We have shown that thallium SPECT remains, an effective tool in the management of gliomas in spite of

its limited resolution. Thallium uptake was highly correlated with the tumour histological grade, and this modality appears to be efficient for discrimination between high grade and low grade gliomas. Furthermore, the preoperative thallium uptake by gliomas was found to be predictive of the overall survival, whatever the histological grade. High thallium uptake in low grade gliomas was predictive of a poor outcome. Taking into account its availability and its relative low cost compared with PET imaging, thallium SPECT certainly has its place in neuro-oncology and should be included in a management strategy for patients with gliomas.

Acknowledgements The authors gratefully acknowledge Dr Sharon Lynn Salhi for constructive comments and presubmission editorial assistance. The authors also thank Dr Eric Barbotte for the statistical analysis.

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14 Taki S, Kakuda K, Kakuma K, Kobayashi K, Ohashi M, Ito S, et al. 201Tl SPECT in the differential diagnosis of brain tumours. Nucl Med Commun 1999; 20: 637–645. 15 Kosuda S, Fujii H, Aoki S, Suzuki K, Tanaka Y, Nakamura O, et al. Reassessment of quantitative thallium-201 brain SPECT for miscellaneous brain tumors. Ann Nucl Med 1993; 4:257–263. 16 Hemm S, Vayssiere N, Zanca M, Ravel P, Coubes P. Thallium SPECT-based stereotactic targeting for brain tumor biopsies. A technical note. Sterotact Funct Neurosurg 2004; 82:70–76. 17 Hemm S, Rigau V, Chevalier J, Picot MC, Bauchet L, El Fertit H, et al. Stereotactic coregistration of 201Tl-SPECT and MRI applied to brain tumor biopsies. J Nucl Med 2005; 46:(in press). 18 Martinez del Valle M, Gomez-Rio M, Horcajadas A, Rodriguez-Fernandex A, Muros de Fuentes A, Acosta-Gomez MJ, et al. False positive thallium-201 SPECT imaging in brain abscess. Br J Radiol 2000; 73:160–164. 19 Lam W, Chan K, Wong W, Poon W, Metreweli C. Pre-operative grading of intracranial glioma. Comparison of MR-determined cerebral blood volume maps with thallium-201 SPECT. Acta Radiol 2001; 42: 548–554. 20 Kim KT, Black KL, Marciano D, Mazziotta JC, Guze BH, Grafton S, et al. Thallium-201 SPECT imaging of brain tumors: methods and results. J Nucl Med 1990; 41:965–969. 21 Burkard R, Kaiser KP, Wieler H, Klawki P, Linkamp A, Mittlebach L, Goller T. Contribution of thallium-201-SPECT to the grading of tumorous alterations of the brain. Neurosurg Rev 1992; 15:265–273. 22 Kallen K, Heiling M, Andersson AM, Brun A, Holtas S, Ryding E. Preoperative grading of glioma malignancy with thallium-201 single-photon emission CT: comparison with conventional CT. Am J Neuroradiol 1996; 17:925–932.

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Weckesser M, Matheja P, Rickert CH, Strater R, Palkovic S, Lottgen J, et al. High uptake of L-3-[123I]iodo-alpha-methyl tyrosine in pilocytic astrocytomas. Eur J Nucl Med 2001; 28:273–281. Matheja P, Rickert C, Weckesser M, Palkovic S, Lottgen J, Riemann B, et al. Sequential scintigraphic strategy for the differentiation of brain tumours. Eur J Nucl Med 2000; 27:550–558. Datta NR, Pasricha R, Gambhir S, Prasad SN, Phadke RV. Comparative evaluation of 201Tl SPECT and CT in the follow-up of irradiated brain tumors. Int J Clin Oncol 2004; 9:51–58. Sato N, Suzuki M, Kuwata N, Kuroda K, Wada T, Beppu T, et al. Evaluation of the malignancy of glioma using 11C-methionine positron emission tomography and proliferating cell nuclear antigen staining. Neurosurg Rev 1999; 22:210–214. De Witte O, Goldberg I, Wikler D, Rorive S, Damhaut P, Monclus M, et al. Positron emission tomography with injection of methionine as a prognostic factor in glioma. J Neurosurg 2001; 95:746–750. Weber WA, Wester HJ, Grosu AL, Herz M, Dzewas B, Feldmann HJ, et al. O-(2-[18F]fluoroethyl)-L-tyrosine and L-[methyl-11C] methionine uptake in brain tumours: initial results of a comparative study. Eur J Nucl Med 2000; 27:542–549. Popperl G, Gotz C, Rachinger W, Gildehaus FJ, Tonn FJ, Tatsch K. Value of O-(2-[18F]fluoroethyl)-L-tyrosine PET for the diagnostic of recurrent glioma. Eur J Nucl Med Mol Imaging 2004; 31:1464–1470. Pauleit D, Floeth F, Tellmann L, Hamacher K, Hautzel H, Muller HW, et al. Comparison of O-(2-[18F]fluoroethyl)-L-tyrosine PET and 3-[123I]iodo-amethyl-L-tyrosine SPECT in brain tumors. J Nucl Med 2004; 45:374–381. Pauleit D, Floeth F, Hamacher K, Riemenschneider MJ, Riefenberger G, Muller HW, et al. O-(2-[18F]fluoroethyl)-L-tyrosine PET combined with MRI in the diagnostic assessment of cerebral gliomas. Brain 2005; 128:678–687.

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Original article

Early dynamic 201Tl SPECT in the evaluation of brain tumours Nobuo Sugo, Kyousuke Yokota, Kousuke Kondo, Naoyuki Harada, Yoshinori Aoki, Chikao Miyazaki, Masaaki Nemoto, Toshiyuki Kano, Hitoshi Ohishi and Yoshikatsu Seiki Objective To estimate the usefulness of early dynamic 201 Tl single photon emission computed tomography (SPECT) studies in distinguishing the histological malignancy of brain tumours. Methods Dynamic 201Tl SPECT was performed for 3 min per scan for 15 min immediately after the administration of 201 TlCl in 110 patients with brain tumours (111 lesions). The data obtained each 3 min were used for dynamic SPECT, and the five sets of data obtained were added to acquire static SPECT data. For static SPECT, the static thallium index (STI) was calculated as the ratio of 201Tl uptake in the tumour to that of the contralateral normal brain. The ratio of the 201Tl uptake for each 3 min was defined as the dynamic thallium index (DTI). The dynamic thallium rate (DTR), as a per cent, was calculated as DTR = (DTI for every 3 min)/STI H 100. The five values were approximated as a linear function and the slope (%/min) was calculated. Results In static SPECT, there was no significant difference between the STI of malignant tumours (glioblastoma and anaplastic astrocytoma) and that of benign tumours (lowgrade glioma, meningioma, pituitary adenoma, neurinoma and haemangioblastoma) (3.7 ± 1.5, 5.0 ± 3.5, respectively).

Introduction 201

Tl, which was initially developed for myocardial imaging, was incidentally noted to exhibit uptake in lung carcinoma, liver cancer, and thyroid cancer, and it was demonstrated that these lesions could be visualized by 201 Tl uptake [1]. The usefulness of 201Tl for localizing brain tumours, estimating the extent of residual tumour or recurrence and distinguishing high-grade gliomas from low-grade gliomas has been reported [2–4]. However, the accumulative degree of only 201Tl is not exclusive to malignant tumours, and benign tumours such as meningioma exhibit varying degrees of uptake [5,6]. Over the past few years, it has been found that early (10–30 min) and delayed (3–4 h) 201Tl single photon emission computed tomography (SPECT) imaging is very useful for distinguishing benign from malignant brain tumours [7–12]. This method indicates not only 201Tl uptake but also reveals the dynamics of accumulation, and offers a more accurate diagnosis of brain tumours. On the other hand, two examinations are needed to acquire early and delayed images, which is inconvenient for brain tumour

On dynamic SPECT, DTI increased markedly over 15 min for malignant tumours. In contrast, the DTI of benign tumours increased slightly, steadily or decreased. The slope of the linear functions calculated from the DTRs was much higher in the malignant tumour group than in the benign tumour group (P < 0.001). Conclusions We suggest that the performance of 201Tl dynamic SPECT for 15 min is useful for distinguishing malignant brain tumours from benign brain tumours and reduces the examination stress of patients. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:143–149
Nuclear Medicine Communications 2006, 27:143–149 Keywords: 201Tl, single photon emission computed tomography, benign brain tumours, malignant brain tumours Department of Neurosurgery, Toho University, Omori Medical Center, Tokyo, Japan. Correspondence to Dr Nobuo Sugo, Department of Neurosurgery, Toho University, Omori Medical Center, 6-11-1, Omori-nishi, Ota-ku, Tokyo 143-0015, Japan. Tel: + 0081 3 3762 4151; fax: + 0081 3 3298 4847; e-mail: [email protected] Received 6 May 2005 Revised 2 September 2005 Accepted 21 September 2005

patients, who often have hemiparesis and/or disturbance of consciousness. Recently, the development of a gamma camera for SPECT has resulted in a remarkable increase in data collection capacity, while the modification of peripheral devices and the introduction of supercomputers have considerably reduced data processing times, enabling the routine use of dynamic SPECT. We performed dynamic 201Tl SPECT studies for 15 min on brain tumour patients to estimate the usefulness of 201Tl dynamics in distinguishing the histological malignancy of brain tumours.

Methods Patient population

Dynamic SPECT studies were performed on 123 consecutive patients with primary brain tumours before surgical resection. Of these, in two cases of meningioma, one of haemangioblastoma, eight of neurinoma and two of pituitary adenoma, the accumulation of 201Tl on images was not acceptable because their tumour diameters were small ( < 15 mm), and the data for these cases were

c 2006 Lippincott Williams & Wilkins 0143-3636


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excluded. Therefore, 111 lesions of 110 cases were evaluated retrospectively in this study. The diameter of these tumours was more than 20 mm. All cases were diagnosed by post-operative histological examination. Histological diagnoses were as follows: 22 patients had glioblastoma (13 males, nine females; median age, 56.7 years; range, 35–76), 10 had anaplastic astrocytoma (seven males, three females; median age, 48.5 years; range, 31–64), five had diffuse astrocytoma (one male, four females; median age, 38.0 years; range, 28–52), one had pilocytic astrocytoma (male, age 28 years), one had oligoastrocytoma (male, age 70 years), 38 had meningioma (16 males, 22 females; median age, 57.8 years; range, 26– 82, all meningiomas corresponded to grade I), 19 had pituitary adenoma (five males, 14 females; median age, 47.6 years; range, 22–66), 10 had neurinoma (two males, eight females; median age, 52.1 years; range, 27–69) and four (five lesions) had haemangioblastoma (four males; median age, 43.4 years; range, 23–55). Diffuse astrocytoma, pilocytic astrocytoma and oligoastrocytoma were grouped together as low-grade glioma. The histology of the tumour specimens obtained by surgical resection was determined according to the World Health Organization (WHO) histological classification [13]. SPECT method

To compare the changes in 201Tl dynamics in each histological group, the five DTR data sets were approximated by the linear function and the slope (%/ min) was calculated. Statistical analysis

One-way analysis of variance and the paired t-test were used. P values < 0.05 were considered statistically significant.

Results Static thallium index

The STI ranged from 2.2. to 7.3 in glioblastomas, 1.2 to 4.7 in anaplastic atrocytomas and 1.5 to 3.4 in low-grade gliomas. The STI of oligoastrocytoma was 3.4, which was the highest value in low-grade gliomas. The STI of glioblastomas was higher than that of anaplastic astrocytomas and low-grade gliomas (P < 0.05 and 0.01, respectively: Fig. 1). The STI ranged from 1.7 to 18.7 in meningiomas, 1.6 to 8.6 in pituitary adenomas, 1.3 to 4.8 in neurinomas and 4.6 to 8.0 in haemangioblastomas. The STI of meningiomas and haemangioblastomas were higher than other groups (Fig. 1). An index cutoff of 3.0 for gliomas, 17 of 22 glioblastomas (77.3%) and 10 of 17 of other gliomas (58.8%), such as anaplastic astrocytomas and low-grade gliomas could be distinguished (Fig. 1);

Fig. 1

∗∗ ∗∗∗

∗∗

20

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∗∗∗

∗∗∗

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18 16 14

∗

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12 STI

SPECT scans were performed with a three-detector camera (Prism3000; Picker International, Bedford Heights, Ohio), with high-resolution fanbeam collimators used for imaging. SPECT examination was performed for 3 min per scan for 15 min immediately after the intravenous administration of 111 MBq of 201TlCl. SPECT data were acquired by continuously rotating the detectors by 1201 (40 steps per 1201 per 15 s), and were stored as a 64 64 word matrix in a supercomputer system (Odyssey; Picker International, Bedford Heights, Ohio). The data of each 3 min were used as dynamic SPECT data, and the five sets of data obtained over 15 min were summed and used as static SPECT data. Dynamic and static SPECT images were reconstructed using a filtered back-projection algorithm after preprocessing with a Butterworth filter. In addition, the thickness of the static and dynamic SPECT image was 5 pixels (17.2 mm). No attenuation correction was performed.

tumour to that of the contralateral normal brain over each 3 min was defined as the dynamic thallium index (DTI). The dynamic thallium rate (DTR), as a per cent, was sequentially calculated as DTR ¼ ðDTI for every 3 minÞ=ðSTIÞ100:

10

Semiquantitative analysis

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Regions of interest (ROIs) were selected within the tumour and in the contralateral normal brain from static SPECT images. The static thallium index (STI) was based on the ratio of 201Tl uptake in the tumour to that of the contralateral normal brain on static SPECT images. In the analysis of dynamic SPECT data, the ROI set on static SPECT was copied. The 201Tl uptakes in the tumour and in the contralateral normal brain were measured over each 3 min and five numerical data sets for 15 min were obtained. The ratio of 201Tl uptake in the

6 4 2 0 GB

AA

LG

MM

PA

NE

HB

Relationships of the static thallium index (STI) among histological groups. GB: glioblastoma, AA: anaplastic astrocytoma, LG: low-grade glioma, MM: meningioma, PA: pituitary adenoma, NE: neurinoma, HB: hemangioblastoma. *P < 0.05, ** P < 0.01, ***P < 0.001.

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201

however, the accumulation degree among all tumour groups could not be diagnosed from the index cut-off. The lesions in this study were divided into a malignant tumour group and a benign tumour group. Malignant tumours comprised glioblastomas and anaplastic astrocytomas while benign tumours comprised low-grade gliomas, meningiomas, pituitary adenomas, neurinomas and haemangioblastomas. There was no significant difference between the STI of malignant tumours and that of benign tumours (3.7 ± 1.5, 5.0 ± 3.5; Fig. 2). Dynamic thallium index

In glioblastomas, the DTI increased markedly from just after 201Tl administration to 15 min later (Table 1 and

Fig. 2

NS 20

16 14 12 STI

Fig. 3(a)). There were significant differences among all pairs of the five data sets in DTI (P < 0.05). The DTI of anaplastic astrocytomas also increased markedly (Table 1 and Fig. 3(b)). There were significant differences among all pairs of the five data sets in DTI excluded between 12 min and 15 min (P < 0.05). No change in DTI over 15 min was observed for low-grade gliomas (Table 1 and Fig. 3(c)). In meningiomas, DTI increased slightly at 6 min (P < 0.001) and 9 min (P < 0.05) compared to the initial 3 min (Table 1 and Fig. 3(d)). DTIs of pituitary adenoma also increased at 6 min (P < 0.01), 9 min (P < 0.05), 12 min (P < 0.05) and 15 min (P < 0.05) compared to the initial 3 min (Table 1 and Fig. 3(e)). DTIs of neurinomas were steady (Table 1 and Fig. 3(f)). In contrast, the DTI decreased markedly from just after 201 Tl administration to 15 min later in haemangioblastomas (Table 1 and Fig. 3(g)). There were significant differences among all data set pairs in the DTI (P < 0.05). Dynamic thallium rate

18

10 8 6 4 2 0 Malignant

Benign

Comparison of the static thallium index (STI) between malignant and benign tumour groups. Malignant: malignant tumour group, Benign: benign tumour group. NS: no statistical difference.

Table 1

Tl SPECT for evaluating brain tumours Sugo et al. 145

DTRs in glioblastomas converted to percentages based on STI were 78.7 ± 10.6% (3 min), 91.7 ± 8.1% (6 min), 105.3 ± 12.4% (9 min), 114.8 ± 11.2% (12 min) and 126.7 ± 21.7% (15 min), and increased linearly with statistically significant differences among all pairs of 5 data sets in the DTR (Fig. 4(a)). In anaplastic astrocytomas, DTR also increased markedly (80.0 ± 15.1% (3 min), 90.7 ± 11.5% (6 min), 104.7 ± 10.6% (9 min), 117.0 ± 19.1% (12 min) and 130.3 ± 30.2% (15 min); Fig. 4(b)). The DTR of low-grade gliomas was steady over 15 min (100.8 ± 13.0% (3 min), 95.6 ± 9.9% (6 min), 95.2 ± 14.2% (9 min), 105.4 ± 15.2% (12 min) and 107.7 ± 17.5% (15 min); Fig. 4(c)). Mild increases in DTR were observed in meningiomas (93.6 ± 15.1% (3 min), 100.9 ± 8.4% (6 min), 103.4 ± 6.3% (9 min), 102.0 ± 10.9% (12 min) and 102.3 ± 16.0% (15 min); Fig. 4(d)) and pituitary adenoma (95.5 ± 13.1% (3 min), 100.8 ± 7.9% (6 min), 102.9 ± 6.2% (9 min), 101.4 ± 10.5% (12 min) and 101.9 ± 11.5% (15 min); Fig. 4(e)). The DTR of neurinomas was unchanged over 15 min (96.3 ± 17.9% (3 min), 101.6 ± 12.0% (6 min), 103.3 ± 9.4% (9 min), 99.6 ± 12.5% (12 min) and 98.9 ± 13.4% (15 min); Fig. 4(f)). The DTRs of haemangioblastomas

Dynamic thallium index of 111 lesions

Histology

Glioblastoma Anaplastic astrocytoma Low-grade glioma Meningioma Pituitary adenoma Neurinoma Haemangioblastoma

Dynamic thallium index 3 min

6 min

9 min

12 min

15 min

3.2 ± 1.2 2.2 ± 1.0 2.4 ± 0.5 6.7 ± 4.2 2.9 ± 1.1 2.2 ± 1.1 6.9 ± 1.2

3.8 ± 1.4 2.5 ± 1.1 2.3 ± 0.8 7.1 ± 4.4 3.1 ± 1.6 2.4 ± 1.2 6.6 ± 1.2

4.3 ± 1.8 3.0 ± 1.3 2.4 ± 1.1 7.2 ± 4.1 3.3 ± 2.0 2.4 ± 1.3 6.0 ± 1.3

4.6 ± 1.7 3.4 ± 1.5 2.6 ± 1.1 6.9 ± 3.6 3.2 ± 1.8 2.3 ± 1.2 5.6 ± 1.4

5.1 ± 1.8 3.7 ± 1.7 2.6 ± 0.9 6.8 ± 3.4 3.3 ± 1.8 2.3 ± 1.3 5.4 ± 1.4

Data are expressed as mean ± standard deviation.

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Fig. 3

Typical static and dynamic SPECT images. (a) Glioblastoma, (b) anaplastic astrocytoma, (c) low-grade glioma, (d) meningioma, (e) pituitary adenoma, (f) neurinoma, (g) haemangioblastoma.

were completely different from those of other histological groups, and decreased linearly (112.9 ± 6.1% (3 min), 107.6 ± 3.7% (6 min), 98.1 ± 2.3% (9 min), 90.5 ± 5.2% (12 min) and 88.0 ± 3.9% (15 min); Fig. 4(g)). Slope of dynamic thallium index linear functions

The slopes of linear functions calculated from DTRs were 1.0. to 8.4% per min (4.0 ± 2.3) for glioblastomas, – 2.0 to 9.7% per min (4.2 ± 3.8) for anaplastic astrocytomas and – 2.2 to 3.1% per min (0.8 ± 1.9) for lowgrade gliomas (Fig. 5). In low-grade gliomas, the value for oligoastrocytoma was – 0.24% per min. The slope was from – 5.3 to 5.5% per min (0.6 ± 2.6) for meningiomas, – 3.0 to 4.2% per min (0.5 ± 2.2) for pituitary adenomas and – 2.9 to 6.6% per min (0.1 ± 2.7) for neurinomas (Fig. 5). In haemangioblastomas, the slope was – 3.5 to – 0.9% per min ( – 2.2 ± 0.9) (Fig. 5). Among histological groups, the slopes for glioblastomas and anaplastic astrocytomas were significantly higher than for all other histological groups (Fig. 5). In addition, the slope for haemangioblastomas was the lowest among all histological groups. When these histological groups were divided into a malignant tumour group and a benign tumour group, as in STI analysis, the slope of the malignant tumour group was much higher than that of the benign tumour group (P < 0.001; Fig. 6). Based on a slope cut-off of 2.4% per min, 23 of 32 malignant tumour lesions (71.9%) and 65 of 79 benign tumour lesions (82.3%) could be correctly predicted with an accuracy of 79.3%.

Discussion The neuroradiological diagnosis of brain tumours has advanced considerably with the development of computed tomography (CT) and magnetic resonance imaging (MRI). However, CT and MRI do not allow assessment of tumour proliferative potency and/or viability [1]. Functional imaging with positron emission tomography (PET) has been used to assess tumour viability [14,15], but is available in only a limited number of institutions and the cost is often prohibitive. Since 201Tl uptake does not rely on breakdown of the blood–brain barrier alone, as do iodinated contrast agents in CT or gadolinium in MRI, 201 Tl SPECT has been used widely for the imaging of various brain tumours and to assess tumour viability [2,3,5,16]. In particular, some authors have described that the accumulative degree of 201Tl relates to malignancy in glioma. Kaplan et al. [3] reported that 201Tl imaging offered the most accurate correlation with viable tumours histologically in malignant glioma. Sasaki et al. [15] reported that the 201Tl uptake was found to increase in rank order with the histological grade of brain tumours and was significantly different among the gliomas. Black et al. [2] described 201Tl SPECT as being useful for predicting whether specific gliomas were of high-grade or low-grade malignancy, and to reduce unrecognized sampling errors during needle biopsies of brain tumours. However, 201Tl uptakes occur not only in biologically malignant tumours but also in benign tumours, such as meningiomas, pituitary adenomas and haemangioblastomas [5,6,17,18]. Therefore, it is difficult to estimate the grade of malignancy with the degree of accumulation only in a brain tumour. In response to the need to detect temporal changes in 201Tl in tumours, a method allowing the comparison of early images and delayed images obtained about 10–30 min and 3–4 h later, respectively, has been developed and is considered useful for evaluating tumour malignancy [7,8,10,19]. Ueda et al. [7] reported that serial 201Tl SPECT permitted the determination of the viability and malignancy grade of brain tumours. Dierckx et al. [8] suggested that clinical information and SPECT studies in combination with early and delayed imaging was expected to improve diagnostic ability. The development of SPECT equipment and the modification of peripheral devices have considerably reduced the data processing time, thereby enabling the routine use of dynamic SPECT. As a simple and rapid examination, we performed dynamic 201Tl SPECT studies for 15 min to estimate the usefulness of 201Tl dynamics in distinguishing the histological grade of brain tumours. Previous studies have demonstrated that factors influencing 201Tl accumulation in tumours include sodiumactivated and potassium-activated adenosine triphosphatase (Na + –K + ATPase) activity, tumour vascularity, and the blood–brain barrier [6,20–22]. Among these factors, Na + –K + ATPase activity and tumour vascularity have

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Tl SPECT for evaluating brain tumours Sugo et al. 147

Fig. 4

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Dynamic thallium rate (DTR) in each histological group. (a) glioblastoma, (b) anaplastic astrocytoma, (c) low-grade glioma, (d) meningioma, (e) pituitary adenoma, (f) neurinoma, (g) haemangioblastoma. * P < 0.05, **P < 0.01, *** P < 0.001.

been shown to be most closely associated with 201TlCl uptake [6,20]. Experimental evidence suggests that the ionic movements of thallium and potassium are related to active transport through the Na + –K + ATP pump and that 201Tl uptake is related to cellular growth rates [21,23]. Studies of early and delayed imaging have shown that early imaging acquired about 10–30 min after administration is related to the vascular bed of a tumour and not its malignancy [7,10]. It was reported for delayed images obtained 3–4 h later than the 201Tl accumulation

of high-grade tumours was higher than that of low-grade tumours and that delayed images are related to tumour malignancy [7,8,10]. It has been shown that the retention index, as a change in accumulation with early and delayed images, increased for malignant tumours and was unchanged or decreased for benign tumours [10]. The STI in this study, which was equal to early images, showed increased accumulation with malignancy in the glioma group, but there was no difference in malignancy in the investigation, including other benign tumours. The STI

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Fig. 5

∗

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Slope of DTR (%/minutes)

12

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∗∗

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∗

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∗

10 8 6 4 2 0 −2 −4 −6 GB

AA

LG

MM

PA

NE

HB

Relationships of slope of dynamic thallium rates (DTRs) among histological groups. GB: glioblastoma, AA: anaplastic astrocytoma, LG: low-grade glioma, MM: meningioma, PA: pituitary adenoma, NE: neurinoma, HB: haemangioblastoma. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 6

P< 0.001 12 10

Slope of DTR (%/minutes)

8 6 4 2 0 −2 −4 −6 Malignant

Benign

Comparison of the slope of the dynamic thallium rate (DTR) between malignant and benign tumour groups. Malignant: malignant tumour group, Benign: benign tumour group.

was higher in meningiomas and haemangioblastomas, which have richer vascular beds than other tumours. However, Bradley-Moore et al. [24] found that the halftime of the disappearance of 201Tl in blood was less than 1 min in a study of the dynamics of 201Tl accumulation. Atkins et al. [25] also reported that the disappearance of 201 Tl from the blood was extremely rapid, and that only 5–8% of injected activity remained in the blood at 5 min after intravenous administration. Based on these reports, the accumulation mechanism of 201Tl over 15 min cannot be explained only from the vascular bed of a tumour. Interestingly, DTR increased linearly in glioblastomas and anaplastic astrocytomas, which are histologically malignant. In contrast, the DTRs of the other benign tumours were decreased, steady, or limited to a slight increase. These results were very similar to the retention index calculated from early and delayed images. These results also indicated early images of 201Tl reflected malignancy in accumulation dynamics than in the accumulation degree. It thus seems reasonable to suppose that the degree of accumulation in a brain tumour over the entire 15 min is affected by two main factors: the isotope conformed to the amount of vascular bed and the isotope taken up and retained in tumour cells. In the results of this study, the slope of the linear function of malignant tumours was higher statistically than that of benign tumours. Therefore, it was suggested that dynamic SPECT for 15 min enabled the differentiation of benign and malignant brain tumours, which was difficult from static SPECT alone. The accuracy for distinguishing malignant and benign tumour lesions was 79.3% in this study. In retention indices calculated from early and delayed images, accuracy has been reported as 87%, and was superior to our results [12]. However, our dynamic SPECT study has some advantages compared with the estimation by early and delayed SPECT images. The dynamic SPECT study shows that an evaluation based on an accurate position is possible, so all 15 min scans can be performed without moving the patient. Furthermore, this method makes it possible to reduce the processing time for each image data set, since there is no need to adjust each imaging location given the use of continuous scanning. Another advantage of this method is its rapidity compared to early and delayed SPECT examination. Long examinations are not appropriate for brain tumour patients, who often have hemiparesis and/or disturbance of consciousness. It is thought that this method reduces the physical burden on patients. Accordingly, dynamic SPECT study may be applied as a short-cut method for early and delayed images. More important information for clinicians is that our dynamic SPECT study may be helpful in the discrimination between high-grade gliomas and low-grade gliomas. In this study, the STI of glioblastomas was higher than

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Tl SPECT for evaluating brain tumours Sugo et al. 149

that of anaplastic astrocytomas and low-grade gliomas. On the other hand, the slopes for glioblastomas and anaplastic astrocytomas were significantly higher than for low-grade gliomas, although a marked overlap exists between individual patients which may be due to the small sample size. Consequently, the slopes calculated from dynamic SPECT have potential value to determine the malignancy of gliomas. Oligodendroglial tumours have generated much interest over the past decade, due to their heightened response to chemotherapy [26]. Although only one case was oligoastrocytma in this study, the STI was the highest for low-grade gliomas, and the slope was low. Unlike astrocytic components, oligodendroglial components are well vascularized by a rich network of capillaries [27,28]. Our findings demonstrate the tumour vascularity and low-grade malignancy of oligodendroglial components.

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Conclusion It is thought that 201Tl accumulation in the early phase after administration not only contributes to vascular beds in a tumour, but also has a strong influence on tumour cell uptake and retention. We suggest that dynamic 201Tl SPECT is useful in distinguishing malignant brain tumours from benign tumours and reduces the examination burden on patients.

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Acknowledgements The authors thank T. Tachiki, T. Takano and H. Takahashi of the Section of Radiology at Toho University School of Medicine for their dedicated work on this project.

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References 1

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Original article

Functional interactions between entorhinal cortex and posterior cingulate cortex at the very early stage of Alzheimer’s disease using brain perfusion single-photon emission computed tomography Kentaro Hiraoa,d, Takashi Ohnishia, Hiroshi Matsudaa,b, Kiyotaka Nemotoa,c, Yoko Hirataa, Fumio Yamashitac, Takashi Asadac and Toshihiko Iwamotod Objective The cause of the reduced regional cerebral blood flow (rCBF) in the posterior cingulate cortex in the early stage of Alzheimer’s disease has not been clarified. In Alzheimer’s disease, the posterior cingulate cortex itself shows little neuropathologic degeneration, and a hypothesis explaining such a discrepancy is that the functional impairment in the posterior cingulate cortex reflects remote effects caused by degeneration in distant but connected areas, such as the entorhinal cortex. To test the hypothesis, we investigated the functional connectivity between the entorhinal cortex and posterior cingulate cortex. Methods Sixty-one patients with probable Alzheimer’s disease at a very early stage and 61 age-matched healthy controls underwent both brain structural magnetic resonance imaging (MRI) and single-photon emission computed tomography (SPECT). Voxel-based morphometry was performed on MRI data to identify clusters of significantly reduced grey matter concentration in patients with Alzheimer’s disease relative to controls, which were set as volumes of interest (VOIs) for correlation analyses of SPECT images. We then used adjusted rCBF values in the VOIs as covariates of interest in statistical parametric mapping. Results Voxel-based morphometry demonstrated a significant reduction in grey matter concentration in the bilateral entorhinal cortex in Alzheimer’s disease. A positive correlation between rCBF in the entorhinal cortex as VOI and that in the limbic and paralimbic systems, including the

Introduction Alzheimer’s disease is a neurodegenerative disorder leading to amnesia, cognitive impairment and dementia, and is associated with pathological neuronal changes resulting from the accumulation of b-amyloid plaques and neurofibrillary degeneration (NFD) [1]. Delacourte et al. [2] reported that NFD with paired helical filaments tau was systematically present in varying amounts in the hippocampal region, not only in the very early stage of Alzheimer’s disease, but also in non-demented aged subjects. When NFD was found in other brain areas, it

posterior cingulate cortex, anterior cingulate cortex, lingual gyri and left middle temporal gyrus (P < 0.001), was observed in Alzheimer’s disease. Control subjects also showed a similar correlation in the limbic and paralimbic systems, but not in the posterior cingulate cortex. Conclusion These results indicate that rCBF changes in the posterior cingulate cortex may be closely related to those in the entorhinal cortex in patients with Alzheimer’s disease, thereby supporting the ‘remote effect’ c 2006 hypothesis. Nucl Med Commun 27:151–156 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:151–156 Keywords: Alzheimer’s disease, mild cognitive impairment, regional cerebral blood flow, SPECT a Department of Radiology, National Center Hospital for Mental, Nervous and Muscular Disorders, National Center of Neurology and Psychiatry, Tokyo, b Department of Nuclear Medicine, Saitama Medical School Hospital, Saitama, c Department of Neuropsychiatry, Institute of Clinical Medicine, University of Tsukuba, Tsukuba and dDepartment of Geriatric Medicine, Tokyo Medical University, Tokyo, Japan.

Correspondence to Hiroshi Matsuda MD, Department of Nuclear Medicine, Saitama Medical School Hospital, 38, Morohongo, Moroyama-machi, Iruma-gun, Saitama, 350-0495, Japan. Tel: + 81 49 276 1302; fax: + 81 49 276 1301; e-mail: [email protected] Sponsorship: This study was supported by the Promotion of Fundamental Studies in Health Science of the Organization for Pharmaceuticals and Medical Devices Agency. Received 7 September 2005 Accepted 24 October 2005

was always along a stereotypical, sequential, hierarchical pathway, and the progression was categorized into several stages according to the brain regions affected. According to this report, the posterior cingulate cortex is not affected by NFD at the early stage of Alzheimer’s disease. Morphological magnetic resonance imaging (MRI) studies have demonstrated that higher atrophy rates in the medial temporal regions, such as the entorhinal cortex and hippocampus, are observed in the very early stage of

c 2006 Lippincott Williams & Wilkins 0143-3636

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Alzheimer’s disease [3–6]. Moreover, recent advances in computer-assisted statistical imaging analysis have revealed that subjects with very mild Alzheimer’s disease typically show abnormal metabolic and regional cerebral blood flow (rCBF) patterns even at the preclinical stage. Using glucose metabolism positron emission tomography (PET) with a voxel-by-voxel statistical analysis, Minoshima et al. [7] reported that the earliest changes observed in very mild Alzheimer’s disease occur in the posterior cingulate cortex. This unexpected finding has been replicated by other groups using both glucose metabolism measurements with PET and less sophisticated measurement techniques, such as rCBF measurements with single-photon emission computed tomography (SPECT). Bradley et al. [8] reported that reduced perfusion appeared between the entorhinal and limbic stages pathologically defined by Braak and Braak [1] in the posterior cingulate cortex, as well as in the anterior temporal lobe, subcallosal area and precuneus. Our previous rCBF SPECT studies demonstrated significantly decreased rCBF in the posterior cingulate cortex and precunei bilaterally in patients with mild cognitive impairment (MCI), proposed by Petersen et al. [9], when compared with controls at least 2 years before they satisfied a clinical diagnosis of Alzheimer’s disease [10,11]. We also reported a diagnostic value of reduced rCBF in the posterior cingulate cortex to assist in discriminating between patients with probable Alzheimer’s disease at the very early stage and age-matched controls before and after partial volume correction [12]. Furthermore, a PET study demonstrated hypometabolism of the posterior cingulate cortex in young subjects with a high genetic risk of developing Alzheimer’s disease [13].

The fact that the posterior cingulate cortex itself shows little degeneration neuropathologically despite the significant reduction in its rCBF or glucose metabolism has been attributed to the possibility that the posterior cingulate cortex reflects remote effects caused by degeneration in distant but connected areas, such as the entorhinal cortex. In a non-human study, Baleydier and Mauguiere [14] reported that, in the monkey, the posterior cingulate cortex receives inputs from the parahippocampal gyrus, especially the entorhinal cortex, as well as from the subiculum and presubiculum. Furthermore, Meguro et al. [15] reported that lesions of the entorhinal cortex cause long-lasting, reduced cerebral glucose metabolism in the parietal, temporal and occipital associative cortices, posterior cingulate cortex and the hippocampal regions. Few in-vivo human studies on the functional connections between the entorhinal cortex and the rest of the brain, using neuroimaging techniques, have been published [16], although the association of atrophy of the medial temporal lobe with reduced rCBF in the posterior parietotemporal cortex has been reported

in patients with a clinical and pathological diagnosis of Alzheimer’s disease [17]. In the present study, using MRI and SPECT, we examined the issue of whether functional connectivity exists between the posterior cingulate cortex and entorhinal cortex in humans, and whether the posterior cingulate cortex is subject to remote effects caused by degeneration in distant but connected areas, such as the entorhinal cortex, in the very early stage of Alzheimer’s disease.

Materials and methods We studied retrospectively 61 patients (32 men and 29 women) with MCI who showed progressive cognitive decline and eventually fulfilled the diagnosis of probable Alzheimer’s disease according to the National Institute of Neurological and Communicative Disorders and Stroke and the Alzheimer’s Disease and Related Disorders Association (NINCDS-ADRDA) criteria [18] during the subsequent follow-up period of 2–6 years. They were recruited from 350 patients complaining of memory impairment in an Outpatient Memory Clinic at the National Center Hospital for Mental, Nervous and Muscular Disorders, National Center of Neurology and Psychiatry, Tokyo, Japan. They ranged in age from 48 to 87 years with a mean ± standard deviation (SD) of 70.6 ± 8.4 years. At the first visit, they showed selective impairment in delayed recall (more than 1.5 SD below the age-matched normal mean scores) of the word-list learning test, story recall test or Rey-Osterrieth complex test on neuropsychologic examination, without an apparent loss in general cognitive, behavioral or functional status. They corresponded to the MCI criteria and scored 0.5 in the Clinical Dementia Rating [19]. The MiniMental State Examination (MMSE) score [20] ranged from 24 to 29 (mean, 26.0 ± 1.5) at the initial visit. Sixty-one control subjects (30 men and 31 women; age, 54–86 years; mean, 70.2 ± 7.3 years) were healthy volunteers without memory impairment or cognitive disorders. Specifically, their performance was within normal limits ( < 1 SD) on both the Wechsler Memory Scale-Revised and Wechsler Adult Intelligence ScaleRevised, and their MMSE score ranged from 26 to 30 (mean, 28.7 ± 1.5). They did not differ significantly in age or education from the Alzheimer’s disease patients. Spouses of the patients comprised the control subjects (not only spouses of the present patients but also spouses of other patients with advanced Alzheimer’s disease). None of the control subjects manifested cognitive changes during the follow-up period of more than 2 years. The local ethics committee approved the study for both healthy volunteers and patients with Alzheimer’s disease, all of whom gave informed consent to participate. All

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Perfusion of posterior cingulate cortex in Alzheimer’s disease Hirao et al. 153

subjects were right-handed, were screened by questionnaire with regard to their medical history and were excluded if they had neurological, psychiatric or medical conditions that could potentially affect the central nervous system, such as substance abuse or dependence, atypical headache, head trauma with loss of consciousness, asymptomatic or symptomatic cerebral infarction detected by T2-weighted MRI, hypertension, chronic lung disease, kidney disease, chronic hepatic disease, cancer or diabetes mellitus. All subjects underwent MRI and brain perfusion SPECT within 2 months after the first visit. The MRI data of a gapless series of thin sagittal sections were obtained using a three-dimensional volumetric acquisition of a T1weighted MPRage sequence (1.0 T system; Magnetom Impact Expert; Siemens, Erlangen, Germany; echo time/ repetition time, 4.4 ms/11.4 ms; flip angle, 151; acquisition matrix, 256  256; one excitation; field of view, 31.5 cm; slice thickness, 1.23 mm). For the pretreatment of voxel-based morphometry (VBM) analysis of the two groups (patients with Alzheimer’s disease and controls), image analysis was performed using Statistical Parametric Mapping 2 (SPM2; Wellcome Department of Cognitive Neurology, London, UK) running on MATLAB6.1 (Mathworks, Sherborn, Massachusetts, USA). Using SPM2 software, the original MRI images were first segmented by extraction of only grey matter, and the segmented images were spatially normalized into the standard space of Talairach and Tournoux [21]. Normalized images were then smoothed with a 12 mm full width at half-maximum isotropic Gaussian kernel to accommodate individual variability in the sulcal and gyral anatomy. The VBM analysis between patients with Alzheimer’s disease and controls was performed by group analysis of SPM2 to identify clusters of significantly reduced grey matter concentration in patients with Alzheimer’s disease relative to controls, which were set as volumes of interest (VOIs) for correlation analyses of SPECT images. Before the SPECT scan was performed, all subjects had an intravenous line established. They were injected while lying supine with their eyes closed in a dimly lit quiet room. Each subject received an intravenous injection of 600 MBq of technetium-99m ethyl cysteinate dimer (99mTc-ECD). Ten minutes after the injection of 99mTcECD, brain SPECT was performed using three-head rotating gamma cameras (Multispect3; Siemens Medical Systems, Inc., Hoffman Estates, Illinois, USA) equipped with high-resolution fan-beam collimators. For each camera, projection data were obtained in a 128  128 format for 24 angles at 50 s per angle. A Shepp and Logan Hanning filter was used for SPECT image reconstruction at 0.7 cycles/cm. Attenuation correction was performed using Chang’s method. To calculate rCBF, the linearization algorithm of a curvilinear relationship between the

brain activity and blood flow was applied, as described in previous reports [22]. Partial volume correction was performed for atrophy correction in SPECT images using the above-mentioned three-dimensional volumetric T1-weighted magnetic resonance images, as described in previous studies [12,23]. In summary, partial volume correction was performed by dividing a grey matter SPECT image by a grey matter magnetic resonance image convoluted with equivalent spatial resolution to SPECT on a voxel-byvoxel basis. In the present study, a fully automated program for the partial volume correction, developed using C ++ language, was employed. The SPECT images after partial volume correction were analyzed with SPM2. Using a template for 99mTc-ECD, the SPECT data were transformed into a standard stereotactic space. The spatial normalization algorithm of SPM2 was used for linear and non-linear transformation. A Gaussian filter (12 mm full width at halfmaximum) was used to smooth each image. The effect of global differences in rCBF between scans was removed by proportional scaling with the threshold at 20% of whole brain activity. Using MRIcro (http://www.psychology.nottingham.ac.uk/staff/cr1/mricro.html), we checked the mask image for statistical analysis and verified that medial temporal regions, including the parahippocampal gyrus and hippocampus, were encompassed in the analysis. The rCBF values of the VOIs identified by VBM analysis in SPECT images after partial volume correction were extracted for each subject. The values were then adjusted using the equation, 100  (rCBF of VOI)/(each global cerebral blood flow), and were treated as covariates of interest. Intercorrelations between different brain regions were analyzed using SPM2 to investigate functional interactions according to Horowitz et al. [24].

Results The VBM analysis demonstrated significant reductions of grey matter concentration in the left ( – 16 – 7 15, x y z; Z = 7.46) and right (18 – 9 – 16, x y z; Z = 7.45) entorhinal cortex in the very early stage of Alzheimer’s disease compared with controls (P < 0.001, corrected for multiple comparisons, Fig. 1). These areas were set as VOIs (1.4 cm3 for each hemisphere). We used the adjusted rCBF values in these entorhinal cortex VOIs as the covariates of interest for correlation analysis of rCBF SPECT. Adjusted rCBF values in the entorhinal cortex VOIs ranged from 42.3 to 142.1% (mean ± SD, 83.4 ± 17.6%) and from 67.4 to 112.2% (mean ± SD, 88.4 ± 10.5%) for patients with Alzheimer’s disease and controls, respectively. Patients with Alzheimer’s disease did not show a significant reduction in grey matter concentration in the posterior cingulate cortex compared with controls, even at a lenient threshold (P < 0.01).

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Correlation analysis (P < 0.001, corrected for multiple comparisons) revealed positive correlations between rCBF values in the entorhinal cortex and those in the limbic and paralimbic systems, including the posterior cingulate cortex, anterior cingulate cortex, lingual gyri and left middle temporal gyri, in Alzheimer’s disease. In contrast, control subjects showed positive correlations in the limbic and paralimbic systems, but not in the posterior cingulate cortex (Table 1, Fig. 2).

Discussion The VBM analysis demonstrated a significant reduction in grey matter concentration in the bilateral entorhinal cortex in the very early stage of Alzheimer’s disease compared with controls. The entorhinal cortex is a wellknown site in which pathological changes of Alzheimer’s disease occur, even at a very early stage [25]. This result corresponds to previous VBM studies [3–6]. Therefore, we believe that the results of VBM analysis confirm that the profile of SPECT is suitable for the aim of our study: the investigation of the functional interaction between

the rCBF in the entorhinal cortex and posterior cingulate cortex at the very early stage of Alzheimer’s disease. The more limited spatial resolution of SPECT scanners in comparison with PET does not allow an exact measurement of the local radiotracer concentration in brain tissue, as partial volume effects underestimate the activity in small structures of the brain. As focal brain atrophy accentuates the partial volume effect on SPECT measurements, actual rCBF values could be underestimated in the entorhinal cortex in Alzheimer’s disease. To obtain accurate rCBF correlation between the entorhinal cortex and other brain areas, rCBF was Fig. 2

Healthy volunteers

Fig. 1

L

Alzheimer's disease patients L

R

L

R

L

R R

Orthogonal sections of Statistical Parametric Mapping 2 (SPM2) results for significant decline of grey matter concentration in patients with very early Alzheimer’s disease compared with age-matched healthy volunteers ( – 16 – 7 15, x y z; Z = 7.46; 18 – 9 – 16, x y z; Z = 7.45). These regions correspond to bilateral Brodmann areas 34 (dorsal entorhinal cortex). Height threshold, < 0.001; corrected for multiple comparisons.

Table 1

Maximum intensity projections of Statistical Parametric Mapping 2 (SPM2) results for functional connectivities between the entorhinal cortex and the rest of the brain in healthy volunteers (top) and patients with Alzheimer’s disease (bottom). Height threshold, < 0.001; corrected for multiple comparisons. Local maxima of regions of correlated regional cerebral blood flow (rCBF) are given in Table 1.

Local maxima of brain areas in which regional cerebral blood flow (rCBF) is correlated with that in the entorhinal cortex Coordinates (mm) Structure

x

y

z

Z-score

Healthy volunteers

Left amygdala Right amygdala Right parahippocampal gyrus (BA35) Right superior temporal gyrus (BA21) Left insula Left parahippocampal gyrus (BA36)

– 18 18 24 53 – 40 – 22

–7 –5 – 26 –2 10 – 34

– 15 – 13 – 14 – 10 0 – 10

Infinite Infinite 5.41 4.81 4.8 4.5

Alzheimer’s disease

Left amygdala Right amygdala Right parahippocampal gyrus (BA36) Left parahippocampal gyrus (BA36) Bilateral posterior cingulate cortex (BA23) Right lingual gyrus (BA18) Bilateral dorsal posterior cingulate cortex (BA31) Left lingual gyrus (BA18) Left anterior cingulate cortex (BA24) Left middle temporal gyrus (BA21)

– 18 18 22 – 26 0 12 0 –4 –4 – 53

–7 –3 – 36 – 34 – 63 – 72 – 11 – 72 37 –1

– 15 – 15 – 13 – 13 14 –3 47 –3 0 – 10

Infinite Infinite 6.44 6.03 5.66 5.62 5.34 5.12 4.85 4.81

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Perfusion of posterior cingulate cortex in Alzheimer’s disease Hirao et al. 155

corrected for the partial volume effect in the present study. Although the correction for the partial volume effect has been reported to decrease the regional metabolic or rCBF difference between patients with Alzheimer’s disease and control subjects [26], the decrease in intersubject variations of adjusted rCBF values has been reported to increase the statistical significance [12]. In the present study, correlation analysis showed positive correlations between rCBF values in the entorhinal cortex and in the limbic and paralimbic systems, including the posterior cingulate cortex, anterior cingulate cortex and lingual gyri, in Alzheimer’s disease. In contrast, control subjects showed a correlation in the limbic and paralimbic systems, but not in the posterior cingulate cortex. Meguro et al. [15] reported that lesions of the entorhinal cortex in non-human primates cause a long-lasting reduced cerebral glucose metabolism in the hippocampus, the inferior parietal, posterior temporal and posterior cingulate cortex, and associative occipital cortices. Insausti et al. [27] also reported that the entorhinal cortex has connections to the limbic and paralimbic systems, including the anterior cingulate cortex and posterior cingulate cortex, insula in the temporal lobe, parainsula area in the parietal lobe, dorsolateral frontal cortex and an orbital region in the frontal lobe in the monkey. Our results in patients with Alzheimer’s disease agreed well with these experimental results. With regard to control subjects, who showed similar correlations in the limbic and paralimbic systems, but not in the posterior cingulate cortex, Meguro et al. [15] have demonstrated that the degree of reduced cerebral glucose metabolism in areas that have connections with the entorhinal cortex correlates significantly with the severity of histologically determined damage in the entorhinal cortex. In Alzheimer’s disease, the entorhinal cortex may be more markedly damaged than in controls, as adjusted rCBF values in the entorhinal cortex VOIs were approximately 6% lower on average in patients with Alzheimer’s disease than in controls. Although our correlation analysis showed connections with the entorhinal cortex more strongly in patients with Alzheimer’s disease than in controls, this may simply be due to a smaller range of adjusted rCBF values in the entorhinal cortex VOIs in controls than in patients with Alzheimer’s disease. Although Mosconi et al. [16] reported the loss of entorhinal cortex correlations with cerebral cortices in glucose metabolism in patients with more advanced Alzheimer’s disease, the entorhinal cortex correlation with the posterior cingulate cortex was observed in patients with Alzheimer’s disease, but not in healthy control subjects, in a similar manner to the present study.

cortex and posterior cingulate cortex in Alzheimer’s disease at the very early stage. The results indicate that rCBF changes in the posterior cingulate cortex may positively correlate with those in the entorhinal cortex through this functional connectivity. Taken together, our results may support the existence of a ‘remote effect’.

Acknowledgements We are very grateful to the technical staff of our hospital for the data acquisition of SPECT and MRI, and to Mr John Gelblum for proofreading the manuscript.

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Original article

Cerebral perfusion heterogeneity and complexity in patients with acute subarachnoid haemorrhage Timo Mustonena,b, Timo Koivistoc, Esko Vanninena, Ritva Vanninenb and Jyrki T. Kuikkaa,d Background The pathophysiological mechanisms of impaired perfusion during acute subarachnoid haemorrhage (SAH) are incompletely understood. Cerebral perfusion at the micro vascular level can be assessed by single photon emission computed tomography (SPECT). We used a SPECT approach with 99mTc-ECD to measure the cerebral perfusion heterogeneity and complexity in patients with acute aneurysmal SAH or perimesencephalic non-aneurysmal SAH (PNSAH). Methods The perfusion SPECT data of 61 patients with aneurysmal SAH, 18 patients with PNSAH, and 20 healthy control subjects were analysed by dividing the brain into 384 regions of interest. The magnitude of spatial perfusion heterogeneity was assessed by calculating the relative dispersion (RD = coefficient of variation). The fractal dimension (FD) was used to describe the overall complexity of global cerebral perfusion. Results Patients with aneurysmal SAH (RD = 11.30 ± 2.17, P < 0.001) and PNSAH (10.38 ± 2.27, P = 0.023) had a higher perfusion heterogeneity than control subjects (8.69 ± 0.80). Patients with aneurysmal SAH tended to have a higher perfusion heterogeneity than patients with PNSAH (P = 0.061). Also the overall complexity of cerebral perfusion was decreased in aneurysmal SAH (FD = 1.11 ± 0.06, P < 0.001) and PNSAH (1.11 ± 0.06,

Introduction Subarachnoid haemorrhage (SAH) is a devastating disease with a high morbidity rate despite efforts to improve treatment [1–5]. In about 80% of cases SAH is caused by a rupture of a cerebral artery aneurysm [6,7]. In 10–15% of patients the source of haemorrhage cannot be demonstrated with angiography [8,9]. The prognosis for these patients is significantly better than that for patients with identifiable pathological findings [7–10]. The bleeding pattern of non-aneurysmal SAH can be further divided into subtypes. Two thirds of patients with a negative angiography have perimesencephalic non-aneurysmal subarachnoid haemorrhage (PNSAH) [9,11]. Patients with PNSAH have an even better outcome than the rest of the patients with no identifiable source of the bleeding. They usually recover quickly, and nearly all of them can resume their normal lives [7,9,10,12]. Delayed cerebral ischaemia due to vasospasm is rarely seen in the patients with non-aneurysmal bleeding [9,10,12]. It

P = 0.004) as compared with control subjects (1.17 ± 0.06). Acute SAH causes increased regional cerebral perfusion heterogeneity and decreased overall complexity of global cerebral perfusion. Conclusion Non-invasive assessment of cerebral perfusion characteristics is feasible with SPECT and fractal analysis in patients with acute SAH and may help evaluating micro vascular function in SAH. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:157–164
Nuclear Medicine Communications 2006, 27:157–164 Keywords: cerebral perfusion, fractal analysis, SPECT, subarachnoid haemorrhage aneurysmal, subarachnoid haemorrhage perimesencephalic a

Departments of aClinical Physiology and Nuclear Medicine, bClinical Radiology, Neurosurgery, Kuopio University Hospital and dNiuvanniemi Hospital, Kuopio, Finland. c

Correspondence to Dr Jyrki T. Kuikka, Department of Clinical Physiology and Nuclear Medicine, Kuopio University Hospital, P.O.B 1777, FIN-70211 Kuopio, Finland. Tel: + 00358 17 173262; fax: + 00358 17 173244; e-mail: [email protected] This research has been supported by grants from the Kuopio University Hospital, Finnish Neurosurgical Association, The Finnish Cultural Foundation of Northern Savo and Maire Taponen Foundation. Received 3 August 2005 Accepted 16 November 2005

might be hypothesized that acute abnormalities in cerebral perfusion could explain the different outcomes. We have previously studied early alterations of cerebral perfusion in acute aneurysmal SAH with single photon emission computed tomography (SPECT) [13]. The SPECT data were analysed visually and semi-quantitatively using seven bilateral regions of interest (ROIs). Our previous studies have indicated that SPECT is sensitive in detecting decreased cerebral perfusion, and that abnormalities in regional cerebral perfusion are common in patients with ruptured aneurysms. In those patients, SPECT findings correlate with clinical vasospasm and late ischaemic deficits, but the positive predictive value of SPECT for these adverse outcomes is low [13]. On the other hand, Powsner et al. found a good correlation with hemispheric SPECT perfusion deficits and poor outcome after aneurysmal SAH [14].

c 2006 Lippincott Williams & Wilkins 0143-3636


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In the present study, we used a SPECT approach to evaluate cerebral perfusion heterogeneity and complexity at microcirculatory level in patients with acute SAH. The magnitude of spatial perfusion heterogeneity was assessed by calculating the relative dispersion (RD), which is often used to determine regional heterogeneity of perfusion, metabolism or receptor density [14]. The fractal dimension was used to describe the overall complexity of cerebral perfusion. Fractal dimension is a scale-independent measure capable of identifying subtle abnormalities of cortical perfusion, thus potentially further improving the precision of diagnostic information [15]. The aim of the present study was to compare perfusion heterogeneity and complexity in patients with acute SAH with healthy control subjects, and to compare whether patients with aneurysmal SAH have different perfusion characteristics than patients with PNSAH before treatment of the ruptured aneurysm.

Material and methods Study design

All patients from eastern Finland (population 900 000) with suspected primary SAH are referred to Kuopio University Hospital. During the period between 1 February 1995 and 28 February 1997 all patients with the diagnosis of acute ( < 72 h) SAH, based on computed tomography, were candidates for a prospective randomized study comparing endovascular and surgical treatment of ruptured aneurysms [16,17]. All patients enrolled in this study were also scheduled to undergo SPECT perfusion studies. After obtaining informed consent from the patient or the patient’s closest relative, SPECT was performed before diagnostic angiography and possible surgical or endovascular treatment. The study protocol was approved by the local ethics and radiation safety committees. In the present study, data from the acute SPECT were analysed to assess cerebral perfusion heterogeneity and complexity. We analysed the data according to the haemorrhage type (aneurysmal/non-aneurysmal), severity (Fisher grade 1–2/Fisher grade 3–5) [18], pre-treatment clinical condition (Hunt & Hess grade I–II/Hunt & Hess grade III–V) [19] and clinical outcome (Glasgow Outcome Scale, GOS) [20] after 12 months (GOS 5/GOS 1–4). Patients and control subjects

The formation of the study groups is summarized in Table 1. During the above time period, those consecutive patients whose SPECT was performed before any treatment and whose SPECT image quality was high enough to allow the analysis were included in the reanalysis. Of the 24 original non-aneurysmal SAH patients three proved to have had an aneurysmal-like haemorrhage on computed tomography, one a spontaneous intracerebral haemorrhage and one a traumatic haemorrhage;

these patients were excluded from the analysis. The clinical characteristics of the final patient groups are shown in Table 2. At the acute stage of SAH, each patient was treated in a similar manner with normovolaemia, intravenously administered betamethasone (Betapred; Defiante Farmaceutica Unipressoal Lda., Funchal, Madeira, Portugal), and Table 1

Reasons for exclusion in the original study population

(n = 94) Reason for exclusion

Aneurysmal SAH group (original, n = 70)

Non-aneurysmal SAH group (original, n = 24)

2

0

1 6 4 2

0 0

SPECT performed after treatment Technical error Poor SPECT image quality Movement artefact Insufficient field of view Not PNSAH-type nonaneurysmal bleeding Final study groups

6 61

18

SAH = subarachnoid haemorrhage; PNSAH = perimesencephalic non-aneurysmal subarachnoid haemorrhage; SPECT = single photon emission computed tomography.

Clinical characteristics of patients with subarachnoid haemorrhage (SAH)

Table 2

Variable

Aneurysmal SAH n (%)

Patients Male Female

Mean (range)

61 27 (44) 34 (56)

Age (years)

PNSAH n (%)

P Mean (range)

18 7 (39) 11 (61) 52 (14–75)

0.869

57 (40–73)

0.146

Hunt & Hess grade I–II III IV V

34 14 8 5

(56) (23) (13) (8)

13 (72) 3 (17) 2 (11) 0

Fisher grade I II III IV

5 18 25 13

(8) (30) (41) (21)

6 (32) 9 (47) 3 (16) 0

Hydrocephalus No Moderate Severe

28 (46) 25 (41) 8 (13)

9 (50) 6 (33) 3 (17)

Ventriculostomy before SPECT

10 (16)

2 (11)

0.012

Intracerebral haematoma

11 (18)

0 (0)

0.048

Time interval from SAH to SPECT study (days)

0.170

0.000

0.706

1 (0–5)

1 (0–3)

0.831

Abbreviations as in Table 1. P = statistical significance between study groups.

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Cerebral perfusion heterogeneity in acute SAH Mustonen et al. 159

intravenously administered nimodipine (Nimotop; Bayer AG, Leverkusen, Germany). All patients of Hunt & Hess grades III–V received routinely an external cerebrospinal fluid (CSF) drain. Furthermore, ventricular drainage was performed when clinical deterioration of the patient was related to development of hydrocephalus or monitoring of intracranial pressure was required during therapeutic sedation. The control group, matched for age and gender, consisted of 20 healthy individuals (six men and 14 women; mean age, 51 years; range, 27–74 years). They had reported neither neurological nor psychiatric disorder nor any kind of continuous medication. Single photon emission computed tomography

Regional cerebral perfusion/metabolism was measured with a Siemens MultiSPECT3 gamma camera equipped with fan-beam collimators (Siemens Medical Systems, Inc., Hoffman Estates, Illinois, USA) and with 99mTc labelled ethyl cysteine dimer (99mTc-ECD, Neurolite, DuPont Pharma/Durham APS, Kastrup, Denmark) as the tracer. A dose of 550 MBq was injected into the patient’s antecubital vein 30–45 min before the SPECT acquisition was started. Good grade (Hunt & Hess I–II) patients were not sedated; confused and restless (Hunt & Hess III) patients were partly sedated; and poor grade (Hunt & Hess IV–V) patients were unconscious having respira-

tory treatment. Patients were advised to keep their eyes closed during the injection of tracer. A full 3601 rotation was obtained (40 views/head, each for 35 s, matrix size 128  128), resulting in 6 to 8 million counts. The imaging resolution was 7–8 mm. Transaxial slices (3 mm thick) were reconstructed using a filtered back-projection technique (Butterworth filter: order 6 and cut-off frequency of 0.55 cm – 1). Chang’s correction method of tissue attenuation was applied with a uniform attenuation coefficient of 0.12 cm – 1. Two consecutive slices were combined to obtain a slice thickness of 6 mm and then saved onto a hard disk for further analysis (Fig. 1). Regions of interest (ROIs) were defined by using a semiautomatic brain quantification program (Siemens Medical Systems, Inc., Hoffman Estates, Illinois, USA). First, the slices were rotated and re-aligned so that the transaxial slices (x direction) are parallel to the orbitomeatal plane. Second, the coronal (z direction) slices were reconstructed and each coronal slice was segmented into 24 symmetrical sectors (12 per hemisphere) using a lower threshold of 60% for the mean cortical counts (Fig. 2). The count levels under 60% (based on our previous tested experience [21]) were removed to reduce artefacts of volume averaging errors, and to exclude white matter and background counts. The ROIs were visually inspected and cerebellar regions were not included in the analysis. Each hemisphere contained 192 sub-regions

Fig. 1

Transaxial slices (6 mm thick) in a 71-year-old female with acute SAH affecting perfusion more severely in her right hemisphere. The SPECT perfusion study was performed 45 min after injection of tracer. The colour scale is shown in the left. The patient’s right is the image left. The lower threshold is 5% and the upper threshold 95%, respectively.

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Fig. 2

Three of her 23 coronal slices and their segmentations into 24 sub-regions/each are illustrated. The count density per sub-region ranged from 6100 to 10 300 counts/cm3.

(mean volume of ROIE1 cm3), and the whole cortex contained 384 sub-regions. The fractal analysis applied to the present data has been described previously in detail [21,22]. The observed relative dispersion (RDobserved = coefficient of variation) was calculated from the mean counts per voxel in each sub region. There is a power-law relation between the observed relative dispersion (RDobserved) and the number of sub-regions (N = 3, 6, 12, 24, 48, 96, 192 and 384):   RDobserved ðN Þ N FD1 ¼ ð1Þ RDobserved ðNref Þ Nref where the reference region (Nref = 1) is an arbitrarily chosen unit reference volume of the whole organ (or tissue), and its relative dispersion can be assumed to represent the ‘apparent’ heterogeneity of the whole organ. FD is the fractal dimension. Since methodological errors occur in reconstructed SPECT images, the observed relative dispersion (RDobserved) has to be corrected by the methodological and temporal variation (RDmethod and RDtemporal) in order to achieve ‘true’ spatial dispersion (RDspatial) [21,22]: RD2observed

¼

RD2spatial

þ

RD2method

þ

RD2temporal

ð2Þ

The methodological dispersion is affected, among other things, by imaging resolution, flood field non-uniformity, statistical counting noise, attenuation, scatter, partial volume and reconstruction errors [15,21]. A cylindrical tank phantom (diameter of 20 cm and length of 10 cm) was filled with the uniform aqueous 99Tc solution (20 kBqcm – 3) and was imaged and reconstructed as human studies. The methodological dispersion for each

sub-region N was calculated. Temporal perfusion heterogeneity was assumed to be zero (RDtemporal = 0). Then, the spatial relative dispersion was calculated from Eq. 2 and fractal analysis of the spatial relative dispersion (Eq. 1) was applied for N = 24 (Fig. 3). The numerical values of FDspatial and RDspatial (N = 384) are reported. Statistical analysis

All analyses were performed with the statistical package SPSS for Windows, Version 11.0 (SPSS, Inc., Chicago, Illinois, USA). All values are expressed as mean ± SD, unless otherwise indicated. All continuous variables were tested for normal distribution with the Kolmogorov– Smirnov one-sample test (level of statistical difference at P < 0.1). The Mann–Whitney U test for continuous variables with abnormal distribution or ordinal scale variables was used for group comparisons in the case of two independent samples. Student’s t-test was applied to continuous variables with normal distribution for group comparisons. Differences were considered to be statistically significant at a two-tailed value of P < 0.05.

Results Comparability of the study groups

The aneurysmal and PNSAH patients differed statistically significantly from each other with respect to Fisher grades (P = 0.000), number of patients having received CSF drain before SPECT (0.012) and number of patients with intracerebral haematoma (ICH) (P = 0.048). The groups were comparable with respect to age and gender distribution, and Hunt & Hess grades (Table 2). The time intervals from SAH to the SPECT were comparable between the two groups. Ten aneurysmal SAH and two PNSAH patients received an external CSF drain before

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Cerebral perfusion heterogeneity in acute SAH Mustonen et al. 161

the SPECT study (P = 0.012). After the SPECT study, ventriculostomy was performed in 15 aneurysmal SAH patients and in one PNSAH patient. Eleven aneurysmal SAH patients had an ICH. In seven of them it was located in the frontal and in four in the temporal area. Based on the computed tomography scan, the calculated volume of the ICH ranged between 39 and 572 ml.

Relative dispersion (%)

FDobserved = 1.20

12

FDspatial = 1.16

9 FDmethod = 1.50 6 3 0 0

100 200 300 Number of sub-regions (N)

400

The observed (open circles), methodological (up triangles) and spatial relative dispersions (closed circles) were plotted against the number of sub regions in a SPECT perfusion study of Fig. 1. Fractal analysis of the relative dispersions with N = 24 showed high correlations (R2observed = 0.998, R2spatial = 0.992 and R2method = 0.990, respectively).

Table 3

Both patient groups showed higher relative dispersion than the control subjects (Table 3, Fig. 4). Aneurysmal SAH patients seemed to have a higher relative dispersion than the PNSAH patients, but this difference did not reach statistical significance. In secondary analyses, patients with a poorer Hunt & Hess (grade III–V) tended to have a higher relative dispersion than patients with grade I–II, but the difference did not reach statistical significance (Table 3). In contrast, a significant difference in relative dispersion was found between Fisher grades 1–2 vs. 3–4. When the most extensive haemorrhages according to Fisher’s grade (3–4) were excluded, there was no difference between the two patient groups. The aneurysmal SAH patients with an ICH had a significantly higher relative dispersion than the other aneurysmal SAH patients.

Fig. 3

15

Relative dispersion

Patients with acute hydrocephalus had a tendency for a higher relative dispersion than patients without hydrocephalus in both patient groups, but the difference did not reach statistical significance. When patients with acute hydrocephalus were excluded, relative dispersion yielded a significant difference between aneurysmal SAH and PNSAH patients (P = 0.030) (Table 3). The relative dispersion was significantly related to the clinical outcome at 12 months; patients with poorer clinical outcome (GOS 1–4) had a higher relative

Relative dispersions and fractal dimensions in patients grouped according to the type and severity of haemorrhage P**

Fractal dimension

0.000 0.023 0.000 0.000 0.000 0.000

1.11 ± 0.06 1.17 ± 0.06 1.11 ± 0.06 1.11 ± 0.06 1.11 ± 0.06 1.11 ± 0.06 1.11 ± 0.06 1.11 ± 0.06

0.823

0.000 0.024

1.11 ± 0.06 1.11 ± 0.06

0.952

0.002 0.009

11,98 ± 2,29 10,97 ± 3,76

0.304

0.000 0.361

1.11 ± 0.06 1.09 ± 0.06

0.451

0.001 0.055

34 9

11,52 ± 2,46 10,94 ± 2,25

0.560

0.000 0.007

1.10 ± 0.05 1.10 ± 0.06

0.418

0.000 0.008

27 9 11

11.02 ± 1.76 9.81 ± 2.27 12.90 ± 2.72

0.030

0.000 0.345 0.000

1.12 ± 0.07 1.12 ± 0.05 1.12 ± 0.08

0.741

0.013 0.042 0.038

50

10.95 ± 1.89

0.000

1.11 ± 0.06

Group

N

Relative dispersion (%)

All SAH patients Control subjects Aneurysmal SAH PNSAH Hunt&Hess I–II Hunt&Hess III–V Fisher 1–2 Fisher 3–4 Fisher 1–2 Aneurysmal SAH PNSAH Fisher 3–4 Aneurysmal SAH PNSAH Hydrocephalus Aneurysmal SAH PNSAH No hydrocephalus Aneurysmal SAH PNSAH Aneurysmal SAH with ICH Aneurysmal SAH without ICH

79 20 61 18 47 32 38 41

11.09 ± 2.22 8.69 ± 0.80 11.30 ± 2.17 10.38 ± 2.27 10.68 ± 1.94 11.69 ± 2.48 10.21 ± 1.66 11.90 ± 2.37

23 15

10.18 ± 1.42 10.26 ± 2.03

38 3

P

Mean ± SD

P*

P***

Mean ± SD 0.000 0.061 0.088 0.001

0.020

0.000 0.783 0.480 0.961

0.792

0.000 0.004 0.000 0.003 0.001 0.001

0.000

The relative dispersion is the spatial relative dispersion for N = 384, in %. P = statistical significance of differences of relative dispersion between groups. * P = statistical significance of differences in the fractal dimension between groups. ** P = statistical significance of differences in the relative dispersion between SAH groups and control group. *** P = statistical significance of differences in the fractal dimension between SAH groups and control group; ICH = intracerebral haematoma. Other abbreviations as in Table 1.

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162 Nuclear Medicine Communications 2006, Vol 27 No 2

Fig. 4

Fig. 5

1.3

15

1.2 Fractal dimension

Relative dispersion (%)

20

10

1.1

5 Controls

ANE-SAH

PN-SAH

Scatter plots of relative dispersion with mean ± SD of cerebral perfusion in the study groups. ANE-SAH = aneurysmal subarachnoid haemorrhage; PN-SAH = perimesencephalic non-aneurysmal subarachnoid haemorrhage.

dispersion (11.95 ± 2.44) than patients with good clinical outcome (GOS 5: 10.74 ± 2.04; P = 0.026). When the PNSAH patients were excluded relative dispersion tended to be higher in patients with poorer outcome (11.95 ± 2.44) than patients with good outcome (10.91 ± 1.92; P = 0.068). Fractal dimension

Both patient groups showed a lower fractal dimension than the control subjects (Table 3, Fig. 5). There was no difference in fractal dimension between the patient groups. No significant differences were found when the severity of the haemorrhage, clinical condition, or the existence of ICH was used as a dichotomized variable (Table 3). The fractal dimension was not significantly related to the clinical outcome at 12 months. Reproducibility

Six elderly healthy subjects were scanned twice within a 1 month interval in order to test reproducibility. Mean differences of repeated measurements were

1.0 Controls

ANE-SAH

PN-SAH

Scatter plots of fractal dimension with mean ± SD of cerebral perfusion in the study groups. ANE-SAH = aneurysmal subarachnoid haemorrhage; PN-SAH = perimesencephalic non-aneurysmal subarachnoid haemorrhage.

– 0.4% ± 1.4% (SD) for relative dispersion (384) and – 0.02 ± 0.02 for fractal dimension, respectively.

Discussion In experimental studies, acute SAH causes a major decrease in global cerebral perfusion, with a gradual return to the initial perfusion level within hours or days [23–25]. A previous comprehensive clinical study suggested that global CBF falls progressively during the first 2 weeks after SAH [26]. Early changes in the cerebral circulation have also been found to be related to clinical outcome [26,27]. However, possible changes in relative dispersion and fractal dimension are not known. Patients with an initially low CBF after SAH have an increased probability of developing cerebral ischaemia, especially if diffuse blood collection is shown by computed tomography [28]. Unlike the delayed cerebral ischaemia caused by vasospasm of the main cerebral arteries, the underlying mechanisms of acute decline in

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Cerebral perfusion heterogeneity in acute SAH Mustonen et al. 163

global perfusion are still incompletely understood. Even in patients in good clinical condition without vasospasm, the cerebral metabolic rate can be reduced [27,29,30]. In addition to a variety of metabolic factors, micro-vascular dysfunction may also play a significant role [31]. Earlier experimental studies have suggested that reduced global cerebral perfusion is likely to be caused directly by vasoconstriction [25]. In addition to vasoconstriction, the reduction of perfusion has been related to the volume of haemorrhage [24,32]. Jakobsen et al. [27] suggested that high initial rCBF values in conscious patients could be explained by early cerebral ischaemic damage and a concomitant transient luxury perfusion syndrome. According to them, the lack of luxury perfusion in patients with poor clinical grades might be due to an initially high intracranial pressure counteracting the development of hyperaemia. In our earlier SPECT study we found that both visually and semi-quantitatively detectable regional perfusion abnormalities are very common in patients with acute SAH [13]. In the present study we also included PNSAH patients, who underwent SPECT after informed consent for the whole randomized study protocol, but before diagnostic angiography. Our present results show that relative dispersion (heterogeneity) of cerebral perfusion is increased in patients with both aneurysmal SAH and PNSAH. Furthermore, increased heterogeneity is associated with more severe Fisher grades (P = 0.001), the presence of ICH (P = 0.020) and tends to be associated with more severe Hunt & Hess grades (P = 0.088). Others have found that the changes in CBF occur whether or not the SAH is due to an aneurysm demonstrable by angiography [26]. Likewise, fractal dimension (complexity) is decreased in both patient groups as compared with the healthy control group. Although there was a tendency for the perfusion to be more heterogenic in aneurysmal SAH than in PNSAH patients, there was no difference between the groups in the complexity of cerebral perfusion. This finding might, in part, be related to the fact that patients with SAH of unknown origin have been reported to have late disturbances in diffuse cognitive functions [33]. PNSAH patients in the current study were consecutive patients in the prospective series; accordingly they represent the normal population of PNSAH patients. Actually the group of PNSAH patients may be more heterogenic than believed. The assessment of both relative dispersion and fractal dimension could theoretically help in characterizing the PNSAH patients more precisely. It is now widely accepted that regional blood flow and metabolism in organs and tissues are heterogeneous [21,22,34–36]. The magnitude of spatial heterogeneity of regional perfusion is usually characterized by the relative

dispersion ( = coefficient of variation). In the present study, relative dispersion and fractal dimension were applied to acute SAH. We found that patients with SAH had a substantially greater relative dispersion than control subjects. Ten aneurysmal SAH and two PNSAH patients received an external CSF drain before the SPECT study. It might be suggested that CSF drainage would affect the findings in rCBF SPECT towards a more heterogeneous blood flow. However, CSF drainage should normalize focal blood flow disturbances caused by increased intracranial pressure, and thus CSF drainage probably diminish the findings in SPECT indicating heterogeneous blood flow. The observed values of the relative dispersions depend on the number of the sample pieces or sub-regions, and the results are not comparable between laboratories or imaging devices [14,20,22]. However, the fractal dimension (see Eq. 1) is independent of the scale of magnification and serves as a measure of the overall complexity of the system studied. The fractal dimension is 1.0 for completely homogeneous systems ( = uniform perfusion) and 1.5 for randomly distributed systems [37]. For organ blood flows, fractal dimension is usually between 1.1 and 1.3 [20,22,37], suggesting that the global perfusion heterogeneity is neither random nor uniform. In the present study of cerebral perfusion, fractal dimension was higher in the control subjects when compared to the patients with acute SAH. Bassingthwaighte and colleagues [22,34,37,38] were the first to show that the relative dispersion of blood flow distribution will scale according to Eq. 1. This empirical relationship for organ blood flow heterogeneity (Eq. 1) has also been theoretically proved [39]. The model predicts that organ blood flow, at the micro circulatory level, should also be heterogeneous. This prediction has been recently supported by micro circulatory observations [35]. The overall heterogeneity of brain blood flow measured with SPECT is well characterized by a fractal dimension when keeping in mind the important aspects of the fractal approach in SPECT [21]. Fractal dimension is an extremely compact measure of complexity, condensing all the details into a single numeric value that summarizes the irregularity of the object [40]. According to our findings, it may be too early to suggest fractal analysis as a post-processing method of SPECT images to be used in a routine clinical practise. Whole analysis including all ROIs is time consuming (requiring 60–80 min in a single patient mainly due to visual inspection of ROIs). However, higher relative dispersion was significantly related to the poor clinical outcome at 12 months. Fractal analysis may give additional information from the micro vascular function of the brain early after SAH. These findings are related to the poorly

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known extent of primary impact of SAH to CBF. In the current study SPECT was performed before the treatment of ruptured aneurysm and the possible later complications including intra-arterial vasospasm. Therefore, the value of fractal analysis in predicting outcome is to be proven in a more comprehensive study taking into account the all known factors effecting outcome.

17

18

19

Conclusion A well documented finding of both global and regional decrease in cerebral perfusion in acute SAH is also associated with increased cerebral perfusion heterogeneity and decreased overall complexity of cerebral perfusion. Although perfusion heterogeneity may be greater in aneurysmal SAH than in PNSAH, there were no differences in the fractal dimensions of cerebral perfusion in these two patient groups. Non-invasive assessment of cerebral perfusion characteristics is feasible in patients with acute SAH and may prove to be a valuable tool in the evaluation of micro vascular function in human SAH.

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Original article

The management of differentiated thyroid cancer using for imaging to assess the need for 131I therapy

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Nidhal Alia, Cherry Sebastiana, Rosemary R. Foleya, Iain Murraya, Ana L. Canizalesa, Paul J. Jenkinsb, William M. Drakeb, P. Nicholas Plowmanc, G. Michael Besserb, Shern L. Chewb, Ashley B. Grossmanb, John P. Monsonb and Keith E. Brittona Background Follow-up of 131I whole-body scanning after 131 I ablation is associated with potential stunning. Previous studies have suggested that, for scanning, 123I is more sensitive than 131I in identifying thyroid tissue, but its specificity when positive is less certain. Aim The use of 123I as an imaging agent in place of serial 131I imaging has been evaluated in the surveillance and treatment of differentiated thyroid carcinoma.

uptake of 131I when first treated and she subsequently demonstrated uptake on a second therapy. Conclusion High-dose 123I imaging is the correct predictor of the 131I post-therapy scan findings in most cases, at an administered activity that avoids stunning. As a diagnostic agent it is preferable to 131I in differentiated c 2006 thyroid carcinoma. Nucl Med Commun 27:165–169
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:165–169

Results A total of 186 studies in 136 patients with differentiated thyroid carcinoma were evaluated after total or near total thyroidectomy followed by 131I ablation. In 125 studies 123I scanning was negative and no 131I therapy was given; four patients were positive on 123I scanning but for other reasons no 131I therapy was given. In 48/49 patients a positive 123I scan was followed by positive 131I therapeutic uptake. Only one patient failed to show positive

Keywords: thyroid cancer, papillary, follicular, radioiodine,

Introduction

inherent radiation dangers, should only be given to patients in whom there is evidence of the avidity of their tumour for the chosen radiotherapeutic agent. For thyroid cancer radioiodine is used to image the patient to demonstrate iodine avidity before the prescription of radioiodine therapy.

Differentiated thyroid cancers (DTCs) include papillary and follicular cancer and their variants (including Hurthle cell neoplasm). Although DTC constitutes less than 1% of all human cancer, it is the most common endocrine cancer and is potentially curable. Patients with DTC have a 10-year cancer-specific mortality rate of less than 10% [1]. Early intervention with surgery and 131I ablation enhances remission and has the potential to reduce mortality [2]. Patients who have undergone total thyroidectomy are usually referred for ablative 131I therapy to destroy any remaining thyroid tissue. This is both to aid in the use of subsequent 131I therapy as the more avid normal thyroid is removed and also assists in determining the presence of residual and/or recurrent tumour. Ablative radioiodine therapy reduces local recurrence and improves survival [3]. Removal of all normal thyroid tissue also allows the measurement of serum thyroglobulin (Tg) to be used as an important means of monitoring the patient with thyroid cancer during follow-up. It is a basic principle of radiotherapeutic practice using unsealed sources that a radioactive therapy dose, with its

a

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Departments of Nuclear Medicine, Endocrinology and Radiotherapy, St. Bartholomew’s Hospital, London, UK. Correspondence to Professor A.B. Grossman, Department of Endocrinology, St Bartholomew’s Hospital, London EC1A 7BE, UK. Tel: + 0044 (0)207 601 8343; fax: + 0044 (0)207 601 8505; e-mail: [email protected] Received 17 October 2005 Accepted 18 November 2005

A whole-body diagnostic scan is thus undertaken to evaluate residual thyroid tissue and identify metastatic sites: 131I tracer imaging has generally been used for follow-up in most centres. The disadvantages of 131I as a diagnostic agent are two-fold. First, 131I is a poor imaging agent for the modern gamma camera because of its physical characteristics (a gamma photo peak of 364 keV). In order to compensate for the low sensitivity of the camera at this energy, the administered dose of 131I has been increased to 370 MBq (10 mCi), for example, for better imaging. Unfortunately, this causes ‘stunning’ of functioning thyroid cells as a consequence of the beta emission, which decreases the impact of a subsequent therapeutic dose of 131I administered after a diagnostic dose of this radionuclide [4–10]. It has been shown that stunning is less likely to occur with doses below 185 MBq, and thus the typical tracer dose for 131I follow-up has

c 2006 Lippincott Williams & Wilkins 0143-3636


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been in the region of 74 MBq (2 mCi). This dose, however, is too low for optimal imaging. An alternative approach, therefore, is to use an iodine radionuclide with better imaging properties, specifically 123 I, which is a pure gamma emitter with an energy of 159 keV, close to that of 99mTc. The combination of the modern gamma camera with a low energy collimator and 123 I imaging gives a count rate up to 20-fold greater and a detectability 6-fold greater compared with an equivalent activity of 131I [11,12]. A dose of 185 MBq (5 mCi) of 123I is thus equivalent to almost 2 GBq (100 mCi) of 131I. Furthermore, the radiation dose to the thyroid from this activity of 123I is less than one fifth of that due to 74 MBq (2 mCi) 131I. Thus, stunning is unlikely to occur using 123 I [13,14] and the quality of imaging is comparable to that following an 131I therapy dose. For this reason its use is increasingly being advocated as a substitute for 131I tracer imaging in the follow-up of thyroid cancer [15–21]. We have previously evaluated this technique in a pilot study [8,9] and have since adopted a protocol in which follow-up of DTC is undertaken using 123I scanning and serum Tg measurements. It was intended to avoid the prescription of 131I therapy to non-iodine avid disease as judged by high-dose 123I imaging and scans obtained after 131 I therapy. This is to avoid unnecessary radiation exposure and isolation of patients, reduce costs, and to meet the justification requirements of the UK’s Ionising Radiation (Medical Exposure) Regulations (2000). Alternatives to 131I and 123I for use in diagnostic scanning are available. The radiopharmaceuticals 201Tl [22], 99mTc sestamibi [23,24] and 99mTc tetrofosmin [25] have been used, usually in the context of non-iodine avid disease, although none has found widespread acceptance. Some authorities claim that positron emission tomography (PET) using 18F-FDG, when available, is superior to any of these [26]. The technique assesses a different physical attribute of these tumours, however, and is not appropriate in the decision as to whether patients may be suitable for radioiodine therapy. We now present the results of our experience with 123I scanning instead of using 131I tracer imaging over a period of 3 years.

Material and methods Patients

One hundred and eighty-six studies were evaluated in 136 patients with DTC (98 female, 38 male), age range 18–81 years, mean 47 years. The histopathological results revealed that 89 had papillary cancer, 36 had follicular cancer and six had Hurthle cell cancer, while five had a follicular variant of papillary cancer. The case note review was permitted by the appropriate departmental committees.

Each patient had undergone a near-total thyroidectomy or total thyroidectomy followed by ablation therapy with 3 GBq (80 mCi) of 131I usually 1–2 weeks postoperatively, followed by imaging of the head, neck and whole body 2–4 days later. While this is relatively early, the dose of radioiodine is predominantly designed to ablate the normal thyroid, so waiting for an elevated serum TSH is not thought necessary. The patients were commenced on doses of liothyronine (T3) 20 mg two or three times per day, 72 h after the ablative 131I therapy, and thereafter adjusted to be sufficient to suppress serum TSH. 123

I scintigraphy

Six months after the ablation therapy and 8 days prior to 123 I imaging, patients discontinued T3 and were asked to follow a low iodine diet and avoid iodine-containing agents. No patient was on thyroxine replacement or required exogenous recombinant TSH therapy. On the day of imaging, serum was drawn for serum Tg and thyroid function tests including serum TSH, and a pregnancy test for women of reproductive age. If the serum TSH was < 30 mUl – 1, TSH-releasing hormone (TRH; 200 mg) was administered intravenously 1 h prior to 123I scanning to increase the serum TSH concentration; if the serum TSH was > 30 mUl – 1, no TRH was given. Thus, in every case the serum TSH was elevated at the time of Tg measurement and radioiodine scanning. Head and neck, and anterior and posterior whole-body images were obtained 2 h and 24 h after administration of 123 I (185–270 MBq) intravenously. Early studies used a dose of 185 MBq, but as single photon emission computed tomography (SPECT) became more commonly used we increased the dose to 270 MBq. Similar views were taken 4–7 days post-therapy following an oral dose of 5.55 GBq of 131I when that was prescribed. The patients were imaged supine using a Millennium dual-headed Gamma Camera (G.E. Medical Systems, Milwaukee, USA) with a low energy, general purpose collimator for 123I imaging and high energy collimator for 131I post-therapy imaging, peaked appropriately for each radionuclide with ± 10% windows. For 123I a total of approximately 4 000 000 counts per image over 20 min were obtained. For the scan obtained after 131I therapy a total of 2 000 000 counts per image, over 10–12 min, were obtained. Both 123I and 131I are manufactured as totally carrier-free; 123I is manufactured in a cyclotron and 131I in a reactor. The products are manufactured by GE Healthcare (formerly Amersham Healthcare). The 123I whole-body scan was reported before any 131I therapy was given. This and the 131I post-therapy scan were reported by a single experienced nuclear medicine physician (K.E.B.).

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Thyroglobulin assay

The thyroglobulin assay is performed using DPC (EURO/ DPC, Los Angeles) Immulite 2000 analyser. This method is a solid-phase chemiluminescent–immunometric assay. Within-batch and between-batch percent coefficient of variation across the range is between 5.6 and 7.2%. We did not routinely test for anti-Tg antibodies. Follow-up protocol

This may be summarized as follows. Low risk patients (female, < 40 years of age, papillary thyroid cancer with unicentric disease and no lymph node metastases) were followed clinically after 131I ablation therapy with serial 6-monthly serum Tg (off T3 for 8 days). If serum Tg rose, an 123I study was performed with imaging at 2, 24 and if necessary 48 h after confirming that serum TSH was > 30 mUl – 1. High-risk patients (i.e., all others not fulfilling the above criteria) after 131I ablation therapy routinely had imaging with 123I at 6 months and serum Tg measurement (off T3 for 8 days, TSH > 30 mUl – 1). If imaging and serum Tg were both normal (Tg < 1 ngml – 1), follow-up was with serum Tg (off T3 for 8 days) twice at 6-monthly intervals. If serum Tg subsequently rose a further 123I study was performed: a positive 123I study led to 131I therapy which was repeated 6-monthly until the scan after 131I therapy was normal or until 45 GBq (1.2 Ci) was reached. At that point we usually limited any further treatment as we would then be concerned about the increasing risk of myelodysplasia, but each case was considered individually at a weekly multidisciplinary meeting. When the scan after 131I therapy became negative, follow-up was with serum Tg (off T3 for 8 days) alone. If 123I imaging was negative and serum Tg rose, then the 123 I scan was repeated and, if still negative, a conventional search for non-iodine avid disease was made leading to surgery, radiotherapy or medical treatment as appropriate. In exceptional cases, individual clinicians decided to initiate 131I therapy in the presence of detectable Tg levels but a negative 123I scan (see below). Localization of non-iodine avid thyroid disease was previously undertaken using 201Tl, but is now either undertaken by 99mTcMIBI or 18F-FDG PET in combination with radiological techniques, particularly MRI (for the neck) and CT (for lung fields). These agents help to localize the site of the recurrence but are not specific to thyroid cancer and do not lead to specific radionuclide therapy.

Results As previously noted, 186 studies were evaluated in 136 patients. Of these studies 125 were 123I negative and not followed by 131I therapy; four further studies were 123I positive but no 131I therapy was administered as a maximum dose of therapy had previously been delivered

I radioiodine imaging of thyroid cancer Ali et al. 167

(two patients), or further surgery had been advised (one patient), while one patient was lost to follow-up. In 50 studies a positive 123I scan was followed by 131I therapy, and in these, 49/50 were positive for both 123I imaging and the 131I post-therapy scan. However, in one of these patients a review of the 123I scan suggested that the uptake was due to oesophageal contamination while the 131I therapy scan showed uptake in the thyroid bed not seen on the diagnostic scan. This patient had an undetectable serum thyroglobulin, and was reclassified as 123 I scan negative. In a single patient the 123I scan was positive, while the first dose of 131I therapy was negative on imaging, but a subsequent second ‘tail-end’ posttherapy image was positive. Thus, there was positive concordance in 48/49 123I positive patients (Table 1). In eight of these patients serum Tg was < 2 ngml – 1, of which five showed levels < 1 ngml – 1. In eight patients the 123I scan was judged negative but the clinician decided on an individual basis to administer 131 I therapy as the serum thyroglobulin was clearly detectable. In five of these patients the 131I therapy imaging was negative, and no further radioactive iodine was given. However, in three patients 131I post-therapy imaging was clearly positive and the patients went on to receive further doses of 131I at 6-monthly intervals with maintained 131I imaging positivity (Table 1). Tg levels are available for two of these patients: they were 0.7 and 31 ngml – 1.

Discussion Radioiodine has played an important role in the management of thyroid cancer for more than four decades [27]. Trapping of radioiodine by thyroid tissue allows acquisition of scans used for the detection of residual thyroid after surgical resection and identification of local and metastatic thyroid cancer. Therapeutic doses of 131I are delivered to ablate residual thyroid tissue as well as recurrent or metastatic thyroid cancer. However, even after successful therapy patients with thyroid cancer have a life-long risk of recurrence, and 20% of them will develop local, regional (lymph node) and, less commonly, distant metastases. Patients who have undergone total thyroid ablation are followed up by diagnostic radioiodine whole-body scans and measurement of circulating thyroglobulin. The radioiodine whole-body scan requires the discontinuation 123 Table 1 I diagnostic and 131I post-therapy scans in differentiated thyroid carcinoma 131 123

I+ 123 I– Total

I+

48 3 51

131

I–

1 5 6

Total 49 8 57

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of thyroid hormones to increase serum TSH levels, although recently recombinant human TSH (rTSH) has been used in some patients instead of hormone withdrawal, the durability of the increase in serum TSH may be less than in endogenous thyroid hormone deprivation and the importance of this remains controversial. The best protocol for a diagnostic study remains unresolved. This is mainly due to the concern that the diagnostic dose of 131I could reduce the efficacy (uptake) of the subsequent therapy dose – the phenomenon of thyroid stunning [6–12]. The dose–response of stunning creates a clinical dilemma: diagnostic scans using higher doses of 131 I (185–370 MBq (5–10 mCi)) results in greater scan sensitivity but are also more likely to reduce the efficacy of therapeutic doses. 123I imaging is designed to avoid giving 131I therapy in the absence of evidence of iodineavid disease. The hypothesis that 123I imaging is more sensitive than an 131I tracer in detecting thyroid avid recurrence is based on the following rationale: the count rate of 131I tracer is 30–75 times less than that of imaging 4 days after 131I therapy; 123 K I is about six times more detectable than 131I for
the same administered activity using a figure of merit, S pffiffiffi b (where S is the signal and b the background); 123 K 185 MBq I gives less than one fifth of the radioactive absorbed dose to the thyroid compared to that from 74 MBq 131I and therefore stunning is unlikely to occur; 123 K 185 MBq I is the imaging equivalent of 3.7 GBq of 131 I (the dose given after 131I therapy). K

We now use 275 MBq 123I, which is equivalent to 5.5 GBq (150 mCi) 131I. When using SPECT this is increased to 370 MBq 123I. Thus, 123I scanning should be at least as sensitive as 131I with fewer disadvantages. However, 123I imaging does require careful attention to various details. The 123I dose contains less iodine than 131I tracer and appears to be more sensitive to iodine interference. Therefore, a particularly careful history of iodine exposure is required. Oesophageal activity from saliva may affect interpretation; liberal swallowing of water before imaging can be helpful in mitigating this problem, and SPECT can be useful. 123I uptake may also be particularly sensitive to a lack of TSH stimulation, so it is important to be certain that the serum TSH is > 30 mUl – 1. Several previous studies have indeed suggested that it may be at least as accurate in detecting disease as 131I [28–31], and it is thus highly sensitive. The results of the present study show that positive 123I imaging was a correct predictor of the 131I post-therapy scan findings in 98% of cases, and thus is highly specific as well as sensitive. In only one case was 123I scanning predictive of residual disease which was not apparent on subsequent 131I therapy, thus leading to very little unnecessary therapy, and even in this sole case subsequent 131I therapy was effective in terms of imageable uptake. There is no immediate explanation for this discrepancy between the two treatment doses. Thus,

where scanning with radioiodine is used, be the agent of choice.

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However, while we did not routinely treat 123I imaged negative patients with 131I therapy, this was carried out in eight cases in this retrospective series, and the results are illuminating. In five cases the negative 123I scan correctly predicted an absence of response to iodine therapy. Unexpectedly, in three cases overall there was clear evidence of effective radioiodine uptake therapeutically where the 123I scan had either been negative or, in one patient, was probably falsely positive with 131I uptake in 123 I scanned negative areas. In one patient, contamination by free iodine was thought to be the explanation and in the other there was a delay of over 3 months between the 123I scan and the dose after 131I therapy. This is clearly of considerable concern if 123I scanning is to be used as the sole determinant of 131I therapy. This finding may be contrasted with the 10/10 131I tracer negative patients with positive 123I scans and positive scans after 131 I therapy in the pilot study [15,16]. Serum Tg has become an important part of thyroid cancer monitoring. The amount of circulating Tg usually correlates with the extent of the disease and the tumour burden, making measurement of the Tg level helpful in determining the extent of disease. Some studies have demonstrated that a TSH-stimulated Tg test using a Tg cut-off of 2 ngml – 1 is sufficiently sensitive to be used as the principal test in the follow-up management of lowrisk patients with DTC, and have therefore suggested that the routine use of diagnostic whole body scanning with 131I is no longer necessary [32]. However, while the current study was not specifically designed to investigate the utility of serum Tg in the management of DTC, it is of some concern that eight patients in our cohort demonstrated uptake of therapeutic 131I in the face of serum Tg < 2 ngml – 1. These results, together with other published data, suggest that sole reliance on serum Tg as a predictor of residual radioiodine-sensitive disease is problematic. The results of our 3 years’ experience suggest that 123I imaging can replace serial 131I tracer studies in the followup of DTC, with the advantages of eliminating stunning, increasing sensitivity in the detection of iodine-avid recurrence and reducing unnecessary radiation exposure. However, while 123I scanning is highly specific in demonstrating radioiodine sensitivity, there remain a few patients (3/136 in this study) in whom 123I negativity is associated with positive therapeutic 131I uptake. Since we did not routinely treat patients who showed negative results after 123I scanning, this proportion may be higher. All but five patients in our series had detectable ( > 1 ngml – 1) serum Tg levels. To maximize the therapeutic potential of 131I, our data suggest that 131I therapy should be considered for all patients showing a

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positive 123I scan. In general, most regulatory authorities recommend the use of therapeutic radiotherapy only where clearly indicated, such as following a positive diagnostic scan. Currently, however, our pragmatic approach to patients showing no 123I scanning positivity is to avoid the use of 131I therapy unless there is evidence of a detectable and rising serum Tg, preferably with anatomical evidence of disease, while recognizing that some such patients would receive 131I therapy without benefit.

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In summary, our data indicate that diagnostic scanning with 123I is almost always predictive of subsequent therapeutic 131I uptake, and is, in our opinion, the tracer imaging technique of choice in patients with differentiated thyroid cancer.

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Acknowledgements We gratefully acknowledge the support of Cancer Research, UK, and Queen Mary’s School of Medicine and Dentistry, University of London.

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Morris LF, Waxman AD, Braunstein GD. Thyroid stunning. Thyroid 2003; 13:333–340. Cohen JB, Kalinyak JE, McDougall JR. Clinical implications of the differences between diagnostic 123I and post-therapy 131I scans. Nucl Med Commun 2004; 25:129–134. Britton KE, Foley RR, Siddiqi A. I-123 imaging for the prediction of 131I therapy for recurrent differentiated thyroid cancer, RDTC, when 131I tracer is negative but raised thyroglobulin,Tg. Eur J Nucl Med 1999; 26:1013. Siddiqi A, Foley RR, Britton KE, Sibtain A, Plowman PN, Grossman AB, et al. The role of 123I-diagnostic imaging in the follow-up of patients with differential thyroid carcinoma compared to 131I-scanning. Avoidance of negative therapeutic uptake due to stunning. Clin Endocrinol 2001; 55:515–521. Park HM. Invited commentary: I-123 almost a designer radioiodine for thyroid scanning. J Nucl Med 2002; 43:77–78. Shankar LK, Yamamoto AJ, Alavi IA, Mandel SJ. Comparison of 1-123 scintigraphy at 5 and 24 hours in patients with differentiated thyroid cancer. J Nucl Med 2002; 43:72–76. Britton KE, Foley RR, Chew SL. Should hTG levels in the absence of iodine uptake be treated? Eur J Nucl Med Mol Imaging 2003; 30:794–795. Gulzar Z, Jana S, Young I, Bukberg P, Yen V, Naddaf S, Abdel-Dayem HM. Neck and whole body scanning with 5-mCi dose of (123)I as diagnostic tracer in patients with well-differentiated thyroid cancer. Endocr Pract 2001; 7:244–249. Alzahrani AS, Bakheet S, Al Mandil M, Al-Hajjaj A, Almahfouza A, Al Haj A. 123 I isotope as a diagnostic agent in the follow up of patients with thyroid cancer: comparison with post 131I therapy whole body scanning. J Clin Endocrinol Metab 2001; 86:5294–5300. Lida Y, Hidaka A, Hatabu H, Kasagi K, Konishi J. Follow-up study of postoperative patients with thyroid cancer by thallium-201 scintigraphy and serum thyroglobulin measurement. J Nucl Med 1991; 32:2098–2100. Yen T-C, Lin H-D, Lee C-H, Change SL, Yeh SH. The role of technetium-99m sestamibi whole body-scan in diagnosing metastatic Hurthle cell carcinoma of the thyroid gland after total thyroidectomy: a comparison with iodine-131 and thallium-201 whole body scans. J Nucl Med 1994; 21:980–983. Rubello D, Mazzarotto R, Casara D. The role of technetium-99m methoxyisobutylisonitrile scintigraphy in the planning of therapy and followup of patients with differentiated thyroid carcinoma after surgery. Eur J Nucl Med 2000; 27:431–440. Gallowitsch H, Mikosch P, Kresnik E, Unterweger O, Gomez I, Linda P. Thyroglobulin and low dose iodine-131 and technetium-99m tetrofosmin whole-body scintigraphy in differentiated thyroid carcinoma. J Nucl Med 1998; 39:870–875. McDougall IR, Davidson J, Segall GM. Positron emission tomography and the thyroid with an emphasis on thyroid cancer. Nucl Med Commun 2001; 22:485–492. Schlumberger MJ. Papillary and follicular thyroid carcinoma. New Engl J Med 1998; 338:297–306. Yaakob W, Gordon L, Spicer KM, Nike SJ. The usefulness of iodine-123 whole-body scans in evaluating thyroid carcinoma and metastases. J Nucl Med Technol 1999; 72:279–281. Mandel SJ, Shanker LK, Bernand F, Alvai A. Superiority of iodine-123 compared with iodine-131 scanning for thyroid remnant patients with differentiated thyroid cancer. Clin Nucl Med 2001; 26:6–9. Gerard SK, Cavalieri RR. 123I diagnostic thyroid tumour whole-body scanning with imaging at 6, 24 and 48 hour. Clin Nucl Med 2002; 27:1–8. De Geus-Oei LF, Oei HY, Henneman G, Krenning EP. Sensitivity of 123I whole body scan and thyroglobulin in the detection of recurrent differentiated thyroid cancer. Eur J Nucl Med Mol Imaging 2002; 29:768–774. Mazzaferri E, Robbins RJ, Spencer CA, Braverman LE, Pacini F, Wartofsky L, et al. A consensus report of the role serum thyroglobulin as a monitoring method for low risk patients with papillary thyroid carcinoma. J Clin Endocrinol Metab 2003; 88:1433–1441.

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Original article

Localization of 131I-labelled monoclonal antibody ERIC1 in a subcutaneous xenograft model of neuroblastoma in SCID mice Christina Ottoa, Markus Jensenb,c,d, Markus Dietleina, Thomas Fischera, Matthias Schmidta, Samir Tawadrosd, Sarah Maria Bo¨rnera, Sebastian Alfred Webera, Ru¨diger Spitzb,c, Wilhelm Bloche, Frank Bertholdb,c, Harald Schichaa and Klaus Schoma¨ckera Purpose To evaluate a novel strategy of immunolocalization of human neuroblastoma by targeting the neural cell adhesion molecule (NCAM), which is over-expressed on neuroblastoma. Methods NCAM expression on the cell surface of established neuroblastoma cells was shown by flow cytometry. A SCID mouse model using IMR5-75 neuroblastoma cells to induce subcutaneous tumour growth was established. 131I was used to label monoclonal NCAM specific ERIC1 antibodies generating the 131I-ERIC1 antibody, which showed a high affinity to NCAM also after labelling (KD = 9 ¾ 10 – 8 mol
l – 1). Results Measurement of organ-specific radioactivity showed low organ-specific uptake (5.33%ID/g (percent of injected dose per gram of tissue) after 72 h), which continuously decreased over the 96 h investigation period, demonstrating clearance of radioactivity. In contrast, tumours accumulated radioactivity continuously up to a peak of 42.07%ID/g at the 96 h time point (31.07%ID/g at 72 h). This specific uptake could be blocked by application of unlabelled ERIC1 antibodies. Measurement of blood specific radioactivity revealed a characteristic clearance

Introduction Neuroblastoma is the most frequent solid extracranial childhood tumour [1] and also the most common neoplasm in the first year of life [2]. The most frequent metastatic sites are bone, bone marrow and liver [3]. By a multidisciplinary therapeutic approach, based mainly on poly-chemotherapy, an overall 5 year survival rate of approximately 67% can be reached [4]. Children with certain risk factors, such as amplification of neural myc gene, age at diagnosis > 2 years or relapsed disease, have a fundamentally poorer prognosis [4–6]. New diagnostic and treatment modalities are urgently required to offer these children a better chance of a cure. Tumour targeting with meta-[131I]iodobenzylguanidine (131I-MIBG) is a well-established method for tumour imaging and for treatment of relapsed disease. 131I-MIBG

over the first 72 h. With 37 Gy, tumour-specific radioactivity reached therapeutic doses after 96 h. Conclusions These results indicate that 131I-labelled ERIC1 has the ability to probe NCAM-expressing tumour cells in vivo with high efficiency and is a promising reagent for the diagnosis and treatment of NCAM-positive human tumours, especially for neuroblastoma. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:171–178 Nuclear Medicine Communications 2006, 27:171–178 Keywords: neuroblastoma, antibody ERIC1, anti-NCAM, radioimmunoscintigraphy, SCID mouse model Departments of aNuclear Medicine, bPaediatric Oncology and Haematology, c Centre for Molecular Medicine Cologne (CMMC), dMolecular Tumour Biology and Tumour Immunology (MTBTI) at the Department I of Internal Medicine, University of Cologne and eDepartment of Molecular and Cellular Sport Medicine, German Sport University, Cologne, Germany. Correspondence to Dr Klaus Schoma¨cker, Department of Nuclear Medicine, University of Cologne, Kerpener Strasse 62, 50924 Cologne, Germany Tel: + 0049 221 478 5977; fax: + 0049 221 478 7635; e-mail: [email protected] Received 29 September 2005 Accepted 17 November 2005

therapy can achieve an objective tumour response rate of 35%, but in fact its role is palliative [7]. Unfortunately, side effects, including dose-limiting thrombocytopenia, detract from the clinical usefulness, and about 10% of all tumours investigated showed no MIBG uptake at all and could not be treated [7]. Radioimmunotherapy with radioactively labelled antibodies directed against cell surface localized antigens on neuroblastoma might present an alternative strategy to 131 I-MIBG treatment and could also be of use in tumour imaging for detection of relapse or metastases. Several monoclonal antibodies (MoAbs) against neuroblastoma have been tested for tumour imaging and therapy in the past, some with certain success. These include antibodies against the GD2 ganglioside (MoAb 3F8 [8], 14G2A [9], ch14.18 [10]) and the L1-CAM (MoAb chCE7 [11]). For these antibodies successful targeting of tumour lesions

c 2006 Lippincott Williams & Wilkins 0143-3636

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172 Nuclear Medicine Communications 2006, Vol 27 No 2

has been reported in different cases but often the interpretation of results is inconsistent. The anti L1CAM antibody chCE7, for example, showed good therapeutic effects with the 131I-labelled antibody in mice, but turned out to have some problems with false negative results in tumour imaging in patients [11]. AntiGD2 ganglioside antibodies like ch14.18 demonstrated good and specific tumour accumulation, but therapeutic side effects; for example, fever in many cases, eruption and intense immediate neuropathic pain [12], limited the clinical usefulness. The neural cell adhesion molecule (NCAM), which is constitutively expressed on nearly 100% of all neuroblastomas [13] likewise has been successfully used as a target for immunotoxins and bispecific antibodies in small-cell lung cancer [13], glioma [14,15] and neuroblastoma [16], in vitro and in vivo.

hybridoma supernatants (Hybridoma Express medium, PAA, Linz, Austria) as described elsewhere [21]. Labelling and quality control

For this test, the antibody ERIC1 was radioiodinated with 131 I to a radiochemical concentration of 20 MBq
ml – 1 using a variation of the chloramine T method [22]. For purification a NAP-5 column (Amersham Biosciences; Sephadex G-25 medium) was used. Quality control was carried out using size exclusion HPLC (Columns TosoHaas TSKgel 2000 SK; HPLC-system: Knauer, Berlin, Germany; radioactivity detector: model ‘steffi’, Raytest, Straubenhardt, Germany). The in-vitro stability of 131I-ERIC1 was tested with HPLC 6 h after the labelling procedure. Cell line

Studies from the mid-1980s on neuroblastoma using radioiodine-labelled UJ13A, a MoAb specifically binding to NCAM, demonstrated selective uptake into neuroblastoma xenografts in SCID mice. Maximal tumour uptake in mice (9%ID/g) was observed at 24 h following injection and fell again thereafter. Results from a small cohort of children with neuroblastoma, treated with the same antibody, showed successful tumour localization [17–19]. These data were definitely encouraging and justify further work on anti-NCAM immunoscintigraphy and therapy.

IMR5-75, a human neuroblastoma cell line, was cultured in RPMI 1640 media (PAA Laboratories GmbH, Linz, Austria) supplemented with 10% fetal calf serum (PAA) and 0.5% ciprofloxacine (Bayer, Leverkusen, Germany). Medium was replaced three times weekly.

In the present study the 131I-labelled anti-NCAM antibody ERIC1 was investigated in a human neuroblastoma xenograft SCID mouse model for the first time. The main interest focused on the potential of the radioimmunoconjugate for radioimmunotherapy. Therefore, biodistribution studies were carried out to serve as basis for dose calculations.

Cell experiments Flow cytometry to detect NCAM expression on IMR5-75 cells

For a future therapeutic application of the antibody the choice of the labelling nuclide is important. The selection of radionuclide (Auger-electron emitter versus beta emitter) is determined by the cellular localization of the radioactive antibody. Consequently, internalization studies were performed for this purpose.

Material and methods

Animals

Female SCID mice (C.B-Igh-1b/IcrTac-Prkdcscid) 6–8 weeks of age, supplied by Taconic M&B, Bomholt, Denmark, and with no T and B lymphocytes but an intact NK (natural killer) cell system were used.

IMR5-75 cells were incubated with 5 mg
ml – 1 ERIC1 antibodies or an isotype control (Pharmingen, San Diego, California, USA). Bound antibodies were detected by biotinylated goat anti-mouse IgG antibodies and streptavidine–FITC (both supplied by Southern Biotechnology, Birmingham, Alabama, USA). Cells were measured in a FACSCalibur flow cytometer (Becton Dickinson, Heidelberg, Germany). Internalization studies

Sandwich methods were used, based on chemical binding between the cellularly bound ERIC1 and an anti-ERIC1 antibody linked with a chromatophore, to visualize the cellular localization of ERIC1.

Radiopharmaceuticals

[131I]iodide obtained from Tyco Healthcare GmbH, Neustadt, Germany, was used for the experiments. Antibodies

ERIC1 is a murine monoclonal antibody of the IgG1 isotype, which recognizes the 140 and 180 isoforms of the NCAM, which are the most frequent isoforms of NCAM on neuroblastoma [20]. ERIC 1 antibodies were purified by protein G affinity chromatography from serum-free

For the tri-step sandwich method cells were incubated with the primary antibody (ERIC1) for 15, 30, 45, 60, 120, 180 or 240 min in normal medium at 371C and 7% CO2 to allow antibody receptor complexes to bind and eventually internalize. Negative controls were performed by omitting the primary antibody. As secondary antibody, biotinylated goat anti-mouse Ig (1:150; DAKO, Glostrup, Denmark) was used (avidin–biotin–peroxidase complex method) [23].

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131

After fixing in 4% paraformaldehyde/phosphate-buffered saline (PBS; pH 7.4), cells were washed three times with PBS, brought onto clean glass slides coated with poly-Llysine (polysine slides, MICROM International, Walldorf, Germany) and air dried at room temperature. Cells were washed again with PBS and incubated for 5 min with 0.5 M ammonium chloride dissolved in tris-buffered saline (TBS) plus 0.25% Triton X. Cells were washed, incubated with secondary antibodies for 60 min and again washed three times. For the colour reaction, slides were coated with horseradish–peroxidase complex in TBS (1:150, self-produced) for 30 min, washed and developed for in 5–7 min under a light microscope with diaminobenzidine solution (selfproduced). Drainage followed in alcohol (70%/96%/100%/ xylol) before covering with Entellan (Merck, Darmstadt, Germany). Slides were examined by using a light microscope (Axiophot; Zeiss, Oberkochen, Germany). As an alternative method (two-step sandwich method) goat anti-mouse Ig conjugated with Cy3 fluorochrome (1:1500; Rockland, Pennsylvania, USA) was used for analyses by fluorescence microscopy. For visualization of the Cy3 conjugated antibodies, a confocal laser scanning microscope (LSM 510 Meta; Zeiss, Oberkochen, Germany) was applied. The configuration of the LSM used was wavelength 543 nm, beam splitter HFT 488/543, filters LP560 for Cy3. All steps were performed at room temperature (both methods) and in the absence of light (two-step method) to avoid bleaching of the fluorochrome. Scatchard analysis to determine binding affinity of the radioimmunoconjugate

Binding of 131I-ERIC1 was measured by incubating duplicate samples of an average of 2.3  106 cells/ml medium with increasing concentrations of 131I-ERIC1 for 30 min at room temperature. After cleaning and removal of solution, radioactivity levels within the cells were measured in a well counter. Negative controls were performed by blocking the cells with unlabelled ERIC1 before application of the 131I-ERIC1. All data were analysed by the Scatchard method [24]. The dissociation constant KD was calculated using the formula B/F = B/ KD + R0/KD, where R0 is the receptor concentration, KD the equilibrium dissociation constant, and F the free radioligand concentration. F was calculated according to F = L0 – B, where L0 is the initial radioligand concentration and B is the concentration bound specifically to receptor.

I-labelled monoclonal antibody ERIC1 Otto et al. 173

Growth of tumour xenografts in SCID mice

Mice were given intravenous injections of 20 ml anti-asialo GM1 (anti-ASGM1) rabbit antiserum (WAKO Chemicals, Du ¨sseldorf, Germany; dose recommended by manufacturer) to deplete murine natural killer cells before the subcutaneous tumour-cell challenge. Anti-ASGM1 was applied to optimize IMR5-75 growth [16] and to obtain groups of mice with homogenous tumour growth. Mice were subcutaneously injected with (2–4)  107 cells from the human neuroblastoma cell line IMR5-75 [16,25] to induce tumour xenograft growth. Anti-ASGM1 treatment was repeated 5 days after tumour cell injection. From 4 to 7 weeks after tumour cell challenge tumours became macroscopically visible and the mice could then be used for experiments. Biodistribution of radioimmunoconjugate in neuroblastoma-bearing mice

Groups of five to six mice received intravenous injections of 3–9 MBq of 131I-ERIC1 on day 0 into the tail vein. At time points 2.5, 24, 48, 72 and 96 h post-injection mice were killed by cervical dislocation. Blood, liver, spleen, kidneys, muscle, femur, thyroid, intestines, lung, tumour and urine were removed and weighed. Organ-specific radioactivity was measured in a well counter in adddition to that of the tails and remnant cadavers. Data were calculated as dose per gram of tissue (%ID/g) and tumour to non-tumour ratios. To confirm antigen specificity of 131I-ERIC1 uptake, a control group of three mice was pre-injected (i.v.) with 140 mg of unlabelled ERIC1 antibodies 24 h before injection of radioactively labelled ERIC1 to block antigen specific uptake. This group was investigated in parallel to the normal 72 h group and compared with them. Gamma camera imaging of tumour in mice

Mice were injected i.v. with approximate 4 MBq of 131Ilabelled ERIC1. To monitor biodistribution of the labelled MoAb in mice, whole-body gamma camera imaging studies were performed 72 h after injection using a Picker Corporation gamma camera (model number 882696; software from Inter Medical (aquisition program for NT-CAM, version 1.7), Espelkamp, Germany) with a 131 I collimator (static whole-body scan over 4.5 h).

Results Results of labelling

Labelling of ERIC1 was effective, and reached a yield of approximate 50–60%. Radiochemical purity was more than 95%, in-vitro stability was good with a de-iodination value of less than 5% over 6 h. Specific activity was high (32 TBq
mmol – 1).

Animal experiments

Cell experiments Expression of NCAM on human neuroblastoma

All experiments were performed in accordance with national and institutional guidelines.

The principal expression of NCAM at protein level in the neuroblastoma line used in this study could be proved by

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174 Nuclear Medicine Communications 2006, Vol 27 No 2

Internalization studies

A strong staining at the cell surface (Fig. 1(b)) after the longest observation time of 240 min could be found. Confocal images of the cells demonstrate the lack of intracellular localization of ERIC1 (Fig. 1(c)). The negative control without ERIC1 treatment does not reveal an immunostaining (Fig. 1(a)). Binding studies of

131

specific results see Table 1. The highest values in the lung tissue (17.51%ID/g) were seen 2.5 h after 131IERIC1 injection. Accumulation of radioactivity in the lung then decreased rapidly, reaching values of 8.74%ID/g at 72 h and 8.96%ID/g at 96 h. For liver and spleen, Fig. 2

Bound radioligand/free radioligand

means of flow cytometry unambiguously (unpublished data).

I-labelled ERIC1 to NCAM

The dissociation constant, KD, calculated using 10 single values by means of Scatchard plot, was 9  10 – 8 mol
l – 1 (Fig. 2). The maximum number of binding sites on each IMR5-75 cell was found to be approximately 1.9  105. Animal experiments Biodistribution studies

The radioactivity concentration in the different organs changed only slightly depending on time post-injection. Organ-specific median radioactivity in the organs measured (lung, liver, spleen, kidney, muscle, femur, thyroid gland, intestine) was 6.96, 7.19, 5.32, 5.33 and 5.81%ID/g at 2.5, 24, 48, 72 and 96 h after application of the radioiodinated MoAb, respectively (Fig. 3(a)). For organ-

0.040

Median KD = 9 × 10−8 mol.l−1, n = 10

0.035 0.030 0.025 0.020 0.015 0

200 400 Bound radioligand (pmol/l)

600

Dissociation constant of ERIC1 after radioiodination. Data were transformed using the Scatchard method. The experiment was repeated four times to obtain the median equilibrium dissociation constant, KD, as indicated on the graph.

Fig. 1

(a)

(b)

10 µm

10 µm

(c)

5 µm Internalization studies of ERIC1 on IMR5-75 cells. Cells were stained by either enzyme (HRP/DAB) or fluorescence (Cy3) detection method. Representative cuttings are shown. (a) Negative control of enzyme staining. (b) Strong staining was found at the cell surface of enzyme stained cells. (c) An inverted confocal image of a cell (fluorescence staining) demonstrating the lack of intracellular localization of ERIC1. No active internalization was visible even after the longest observation time of 240 min.

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131

I-labelled monoclonal antibody ERIC1 Otto et al. 175

Fig. 3

(a)

(b) 60 40

Tumour

Grey = tumour 50

30 %ID/g

%ID/g

40

20

30

20 Solid organs 10 10 White = blood 0

0

20

40 60 Hours

80

0

100

2.5

24 48 72 Time points (h)

96

Biodistribution of 131I-ERIC1 in IMR5-75 tumour-bearing SCID mice. The mice were killed 2.5, 24, 48, 72 or 96 h after injection of 131I labelled ERIC1 antibodies and organ-specific radioactivity was measured in a scintillation counter. (a) Closed circles = tumour; open circles = solid organs (kidney, liver, thyroid gland, muscle, femur, intestines, spleen, lung). (b) Box plot showing median and range of tumour specific radioactivity and radioactivity in the peripheral blood; for exact numerical values see Table 1. Grey boxes = tumour; white boxes = peripheral blood.

Organ-specific enrichment of radioactivity (as percent injected dose per gram of tissue) measured 2.5, 24, 48, 72 and 96 h after injection of 131I-ERIC1 in tumour-bearing SCID mice. Median values, as measured for five to six mice per time point, are indicated

Table 1 Organ

Percent injected dose per gram of tissue ( ± SD) at various times after injection 2.5 h, (n = 2)

Lung Liver Spleen Kidney Muscle Femur Thyroid Intestines Tumour Blood

17.51 9.65 7.98 9.37 1.00 3.37 4.22 2.6 3.66 44.86

(1.08) (1.40) (0.05) (1.78) (0.11) (1.31) (0.35) (0.29) (0.26) (2.53)

24 h, (n = 6) 11.6 6.23 9.82 6.82 2.85 3.98 11.14 5.15 17.19 28.9

(2.60) (1.88) (3.07) (1.69) (1.01) (0.91) (2.46) (2.17) (6.66) (10.20)

48 h, (n = 5)

72 h, (n = 5)

9.83 5.79 8.82 4.88 2.61 3.61 5.47 1.56 23.33 18.54

8.74 4.55 8.54 5.23 2.15 3.93 7.71 1.85 31.37 12.96

(1.90) (1.48) (2.01) (1.10) (1.04) (1.59) (1.90) (1.00) (3.16) (5.59)

(2.78) (1.98) (2.04) (2.43) (0.50) (0.49) (2.46) (1.19) (4.45) (1.63)

96 h, (n = 5) 10.67 9.21 8.85 6.46 4.30 4.71 6.90 3.19 42.07 28.48

(2.57) (4.00) (4.05) (2.12) (0.64) (1.80) (2.37) (1.34) (20.37) (12.09)

Blocked mice, t = 72 h (n = 3) 31.45 11.85 12.33 11.70 3.78 9.42 11.49 5.49 11.75 51.77

(1.53) (0.89) (1.39) (4.16) (2.26) (1.78) (3.60) (2.25) (1.32) (7.72)

4.55–9.65%ID/g and 7.59–9.82%ID/g, respectively, were detected. Other organs showed significantly lower values of accumulated radioactivity.

continuously over the next 3 days reaching 12.96%ID/g at 72 h, thus showing a similar clearance curve to that seen in other radiolabelled antibodies.

Tumour-specific radioactivity was initially 3.66%ID/g (2.5 h value) and then increased continuously and rapidly, reaching a maximum of 42.07%ID/g at the 96 h time point. This is 6.9 times the median value for all organs measured. Tumour-specific enrichment of radioactivity does not seem to peak at the 96 h time point since the curve (Fig. 3(a)) did not reach a plateau, but investigations were not extended to include later time points. Radioactivity levels in the blood were high (44.86%ID/g) 2.5 h after i.v. injection of 131I-ERIC1 but decreased

Unexpectedly, at the 96 h time point deviations in the %ID/g values increased drastically in tumour tissue (range 18.63–73.15%ID/g) and in blood (range 15.77–45.26%ID/g) but not in any of the organs or the remnant cadavers (see Fig. 3(b) and Table 1). In blood the tendency to an increase of radioactivity 96 h post-injection compared to 72 h post-injection4 could be observed.

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176 Nuclear Medicine Communications 2006, Vol 27 No 2

Table 2

Arithmetic means of tumour to non-tumour ratios from Table 1

Organ

Lung Liver Spleen Kidney Muscle Femur Thyroid gland Intestines Blood

Tumour-to-non-tumour ratio at various times after injection

72 h blocked

2.5 h

24 h

48 h

72 h

96 h

0.21 0.38 0.46 0.39 3.65 1.09 0.87 1.41 0.08

1.48 2.76 1.75 2.52 6.03 4.31 1.54 3.34 0.59

2.37 4.03 2.65 4.78 8.94 6.47 4.27 14.95 1.26

3.59 6.89 3.67 6.00 14.62 7.98 4.07 17.00 2.42

3.99 4.54 4.71 6.39 9.53 8.81 6.05 13.00 1.41

Interestingly, a single mouse, killed 72 h post-injection, had one metastasis next to the vertebral column. This metastasis was measured too, and showed very high uptake, with 51.5%ID/g. In general, tumour-to-normal tissue ratios increased continuously throughout the study (Table 2) and median ratios for the organs were 1.06, 2.96, 6.05, 7.97 and 7.12 at the time points 2.5, 24, 48, 72 and 96 h respectively. For organ-specific values see Table 2. Results of the blocking experiments are shown in Table 1. Tumour uptake in this mouse was only 11.75%ID/g (average). This value was close to the calculated median for organ-specific uptake and significantly lower (P = 0.0003) than in unblocked mice (31.37%ID/g). In blocked mice organ-specific radioactivity was marginal while blood-specific radioactivity (51.77%ID/g) strongly increased (Table 1).

0.37 0.99 0.95 1.00 3.11 1.25 1.02 2.14 0.23

Fig. 4

< Cranial

< Thyroid

< Chest

< Tumour

Gamma camera imaging

The scintigram (Fig. 4) shows a clear radioactive uptake in the right flank of a mouse corresponding to the tumour localization.

< Caudal

Discussion This study represents the first neuroblastoma tumour targeting with 131I labelled ERIC1 monoclonal antibodies. MoAb ERIC1 was originally generated by immunizing mice with human retinoblastoma tissue taken from a patient during surgery [20]. ERIC1 was used as carrier for the radioactivity because of the good results in clinical studies with NCAM positive gliomata [14,15]. High accumulation of activity was measured at the walls of tumour cavities in comparison to normal tissue doses, and HAMA development was low. These promising results in both studies show the good potential for NCAM as a target structure for radioimmunotherapy. For clinical application of radiolabelled antibodies, crossreactivity with normal human tissues is an important factor, because the aim is to limit the toxicity of radiation delivered by a tumour-specific antibody to normal tissue.

Whole-body gamma camera imaging of a neuroblastoma-bearing SCID mouse 72 h after i.v. injection of 3.8 MBq of 131I-ERIC1, demonstrating the tumour in the flank of the SCID mouse. Static recording over 4.5 h.

A weak cross-reaction of ERIC1 with normal fetal human tissue of neuroectodermal origin such as fetal muscle and kidney was reported [20], but no binding to pediatric or adult kidney and muscle was found. In these organs, the expression of ERIC1 antigen (NCAM) seems to be developmentally regulated [20]. NCAM is also expressed in adult human brain [20]. Since systematically applied radiolabelled antibodies do not cross the blood–brain barrier [11], expression of NCAM in normal brain does not impair the usefulness of the antibody in

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131

I-labelled monoclonal antibody ERIC1 Otto et al. 177

targeting tumours outside the brain. So NCAM, which is constitutively expressed on nearly 100% of all neuroblastomas [13], seems to be a promising antigen for targeted radiotherapy of human neuroblastoma.

The relatively high value of the lung tissue (17.51%ID/g 2.5 h after 131I-ERIC1 injection) was outstanding. This probably reflects antibody retention in lung capillaries as a kind of ‘first pass’ effect after i.v. injection.

With 131I a strong and stable labelling of MoAb ERIC1 was observed. Radiochemical purity reached good values ( Z 95%; specific activity 32 TBq
mmol – 1) and invitro stability of radioimmunoconjugate (RIC) was satisfactory.

Uptake to the tumour could be blocked by application of unlabelled ERIC1 antibodies (Table 1). In blocked mice organ-specific radioactivity was marginal while bloodspecific radioactivity (51.77%ID/g 72 h p.i.) strongly increased, which was to be expected when absorption of the 131I-MoAb by the tumour had been reduced (Table 1). In this case most of the antigen molecules (NCAM) expressed on the surfaces of the tumour cells were saturated with the premature injected unlabelled ERIC1, so 131I-ERIC1 could not bind there.

The immunohistochemical data demonstrated no active internalization of unlabelled ERIC1 antibody after binding to IMR5-75 cells. This observation does not exclude a possible internalization of antigen–antibody complexes via cell surface renewal. Because of the observed strong staining at the cell surface I was used for radiolabelling of the MoAb. Currently, 131 I is the radionuclide that is usually used for radioimmunotherapy because well-defined labelling methods are available for it. Additionally, it is abundant and relatively cheap [25]. Its beta emission is good for therapy, while its gamma emission allows its in-vitro and in-vivo distribution to be traced by using external camera imaging. Beta particles emitted by 131I have a mean range of 3 mm, allowing a ‘cross-fire’ effect between cells that bind 131I-ERIC1 and adjacent cells that do not bind the radionuclide. Similarly, it could be potentially disadvantageous in the case of marrow infiltration where adjacent haematopoietic cells would be within range. 131

For our studies SCID mice xenografted with human neuroblastoma cells of the IMR5-75 species were used, which is the neuroblastoma cell line that is usually used [16,26]. To confirm that NCAM is expressed on this cell line, too, flow cytometry investigations were undertaken with IMR5-75 and other human neuroblastoma cell lines (unpublished data). The results demonstrate expression of NCAM on all investigated cell lines. Antigen expression on IMR5-75 is at the same average level of other cell lines. This justifies the work with IMR5-75 cells, because they certainly are not biased in favour of high antigen expression. With approximately 190 000 binding sites the expression of sites seems to compare tolerably well with the neuroblastoma cell line SK-N-AS (40 000 binding sites) and SK-N-BE2 (200 000 binding sites) [11]. The dissociation constant, 9  10 – 8 mol
l – 1 (Fig. 2), is a good value and comparable to the KD of 131I rituximab (KD = 3.44  10 – 8 mol
l – 1) [27]. Accumulation of radioactivity in the investigations was both tumour-specific and antigen-specific, as demonstrated by the strong uptake of radioactivity in the tumour tissue with favourably low levels in normal tissues (Fig. 3).

Surprisingly, 5 days after i.v. application of 131I-ERIC1 an average cumulative dose of 37 Gy (values between 16 and 111 Gy) was reached in the tumour tissue. This value was calculated with a variation of the Marinelli formula [28] and is expected to be a tumour toxic dose. The great deviation of radioactivity uptake to the tumours of the 96 h cohort could be interpreted as an initial sign of tumour decay. It suggests that radioactivity is released again from the tumour and reappears in peripheral blood through decay of tumour cells, so with exception of the 96 h values radioactivity levels in the blood showed a similar clearance curve to that seen in other radiolabelled antibodies. Then, activity in blood increases again because of the release of activity from the tumour (at 96 h). Tumour uptake is still rising at this time point, too. One possible explanation could be the radiation-induced loss of tumour mass. If the tumour becomes smaller, according to the time, but still retains at least most of the radioactivity, then the %ID/g would rise. If some tumours become smaller but others do not then the large increase in variation in tumour uptake is explained. But it was not the aim of this study to supervise the growth of the tumours according to the time of treatment, so this explanation has not been proven. These observations have to follow in future studies. Although it was not the aim of the study to destroy the tumour, the high tumour-specific doses reached are sufficiently encouraging to justify further study of 131 I-ERIC1 as the therapeutic vehicle for human neuroblastoma. Future studies in mice will be needed to establish whether results obtained with the IMR5-75 line can also be obtained in other neuroblastoma cell lines or other NCAM-expressing tumour entities, as is expected. Since the doses applied in the present study were relatively high and tumour uptake was indeed good, a possible dose

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178 Nuclear Medicine Communications 2006, Vol 27 No 2

reduction could be attempted to further reduce nontumour doses. Therapeutic studies, including toxicity monitoring, still need to be carried out in mice. The use of metallic radionuclides such as 90Y, a betaparticle emitter with higher emission energy than 131I, should be tested. 90Y is preferentially used for therapeutic applications in humans. The present data on 131I-labelled ERIC1 are definitely encouraging. Future studies along the lines discussed above will improve our understanding of the mechanisms underlying 131I-ERIC1 therapy and help to optimize its use.

References Grovas A, Fremgen A, Rauck A, Ruymann FB, Hutchinson CL, Winchester DP, et al. The National Cancer Data Base report on patterns of childhood cancers in the United States. Cancer 1997; 80:2321–2332. 2 Gurney JG, Ross JA, Wall DA, Bleyer WA, Severson RK, Robinson LL. Infant cancer in the U.S.: histology-specific incidence and trends, 1973 to 1992. J Pediatr Hematol Oncol 1997; 19:428–432. 3 Kramer K, Kushner B, Heller G, Cheung NK. Neuroblastoma metastatic to the central nervous system. The Memorial Sloan-Kettering Cancer Center Experience and a literature review. Cancer 2001; 91:1510–1519. 4 Berthold F, Hero B. Neuroblastoma: current drug therapy recommendations as part of the total treatment approach. Drugs 2000; 59:1261–1277. 5 Raggi CC, Maggi M, Renzi D, Calabro` A, Bagnoni ML, Scaruffi P, et al. Quantitative determination of sst2 gene expression in neuroblastoma tumor predicts patient outcome. J Clin Endocrinol Metab 2000; 85:3866–3873. 6 Kushner BH. Neuroblastoma: A disease requiring a multitude of imaging studies. J Nucl Med 2004; 45:1172–1188. 7 Hoefnagel CA. Nuclear medicine therapy of neuroblastoma. Q J Nucl Med 1999; 43:336–343. 8 Cheung NKV, Neely JE, Landmeier B, Nelson D, Miraldi F. Targeting of ganglioside GD2 monoclonal antibody to neuroblastoma. J Nucl Med 1987; 28:1577–1583. 9 Handgretinger R, Baader P, Dopfer R, Klingebiel T, Reuland P, Treuner J, et al. A phase I study of neuroblastoma with the anti-ganglioside GD2 antibody 14.G2a. Cancer Immunol Immunother 1992; 35:199–204. 10 Mueller BM, Romerdahl CA, Gillies SD, Reisfeld RA. Enhancement of antibody-dependent cytotoxity with a chimeric anti-GD2 antibody. J Immunol 1990; 4:1382–1386. 11 Hoefnagel CA, Rutgers M, Buitenhuis CKM, Smets LA, de Kraker J, Meli M, et al. A comparison of targetting of neuroblastoma with mIBG and anti L1-CAM antibody mAb chCE7: therapeutic efficacy in a neuroblastoma xenograft model and imaging of neuroblastoma patients. Eur J Nucl Med 2001; 28:359–368. 12 Simon T, Hero B, Faldum A, Handgretinger R, Schrappe M, Niethammer D, et al. Infants with stage 4 neuroblastoma: The impact of the chimeric

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anti-GD2-antibody ch14.18 consolidation therapy. Klin Pa¨diatr 2005; 217: 147–152. Roy D, Ouellet S, Le HC, Ariniello P, Perreault C, Lambert J. Elimination of neuroblastoma and small-cell lung cancer cells with an anti-neural cell adhesion molecule immunotoxin. J Natl Cancer Inst 1996; 88:1136–1145. Papanastassiou V, Pizer BL, Coakham HB, Bullimore J, Zananiri T, Kemshead JT. Treatment of recurrent and cystic malignant gliomas by a single intracavity injection of 131I monoclonal antibody: feasibility, pharmacokinetics and dosimetry. Br J Cancer 1993; 67:144–151. Hopkins K, Chandler C, Bullimore J, Sandeman D, Coakham H, Kemshead JT. A pilot study of the treatment of patients with recurrent malignant gliomas with intratumoral yttrium-90 radioimmunoconjugates. Radiotherapy Oncol 1995; 34:121–131. Jensen M, Tawadros S, Sedlacek HH, Schultze JL, Berthold F. NK cell depletion diminish tumour-specific B cell responses. Immunol Lett 2004; 93:205–210. Lashford L, Jones D, Pritchard J, Gordon I, Breatnach F, Kemshead JT. Therapeutic application of radiolabeled monoclonal antibody UJ13A in children with disseminated neuroblastoma. Natl Cancer Inst Monogr 1987; 3:53–57. Goldman A, Vivian G, Gordon I, Pritchard J, Kemshead J. Immunolocalization of neuroblastoma using radiolabeled monoclonal antibody UJ13A. J Pediatr 1984; 105:252–256. Jones DH, Lashford LS, Dicks-Mireaux C, Kemshead JT. Comparison of Pharmacokinetics of radiolabeled monoclonal antibody UJ13A in patients and animal models. Natl Cancer Inst Monogr 1987; 3:125–130. Bourne S, Patel K, Walsh F, Popham C, Coakham H, Kemshead JT. A monoclonal antibody (ERIC-1), raised against retinoblastoma, that recognizes the neural cell adhesion molecule (NCAM) expressed on brain and tumours arising from the neuroectoderm. J Neurooncol 1991; 10:111–119. Jensen M, Ernestus K, Kemshead J, Klehr M, von Bergwelt-Baildon MS, Schinko¨the T, et al. The bi-specific CD3  NCAM antibody: a model to preactivate T cells prior to tumour cell lysis. Clin Exp Immunol 2003; 134:253–263. Greenwood FC, Hunter WM, Glover JS. The preparation of I-131-labelled human growth hormone of high specific radioactivity. Biochem J 1963; 89:114–123. Hsu SM, Raine L, Fanger H. Use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures. J Histochem Cytochem 1981; 29:577–580. Scatchard G. The attraction of proteins for small molecules and ions. Ann NY Acad Sci 1948; 51:660–672. Cardillo TM, Ying Z, Gold DV. Therapeutic advantage of 90yttrium- versus 131 iodine-labeled PAM4 antibody in experimental pancreatic cancer. Clin Cancer Res 2001; 7:3186–3192. Fichtner I, Lemm M, Becker M, Berthold F. Effects of amifostine (WR-2721, ethyol) on tumor growth and pharmacology of cytotoxic drugs in human xenotransplanted neuroblastomas. Anticancer Drugs 1997; 8:174–181. Fischer T, Zimmermanns B, Bo¨rner S, Pogge E, Tawadros S, Engert A, Schoma¨cker K. New anti-Hodgkin-radioimmunoconjugates (RIC): Consequences of variation of labelling nuclide and RIC-size to biodistribution. Nuklear Medicine 2004; 43:A132. Marinelli LD. Dosage determination in the use of radioactive isotypes. J Clin Invest 1949; 28:1271–1280.

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Original article 99m

Tc-HMPAO labelled leucocyte scintigraphy in the diagnosis of pelvic inflammatory disease Hatice Uslua, Erhan Varoglua, Sedat Kadanalib, Mustafa Yildirima, Rezzan Bayrakdara and Ayten Kadanalic Background Scintigraphy using leucocytes labelled with 99m Tc hexamethylpropyleneamine oxime (99mTc-HMPAO) is widely used for the localization of inflammatory foci and abscesses in cases of acute pelvic inflammatory disease, which is one of the serious health problems of women of child-bearing age. Early diagnosis and effective management of this condition can preserve fertility and prevent serious complications, such as peritonitis and sepsis. Aim To evaluate the importance of scintigraphy using 99m Tc-HMPAO labelled leucocytes in the early diagnosis of patients with pelvic inflammatory disease. Methods Fifteen women (mean age 29.2 ± 8 years, range 25–46 years) with suspicion of pyogenic pelvic inflammatory disease based on gynaecological examinations, clinical findings and blood tests were included in this study. The patients received 555 MBq 99mTc-HMPAO labelled leucocytes, by injection, and were scanned by scintigraphy 0.5–1, 3 and 24 h later in the anterior abdominal projection. Ten of the patients were then evaluated by abdominal or transvaginal ultrasonography, four by computed tomography and two by both ultrasound and computed tomography. The final diagnosis was made by surgical intervention. Results Scintigraphy detected pelvic inflammatory disease in five of the patients. In three of them the disease was apparent on the scans taken at 0.5–1 h, and in the other two it was apparent at 3 h. There were no false negative results,

Introduction Scintigraphy using leucocytes labelled with 99mTc hexamethylpropyleneamine oxime (99mTc-HMPAO) is widely used for the localization of inflammatory foci and abscesses in acutely ill patients [1–3]. Pelvic inflammatory disease (PID) is a serious health problem in women of childbearing age. The primary causes of PID are Neisseria gonorrhoea and Chlamydia trochomatis. PID is actually a spectrum of diseases, beginning with cervicitis and progressing to endometritis and eventually salphingitis. Sequelae of PID include ectopic pregnancy, infertility, chronic pelvic pain, hydrosalphyinx and tubo-ovarian abscesses (TOA). Early diagnosis and effective management of this condition preserves fertility and also prevents serious complications like peritonitis and sepsis [3,4]. Ultrasonography and computed tomography (CT) are rapid imaging modalities, but they are non-specific

and one false positive result. The scan accurately reflected the absence of pelvic inflammatory disease in nine patients showing non-pathological tracer uptake in the lower abdominal region. Conclusion We showed that scintigraphy with 99mTcHMPAO labelled leucocytes had a sensitivity of 100%, specificity of 90%, overall accuracy of 93%, positive predictive value of 83%, and negative predictive value of 100%. Therefore, we conclude that 99mTc-HMPAO labelled leucocyte scans provide a rapid and highly accurate method for diagnosing pelvic inflammatory disease in women of child-bearing age. This adds an important contribution to the diagnosis of infection and helps determine further operative or conservative treatc 2006 Lippincott ment. Nucl Med Commun 27:179–183 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:179–183 Keywords: pelvic inflammatory disease, scintigraphy, abdominal abscesses

99m

Tc-HMPAO labelled leucocyte

Departments of aNuclear Medicine, bObstetrics and Gynecology and cInfection Diseases, Ataturk University Medical Faculty, Erzurum, Turkey. Correspondence to Dr Hatice Uslu, Tosunpasa sok, No:13/19, 34672/Uskudar, Istanbul, Turkey. Tel: + 0090 532 436 3976; fax: + 0090 216 346 0582; e-mail: [email protected] Received 28 July 2005 Accepted 4 November 2005

and insensitive for the diagnosis of many abdominal inflammations, such as inflammatory bowel disease, appendicitis, and PID. 99mTc-HMPAO labelled leucocyte scintigraphy can localize inflammatory foci and abscesses within 4 h and even as early as 30 min after injection [2,5] and appears to be an accurate and rapid method of diagnosing abscesses and inflammations. The aim of this study was to evaluate the diagnostic importance of 99mTc-HMPAO labelled leucocyte scintigraphy in the early diagnosis of patients with PID.

Materials and methods Fifteen women (mean age 29.2 ± 8 years, range 25–46 years) in whom pyogenic PID was suspected based on the results of gynaecological examinations, clinical findings and blood tests were included in this study. These

c 2006 Lippincott Williams & Wilkins 0143-3636

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180

Nuclear Medicine Communications 2006, Vol 27 No 2

patients underwent 99mTc-HMPAO labelled leucocyte scintigraphy and were subsequently examined by abdominal or transvaginal ultrasonography (10 patients), CT (four patients) and both ultrasound and CT (two patients). The final diagnosis was made by laparoscopy in nine patients and by laparotomy in six. Labelling of leucocytes with

99m

Tc-HMPAO

Mixed leucocytes were isolated and labelled as described previously [6]. A 40 ml sample of venous blood was taken into a 60 ml syringe containing 10 ml of acid citrate dextrose and 10 ml of 6% hydroxyethyl starch. After sedimentation for 1 h at room temperature, the supernatant was centrifuged in sterile tubes at 100  g for 5 min. The leucocytes were resuspended in 1 ml of cellfree autologous plasma. 99mTc-HMPAO was prepared according to the instructions supplied (Ceretec, Amersham International). Five millilitres (555 MBq) of 99mTcHMPAO complex were added to the leucocyte suspension and incubated for 10 min at room temperature. The suspension was centrifuged again at 100  g for 5 min, the supernatant was discarded, and the cells were resuspended in 5 ml of cell-free plasma and re-injected intravenously. The mean cell labelling efficiency was 42% (range 22–76%) and the mean dose injected was 180 MBq (range 80–350 MBq). Scans were performed 0.5–1, 3 and 24 h in the anterior abdominal projection after injection. In addition to the posterior abdominal view, right or left anterior oblique views were often used to localize the tracer uptake. The study was performed using single-head gamma camera system (GE 4000 XC/T) equipped with a low energy, all purpose collimator interfaced to a General Electric Computer system. Images were obtained in a 256  256 matrix and a 1.33 zoom was selected. All images were examined by two experienced nuclear medicine physicians who were blinded to each other and the clinical history and diagnosis. They evaluated the pathological and non-pathological tracer uptake as positive or negative.

Results Positive scintigraphic uptake showed circular shaped areas of increased accumulation of leucocytes in the genital region compatible with inflammation. Pelvic inflammatory disease was found in five patients. For three of the patients the disease was apparent on the scans taken at 0.5–1 h, while for the other two patients the disease was apparent on the 3 h scan. The abnormal activity increased with time (Fig. 1(a–d)). However, images obtained at 3 h did not give additional information compared with the 1 h images in any patient, with the exception of true negative images at 3 h. In patients with positive scintigraphic images, TOA was demonstrated by

CT (Fig. 2), and ultrasonography (Fig. 3). Laparoscopy or laparotomy confirmed that all five were true positive cases. Thus, four were found to have TOA and one pyosalpinx. One false positive result occurred in a patient who had chronic inflammatory bowel disease of the ulcerative colitis type (Fig. 4). Verification of an infection or inflammation in this patient was obtained by surgical and pathological diagnosis. Nine cases that showed no pathological uptake of tracer in the lower abdominal region for PID were scintigraphically true negatives, as subsequently corroborated by laparoscopy or laparotomy. In six of these patients there was no pathological pelvic mass. The other three patients had non-infected ovarian cysts. In our study, 99mTc-HMPAO labelled leucocyte scintigraphy yielded no false negative results for pelvic inflammatory diseases. The results of different diagnostic procedures in the patients are presented in Table 1. In this study, 99mTcHMPAO labelled leucocyte scintigraphy had a sensitivity of 100%, specificity of 90%, overall accuracy of 93%, positive predictive value of 83% and negative predictive value of 100% (Table 2).

Discussion The preparation of 99mTc-HMPAO labelled leucocytes in vitro was first described in 1986 [7]. Since then scintigraphy using 99mTc-HMPAO labelled leucocytes has been used successfully for imaging a wide variety of inflammatory diseases [7,8]. For example, it has been widely used to localize sites of leucocyte infiltration and inflamed tissues in abdominal infections [4,9], ulcerative colitis [10], acute pancreatitis [11], pelvic inflammatory disease [3,12], bone infection, acute appendicitis [13] and undiagnosed fever, but has not been as useful for detecting chronic inflammation [14]. The 99mTc-HMPAO leucocyte method has some advantages compared with 67Ga citrate and 111In labelled agents, and with 99mTc labelled human immunoglobulin G (HIG). The main disadvantage of these agents has been the diagnostic delay, which can be at least 24 h. There have been reports indicating that abdominal infections can be detected as early as 0.5 h following injection of 99mTc-HMPAO leucocytes [7,8] and we successfully detected infections in three patients in 0.5–1 h. Accumulation of 67Ga and macromolecules such as HIG on the basis of increased endothelial permeability may be preferable to labelled leucocytes in chronic inflammation [15]. Other advantages of the 99mTcHMPAO leucocyte procedure relative to 111In labelled leucocytes and 67Ga citrate includes the considerably lower radiation dose, ready availability and superior images [16,17]. A significant disadvantage of labelled leucocytes is the need for in-vitro isolation of blood cells, which can expose the patients and laboratory technicians to the hazards of infection.

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Scintigraphy for diagnosing pelvic inflammatory disease Uslu et al. 181

Fig. 1

Anterior abdominal images at 0.5 h (a), 1 h (b), 3 h (c), and 24 h (d) after the injection of 99mTc-HMPAO labelled leucocytes in a patient with a tuboovarian abscess.

Fig. 2

A computed tomography image that shows a tubo-ovarian abscess.

The non-specific accumulation in the bowel which appears after 4 h following injection has been mentioned as a potential disadvantage of the 99mTc-HMPAO labelled

Fig. 3

The tubo-ovarian abscess is shown by the ultrasonography image.

leucocyte procedure [17], but, the physiological bowel activity did not interfere with accurate diagnosis (Fig. 5). The pathological uptake was always more intense than

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182 Nuclear Medicine Communications 2006, Vol 27 No 2

the bowel activity, and the intensity increased over time (Fig. 1(a–d)) [3]. In the case of inflammatory bowel disease, the activity was intense and the location did not change [18]. This situation sometimes causes false positive results, as observed in our study (Fig. 4). Moreover, it is suggested that the other lower abdominal Fig. 4

infections (i.e., appendicitis, pelvic abscess) should be ruled out. Pelvic inflammatory disease is one of the most infectious diseases of women at a reproductive age. Early and accurate diagnosis and successful treatment of PID can have an important impact on a woman’s health. Early diagnosis of PID may prevent serious surgical complications such as peritonitis and sepsis. Computed tomography and magnetic resonance imaging (MRI) are appropriate methods for imaging patients with an intra-abdominal abscess. However, scintigraphic techniques have an advantage over CT and MRI because the former evaluates the entire body for infection, while radionuclides can differentiate an infectious locus from non-infectious fluid collections [16]. Also, since the whole body is imaged by scintigraphy, unsuspected sites of infection may be found [19]. Scintigraphy with 99mTcHMPAO labelled leucocytes offers an alternative and rapid method by which images can be obtained within 3–4 h. Although the labelling is time-consuming, the diagnosis is usually available in less than 2 h. There were no false negative results, but one false positive result in our group of patients with PID. A similar result has been reported by Rachinsky et al. [3]. They found that the overall sensitivity, specificity, accuracy, and positive and negative predictive values were 100%, 91.6%, 95%, 89% and 100%, respectively, for scintigraphy with Results of Tc-99m HMPAO labeled leukocyte scintigraphy imaging methods in patients with PID

Table 2

Abnormal activity is seen in the right side of lower abdomen in patients with inflammatory bowel disease (ulcerative colitis).

Table 1

Sensitivity Specificity Overall accuracy Positive predictive value Negative predictive value

100% 90% 93% 83% 100%

Summary of the patients

Patient number

Age (years)

PV

Scintigraphy

Computed tomography

Ultrasound

Final diagnosis Laparoscopy

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

30 28 33 40 42 38 37 45 25 37 36 29 46 27 33

+ + + + + + + + + + + + + + +

+ – + – – – + – + + – – – + –

+

–

+ ± + – – +

–

Laparotomy TOA

NP PS NIOC NP NIOC TOA NP

±

IBD TOA

+ –

NP NP

+ +

NIOC TOA NP

TOA: tubo-ovarian abscess; PS: pyosalpinx; NP: no pathology; NIOC: non-infected ovarian cyst; IBD: inflammatory bowel disease; PV, bimanual vaginal examination.

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Scintigraphy for diagnosing pelvic inflammatory disease Uslu et al. 183

Fig. 5

quality and makes an important contribution to the diagnosis of infection, which determines further operative or conservative treatment. Therefore, we recommend this scan as one of the steps for rapid and accurate diagnosis of pelvic inflammatory disease.

References 1

2 3

4

5

6

7

An example of no abnormal uptake. The scan demonstrates nonspecific accumulation of the tracer in the bowel.

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9 99m

Tc-HMPAO labelled leucocytes in the diagnosis. They also did not obtain false negative scintigraphic findings. In the other study, Lantto [2] reported that the sensitivity, specificity and accuracy rates were 90%, 91% and 91%, respectively, in patients with suspected intraabdominal abscesses. Their sensitivity rate was lower than our value. Thus, we had a sensitivity of 100%, specificity of 90%, overall accuracy of 93%, positive predictive value of 83% and negative predictive value of 100%. Importantly, we found that the correlation between scintigraphic and laparoscopic findings was sufficiently close to potentially reduce the need for CT and diagnostic laparoscopy. A negative finding by using 99m Tc-HMPAO leucocyte scanning might prevent unnecessary invasive surgical interventions.

10

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As a result of the findings of our study, we concluded that 99m Tc-HMPAO labelled leucocyte scans provide a rapid and highly accurate method for diagnosing PID in women of child-bearing age. Early diagnosis and effective management of PID preserves fertility and also prevents serious complications like peritonitis and sepsis. 99mTcHMPAO labelled leucocyte scintigraphy is a non-invasive, safe and accurate procedure with excellent imaging

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Lantsberg S, Rachinsky I, Boguslavsky L. Tubo-ovarian abscess: Tc-99m hexamethylpropylene amine oxime leukocyte scintigraphy and correlative imaging. Clin Nucl Med 1999; 24:796–797. Lantto EH. Leucocytes labelled with 99mTc-HMPAO in the detection of abdominal abscesses. Eur J Surg 1991; 157:469–472. Rachinsky I, Boguslavsky L, Goldstein D, Golan H, Pak I, Katz M, Lantsberg S. Diagnosis of pyogenic pelvic inflammatory diseases by 99mTc-HMPAO leucocyte scintigraphy. Eur J Nucl Med 2000; 27:1774–1777. Agranovich S, Rachinsky I, Pak I, Benkovich E, Lantsberg S. The usefulness of Tc-99m-HMPAO-labeled leukocyte scintigraphy in the diagnosis of multiple intra-abdominal abscesses following in vitro fertilization (IVF) procedure. Eur J Obstet Gynecol Reprod Biol 2004; 116:103–105. Lantto EH, Lantto TJ, Vorne M. Fast diagnosis of abdominal infections and inflammations with technetium-99m-HMPAO labeled leukocytes. J Nucl Med 1991; 32:2029–2034. Vorne M, Soini I, Lantto T, Paakkinen S. Technetium-99m HM-PAO-labeled leukocytes in detection of inflammatory lesions: comparison with gallium-67 citrate. J Nucl Med 1989; 30:1332–1336. Peters AM, Osman S, Henderson BL. Clinical experience with 99m-Tchexamethyleneamineoxime for labelling leukocytes and imaging inflammation. Lancet 1986; 2:946–949. Vorne M, Soinin I, Lantto T. Technetium-99m-HM-PAO-labeled leukocytes in detection of inflammatory lesions: comparison with gallium-67-citrate. J Nucl Med 1989; 30:1332–1336. Lantto EH, Lantto TJ, Vorne M. Fast diagnosis of abdominal infections and inflammations with technetium-99m-HMPAO labeled leukocytes. J Nucl Med 1991; 32:2029–2034. Amer S, Bodemar G, Lindstrom E. Air enema radiology compared with leukocyte scintigraphy for imaging inflammation in active ulcerative colitis. Eur J Gastroenterol Hepatol 1995; 7:59–64. Scho¨lmerich J, Schu¨michen C, Lausen M, Gross V, Leser HG, Lay L, et al. Scintigraphic assessment of leukocyte infiltration in acute pancreatitis using technetium-99m-hexamethyl propylene amine oxine as leukocyte label. Dig Dis Sci 1991; 36:65–70. Uslu H, Varoglu E, Kadanaly´ S, Yildirim M, Demirci M, Sahin A. Tc-99m HMPAO labeled leukocytes scintigraphy in the diagnosis of pelvic inflammatory disease [Abstract]. Eur J Nucl Med 2000; 27(8 suppl):1088. Varoglu E, Polat KY, Tastekin G, Akc¸ay F, Polat C. Diagnostic value of Tc99m HIG scintigraphy in the detection of acute appendicitis. Clin Nucl Med 1996; 21:645–647. Peters AM. The use of nuclear medicine in infections. Br J Radiol 1998; 71:252–261. Oyen WJG, Claessens RAMJ, van der Meer JWM. Detection of subacute infectious foci with In-111-labelled autologous leukocytes and In-111labeled human nonspecific immunoglobulin G: a prospective comparative study. J Nucl Med 1991; 32:1854–1860. Datz FL. Abdominal abscess detection: gallium, 111In-, and 99mTc-labeled leukocytes, and polyclonal and monoclonal antibodies. Semin Nucl Med 1996; 26:51–64. Hotze A, Kropp J, Kozak B. 99m-Tc-HMPAO leukocyte imaging. First clinical results and possible problems in inflammatory abdominal diseases. J Nucl Med 1988; 27:115–116. Li DJ, Middleton SJ, Wright EP. 99m-Tc-HMPAO-labelled leukocyte scintigraphy in inflammatory bowel disease. Nucl Med Commun 1990; 12:889. Seabold JE, Wilson DG, Lieberman LM, Boyd CM. Unsuspected extraabdominal sites of infection: scintigraphic detection with indium-111-labeled leukocytes. Radiology 1984; 151:213–217.

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Original article

Localized internal radiotherapy with 90Y particles embedded in a new thermosetting alginate gel: A feasibility study in pigs Øyvind Holtea, Arne Skrettingb, Tore Bach-Gansmoc, Per Kristian Hold, Kjersti Johnsrudc, Hanne Hjorth Tønnesena and Jan Karlsena Background Internal radiotherapy requires the localization of the radionuclide to the site of action. A new injectable alginate gel formulation intended to undergo immediate gelation in tissues and capable of encapsulating radioactive particles containing 90Y was investigated. Methods The formulation was injected intramuscularly, into the bone marrow compartment of the femur and intravenously, respectively, in pigs. The distribution of radioactivity in various tissues was determined.

intravenous injection, 80–90% of the radioactivity was found in the lungs. Conclusion The present formulation appears suitable for localized radiotherapy in organs and tissues having c 2006 low perfusion. Nucl Med Commun 27:185–190 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:185–190 Keywords: yttrium-90 silicate, alginate, animal experiments, injectables, gels, localized radiation therapy a

Results Following intramuscular injection, more than 90% of the radioactivity was found at the site of injection. Following injection into bone marrow, 30–40% of the radioactivity was retained at the site of injection, but a considerable amount of radioactivity was also detected in the lungs (35–45%) and the liver (5–18%). Following

Introduction The use of radioactive particles for internal radiation therapy is an attractive technique for localized destruction of malignant cells and tissue [1]. The specific localization of the radionuclide to the site of action is critical for treatment efficacy and necessary to avoid unwanted side effects. Only body compartments capable of retaining particulate material are suitable for such radiotherapy. Examples include the peritoneal cavity and the synovia of joints [1–5]. Radiotherapy of tumours in the liver has been performed by selective injection into a hepatic artery of radioactive particles [6–11] or radioactive oil [12–14]. The formation of complexes with the gel-forming agent chitosan has enabled the retention of certain radionuclides in the knee joint [15,16], the prostate [17] or the liver [18]. In previous work, we described the development of a new injectable pharmaceutical formulation making possible the use of radioactive particles in a wider range of organs and tissues [19]. In that work, the radioactive particles containing 90Y were encapsulated in a biocompatible alginate gel matrix, thus preventing the particles from leaking from the site of application. The in-vitro experiments demonstrated that the formulation is stable at least 1 week. This corresponds to almost three times the halflife of 90Y. The gel formulation is a viscous fluid when injected, and solidifies to a gel when exposed to normal

School of Pharmacy, University of Oslo, and, Departments of bMedical Physics, Nuclear Medicine and dThe Interventional Centre, RikshospitaletRadiumhospitalet Health Enterprise, Oslo, Norway. c

Correspondence to Øyvind Holte, University of Oslo, School of Pharmacy, P.O. Box 1068 Blindern, N-0316 Oslo, Norway. Tel: + 0047 228 56589; fax: + 0047 228 54402; e-mail: [email protected] Received 3 October 2005 Accepted 18 November 2005

body temperature. With this technique, it would not be necessary to perform surgery in order to place the gel formulation at the intended site, but the formulation may be injected through 18G needles. Thus, it should be possible to easily place the formulation into or adjacent to a wide range of organs and tissues. The gel formulation is a mixture of calcium chloride encapsulated in liposomes, sodium alginate solution and particulate 90Y silica (YMM1). Sodium alginate is a linear polysaccharide composed of uronic acid residues. The polymer is water soluble at neutral pH, but forms a gel when mixed with certain divalent cations like Ca2 + [20]. The main phase transition temperature of the liposomes is 361C; thus they retain the encapsulated calcium chloride at room temperature, but on heating to normal body temperature, calcium chloride is released to the liposomes’ external phase. Rapid gel formation occurs, and the radioactive particles are trapped in the gel formed at the site of injection. Alginate gels are biocompatible [21–23]. YMM-1 is a commercially available preparation of 90Y3 + physically linked to silica particles having a diameter of approximately 2 mm. One established application of the YMM-1 is the treatment of synovial hypertrophy in the knee joints [4]. The radioactive dose delivered to non-target organs as a result of YMM-1 leaking unintentionally from the formulation can be estimated as follows. Assume that the beta emitter is uniformly distributed throughout a gel

c 2006 Lippincott Williams & Wilkins 0143-3636

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or tissue that is larger than the beta particle range. The maximum absorbed dose to that gel or tissue is simply the product of the mean beta energy and the time integral of disintegrations per mass unit gel or tissue. If there is no clearance of the radionuclide from the gel or tissue, the absorbed dose can be easily calculated [24]. Such calculations are equivalent to the more generalized MIRD scheme that would also take into account dose contributions from photons and radionuclide residence times for each organ [25]. In this paper, in-vivo experiments with this formulation are reported. The local retention of the radioactivity as well as the distributions of the radioactivity to the liver, spleen, lungs and kidneys after injection at three different sites are reported, and the future prospects of the formulation are discussed.

Materials and methods Animals

The experiments were approved by the local ethics committee. The experiments were performed in six female outbred ‘Norsvin’ pigs supplied from Løken ga˚rd, Hærland, Norway, weighing 32–36 kg. The pigs were housed in separate steel cages (2.4 m2), in a 45 m2 room with regulated temperature and relative humidity (20 ± 11C, 55 ± 10%), and 18 air changes per hour. Artificial lighting was 100 lx, 12/12 h light and dark periods, 1 h dusk and dawn. The pigs were fed standard pig feed (FORMAT) supplied by Felleskjøpet, Norway. Drinking water was untreated mains water ad libitum. The animals were given atropine, azaperon and ketamine i.v. as premedication 1 h before the surgical procedure. They were asleep during transportation to the surgery. Anaesthesia was induced by using pentobarbital (5–15 mg
kg – 1) i.v. as required. Respiration, ECG, O2 saturation and circulation were monitored during anaesthesia. An infusion (1 ml
h – 1) of morphine 10 mg
ml – 1 was given as analgesic during the surgery. While anaesthetized, the animals were killed with 80 mmol KCl and 1 g pentobarbital i.v. Materials

The sodium alginate was Protanal SF 120 (FMC BioPolymer, Drammen, Norway), FG = 0.694, MW 237 000. A 5% solution of alginate was prefiltered using a 0.7 mm filter disc (GF/F; Whatman, Maidstone, UK) and aseptically filtered through a 0.22 mm sterile polycarbonate filter (MFS, Japan) in a Microflow LAF-bench with horizontal air-flow (Bioquell PLC, Hampshire, UK) into sterile sealable flasks. The concentration of alginate in the solution was approximately 2.3% as determined by rotational viscosimetry (Bohlin VOR, Lund, Sweden). The 90Y colloid suspension was YMM-1 (CIS bio international, Gif-sur-Yvette Cedex, France). The phos-

pholipids DPPC and DMPC were supplied by Lipoid, Ludwigshafen, Germany; sodium chloride by NMD, Oslo, Norway; and calcium chloride, chloroform and methanol by Merck, Darmstadt, Germany. Production of liposomes

All glassware used was heat sterilized before use. Calcium chloride and sodium chloride solutions were aseptically filtered through a 0.22 mm sterile polycarbonate filter (MFS) in a Microflow LAF-bench with horizontal air-flow (Bioquell PLC, Hampshire, UK) into sterile sealable flasks and autoclaved (Astell Sterilizer AVX 030, Astell Scientific, Kent, UK) before use. Liposomes were prepared by the film method [26]. All work involving the exposure of the product to air was performed aseptically in a LAF-bench with horizontal air-flow. The phospholipids (90 mol% DPPC, 10 mol% DMPC) were dissolved in chloroform/methanol (9:1). A thin lipid film was prepared in a 500 ml round flask as the solvent was evaporated for 10 min at 100 mbar and 501C using a rotavapor (Vacuubrand PC 511, Wertheim, Germany). The lipid film obtained was hydrated with 133 mM CaCl2 at 501C for 2 h with occasional stirring. The liposomes were subjected to three freeze–thaw cycles employing liquid nitrogen and a water bath (501C) followed by subsequent extrusion (Lipex extruder, Biomembranes Inc., Vancouver, Canada) through 2 mm and 800 nm polycarbonate membranes (Nucleopore, Costar Corp., Cambridge, USA). The liposomes were centrifuged for 10 min in an IEC Centra MP4 centrifuge (International Equipment, Needham Heights, Massachusetts, USA) (3000 rpm, rotor 224, maximum relative centrifugation force 1300  g). The temperature in the centrifuge did not exceed 301C at any time. The supernatant was removed and replaced with 200 mM NaCl. The concentration of calcium in the liposome suspension was determined by titration of the suspension with 2 mM EDTA in the presence of Calcon colour indicator. Centrifugation and supernatant replacement was repeated until calcium concentration in the supernatant was sufficiently low ( < 5 mM). Thus, in the external phase of the final liposome suspension, the calcium concentration was below 5 mM. A small amount of the liposome suspension was isolated for analysis of calcium content and lipid concentration. The encapsulated calcium was liberated from the liposomal interior by heating the suspension to 371C. The calcium concentration in the heated liposome suspension was determined to be 28 mM. The concentration of phospholipids in the liposome suspension was 20.3 mM as determined by the phosphate analysis described by Rouser et al. [27]. Preparation of the formulation

YMM-1 (0.5 ml) was inserted through the rubber stopper of the sealed flask of sterile alginate solution. This mixture was used as stock solution for all the six

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90

experiments. All the experiments were performed within 24 h. The radioactive concentration of the mixture was approximately 10 MBq
ml – 1 of 90Y at the time of injection. Shortly before injection of the formulation, the liposomes were mixed with the alginate/YMM-1 mixture. Liposome suspension (1 ml) and alginate/YMM-1 (1 ml) were inserted through the rubber stopper of a sealed, sterile flask containing a stirring magnet and aseptically mixed using a magnet stirrer.

Y particles in alginate gel Holte et al. 187

slices of the muscle surrounding the site of injection using a scalpel. Each slice was further divided into pieces of approximately 1 g and analysed. The longitudinal distribution of yttrium in the bone marrow compartment following intrafemural injection was investigated by preparing 5 mm thick slices of the femur while frozen, using a hacksaw. The bone marrow was extracted from each slice and analysed. Detection of radioactivity in tissue samples

Injection of the formulation into pigs

Three different experiments were performed to evaluate the usefulness of the preparation. Biodistribution of 90Y and animal behaviour was observed following intramuscular (pigs 1 and 2), intrafemural (pigs 3 and 4) or intravenous injection (pigs 5 and 6). Intramuscular and intrafemural injections were performed to determine the behaviours of the formulation in tissue experiencing various levels of perfusion. One possible application of the 90Y gel would be to inject it into the bone marrow compartment adjacent to metastases from mammary or prostate cancer, for example. We therefore investigated the behaviour of the formulation in the bone marrow compartment of the femur. Treatment of an external bone surface could involve an intramuscular injection between the bone and a muscle. The behaviour of the formulation was therefore investigated after an intramuscular injection. Intravenous injection was performed to examine the consequences of an inadvertent injection into the circulation. Approximately 1 ml of the preparation (corresponding to 5 MBq of 90Y) was injected in each experiment. Physiological saline was injected through the needle before removal from the site of injection. Two parallels of each experiment were performed. Intramuscular injections were performed into musculus quadriceps femoris. Before intrafemural injection, an awl was used to bore a small hole centrally in the femur, through which the formulation was injected. Proper positioning of the needle within the femur was verified by X-ray fluoroscopy before injection. Intravenous injection was performed into the great saphenous vein. After intramuscular and intrafemural injection, the animals lived 4 days before they were sedated and killed, and autopsy was performed immediately. No animal suffered unexpected pain leading to pre-scheduled termination of the experiment. In the cases of intravenous injection, the animals were killed 1 h after the injection while still sedated and anaesthetized.

For quantification of 90Y in tissue samples, measurement of Bremsstrahlung radiation in a scintillation well counter was preferred due to the cumbersome sample preparation required for the more precise liquid scintillation counting. Tissue samples were weighed and filled in 20 ml plastic counting vials with screw caps. Water was added to the samples to adjust sample volumes and to ensure as far as possible equal conditions for Bremsstrahlung generation throughout the sample volume. Samples were analysed using a 1480 Wallac Wizard 3’’ automatic gamma counter (PerkinElmer). The photon counts from 90Y in each sample were multiplied with organ weight to estimate the total amount of 90Y in the actual organ. The remainders of the six preparations were weighed and treated in the same matter as the tissue samples, and the Bremsstrahlung was determined to estimate the total amounts of 90Y injected in each of the six pigs. The estimated total amounts of 90Y in the various organs were compared with the estimated amounts of 90Y injected in each pig. Microscopic analysis of lung tissue samples

Lung samples from pigs given the formulation by intravenous injection (pigs 5 and 6) were investigated by microscopy to detect possible gel deposition in the lung capillaries. The specimens were fixed in formalin. All specimens were embedded in paraffin wax using a Tissue-Tex VIP Vacuum Infiltration Processor (Sakura Finetex USA Inc., Torrance, California, USA) and a paraffin dispenser. The paraffin wax blocks were cut at 3 mm, dried at 601C for 45 min and de-waxed in xylene. Subsequently, sections were rehydrated through graded ethanol in water, and washed in water. The slides were counter-stained in haematoxylin & eosin and rinsed, followed by dehydration and mounting in Diatex mounting media.

Preparation of tissue samples

From each animal, tissue samples were collected from the site of injection, in addition to liver, spleen, kidney, lung and blood. Urine or faeces was not collected. The spatial distribution of yttrium in the muscle following intramuscular injection was investigated by preparing 6 mm thick

Results The distribution of 90Y detected in various tissues following intramuscular injection is displayed in Table 1. In both animals, a large proportion of the recovered 90Y was found at the site of injection. More than 80% of the

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radioactivity detected in the muscle was found within about 9 cm3 of tissue.

Fig. 1

40

The distribution of 90Y detected in various organs following intravenous injection is displayed in Table 3. The calculated amount of 90Y detected in the lungs of pig number 6 is obviously overestimated. This could be due

30

%

The distribution of 90Y detected in various tissues following intrafemural injection is displayed in Table 2. Less than 40% of the injected 90Y was recovered from pig number 3. This could be partially explained by difficulties experienced during the injection procedure. The needle was blocked by bone debris, resulting in the leakage of some of the preparation from the syringe. Twenty-eight to 40 percent of the recovered 90Y was found at the site of injection. The intrafemural distribution of 90Y is presented in Fig. 1. 90Y was detected throughout the bone marrow, which constitutes approximately the middle third of the bone length. The peripheral third of the bone is composed of highly trabecular bone tissue, not allowing for significant liquid flow. No 90Y was detected in these peripheral parts of the bone. Most of the extrafemural radioactivity was found in the lungs, but significant amounts were also detected in the liver and blood.

20

10

0

−3

−2

−1

0

1 cm

2

3

4

5

Longitudinal relative distribution of 90Y within the bone marrow compartment. The grey and black bars represent numbers obtained for pigs 3 and 4, respectively. The zero along the abscissa indicates the site of injection.

Fig. 2

90 Y detected in various tissues 4 days after intramuscular injection of the gel formulation

Table 1 Organ

Percent of recovered

Y

Percent of injected 90Y

Pig 1

Pig 2

Pig 1

Pig 2

0.3 0.0 0.0 0.3

3.6 0.3 0.1 0.7

0.3 0.03 0.04 0.2

0.6 0.05 0.02 0.1

0.2 0.0 99.1

0.7 5.4 89.2

0.1 0.00 79.3

0.1 0.9 14.6

80.0

16.4

Liver Spleen Kidney Central lung, close to main bronchi Peripheral lung Blood Muscle (site of injection) Total

90

100

100

Temporary urticaria experienced by the pigs shortly after intravenous injection of the gel formulation. 90 Table 2 Y detected in various tissues 4 days after intrafemural injection of the gel formulation

Organ

Liver Spleen Kidney Central lung, close to main bronchi Peripheral lung Blood Bone marrow compartment (site of injection) Total

Percent of recovered

90

Y

Percent of injected

90

Pig 3

Pig 4

Pig 3

Pig 4

17.5 2.5 0.4 25.1

6.0 0.5 0.1 43.0

6.8 1.0 0.2 9.7

5.7 0.5 0.1 40.5

10.6 15.3 28.6

5.0 5.1 40.2

4.1 5.9 11.1

4.7 4.8 37.9

38.6

94.1

100

100

Y

to lung samples not being representative of the entire lungs. A large proportion ( > 80%) of the 90Y detected was found in the lungs. Both pigs experienced an anaphylactic reaction to the injection, with temporary urticaria all over the body (Fig. 2). Monitoring pulse and oxygen saturation was not possible during the event due to a markedly reduced peripheral blood pressure. An ST elevation was observed on the ECG. This event occurred 2 or 3 min after the injection, respectively. Twenty minutes after the injection, the pigs did not show any sign of the inflammatory response. Microscopic analysis of the lung tissue did not reveal any unusual features.

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90

90 Table 3 Y detected in various tissues 1 h after intravenous injection of the gel formulation

Organ

Liver Spleen Kidney Central lung, close to main bronchi Peripheral lung Blood Total

Percent of recovered

90

Y

Percent of injected

90

Pig 5

Pig 6

Pig 5

Pig 6

8.6 1.9 0.2 66.9

0.7 0.3 0.0 90.3

7.2 1.6 0.2 55.9

2.2 1.0 0.1 302.0

16.2 6.3

2.5 6.2

13.5 5.2

8.4 20.6

83.6

334.4

100

100

Y

Discussion The significant variation in total recovery of radioactivity is surprising, and may reflect an inhomogeneous distribution of radioactive particles within the tissues investigated. However, the distributions of the recovered radioactivity reveal important knowledge about the formulation and its behaviour in various tissues. Femoral muscle perfusion is reported to range from 3 to 17 ml
min – 1 per 100 g at rest [28–31]. In contrast, perfusion to the femoral bone is reported to range from 11 to 40 ml
min – 1 per 100 g [30–32]. Muscular perfusion is highly dependent on exercise and medicaments, and perfusion is different in various parts of the femur, but it seems that the muscular perfusion is lower than that of the femur at rest. Previous experiments injecting a viscous contrast agent into the bone marrow demonstrated a rapid removal of the contrast agent from the femoral compartment (data not shown). The different behaviour of the formulation in muscle and bone marrow probably results from the greater perfusion experienced by the formulation in the bone marrow. Østensen et al. [33] also reported the accumulation of intravenously injected particulate material in the lungs of pigs, leading to acute pulmonary hypertension and systemic hypotension. This was not observed following intravenous injection in rabbits, monkeys or man, and was presumed to be a species specific effect. The effect is probably associated with an abundance of macrophages in the lung capillaries of the pig. In the present study, the anaphylactic reaction observed could be triggered by circulating liposomes, YMM-1 particles or calcium alginate gel. The possible deposition of gel in the lung capillaries should be visible in the microscope, and if such deposition is harmful, a permanent clinical response should be expected, rather than the temporary systemic reaction observed. When injected intravenously, the formulation is probably quickly diluted, thus preventing any gel formation. Thus, the anaphylactic reaction observed following the intravenous injection is probably

Y particles in alginate gel Holte et al. 189

due to circulating YMM-1 particles or liposomes. The 90Y detected in the lungs was probably due to circulating YMM-1 particles retained by pulmonal macrophages, rather than gel encapsulated material. Intramuscular or intrafemural injection did not lead to any inflammatory response in the pigs. Possibly, the slower leakage of particles from the muscle or bone marrow does not trigger the inflammatory response. This allows the particles to pass through the lung capillaries and to be distributed to the liver. The complete retention of the radionuclide at the site of injection was not obtained following intramuscular or intrafemural injection. For therapeutic purposes, however, a certain leakage of radioactivity from the gel formulation might be acceptable. A realistic estimation of absorbed doses in non-target organs may be obtained, assuming that most circulating YMM-1 particles will be distributed to the liver: the maximum applied volume of the gel formulation is probably about 200 ml. A suitable therapeutic dose would aim at delivering 100 Gy to the gel. The dose delivered at the gel/tissue boundary will be lower, and depending on the type of tissue (e.g., bone or soft tissue). This would require a total activity of 2 GBq of 90Y in the gel. If 10% of this activity ends up homogenously distributed in a liver weighing 1.9 kg, the resulting absorbed dose would be 0.55 Gy, which is an acceptable liver dose. In order to retain the particulate material, the gel formulation must set quickly to avoid being diluted and drained by circulating fluids. Thus, the present formulation appears to be suitable for the application to organs and tissues having low perfusion, like muscle or adipose tissue. However, the formulation has to be improved to make it useful in environments with higher perfusion. Such improvements should aim at making the final gel more sustainable, or accelerating the gelling process. The former could be achieved by increasing the calcium and alginate concentrations. The latter could be achieved by changing the lipid composition of the liposomes, resulting in a more suitable phase transition temperature of the liposomes. Alternatively, the formulation could be pre-heated shortly before injection, reducing the time required to heat the formulation to the phase transition temperature of the liposomes after injection.

Conclusion A new alginate gel formulation suitable for localized radiotherapy in organs and tissues having low perfusion has been developed. The formulation is an injectable liquid, and sets quickly to a gel after injection. Radioactive particles are thus encapsulated in the gel matrix and retained at the injection site.

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Original article

Reproducibility of technetium-99m mercaptoacetyltriglycine clearance in patients with impaired renal function Tanju Yusuf Erdila, Fuat Dedea, Serhan Tuglularb, Feyza Sena, Tunc Onesa, Zeynep Farsakoglua, Sabahat Inanira, Cetin Ozenerb, Emel Akoglub and Turgut Turoglua Background The aim of this study was to determine the reproducibility of technetium-99m mercaptoacetyltriglycine (99mTc-MAG3) clearance in patients with a 99mTc-MAG3 clearance below 100 ml/min/1.73 m2. Methods Two separate multi-sample clearance studies were performed in 16 patients at a 1 week interval. The clearances were calculated according to the open twocompartment model of Sapirstein et al., accepting the 90, 120 and 180 min samples as the last points of the biexponential curve. The clearance measurements were also performed according to the single-sample methods of Russell et al. and Bubeck using the fitted value at 44 min.

Conclusion Single-sample methods give very poor reproducibility and accuracy and should not be used in patients with poor renal function. The reproducibility of 99mTc-MAG3 clearance using the multi-sample method (90 min) in patients with impaired renal function is 12.6%, which is similar to that in patients with good renal function and that obtained with other tubular agents. Whether this level of reproducibility is satisfactory for documenting serial changes in an individual patient with a 99m Tc-MAG3 clearance below 100 ml/min/1.73 m2 depends on the expectation of the clinician. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:191–196
Nuclear Medicine Communications 2006, 27:191–196

Results There was no significant difference between the two clearance measurements for all five samples (P > 0.05). There was a systematic increase in clearance measurements of 8.0 ± 2.7% from the 180 to 120 min samples and 4.8 ± 2.0% from the 120 to 90 min samples. Both single-sample methods (Bubeck and Russell et al.) gave more divergent results than multi-sample methods. The mean and standard deviation (%) of the normalized differences between two successive tests were – 3.9 ± 12.6, – 2.4 ± 13.1, – 1.9 ± 14.9, – 4.1 ± 53.5 and – 13 ± 82.1 for 90, 120 and 180 min samples and the Russell et al. and Bubeck methods, respectively.

Introduction Technetium-99m mercaptoacetyltriglycine (99mTcMAG3) has become the radiopharmaceutical of choice for renal scintigraphy in the evaluation of nephrourological diseases because of the favourable dosimetry compared with iodine-131-orthoiodohippurate ([131I]OIH) and the superior imaging quality compared with both [131I]OIH and any glomerular tracer [1–3]. The utility of 99mTc-MAG3 clearance has been extensively investigated in several studies [4–6], and it has been shown that the clearance of 99mTc-MAG3 is closely correlated with the clearance of [131I]OIH [6–9]. However, it has generally been reported that, although 99m Tc-MAG3 has very good accuracy [7–12], the results in terms of reproducibility are still controversial [13–19]. The reproducibility of the study is especially important in patients with renal failure in order to determine whether changes in renal function in serial measurements are

Keywords: clearance, impaired renal function, reproducibility,

99m

Tc-MAG3

a Department of Nuclear Medicine, Marmara University Hospital and bSection of Nephrology, University Hospital, Istanbul, Turkey.

Correspondence to Tanju Yusuf Erdil MD, Kozyatagi mah, Ziraat Bank, Lojmanlari, A Blok D:22, Kozyatagi, Istanbul 34742, Turkey. Tel: + 90 216 327 69 57; fax: + 90 216 327 69 56; e-mail: [email protected]

Received 19 August 2005 Accepted 16 November 2005

related to progression, regression, resolution of the disease or poor reproducibility of the test. The aim of this study was to determine the reproducibility of 99m Tc-MAG3 clearance in patients with chronic renal failure.

Materials and methods Patients

Sixteen patients (eight females, eight males; age range, 42–68 years; mean, 54 years) with a history of stable chronic renal failure (plasma creatinine level of 3.4 ± 0.34 mg/dl) were included in the study. The patients were not on dialysis during the study. There was no intervention, nephrotoxic therapy or contrast medium administration between the two clearance studies. None of the patients had oedema or ascites. The serum creatinine levels were determined on the same day after both studies. Blood pressure and heart rate

c 2006 Lippincott Williams & Wilkins 0143-3636


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192 Nuclear Medicine Communications 2006, Vol 27 No 2

were examined before, during and after the study. The local ethical committee of our hospital gave permission for the study, and written informed consent was obtained from each patient.

solution. The syringes used for standard aliquots were also weighed and counted before and after injection to calculate the net amount of the tracer. After shaking the flask, 1 ml samples were pipetted into preweighed and labelled counting tubes.

Patient preparation

After a light breakfast early in the morning, all patients drank 500–750 ml of water 30 min before the examinations (between 09.00 and 10.00 h). For tracer injection and blood sampling, venous access was provided into both arms at least 5 min before injection. Hydration was continued by regular fluid intake during the study. Before starting the study, the bladder was emptied. The patients remained in the department throughout the study. The whole procedure was repeated 1 week later under the same conditions.

Data analysis

For the clearance measurements, the open two-compartment model described by Sapirstein et al. [20] was used. The data were plotted against time and the rate constants (a and b) and intercepts (A and B) for slow and fast components were calculated using biexponential curve fitting analysis with 90, 120 and 180 min samples (as the last point of the slow component). Clearance measurements were also performed with the methods of Russell et al. [21] and Bubeck [22] using the fitted value at 44 min. All results were corrected to a body surface area of 1.73 m2.

Radiopharmaceutical

In a maximum volume of 2 ml, 185 MBq of 99mTc-MAG3 (Mallinckrodt Inc., Petten, The Netherlands), which was freshly prepared in accordance with the recommendations of the manufacturer, was injected as a bolus via a three-way stopcock connected to a butterfly line, and the empty syringe was flushed with 10 ml of saline. Radiopharmaceutical purity was determined sporadically at the time of injection as recommended by the manufacturer, and the labelling efficiency was found to be greater than 96% in each case. Gamma camera imaging was also carried out in the supine position simultaneously with the clearance measurement for each patient. At the end of the acquisition, the injection site was imaged for poor injections, and infiltration exceeding 0.5% was not found at any injection site. Clearance measurement

Following the injection, 1 ml washings from the injection syringe and butterfly infusion set were obtained and transferred into preweighed and labelled counting tubes to determine the residual activity in the syringe. Subsequently, 10 blood samples (5, 12, 17, 27, 37, 44, 60, 90, 120 and 180 min post-injection) of 5 ml were drawn into heparinized tubes through an indwelling catheter with a three-way stopcock from the contralateral arm; the catheter was rinsed with saline after each blood sampling. At each time point, before actual sampling, 5 ml of blood was drawn into a heparinized syringe, rinsed with saline and then reinjected into the patient after actual sampling. After centrifugation for 10 min at 1500 g, 1 ml samples were pipetted into preweighed and labelled counting tubes. All counting tubes were reweighed and each sample was counted twice for 1 min at the 99mTc peak in a well-type gamma counter. The relative counting error was below 1%. The injected dose was estimated from the weight difference of the syringes before and after injection. The standard was prepared at the time of injection by diluting an aliquot equal to the injection dose into a 1000 ml volumetric flask filled with 0.09% NaCl

Statistical analysis

The reproducibility was calculated according to the method described by Bland and Altman [23]. For each patient, the difference between the two clearance measurements was expressed as a percentage of the mean value of the two studies. The mean of these normalized differences represents the systematic bias, whereas the standard deviation (SD) represents the reproducibility of the technique. For the single-sample methods of Russell et al. [21] and Bubeck [22], accepting the 90 min measurement as the reference method, the reproducibility (corrected) measured using
p 2 1was2 also the formula SC ¼ SD þ =4S1 þ 1=4S22 (where SC is the corrected SD of the differences, SD is the SD of the differences between the means of each method and S1 and S2 are the SDs of the differences between repeated measurements for each method separately) of Bland and Altman [23] for repeated measurements by each of two methods on the same subjects. Statistical analysis was performed using the paired t-test, and the results were considered to be significant when the P value was below 0.05. The correlation between the mean 99m Tc-MAG3 clearance and the normalized difference was performed using linear regression analysis.

Results The results of the clearance measurements and statistical analysis are summarized in Table 1. There was no significant difference between the two creatinine values (P = 0.13). There was a systematic increase in clearance measurements of 8.0 ± 2.7% (P = 1.56E-09) from 180 to 120 min samples and of 4.8 ± 2.0% (P = 1.23E-09) from 120 to 90 min samples. The smallest SD between singlesample (methods of both Bubeck [22] and Russell et al. [21]) and multi-sample clearances was obtained using the 90 min samples for the two clearance measurements (Table 2). However, the methods of both Bubeck and Russell et al. (especially Bubeck) gave more divergent

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13.6 59.3 82.6 – 18.7 18.6 40.2 – 14.8 – 34.1 – 58.2 – 19.7 – 32.2 – 34.3 14.3 25.5 30.2 5.1 17.8 20.0 13.5 73.3 81.0 13.6 – 40.9 – 69.5 – 8.3 20.7 22.2 – 5.3 – 45.9 – 53.4 – 16.7 – 80.8 – 186.0 16.7 30.7 38.9 – 14.1 – 100.8 – 146.1 18.8 52.3 75.0 – 16.1 39.3 46.6 – 12.6 – 67.9 – 96.8 – 1.9 – 4.1 – 13.0 14.9 53.5 82.1 13.9 – 12.3 – 18.0 – 17.8 13.4 3.5 11.9 9.9 – 11.3 – 3.4 – 17.5 14.7 – 17.1 10.6 – 7.7 – 10.8 – 2.4 13.1 15.3 – 11.1 – 19.9 – 17.9 11.3 2.1 8.7 6.1 – 13.1 – 7.7 – 16.7 11.6 – 18.8 9.1 – 8.6 – 12.7 – 3.9 12.6 47.3 11.8 – 25.1 – 30.2 23.8 17.0 74.1 – 27.7 19.8 – 41.5 – 50.6 25.7 – 86.8 43.1 35.3 – 46.8 – 0.7 44.1 34.0 7.2 – 16.5 – 25.8 19.0 14.1 63.9 – 18.0 16.9 – 33.3 – 32.2 19.3 – 63.1 30.8 28.0 – 34.7 0.6 33.5

B R 180

(%)

8.7 – 6.6 – 5.1 – 14.0 9.5 3.3 8.3 5.1 – 4.8 – 3.2 – 6.4 9.5 – 8.4 10.5 – 8.6 – 6.7 – 0.6 8.2

180 120

9.4 – 4.8 – 6.9 – 13.5 9.4 2.4 7.8 3.9 – 7.0 – 2.3 – 7.2 9.0 – 10.9 6.6 – 4.5 – 6.2 – 0.9 7.8 10.9 – 4.6 – 8.1 – 14.2 8.2 1.6 6.0 2.6 – 8.8 – 5.4 – 7.5 7.4 – 12.6 6.0 – 5.3 – 7.7 – 2.0 8.0 33.6 23.4 55.7 103.1 66.8 76.3 54.4 53.7 79.1 98.4 52.5 53.1 102.7 35.9 58.1 71.6 63.7 23.9 40.3 35.0 56.7 92.9 65.1 71.9 55.2 53.1 72.8 89.3 56.0 53.3 94.1 43.5 57.3 68.5 62.8 17.9 59.2 38.7 36.9 77.9 61.8 63.3 57.2 34.6 59.6 62.5 41.3 51.9 63.9 50.7 57.9 56.8 54.6 11.7 63.2 41.0 41.6 82.7 65.9 68.2 61.8 37.8 66.1 67.6 44.7 56.3 69.2 59.2 61.1 60.8 59.2 12.2 65.7 43.7 44.8 86.6 69.0 72.0 65.6 41.8 72.0 72.3 48.5 59.7 73.3 62.9 63.7 64.5 62.9 12.5 80.9 35.1 30.6 72.9 90.6 93.3 128.4 26.0 98.9 56.9 1.9 78.8 16.0 79.1 93.4 24.9 63.0 36.3 74.3 42.2 40.2 67.1 84.1 85.9 119.1 35.1 89.7 56.0 23.8 72.6 31.0 74.3 85.4 33.8 63.4 27.0

(ml/min/1.73 m )

R

B

90

120

180

2

(ml/min/1.73 m )

R

B

90

Difference

2

(ml/min/1.73 m )

R

B

90

120

Difference

Tc-MAG3 clearance Erdil et al. 193

67.9 32.1 31.8 63.9 71.4 66.6 65.5 39.6 54.8 59.2 34.9 61.4 55.5 61.3 49.3 50.1 54.1 13.2 72.7 36.2 34.8 69.2 75.4 70.6 69.7 41.8 59.0 65.3 37.5 65.2 58.3 65.8 56.6 54.6 58.3 13.7 Cr, creatinine; F, female; M, male.

M M F M M F F M M F M F F M F M 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 Mean SD

64 49 68 63 64 54 48 52 60 45 51 53 42 48 53 54 54.3 7.5

3.1 3.8 3.9 3.2 3.1 3.1 3.3 3.9 3.5 3.2 4.1 3.3 3.1 3.3 3.4 3.4 3.4 0.3

3.1 3.6 3.7 3.3 3.2 3 3.1 3.7 3.6 3.2 3.9 3.4 3.1 3.2 3.5 3.3 3.4 0.3

76.6 39.1 36.7 72.4 77.2 73.6 71.6 44.4 63.1 66.9 41.0 67.1 60.7 68.8 58.4 56.8 60.9 13.7

180 120 90 (mg/dl)

Cr 1st Cr 2nd Age

(years) Sex Patient

In our study, the reproducibility of 99mTc-MAG3 clearance was found to be 12.6% (8.0 ml/min/1.73 m2). This finding is in agreement with that of Kotzerke et al. [14],

2nd study

Accurate measurement of renal function is very important in clinical situations in which it is necessary to detect the presence and to determine the severity of renal function impairment. However, in serial measurements of renal function, especially in patients with renal failure, good reproducibility of the test is vitally important for monitoring renal function and to ascertain whether changes in renal function are related to disease or to the limited reproducibility of the study. Inconsistent results of the reproducibility of 99mTc-MAG3 clearance have been reported, ranging from 6.3 to 40.4% depending on the level of renal function, the clearance measurement technique applied and the time interval between the two studies [13–19,24]. Furthermore, almost the same level of reproducibility with a relative SD of around 12% was accepted as adequate for monitoring renal function by Werner et al. [15], Russell and Dubovsky [16] and Kanazawa et al. [17], whereas it was found to be inadequate by Kotzerke et al. [14], Moller and Widding [18] and Sadeleer [19]. However, most of the patients in these studies had normal renal function. Only the results of three patients with a 99mTc-MAG3 clearance lower than 100 ml/min/1.73 m2 were reported and showed very high SDs (56, 68 and 100%) [14]. The assumption that the reproducibility is worse in patients with poor renal function is based mainly on these three patients, and has not been substantiated in a large group of patients [14,15]. We therefore aimed to determine the reproducibility of 99mTc-MAG3 clearance in patients with stable chronic renal failure with a 99mTc-MAG3 clearance lower than 100 ml/min/1.73 m2.

99m

2

Discussion

1st study

results (from – 100.8% to 73.3% for Russell et al. and from – 186% to 82.6% for Bubeck) compared with the multisample method. There was no significant difference between the two consecutive clearance measurements for all five samples (P > 0.05). The lowest mean and SD of the normalized differences were found with the 180 min ( – 1.9%; – 0.6 ml/min/1.73 m2) and 90 min (12.6%; 8.0 ml/ min/1.73 m2) samples, respectively. The corrected SDs of the differences of the single-sample methods, accepting the 90 min measurement as the reference method, were 19.2 ml/min/1.73 m2 for Russell et al. and 25.8 ml/min/ 1.73 m2 for Bubeck, which were significantly lower than the non-corrected SDs of the differences (33.5 ml/min/ 1.73 m2, 53.5% for Russell et al. and 44.1 ml/min/1.73 m2, 82.1% for Bubeck). No correlation was found between the mean 99mTc-MAG3 clearance and the normalized difference between the two measurements (r = 0.30). There was no significant change in blood pressure and heart rate during the study.

99m Table 1 Technetium-99m mercaptoacetyltriglycine ( Tc-MAG3) clearance values of two separate multi- and single-sample (R, Russell et al. [21]; B, Bubeck [22]) measurements, and the systematic bias (mean difference) and reproducibility (standard deviation, SD) of the study expressed in ml/min/1.73 m2 and as a percentage of the mean value of the two studies

Reproducibility of

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Table 2

Mean and standard deviation (SD) of the normalized differences between single- and multi-sample clearance measurements Russell et al. [21](44 min)

90 min 120 min 180 min

Bubeck [22](44 min)

1st study (%)

2nd study (%)

1st study (%)

2nd study (%)

– 2.17 ± 32.11 – 6.91 ± 33.10 – 15.71 ± 36.58

– 0.25 ± 20.74 – 6.86 ± 23.08 – 16.01 ± 25.56

1.85 ± 48.05 – 2.46 ± 49.81 – 10.93 ± 54.25

– 0.33 ± 28.15 – 6.94 ± 30.65 – 16.07 ± 33.51

who found an SD of normalized clearance differences of 16% for repeated measurements of 99mTc-MAG3 clearance at a 1 week interval after exclusion of three patients with a clearance lower than 100 ml/min/1.73 m2. However, the SD increased to 40.4% when these three patients were also included in the calculation [14]. No correlation was found between the level of renal function and the mean normalized difference in the study of Kotzerke et al. [14], which is in agreement with our study, indicating that the reproducibility of the clearance measurement is independent of the level of renal function regardless of whether it is poor or good. This increase might be related to the single-sample method (Bubeck) applied for the calculation of clearance [14]. We also found very poor reproducibility using the single-sample methods of Russell et al. [21] (53.5%) and Bubeck [22] (82.1%). The corrected SDs of the differences of the methods of Russell et al. and Bubeck decreased from 33.5 to 19.2 ml/ min/1.73 m2 and from 44.1 to 25.8 ml/min/1.73 m2, respectively, accepting the 90 min measurement as the reference method. However, these values are still high and not acceptable for routine clinical use. Furthermore, because of the need for the multi-sample method for the measurement of the corrected SD of the differences, it is not suitable to use single-sample methods in clinical practice. Russell et al. [7] demonstrated that singlesample methods tend to be inaccurate and show increased random error at low clearance values. Rehling and Nielsen [25] found a large difference between the clearance obtained by a single-sample method and the reference multi-sample method in patients with a plasma 99m Tc-MAG3 clearance of less than 75 ml/min, and concluded that single-sample methods should not be used in these patients. Using the single-sample methods of Russell et al. and Bubeck, in agreement with these studies, we also found very large differences compared with the multi-sample method (poor accuracy) and between two consecutive clearance measurements (poor reproducibility). Urine collection or late blood sampling is recommended in patients with poor renal function [3,5,7,26]. However, perfect urine collection is far from easy to achieve and, even though urinary clearances are accurate, they are not reproducible [3,4,27]. With urine collection, the reproducibility was poor with an SD of 8.9% for normal volunteers even in expert hands [28,29]. For routine clinical use, urine collection seems to be necessary only in patients

with extremely low clearance values [26,30]. In addition, none of our patients had oedema or ascites, for which urine collection is necessary [3,4]. Therefore, we did not use a urinary clearance method. Instead, we employed late blood sampling (3 h), which has been recommended in patients with poor renal function [28]. It has been shown that the ratio between the plasma clearance of 99m Tc-MAG3 and [131I]OIH is constant irrespective of the glomerular filtration rate (GFR), and that the extrarenal clearance of 99mTc-MAG3 is smaller than that of 51Cr-ethylenediaminetetraacetate (51Cr-EDTA), and it has been concluded that the plasma clearance of 99mTcMAG3 can be used as a measure of renal tubular function in all patients regardless of the GFR level [12]. Russell [28] reported that sampling must begin by 5 min and continue for at least 90 min, and that at least six samples must be obtained for single-injection multisample plasma clearance methods for tubular agents. In our study, as recommended by Russell [28], we took 10 samples beginning at 5 min and continuing for 3 h, and calculated the clearance measurement according to 90, 120 and 180 min samples (as the last point of sampling). We observed a progressive and systematic decrease in the clearance value depending on the choice of the last point of the curve, as in the study of Piepsz et al. [13]. However, it is not clear which sampling time gives the most accurate clearance measurement. Protein binding and even very small amounts of radiochemical impurities (1– 3%) in the preparation influence considerably the late part of the plasma curve and introduce an additional significant exponential, thus lowering the clearance value significantly depending on the duration of sampling [13]. However, Russell [31] reported that a three-compartment model for 99mTc-MAG3 clearance leads to lower and probably more accurate clearance values than the conventional two-compartment model, assuming that the radiochemical impurities in commercial preparations of 99mTc-MAG3 do not cause appreciable errors in clearance measurements. In our study, any decrease in the sampling time interval increased the mean of the normalized differences ( – 3.9, – 2.4 and – 1.9% for 90, 120 and 180 min, respectively), but improved the reproducibility (12.6, 13.1 and 14.9% for 90, 120 and 180 min, respectively) of the clearance measurement, even though the results were not significantly different from each other. In our study, only patients with stable chronic renal failure were included and no significant

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Reproducibility of

change in renal function was determined throughout the study. However, we cannot exclude that, for some reason, a real change occurred between the two tests and therefore the true reproducibility could be better than estimated, as demonstrated in the study of Russell and Dubovsky [16].

99m

Tc-MAG3 clearance Erdil et al. 195

Our results were not very different from those of other authors, who estimated the reproducibility of 99mTcMAG3 single- or two-sample clearance with an SD of around 12% [14–17]. Kotzerke et al. [14], Werner et al. [15] and Russell and Dubovsky [16] performed the study retrospectively in different patient populations using a single-sample method, and calculated the reproducibility as 11.7, 11.7 and 12%, respectively. Kanazawa et al. [17] carried out their study prospectively in healthy volunteers with the two-sample method at a 1 month interval, and the reproducibility was found to be 11.1%, which is in agreement with that of other studies [14–16]. In the same study, it was shown that the reproducibility of [131I]OIH (13.6%) was very similar to that of 99m Tc-MAG3 [17]. Russell and Dubovsky [16] also found no significant difference between the reproducibility of 99m Tc-MAG3 and [131I]OIH. In normal volunteers at a 1 week interval, the other technetium-labelled tubular agent, 99mTc-ethylenedicysteine, also gave almost the same level of reproducibility (12.7%) as both 99mTcMAG3 and [131I]OIH [24]. Moller and Widding [18] performed the gamma camera method for 99mTc-MAG3 clearance estimation, and the reproducibility was found to be 12.4%, consistent with that of single- and twosample methods.

[16], this large systematic bias was not random; rather, it was systematic, suggesting an explanation not in the measurement technique but in the study design. This discrepancy has been explained by psychogenic stress: the volunteers may have been more apprehensive for the first measurement than for the second, leading to systematically lower results for the first measurement [13,16]. Monitoring the blood pressure during the study and performing venepuncture several minutes before tracer injection have been advised to prevent psychogenic stress [16]. In our routine practice, venous access is provided at least 5 min before the injection, and there was no case of hypotension during the study. Furthermore, the patients in our study were very familiar with nuclear medicine and other laboratory examinations. Werner et al. [15] have proposed that variable patient hydration was one of the main reasons for the poor reproducibility of the study by Piepsz et al. [13]. Hydration with more than 10 ml/kg body weight was recommended by Werner et al. [15] and Frokiaer et al. [32]. However, the main problem in the study by Piepsz et al. [13] was the large systematic bias, not the poor reproducibility. Different degrees of hydration may cause random, but not systematic, discrepancy. Furthermore, as recommended by the Nephro-urology Committee on Renal Clearance of the Society of Nuclear Medicine, approximately 5–6 ml/kg of fluid intake before a clearance study is sufficient for good hydration [26]. Therefore, the standard oral preparation of 0.5 l of fluid intake employed in the study by Piepsz et al. [13] must have been sufficient for good hydration in patients with a body weight of up to 100 kg.

In contrast with the above-mentioned studies, Piepsz et al. [13] and Sadeleer [19] found very poor reproducibilities in healthy volunteers. Sadeleer [19] performed three successive tests at a 1 week interval, and found the SDs of the differences between two clearance measurements to be 35.8, 47.7 and 57.7 ml/min/1.73 m2 using the singlesample method of Russell et al. [21]. Piepsz et al. [13] also found a very large SD of 25% for paired measurements in 12 young healthy volunteers using a multi-sample 99mTcMAG3 clearance method at a 1 week interval. However, there was a significant difference between the two measurements of an average of 20%; the calculation of reproducibility is not possible in the presence of this kind of large systematic bias [13,15,23]. If the mean difference is significantly different from zero, the data cannot be used to assess the reproducibility, because a knowledge of the first measurement may affect the second or the process of measurement may alter the quantity [23]. Methodological (impurities contained in the commercial kit, variable protein binding, labelling efficiency) and physiological (pH of urine, sodium load, protein intake, fluctuation in tubular function and stress) factors have been proposed to explain this large systematic bias [13]. However, as stated in the study by Russell and Dubovsky

Glomerular agents show less day-to-day variation than tubular agents [3,4,13,15,18]. Although the reproducibility in one single person has no clinical meaning, Donath [33] reported that, in three children, renal plasma flow (RPF) measurements with [125I]OIH showed good reproducibility with SDs of 1.7 (five times within 12 days), 2.3 and 0.9% (once a day for 4 days) versus GFR with 51Cr-EDTA with SDs of 5.3, 5.2 and 6%, respectively. In contrast with RPF, GFR is maintained over a large range by regulation mechanisms that vary the resistances of the afferent and efferent arteries around the glomerulus [3]. RPF is therefore an input parameter to the kidney and does not necessarily reflect the renal function [3]. For this reason, in clinical practice, GFR is more familiar to most clinicians and is considered to be the more reliable index of renal function [3,4,26,34]. 51 Cr-EDTA is the radiopharmaceutical of choice for GFR [3,26]. However, it is not suitable for imaging and is not readily available in some countries. 99mTcdiethylenetriaminepentaacetate (99mTc-DTPA) can also be used for the estimation of GFR, but is not as accurate as 51Cr-EDTA because of its pharmacokinetic properties [26]. 99mTc-MAG3 has gained wide acceptance for both imaging and clearance studies and is preferred to 99mTc-

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DTPA and [131I]OIH, especially in patients with renal failure, by virtue of its higher extraction rate, superior image quality, faster clearance and favourable dosimetry [1–4,7,15,16,26]. In conclusion, the reproducibility of 99mTc-MAG3 clearance (12.6%, 90 min sample) using the multi-sample method in patients with impaired renal function is similar to that in patients with good renal function using singleor two-sample methods and that obtained with other tubular agents. However, the difference between two consecutive measurements must be higher than 25–30% in order to be recognized as a real change in renal clearance with a confidence interval of 95%. Whether this level of reproducibility is acceptable for routine clinical use to monitor renal function in patients with a 99m Tc-MAG3 clearance lower than 100 ml/min/1.73 m2 depends on the expectation of the clinician. Singlesample methods, which are the methods of choice in routine practice, give very poor reproducibility and accuracy, and should not be used in patients with poor renal function.

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Rehling M, Nielsen BV, Pedersen EB, Nielsen LE, Hansen HE, Bacher T. Renal and extrarenal clearance of 99mTc-MAG3: a comparison with 125I-OIH and 51Cr-EDTA in patients representing all levels of glomerular filtration rate. Eur J Nucl Med 1995; 22:1379–1384. Piepsz A, Tondeur M, Kinthaert J, Ham HR. Reproducibility of technetium99m mercaptoacetyltriglycine clearance. Eur J Nucl Med 1996; 23: 195–198. Kotzerke J, Glatz S, Grillenberger K, Kleinschmidt K, Reske SN. Reproducibility of a single-sample method for 99Tcm-MAG3 clearance under clinical conditions. Nucl Med Commun 1997; 18:352–357. Werner E, Blasl C, Reiners C. Reproducibility of technetium-99m-MAG3 clearance using the Bubeck method. J Nucl Med 1998; 39:1066–1069. Russell CD, Dubovsky EV. Reproducibility of single-sample clearance of 99mTc-mercaptoacetyltriglycine and 131I-orthoiodohippurate. J Nucl Med 1999; 40:1122–1124. Kanazawa T, Shimizu M, Seto H. Reproducibility of 99Tcm-MAG3 clearance in normal volunteers with the two-sample method: comparison with 131I-OIH. Nucl Med Commun 1998; 19:899–903. Moller ML, Widding A. Reproducibility of 99mTc-MAG3 compared to 99mTcDTPA renography: day to day variation in estimates of renal function. Eur J Nucl Med 1996; 23:1189. Sadeleer CD. Reassessment of the reproducibility of technetium-99m mercaptoacetyltriglycine clearance. Nucl Med Rev Cent East Eur 2005; 8:15–20. Sapirstein LA, Vidt DG, Mandel MJ, Hanusek G. Volumes of distribution and clearances of intravenously injected creatinine in the dog. Am J Physiol 1995; 181:330–336. Russell CD, Taylor A, Eshima D. Estimation of technetium-99m-MAG3 plasma clearance in adults from one or two blood samples. J Nucl Med 1989; 30:1955–1959. Bubeck B. Renal clearance determination with one blood sample: improved accuracy and universal applicability by a new calculation principle. Semin Nucl Med 1993; 23:73–86. Bland M, Altman D. Statistical methods for assessing agreement between two methods of clinical measurements. Lancet 1986; 11:307–310. Kabasakal L, Halac M, Aklan E, Ozcelik N, Uslu I. Reproducibility of technetium-99m ethylenedicysteine clearance. Eur J Nucl Med 1999; 26:900–902. Rehling M, Nielsen LE. Plasma clearance of 99mTc-MAG3: accuracy of five single-sample methods. Nucl Med Commun 2000; 21:193–197. Blaufox MD, Aurell M, Bubeck B, Fommei E, Piepsz A, Russell C, et al. Report of the radionuclides in nephrourology committee on renal clearance. J Nucl Med 1996; 37:1883–1890. Levey AS. Use of glomerular filtration rate measurements to assess the progression of renal disease. Semin Nephrol 1989; 9:370–379. Russell CD. Optimum sample times for single-injection, multisample renal clearance methods. J Nucl Med 1993; 24:1761–1765. Smith HW, Goldring W, Chasis H. The measurement of tubular excretory mass, effective blood flow and filtration rate in the normal human kidney. J Clin Invest 1938; 17:263–278. Russell CD, Dubovsky EV. Comparison of single-injection multisample renal clearance methods with and without urine collection. J Nucl Med 1995; 36:603–606. Russell CD. A bayesian 3-compartment model for 99mTc-MAG3 clearance. J Nucl Med 2003; 44:1357–1361. Frokiaer J, Knudsen L, Flo C, Hansen HH, Eika B, Mortensen J, et al. Reproducibility of iodine-123-hippuran renoscintigraphy in normal pig at various flow rates. Scand J Urol Nephrol 1989; 125 (Suppl):87–93. Donath A. The simultanous determination in children of glomerular filtration rate and effective renal plasma flow by the single injection clearance technique. Acta Paediatr Scand 1971; 60:512–520. Swan SK. The search continues – an ideal marker of GFR [editorial]. Clin Chem 1997; 43:913–914.

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Original article

Preparation of 99mTc-MAG3: radiochemical purity is affected by the residence time of sodium chloride injection in a three-part syringe Clinton C. Waight, Lynn A. Beattie, Lesley M. O’Brien and Alistair M. Millar Background Routine technetium-99m mercaptoacetyltriglycine (99mTc-MAG3) radiochemical purity measurements have revealed occasional unacceptably low values. Preliminary investigations suggested a causal link with the residence time of sodium chloride injection in the syringe used to reconstitute the MAG3 kit. Objectives To investigate the cause of this phenomenon, determine how it can be avoided and establish whether it occurs with other 99mTc radiopharmaceuticals.

chloride injection treated with lubricated and unlubricated syringe rubber plunger ends had radiochemical purities of 85.3 ± 6.6% and 82.1 ± 6.5% (n = 5), respectively. The radiochemical purities of other 99mTc radiopharmaceuticals prepared using sodium chloride injection incubated for 0 or 15 min in Plastipak syringes were as follows: 99mTcexametazime, 95.3 ± 0.6% and 94.5 ± 1.8%; 99mTc-sestamibi, 98.0 ± 0.6% and 97.7 ± 0.6%; 99mTc-tetrofosmin, 96.5 ± 0.2% and 97.0 ± 0.4%. None of the differences was significant (P > 0.05, n = 5).

Methods 99mTc-MAG3 was prepared by drawing sodium chloride injection into a lubricated, three-part, 10 ml Plastipak syringe and using it to reconstitute a MAG3 kit immediately or after a 15 min incubation period. The radiochemical purity was measured by high-performance liquid chromatography. The experiment was repeated using lubricant-free, two-part, Norm-Ject syringes and lubricated, two-part, Monoject syringes (15 min incubation only). To investigate the influence of Plastipak’s rubber components on the radiochemical purity, samples were prepared using sodium chloride injection that had been incubated with lubricated or lubricant-free rubber plunger ends. Similar experiments were performed to determine the effect of Plastipak on 99mTc-exametazime, 99mTcsestamibi and 99mTc-tetrofosmin.

Conclusions A lipophilic impurity, originating from the rubber plunger of a three-part Plastipak syringe, is formed in 99mTc-MAG3 when the sodium chloride injection used to reconstitute the kit is in the syringe for a prolonged time. The effect is eliminated by using a two-part syringe or by injecting the sodium chloride injection into the kit immediately. The phenomenon does not occur with 99mTcexametazime, 99mTc-sestamibi or 99mTc-tetrofosmin. Nucl c 2006 Lippincott Williams & Med Commun 27:197–200 Wilkins.

Results The radiochemical purities of 99mTc-MAG3 prepared with sodium chloride injection incubated for 0 and 15 min in Plastipak syringes were 96.4 ± 0.5% and 89.4 ± 5.5%, respectively. The difference was significant (P < 0.05, n = 10). With Norm-Ject syringes, the radiochemical purities were 96.5 ± 0.5% and 96.6 ± 0.5%, respectively. The difference was not significant (P > 0.05, n = 10). With Monoject syringes, the radiochemical purity was 96.6 ± 0.4% (n = 10). 99mTc-MAG3 prepared using sodium

Radiopharmacy, The Royal Infirmary of Edinburgh, Edinburgh, UK.

Introduction

the radiochemical purity of 99mTc-MAG3 prepared in our department revealed an occasional unacceptably high level of a lipophilic impurity. The levels of impurity were such that the products did not achieve the minimum radiochemical purity of 94% that is specified for 99mTcMAG3 in the European Pharmacopoeia [9], and the products were therefore unacceptable for administration to patients. Investigations into the possible causes of this

99m

Technetium-99m mercaptoacetyltriglycine ( TcMAG3) is a robust radiopharmaceutical of high radiochemical purity when prepared using various techniques and conditions [1–3]. There have, however, been reports of ancillary materials, such as sanitizing agents and sodium chloride injection from plastic ampoules, causing low radiochemical purity [4–8]. Routine measurements of

Nuclear Medicine Communications 2006, 27:197–200 Keywords: preparation, radiochemical purity,

99m

Tc-MAG3

Correspondence to Mr C. C. Waight, Department of Pharmacy, The Royal Infirmary of Edinburgh, Little France Crescent, Edinburgh EH16 4SA, UK. Tel: + 44 131 242 2930; fax: + 44 131 242 2931; e-mail: [email protected]

Received 30 August 2005 Accepted 15 November 2005

c 2006 Lippincott Williams & Wilkins 0143-3636

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198

Nuclear Medicine Communications 2006, Vol 27 No 2

phenomenon suggested a correlation between the appearance of the impurity and the residence time of sodium chloride injection in the syringe used for reconstitution of the kit. The aims of this work were to investigate the cause of this phenomenon, determine how its recurrence could be avoided and establish whether the effect was relevant to other radiopharmaceuticals.

care), 99mTc-sestamibi (Cardiolite, Bristol-Myers Squibb, Brussels, Belgium) and 99mTc-tetrofosmin (Myoview, GE Healthcare) were prepared using Plastipak 10 ml syringes to inject the sodium chloride injection into the kits. As above, samples produced by immediate transfer of the sodium chloride injection and samples produced following 15 min of incubation in the syringe were prepared for comparison. Measurement of radiochemical purity

Materials and methods Preparation of radiopharmaceuticals 99m

Tc-MAG3 was prepared by injecting Sodium Chloride Injection BP (Braun, Melsungen, Germany) followed by Sodium Pertechnetate [99mTc] Injection BP from a Drytec generator (GE Healthcare, Amersham, Buckinghamshire, UK) into a Technescan MAG3 kit (Mallinkrodt Medical, Petten, the Netherlands) to give a radioactive concentration of 500 MBq/10 ml. The kit was heated for 10 min in a boiling water-bath and then cooled in a room temperature water-bath for 10 min. Ten samples were prepared by drawing the sodium chloride injection into a three-part 10 ml Plastipak syringe (Product code 300912, Becton Dickinson, Franklin Lakes, New Jersey, USA) and injecting it immediately into the MAG3 kit. The Sodium Pertechnetate [99mTc] Injection was injected into the kit using a 3 ml Plastipak syringe (Product code 300910). A further 10 samples were prepared using an identical technique, except that the sodium chloride injection was incubated in the syringe for 15 min before being injected into the kit. The above experiment was repeated using lubricant-free, two-part, Norm-Ject syringes (Henke Sass Wolf, Tuttlingen, Germany) and lubricated, two-part, Monoject syringes (Tyco Healthcare UK Ltd, Gosport, Hampshire, UK) with 15 min incubation only.

The radiochemical purity of 99mTc-MAG3 was measured by high-performance liquid chromatography (HPLC) using a method reported previously [1]. Briefly, a 20 ml sample of test material was injected onto a 250  4.6 mm Hypersil ODS column (Thermo Electron Corporation, Bellefonte, Pennsylvania, USA) via a loop valve injector. The column was eluted at 1 ml/min with a mobile phase containing ethanol and 10 mM phosphate buffer pH 6 (5 : 95), for 7 min and then methanol and water (90 : 10) for 8 min. These conditions gave a chromatogram with a group of hydrophilic 99mTc impurities with retention times of 2–4 min, a 99mTc-MAG3 peak with a retention time of 5 min and a group of lipophilic 99mTc impurities with retention times of 7–12 min. Samples were tested 1 h after preparation. The radiochemical purity of 99mTc-exametazime was measured by HPLC using a modification of the method of Neirinckx et al. [10]. Analysis was performed using gradient elution of a 150  4.6 mm PRP-1 column (Hamilton, Reno, Nevada, USA) fitted with a 25  3 mm guard column. The column was eluted with 20 mM phosphate buffer pH 7.4, at a flow rate of 2 ml/min. Immediately after injection of the sample, tetrahydrofuran was introduced into the mobile phase in a linear gradient of 0–25% over 6 min. The radiochemical purity of 99mTc-tetrofosmin was measured by thin-layer chromatography (TLC) on an ITLC-SG plate (Pall, Ann Arbor, Michigan, USA) with a mobile phase containing acetone and dichloromethane (35 : 65) [11].

To determine whether the incidences of low radiochemical purity were attributable to the black rubber plunger ends of the Plastipak syringes, the rubber components were removed from five syringes. These were immersed in 10 ml of sodium chloride injection in a glass vial. The solution was agitated on a vortex mixer for 15 min and then drawn into a Norm-Ject, lubricant-free, two-part syringe and used in the preparation of 99mTcMAG3. To determine whether the effect was caused by the lubricant in the syringe, the experiment was repeated with rubber plunger ends from which the lubricant had been removed by wiping with tissue before being immersed in the sodium chloride injection.

The radiochemical purity of 99mTc-sestamibi was measured by a combination of TLC on a 5  10 cm RP-18 F254s plate (Merck, Darmstadt, Germany) with a mobile phase containing tetrahydrofuran, ammonium actetate 38.5 g/l, methanol and acetonitrile (10 : 20 : 30 : 40) and HPLC on a 5 mm Hypersil BDS column (Thermo Electron Corporation) with a mobile phase containing acetonitrile, ammonium sulphate 6.6 g/l and methanol (20 : 35 : 45) [9].

To determine whether other radiopharmaceuticals were susceptible to the effect observed with 99mTc-MAG3, samples of 99mTc-exametazime (Ceretec, GE Health-

The eluates from the HPLC columns were monitored with an on-line scintillation detector. The distribution of 99m Tc on TLC plates was measured with a scintillation

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Effect of syringes on the radiochemical purity of

detector on a Mini-Scan radiochromatogram scanner (Bioscan, Washington DC, USA). Chromatograms were recorded and analysed with LauraLite 3 radiochromatography software (LabLogic, Sheffield, Yorkshire, UK). The radiochemical purity was calculated by expressing the counts in the principal peak as a percentage of the total counts in the chromatogram. The statistical significance between values of radiochemical purity obtained under different conditions was measured using the paired t-test. P < 0.05 was taken to demonstrate a significant difference.

Results The results of the radiochemical purity measurements on 99m Tc-MAG3 are summarized in Table 1. All 10 samples prepared by immediate transfer of the sodium chloride injection with a Plastipak syringe had high radiochemical purities that were typical of 99mTc-MAG3. However, when the sodium chloride injection was incubated in the Plastipak syringe for 15 min before being injected into the kit, a wide variation in radiochemical purity was obtained. Seven of the 10 samples had radiochemical purities lower than the 94% limit and the mean was also below the limit. The difference between the results from the two incubation times was statistically significant. The samples contained normal levels of hydrophilic impurities (B1%), but high levels of lipophilic impurities, which were responsible for the low radiochemical purities. In contrast, all samples prepared using Norm-Ject, lubricantfree, two-part syringes and Monoject, lubricated, two-part syringes had high radiochemical purities. The difference between the results from the Norm-Ject syringes at the two incubation times was not statistically significant. In the experiments to investigate the role of the Plastipak rubber plunger end and lubricant, all 10 samples prepared with sodium chloride injection that had been incubated with either lubricated or lubricant-free rubber plunger ends contained high levels of lipophilic impurities.

Tc-MAG3 Waight et al. 199

difference between the results from the two incubation times was not statistically significant.

Discussion The radiochemical purity of 99mTc-MAG3 is affected by surprising factors. The use of a bacteriostatic guard vial on the 99mTc generator [6,7], a disinfectant alcoholic spray containing hydrogen peroxide [6,7], the disinfectant Biocide B (M. Slade, Taunton, Somerset, UK personal communication, 2005) and sodium chloride injection from plastic ampoules [5,8] can all have deleterious effects on the radiochemical purity of 99m Tc-MAG3. Our current finding that low radiochemical purity can be caused by reconstituting the MAG3 kit with sodium chloride injection that has been allowed to stand in a plastic syringe is another apparently innocuous act that can affect the radiopharmaceutical. A similar effect has been reported previously for the liver imaging radiopharmaceutical 99mTc-tin colloid [12]. When this radiopharmaceutical was prepared using a specific brand of syringe, an unacceptably high level of a radiochemical impurity was created. The impurity was shown to originate from the rubber plunger end of the syringe. The chromatograms from the 99mTc-MAG3 samples that were prepared with sodium chloride injection that had been incubated in Plastipak syringes revealed that the cause of the low radiochemical purity was a lipophilic impurity. The presence of a high level of lipophilic impurity in 99mTc-MAG3 has not been reported previously. All previous incidences of low radiochemical purity for 99mTc-MAG3 have been due to the presence of high levels of hydrophilic impurities. Table 2 Radiochemical purities of radiopharmaceuticals prepared with sodium chloride injection incubated in a syringe for different times Radiochemical purity (%) Radiopharmaceutical 99m

Tc-exametazime Tc-sestamibi 99m Tc-tetrofosmin 99m

The results for the other 99mTc radiopharmaceuticals are summarized in Table 2. For each radiopharmaceutical, the

99m

0 min incubation

15 min incubation

95.3 ± 0.6 98.0 ± 0.6 96.5 ± 0.2

94.5 ± 1.8 97.7 ± 0.6 97.0 ± 0.4

Values of radiochemical purity are shown as mean ± standard deviation (n = 5).

99m Table 1 Levels of radiochemical species in technetium-99m mercaptoacetyltriglycine ( Tc-MAG3) resulting from various sodium chloride injection incubation conditions

Incubation condition Plastipak, 0 min Plastipak, 15 min Norm-Ject, 0 min Norm-Ject, 15 min Monoject, 15 min Lubricated rubber tips, 15 min Wiped rubber tips, 15 min Values for

99m

n 10 10 10 10 10 5 5

Hydrophilic

99m

Tc impurity (%)

1.0 ± 0.2 1.1 ± 0.3 1.0 ± 0.2 1.0 ± 0.2 0.9 ± 0.2 1.4 ± 0.4 0.8 ± 0.3

Lipophilic

99m

Tc impurity (%)

2.6 ± 0.3 9.5 ± 5.3 2.5 ± 0.4 2.7 ± 0.4 2.6 ± 0.2 13.4 ± 6.4 17.0 ± 6.5

99m

Tc-MAG3(%)

Number of fails

96.4 ± 0.5 89.4 ± 5.5 96.5 ± 0.5 96.3 ± 0.5 96.6 ± 0.4 85.2 ± 6.6 82.2 ± 6.5

0 7 0 0 0 5 5

Tc species are shown as mean ± standard deviation. Number of fails = number of samples with radiochemical purity of less than 94%.

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200 Nuclear Medicine Communications 2006, Vol 27 No 2

The Norm-Ject syringe was studied as a possible solution to the low radiochemical purity as, unlike the Plastipak, it has a two-piece construction, that is, there is no rubber end on the plunger and it does not contain lubricant. The use of the Norm-Ject syringe was successful in eliminating the low radiochemical purity that resulted from incubation of sodium chloride injection in the syringe before it was injected into the MAG3 kit. This finding suggested that the origin of the impurity was either the rubber end on the plunger or the presence of lubricant in the Plastipak syringe. This theory was strengthened by the high level of lipophilic impurity detected when 99mTcMAG3 was prepared with sodium chloride injection in which the rubber plunger ends from five Plastipak syringes had been incubated for 15 min. In an attempt to determine whether the rubber or lubricant was responsible for the phenomenon, the experiment was repeated with tips from which the lubricant had been wiped as thoroughly as possible. Similar levels of lipophilic impurity were detected, suggesting that the rubber, not the lubricant, caused the effect. Wiping was chosen as the means of removing the lubricant as the use of a solvent, such as dichloromethane, might have leached any causative agents from the rubber. Unfortunately, wiping cannot be guaranteed to remove all traces of lubricant. Conclusive proof would have required the use of sodium chloride injection that had been adulterated with lubricant, but the Plastipak syringe manufacturer declined to provide a sample of lubricant for the experiment. The lubricant in Monoject syringes is a silicone fluid (Dow Corning 360 Medical Fluid). Although we have been unable to establish the nature of the lubricant in Plastipak syringes, it is likely to be of a similar composition. The satisfactory results obtained after incubation of the sodium chloride injection in Monoject syringes provides further evidence that the lubricant is not responsible for the formation of the lipophilic impurity. Having identified this potential problem with 99mTcMAG3, we decided that other radiopharmaceuticals should be studied to determine whether the effect was relevant to them. 99mTc-exametazime, 99mTc-sestamibi and 99mTctetrofosmin were chosen for two reasons. Firstly, they are all lipophilic compounds and, as the impurity detected in 99m Tc-MAG3 is lipophilic, they might be susceptible to the same effect. Secondly, for each there is an analytical technique that has the specificity to detect lipophilic impurities. None of these radiopharmaceuticals was found to be susceptible to the effect observed with 99mTc-MAG3. Dispensing sessions in most hospital radiopharmacies are busy periods of activity with a large number of injections being prepared in a short time. Time-saving measures are always being sought. The results of this work demonstrate that the drawing up of the sodium chloride

injection in anticipation of it being used to reconstitute a radiopharmaceutical kit is one such seemingly innocuous time-saving measure that can have serious consequences for the quality of 99mTc-MAG3. Also, during the preparation of 99mTc-MAG3, there is the possibility that the sodium chloride injection might be held in a syringe whilst the activity of sodium pertechnetate [99mTc] that is to be used in the reconstitution of the kit is measured, or until the water-bath reaches boiling point. To ensure the preparation of high radiochemical purity 99mTc-MAG3, this situation should be avoided. The most effective means of eliminating the risk is to use two-part syringes for the preparation of 99mTc-MAG3.

Conclusions (1) When reconstituting a MAG3 kit with a three-part syringe, inject the sodium chloride injection into the kit immediately to prevent the formation of a product of low radiochemical purity. (2) The low radiochemical purity is caused by a species that leaches from the rubber plunger end of the syringe. (3) The effect can be eliminated using a two-part syringe. (4) 99mTc-exametazime, 99mTc-sestamibi and 99mTctetrofosmin are not affected by the phenomenon.

Acknowledgements We are grateful to Amanda Bryan for her assistance with the analysis of the 99mTc-exametazime samples and to Dr Hector Knight of Mallinckrodt Medical BV, Petten, the Netherlands for supplying Technescan MAG3 kits.

References Millar AM, Wilkinson AG, McAteer E, Best JJK. 99mTc-MAG3: in vitro stability and in vivo behaviour at different times after preparation. Nucl Med Commun 1990; 11:405–412. 2 Millar AM, O’Brien L. An investigation of factors that might influence the radiochemical purity and stability of 99mTc-MAG3. Eur J Nucl Med 1990; 16:615–619. 3 Millar AM, O’Brien L. Preparation of 99mTc-MAG3: no confirmation that sodium chloride injections from plastic containers affect radiochemical purity. Nucl Med Commun 1998; 19:475–477. 4 Stringer RE, Maltby PJ. A 1 year study of factors affecting 99mTc-MAG3 kit failure. Nucl Med Commun 1996; 17:993. 5 Mallinckrodt Medical. Use of saline in ‘plastic bottles’ for dilution of technetium kits. Letter to customers. Petten: Mallinckrodt Medical; 1996. 6 Stringer RE, Schroeder NE, Maltby PJ. MAG3 failure is due to inadvertent oxidant contamination. Nucl Med Commun 1997; 18:294. 7 Maltby P, Stringer R. Reconstitution of MAG3. Pharm J 1997; 258:839. 8 Millar AM, Hesslewood SR. A comparison of SepPak and high-performance liquid chromatography as techniques for measuring the radiochemical purity of 99mTc-MAG3. Nucl Med Commun 2004; 25:1049–1051. 9 European pharmacopoeia. 4th ed. Strasbourg: European Directorate for the Quality of Medicines; 2002. 10 Neirinckx RD, Canning LR, Piper IM, Nowotnik DP, Pickett RD, Holmes RA, et al. Technetium-99m d,l-HM-PAO: a new radiopharmaceutical for SPECT imaging of regional cerebral blood perfusion. J Nucl Med 1987; 28:191–202. 11 GE Healthcare. Myoview package insert. Amersham, Buckinghamshire: GE Healthcare; March 2003. 12 Slater DM, Anderson M, Garvie NW. Syringe extractibles: effects on radiopharmaceuticals. Lancet 1983; ii:1431–1432. 1

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NEWS AND VIEWS February 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: + 44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: + 44 (0)113 206 6930, E-mail: [email protected] Nuclear Medicine Communications, 2006, 27:201–202

Cracking the codes

In previous issues we have touched on NPfIT and examination coding and would now like to share the current state of development, as it will be of great importance to both the smooth workflow and finance of nuclear medicine departments in the near future. There are four levels of interconnected coding systems that are important to the NHS. It is easiest to think of them in terms of functionality and what better way than to follow the patient pathway. CRS codes

At the referral, a code system is required to identify the test being requested, this is the care records service (CRS) code. This tag will eventually be used in the picture archiving and communication system (PACS) index for serving up the results to those wishing to view the images (i.e. to identify the bone scan images amongst all the other radiological imaging done on the same patient). Let’s use as an example the bone scan. The raw CRS code is translated into descriptive text for human users as ‘Radionuclide bone study’. RIS codes

At the imaging dept the basic CRS code needs to be elaborated on so that the nuclear physician (practitioner in IRMER-speak) can indicate the entire procedure he/she wishes to take place, e.g. it could be three-

phase or whole-body plus SPECT. This will typically be fed into the radiology information system (RIS) which, depending on its complexity, can perform many tasks for the imaging department automatically, e.g. room scheduling, ordering tracers, generation of workload statistics. Here is the code for a wholebody bone scan (uptake phase only): NBONW. RIS and CRS codes are still under development and are regularly discussed and updated on the PACS and Teleradiology Group web site: www.pacsgroup.org.uk. Navigate to ‘Questions and Answers’ > ‘Examination Coding’ > ‘RIS coding and DESCRIPTORS + SNOMED’. SNOMED codes

After the scan, when the report is made, it may be useful to keep a code indicating the basic diagnosis. This is the SNOMED code which would be generated from the CRS code with additional suffixes indicating the referral criteria, diagnosis and much more. It is much the most complex of the four coding systems and there is some guidance at http://www.snomed.org/snomedct/what_is.html. The purpose of SNOMED is principally as a tool for evidence based data analysis, but being the most detailed record could be put to a wide variety of uses. It has to be said that the SNOMED code is not the most

readable by humans and would generally require a program between the user and the data. Here’s an example: 184681000000103 for a whole-body bone scan with no diagnosis yet included. Healthcare resource groups (HRGs)

These will also be generated from the CRS code. They are few and they are simple. Each nuclear medicine test will fall into one of only about five categories (levels 1 to 5) with, for example, GFR and red cell mass being level 1 and DATscan, MIBG, Octreoscan, and PET being level 5. These will be generated by the hospital computer systems and be sent to the referrer’s organization for reimbursement. This is of course very similar to Korner grouping but it is believed that for the time being both will exist in parallel, which is a shame since in the authors’ opinions Korner groups for nuclear medicine started off with illogical ranking and never got any better. RIS codes have been mapped to Korner codes. More information about HRGs can be found at http://www.icservices.nhs.uk/ casemix/pages/default.asp. Information about Korner codes is at http://www.dh. gov.uk/PolicyAndGuidance/Organisation Policy/FinanceAndPlanning/NHSCostingManual/fs/en. Meeting Announcements

PET/CT and SPECT/CT: Practical Issues and Applications

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Date: 21 February 2006 Venue: Geological Society, London Website: www.ipem.ac.uk Royal Marsden PET/CT Symposium Dates: 23 and 24 February 2006 Venue: London, UK Email: [email protected] Annual clinical PET CT course – a user’s guide Dates: 8–10 March 2006 Overview of basic science, PET CT imaging and interactive teaching. Contact for information and registration: [email protected] BNMS Spring Meeting Dates: 27–29 March 2006

Venue: Manchester, UK Website: www.bnms.org 2nd European IRPA Congress on Radiation Protection Dates: 15–19 May 2006 Venue: Paris, France Website: www.irpa2006europe.com BNMS Autumn Meeting Dates: 4–5 September 2006 Venue: Cambridge, UK Website: www.bnms.org.uk EANM Annual Meeting Dates: 29 September to 4 October 2006 Venue: Athens, Greece Website: www.eanm.org

9th World Congress of Nuclear Medicine and Biology Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/congress2006/ index02.htm Education and Training

EANM Learning Courses Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php E-mail: [email protected]

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Editorial

Changes to ARSAC: new notes for guidance and changes to operational arrangements Steven Ebdon-Jackson, J.H. McKillop and Thomas O. Nunan

Nuclear Medicine Communications 2006, 27:203–204

Correspondence to Steve Ebdon-Jackson. e-mail: [email protected]

Steve Ebdon-Jackson leads the ARSAC Secretariat, Jim McKillop is a previous chairman of the ARSAC, and Tom Nunan is the present chairman of the ARSAC.

Received 12 December 2005 Accepted 12 December 2005

The new notes for guidance The last revision of the Administration of Radioactive Substances Advisory Committee (ARSAC) Notes for Guidance on the Clinical Administration of Radiopharmaceutical and Uses of Sealed Radioactive Sources was published in December 1998 [1]. Over the past 2 years or so the committee has been reviewing this document in the light of changes to the legislation and changes in clinical practice. This editorial highlights the significant changes. The new document will not be made generally available in paper format, but can be reviewed on a new ARSAC website at www.arsac.org.uk. A single hard copy will also be available per site, on request. During the same period, there have been some changes to the structure of the ARSAC Secretariat and these will be covered in the second part of this editorial. Significant changes are as follows. Applicants who should apply and who need not apply

Further guidance regarding the prolonged absences of certificate holders, the issue of renewals, extensions and variation of certificates is included in the new notes. Applicants applying for renewal of current certificates are reminded that they should only apply for certificates for serials for diagnosis or treatment for which they have maintained clinical competence. Issues relating to diagnostic certificates

Further advice is given regarding administration of activities that vary from the diagnostic reference levels. This relates to occasions when the activity might be adjusted downwards and upwards.

should write to the ARSAC Support Unit by letter rather than submitting a new application. This letter should include the basis for the extension or changes. Investigations in children and young persons

This section has been shortened. In the previous version of the notes for guidance it was felt appropriate to provide a significant amount of detail regarding practical issues relating to paediatric nuclear medicine. This detail is now available at the European Association of Nuclear Medicine website (www.eanm.org) and hence does not need to be included in the notes for guidance. One new table in this section addresses minimum activities that should be administered to children requiring radionuclide investigations. Appendices

The most obvious change for those who use the notes for guidance frequently is the layout of part A. This is now in landscape format and includes a reference to the appropriate functional group for each radiopharmaceutical. There has been a review of all products in the list and many products which are no longer used routinely in clinical practice have been removed. Products which have come into routine use since the revision have been added. Sentinel node imaging was reviewed and separate serials for breast and melanoma sentinel node studies have been issued. If other tumour types, such as head and neck, cervical or uterine cancers also become a routine indication for sentinel nodes studies, then separate serials are likely to be issued for these as well.

Certificates for research

Functional groups, which were introduced in the last notes for guidance have caused some confusion to applicants. The new notes aim to explain how to apply for them. Functional groups for therapeutic administration were felt not to have worked well and these have been removed. In addition, information regarding the radiopharmaceutical and the clinical indication is included for each serial listed in the group tables.

Research certificates are valid for 2 years only. If the research project over-runs, or if the original parameters of the research are changed in a minor fashion, the applicant

A new appendix has been added covering training and experience. The purpose of this is to assist applicants

Certificates for therapy

The increasing use of multidisciplinary meetings and collaboration is recognized and, if appropriate, applicants are asked to provide evidence of participation in these teams.

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who have not undergone recognized training programmes such as those provided by the royal colleges. A new section on training and experience required for positron emission tomography has been added. A further section covers issues that occur when new techniques are developed and relate to training requirements for the support staff. A section has also been added on requirements of those working under the written directions of a certificate holder. This was written particularly with sentinel node biopsy procedures in mind, but the principles apply to other techniques that might be developed. In all issues relating to training, it is easier for the committee to advise health ministers that applicants have undergone appropriate training if that training has been undertaken under the auspices of a recognized provider such as a royal college. Those applicants who acquire training and experience through other routes will need to provide additional information to the committee in order that the committee can be satisfied. Two new appendices have been added: appendix 6 informs applicants of the process of certification and appendix 7 is intended to assist applicants with the communication of the risk associated with radionuclide procedures, to ethics committees, patients and research subjects.

Changes to operational arrangements The publication of the new notes for guidance coincides with changes within the Department of Health (DH) and the expansion of the Health Protection Agency (HPA). Since April 2005, the HPA has incorporated the National Radiological Protection Board (NRPB) and provides advice on radiation matters. In broad terms, while the responsibility for government policy on health matters remains with DH, advice on

radiation protection and radiological practice issues and many of DH’s operational functions are now provided by the HPA Radiation Protection Division (HPA-RPD). With regard to ARSAC, health ministers will continue to issue certificates and remain responsible for the appointment of ARSAC members, as stated in regulations and as is appropriate for a statutory advisory committee. Patricia Brown, who has been involved with ARSAC for many years, remains in DoH where among other things she will provide ARSAC’s interface with government. She has become the DoH observer to the committee. All ARSAC Secretariat functions have transferred to the Medical Exposure Department of HPA-RPD, based at Chilton. A number of staff have transferred from DoH to HPA-RPD, including Steve Ebdon-Jackson who leads the new ARSAC Secretariat. Staff of the ARSAC Support Unit are now an integral part of the Medical Exposure Department. A new database will come on-line in the spring of 2006, following the introduction of the new ARSAC website. The most significant change for certificate holders will be changes to the format of schedules to certificates; these will reflect applications which take advantage of the functional group facility. Further information and examples of these formats will be included on the website. In most other respects, the certification process will remain the same. For the forseeable future, applications with written signatures will still be required and certificates will be signed and distributed in hardcopy format.

References 1

Notes for guidance on the clinical administration of radiopharmaceuticals and use of sealed radioactive sources. NMC 2000; 21 (Suppl).

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Review paper

Fever of unknown origin: a systematic review of the literature for 1995–2004 Giovanni B. Gaeta, Francesco M. Fusco and Salvatore Nardiello Background Fever of unknown origin (FUO) identifies a pattern of fever with temperature higher than 38.38C on several occasions over more than 3 weeks, in which the diagnosis remains uncertain after an initial diagnostic work-up. The identification of the cause of FUO is a challenge in clinical practice despite recent advances in diagnostic techniques. There are more than 200 reported causes of FUO and they can be classified in four diagnostic categories: infections, neoplasms, non-infectious inflammatory diseases and miscellaneous. Methods We performed a systematic research of the literature on classical FUO to retrieve the review articles and case series published from 1995 to 2004, including articles from developing countries. The case series were reviewed to identify the tests commonly used both to qualify a fever as FUO and to determine the cause of the FUO, and to design an updated flow chart for the diagnosis of classical FUO. Results and Conclusions No standardized diagnostic strategy could be determined. The diagnostic process

Introduction Fever of unknown origin (FUO) identifies a pattern of fever that does not resolve spontaneously, in which the diagnosis remains uncertain after an initial diagnostic work-up. In 1961, FUO was defined by Petersdorf and Beeson as an illness of more than 3 weeks’ duration, with fever greater than 38.31C (1011F) on several occasions, the cause of which is uncertain after 1 week of in-hospital investigations. The first two criteria allow the elimination of most of the acute, self-limited diseases such as viral infection [1]. In 1991, Durak and Street proposed two major changes to the original definition by Petersdorf and Beeson. The first was the distinction between classical FUO and three other types, named nosocomial FUO, neutropenic FUO and HIV-associated FUO: indeed, in the latter three types of FUO both the spectrum of the underlying diseases and the clinical approach are different from classical FUO. The second change proposed by Durak and Street was the required duration of investigation before qualifying a fever as FUO, i.e., at least 3 days in hospital or at least three outpatient visits [2]: this reflects the greater importance given to outpatient investigation. Evolving knowledge and the improvement in diagnostic methods, including new microbiological techniques and

should be guided by the potential diagnostic clues (PDCs) emerging from the history, physical examination and baseline tests. A standardized flow chart can be applied only in absence of PDCs or when the PDCs are contradictory. Nuclear medicine techniques are a valuable aid in the search for the origin of FUO due to bacterial infections or in the absence of PDCs. Nucl Med Commun 27:205–211

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:205–211 Keywords: FUO, fever of unknown origin, diagnostic work-up, diagnostic clues Department of Infectious Diseases, Second University of Naples, Italy. Correspondence to Prof. Giovanni B. Gaeta, Dipartimento di Malattie Infettive, Seconda Universita` di Napoli, c/o osp. Gesu` e Maria, Via Cotugno 1, 80135 Naples, Italy. Tel: + 39 815666208; fax: + 39 815666206; e-mail: [email protected] Received 22 June 2005 Accepted 6 September 2005

new instrumental procedures, necessitate a constant update of the tests included in a minimal diagnostic workup to qualify a fever as FUO. In addition, the identification of the cause of FUO remains a challenge in clinical practice. A long list of diseases (more than 200) can cause FUO. The diagnostic work-up is frustrating for patients and physicians because it involves several non-invasive and invasive procedures, and fails to reach a final diagnosis in about 25% of cases [3–5]. We performed a systematic review of the literature on classical FUO published from 1995 to 2004 to identify the tests commonly used by different authors to K K

qualify a fever as FUO, determine the cause of the FUO.

Our purpose was to design an updated flow chart for the diagnosis of classical FUO.

Methods The MEDLINE database was searched for the following keywords: fever of unknown origin, FUO, pyrexia of unknown origin, PUO. The limits applied were: English language, publication date from 1995 to 2004, keywords

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present in the title/abstract. Further research was done on the Cochrane database and Embase. This systematic approach resulted in 705 articles. We excluded the articles focusing on neutropenic patients (108 titles), paediatric patients (48 titles), and HIV + patients (35 titles). The remaining 514 titles were checked independently by two of us in order to retrieve case series and review articles: 11 case series [6–17] and 18 review articles [3–5,18–32] were found. A check of the references of the review articles did not afford any further case series. An assessment of the quality of the 11 case series was performed according to the Newcastle–Ottawa Scale (NOS) for case–control studies [33], limited to the items concerning ‘adequacy of the case definition’ and ‘representativeness of the case’. The data of the case series on FUO were analysed in order to investigate their general characteristics, the diagnostic work-up performed to qualify a fever as FUO and the diagnostic procedures used to reach a final diagnosis. The chi-squared test was used to compare prevalences and 5% was considered as significance level.

Results Using the NOS criteria, the ‘representativeness of the case’ was adequate for all 11 case series, while the ‘case definition’ was adequate for all but three case series [10,13,15], in which the recruitment of the patients was not made according to independent validation. The series with inadequate ‘case definition’ included 558 patients out of 1488 (37.5%). In these patients, no statistically significant differences were found in the final diagnoses, except for the category of diagnosis ‘miscellaneous’, which is higher in the series with an inadequate assessment of quality (14.9% vs. 7.7%, P = 0.000). The main characteristics of the 11 series reporting 1488 cases of classical FUO are presented in Table 1. Among the 11 case series, three were from Europe (two Table 1

from the Netherlands and one from Belgium), three were from the Middle East (two from Turkey and one from Israel) and five from the Far East (three from India, one from China and one from Taiwan). Six studies were retrospective and five were prospective. There were no differences in diagnosis between prospective and retrospective series. The period of recruitment ranged from 8 months to 27 years. All series included more than 50 patients; eight of them included at least 100. Men were prevalent in all series except three and constituted 56.2% of cases. The mean age of patients ranged from 29.3 to 53.7 years. Six case series adopted the Petersdorf & Beeson criteria for the case definition, three adopted the modified criteria of Durak & Street and two studies adopted personal nonstandardized criteria. The minimal diagnostic work-up carried out to qualify a fever as FUO was reported in six case series [7–9, 11–13,15]. Routine blood tests, haemogram, urinalysis, blood cultures and chest X-ray were performed as initial screening procedures in all six series (Table 2). The majority of the authors (four out of six) included urine culture. The other diagnostic procedures were performed in half or fewer of the studies. In particular, an electrocardiogram and an HIV test were performed only in series from the Far East, while a large number of other tests were performed only in the series from Europe and Turkey (Fig. 1). Table 3 shows the categories of diagnoses in the 11 case series. The most frequent diagnostic category was infection (36.6%), followed by non-infectious inflammatory disease (15.9%) and by neoplasm (11.2%). In 10% of FUO the diagnosis could not be included in any of the previous categories, and in about 25% of the cases FUO remained undiagnosed. The case series from the Netherlands and from Belgium reported the lowest prevalence of infections. The same case series reported the highest prevalence of undiagnosed FUO.

Characteristics of 11 case series on fever of unknown origin published between 1995 and 2004

First author, year and reference

Country

Model of study

Study period

Number of patients

M/F

Mean age (years)

Definition criteria

de Kleijn, 1995 [6] Handa, 1996 [7]

Netherlands India

Retrospective Prospective

53 121

25/28 64/57

51 29.5

P–B Personal

de Kleijn, 1997 [8,9] Jung, 1999 [10] Kejariwal, 2001 [11] Vanderschueren, 2003 [12] Zhiyong, 2003 [13] Zamir, 2003 [14] Tabak, 2003 [15] Liu, 2003 [16] Saltoglu, 2004 [17] Total

Netherlands India India Belgium

Prospective Retrospective Prospective Prospective

1988–1992 not specified (lasting 2 years) 1992–1994 1993 1998–2001 1990–1999

167 233 100 223

80/87 150/83 59/41 131/92

49.6 29.3 32.4 53.7

P–B Personal P–B D–S

China Israel Turkey Taiwan Turkey

Retrospective Retrospective Retrospective Retrospective Prospective

1997–2001 1972–1998 1990–2001 1999–2002 1994–2002

208 101 117 78 87 1488

119/89 54/47 51/66 42/36 61/26 836 (56.2%)/652 (43.8%)

35 53 35 49 38.5 40.6

P–B D–S P–B D–S P–B

P–B: criteria as defined by Petersdorf and Beeson. D–S: criteria as defined by Durak and Street.

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A literature review of fever of unknown origin Gaeta et al. 207

Table 2

Diagnostic procedures performed in case series on fever of unknown origin published between 1995 and 2004 to qualify a fever as

FUO Diagnostic procedures

Handa, 1996 [7]

de Kleijn, 1997 [8,9]

Kejariwal, 2001 [11]

Vanderschueren, 2003 [12]

Zhiyong, 2003 [13]

Tabak, 2003 [15]

x x x x x x x

x x x x x

x x x x x x

x x x x x

x x x x x

x x x x x x x

Routine blood test Haemogram Urinalysis Haemoculture Chest X-ray Smears for malaria Urine culture Abdominal ultrasound Tine test Faecium culture Widal reaction or infectious serology Occult blood in stool ECG Sputum culture HIV test Antinuclear antibodies Rheumatoid factor

x x x x

x x

x x

x x

x

x x

x

x

x x

x

x

x

x x

x x

Fig. 1

3 Europe and Turkey Asia

Series

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Method performed Comparison between minimal diagnostic work-up performed in case series from Europe and Turkey (’) versus case series from Asia (&). US, ultrasound; ECG, electrocardiogram.

The diagnostic procedures performed to reach the final diagnosis are clearly reported in five of the 11 case series (594 patients out of 1488, 36.3%). Diagnosis was made in 408 of the 594 cases (Table 4). Some case series [8,9,17]

reported only the methods that were decisive for the final diagnosis, while others [6,11,12] reported all the methods contributing to the final diagnosis. Among the procedures performed to obtain the diagnosis 69.2% were

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Table 3

Diagnostic categories of fever of unknown origin in 11 case series published between 1995 and 2004

First author, year and reference

Number of patients

Infection (%)

Neoplasm (%)

NIID (%)

Miscellaneous (%)

No diagnosis (%)

de Kleijn, 1995 [6] Handa, 1996 [7] de Kleijn, 1997 [8,9] Jung, 1999 [10] Kejariwal, 2001 [11] Vanderschueren, 2003 [12] Zhiyong, 2003 [13] Zamir, 2003 [14] Tabak, 2003 [15] Liu, 2003 [16] Saltoglu, 2004 [17] Total

53 121 167 233 100 223

20.7 43.8 25.7 49.5 53.0 14.3

18.9 8.3 12.6 0.9 17.0 10.8

22.6 15.7 24.0 0.4 11.0 20.6

7.5 13.2 7.8 25.8 5.0 10.3

30.2 19.0 29.9 26.7 14.0 43.9

208 101 117 78 87 1488

31.7 54.5 34.0 42.3 58.6 36.6

16.8 7.9 19.0 6.5 13.8 11.2

22.1 2.0 23.0 20.5 18.4 15.9

5.3 2.9 10.0 7.7 2.3 10.4

24.0 32.7 14.0 23.1 6.9 25.9

NIID, non-infectious inflammatory disease.

Table 4

Methods performed to obtain the diagnosis in case series on fever of unknown origin published between 1995 and 2004

Author, year and reference

Number of diagnoses/ total of patients (%)

De Kleijn, 1995 [6] De Kleijn, 1997 [8,9] Kejariwal, 2001 [11] Vanderschueren, 2003* [12] Saltoglu, 2004 [17] Total

Number of non-invasive methods performed to obtain diagnosis

Number of invasive methods performed to obtain diagnosis

History and evolution

Laboratory tests

Instrumental tests

Biopsy

Other

(69.8) (70.1) (86) (56.1)

5 0 16 24

11 39 27 18

5 87 9 16

12 27 26 42

4 1 8 5

81/87 (93.1) 408/594

22 67

43 138

53 170

34 141

8 26

37/53 117/167 86/100 125/223

*

Twenty diagnoses obtained with unspecified combinations of methods.

Table 5 Number of diagnoses obtained in patients with or without potential diagnostic clues (PDCs) in two case series on fever of unknown origin Author, year and reference de Kleijn, 1995 [6] de Kleijn, 1997 [8,9] Total

Number of diagnoses/total of patients (%)

Number of diagnoses in patients with PDCs (%)

Number of diagnoses in patients without PDCs (%)

37/53 (69.8) 117/167 (70.1) 154/220 (70)

37/48 (77.1) 114/162 (73) 151/210 (72)

0/5 (0) 3/5 (60) 3/10 (30)

non-invasive and 30.8% were invasive. Biopsy was strongly prevalent among the invasive methods (84.4%). Only two case series, both by the same author [6,8,9], highlight the importance for the diagnosis of the potential diagnostic clues (PDCs), which are based on the history, physical examination and tests performed (Table 5). In these two studies a total of 220 patients were enrolled: PDCs were present in 210 of them (95.4%). The final diagnosis was reached in 151 of the patients presenting with PDCs (72%) and in three of the patients without PDCs (30%, P = 0.013).

Discussion This systematic review retrieved 11 case series of FUO published in the period 1995–2004 and included 1488 patients. Our review considered only English language literature. Five case series were from eastern Asia, three from the Middle East and three from northern Europe.

The definition of FUO was based on standard criteria [1,2] in nine series although only six papers specified the investigations performed to qualify a fever as FUO (minimal diagnostic work-up). The period of recruitment of these case series ranged from 1990 to 2001. Therefore, these studies are comparable regarding the availability of the diagnostic methods. A minimal diagnostic work-up included routine blood tests, haemogram, urinalysis, blood culture and chest X-ray. More specialized examinations were performed differently in the case series on the basis of the local epidemiological data and the availability of costly diagnostic facilities. For instance, cultures of materials other than blood or urine and screening for auto-immunity or occult blood in the stool were performed only in series from Europe and Turkey, justified by the greater availability of resources and by the high prevalence of non-infectious inflammatory disease or neoplasm in these areas. Instead, the routine use of peripheral blood smears for malaria is more frequent for countries where the disease is common.

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A literature review of fever of unknown origin Gaeta et al. 209

Fig. 2

Fever greater than 38.3°C on several occasions, lasting from at least 3 weeks

Minimal diagnostic work-up (routine blood tests, haemogram, urinalysis, blood culture, chest X-ray, urine culture, tine-test, abdominal and pelvic ultrasound, infectious direct and serological methods based on local epidemiological data, discontinuation of non-essential drugs)

Diagnosis? No FUO

No diagnosis? FUO

Oriented history (in specialist units) Complete physical examination

Potential diagnostic clues

No diagnostic clues

First Line. Further investigations for: infections: direct tests or serology for unusual presentation of the common disease, WBC scan neoplasm: screening laboratory tests (neoplastic markers), imaging (CT, MRI) NIID: screening laboratory tests Other: TSH -

Diagnosis? OK!

Oriented diagnostic procedures (especially biopsies and other investigations such as cultures, etc.)

No diagnosis?

Diagnosis? OK!

No diagnosis?

Second Line. Further investigations: direct tests or serology for rare diseases, cultures for hard-to-grow pathogens (HACEK) nuclear medicine (whole body imaging with non-specific agents – 67Ga or 18F-FDG scan)

Diagnosis? OK!

No diagnosis?

-

Wait and see strategy Therapeutic trials

Diagnostic flow chart for fever of unknown origin (FUO). CT, computed tomography; MRI, magnetic resonance imaging; NIID, non-infectious inflammatory disease.

The application of a different minimal diagnostic work-up may lead to the inclusion of different populations of patients even though selected using the same criteria;

thus, the comparability of the clinical series of FUO should take into account the geographical origin and the choice of baseline diagnostic examinations. Comparability

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210 Nuclear Medicine Communications 2006, Vol 27 No 3

of clinical series on FUO would be increased using a standardized minimal diagnostic work-up. Among the tests performed in the case series, urine culture and tinetest are low-cost tests and should be included in a standardized minimal diagnostic work-up before qualifying a fever as FUO. Abdominal and pelvic ultrasound requires specialized operators and, if available, should be used in the minimal diagnostic work-up. Likewise, direct and serological tests for the detection of infections based on local epidemiological data should be included. Overall, infections were the most frequent ascertained diagnoses in FUO patients. Despite reports that the prevalence of infections is declining over time as a cause of FUO, the prevalence of infections in series reviewed in our paper is higher than in case series published from 1961 to 1995 (reviewed by Knockaert et al. [3] and Arnow and Flaherty [4]). The high prevalence of infections in our series is due to the inclusion of case series from developing countries. Indeed, dividing the diagnoses according to geographical origin reveals that infections were more frequent in the series from the Far East (43.2%) and Middle East (47.8%) than those from Europe (19.4%). The prevalence of neoplasm in the case series included in our review was lower than in previous case series [3,4]. This trend may be due to improvements in diagnostic methods, which allow an early detection of neoplasms. The prevalence of undiagnosed FUO was high in case series in this study. The undiagnosed cases of FUO are increasing over time [3–5]. Therefore, only difficult-to-diagnose diseases are qualified as FUO, due to the increasing availability of diagnostic facilities, both in hospital and outpatient settings. The analysis of the procedures performed to identify the cause of FUO shows that non-invasive methods led to most of the diagnoses, whereas biopsy contributed to a high percentage to the final diagnoses. Thus, both noninvasive methods, such as well-conducted clinical history and well-chosen laboratory and instrumental methods, and invasive methods, especially biopsy, may contribute to the final diagnosis of FUO. However, an early use of biopsy may be encouraged in the presence of potential diagnostic clues.

Our systematic review analysed studies published over a 10-year period during which new diagnostic techniques have been successfully applied in selected patients. These techniques show high sensitivity and specificity in identifying some conditions that are a frequent cause of FUO. In particular, several studies focused on the application of nuclear medicine to detect the cause of FUO. 67Ga citrate, labelled leukocytes (111In, 99mTc), labelled human immunoglobulin (111In, 99mTc) and 2-[18F]fluoro-2-deoxy-D-glucose (FDG) represent the most widely investigated agents in the field of FUO [34–40]. Current clinical practice should take these results into account, even though no extensive randomized trials have been performed to provide high degree evidence. Nuclear medicine methods should be used as second-line tests to identify a potential cause of FUO in the absence of PDCs or in the presence of misleading PDCs. In these cases, whole-body imaging with nonspecific agents, i.e. agents that may point to infection, neoplasm or inflammation (67Ga citrate, FDG scan) is recommended. If a potential cause is identified, it can be further investigated using instrumental methods or biopsy. In conclusion, the diagnosis of FUO may not be a problem of evidence-based medicine. The main target of a diagnostic flow chart is to identify the PDCs. A standardized flow chart should be used only in the absence of PDCs, or when the PDCs are misleading. A diagnostic flow chart for FUO is presented in Fig. 2. The flow chart is based on the review conducted on the case series published from 1995 to 2004 and on some considerations, concerning both the geographical origin of the patients and the rapidly evolving diagnostic methods. The flow chart proposes a homologation of the minimal diagnostic work-up to achieve a more standardized definition of FUO. Moreover, the proposed flow chart allows a rational approach to the several laboratory, instrumental and invasive methods used to reach a final diagnosis.

References 1 2 3

The diagnostic process to identify the cause of FUO should be guided by the PDCs. The diagnoses obtained in patients presenting PDCs were significantly higher than in patients without PDCs (72 vs. 30%, P = 0.013). Therefore the first aim of physicians approaching patients with FUO must be to find PDCs through the medical history and physical examination. Only when no PDCs are found, or when the PDCs do not reveal the cause of FUO, a standardized step-to-step approach should be applied. All the PDCs that emerge from the standardized approach should be given credit.

4 5 6 7 8

9

Petersdorf RG, Beeson P. Fever of unexplained origin: report on 100 cases. Medicine (Baltimore) 1961; 40:1–30. Durak DT, Street AC. Fever of unknown origin – re-examined and refined. Curr Clin Top Inf Dis 1991; 11:35–51. Knockaert DC, Vanderschueren S, Blockmans D. Fever of unknown origin in adults: 40 years on. J Intern Med 2003; 253:263–275. Arnow PM, Flaherty JP. Fever of unknown origin. Lancet 1997; 350:575–580. Hirschmann JV. Fever of unknown origin in adults. Clin Infect Dis 1997; 24:291–302. de Kleijn EM, van der Meer JW. Fever of unknown origin (FUO): report on 53 patients in a Dutch university hospital. Neth J Med 1995; 47:54–60. Handa R, Singh S, Singh N, Wali JP. Fever of unknown origin: a prospective study. Trop Doct 1996; 26:169–170. de Kleijn EM, Vandenbroucke JP, van der Meer JW. Fever of unknown origin (FUO). I. A prospective multicenter study of 167 patients with FUO, using fixed epidemiologic entry criteria. The Netherlands FUO Study Group. Medicine (Baltimore) 1997; 76:392–400. de Kleijn EM, van Lier HJ, van der Meer JW. Fever of unknown origin (FUO). II. Diagnostic procedures in a prospective multicenter study of 167 patients.

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10 11

12

13

14 15 16

17

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The Netherlands FUO Study Group. Medicine (Baltimore) 1997; 76: 401–414. Jung A, Singh MM, Jajoo U. Unexplained fever-analysis of 233 cases in a referral hospital. Indian J Med Sci 1999; 53:535–544. Kejariwal D, Sarkar N, Chakraborti SK, Agarwal V, Roy S. Pyrexia of unknown origin: a prospective study of 100 cases. J Postgrad Med 2001; 47:104–107. Vanderschueren S, Knockaert D, Adriaessens T, Demey W, Durnez A, Blockmans D, Bobbaers H. From prolonged febrile illness to fever of unknown origin: the challenge continues. Arch Intern Med 2003; 163:1033–1041. Zhiyong Z, Bingjun L, Xiaoju L, Xinjian F, Ping F, Wenya W. Fever of unknown origin: a report from China of 208 cases. Int J Clin Pract 2003; 57: 592–596. Zamir D, Leibovitz I, Polychuck I, Reitblat T, Weiler Z, Zamir C. Fever of unknown origin in Israel. Acta Clin Belg 2003; 58:356–359. Tabak F, Mert A, Celik AD, Ozaras R, Altiparmak MR, Ozturk R, Aktuglu Y. Fever of unknown origin in Turkey. Infection 2003; 31:417–420. Liu KS, Sheng WH, Chen YC, Chang SC, Hsieh WC. Fever of unknown origin: a retrospective study of 78 adult patients in Taiwan. J Microbiol Immunol Infect 2003; 36:243–247. Saltoglu N, Tasova Y, Midikli D, Aksu HS, Sanli A, Dundar IH. Fever of unknown origin in Turkey: evaluation of 87 cases during a nine-year period of study. J Infect 2004; 48:81–85. Wong SY, Lam MS. Pyrexia of unknown origin – approach to management. Singapore Med J 1995; 48:204–208. Jayaram BM, Kantharaj L, Gopinath R, Nagaraj BS. Pyrexia of unknown origin. Natl Med J India 1996; 9:28–31. Norman DC, Yoshikawa TT. Fever in the elderly. Infect Dis Clin North Am 1996; 10:93–99. Cunha BA. Fever of unknown origin. Infect Dis Clin North Am 1996; 10:111–127. Konecny P, Davidson RN. Pyrexia of unknown origin in the 1990s: time to redefine. Br J Hosp Med 1996; 56:21–24. Adhikari PM. Fever of unknown origin – past, present and future. Indian J Med Sci 1998; 52:333–340. Volker D. Fever of unknown origin. Nurse Pract Forum 1998; 9:70–76.

25 Norman DC. Fever in the elderly. Clin Infect Dis 2000; 31:148–151. 26 Davies GR, Finch RG. Fever of unknown origin. Clin Med 2001; 1:177–179. 27 Tal S, Guller V, Gurevich A, Levi S. Fever of unknown origin in the elderly. J Intern Med 2002; 252:295–304. 28 Mourad O, Palda V, Detsky AS. A comprehensive evidence-based approach to fever of unknown origin. Arch Intern Med 2003; 253:263–275. 29 Morgan J, Whelan L. Problem based learning: pyrexia of unknown origin? Accident Emerg Nurs 2003; 11:131–140. 30 Amin K, Kauffmann CA. Fever of unknown origin. A strategic approach to this diagnostic dilemma. Postgrad Med 2003; 114:69–75. 31 Roth AR, Basello GM. Approach to the adult patients with fever of unknown origin. Am Fam Phys 2003; 68:2223–2228. 32 Woolery WA, Franco FR. Fever of unknown origin: keys to determining the etiology in older patients. Geriatrics 2004; 59:41–45. 33 Wells GA, Shea B, O’Connell D, Peterson J, Welch V, Losas M, Tugwell P. The Newcastle–Ottawa Scale (NOS) for assessing the quality of nonrandomized studies in meta-analyses. www.ohri.ca/programs/ clinical_epidemiology/oxford.htm 34 Peters AM. Nuclear medicine imaging in fever of unknown origin. Q J Nucl Med 1999; 43:618–620. 35 Corstens FHM, van der Meer JWM. Nuclear medicine’s role in infection and inflammation. Lancet 1999; 354:765–770. 36 Kjaer A, Lebech AM. Diagnostic value of (111)In-granulocyte scintigraphy in patients with fever of unknown origin. J Nucl Med 2002; 43:140–144. 37 Lorenzen J, Buchert R, Bohuslavizki KH. Value of FDG PET in patients with fever of unknown origin. Nucl Med Commun 2001; 22:779–783. 38 Blockmans D, Knockaert D, Maes A, De Caestecker J, Stroobants S, Bobbaers H, Mortelmans L. Clinical value of [(18)F]fluoro-deoxyglucose positron emission tomography for patients with fever of unknown origin. Clin Infect Dis 2001; 32:191–196. 39 Meller J, Ivancevic V, Conrad M, Gratz S, Munz DL, Becker W. Clinical value of immunoscintigraphy in patients with fever of unknown origin. J Nucl Med 1998; 39:1248–1253. 40 Kjaer A, Lebech AM, Eigtved A, Hojgaard L. Fever of unknown origin: prospective comparison of diagnostic value of 18F-FDG PET and 111 In-granulocyte scintigraphy. Eur J Nucl Med Mol Imaging 2004; 31: 622–626.

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Review paper

Fever of unknown origin, infection of subcutaneous devices, brain abscesses and endocarditis Giuseppe Lucio Cascinia, Diego De Palmab, Federica Matteuccic, Alberto Biggid, Pier Francesco Rambaldie, Alberto Signoref and Luigi Mansie The term ‘fever of unknown origin’ includes a wide range of conditions that often remain undiagnosed. The possibility of an infection must be promptly diagnosed in order to begin appropriate therapy. Imaging with radiopharmaceuticals, computed tomography, magnetic resonance imaging and ultrasound are the most commonly applied techniques, usually performed in addition to blood tests, biopsies or tissue cultures when required. The lack of comparative studies investigating the accuracy of each radiopharmaceutical for the study of fever of unknown origin was the incentive to perform a meta-analysis of peer articles published between 1981 and 2004 (33 papers) describing the use of nuclear medicine imaging for this purpose. Furthermore, infection of subcutaneous devices, brain abscesses and endocarditis must be considered amongst the causes of fevers of unknown origin. Reviews of 23, 10 and 10 papers, respectively (from 1976 to 2005), were performed on these specific topics. The results may be a useful guide for the choice of the optimal

radiopharmaceutical(s) and diagnostic strategy to be applied in each clinical condition and for different c 2006 Lippincott aims. Nucl Med Commun 27:213–222
Williams & Wilkins.

Fever of unknown origin

The prevalence of the diseases underlying FUO has changed over the last 50 years. For instance, systemic lupus erythematosus, a common cause in the past, is now more easily diagnosed by serological tests and rarely appears as FUO. Many diseases that previously caused FUO no longer do so because of the dramatic improvement in diagnostic imaging in recent years.

Introduction

The term ‘fever of unknown origin’ (FUO) is used to define a fever of more than 38.31C, lasting for longer than 3 weeks, with no established diagnosis despite appropriate investigations for 1 week. Clinical and anamnestic aspects, such as previous surgical intervention and trauma, are not listed in this definition, therefore not permitting a clear distinction to be made between nonlocated highly suspected infections (occult infections) and FUO. Many different diseases may cause FUO. Infections, neoplasms and inflammatory non-infective diseases, with particular regard to connective disorders and granulomatous diseases, are most prevalent. Other conditions, defined as miscellaneous disorders, including drug-related fevers, factitious fevers and habitual hyperthermia, are minor categories, but should not be underestimated. A variable number of patients affected by FUO still remain undiagnosed after an intensive diagnostic work-up. Moreover, the incidence of the previously described aetiopathogenetic category is extremely variable and is dependent on factors such as the patient’s age, geographical and economic conditions, the suitability of the diagnostic approach and the immune response (fever in immunocompromised patients is more likely to be due to tumours or infectious causes).

Nuclear Medicine Communications 2006, 27:213–222 Keywords: FDG-PET scan, fever of unknown origin, gallium scan, leucocyte scintigraphy, occult infection a

Nuclear Medicine, University Magna Graecia, Catanzaro, bNuclear Medicine, Ospedale di Circolo, Varese, cNuclear Medicine, AUSL Ravenna, P.O. Faenza, d Nuclear Medicine Service, S. Croce General Hospital, Cuneo, eNuclear Medicine and Centro di Eccellenza in Malattie Cardiovascolari, Second University of Naples, Naples and fNuclear Medicine, 2nd Faculty of Medicine, University ‘La Sapienza’, Rome, Italy. Correspondence to Luigi Mansi, Centro di Eccellenza in Malattie Cardiovascolari, Second University of Naples, Naples 80135, Italy. Tel: + 39-081-5665190; fax: + 39-081-5665202; e-mail: [email protected] Received 23 June 2005 Accepted 28 September 2005

The causes of FUO differ between various patient groups. For example, self-limited viral syndromes are an uncommon cause of FUO in older patients, whereas temporal arteritis, tumours and tuberculosis are more likely causes in older patients than in younger patients. Furthermore, infections are commonly detected in young patients, but seldom present as a cause of FUO in patients over 65 years. Similarly, the percentage of patients with undiagnosed fever after an intensive work-up varies with age (from 40% in paediatric patients to 8% in older patients) [1]. In addition to the prevalence of different causes underlying FUO, extensive diagnostic work-up should be directed by the clinician on the basis of potential diagnostic clues. The search for clues is crucial in establishing a diagnosis in patients with FUO, although, in many cases, they may be misleading [2]. The

c 2006 Lippincott Williams & Wilkins 0143-3636


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214 Nuclear Medicine Communications 2006, Vol 27 No 3

diagnostic strategy in establishing the cause of FUO is based on a multi-level approach, focused on the clues or abnormalities observed. The first step usually includes a complete blood cell count, routine chemistry tests, urinalysis, urine culture, blood cultures and a chest radiograph. Subsequent work-up should be guided by abnormalities noted on the physical examination, the age of the patient, anamnesis and results of the initial laboratory tests. Several imaging procedures have been proven to be useful in defining the cause of a fever. Abdominal ultrasound is a low-cost test that can detect abnormalities of the hepatobiliary and genitourinary systems, as well as fluid collections elsewhere in the abdomen. Computed tomography (CT) of the chest, abdomen and pelvis is helpful in patients with abscesses, haematomas, malignancies and lymphadenopathies related either to lymphoma or to granulomatous diseases. The findings obtained by ultrasound and CT can be non-conclusive, frequently requiring a subsequent biopsy or aspiration for a definitive diagnosis. These imaging procedures should be part of the routine work-up for patients with suspected FUO [3]. In patients with a diagnosis of FUO not well established after the initial workup, more detailed second-level examinations are required to search for a cause and to explain the abnormalities found in the previous tests, with particular regard to repeated blood cultures, biopsies and echocardiography, in addition to other nuclear medicine procedures. The role of nuclear medicine

Different nuclear medicine methods and agents have demonstrated a potential role in patients with FUO as integrative and second-line diagnostic procedures. In particular, labelled leucocyte (white blood cell, WBC) scan, 67Ga-citrate (67Ga), human polyclonal immunoglobulins (HIGs), labelled monoclonal antibodies (MoAbs) and [18F]fluorodeoxyglucose (FDG) have been widely tested, achieving unequivocal results in this challenging setting. The choice of a radionuclide procedure should be guided by the pretest likelihood of detecting an inflammatory or tumour cause of FUO. Moreover, nuclear medicine can be introduced into the diagnostic work-up when no clues are found or when a clue is of no use in the identification of the cause. Whole body imaging is suitable for the localization of a potential cause, which should be successively investigated by ultrasound, CT, magnetic resonance imaging (MRI), endoscopy and biopsy. To reach this goal, non-specific radiocompounds able to detect a wide disease spectrum are preferred. The use of WBC scintigraphy is based on the accumulation of leucocytes at the inflammatory site, which is regulated by adhesion molecules expressed on the surface of the vascular endothelium. WBCs can be labelled with

both 111In and 99mTc. Although 111In-WBCs can permit scanning up to 24 h after administration, which is helpful in detecting low-activity infections, at present 99mTcWBCs are the most commonly used radiolabelled form. WBC scan is widely used to detect acute inflammatory processes with an intense neutrophilic infiltrate, as in pyogenic infections, because the predominantly labelled cell population is made up of neutrophils. Unfortunately, most causes of FUO, also infective, have a monocytic or lymphocytic infiltrate; consequently, WBC scan is not expected to show significant uptake or diagnostic results in many of these patients. Therefore, WBC scintigraphy has been demonstrated to be effective in several series of FUO patients only when an infection, especially pyogenic, is assumed [4,5]. Thus, a high diagnostic accuracy has been reported in occult infections rather than in classical FUO [6,7]. Although inadequate results of WBC scintigraphy have been reported in many series of FUO patients, infection still remains the most important cause, and its detection represents a crucial point in the work-up of patients with prolonged fever; in particular, the negative predictive value of WBC scan may be effective in ruling out an infectious cause of FUO. HIGs, labelled with 111In or 99mTc, were introduced for the imaging of inflammation on the basis of their binding to Fc receptors at the site of inflammation. The specific uptake mechanism encouraged some authors to apply HIGs in patients with infection and/or FUO. Although HIGs are mainly a marker of increased permeability and are non-specifically accumulated in the interstitial space, they demonstrate almost the same diagnostic accuracy as the WBC scan [8,9]. Many labelled MoAbs have been proposed to detect infection and inflammation. 99mTc-labelled MoAbs (BW 250/183), Fab fragments and, more recently, antibodies against granulocyte antigens have been used to label neutrophils at the level of the inflammatory site. The uptake mechanism is due to MoAb-specific binding and non-specific accumulation in the interstitial space. One of the main disadvantages in their clinical use is the production of human anti-mouse antibodies, which seems to be dose dependent. The diagnostic accuracy of MoAbs has been demonstrated to be high, with an overall specificity close to 95% in infective patients and a high sensitivity due to their capability to detect more pathological sites than WBC scan, because the uptake mechanism also depends on the vascular permeability. 67

Ga is accumulated in abnormal areas by different nonspecific mechanisms, including increased vascular permeability and binding to extravascular transferring receptors.

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Fever of unknown origin Cascini et al. 215

67

Ga is still the most widely used agent to identify a cause in FUO, because of its satisfactory accuracy, low cost and wide availability. Different pathological conditions, such as abscesses, bacterial pneumonias, granulomatous diseases and neoplasms, may be detected. The diagnostic disadvantage in abdominal disease reported in several series is generally reduced by single photon emission tomography (SPET) and patient preparation. 67Ga is often helpful in patients with prolonged fever, especially if the preliminary work-up has not drawn attention to a specific disease. Its diagnostic efficacy is generally high in patients affected by FUO or occult infections, in which the fever has lasted for more than 2 weeks. Therefore, 67 Ga scan, which can identify subacute or chronic infections in addition to some inflammatory non-infective diseases and tumours, should be considered, at present, as the first-line diagnostic method in the nuclear medicine scenario, awaiting a wider availability and more extensive experience with FDG. FDG has been successfully used in the management of a variety of malignancies and, today, is a major tool in oncology. Multiple reports have shown that lesions with a high concentration of inflammatory cells, such as neutrophils and activated macrophages, may be visualized during a PET scan. Autoradiographic studies have demonstrated that the granulation tissue surrounding the tumour can present a significant FDG uptake. Although the inflammatory FDG uptake represents a disadvantage in nuclear oncology, it becomes a diagnostic benefit in the FUO setting. The percentage of FDG PET scans that are helpful in the diagnostic process in patients with FUO, as reported in the literature, varies from 41% to 69% [10]. Emerging data on FDG PET sensitivity, not only in the diagnosis of malignancies, but also in the detection of infective and inflammatory diseases, strongly support its prominent role in the management of FUO patients in the near future. Meta-analysis

Thirty-three series of patients with FUO (published from 1981 to April 2005), evaluated with nuclear medicine procedures, were analysed. Some reports were excluded because they were not directly focused on the FUO problem. Furthermore, difficulties in evaluation were encountered when analysing many of the selected papers; for example, patients with prolonged fever were frequently enrolled without excluding underlying diseases, previous interventions or trauma. Moreover, a final diagnosis was not always available and, in some cases, scintigraphic abnormalities were only assumed to be correct on the basis of the clinical evolution. The major question concerns which end-point should be used to evaluate the diagnostic accuracy of a scintigraphic procedure in this diagnostic scenario. Other problems were derived from the very low percentage of prospective studies in the evaluated papers and from the large

variability in defining the level of examinations during work-up. The comparison between nuclear medicine and standard imaging (ultrasound or CT) was not always reported and mainly in non-topical papers. Starting from these premises, 14 of the 33 series examined in the metaanalysis [4–9,11–31] were classified as occult infection, and 19 were defined as FUO. The meta-analysis revealed a high diagnostic accuracy for 99m Tc-WBC scan in patients with FUO (Table 1). These results, obtained exclusively in one paper [18], seem to differ from the diagnostic performance achieved using 111 In-WBCs, especially in terms of sensitivity. Data analysis demonstrated a good overall accuracy for all nuclear medicine methods, suggesting a second-line role in the diagnostic work-up of FUO patients. Moreover, specific agents, such as 111In-WBCs and 99mTc-MoAbs, showed a higher specificity than 67Ga and FDG, which were affected by a lack of specificity. A high rate of false negative results was observed for all procedures used in the FUO group, probably due to a non-infective end-point. Conversely, the overall accuracy was increased with both specific and non-specific agents in patients with occult infections (Table 2). In this setting, 111In-WBCs and FDG PET should be considered the procedures of choice, in particular due to the high negative predictive value. The comparison between radiological techniques and nuclear medicine is not reported, because both procedures were tested in only a few case reports. Clinical indications

In many cases, the choice of nuclear medicine technique in the infective disease scenario represents a dilemma, in which the economic context and the technological availability play an important role.

Table 1 Results of meta-analysis concerning patients with fever of unknown origin Procedure 111

In-WBCs Tc-WBCs 99m Tc-MoAbs 67 Ga FDG PET 99m

No. of patients

Se (%)

Sp (%)

224 26 115 76 168

63 90 66 60 78

87 87 92 70 59

PPV (%) NPV (%) 75 83 88 71 69

73 93 74 62 69

Acc (%) 71 88 78 61 77

Acc, accuracy; FDG PET, [18F]fluorodeoxyglucose positron emission tomography; MoAb, monoclonal antibody against granulocyte antigens; NPV, negative predictive value; PPV, positive predictive value; Se, sensitivity; Sp, specificity; WBC, white blood cell. All parameters have been weighted for the number of patients in each study. Numbers in bold refer to the best performing imaging tool.

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216 Nuclear Medicine Communications 2006, Vol 27 No 3

Table 2

Results of meta-analysis concerning patients with occult

infection Procedure 111

In-WBCs Tc-WBCs In-HIGs 99m Tc-HIGs 67 Ga FDG PET 99m 111

No. of patients

Se (%)

Sp (%)

492 100 110 75 310 112

86 89 82 55 87 95

92 94 89 85 81 88

PPV (%) NPV (%) 88 92 82 58 59 91

95 90 87 84 96 95

Acc (%) 93 92 85 77 85 92

Acc, accuracy; FDG PET, [18F]fluorodeoxyglucose positron emission tomography; HIG, human immunoglobulin; NPV, negative predictive value; PPV, positive predictive value; Se, sensitivity; Sp, specificity; WBC, white blood cell. All parameters have been weighted for the number of patients in each study. Numbers in bold refer to the best performing imaging tool.

In a recent review focused on the effectiveness of FDG PET in the inflammatory setting, the authors made some suggestions:

specific agents. 67Ga, FDG and WBCs revealed a high negative predictive value, strictly required to rule out an infective cause of fever. A second-line role was also claimed in paediatric, neutropenic and human immunodeficiency virus-positive patients, in whom infections are prominent.

Infection of subcutaneous devices Introduction

Infection of subcutaneous devices must be considered as a cause of FUO. These devices are increasingly being used in the treatment of patients affected by dangerous arrhythmias, disturbances of intracardiac electrophysiological conduction, central pain syndromes or under continuous infusion therapy. For example, it is estimated that every year in the USA 30 000–60 000 new pacemakers are implanted.

(1) infection may be assessed by FDG PET, but evidence to support the routine use of the technique is lacking; (2) FDG PET should be limited to patients with FUO and a low probability of infection; (3) in patients with FUO and a high probability of infection, WBC scan is more accurate [10,32].

The number and incidence of complications increase with the increase in use, and include infections of both bodies and wires [33–38]. Such infections are mainly due to bacteraemia and their incidence is reported to be in the range 1–9% [39,40]. They are extremely dangerous, with an associated morbidity and mortality [41], often due to the difficulty in removing the device, which is nevertheless mandatory.

On the basis of these suggestions, we have distinguished between patients with FUO and those with occult infection.

There are no guidelines on this subject provided by any cardiological or anaesthesiologist’s scientific society.

Patients with FUO

The diagnostic approach to patients with FUO is strictly dependent on symptoms. In patients without clues after a negative first-level work-up, three main approaches should be considered: wait and observe, whole body scintigraphy and therapeutic trials. The choice of a specific second-line strategy is generally defined on the basis of the clinical and economic scenario rather than technological availability. In particular, the choice of infective-specific scintigraphy (WBCs and MoAbs) should be made when an infective cause for FUO is supposed. Although the results of meta-analysis did not demonstrate a significant difference between specific and non-specific agents, in clinical practice, 67Ga and, more recently, FDG scan, when available, should be preferred.

In clinical cardiology, the normal flow chart requires clinical examination, cultural samples of the skin pocket and ultrasound by a transthoracic or transoesophageal approach to look for vegetations on the wires. In a recent paper, however, this approach was found to be effective in only 67% of patients [42]. The clinical/cultural approach is also mandatory for other kinds of devices, in which experience is more limited, and there is no agreement about the evaluation of parts in proximity to or entering the central nervous system [36,37]. Today, many radionuclide imaging techniques are available to evaluate the presence of infection of subcutaneous devices; in the literature, they are presented mainly as case reports, using as tracers 67Ga or labelled WBCs [43–53].

A fever with a duration of less than 3 weeks, or arising after trauma or intervention, is a fruitful field to apply nuclear medicine techniques.

In 1994, an interesting paper was published about the use of monoclonal anti-granulocyte antibodies (AGABs) in identifying the intracardiac vegetations in endocarditis, but this did not include the diagnosis of infected intracardiac electrode wires [54].

The results of meta-analysis demonstrated a high overall accuracy in this setting for both WBC scan and non-

Radionuclide imaging techniques seem to have no major role in studying infected subcutaneous devices, and

Patients with occult infection

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Fever of unknown origin Cascini et al. 217

Results of meta-analysis concerning infection of subcutaneous devices

Table 3

Scan type 67

Lesions

Se (%)

Sp (%)

Acc (%)

3 11

100 36.4

NA 100

100 36.4

Ga Tc-WBCs

99m

PPV (%) NPV (%) 100 100

NA 0

Acc, accuracy; NA, non-available data; NPV, negative predictive value; PPV, positive predictive value; Se, sensitivity; Sp, specificity; WBC, white blood cell. All parameters have been weighted for the number of patients in each study. Numbers in bold refer to the best performing imaging tool.

preliminary experience exists only for peritoneal dialysis catheters [55]. Recent papers have reported a high incidence of dubious diagnoses of infection of the skin pocket [56,57], where germs may stay nested without clinical signs for long periods of time and then cause a later infection. There are as yet no papers reporting the usefulness of nuclear medicine in this field. Meta-analysis

Twenty-three original papers and four abstracts referring to the imaging of the infection of subcutaneous devices were published in the period between 1985 and 2005, and are reviewed [33–59]. The main end-point emerging from an analysis of these scientific papers is that only seldom is radionuclide imaging helpful (Table 3). Moreover, especially in the case of pain palliation devices, the problem of occult infection is increasing because of the increasing number of implants. It was impossible to obtain data on the sensitivity and specificity from most papers because of the small number of cases considered. Indications 67

Ga scan is useful in the case of dubious (i.e. clinically non-evident) residual infection of the skin pocket, especially if surgical removal of the device is in question. Nevertheless, it should be noted that the results of this meta-analysis were limited by the small number of publications and patients available. Other radiological and clinical examinations must be considered in the evaluation of infected subcutaneous devices.

Brain abscesses Introduction

Brain abscesses usually manifest with subacute progression of focal neurological signs, altered mental status and headache, with mild or absent signs of meningeal irritation. The clinical differential diagnosis of brain abscesses remains difficult and includes, amongst other conditions, brain tumours. Generally, brain CT will depict a contrastenhancing intracerebral mass lesion; however, cystic tumours or infarcts with rims due to neovascularization may give a similar ring-enhancing appearance on contrast

CT. Moreover, in the pre-encapsulated stage of abscess evolution, CT findings may be atypical and indistinguishable from brain oedema due to any cause. Therefore, additional diagnostic modalities are required to establish more precisely the nature of the lesion, and also to acquire reliable information to determine whether or not to operate on the patient and which type of surgery to perform. Nowadays, nuclear magnetic resonance (NMR) is the most useful and most commonly used diagnostic modality; nevertheless, it can be important to combine it with nuclear medicine imaging in non-diagnostic cases. The aim of this meta-analysis was to provide indications on what nuclear medicine diagnostic tool should be used together with NMR. The role of nuclear medicine

Over the past two decades, several studies have been published that have addressed the role of WBC scintigraphy for the purpose of differentiating cerebral abscesses. Irrespective of the type of WBC labelling, the overall reported accuracy obtained in these series is in the range 96–100%. The accuracy of WBC scintigraphy appears to be related to three major variables. (1) The use of a grading system and the selection of the reference region to classify the lesions as either positive or negative: a moderate uptake of labelled leucocytes, normally weaker with respect to abscesses, may be observed in tumours, consistent with the presence of small amounts of leucocytes in neoplastic lesions. (2) The location of the lesion: lesions located next to the base of the skull may be very difficult to detect and characterize compared with lesions positioned in the parenchyma or next to the calvarium. (3) The type of medical treatment received by patients at the time of the WBC study, with main reference to the administration of steroids. Inconsistent findings have been reported in patients submitted to treatment with steroids. It should be noted that, although steroid administration significantly reduces the degree of WBC accumulation, slight activity remains and may be detected. The Editorial, ‘The role of nuclear medicine imaging in routine assessment of infectious brain pathology’, describes how WBC scintigraphy needs to be analysed for a reliable differential diagnosis of brain abscesses [60]. Meta-analysis

Ten papers on the imaging of brain abscesses (published from 1985 to the present) were analysed [61–70]. In Table 4, the accuracy of nuclear medicine techniques is reported. In the large majority of cases, the gold standard

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218 Nuclear Medicine Communications 2006, Vol 27 No 3

Table 4

Results of meta-analysis concerning brain abscesses

Tracer

No. of scans

Se (%)

Sp (%)

Acc (%)

159 102 26

93 100 100

97 100 83

98 100 NA

111

In-WBCs Tc-WBCs Tc-MoAbs

99m 99m

Acc, accuracy; MoAb, monoclonal antibody against granulocyte antigens; NA, non-available data; Se, sensitivity; Sp, specificity; WBC, white blood cell.

was considered to be evidence of pus at the time of surgery, or on microscopic examination of the surgical specimens when the pus was not visually evident at the intervention. Clinical indications for WBC scan

The main clinical indication for WBC scintigraphy is the differential diagnosis of intracerebral cystic lesions, hypodense at CT and non-conclusive at MRI. In assessing the post-operative course of patients treated for brain abscesses, WBC scintigraphy may be useful in follow-up, in association with CT. Recurrent brain abscesses appear as a hypodense area at CT, which is not easily distinguishable from ischaemia, oedema or post-surgical sequelae. Conversely, an intense uptake of WBCs strongly supports the diagnosis of recurrent abscess.

Endocarditis Introduction

Infective endocarditis represents a severe event requiring early diagnosis and adequate therapy. It is commonly classified into acute and subacute-chronic endocarditis on the basis of the severity of clinical presentation and progression of the untreated disease. The characteristic lesion, a vegetation made up of fibrin, platelets, microorganisms and inflammatory cells, involves, most frequently, both native and prosthetic heart valves, and is also found on the septum, tendinous chords or mural endocardium. The epidemiological features have changed, mainly as a consequence of the increasing longevity of patients, the larger number of patients undergoing valvular surgical replacements and drug addicts; currently, in the USA and Western Europe, the incidence of the disease is 1.7–6.2 cases per 100 000 inhabitants per year [71,72]; men are more frequently affected than women (male-to-female ratio, 1.7 : 1) in the age range between 47 and 69 years [72,73]. In drug addiction by intravenous injection, the estimated incidence reaches 150–2000 cases per 100 000 inhabitants per year [74]. The most common agent causing infective endocarditis is Staphylococcus aureus. In patients presenting with early prosthetic valve endocarditis, the infection is frequently supported by coagulase-negative staphylococcus [75].

The clinical onset of infective endocarditis frequently includes extracardiac manifestations or findings associated with the intracardiac extension of the infection; the main sign is fever, sometimes absent or minimal in patients with congestive heart failure or chronic renal or liver failure. Other common symptoms are anorexia and weight loss. The large majority of patients with infective endocarditis present with a heart murmur (most commonly pre-existing), petechiae on the skin, conjunctiva or oral mucosa and splenomegaly. The role of nuclear medicine

The diagnosis of infective endocarditis requires, as a first step, the integration of clinical, laboratory and echocardiographic data, which also play a fundamental role in follow-up. The Duke University criteria for assessing patients with suspected infective endocarditis, proposed in 1994 [76], still remain a reference in the field today because of the very high specificity and negative predictive value, up to 99% and 92%, respectively, demonstrated in several studies [77,78]. Transthoracic echocardiography (TTE) is a rapid and non-invasive method showing a high specificity for the diagnosis of vegetations (up to 98%), but is affected by an unsatisfactory sensitivity (60–70%) [79]. This procedure may be inadequate in up to 20% of adults because of obesity, chronic obstructive pulmonary disease and chestwall deformations. Using transoesophageal echocardiography (TEE), a higher sensitivity in detecting vegetations, with no loss in specificity (75–95% and 85–98%, respectively) relative to TTE [80,81], with a negative predictive value up to 92%, is obtained [78]. Although this procedure is high cost and invasive, it can be helpful in patients with prosthetic valves and in the evaluation of myocardial involvement. A major problem for both TTE and TEE is the unreliability in differentiating florid active vegetations from post-therapeutic outcomes at the level of previously affected valves. This is an important clinical challenge, as more than 60% of vegetations remain morphologically unchanged despite successful antibiotic therapy. At present, nuclear medicine procedures are not considered in the diagnostic flow chart [82], although several reports support a clinical role, mainly in cases uncertain at echocardiography. In the diagnosis of infective endocarditis, many radiocompounds, such as 67Ga, WBCs radiolabelled with 111In or 99mTc and 99mTc-AGABs, have been proposed. The possible role of FDG PET, as a result of its ability to diagnose inflammatory and/or infective diseases [83–86], is under evaluation. In a recent paper

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Fever of unknown origin Cascini et al. 219

[87], FDG PET demonstrated a satisfactory accuracy, comparable to that obtained by echocardiography, in detecting endocarditis. The physiological uptake by the left ventricle does not represent a disadvantage as it does not interfere with the detection of endocarditis. However, in the presence of active disease, an easier diagnosis, due to better localization of the pathological site, is obtained.

111

In-WBCs present some advantages with respect to Tc-WBCs. Because of the possibility of more delayed scans, when a lower blood pool activity is present, a more favourable lesion-to-background ratio can be achieved. Nevertheless, recent case reports encourage the use of 99m Tc-WBCs in the diagnosis of active infective endocarditis, mainly because of the higher count rate achievable, allowing a more accurate SPET acquisition at an early period. 99m

Meta-analysis

To evaluate the possible role of nuclear medicine in endocarditis, 10 original papers and two reviews, published in the period 1976–2005 and referring to a total of 378 patients (380 lesions), were considered (Table 5) [9,54,88–97]. At the meta-analysis, TTE showed a lower sensitivity and specificity than TEE, with a better accuracy and positive predictive value when both procedures were performed together. 67Ga was affected by a low sensitivity (45%), but its uptake strongly suggested the presence of endocarditis. These data had to be evaluated carefully, because they were obtained from old studies containing a small number of patients and from scans partially performed with obsolete technologies. Some evidence suggests that better results could be obtained using more recent instruments and a more accurate methodology, including the use of all three photo-peaks of 67Ga, a wider use of SPET acquisitions and more images obtained with delayed scans beyond 48 h post-injection, when a lower blood pool activity remains.

A very high accuracy, both sensitivity (up to 100%) and specificity (up to 86%), can be attained when 99mTcAGABs are used in association with echocardiography. Echocardiography is indispensable in evaluating morphological and functional changes in affected cardiac valves, whereas 99mTc-AGABs seem to better assess the activity of the underlying inflammatory process. As described previously, a limitation for this procedure is the possible production of human anti-mouse antibodies, measurable in 4.5–30% of patients undergoing repeated injections.

Acknowledgements

99m

Tc-pyrophosphate has been shown to accumulate in experimentally induced infective endocarditis. In clinical practice, as reported in the only study published, no pathological uptake was detected in 28 patients with a diagnosis of endocarditis of the native valve.

At scintigraphy with 111In-labelled WBCs, a higher sensitivity (67%) than TTE and a higher specificity (95%) than TEE have been reported. These data must be further confirmed, however, because they were derived from studies performed in a small series of patients. It is interesting to note that, in the diagnosis of endocarditis, Table 5

Very promising results have been obtained using immunoscintigraphy with 99mTc-AGABs. In an analysis of five papers, comparing this procedure with TTE and TEE in 194 lesions, 99mTc-AGABs showed a higher sensitivity (65%) than TTE and a better specificity (86%) than TTE (62%) and TEE (76%), comparable with that obtained when both ultrasound procedures were combined.

This work is part of a large multi-centre study, conducted by the Italian Study Group on Inflammation–Infection Imaging of the Italian Society of Nuclear Medicine (AIMN), co-ordinated by Dr Alberto Signore. Members of the group include: Marco Agnolucci, Alessio Annovazzi, Giorgio Ascoli, Carla Augeri, Bruno Bagni, Marilena Bello`, Sergio Bissoli, Nicola Boccuni, Sergio Boemi, Paolo Braggio, Luca Burroni, Dario Cantalupi, Gabriela Capriotti, Giuseppe Cascini, Marco Chianelli, Arturo Chiti, Micaela D’Alberto, Diego De Palma, Giovanni D’Errico, Narcisa De Vincentis, Salvatore di

Results of meta-analysis concerning infective endocarditis

Procedure TTE + TEE TTE TEE 99m Tc-AGABs AGABs + echo 67 Ga 99m Tc-Pyr 111 In-WBCs

Lesions

SE

SP

Lesions

ACC

Lesions

PPV

NPV

166 102 21 194 160 70 31 30

88.0 44.2 100.0 65.3 100.0 45.5 0.0 67.0

86.6 62.3 76.0 86.0 85.7 95.4 0.0 95.0

136 104 21 194 160 70 31 30

90.4 66.0 81.0 79.9 92.2 52.9 0.0 86.0

166 104 21 194 160 42 31 30

78.0 65.5 64.0 83.6 88.8 89.0 0.0 86.0

88.8 57.4 100.0 80.1 100.0 82.8 0.0 86.0

Acc, accuracy; AGABs, anti-granulocyte antibodies; echo, echocardiography; NPV, negative predictive value; PPV, positive predictive value; Pyr, pyrophosphate; Se, sensitivity; Sp, specificity; TEE, transoesophageal echocardiography; TTE, transthoracic echocardiography; WBC, white blood cell. All parameters have been weighted for the number of patients in each study. Numbers in bold refer to the best performing imaging tool.

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220 Nuclear Medicine Communications 2006, Vol 27 No 3

Rosa, Paola Erba, Antonio Ferrarese, Guido Ferretti, Chiara Gallini, Elena Lazzeri, Lorenzo Maffioli, Giulia Manfredini, Rita Mannino, Luigi Mansi, Giuliano Mariani, Mario Marinelli, Pietro Marinelli, Luigi Martino, Federica Matteucci, Marino Mele, Angelo Mita, Monica Mori, Maria Gemma Parisella, Valentina Picardi, Carlo Poti, Michele Povolata, Napoleone Prandini, Pierfrancesco Rambaldi, Brunella Rossi, Domenico Rubello, Vittoria Rufini, Orazio Schillaci, Alberto Signore, Alberto Spina, Luca Tagliabue, Maria Cristina Tappa, Daniela Turrin, Venanzio Valenza, Anna Viglietti and Alberto Vignati. Our gratitude and acknowledgements extend to all members for their collaboration and helpful discussions in the preparation and progress of this study.

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Review paper

Clinical applications of 188Re-labelled radiopharmaceuticals for radionuclide therapy Bieke Lamberta and John M.H. de Klerkb 188

Re is a radionuclide in which there is widespread interest for therapeutic purposes because of its favourable physical characteristics. Moreover, it can be eluted from an on-site installable 188W/188Re generator, which has a useful shelf-life of several months. Most of the clinical experiences gained with 188Re concern the use of 188 Re-1,1-hydroxyethylidenediphosphonate (188Re-HEDP) for bone pain palliation in patients suffering prostate cancer. The maximum tolerated activity was 3.3 GBq 188 Re-HEDP and if the platelet count exceeded 200 ¾109 l – 1, the administration of 4.4 GBq appeared safe. Evidence for repeated administrations of 188Re-HEDP rather than single injections was established. In general, pain palliation occurs in 60–92% of patients with only moderate transient toxicity, mainly related to changes in blood counts. Also in haematology, radioimmunotherapy by means of 188Re might play a role by selectively targeting the bone marrow in patients undergoing conditioning prior to haematopoetic stem cell transplantation. The feasibility of such an approach was proven using a 188Re-labelled monoclonal antibody directed toward the CD66-antigen. More recently, encouraging safety data on locoregional treatment of primary liver tumours using 188Re-labelled lipiodol were reported. The normal organs at greatest risk for toxicity are

the normal liver and the lungs. About 50% of the patients reported mild and transient side effects, mainly consisting of low grade fever, right hypochondrial discomfort or aggravation of pre-existing liver impairment. Besides the applications in oncology 188Re-based therapies have also been pioneered for benign condition such as prevention of re-stenosis following angioplasty and for radiosynovectomy in cases of refractory arthritis. c 2006 Lippincott Williams Nucl Med Commun 27:223–229
& Wilkins.

Introduction

Similar to the introduction of the in-house use of the 99 Mo/99mTc generator in a radiopharmacy for the radiolabelling of a wide variety of diagnostic agents, generatorderived therapeutic isotopes would contribute to the further development and implementation of radionuclide therapies. 188Re is eluted from a 188W/188Re generator, which has a long useful shelf-life of several months [1]. 188 Re is of widespread interest for therapeutic applications because of its high-energy beta emission (maximum energy 2.12 MeV). Its gamma emission of 155 keV, with an abundance of 15%, and relatively short half-life of 16.9 h, limit radioprotection problems and represent a firm advantage in comparison with the now widespread use of 131I for various therapeutic radiopharmaceuticals.

Besides its merits in the management of benign conditions, such as thyroid disease and chronic joint inflammation, radionuclide therapy has shown its usefulness in oncology, for instance in the management of thyroid cancer, metastatic neuro-endocrine tumours, bone pain palliation and locoregional treatment for primary liver cancer or liver metastasis. More recently, promising progress has been established in the field of haematology. However, radionuclide therapy remains a relatively unknown treatment modality for many colleagues, even those working in the fields of oncology and nuclear medicine. Despite the available literature underscoring the clinical benefits of radiolabelled diphosphonates, meta-iodobenzylguanidine, somatostatin analogues and antibodies, the application of these radiopharmaceuticals remains limited to a few expert centres in welldeveloped countries. Radioprotective concerns, limited availability and high costs of some radionuclides, as well as compounds such as monoclonal antibodies and peptides, have been a drawback for widespread implementation of radionuclide based treatment strategies.

Nuclear Medicine Communications 2006, 27:223–229 Keywords: rhenium-188, radionuclide therapy, radiopharmaceuticals a Division of Nuclear Medicine, Ghent University Hospital, Belgium and bDivision of Nuclear Medicine, Meander Medical Centre, Amersfoort, the Netherlands.

Correspondence to Dr Bieke Lambert, Division of Nuclear Medicine, Ghent University Hospital, De Pintelaan 185, B-9000 GENT, Belgium. Tel: + 0032 9 240 3028; fax: + 0032 9 240 3807; e-mail: [email protected] Sponsorship: Bieke Lambert is sponsored by a grant from the Bijzonder Onderzoeksfonds of the Ghent University (011D9501). Received 26 April 2004 Accepted 23 November 2005

188

Re in oncology

Bone pain palliation

The two radiopharmaceuticals, strontium-89 chloride and phosphorus-32 phosphate, which have been used in the last decades for treatment of bone metastasis, are based on relatively long-lived beta emitters, lacking suitable gamma emissions for high-resolution imaging. The more

c 2006 Lippincott Williams & Wilkins 0143-3636


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224 Nuclear Medicine Communications 2006, Vol 27 No 3

recently introduced 153Sm on the other hand, allows for imaging and has a shorter half-life. Since it is produced in a reactor, it is not readily available in certain areas. The beta emitter 186Re is also produced in a reactor but has a somewhat longer half-life than 153Sm. The encouraging results obtained with 186Re-labelled hydroxyethylidene diphosphonate (HEDP), inspired others to develop a 188 Re-labelled analogue [2–4]. Diphosphonates such as methylene diphosphonate (MDP) and hydroxymethane diphosphonate (HDP) are well known bone seeking agents applied for imaging with 99mTc, but labelling of HDP and MPD with 188Re does not yield satisfying uptake in the skeleton [5]. Significant higher skeletal uptake was obtained with HEDP labelled with 188Re [5]. It was shown that the total mass of rhenium in the radiolabelling procedure of HEDP with 188Re is essential for obtaining a satisfactory biodistribution comparable with that of 186Re-HEDP [6,7]. The initial clinical results with 188Re-labelled HEDP were established by the University of Bonn [8]. This group performed a subsequent study in which increasing activities of carrier added 188Re- HEDP were administered to a group of hormone refractory prostate cancer patients suffering multifocal bone metastasis. Only patients with a white blood cell count exceeding 3  109 l – 1, over 100  109 l – 1 platelets, a serum creatinine below 1.4 mgdl – 1 and without previous chemotherapy or external-beam hemibody irradiation were eligible. Pain relief was reported by 64% of patients with a mean duration of 7.5 weeks. Higher administered activities seemed to induce more frequent responses. The maximum tolerated activity was 3.3 GBq of carrier added 188Re-HEDP and in patients with baseline platelet counts exceeding 200 109 l – 1, the administration of 4.4 GBq appeared safe [9]. More recently, the same group established evidence for repeated administrations of 188Re-HEDP instead of a single injection. The effect of pain palliation after a single injection of 188Re-HEDP, which produced a response rate of 60%, could be increased to 92% after repeated injection 8 weeks later. The duration of pain relief was also prolonged from 2.6 after a single injection to 5.7 months after repeated injection. They observed that a second treatment with 188Re-HEDP can lead to effective pain therapy, even if the first injection did not induce pain relief. Moreover, significant reduction in the levels of the prostate specific antigen (PSA) were documented following repeated injections whereas this effect was not present in a single injection set-up [10,11]. Other groups have focused on the impact of this radionuclide therapy on the general status of the patient: a statistical significant increase in the Karnofsky performance score of 74 ± 7% before therapy to 85 ± 9% at 12 weeks after a single injection was observed [12]. Initial trials focused mainly on patients suffering prostate cancer and in some cases breast cancer patients were included. However, evidence is available that patients suffering

bone metastasis due to other malignancies such as lung, renal, rhinopharyngeal and bladder cancer would also benefit from 188Re-HEDP treatment [13]. In general, only moderate transient toxic effects, mainly related to changes in blood counts, were described. A flare of bone pain occurred in a minority of patients and does not seem to predict refractory bone pain. Li and coworkers applied repeated administrations of relatively low (1.1 GBq) activities of 188Re-HEDP and no haematological toxicity was observed. Within the first 48 h, a small subset of patients experienced low-grade fever, hidrosis, nausea, vomiting and joint pain. All complaints resolved spontaneously. It was shown that 188Re-HEDP treatment is feasible on an outpatient basis in many countries [6]. A 188Re(V)-dimercaptisuccinic acid (188Re(V)-DMSA) kit was developed by Blower et al. [14,15] for bone pain palliation and it was shown that 99mTc(V)-DMSA quantitatively predicts 188Re(V)-DMSA distribution in patients with prostate cancer metastatic to the bone. The clinical experience with 188Re(V)-DMSA is still limited and possibly this agent might rather play a role in treatment of soft tissues tumours or medullary thyroid cancer. In conclusion, encouraging results were reported for the use of 188Re-HEDP but various aspects of this treatment modality remain unclear. Some authors prefer the use of emitters with a low beta energy for treatment of painful bone metastasis, because it is assumed that such radionuclides would reduce bone marrow toxicity. However, little evidence for such a strategy is available. 188Re-HEDP and 153Sm-ethylenediamine-N,N,N0 ,N0 -tetrakis(methylene phosphonic acid) (153Sm-EDTMP), show similar pain palliation effect in patients suffering from breast and prostate cancer. There were no differences in bone marrow toxicity between the higher beta energy 188Re-HEDP and the lower beta energy 153Sm-EDTMP in a comparative study [16]. Other authors suggest enhanced antitumoral effects if high-energy beta emitters are used and propose aggressive treatment in an earlier disease stage instead of using these radiopharmaceuticals only in end-stage patients suffering intractable bone pain. The electrons emitted by 188Re have a range of 3–5 mm in osseous tissue and thus target tumour as well as surrounding bone trabeculae and periostium. This would lead to cytotoxic effects in the outer layers of the tumour and repeated treatment with short intervals would prevent new tumour growth and eradication of deeper tumour layers [10]. Another approach consists of including other treatment modalities such as autologous stem cell rescue and chemotherapy to a radionuclide treatment scheme [17]. Future research should focus on the potential antitumoral effects of such aggressive treatment strategies and their impact on quality of life as well as survival.

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Clinical applications of

So far, no studies have been carried out in patients with compromised bone marrow or impaired renal function. However, in daily practice, a considerable number of patients presenting with bone pain would not meet the inclusion criteria of the published phase I studies and it remains unclear whether they would benefit from this treatment. Additional research focussing on this subset of patients is needed. In addition, the efficacy of 188ReHEDP for bone metastasis originating from other malignancies then prostate and breast cancer should be investigated. Haematological malignancies

Haematopoietic stem cell transplantation (HSCT) was developed as a means to overcome otherwise lethal myeloablation in intensive treatment regimes. Eligible patients are often in partial or complete remission, thus presenting with a small tumour load. Standard conditioning regimens for HSCT consist of combinations of cytotoxic drugs with or without total body irradiation and often induce gastro-intestinal, pulmonary and renal toxicity. Higher total-body radiation doses tend to reduce the relapse rate but a potential survival benefit is offset by a steep rise in toxicity related deaths with increasing doses [18]. A potential role for radionuclide therapy may consist of selective bone marrow targeting by means of radiolabelled antibodies that bind to normal and eventually malignant haematopoietic cells but not to nonhaematopoietic cells. Among the potential targets, CD45 (pan leukocyte antigen), CD33 and CD66 (myeloid antigen) are of great interest in case of leukaemia. These antigens are present at high density on the cell surface. However, they are not tumour specific and may or may not be expressed by neoplastic cells. Since HSCT is mainly attempted in patients with limited tumour load, residual tumour cells are expected to be killed by cross-irradiation from normal cells in the marrow [18]. Most research with 188Re-labelled antibodies so far has been done by Kotzerke et al. [19]. Very interesting results were obtained using a 188Re-labelled monoclonal antibody directed toward the CD66-antigen (Antigranulocyte; Syntec Diagnostics, Zug, Switzerland). Only patients with favourable biodistribution following a 99mTc-Antigranulocyte scan were eligible for subsequent dosimetry following 1.2 GBq 188Re-labelled Antigranulocyte. It was shown that despite the availability of 99mTc-Antigranulocyte additional dosimetry by means of the 188Re-labelled antibody remained essential since the biological half-life and bone marrow uptake is lower for 188Re-labelled than for 99mTc-labelled Antigranulocyte. Patients without ‘pathological’ uptake suggesting inflammatory lesions and significant higher uptake in the bone marrow, compared to liver, lungs and kidneys were eligible for subsequent radioimmunotherapy. This was the case in all

188

Re for radionuclide therapy Lambert and de Klerk 225

assessed patients. Fifty patients with a high risk leukaemia for relapse following classic HSCT received about 10 GBq of the 188Re-labelled monoclonal antiCD66 and full dose conditioning including 12 Gy totalbody irradiation. The absorbed dose to the bone marrow following radioimmunotherapy was 13.9 ± 4.6 Gy. The kidney was the non-target organ receiving the highest dose (0.7 ± 0.2 GyGBq – 1). All 50 patients proceeded to the transplant phase of the study and showed primary engraftment. No increase in transplant related toxicity was observed and only mild adverse events, mainly consisting of nausea and stomatitis in the majority of patients after radionuclide therapy were reported [20]. In an earlier report of the same group, five out of 36 patients developed the clinical syndrome of bone marrow radiation nephropathy with hypertension, anaemia and raised serum creatinine. The serum creatinine was elevated in 17% of patients 6–12 months following transplantation. Further optimization of conditioning regimes should pay attention to the potential detrimental effects of combining particular cytotoxic drugs, external-beam total body irradiation and radioimmunotherapy [21]. Other authors have pointed out the risk of enhanced acute intestinal graft-versus-host disease and transplant related mortality following conditioning with 188Re-anti-CD66 monoclonal antibody. Possibly these events were related to differences in immunosuppressive schemes and the use of unmanipulated grafts instead of T-cell depleted grafts, as used by Bunjes and co-workers [22]. Solid tumours

Several antibodies have been successfully labelled with 188 Re. These direct labelling techniques do not yield identical in-vitro stability as the 99mTc-labelled counterparts but animal studies failed to show significant differences in renal clearance [23]. Juweid and coworkers [24] conducted a phase I ‘dose escalation’ study using 188Re-labelled anti-CEA monoclonal IgG (188ReMN-14) in patients suffering advanced gastro-intestinal cancer. Besides myelosuppression, no organ toxicity was observed and only activities exceeding 2.2 GBqm – 2 induced grade 3 thrombocytopenia. Compared to its 131 I-labelled counterpart, the renal uptake of 188Re-MN-14 was higher and its stability was unsatisfactory. In addition, the ideal molecular weight of the 188Re-labelled antibodies or antibody fragments is a debatable issue in respect to the relatively short physical half-life of 188Re. The use of low molecular weight fragments with fast tumour uptake is appealing but this advantage is counterbalanced by the higher renal accumulation of these fragments. Although various animal experiments on the subject were published, no recent clinical advances using 188Relabelled somatostatin analogues or other peptides were reported [25–29].

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226 Nuclear Medicine Communications 2006, Vol 27 No 3

In contrast to the only modest advances made in systemic 188 Re based radionuclide therapy for solid tumours, encouraging results for locoregional treatment of hepatocellular carcinoma (HCC) were reported. HCC is the most prevalent primary liver cancer [30]. It constitutes the third cause of cancer related deaths, responsible for more than 500 000 deaths worldwide annually [31]. Surgery, either by means of hepatectomy or liver transplantation, is the mainstay of curative treatment. Overall, if the degree of underlying liver dysfunction and the tumour load is taken into consideration, the vast majority of patients are not eligible for surgery. Among the few treatment modalities producing response rates exceeding 20% in inoperable patients are the use of Lipiodol mixed with chemotherapeutic agents or radiolabelled with 131I without subsequent embolization [32]. Encouraging results have been obtained using 131 I-Lipiodol, but the use of 131I has hampered its routine clinical implementation [33]. During the past 10 years, promising pre-clinical results using 188Re-labelled Lipiodol for treatment of HCC have been reported [34,35]. Among several synthesized compounds, the Lipiodol solution of 188Re-HDD (4-hexadecyl-TDD) had the most favourable tumour retention properties and dosimetric characteristics [36,37]. Only recently, however, were the first clinical results using 188Re-4hexadecyl-1-2,9,9-tetramethyl-4,7-diaza-1,10-decanethiol/ Lipiodol (188Re-HDD/Lipiodol) reported by Sundram and co-workers [38]. In a multi-centre study the safety of transarterial 188Re-HDD/Lipiodol for treatment of inoperable HCC was assessed. Sixteen patients were treated in a pilot study with administered activities ranging from 1.8 GBq to 7.7 GBq. The level of radioconjugate activity was based on radiation absorbed dose to critical organs, calculated following a ‘scout dose’ of radioconjugate. The normal organs at greatest risk for toxicity are the normal liver and the lungs. Due to arterio-venous shunting in the cirrhotic liver or tumour and the embolizing feature of Lipiodol, varying uptake in the lungs is observed. In a subsequent study a further 54 patients were treated with a mean administered activity of 4.6 GBq. About 50% of the patients reported mild and transient side effects, mainly consisting of low grade fever and right hypochondrial discomfort. Two patients developed pleural effusion [39]. Besides liver and lung uptake, post therapy scintigraphies showed a faint visualization of the kidneys and in a limited number of cases gastro-intestinal activity was reported [39,40]. A biodistribution study carried out by Lambert et al. found that a mean of 44% of administered 188Re-HDD is excreted in the urine within 76 h and that a small part of the tracer is likely to be cleared via the biliary tract [40]. Further treatment of larger numbers of patients, however, is required to assess the therapeutic efficacy of 188ReHDD/Lipiodol. Given the labelling yield of 188Re-HDD/Lipiodol ranges between 50 and 70%, which may pose a problem for the

synthesis of high therapeutic activities, a number of authors have focused on the development of new derivatives. These include 188Re-nitrido bis-(diethyldithiocarbamato) Lipiodol (188ReN-DEDC) and 188Re(S2CPh)(S3CPh)2 Lipiodol (188Re-SSS Lipiodol) [41]. While for the 188ReN-DEDC version feasibility was shown for intra-arterial treatment of unresectable HCC, either alone or in combination with transarterial chemoembolization, in a series of nine patients, data on 188ReSSS Lipiodol in HCC patients are currently lacking [42]. Other preclinical research focussed on rhenium-labelled glass microspheres or ethyl cyteinate dimer (ECD) extracted into the Lipiodol phase [43,44]. 188

Re for benign diseases

Intracoronary radionuclide therapy

Restenosis is a frequent complication following percutaneous transluminal coronary angioplasty and occurs within 6 months in 40–60% of all cases. Stent implantation reduces the re-stenosis rate by preventing constrictive remodelling and elastic recoil of the artery but fails to prevent neointimal hyperplasia. Various preclinical studies have shown that beta or gamma irradiation can inhibit neointimal proliferation and therefore help to prevent in-stent re-stenosis after angioplasty. In contrast to a radioactive wire, a balloon filled with a beta-emitting radionuclide provides a radiation field that conforms to the vessel geometry in an optimal fashion, irrespective of cardiac motion. The main disadvantage is the risk of balloon rupture. Therefore bone-seeking radionuclides such as 90Y and 32P should be avoided. In the case of a 188 Re-filled balloon, perchlorate blocking prior to the intervention or linking the radionuclide to urinary cleared chelating agent avoids accumulation of the radionuclide in the thyroid and gastric mucosa in the event of balloon rupture [45]. Ho¨her and co-workers reported that coronary irradiation with a 188Re-filled balloon is technically feasible and safe, requiring only standard percutaneous transluminal coronary angioplasty techniques [46]. Although encouraging initial experiences were reported, long-term safety and efficacy of intracoronary radiotherapy remains uncertain [46–48]. In a study conducted by Kim et al. [49] 187 patients were randomly assigned to receive either intracoronary beta irradiation with a 188Refilled balloon (n = 104) or to no additional treatment (n = 83) after successful catheter-based intervention for de-novo or re-stenotic lesions. At 9 months angiographic re-stenosis was significantly reduced with intracoronary irradiation (18.9% vs. 45.9%, P < 0.001), but the incidence of major adverse cardiac events by 3 years showed no significant difference. Lack of clinical benefit might be related to mismatched balloon positioning in respect to the lesion, balloon-induced unhealed dissection and late thrombosis. By refining the technique and postinterventional medication, these complications are avoid-

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Clinical applications of

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Re for radionuclide therapy Lambert and de Klerk 227

able and thus long-term outcomes could be further improved. The results of this study are difficult to interpret since the proportion of patients suffering restenosis was significantly higher in the irradiated group compared to the control patients and this difference might have affected late outcome.

study in which 188Re-labelled monoclonal antibodies targeting fungal antigens were shown to prolong survival of infected mice [53]. Once more the physical half-life of 188 Re might be too short for combination with high molecular weight antibodies, especially if abscesses are present.

In conclusion, little evidence is available on the long-term benefit of intracoronary 188Re-based therapy for the various kind of lesions. Recently, drug-eluting stents based on sirolimus or paclitaxel have been generally accepted as a new tool to reduce in-stent stenosis. Therefore, it is foreseen that the indication for intracoronary radionuclide therapy will decrease rapidly in the near future [50].

Conclusion

Radiation synovectomy

Rheumatoid arthritis causes chronic synovial inflammation which can lead to destruction of the articular cartilage, followed by progressive loss of joint function. Radiation synovectomy has been developed as an alternative to surgical synovectomy for the treatment of rheumatoid arthritis. This procedure consists of an intraarticular injection of beta-emitting radiopharmaceuticals into a joint to counteract and control synovial inflammation. This technique has been used extensively in some European countries for more than 30 years. However, it has not gained widespread acceptance. Radioactive leakage from the treated joints can be minimized by immobilizing the treated joint for the first 48 h after administration, by using radioactive particles of an appropriate size (1–20 mm) and by choosing a radioisotope with a short half-life. 188Re is suitable for treatment of the knee owing to its deep tissue penetration (maximum 11 mm, average 3.8 mm) and its short half-life can effectively reduce the hazard of systemic radiation secondary to leakage [51]. A 188Re–Sn colloid was developed by the Seoul National University and its efficacy and safety was investigated in 21 rheumatoid arthritis patients refractory to intra-articular corticosteroid injection. Activities ranging from 3.7 to 1.1 GBq were injected in the joint cavity of the knee. Patients were followed for 1 year and pain decreased in 86%, joint tenderness improved in 64% and joint swelling was reduced in all cases. Transient reactive synovitis was observed in 82%. No clinical side-effects or abnormalities in leukocyte count, platelet count, liver function tests or urine analysis were observed in any patient [52]. Future prospects

In the future, antimicrobial therapies by means of radioimmunotherapy could play a role. Fungal infections are difficult to treat and currently constitute a major clinical problem, in particular in the light of the overwhelming number of HIV-positive patients. Dadachova and co-workers reported an interesting animal

The use of an on-site installable 188W/188Re generator provides a good yield of carrier-free 188Re routinely for radionuclide therapy. The main advantages of 188Re are the relatively short physical half-life allowing high dose rates and repeated treatments and the high energy of the beta particles. Compared to the use of 131I, fewer radioprotective concerns are involved with 188Re. Most of the clinical experiences to date have been established with HEDP labelled with 188Re for treatment of painful bone metastases. Encouraging results were reported using repeated dosages. Besides good palliative effects, anti-tumour effects in terms of decreases in PSA levels were reported. In the field of haematology 188Re has been used in an innovative setting in which 188Re-based radioimmunotherapy was used as a part of the conditioning regimes prior to HSCT. In contrast to the only modest advances made in systemic 188 Re-based radionuclide therapy for solid tumours, encouraging results for loco-regional treatment of HCC were reported. Further optimization of the applied radiopharmaceuticals is under way. Besides several indications in clinical oncology, 188Re has also been proposed for benign disorders. Most research was performed in the area of prevention of stenosis following coronary angioplasty. However, progress in the development of drug-eluting stents will probably reduce the indication for intracoronary radionuclide therapy in the future. Another indication for 188Re-based therapy is the intra-articular application for radiosynovectomy in refractory arthritis. In conclusion, a considerable number of clinical trials, directed to the application of 188Re in oncological as well as benign conditions, have demonstrated its feasibility. However, most clinical know-how remains limited to a number of expert centres in Germany and Asia. In the future, efforts should be made to facilitate innovative research using a 188W/188Re generator and the implementation of 188Re-based therapies should be encouraged in other centres.

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Original article

FDG PET positive lymph nodes are highly predictive of metastasis in breast cancer Rakesh Kumar, Hongming Zhuang, Mitchell Schnall, Emily Conant, Stephanie Damia, Susan Weinstein, Prem Chandra, Brain Czerniecki and Abass Alavi Aim To determine whether or not fluorodeoxyglucose positron emission tomography (FDG PET) imaging when positive could obviate the necessity for sentinel lymph node biopsy and for complete axillary node dissection in patients with breast cancer. Methods A total of 80 female patients with a histological diagnosis of breast cancer and clinically negative axillary nodes underwent an FDG PET and sentinel lymph node biopsy (SLNB) or total axillary dissection for staging of axilla. Both SLNB and axillary dissection were performed in 72 patients, while eight patients had total axillary dissection without SLN biopsy. Results Of the 80 patients, 36 had lymph node metastasis on histopathology. SLNB was positive for metastasis in 35 (97%) of 36 patients (29 macrometastasis and seven micrometastasis). In the patient with false negative SLNB, the lymph node was completely replaced by the tumour. The FDG PET was true positive in 16 of 36 patients (sensitivity, 44%). There were two false positive studies with FDG PET, resulting in a specificity of 95%. The positive predictive value and accuracy of FDG PET for the detection of axillary lymph node metastasis were 89% and 72%, respectively. Univariate analysis revealed that higher grade of tumour, increased size and number of axillary lymph nodes were significantly associated with positive FDG PET results for axillary staging.

Introduction Accurate staging is important for selecting an appropriate treatment plan for all malignancies, including breast cancer. Axillary lymph node metastasis is the single most important factor in determining prognosis and for deciding about adjuvant chemotherapy [1]. Presently, sentinel lymph node biopsy (SLNB) is a well-established procedure for detecting axillary lymph node metastasis from primary breast cancer [2,3]. This procedure has a high accuracy and can therefore separate those patients who should undergo total axillary dissection, which has high morbidity from those who can be managed without axillary dissection. However, false negative axillary results can occur in a variable percentage of patients. In addition, there is also concern that upstaging of the disease may

Conclusion FDG PET cannot replace histological staging using SLNB in patients with breast cancer. However, FDG PET has a high specificity and positive predictive value for staging of the axilla in these patients. The patients with higher grade of tumour, larger size and higher number of axillary lymph nodes may be considered for FDG PET scan for axillary staging. Nucl Med Commun 27:231–236

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:231–236 Keywords: FDG PET, sentinel lymph node biopsy, breast cancer, staging, axillary dissection Division of Nuclear Medicine, Department of Radiology, Hospital of the University of Pennsylvania, Philadelphia, USA. Correspondence to Dr Abass Alavi, Division of Nuclear Medicine, Hospital of the University of Pennsylvania, 110 Donner Building, 3400 Spruce Street, Philadelphia, PA, 19104, USA. Tel: + 001 215 662 3069; fax: + 001 215 349 5843; e-mail: [email protected] Sponsorship: This work was supported by Public Health Services Research Grant M01-RR00040 from NIH. Dr Rakesh Kumar was financially supported by the International Union Against Cancer, Geneva, Switzerland, under an ACSBI fellowship. Received 15 July 2005 Accepted 6 September 2005

occur in up to 46% of patients with breast cancer when SLNB is coupled with very thorough pathological examination (immunohistochemical analysis and multistep sectioning of the sentinel node), which may detect micrometastasis. However, the prognostic and therapeutic relevance of these micrometastasis remains to be established [4–6]. Step sectioning and immunohistochemistry (IHC) can detect micrometastasis, but these techniques are time-consuming and laborious [7]. Therefore, these patients would undergo a two-step surgical procedure, depending upon the histopathology and immunohistochemistry results from sentinel lymph node biopsy. SLNB in the detection of possible metastasis in the internal mammary and supraclavicular lymph nodes is not clear.

c 2006 Lippincott Williams & Wilkins 0143-3636

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232 Nuclear Medicine Communications 2006, Vol 27 No 3

Many investigators have assessed the role of positron emission tomography (PET) using [18F]fluorodeoxyglucose (FDG) in the management of patients with breast cancer [8–10]. FDG PET has a relatively high sensitivity and specificity with regards to the status of axillary lymph nodes, distant metastasis, and recurrence as compared to conventional morphological imaging modalities [9–11]. But many recent studies have yielded conflicting results, wherein the accuracy of FDG PET has been compared to the accuracy of sentinel lymph node biopsy or axillary node dissection [12–16]. A recent study by Lovrics et al. showed that FDG PET had limitations in detecting lymph node metastasis when compared to sentinel lymph node biopsy although the technique was highly specific for axillary staging [16]. This study was designed to determine whether a positive FDG PET scan could be used to guide complete axillary dissection without the need for sentinel lymph node biopsy in patients with breast cancer.

Materials and methods Patient population

A total of 80 female patients with a histological diagnosis of breast cancer and clinically negative, non-palpable axillary lymph nodes were recruited for this study. The mean age was 52 ± 11 years and age range was 32–79 years. All patients underwent multiple imaging techniques, including FDG PET, as part of an NIH funded project for characterizing primary breast lesions and for locoregional staging. This study was conducted after obtaining ethics clearance from the Institutional Review Board, University of Pennsylvania, Philadelphia. In this report we describe only the results of FDG PET imaging and sentinel lymph node biopsy and axillary dissection. Of the 80 patients SLNB plus axillary dissection was carried out in 72 patients, while the remaining eight patients had a total axillary dissection without SLNB. The final diagnosis and staging were determined on the basis of histopathology of the breast lesions, sentinel nodes, and other axillary lymph nodes. 18

F-FDG positron emission tomography imaging

Patients fasted for a minimum of 4 h and had a normal serum glucose level (less than 150 mg
dl – 1) which was verified before the administration of 5.2 MBq
kg – 1 (0.14 mCi
kg – 1) FDG through a peripheral vein. Sequential overlapping emission scans of the neck, chest, abdomen, and pelvis were acquired on a dedicated wholebody PET scanner (Allegro Philips Medical System, Philadelphia, PA) 60 min after the administration of the radiopharmaceutical. Transmission scans using 137Cs point sources were interleaved between the multiple emission scans to correct for non-uniform attenuation. Images were reconstructed using an iterative reconstruction algorithm, and both attenuation corrected and nonattenuation corrected images were interpreted.

Radiotracer injection and lymphoscintigraphy

Radiotracer injection was performed on the day of surgery using 37 MBq (1.0 mCi) of 99mTc labelled sulfur colloid filtered through a 0.22 mm filter. For this purpose, a single injection of 0.1 ml of radiotracer was given in the subareolar region of the affected breast. In cases where the tumour was not palpable, image guided needle localization and radiotracer injection were performed using ultrasonography. For radiotracer injection around the site of a prior excision biopsy, ultrasound localization was used to prevent injection into the biopsy cavity. Lymphoscintigraphy was performed using a large field of view gamma camera with a high-resolution collimator (Model 2000; General Electric Medical Systems, Waukesha, Wisconsin). Static images were obtained using both anterior and lateral projections for 1–2 h following radiotracer administration. If necessary, an external source of radiotracer was used to map the contour of the shoulder region in order to determine the relative location of sites visualized during scanning. All the draining lymph nodes identified on scintigraphy were marked on the overlying skin with indelible ink. If nodal drainage was not identified after 2 h, the case was considered to be a lymphoscintigraphy failure. Blue dye injection and operative procedure

Following the induction of general anaesthesia, all patients underwent sub-areolar injection of 3 ml 1% isosulfan blue dye (Lymphazurin; US Surgical Corporation, Norwalk, Connecticut). Within 5–10 min of blue dye administration, the SLN mapping procedure was initiated. A gamma-detecting probe (Norwalk, Connecticut) was used intra-operatively to identify isotope positive nodes. An initial background axillary count was taken with the gamma-detecting probe, and then the axillary site with the highest number of counts was located. The axillary skin incision was made directly over this site. Careful dissection was carried out to identify all blue lymphatics and blue nodes (defined as any node that stained blue). Additional nodes were identified for dissection by using the gamma-detecting probe to locate isotope positive nodes. These were defined as nodes with counts at least three times the background or ten times ex-vivo fat. Isotope positive nodes were removed until the background axillary counts were < 10% of the most radioactive node removed. After completion of the SLN mapping portion of the procedure, removal of the primary tumour was performed. Pathological evaluation

Axillary lymph nodes were identified as SLN or non-SLN and were formalin fixed and paraffin embedded. All SLNs were evaluated by haematoxylin and eosin (H&E) staining and by immunohistochemistry using a cytokeratin antibody (AE1/3; monoclonal antibody, Boehringer

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FDG PET positive lymph nodes predict metastasis in breast cancer Kumar et al. 233

Mannheim, Indianapolis, Indiana). Non-SLNs were evaluated by H&E staining only. Statistical analysis

The unpaired Student’s t-test was used to see the mean significant difference between true positive and false negative FDG PET results for axillary lymph node metastasis among various quantitative variables such as age and tumour size. For skewed data, corresponding nonparametric test, i.e, Mann–Whitney U statistics was applied. The chi-squared test was used to determine the association between various qualitative variables such as tumour type and tumour grade with true positive and false negative FDG PET results. A P value < 0.05 was considered significant. All the statistical analysis was carried out by using statistical software SAS 8.2.

detection of lymph node metastasis was 44%. The characteristics of patients with true positive FDG PET results for axillary lymph node metastasis are given in Table 2. There were two false positive results with FDG PET. The overall specificity of FDG PET for the detection of lymph node metastasis was 95%. The positive predictive value and accuracy of FDG PET for the detection of axillary lymph node metastasis were 89% and 72%, respectively. Interestingly, the patient who had a false negative SLNB had positive result for lymph node metastasis on the FDG PET scan. In one patient, FDG PET showed axillary and internal mammary lymph node metastasis, while SLN imaging showed a single lymph node in the axilla. This axillary node was later confirmed to be a metastasis by SLNB but no surgery was performed to find the internal mammary lymph node, which was positive according to FDG PET imaging.

Results The demographic and clinical characteristics of all 80 patients are summarized in Table 1. Of the 80 patients who underwent SLNB plus axillary dissection or complete axillary dissection, 36 had lymph node metastasis. Of these 36 patients, 29 were noted to have macrometastasis and the remaining seven had micrometastasis. The SLNB was positive for metastasis in 35 of 36 (97%) patients. One patient had a false negative SLNB, resulting in a false negative rate of 3%. The FDG PET was true positive in 16 of 36 patients. The smallest lymph node detected by FDG PET was 8 mm in size. The overall sensitivity of FDG PET for the Demographic and clinical characteristics of the 80 patients with clinically negative axillary lymph nodes

Table 1

Characteristic Age (years) Mean Range Breast (right/left) Menopausal status Premenopausal Perimenopausal Postmenopausal

52 ± 11 32–79 40/40 33 11 36

Tumour size (mm) Mean Range

16.4 ± 13.5 2–69

Tumour type Ductal Lobular Ductal + lobular Others

63 6 9 2

Tumour grade High Moderate Low

29 22 29

Standardized uptake values Maximum Average

2.9 ± 3.1 2.3 ± 2.5

FDG PETwas false negative for lymph node metastasis in 13 of 29 macrometastasis and in all seven patients with micrometastasis. The characteristics of patients with false negative FDG PET results for axillary lymph node metastasis are given in Table 3. The lymph nodes, which were positive for micrometastasis, were found to harbour one to 20 malignant cells and size of the tumour ranged from 0.5 mm to 2 mm in size. Among the 13 patients who had macrometastasis and false negative FDG PET results, the size of the lymph nodes varied from 5 mm to 20 mm. Univariate analysis revealed significant differences in patients with true positive and false negative results of FDG PET for axillary lymph node metastasis among the tumour grade, maximum and average SUV and size and number of lymph nodes (P < 0.05). Higher grade of tumour, high maximum and average SUV and increased size and number of lymph nodes were significantly associated with positive FDG PET results. Increased size of the primary tumour was not significantly associated with positive results of FDG PET for axillary lymph node metastasis.

Discussion Axillary lymph node metastasis is the most important prognostic factor for patients with breast cancer [1]. SLNB has improved patient management in several types of malignancies, among which breast cancer stands out as one of the most influenced by this technique. SLNB is best indicated in smaller tumours with clinically nonpalpable axillary lymph nodes. In the present study most of the patients (74/80) had primary tumour size of 3 cm. SLNB has certain limitations, including a false negative rate of 1–15% as previously reported [17–21]. The common causes of false negative SLNB results have been reported to be due to replacement of the entire lymph node with tumour cells, blockage of lymphatic ducts by infiltrating tumour cells, and post-surgical or

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Table 2

The characteristics of patients with true positive FDG PET results for axillary lymph node metastasis

S. no. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 Mean ± SD

Age (years)

LN size (mm)

LN number

Micro/macro metastases

Tumour size (mm)

Tumour type

Tumour grade

Uni/multi focal

46 49 35 38 66 51 40 48 44 73 51 46 58 50 54 38

10, 9, 5 20, 15, 10 15, 10 20 NA 30, 8 16, 17 NA NA 10 NA 18 NA NA NA NA

3 10 6 3 1 2 1 2 3 1 2 1 5 6 1 3

Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets Macromets

45, 20, 12 10 12 25 9 5 15, 12 5 19 3, 2, 3 47 5 60 40 6 15

ILC IDC IDC IDC IDC IDC IDC IDC IDC IDC + ILC IDC IDC IDC IDC IDC IDC

High High High High Low Low High Low High Low Moderate High High Moderate High High

Multifocal Unifocal Unifocal Unifocal Unifocal Unifocal Multifocal Unifocal Unifocal Multifocal Unifocal Unifocal Unifocal Unifocal Unifocal Unifocal

49 ± 10

15.8 ± 5.1

3.1 ± 2.5

18.8 ± 16.8

Tumour SUV Tumour SUV (max.) (avg.) 1.9 2.3 6.5 2.5 1.3 2.1 2 1.5 8.9 0.8 7.5 3.1 4.5 9.2 2.5 9.7

1.7 1.8 4.5 2.1 1.2 1.8 1.4 1.2 7.4 0.7 5.2 2.5 2.7 6.2 2.1 6.7

4.1 ± 3.1

3.1 ± 2.2

NA, not available; IDC, invasive ductal carcinoma; ILC, invasive lobular carcinoma; LN, lymph node; SUV, standardized uptake value.

Table 3

The characteristics of patients with false negative FDG PET results for axillary lymph node metastasis

S. no. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Mean ± SD

Age (years)

LN size (mm)

LN number

Micro/macro metastasis

Tumour size (mm)

Tumour type

Tumour grade

Uni/multi focal

44 57 59 57 74 49 52 56 64 72 51 42 49 43 50 41 53 53 40 47

12 7 2 NA NA NA 5, (20 cells) NA 1 5, (20 cells) 12 20 NA 14 1 cell < 10 NA NA NA NA

1 1 1 5 2 2 1 2 1 1 1 1 2 1 1 1 4 3 3 2

Macromets Macromets Micromets Macromets Micromets Macromets Micromets Macromets Micromets Micromets Macromets Macromets Macromets Micromets Micromets Macromets Macromets Macromets Macromets Macromets

8 5 9 20 9, 14 2 9 24 10 40 30 6 15 29 30 16 12 69 5 22

IDC IDC IDC IDC ILC + IDC IDC ILC IDC IDC IDC + ILC IDC + ILC IDC IDC + ILC IDC IDC IDC IDC IDC IDC IDC

Low Low Low Moderate Moderate Low Low Moderate Moderate Moderate Moderate Low Moderate Moderate High Moderate Moderate Moderate Low High

Multifocal Unifocal Unifocal Multifocal Multifocal Unifocal Unifocal Unifocal Unifocal Unifocal Multifocal Unifocal Unifocal Unifocal Multifocal Unifocal Unifocal Unifocal Unifocal Unifocal

53 ± 9

8.1 ± 6.1

1.8 ± 1.2

19 ± 16

Tumour SUV Tumour SUV (max.) (avg.) 1.2 1.5 1.3 2.5 0.7 1.2 2.2 2.7 1.6 2.9 0.9 1.4 1.5 3 3.8 1.2 2.1 1.2 1.5 2.5

1 1.3 1.2 2.3 0.5 1.1 1.7 2.5 1.3 2.4 0.8 1.2 1.2 2.2 3.2 1 1.5 0.9 1.2 2.2

1.8 ± 0.8

1.5 ± 0.7

Abbreviations as in the footnote to Table 2.

technical limitations (injection of the wrong radiotracer, improper injection techniques). A recent multi-centre study involving 4117 patients with 106 false negative results demonstrated that a tumour size < 2.5 cm, upper outer quadrant tumour location, removal of only a single SLN, minimal surgeon experience, presence of a single positive axillary lymph node, and use of immunohistochemistry for SLN analysis were independently associated with an increased risk of false negative results [22]. In the present study, only one patient (3%) had false negative SLNB results. Histopathology in that case showed replacement of the entire lymph node by tumour cells, along with central necrosis. FDG PET was true positive in this patient. FDG PET imaging appears to be a promising technique for the detection of axillary involvement, especially in the setting described above

where SLNB may possibly fail and therefore adversely affect patient management.

FDG PET reflects the glycolytic activity of cells, which is higher in cancer cells than in non-neoplastic tissues. Because of its unique metabolic imaging capabilities, FDG PET has proven to be very successful as a noninvasive diagnostic method for the management of numerous types of malignancies, including breast cancer. Several investigators have assessed the sensitivity and specificity of FDG PET for determining the status of axillary lymph nodes in patients with breast cancer. These investigations have yielded widely variable and conflicting results. Most of the earlier and only a few recent studies have reported that FDG PET imaging can

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FDG PET positive lymph nodes predict metastasis in breast cancer Kumar et al. 235

adequately replace the invasive SLNB procedure [13,14,23–27], while most of the more recent studies have revealed controversial results about the accuracy of FDG PET for the assessment of axillary nodes in breast cancer patients [12,15,28–33]. In a recent multi-centre and prospective study Wahl et al. demonstrated the mean sensitivity and negative predictive value of PET as 61% and 79%, respectively [34]. However, they found FDG PET as highly predictive for nodal tumour involvement, when multiple intense foci of tracer uptake were identified. This poorer performance of PET might be explained by the different characteristics of the patients studied, such as the prevalence of small and less easily detectable axillary metastasis, or the protocols used to reconstruct PET images. In contrast to the variable sensitivity of FDG PET as reported in the literature, almost all studies have shown specificities higher than 90% and some approaching 100% for FDG PET in determining the status of axillary lymph nodes (Table 4). In other recent studies, the specificity of FDG PET for axillary staging has been reported to vary from 93% to 98% [16,35,36]. In our study, the combined radiocolloid and blue dye method was used to identify SLNs in all patients. Among all patients who had axillary lymph node dissection with or without SLN detection, 45% had lymph node metastasis. In this study, FDG PET results included only two false positive results for the detection of axillary lymph node involvement. Both of these patients had recent excision biopsy before FDG PET study and histopathology results were consistent with inflammation.

Specificity of FDG PET scanning in detection of sentinel lymph node and axillary lymph node metastasis in breast cancer patients

Table 4

Author

Year

Number of patients

Sensitivity (%)

Specificity (%)

Present study Lovrics et al. [16] Zornoza et al. [36] Fehr et al. [35] Wahl et al. [34] Barranger et al. [12] Van der Hoeven et al. [15] Kelemen et al. [30] Guller et al. [28] Yang et al. [33] Schirrmeister et al. [32] Greco et al. [13] Yutani et al. [37] Crippa et al. [40] Noh et al. [25] Smith et al. [26] Crippa et al. [39] Adler et al. [23] Avril et al. [29] Scheidhauer et al. [27] Utech et al. [14] Crowe et al. [24] Adler et al. [38]

2005 2004 2004 2004 2004 2003 2002

80 98 200 24 360 32 70

44 40 84 20 61 20 25

95 97 98 93 80 100 97

2002 2002 2001 2001 2001 2000 1998 1998 1998 1997 1997 1996 1996 1996 1994 1993

15 31 18 117 167 38 72 27 50 82 50 41 18 124 20 20

20 43 50 79 94 50 85 93 90 84 95 79 100 100 90 90

90 94 100 93 86 100 91 100 97 85 66 100 89 75 100 100

Our results are similar to most of the other studies, which also showed high specificities (Table 4). FDG PET scans were false negative in all seven patients with micrometastasis and 13 of 29 patients with macrometastasis. This finding is also comparable to the results published in the literature, wherein false negative rates have been reported to vary between 0% and 80% [15,24–26,28,37]. The main reasons for higher FDG PET false negative results for axillary staging include the limited spatial resolution of the technique, and fewer metabolically hyperactive cells in patients with micrometastasis. In the present study, the mean size of the lymph node was 8.1 mm in false negative cases while it was 15.8 mm in patients with true positive results. In patients with micrometastasis, there were few tumour cells (one to 20 cells) on microscopy. We also found other factors which were associated with false negative FDG PET results, i.e., malignancy with moderate and lower histological grade, lower standardized uptake value of primary tumour and fewer lymph nodes harbouring metastasis. We did not find any significant association between primary tumour size and false negative FDG PET results, which was reported by Martin et al. [22]. Together, the results of the present study and a review of the literature have revealed a high specificity of FDG PET for the detection of axillary lymph node metastasis. Therefore, FDG PET may be warranted for use in preoperative staging of patients with breast cancer especially in patients presenting with higher grade of primary tumour, increased size and number of axillary lymph nodes. In addition, FDG PET also provides additional information about the extent of disease, such as metastasis to lymph nodes outside the axillary region, as well as distant metastasis to the bone marrow and the liver [9]. In this study, FDG PET imaging showed an increased FDG uptake in the internal mammary lymph node region (in addition to the axilla) in one patient. However, SLN imaging showed only the lymph node in the axilla and SLNB of this node confirmed metastatic involvement. Although no surgical intervention was performed to find the internal mammary lymph node, which appeared on the FDG PET scan, the possibility of metastatic involvement of the internal mammary lymph node cannot be excluded. This may be due to the fact that in the present study we did not use a deep injection of the radiotracer, which may prevent the visualization of SLNs in the internal mammary region. FDG PET imaging may have a potential role in identifying extraaxillary lymph node metastasis which may be missed by SLN imaging. The study concludes that FDG PET cannot replace histological staging using SLNB in patients with breast cancer. However, FDG PET has a high specificity and positive predictive value for staging of the axilla in these

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patients. The patients with higher grade of tumour, larger size and higher number of axillary lymph nodes may be considered for FDG PET scan for axillary staging.

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Krag D, Weaver D, Ashikaga T, Moffat F, Klimberg VS, Shriver C, et al. The sentinel node in breast cancer – a multicenter validation study. N Engl J Med 1998; 339:941–946. Borgstein PJ, Pijpers R, Comans EF, van Diest PJ, Boom RP, Meijer S. Sentinel lymph node biopsy in breast cancer: guidelines and pitfalls of lymphoscintigraphy and gamma probe detection. J Am Coll Surg 1998; 186:275–283. Martin 2nd RC, Chagpar A, Scoggins CR, Edwards MJ, Hagendoorn L, Stromberg AJ, et al. Clinicopathologic factors associated with false-negative sentinel lymph-node biopsy in breast cancer. Ann Surg 2005; 241: 1005–1012. Adler LP, Faulhaber PF, Schnur KC, Al-Kasi NL, Shenk RR. Axillary lymph node metastasis: screening with [F-18]2-deoxy-2-fluoro-D-glucose (FDG) PET. Radiology 1997; 203:323–327. Crowe Jr JP, Adler LP, Shenk RR, Sunshine J. Positron emission tomography and breast masses: comparison with clinical, mammographic, and pathological findings. Ann Surg Oncol 1994; 1:132–140. Noh DY, Yun IJ, Kim JS, Kang HS, Lee DS, Chung JK, et al. Diagnostic value of positron emission tomography for detecting breast cancer. World J Surg 1998; 22:223–227. Smith IC, Ogston KN, Whitford P, Smith FW, Sharp P, Norton M, et al. Staging of the axilla in breast cancer: accurate in vivo assessment using positron emission tomography with 2-(fluorine-18)-fluoro-2-deoxy-D-glucose. Ann Surg 1998; 228:220–227. Scheidhauer K, Scharl A, Pietrzyk U, Wagner R, Gohring UJ, Schomacker K, et al. Qualitative [18F]FDG positron emission tomography in primary breast cancer: clinical relevance and practicability. Eur J Nucl Med 1996; 23: 618–623. Guller U, Nitzsche EU, Schirp U, Viehl CT, Torhorst J, Moch H, et al. Selective axillary surgery in breast cancer patients based on positron emission tomography with 18F-fluoro-2-deoxy-D-glucose: not yet!. Breast Cancer Res Treat 2002; 71:171–173. Avril N, Dose J, Janicke F, Ziegler S, Romer W, Weber W, et al. Assessment of axillary lymph node involvement in breast cancer patients with positron emission tomography using radiolabeled 2-(fluorine-18)-fluoro-2-deoxy-Dglucose. J Natl Cancer Inst 1996; 88:1204–1209. Kelemen PR, Lowe V, Phillips N. Positron emission tomography and sentinel lymph node dissection in breast cancer. Clin Breast Cancer 2002; 3:73–77. Kumar R, Alavi A. Fluorodeoxyglucose-PET in the management of breast cancer. Radiol Clin North Am 2004; 42:1113–1122. Schirrmeister H, Kuhn T, Guhlmann A, Santjohanser C, Horster T, Nussle K, et al. Fluorine-18 2-deoxy-2-fluoro-D-glucose PET in the preoperative staging of breast cancer: comparison with the standard staging procedures. Eur J Nucl Med 2001; 28:351–358. Yang JH, Nam SJ, Lee TS, Lee HK, Jung SH, Kim BT. Comparison of intraoperative frozen section analysis of sentinel node with preoperative positron emission tomography in the diagnosis of axillary lymph node status in breast cancer patients. Jpn J Clin Oncol 2001; 31:1–6. Wahl RL, Siegel BA, Coleman RE, Gatsonis CG. PET Study Group. Prospective multicenter study of axillary nodal staging by positron emission tomography in breast cancer: a report of the staging breast cancer with PET Study Group. J Clin Oncol 2004; 22:277–285. Fehr MK, Hornung R, Varga Z, Burger D, Hess T, Haller U, et al. Axillary staging using positron emission tomography in breast cancer patients qualifying for sentinel lymph node biopsy. Breast J 2004; 10:89–93. Zornoza G, Garcia-Velloso MJ, Sola J, Regueira FM, Pina L, Beorlegui C. 18 F-FDG PET complemented with sentinel lymph node biopsy in the detection of axillary involvement in breast cancer. Eur J Surg Oncol 2004; 30:15–19. Yutani K, Shiba E, Kusuoka H, Tatsumi M, Uehara T, Taguchi T, et al. Comparison of FDG–PET with MIBI-SPECT in the detection of breast cancer and axillary lymph node metastasis. J Comput Assist Tomogr 2000; 24:274–280. Adler LP, Crowe JP, al-Kaisi NK, Sunshine JL. Evaluation of breast masses and axillary lymph nodes with [F-18] 2-deoxy-2-fluoro-D-glucose PET. Radiology 1993; 187:743–750. Crippa F, Agresti R, Donne VD, Pascali C, Bogni A, Chiesa C, et al. The contribution of positron emission tomography (PET) with 18 F-fluorodeoxyglucose (FDG) in the preoperative detection of axillary metastasis of breast cancer: the experience of the National Cancer Institute of Milan. Tumouri 1997; 83:542–543. Crippa F, Agresti R, Seregni E, Greco M, Pascali C, Bogni A, et al. Prospective evaluation of fluorine-18-FDG PET in presurgical staging of the axilla in breast cancer. J Nucl Med 1998; 39:4–8.

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Original article

Three-dimensional positron emission tomography imaging with 124I and 86Y Stefaan Vandenberghe Background Impure positron emitters have physical characteristics that degrade image quality compared to conventional positron emitters like 18F. Two impure positron emitters with potentially interesting applications are 124I and 86Y. The degradation in image quality due to the imperfection of these isotopes is quantified for a human three-dimensional (3-D) positron emission tomography (PET) system. An acquisition protocol to obtain similar image quality as for 18F imaging is determined by Monte Carlo simulations. Methods The effects of larger positron range, associated singles and the other decay modes on image quality are determined by extensive Monte Carlo simulations of the Allegro scanner. Spatial resolution was evaluated for both isotopes and compared to spatial resolution of 18F. The loss in sensitivity due to triple coincidences was determined as a function of the axial acceptance angle of the PET scanner. The performance of the scanner at low count rates was studied by determining the noise equivalent count (NEC) values for different upper energy thresholds. The image degrading effect of spurious coincidences is taken into account by adding another factor to the NEC calculation. This allowed the contribution of spurious coincidences to be minimized by using a setting for the appropriate energy window. For this optimal energy window the amount of spurious and scattered coincidences was quantified. Simulations of count rate performance were also done to determine the peak NEC and the activity at which the maximum occurred. Results Spatial resolution degradation, compared to 18F, is about 0.5 mm for 86Y and 1 mm for 124I. Associated singles have a similar effect as scattered coincidences, as they also add a background to the image. The effect, however, is

Introduction In addition to thyroid cancer therapy with Na131I, which is approved by the US Food and Drug Administration (FDA), there are other agents that are being considered for radionuclide therapy. Various alpha and beta emitters have the potential to be successful agents for radionuclide therapy. Recently, treatments with Zevallin (90Y) and Bexxar (131I) have received FDA approval. The current state of the art in radionuclide therapy is well described by Larson and Krenning [1].

less important than the effect of scatter. The fraction of triple coincidences is quite small for a 3-D PET scanner used for humans as the axial acceptance angle is still moderate. For the Allegro with an energy resolution of 18% the optimal upper energy threshold was determined at 600 keV. For 124I this leads to 2.5% extra contamination that needs to be added to the scatter fraction. For 86Y this fraction is about 5.5%. Conclusion 3-D PET images of 124I and 86Y have lower spatial resolution. For PET scanners used for humans the difference is not as important as for scanners used for animals. The limited positron decay fraction of both isotopes can be compensated by increasing the imaging time by a factor of 3–5 (same activity). A short coincidence window limits the contamination from other decay modes. Good energy resolution allows setting a selective upper energy threshold to limit the effect of spurious coincidences. With an appropriate setting of the energy window it should be possible to obtain good image quality in a relatively short time because of the high sensitivity of 3-D PET scanners. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:237–245
Nuclear Medicine Communications 2006, 27:237–245 Keywords: positron emission tomography, immuno-PET, non-conventional isotopes ELIS/MEDISIP Ugent, Belgium, formerly with Philips Research USA, Briarcliff, New York, USA. Correspondence to Dr Stefaan Vandenberghe, ELIS/MEDISIP Ugent, St-Pietersnieuwstraat 41, 9000 Gent, Belgium. Tel: + 0032 9 264 6628; fax: + 0032 9 264 3594; e-mail: [email protected] Received 13 October 2005 Accepted 25 November 2005

One important step in radionuclide therapy is to have an image of the distribution of the agent in the body before and during the therapy. This makes it possible to estimate the absorbed dose before and during therapy. Currently, this is often done using scintigraphic wholebody planar images. For a limited number of agents (mostly 131I), radiation produced by the isotope can be used for imaging, but the images obtained with single photon emission computed tomography (SPECT) scanners are often of poor quality for a variety of reasons [2,3].

c 2006 Lippincott Williams & Wilkins 0143-3636


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Often it is necessary to use high-energy collimators which have poor spatial resolution: sometimes there is also contamination from higher energy peaks. Another reason is that SPECT images are noisy due to the low number of counts that are acquired. There are some impure positron emitters available, however, with potential applications for dosimetry, which can be used for Positron emission tomography (PET) imaging [4]. Their longer half-lives (compared to 18F) can make their use practical and allows processes to be followed in the body over different days. The advantage of these isotopes (compared to their equivalent SPECT isotopes) is that the annihilation photon emissions allow electronic collimation by PET, which results in a higher quality scan than single-photon imaging devices. PET systems have a much higher sensitivity, a better spatial resolution and it is easier to obtain quantitative data. It is also easier to obtain tomographic whole-body images in a reasonable time (30–40 min). Furthermore, almost all new PET systems are hybrid devices and the attached or integrated computed tomography (CT) also delivers a high quality whole-body CT in a very short time (a few minutes) [5]. The CT makes it possible to segment (manually or with dedicated algorithms) the different organs accurately. The direct availability of the accompanying emission distribution will make it possible to perform more accurate dosimetry. Due to the direct availability of a transmission map, the simpler attenuation correction and the better spatial resolution quantification in PET is also easier than for SPECT imaging. These are all interesting properties for dosimetric applications Most isotopes for PET produce only a single positron that annihilates with an electron into two 511 keV gamma photons. Impure positron emitters, however, emit additional gamma photons. These gamma photons are sometimes even in cascade with the positron emission. This process takes place within a time interval that is smaller than the timing window of current PET scanners. Therefore the system will not always detect the true coincidences and it can detect combinations of the cascade single with one of the annihilation photons. These are called contaminated or spurious coincidences [6]. As there is no angular correlation between the cascade single and the annihilation photon, the recorded line of response will not be correct. In general, it would be expected that a three-dimensional (3-D) PET system would detect more of these spurious coincidences than two-dimensional (2-D) PET systems due to a larger solid angle. As it is well known that 3-D PET systems have a higher sensitivity, both factors have to be taken into account when considering image quality. The latest generation of 3-D PET scanners have an improved energy resolution and the size of their energy window is reduced compared to older PET scanners [7,8]. The good energy resolution allows the upper and lower thresholds to be

adjusted more strictly to limit the number of contaminated coincidences while the majority of true coincidences are maintained. The focus in this work is on two isotopes: 124I and 86Y. However, the methods described below can be applied to other impure positron emitters like 76Br, 60Cu, 61Cu, 64Cu, 66 Ga and 94Tc, which can be used for PET imaging [6]. It is important to understand how the physical characteristics of these isotopes have an impact on the imaging performance. They make it quite challenging for a PET system to acquire the correct coincidences. The study of these isotopes on a real system is often difficult due to their limited availability. Therefore the use of accurate Monte Carlo simulations was chosen as this allows the history of all detected coincidences to be documented. Another advantage of Monte Carlo techniques is that it makes it possible to determine how many true coincidences were detected and to determine to which group a detected coincidence (true, scattered, random, contaminated) belongs. Even with special phantoms (e.g., line source) it is not possible to differentiate whether a measured coincidence was a scattered, a random or a contaminated coincidence in PET.

Materials and methods Physical characteristics of therapy isotopes 131

131

I and

90

Y

–

I is b emitter with a maximum energy of 606 keV and a mean energy of 191 keV [9]. The b – emission is followed by emission of gamma rays (mainly 364 keV and 637 keV). SPECT imaging of 131I with high-energy collimators delivers low count images with poor contrast. The high-energy collimator limits the spatial resolution, the 637 keV high-energy photons add a background to the image and the high dose administered to the patient results in a count rate that is higher than the normal range at which most gamma cameras normally operate. 90

Y does not emit any gamma photons that can be used for gamma camera imaging. It emits high-energy b – particles (maximum 2.27 MeV) useful for dosimetric purposes [9]. These emissions have a maximum range of approximately 10 mm (mean range of 3.9 mm) in soft tissue. The whole-body radiation absorbed dose burden is small due to the fact that no other decays are present. Physical characteristics of imaging isotopes 124I and 86Y

The decay scheme of 124I is rather complex [9]. It has different decay modes and only in 11% of the decays is there a pure positron emission. However, in about another 12% of the decays it also emits a positron that is directly (within a few picoseconds) followed by a single photon of 602 keV. Compared to 18F it emits positrons with a higher energy. This leads to degradation of spatial resolution. In the other 75% of the decays, it decays by a cascade of gamma rays (often ending with a 602 keV photon).

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3-D PET imaging with

Fig. 1

(a)

I-124

Non positron decays 76%

β+ + 602 kev 12%

... 602 kev

+ β 12%

0 kev Y-86

(b)

β+ + 627 kev 6.4%

Non positron decays 68% ...

+ β + 443 kev 6.4%

627 kev 443 kev

+ β 20%

0 kev Simplified decay schemes for the isotopes

124

I and

86

Y.

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I and

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Y Vandenberghe

239

medicine. Due to the use of the Geant 4 libraries it also makes it easy to use any isotope. The scanners above were modelled with the GATE tool. We have used an accurate model (a GATE macro) describing the geometry, shielding, covers and crystals [11]. In this paper the model was validated versus measured data and obtained good agreement at low rates for sensitivity and resolution. Dead-time behaviour of PET scanners is influenced by different factors (scintillator/electronics). There is a continuing progress in electronics to achieve lower dead-times in different components of the electronics chain. Therefore it is difficult to speak of one dead-time for a certain type of scanner. The exact performance is dependent on the exact implementation/release of the scanner. Obtaining a perfect match requires quite some fine tuning of the post-processing dead-times in the simulator. We have chosen to use a relatively simple deadtime (singles paralysable) that results in a realistic NEC performance of the scanner close to the performance of the Allegro scanner. The system was modelled with a paralysable singles dead-time of 210 ns, resulting in a peak true rate of 87.6 kcps at 13 kBq  ml – 1 and a peak NEC [12] of 38.8 kcps at 9.8 kBq  ml – 1. The values from a measurement on the first Allegro scanner [7] are a peak true rate of 83 kcps at 13.7 kBq  ml – 1 and a peak NEC rate of 30 kcps at 9.25 kBq  ml – 1. Image quality

86

The decay scheme of Y is also quite complex [9]. It is a positron emitter with a half-life of 14.7 h. Like 124I it has different decay modes and only in 32% of the decays is there a positron emission. All of these are followed by one or more gamma rays in cascade. The other 67% of transitions are via electron capture. All of these are followed by two or more gamma rays in cascade. Simplified decay schemes for both 124I and 86Y are shown in Fig. 1. Scanners

The Philips Allegro scanner is a PET scanner designed for human whole-body imaging [7]. It is composed of 28 flat modules of a 22  29 array of gadolinium orthosilicate (GSO) crystals with dimensions of 4  6  20 mm3. The diameter is 82 cm and the scanner has an axial field-ofview (FOV) of 18 cm. Because GSO has a good energy resolution and a fast decay time it can be used in fully 3-D mode [8]. This leads to a high sensitivity and countrate performance. The patient port is 56 cm, and 2.8575 cm thick lead shielding limits the influence of activity outside the FOV. GATE Monte Carlo simulation

As a Monte Carlo simulation tool, GATE (the GEANT4 Application for Tomographic Emission), was used [10]. It encapsulates the GEANT4 libraries and is a versatile simulation toolkit adapted to the field of nuclear

The non-ideal decay of 124I and 86Y has an impact on the image quality. The following characteristics of the decay of 124I are expected to influence different parameters: 1. The positron range is larger than for 18F and this will affect spatial resolution. 2. Only 23% of decays of 124I are pure positron emission (30% for 86Y), which will lead to lower sensitivity and inferior count-rate performance. 3. Fifty percent of the positron decay is followed by a single photon of 602 keV, which can lead to spurious and triple coincidence. This affects the sensitivity and noise equivalent counts. 4. There are singles of different energies from nonpositron decay modes. This can lead to an increased randoms fraction. The same problems exist for 86Y: a positron is generated only in 33% of decays. It can be followed in cascade with 627 keV or 443 keV. These effects are investigated in detail by extensive Monte Carlo simulations. Spatial resolution

The larger positron range will lead to a degradation of spatial resolution. The absolute effect will be different for small animal imaging systems and human imaging systems due to the fact that resolutions add in quadrature. Here we limited our simulations to the Allegro scanner.

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Three simulations were performed. A line source was filled with either 18F, 124I or 86Y (1 MBq). This line was placed on the central axis of the scanner. One million detections were simulated. The axial sum of all rebinned sinograms (single-slice rebinning) generates a straight line. Adding all angles together delivers a Gaussian profile. From this profile the full width at half maximum(FWHM) was determined with a Matlab function written in-house. The profiles are shown in Fig. 2 and the FWHM and FWTM for the three isotopes are shown in a bar plot in Fig. 3.

Fig. 3

14 FWHTM FWHM

12 10 8 6 4

Sensitivity

The low abundance of positron decay leads to a lower sensitivity and consequently higher noise for the same activity in the FOV. A point source (respectively filled with 18F, 124I and 86Y) at the centre of the FOV was simulated and the number of detected coincidences was determined (Fig. 4). The single photons following the positron decay can lead to contamination of the coincidence measurement. The contamination can result in triple coincidences or contaminated coincidences (Fig. 5). While the contaminated coincidences have no effect on the sensitivity, in most PET scanners triple coincidences will be discarded and therefore this will lead to an extra loss of sensitivity. Given that a coincidence was detected, the probability (at low rates) of having a triple coincidence is equal to the probability of detecting the contamination single (602 keV for 124I and 627 or 443 keV for 86Y). This probability becomes larger when in changing from 2-D to 3-D PET and increases further when 3-D PET with a larger acceptance angle is used. To

2 0 I-124

Y-86

F-18

Full width at half maximum and full width at tenth maximum of and 86Y.

18

F,

124

I

Fig. 4

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9000 8000 7000 6000 5000 4000 3000 2000 1000 0

Fig. 2

Sensitivity of the different isotopes.

0.18 0.16 F-18 Y-85 I-124

0.14

quantify this effect we simulated the same point source in 3-D PET scanners (based on Allegro geometry, but with less or more rings) with an increasing number of axial rings. The result is shown in Fig. 6.

0.12 0.1

Optimization of upper energy threshold for

0.08 0.06 0.04 0.02 0 240

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Pixels (1 mm pixel size) Summed profile (over all angles) of the sinogram of a line source filled with the different isotopes.

124

I

The default upper energy threshold on the Allegro scanner is 670 keV. While this is optimal for maximizing the sensitivity of 18F studies, it is not optimal when spurious coincidences between a 511 keV and a 602 keV photon occur. Due to the good energy resolution (15– 18%) the upper energy threshold for 124I can be further optimized. The figure of merit used here is the NEC at low rates. After estimating the true, scattered and contaminated coincidences, the NEC [12] was calculated using the equation NEC ¼

T T T þSþC

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3-D PET imaging with

Fig. 5

124

I and

86

Y Vandenberghe 241

Fig. 7

300 602 keV

602 keV 511 keV 511 keV 511 keV

511 keV

Triple coincidence

Spurious coincidence

Triple and spurious coincidences shown in the simulated PET scanner.

Coincidence count-rate (cts/s)

250

200

150

100

Trues Scatter NEC = T(1-SF-CF) Contamination

50

0 500

Fig. 6

% Coincidences detected

550 600 650 Upper energy threshold (keV)

700

12 Scattered true and contaminated counts and noise equivalent count as a function of upper energy window threshold.

511 keV (600 keV upper) 511 keV (triple corrected)

10 8 6 4 2 0 0

5

15 20 10 Axial opening angle (degrees)

25

30

This curve shows the percent of coincidences detected in a PET scanner with different axial opening angle. The lower curve is for 124I and is without triple coincidences. The upper curve is for a conventional isotope (no contaminated coincidences possible).

where T is the true, S is scatter and C is the contamination. Similar to the NEC-2001 scatter fraction a line source filled with 124I was placed at a 4.5 mm radial offset from the centre of a solid polyethylene cylinder (diameter = 20 cm, length = 70 cm) with a water equivalent density of 1 g  ml – 1. The number of scattered, true and spurious coincidences and the NEC value is plotted as a function of the varying upper energy threshold in Fig. 7. Scatter and contamination after energy window optimization

The scatter and contamination fraction for the Allegro scanner (energy resolution of 15% and energy window of 434–600 keV) was simulated following the NU 2 2001 procedure. This uses a line source filled with 18F, placed at a 4.5 mm radial offset from the centre of a solid polyethylene cylinder (diameter = 20 cm, length = 70 cm)

with a water equivalent density of 1 g  ml – 1. The length of tube is 70 cm and a volume of approximately 5.7 ml. The data were acquired at low count rates and rebinned using single-slice rebinning [13]. The sinogram profile was used to calculate the number of scatter events within a diameter of 24 cm (4 cm larger than the phantom diameter) and the number of trues within a 2 cm radius of the source. The activity was chosen at 1 MBq to ensure a very small fraction of random coincidences. The line source was filled with the three different isotopes and information provided in the output allowed us to determine whether the detected event was true or scattered or contaminated. The scatter (+ contamination) fraction for the different isotopes is shown in Fig. 8. Two-dimensional versus three-dimensional PET

An interesting problem when using impure isotopes is whether to choose 2-D PET or 3-D PET. In most applications 3-D PET has become the preferred option because of its higher NEC values for the same activity. Here we determine the number of contaminated and scattered photons and the resulting NEC value (at low rates) for a 2-D and 3-D PET configuration. For the 3-D PET scanner we used the Allegro mode; the 2-D PET scanner was obtained by adding septa (in the simulation set-up) to this system. The number of true coincidences, scattered and contaminated coincidences was determined using the same 70 cm phantom. This is shown in Fig. 9. The sensitivity of the 2-D PET was normalized to 1 and the same normalization was used for the 3-D PET results.

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242 Nuclear Medicine Communications 2006, Vol 27 No 3

Count-rate performance

The count-rate performance in a whole-body imaging situation was also simulated. Coincidences were also classified in true, random, scattered and contaminated coincidences using the information in the Monte Carlo output. The randoms are formed by the GATE software that pairs singles that occur in the given coincidence window and energy window. It also includes an estimation of randoms. Spurious coincidences are treated equally as scattered coincidences and included in the calculation of the NEC. The same study was repeated for 18F, 124I, 86Y. Count rates between 0 and 20 MBq  ml – 1 were simulated. The results for 124I and 86Y are shown in Fig. 10 and

Scatter + contamination fraction

Fig. 8

0.5 0.4

F-18 I-124 Y-86

0.3 0.2 0.1

11, respectively. The number of singles (in the energy and coincidence window) is shown in Fig. 12.

Results Spatial resolution

Figures 2 and 3 summarize the spatial resolution simulations for the Allegro scanner. The spatial resolution at the centre is 4.8 mm in the transverse direction for 18F, which is in good agreement with recent publications [14] where a value of 4.9 mm was reported for a point in the centre. The FWTM is 9.4 mm. The same simulation with 124 I gives a FWHM of 6.1 mm and a FWTM of 12.4 mm. 86 Y gives slightly better results: FWHM of 5.7 mm and FWTM of 12 mm. In PET scanners with square detector pixels, the axial resolution is better than the resolution in plane. For this particular scanner, however, the detector pixel size is not square but rectangular: 4 mm in the radial direction and 6 mm in the axial direction. Therefore the resolution values are less good in this direction, as shown by Surti and Karp [7]. The resolution loss due to the increased positron range of 124I and 86Y is isotropic (positron emission is isotropic) and adds in quadrature to the axial resolution. As the axial resolution is worse (for this particular scanner) the relative change due to positron range compared to 18F will be smaller.

0

Isotope Scatter and contamination fraction for the different isotopes for an optimized energy window of 434–600 keV.

Sensitivity

The sensitivity is approximately proportional to the fraction of positron decays. An extra loss in sensitivity occurs due to the fact that triple coincidences can be

5 4.5 4 3.5 3 2.5 2 1.5 1 0.5 0

Scatter + contamination

20 PET

30 PET

5 4.5 4 3.5 3 2.5 2 1.5 1 0.5 0

NEC = T(1-SF-CF)

Trues

Fig. 9

5 4.5 4 3.5 3 2.5 2 1.5 1 0.5 0 20 PET

30 PET

Factor of 3.65 more effective counts at low rates 20 PET

30 PET

Scatter ( + spurious), true coincidences and noise equivalent count for 3-D and 2-D PET imaging of

124

I.

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3-D PET imaging with

Fig. 10

I and

86

Y Vandenberghe 243

Fig. 11

(a)

(a)

Trues F-18 Randoms F-18 Scatter F-18 Trues I-124 Randoms I-124 Scatter I-124

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80 60 40 20 0

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(b) 45 40 35 30 25 20 15 10 5 0

Trues F-18 Randoms F-18 Scatter F-18 Trues Y-86 Randoms Y-86 Scatter Y-86

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Count rate curves obtained by Monte Carlo simulations of 18F and 124I.

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Concentration (kBq/ml) Count rate curves obtained by Monte Carlo simulations of 18F and 86Y.

Optimization of upper energy threshold for

124

I

From Fig. 7 it is clear that scatter increases continuously with an increase in upper energy threshold while contaminated coincidences increase more steeply above 575 keV. Overall, the behavior of the NEC is a curve with a smooth maximum around 600 keV. The smooth maximum suggests that the difference in image quality for images acquired at different upper energy thresholds will be moderate. Scatter and contamination after energy window optimization

The scatter fraction from 18F (Fig. 8) is about 30%. This is clearly lower than the combined scatter and contamination fraction of the other two isotopes. The scatter ( + contamination) fraction is simulated for 124I (32.5%) and 86Y (35.5%).

Fig. 12

Singles rate after deadtime + energywindow 14 Count-rate (Mcps)

measured. For whole-body PET scanners (similar to Allegro and with an optimized energy window) the fraction of triple coincidences (124I) is determined by Monte Carlo simulations to be only 4% of the total detected coincidences. The curve in Fig. 6 shows the expected behaviour of scanners with a larger axial opening angle. The probability of triple coincidences increases clearly with the solid angle.

12 10 8 6 F-18 I-124 Y-86

4 2 0 0

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10 15 Activity (kBq/ml)

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The number of singles in the energy and coincidence window.

Two-dimensional versus three-dimensional PET

In Fig. 9 shows that the sensitivity of 3-D PET is a factor of 4.7 higher than 2-D PET (same scanner, but with axial septa). While the fraction of contamination coincidences is clearly higher in 3-D, the NEC (including contamination) is still significantly higher (3.65) than for 2-D PET.

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Count-rate performance

Figures 10 and 11 show the results of the count-rate simulations. The peak NEC for 18F is 37 kcps. Due to the lower positron abundance the peak NEC is lower: 14 kcps for 124I and 16.5 kcps for 86Y. It is also interesting to note that the NEC for 124I and 86Y peaks at higher activities than for 18F. The number of randoms is not high for 124I and 86Y. As randoms are proportional to the square of the singles a plot (Fig. 12) of the singles rate versus activity for the different isotopes is drawn. As shown, the number of singles hitting the detector, and which are in the energy and coincidence window, is lower than for 18F (same activity).

Discussion Impure positron emitters with associated singles (dirty isotopes) require good performance from 3-D PET scanners. The energy resolution has always been a key parameter in 3-D PET as it allows a reduction of the scatter fraction. Typically, whole-body PET scanners have an energy resolution in the range 15–25%, resulting in scatter fractions of about 30–45% [12] (3-D PET). To minimize the contribution of the spurious coincidences it becomes even more important to have good energy resolution. The most recent 3-D PET scanners are capable of 15% energy resolution after advanced calibration. It is expected that scintillators with even better energy resolution will become available. A prototype 3-D PET scanner with LaBr3 (6–8% energy resolution) is now being developed at the University of Pennsylvania [15]. With such an energy resolution it will be possible to eliminate even more spurious coincidences. Recent work [16] has shown that the current techniques of scatter background subtraction [17] eliminate the background due to spurious coincidences quite efficiently. Another important factor is the axial acceptance angle of 3-D PET scanners. The probability of triple coincidences increases clearly with the solid angle, but the sensitivity increases also. In human PET there has been a trend to extend the scanners with more axial coverage. Compared to other available isotopes (e.g., 123I), 124I has the disadvantage of its lack of purity. However, the sensitivity of 2-D PET is already clearly higher (about factor 10) than of SPECT. By going to 3-D PET the sensitivity increases by another factor of 5. This should allow whole-body tomography to be performed in a reasonable time frame. Also, the spatial resolution is clearly better for PET (5 mm) compared to SPECT (8 mm). In PET some progress has been made towards higher sensitivity by using more axial rings. Recently, there is a new development of time-of-flight PET systems [15]. Due to the time-of-flight information (Gaussian distribution instead of uniform distribution along a line) the same number of measured counts will lead to an improved image quality for larger objects. So,

for large objects time-of-flight PET will lead to a higher effective sensitivity. On the other hand it is still not clear how to gain an important factor of sensitivity in SPECT. The situation is different for small-animal PET systems. Solid angle coverage is larger than for human PET scanners resulting in a clearly higher sensitivity. Because of the small amount of scatter in small animals the energy resolution of small-animal PET systems is not optimized as in human PET scanners. Energy windows are rather large in order to maximize sensitivity. For dirty isotopes the larger solid angle and the poorer energy resolution will lead to an even higher fraction of triple coincidences and spurious coincidences. The fact that the NEC peaks at a higher concentration can be used to increase the image quality. Higher activities can be injected to obtain more counts in a given time, but as the maximum NEC for 124I or 86Y is lower than the peak NEC for 18F this is not sufficient. Longer imaging times will be needed to compensate for the low positron abundance. The dose delivered for the same activity of 124I or 86Y is clearly higher than for the same dose of 18F. Therefore, because of dose limitations, the administration of activities in the range of maximal NEC will not be allowed in patients who do not require radionuclides for therapeutic purposes. For patients receiving radionuclide therapy it may be more acceptable to increase the dose to obtain higher image quality.

Conclusion The upper energy window of a simulated 3-D PET scanner has been optimized for 124I to 600 keV using the NEC parameter. It is shown that with this optimal setting 3-D PET imaging of 124I and 86Y leads to a small increase of the scatter fraction with spurious coincidences (from 30% to 32.5% for 124I and 35.5% for 86Y). The main problem with 124I and 86Y imaging is the lower positron abundance, resulting in a lower number of coincidences detected for a given activity. Therefore it is even more important to use 3-D PET, which has a high sensitivity. The activity has to be increased by a factor somewhat higher than the inverse of the positron abundance to obtain similar background noise as with 18 F. It has to be a little higher to compensate for the noise contribution from the spurious coincidences and the small degradation in spatial resolution. In clinical studies biological uptake is also important in determining image quality. Count rate curves show that it is possible to scan at higher activities than for 18F. The peak NEC values (14 kcps for 124I and 16.5 kcps for 86Y) are, of course, lower due to the low positron abundance but are still high

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3-D PET imaging with

enough to obtain a sufficient number of counts in a reasonable time.

7 8

Acknowledgements We would like to thank Suleman Surti and Joel Karp from the University of Pennsylvania, and Daniel Gagnon, Jeff Kolthammer and Nicholas Salem from Philips Medical Systems, Bernd Schweizer and Heinrich von Busch from Philips Research Aachen for exchange of results and useful discussions on this topic.

References 1 2

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Larson SM, Krenning EP. A pragmatic perspective on molecular targeted radionuclide therapy. J Nucl Med 2005; 46:1S–3S. Autret D, Bitar A, Ferrer L, Lisbona A, Bardie`s M. Monte Carlo Modeling of gamma cameras for 131I imaging in targeted radiotherapy. Cancer BiotherRadiopharm 2005; 20:77–84. Bice AN, Eary JF, Nelp WB. Quantification of iodine-131 distribution by gamma camera imaging. Eur J Nucl Med 1991; 18:142. Verel I, Visser GWM, van Dongen GA. The promise of immuno-PET in radioimmunotherapy. J Nucl Med 2005; 46:164S–171S. Townsend DW, Carney JPJ, Yap JT, Hall NC. PET/CT today and tomorrow. J Nucl Med 2004; 45:4S–14S. Beattie BJ, Finn RD, Rowland DJ, Pentlow KS. Quantitative imaging of bromine-76 and yttrium-86 with PET: A method for the removal of spurious activity introduced by cascade gamma rays. Med Phys 2003; 30: 2410–2423.

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Original article

The effect of verapamil on the restoration of myocardial perfusion and functional recovery in patients with angiographic no-reflow after primary percutaneous coronary intervention Shigeo Umemuraa, Seishi Nakamuraa, Tetsuro Sugiurab, Yoshiaki Tsukaa, Keisuke Fujitakaa, Susumu Yoshidaa, Masato Badena and Toshiji Iwasakac Objective Angiographic thrombolysis in myocardial infarction (TIMI) flow grade r 2 after primary percutaneous coronary intervention (PCI), defined as angiographic no-reflow, predicts poor functional recovery in patients with acute myocardial infarction. We investigated the effect of verapamil on the restoration of myocardial perfusion and functional recovery in patients with angiographic no-reflow after PCI. Methods 99mTc tetrofosmin single photon emission computed tomographic (SPECT) imaging was performed (before, immediately after and 1 month after PCI) in 101 consecutive patients with acute myocardial infarction. The defect score was calculated as the sum of perfusion defect in a 13-segment model (scores of 3, complete defect to 0, normal perfusion). The asynergic score, defined as the number of asynergic segments, was assessed by echocardiography before and 1 month later. Multiple logistic regression analysis was performed to elucidate the effect of verapamil administration. Results Of 101 patients, 32 (31%) had angiographic no-reflow and were divided into two groups: 18 patients with verapamil (group 1) and 14 patients without verapamil (group 2). Sixty-nine patients had TIMI grade 3 reflow after PCI (group 3). The change in the defect score 1 month after PCI in group 1 was significantly larger than that in group 2 (P = 0.003). The asynergic score improved more at 1 month in group 1 compared to that in group 2 (P = 0.007).

Introduction The mechanical restoration of epicardial coronary artery obstruction is the goal of primary percutaneous coronary intervention (PCI) in acute myocardial infarction. However, profound and broad microvascular damage in the reperfused myocardium after reperfusion therapy is increasingly recognized after primary PCI for acute myocardial infarction. Reduced epicardial flow (angiographic thrombolysis in myocardial infarction (TIMI) flow grade r 2) after primary PCI, without any mechanical causes of flow reduction, is defined as angiographic no-reflow [1]. A recent study reported that angiographic no-reflow predicts poor left ventricular functional recovery and survival in patients with acute myocardial infarction [2,3].

Moreover, logistic regression analysis revealed that TIMI grade reflow r 2 after PCI (P = 0.04, OR = 5.51), the defect score before PCI (P = 0.03, OR = 1.15), the asynergic score before PCI (P = 0.01, OR = 0.64) and the administration of verapamil (P = 0.002, OR = 22.4) were independently associated with successful myocardial reperfusion immediately after PCI. Conclusions Intracoronary verapamil restored myocardial perfusion in patients with angiographic no-reflow after PCI and lead to better functional recovery after acute myocardial infarction. Nucl Med Commun 27:247–254
c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:247–254 Keywords: technetium-99m tetrofosmin, acute myocardial infarction, percutaneous transluminal coronary intervention, angiographic no-reflow phenomenon, verapamil a

Division of Cardiology, Takarazuka Hospital, Hyogo, bDepartment of Laboratory Medicine, Kochi Medical School, Kochi and cThe Cardiovascular Center, Kansai Medical University, Osaka, Japan. Correspondence to Dr Seishi Nakamura, The Division of Cardiology, Takarazuka Hospital, 2-1-2 Nogami, Takarazuka, Hyogo, 665-0022, Japan. Tel: + 0081 797 71 3111; fax: + 0081 797 76 3161; e-mail: [email protected] This work was partially funded by a grant-in-aid of Hyogo Prefectural Medical Association (Hyogo, Japan). Received 11 October 2005 Accepted 25 November 2005

When focusing on both the epicardial coronary artery and microvasculature, there is a need for safe and effective treatment for no-reflow. Effective treatment has a clinical impact if it can be applied to acute myocardial infarction in the emergency department. Recently, it has been reported that the administration of verapamil attenuated ischaemic microvascular damage after PCI in patients with angina pectoris or acute myocardial infarction [1,4]. However, there are no systematic studies evaluating the effect of intracoronary verapamil as a rescue strategy for angiographic no-reflow. The prompt assessment of both coronary anatomy and the quality of microvascular perfusion by serial scintigraphic

c 2006 Lippincott Williams & Wilkins 0143-3636


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imaging can aid the diagnosis of microvascular dysfunction and decision making in patients with primary PCI [5–10]. The aim of this study was to determine the effect of verapamil on the restoration of myocardial perfusion and the recovery of left ventricular wall motion in patients with angiographic no-reflow after primary PCI by serial 99mTc tetrofosmin imaging.

Materials and methods Study patients

The study included 136 consecutive patients with their first acute myocardial infarction admitted to our hospital between 1 January 1996 and 31 December 2002 who had 99m Tc tetrofosmin imaging before PCI and met the following criteria: (1) at least 30 min of chest pain; (2) ST segment elevation ( Z 0.1 mV from TP segment) in at least two contiguous leads in the same vascular territory; (3) more than a 2-fold increase in serum levels of creatine kinase; and (4) a successful primary PCI, defined as < 50% diameter stenosis of the infarct-related artery after primary PCI. Thirty-five patients were excluded: (1) five patients in an unsuitable condition (cardiogenic shock) for 99mTc tetrofosmin imaging before primary PCI; (2) 22 patients with TIMI flow grade 3 (spontaneous recanalization) at the time of emergency coronary angiography; (3) six patients unable to receive 99mTc tetrofosmin; and (4) two patients with re-occlusion of the infarct-related artery at a 1 month follow-up angiography. The remaining 101 patients formed the study group: 79 males and 22 females (mean age 65 ± 12 years). The culprit lesion was the left anterior descending coronary artery in 37 patients, the left circumflex coronary artery in 12 patients and the right coronary artery in 52 patients. Seventy-one patients had one-vessel disease, 22 had two-vessel disease and eight had three-vessel disease. The time from the onset of chest pain to primary PCI was 6 ± 4 h (range 2–14 h) and the time from admission to primary PCI was 68 ± 5 min (range 41–90 min). TIMI grade before primary PCI was 0 in 60 patients, 1 in 13 and 2 in 28 [10]. All the patients in this study had sufficient dilatation of the infarct-related artery just after primary PCI without significant restenosis as confirmed by repeat coronary angiography 1 month after primary PCI. SPECT studies

All patients were given a chewable baby aspirin tablet (81 mg) and 3000 units of intravenous heparin and transported promptly to the nuclear medicine laboratory. Initial 99mTc tetrofosmin imaging was performed during the preparation of the catheterization next to the nuclear medicine laboratory. The first tomographic imaging was obtained 15 min after the injection of 99mTc tetrofosmin (370 MBq). To reduce the effect of residual activity from the initial study, 740 MBq of 99mTc tetrofosmin (twice the initial dose) was intravenously administered at the end of the angiographic procedure and the second tomographic image was acquired. The third 99mTc

tetrofosmin imaging procedure was performed 1 month after primary PCI (Fig. 1). Single photon emission computed tomography (SPECT) was performed using a large-field-of-view gamma camera with a high resolution, parallel hole collimator (Shimadzu SNS 400S), rotated 1801 around the long axis of each patient. Thirty-six views every 51 were obtained for 25 s from the 451 left posterior oblique to the 451 right anterior oblique. After correction for non-uniformity and the centre of rotation, images were reconstructed into long-axis and short-axis cuts [11–13]. The study protocol was approved by the Takarazuka Hospital ethics committee on human research. Informed consent was obtained from all patients. Angiographic study

Primary PCI was performed by conventional techniques after the intravenous administration of 7000 units of heparin. The culprit lesion ( > 50% stenosis of infarctrelated artery) was successfully dilated in all patients. Coronary flow of the infarct-related artery before and after primary PCI was graded visually according to the TIMI Study group flow classification. Collateral flow before PCI was graded visually using a classification proposed by Rentrop et al. [14]. Grade 0 collateral flow was defined as the absence of visible collaterals, grade 1 as the filling of side branches only, grade 2 as the filling of side branches and a portion of the main epicardial artery and grade 3 as complete filling of side branches and the epicardial artery beyond the point of the occlusion. Adequate collateral flow was considered present when it was graded 2 or 3. The coronary angiogram was carefully reviewed by two experienced investigators who were unaware of other clinical data. The time from the onset of chest pain to primary PCI was calculated as the time between the onset of chest pain and the first balloon inflation. Coronary angiography was repeated 1 month after primary PCI to estimate the patency of the infarctrelated artery. Fig. 1

Before PCI

Immediately after PCI

Tc-99 m tetrofosmin Tc-99 m tetrofosmin 370 MBq 740 MBq Acute MI

Imaging

Primary PCI

Echocardiography

Imaging

1 month after PCI Tc-99 m tetrofosmin 740 MBq Imaging Echocardiography

99m Tc tetrofosmin imaging protocol. The first study was performed before primary percutaneous coronary intervention (PCI). Images were obtained 15 min after the injection of 99mTc tetrofosmin (370 MBq). Twice the initial dosage of 99mTc tetrofosmin (740 MBq) was administered at the end of angiographic procedure, and the second imaging study was performed. The third study was performed 1 month after primary PCI.

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Verapamil and myocardial salvage after PCI Umemura et al. 249

Analysis of SPECT image

SPECT images were divided into 13 segments: the shortaxis images were obtained at middle and lower ventricular levels, and were divided into six segments. The apex of the left ventricle was obtained from the vertical long-axis image. 99mTc tetrofosmin uptake in each of the 13 segments was graded as follows: 3, complete defect; 2, severely reduced perfusion; 1, mildly reduced perfusion; and 0, normal perfusion. The defect score was calculated as the sum of the perfusion defects. The tomogram was visually interpreted by two independent observers who were unaware of the clinical and angiographic data. Successful myocardial reperfusion immediately after primary PCI was defined as a change (before minus after PCI) of Z 4 in the defect score. Because the mean absolute inter-observer variability in the defect score was 2.0 ± 0.8, an absolute change of Z 4 (mean + 2  standard deviation) was considered to represent a significant difference between the two measurements using the cut-off value of score 4 [11,13]. In addition, the myocardial salvage index (% change in defect score = defect score before minus 1 month after PCI/defect score before PCI  100) was calculated in each patient. Echocardiographic study

Two-dimensional echocardiographic analysis of regional wall motion was performed on admission and 1 month after primary PCI with a Hewlett-Packard Sonos 2500 equipped with a 2.5 MHz transducer. Parasternal longaxis, short-axis and apical two-chamber views were analysed by an experienced echocardiographer who had no knowledge of the clinical and scintigraphic data and were used to assess the regional wall motion at the middle ventricular level, the lower ventricular level and at the apex. The echocardiographic images were divided into 13 segments representing anteroseptal, anterior, lateral, inferolateral (posterior), inferior and inferoseptal walls. The asynergic grade was calculated on a semiquantitative scale from 0 to 3, in increments of 1, with normal wall motion scored as 0 and akinetic/dyskinetic wall motion scored as 3. The asynergic score was defined as the number of segments with advanced asynergy (the asynergic grade Z 2; severe hypokinetic or akinetic). Intracoronary administration of verapamil

Angiographic no-reflow was defined as TIMI flow grade r 2 after primary PCI without any mechanical causes of flow reduction such as apparent epicardial coronary dissection, thrombosis or distal vessel cut-off suggestive of macroembolization [1–3]. When angiographic no-reflow was judged to be present by the consensus of two experienced cardiologists, the initial therapy consisted of a repeat intracoronary dose (200–400 mg) of nitroglycerin. We started the routine intracoronary administration of verapamil as a rescue strategy for angiographic no-reflow on 1 January 1998. Therefore, patients with angiographic no-reflow were divided into two groups: those without (admitted from 1996

to 1997) and those with (from 1998 to 2002) verapamil. Intracoronary verapamil was given until the restoration of epicardial flow as previously described. Briefly, verapamil (5 mg) was diluted with a saline solution to a total volume of 5 ml (1 mg  ml – 1). One millilitre of this solution was further diluted to a total volume of 10 ml (100 mg  ml – 1) and was administered through the guiding catheter or central lumen of the dilating balloon after removal of the guide wire [1]. Intracoronary verapamil administration was stopped when new Z 21 atrio-ventricular block or systemic hypotension (systolic arterial pressure < 90 mmHg) was observed. A temporary pacing was immediately performed in a case of Z 21 atrio-ventricular block. Patients with TIMI grade 3 reflow after primary PCI during the same study period (1996–2002) served as the control group. Statistical analysis

Results are expressed as mean ± standard deviation. Statistical analyses among the three groups were performed by 1-way layout analysis of variance or chi-squared analysis followed by a Scheffe-type multiple comparison method. Changes in the defect score were estimated by 2-way repeated measures ANOVA. Multivariate logistic regression analysis was performed to evaluate the important variables related to the amount of myocardial salvage immediately after primary PCI. A probability value of < 0.05 was considered significant.

Results Patient characteristics

Angiographic no-reflow after primary PCI was observed in 32 patients (31%) and were divided into two groups: 18 patients treated with intracoronary verapamil (group 1) and 14 patients without verapamil (group 2). Sixty-nine patients had TIMI grade 3 reflow after PCI (group 3). Comparison of clinical characteristics is summarized in Table 1. There were no significant differences among the three groups in age, gender, infarct-related artery, the number of diseased vessels, the time from the onset of chest pain to primary PCI and TIMI flow grade before PCI or collateral grade before PCI. There was no significant difference in the frequency of stent usage among the three groups. None of our patients received glycoprotein IIb/IIIa blockers, clopidgrel, nicorandil or thrombolytic therapy. There was no significant difference in the frequency of beta blocker therapy among the three groups (five patients in group 1, four in group 2 and seven in group 3, P = 0.705). Management of angiographic no-reflow

Of the 18 patients in group 1, 11 had TIMI grade flow 2 and seven had TIMI grade flow 1 after primary PCI. The average dosage of intracoronary verapamil was 277 ± 142 mg (100–600 mg). Intracoronary verapamil increased TIMI flow grade in 16 patients (88%) and TIMI grade flow 3 was obtained in 14 patients (77%) (Fig. 2). TIMI flow grade did not improve in two patients because

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Table 1

Comparison of clinical characteristics

Gender (male/female) Age (years) Infarct-related artery (LAD/LCX/RCA) Number of diseased vessels (1/2/3) Time to primary PCI (h) TIMI flow grade before PCI (0/1/2) Collateral grade (present/absent) Stent usage (%) Beta-blocker usage (%)

Group 1 n = 18

Group 2 n = 14

Group 3 n = 69

P value

15/3 66 ± 11 7/0/11 10/6/2 9±6 12/2/4 9/9 8 (44) 5 (28)

11/3 68 ± 8 6/2/6 9/3/2 9±6 9/3/2 9/5 2 (17) 4 (29)

53/16 63 ± 13 32/10/35 52/13/4 8±6 39/8/22 44/25 13 (18) 7 (10)

0.83 0.32 0.48 0.46 0.62 0.60 0.96 0.05 0.71

Group 1, TIMI r 2 patients with verapamil; group 2, TIMI r 2 patients without verapamil; group 3, patients with TIMI 3 grade reflow; LAD, left anterior descending coronary artery; PCI, percutaneous coronary intervention; Time to primary PCI, time from onset of chest pain to primary percutaneous coronary intervention; TIMI flow grade, Thrombolysis in Myocardial Infarction flow grade; LCX, left circumflex coronary artery; RCA, right coronary artery.

Fig. 2

Fig. 3

9

TIMI 1 (4) TIMI 0 (11) Before PCI

2 1 3 1

TIMI 1 (11)

5

2 2

TIMI 1 (4)

TIMI 1 (7)

6 5 After PCI

20

Defect score

TIMI 2 (3)

25

TIMI 3 (14)

15

S S

10

After verapamil

Thrombolysis in myocardial infarction (TIMI) flow grade after intracoronary administration of verapamil.

5

0

of the cessation of intracoronary verapamil injection due to systemic hypotension (one patient) or 21 atrioventricular block requiring temporary pacing (one patient). These two patients in group 1 and nine of 14 patients in group 2 were assisted by intra-aortic balloon counter-pulsation after PCI.

Before PCI

Immediately after PCI

1 month after PCI

Serial changes in the defect score after primary PCI. Unfilled circles, TIMI r 2 patients with verapamil (group 1); filled circles, TIMI r 2 patients without verapamil (group 2); filled squares, patients with TIMI 3 grade reflow (group 3). zP < 0.01 compared with group 2. (Abbreviations as in Figs 1 and 2.)

Serial change in defect score after PCI

A sufficient quality of SPECT image was obtained in all 101 patients. The defect score decreased significantly after primary PCI (before PCI = 13.9 ± 4.4, immediately after PCI = 10.4 ± 4.8 and 1 month after PCI = 7.2 ± 4.8, P < 0.0001) in all patients. A serial change in the defect score after primary PCI is shown in Fig. 3. There was no significant difference in the defect score before primary PCI among the three groups (group 1 = 13.3 ± 3.9, group 2 = 16.1 ± 5.0 and group 3 = 13.7 ± 4.6, P = 0.14). The defect score immediately after PCI in group 1 was significantly smaller than that in group 2 (9.5 ± 3.6 versus 14.5 ± 5.1, P = 0.011). Furthermore, the final infarct size, expressed as the defect score 1 month after primary PCI, was significantly smaller in group 1 than that in group 2 (7.5 ± 4.3 vs. 12.8 ± 4.9, P = 0.003). Moreover, the defect

score immediately after primary PCI and 1 month after primary PCI in group 1 was identical to that in group 3 (defect score = 9.8 ± 4.7, P = 0.77, 6.2 ± 4.2, P = 0.84, respectively). A significantly greater myocardial salvage index was observed in group 1 compared to group 2 (45.6 ± 19.8% vs. 21.2 ± 24.8%, P = 0.004), whereas the myocardial salvage index in group 1 was not significantly different from that in group 3 (57.4 ± 23.7%, P = 0.057, respectively). Serial change in asynergic score after PCI

The change in the asynergic score after primary PCI is shown in Fig. 4. Although there was no significant difference in the asynergic score before primary PCI among the three groups (group 1 = 4.2 ± 1.3, group

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Verapamil and myocardial salvage after PCI Umemura et al. 251

2 = 4.4 ± 1.5 and group 3 = 3.5 ± 1.2, P = 0.09), the asynergic score 1 month after PCI in group 1 was significantly smaller than that in group 2 (1.2 ± 1.3 versus 3.3 ± 1.7, P = 0.007). Moreover, the asynergic score 1 month after primary PCI in group 1 was identical to that in group 3 (asynergic score = 0.8 ± 1.4, P = 0.07, respectively). Multivariate analysis

To determine the important variables to the amount of myocardial salvage after primary PCI, we performed multiple logistic regression analysis. As a result, the defect score before primary PCI, the asynergic score before primary PCI, TIMI grade reflow r 2 after primary PCI and verapamil administration were independent predictors of successful myocardial reperfusion immediately after primary PCI (Table 2).

Fig. 4

6 5

Asynergic score

4

Discussion Angiographic no-reflow phenomenon after primary PCI has been demonstrated to predict poor left ventricular functional recovery, a higher cardiac mortality in the acute phase of myocardial infarction [2] and a higher incidence of long-term cardiac complications [3]. Angiographic noreflow phenomenon is associated with profound and broad microvascular damage in the reperfused myocardium [1,4,15]. A serial 99mTc tetrofosmin study from our laboratory clarified that angiographic no-reflow was a highly specific marker predicting impaired myocardial reperfusion and suboptimal myocardial salvage in the early phase of acute myocardial infarction after primary PCI [5,11]. The defect score before primary PCI was one of the predictors of successful myocardial reperfusion by multivariate analysis. Although the defect score before primary PCI indicates the area at risk, it includes a mildly reduced perfusion area (grade 1). Therefore, when evaluating the amount of myocardial salvage immediately after primary PCI, a high defect score before primary PCI is associated with a larger decrease in the defect score. In contrast, the higher asynergic score before primary PCI, defined as the number of asynergic (severe hypokinetic or akinetic) segments, predicts impaired myocardial reperfusion. These data confirm that the asynergic score before primary PCI is well correlated with the infarct size after primary PCI [7,16,17].

3

S S

2 1 0 −1

Before PCI

1 month after PCI

Serial changes in the asynergic score after primary PCI. Unfilled circles, TIMI r 2 patients with verapamil (group 1); filled circles, TIMI r 2 patients without verapamil (group 2); filled squares, patients with TIMI 3 grade reflow (group 3). zP < 0.01 compared with group 2. (Abbreviations as in Figs 1 and 2.)

Table 2

Several pharmacological interventions have attempted to attenuate ischaemic microvascular dysfunction during primary PCI. Among them, verapamil has a potent vasodilative effect of the distal microcirculation and is also beneficial in preserving post-ischaemic myocardial function from impaired microvascular dysfunction [1,4]. Angiographic no-reflow, refractory to nitroglycerin, responded promptly to intracoronary verapamil, which indicates that a flow-restricting spasm of the distal microvasculature after coronary intervention might have been responsible for angiographic no-reflow in our patients. In a previous clinical study, Piana et al. [1] demonstrated that intracoronary verapamil improved TIMI flow grade after coronary intervention in about 90% of patients with

Multivariate predictors of successful myocardial reperfusion immediately after percutaneous coronary intervention

Defect score before PCI Asynergic score before PCI Time to primary PCI TIMI flow grade r 2 flow before primary PCI TIMI flow grade r 2 reflow after primary PCI Infarct related artery Verapamil treatment Stent usage

Chi-squared

P value

Odds ratio

95% CI

4.578 6.910 1.027 0.226 3.982 1.860 9.433 0.001

0.032 0.009 0.305 0.635 0.046 0.173 0.002 0.974

1.145 0.637 1.080 1.299 0.182 0.248 22.42 0.981

1.01–1.30 0.45–0.89 0.93–1.25 0.44–3.82 0.03–0.97 0.03–1.84 3.08–16.3 0.29–3.28

CI, confidence interval; PCI, percutaneous coronary intervention; Time to primary PCI, time from onset of chest pain to primary percutaneous coronary intervention; TIMI flow grade, Thrombolysis in Myocardial Infarction flow grade.

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252 Nuclear Medicine Communications 2006, Vol 27 No 3

angiographic no-reflow. However, more than 70% of their patients in their study did not have acute myocardial infarction. It is important to evaluate the effect of verapamil on the restoration of coronary flow after primary PCI in patients with acute myocardial infarction, because there is a much higher incidence of angiographic no-reflow and large myocardial damage in acute myocardial infarction (12–25%) [1,2] compared to angina patients (1.5%). In our study, intracoronary administration of verapamil improved TIMI flow grade in 88% with angiographic no-reflow and TIMI grade flow 3 was obtained in 14 patients (77%). Moreover, intracoronary administration of verapamil was the most important predictor of successful myocardial reperfusion by multivariate analysis. These data indicate that intracoronary administration of verapamil augmented coronary blood flow in patients with angiographic no-reflow after primary PCI. Taniyama et al. [4] investigated the acute effect of intracoronary injection of verapamil on microvascular function after primary PCI for acute myocardial infarction with myocardial contrast echocardiography and demonstrated that verapamil attenuated post-ischaemic microvascular dysfunction, leading to augmentation of myocardial blood flow in the reperfused area. However, the effect of intracoronary verapamil on myocardial reperfusion in patients with angiographic no-reflow after primary PCI was not fully investigated because the number of patients (six of 20 patients, 30%) with angiographic no-reflow was relatively small. In this study, a better left ventricular functional recovery (greater improvement of the asynergic score), a larger amount of myocardial salvage (decrease in the defect score immediately after primary PCI) and a smaller final infarct size (smaller defect score 1 month after primary PCI) due to the restoration of coronary blood flow (improvement of TIMI flow grade) were observed by verapamil despite angiographic no-reflow. Several parameters have been proposed for the assessment of reperfusion success in patients with acute myocardial infarction. Parameters that reflect flow restoration of both large epicardial arteries (TIMI flow grade) and microcirculation, an important determinant of myocardial salvage, are being investigated with increasing interest. Based on this concept, several diagnostic techniques such as myocardial contrast echocardiography [15,18], Doppler flow wire [19], myocardial blush grade [20] and corrected TIMI frame count [21,22] have been employed to evaluate tissue-level microvascular perfusion in patients with acute myocardial infarction undergoing reperfusion therapy. A prompt assessment of coronary anatomy, the quality of microvascular perfusion and myocardial viability, may aid in decision making in individual patients. 99mTc tetrofosmin imaging after reperfusion therapy can directly estimate microvascular integrity and myocardial viability of the reperfused area

[11,23,24]. Serial SPECT imaging with 99mTc tetrofosmin during acute myocardial infarction can provide quantitative assessment of the area at risk, the infarct size and myocardial salvage [6–8,11–13]. Several studies have shown that myocardium at risk was estimated after reperfusion therapy because myocardial perfusion at the time of injection can be frozen. However, recent studies reported that redistribution of 99mTc labeled compounds was detected in reperfused myocardium in patients with acute myocardial infarction following thrombolytic therapy [7,25]. Therefore, baseline SPECT acquisition (identification of area at risk) was performed prior to primary PCI in this study. 99mTc tetrofosmin is supplied in a kit form and requires only 15 min incubation at room temperature. Furthermore, compared to 99mTc sestamibi, its rapid accumulation in the myocardium and relatively rapid clearance from the background organs enable imaging as early as 5–15 min after injection [26–28]. Using this advantage, acquisition before primary PCI could be performed safely without time delay. As a result, time between hospital admission to primary PCI in this study was nearly identical to the report by de Boer et al. and Grines et al. at a 24 h coverage interventional cardiology department [29,30] and no deterioration during imaging was observed in all patients. Although assessment of myocardial salvage after reperfusion therapy is essential to determine the relation between coronary intervention and clinical outcome [4,15], the criteria identifying the lack of myocardial salvage has not been well defined. We previously reported that absolute change of defect score Z 4 before and immediately after PCI could discriminate between success and failure of myocardial salvage [11,13]. Therefore, we used these scintigraphic criteria to evaluate the salutary effect of pharmacological treatment in the presence of microvascular dysfunction. In this study, serial 99m Tc tetrofosmin imaging showed a greater improvement of myocardial perfusion immediately after primary PCI by the administration of verapamil in patients with angiographic no-reflow, which indicates that 99mTc tetrofosmin is a useful method to promptly predict functional recovery after acute myocardial infarction. Three limitations of this study should be addressed. First, we used an activity ratio of 1 : 2 (first vs. second) with shorter imaging time interval (1–2 h) between a set of imagings during primary PCI in this study. The residual activity of the first image may have superimposed on the second image and a true second image was not necessarily obtained because of relative smaller difference of radioactive tracer dosage. However, images of sufficient quality were obtained in all 101 patients and a significant change in the defect score was observed during primary PCI. Moreover, we have reported that our study protocol could identify a group of patients with impaired myocardial perfusion immediately after primary PCI and a smaller

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Verapamil and myocardial salvage after PCI Umemura et al. 253

amount of myocardial salvage could lead to a larger infarct size and poorer left ventricular function [11]. Therefore, we believe that our imaging protocol was a useful method to evaluate perfusion change during primary PCI. Second, verapamil administration for angiographic no-reflow was not randomized in this study. The beneficial effect of verapamil to restore antegrade flow in patients with acute myocardial infarction was first reported in 1997 [4]. Since then we have used verapamil routinely as an adjunctive treatment in the presence of angiographic no-reflow. Therefore, we compared two groups of consecutive patients with angiographic no-reflow: before the use of verapamil (1996–1997) and after the routine use (1998–2002). Despite no significant differences in other technical devices and clinical characteristics, intracoronary verapamil improved the grade of TIMI flow and decreased the defect score immediately after and 1 month after primary PCI in patients with angiographic no-reflow. Moreover, multivariate analysis documented that intracoronary administration of verapamil was a best predictor of myocardial salvage. Third, accurate diagnosis of angiographic no-reflow is difficult. The reduction of epicardial flow due to dissection, thrombosis or distal vessel cut-off suggestive of macro-embolization could not be completely eliminated because of variability in visual assessment of TIMI flow grade.

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In conclusion, intracoronary verapamil administration restored myocardial perfusion in patients with angiographic no-reflow after primary PCI and lead to better functional recovery in the convalescence phase of acute myocardial infarction. Serial SPECT imaging using 99mTc tetrofosmin is a useful method to promptly evaluate the salutary effect of intracoronary verapamil in patients with angiographic no-reflow.

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Acknowledgements The authors wish to thank NIHON Mediphysics for technological support. We acknowledge the assistance of Norio Sugimoto in statistical analysis and the secretarial assistance provided by Yoshiko Miura.

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References 1

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Piana RN, Paik GY, Moscucci M, Cohen DJ, Gibson CM, Kugelmass AD, et al. Incidence and treatment of ‘no-reflow’ after percutaneous coronary intervention. Circulation 1994; 89:2514–2518. Morishima I, Sone T, Okumura K, Tsuboi H, Kondo J, Mukawa H, et al. Angiographic no-reflow phenomenon as a predictor of adverse long-term outcome in patients treated with percutaneous transluminal coronary angioplasty for first acute myocardial infarction. J Am Coll Cardiol 2000; 36:1202–1209. Morishima I, Sone T, Mokuno S, Taga S, Shimauchi A, Oki Y, et al. Clinical significance of no-reflow phenomenon observed on angiography after successful treatment of acute myocardial infarction with percutaneous transluminal coronary angioplasty. Am Heart J 1995; 130:239–243. Taniyama Y, Ito H, Iwakura K, Masuya T, Masatsugu H, Takiuchi S, et al. Beneficial effect of intracoronary verapamil on microvascular and myocardial salvage in patients with acute myocardial infarction. J Am Coll Cardiol 1997; 30:1193–1199. Nakamura S, Takehana K, Sugiura T, Hatada K, Hamada S, Asada J, et al. Quantitative estimation of myocardial salvage after primary percutaneous

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transluminal coronary angioplasty in patients with angiographic no reflow. Eur J Nucl Med Mol Imaging 2003; 30:383–389. Wackers FJT, Gibbons RJ, Verani MS, Kayden DS, Pellikka PA, Behrenbeck T, et al. Serial quantitative planar technetium-99m isonitrile imaging in acute myocardial infarction: efficacy for noninvasive assessment of thrombolytic therapy. J Am Coll Cardiol 1989; 14:861–873. Christian TF, Schwartz RS, Gibbons RJ. Determinants of infarct size in reperfusion therapy for acute myocardial infarction. Circulation 1992; 86:81–90. Clements IP, Christian TF, Higano ST, Gibbons RJ, Gersh BJ. Residual flow to the infarct zone as a determinant of infarct size after direct angioplasty. Circulation 1993; 88(pt I):1527–1533. Laster SB, O’Keefe JH, Gibbons RJ. Incidence and importance of thrombolysis in myocardial infarction grade 3 flow after primary percutaneous transluminal coronary angioplasty for acute myocardial infarction. Am J Cardiol 1996; 78:623–626. Karagounis L, Sorensen SG, Menlove RL, Moreno F, Anderson JL. Does thrombolysis in myocardial infarction (TIMI) perfusion represent a mostly patent artery or a mostly occluded artery? Enzymatic and electrocardiographic evidence from the TEAM-2 study. J Am Coll Cardiol 1992; 19:110. Hamada S, Nakamura S, Sugiura T, Murakami T, Fujimoto T, Watanabe J, et al. Early detection of the no-reflow phenomenon in reperfused acute myocardial infarction using technetium-99m tetrofosmin imaging. Eur J Nucl Med 1999; 26:208–214. Hamada S, Nakamura S, Sugiura T, Nishiue T, Watanabe J, Hatada K, et al. Accuracy of technetium-99m tetrofosmin myocardial perfusion imaging in the detection of spontaneous recanalization in patients with acute anterior myocardial infarction. Eur J Nucl Med 2001; 28:327–333. Watanabe J, Nakamura S, Sugiura T, Takehana K, Hamada S, Miyoshi H, et al. Early identification of impaired myocardial reperfusion with serial assessment of ST segments after percutaneous transluminal coronary angioplasty during acute myocardial infarction. Am J Cardiol 2001; 88: 956–959. Rentrop KP, Cohen M, Blanke H, Phillips RA. Change in collateral channel filling immediately after controlled coronary artery occlusion by an angioplasty balloon in human subjects. J Am Coll Cardiol 1985; 5: 587–592. Ito H, Okamura A, Iwakura K, Masuyama T, Hori M, Takiuchi S, et al. Myocardial perfusion patterns related to thrombolysis in myocardial infarction perfusion grades after coronary angioplasty in patients with acute anterior wall myocardial infarction. Circulation 1996; 93:1993–1999. Ito H, Tomooka T, Sakai N, Higashino Y, Fujii K, Katoh O, et al. Time course of functional improvement in stunned myocardial in risk area in patients with reperfused anterior infarction. Circulation 1993; 87:355–362. Sinusas AJ, Trautman KA, Bergin JD, Watson DD, Ruiz M, Smith WH, et al. Quantification of area at risk during coronary occlusion and degree of myocardial salvage after reperfusion with technetium-99m methoxyisobutyl isonitrile. Circulation 1990; 82:1424–1437. Kenner MD, Zajac EJ, Kondos GT, Dave R, Winklelmann JW, Joftus J, et al. Ability of the no-reflow phenomenon during an acute myocardial infarction to predict left ventricular dysfunction at one-month follow-up. Am J Cardiol 1995; 76:861–868. Yamamoto K, Ito H, Iwakura K, Kawano S, Kiushima M, Masuyama T, et al. Two different coronary blood flow velocity patterns in thrombolysis in myocardial infarction flow grade 2 in acute myocardial infarction: insight into mechanisms of microvascular dysfunction. J Am Coll Cardiol 2002; 40:1755–1760. Van’t Hof AW, Liem A, Suryapranata H, Hoorntje JC, de Boer MJ, Zijlstra F. Angiographic assessment of myocardial reperfusion in patients treated with primary angioplasty for acute myocardial infarction: myocardial blush grade. Zwolle Myocardial Infarction Study Group. Circulation 1998; 97:2302–2306. Gibson CM, Cannon CP, Daley WL, Dodge Jr JT, Alexander B, Marble SJ, et al. TIMI frame count. A quantitative method for assessing coronary artery flow. Circulation 1996; 93:879–888. Hamada S, Nishiue T, Nakamura S, Sugiura T, Kamihata H, Miyoshi H, et al. TIMI frame count immediately after primary coronary angioplasty as a predictor of functional recovery in patients with TIMI 3 reperfused acute myocardial infarction. J Am Coll Cardiol 2001; 38:666–671. Ragosta M, Camarano G, Kaul S, Powers ER, Sarembock IJ, Gimple LW. Microvascular integrity indicates myocellular viability in patients with recent myocardial infarction. New insights using myocardial contrast echocardiography. Circulation 1994; 89:2562–2569.

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24 Glover DK, Ruiz M, Koplan BA, Watson DD, Beller GA. 99mTc-tetrofosmin assessment of myocardial perfusion and viability in canine models of coronary occlusion and reperfusion. J Nucl Med 1999; 40:142–149. 25 Miller TD, Christian TF, Hopfenspirger MR, Hodge DO, Gersh BJ, Gibbons RJ. Infarct size after acute myocardial infarction measured by quantitative tomographic technetium-99m sestamibi imaging predicts subsequent mortality. Circulation 1995; 92:334–341. 26 Matsunari I, Tanishima Y, Taki J, Ono K, Nishide H, Fujino S, et al. Early and delayed technetium-99m-tetrofosmin myocardial SPECT compared in normal volunteers. J Nucl Med 1996; 37:1622–1626. 27 Jain D, Wackers FJT, Mattera J, McMahon M, Sinusas AJ, Zalet BL. Biokinetics of technetium-99m-tetrofosmin: myocardial perfusion imaging

agent: Implications for a one-day imaging protocol. J Nucl Med 1993; 34:1254–1259. 28 Sridhara BS, Braat S, Rigo P, Itti R, Cload P, Lahiri A. Comparison of myocardial perfusion imaging with technetium-99m-tetrofosmin versus thallium-201 in coronary artery disease. Am J Cardiol 1993; 72:1015–1019. 29 de Boer MJ, Suryapranata H, Hoorntje JCA, Reiffers S, Liem AL, Miedema K, et al. Limitation of infarct size and preservation of left ventricular function after primary coronary angioplasty compared with intravenous streptokinase in acute myocardial infarction. Circulation 1994; 90:753–761. 30 Grines CL, Browne KF, Marco J, Rothbaum D, Stone GW, O’Keefe J, et al. A comparison of immediate angioplasty with thrombolytic therapy for acute myocardial infarction. N Engl J Med 1993; 328:673–679.

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Original article

Post-test quality control for the single blood sample technique in glomerular filtration rate measurement in children Carlos De Sadeleera, Amy Piepszb and Hamphrey R. Hama Background While the value of the single blood sample (SBS) method for estimating 51Cr-EDTA plasma clearance has been repeatedly demonstrated, some nuclear medicine physicians are still reluctant to use it because of the lack of quality control parameters. Purpose To present a post-test quality control procedure for the SBS technique in children. Methods In addition to the SBS clearance calculated using the specific paediatric SBS method, three artificial slope intercept (ASI) method clearances were calculated by assuming the distribution volume as, respectively, 20%, 25% and 30% of body weight. By dividing the injected activity by the distributional volume, the initial plasma concentrations (A0,30%, A0,25% and A0,20%) were calculated. Using these A0 values and the available single sample, ASI clearances were calculated by using the classical slope–intercept method. The working hypothesis of this approach was as follows. In the absence of significant errors, the three ASI clearance values should be close to that of the SBS method. This hypothesis has been tested using both simulated and patients’ data. Results The results of the simulated study showed that an error in the injected dose produced variable differences

Introduction The value of single blood sample (SBS) method for estimating 51Cr-EDTA plasma clearance has been repeatedly demonstrated and its clinical use is recommended by different guidelines and consensus reports for the calculation of the glomerular filtration rate (GFR) [1– 3]. In children, the method of Ham and Piepsz [4] is recommended. However, some nuclear medicine physicians are still reluctant to use the SBS technique because of a lack of internal quality control procedures. In the British Nuclear Medicine Society (BNMS) guidelines [3], the use of the single blood sample technique is not even recommended. Indeed, when using the conventional slope–intercept method, the slope rate constant of the terminal exponential and the calculated distribution volume can be used as a means of quality control. If three or more blood samples are used, the fit (r2) can also be used as quality control parameter. When using the SBS

between SBS and ASI clearances depending on the clearance values. The effect of an error on the plasma sample also varied as a function of the clearance values. The analysis of patient data revealed that the ASI approach allowed the identification of patients in whom the classical slope–intercept method suggested the presence of a possible error. Conclusion A post-test quality control procedure for the SBS GFR measurement is presented. When the SBS clearance shows a difference with the ASI method ( > 10 ml
min – 1 per 1.73 m2), the presence of an error is highly probable. A smaller difference, however, does not c exclude erroneous data. Nucl Med Commun 27:255–260  2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:255–260 Keywords:

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Cr-EDTA, GFR, quality control, single sample, children

a Department of Nuclear Medicine, UZ Gent and bDepartment of Nuclear Medicine, CHU St Pierre, Brussels, Belgium.

Correspondence to Dr Carlos De Sadeleer, Borrekent 51, 9450 Haaltert (Denderhoutem), Belgium. Tel: + 00 32 54 32 56 07; fax: + 00 32 54 51 81 51; e-mail: [email protected] Received 19 September 2005 Accepted 25 November 2005

method, an error on the injected dose or an error on the plasma sample will directly and proportionally affect the calculated clearance. There is no slope, no initial distribution volume or any other parameter that can be used as an ‘alarm bell’ signalling a possible error. The aim of this work was to present a post-test quality control procedure for the SBS technique in children which can identify cases with an abnormal distribution volume.

Material and methods Principles

In addition to the SBS clearance calculated using the specific paediatric SBS method [4], three artificial slope intercept (ASI) method clearances were calculated by assuming the initial one-compartment distribution volume as, respectively, 20%, 25% and 30% of body weight. By dividing the injected activity by these values of the distribution volume, the initial plasma concentrations

c 2006 Lippincott Williams & Wilkins 0143-3636 

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(A0,30%, A0,25% and A0,20%) were calculated. Using these A0 values and the available single sample, ASI clearances were calculated by means of the classical slope–intercept method.

Errors were then introduced in the injected dose (errors from – 50% to + 50%) and on the 120 min blood sample (errors from – 50% to + 50%), and the SBS and the three ASI clearances were calculated and compared.

The working hypothesis of this approach was as follows. In the absence of significant errors, the three ASI clearance values should be close to that of the SBS method. This hypothesis has been tested using both simulated and patient data.

Patient study

Simulation study

A simulated computer model of a single compartment system was created in order to represent three children aged 3, 6 and 10 years. The three age groups were chosen to simulate patients with different heights and weights, which were taken from the children’s age curves. The heights and weights allowed us to calculate the body surfaces. For each of the three ‘children’, five different clearances values and various distribution volumes (20% of the body weight, 25% of the body weight and 30% of the body weight) were simulated. In each setting the 120 min plasma concentration (expressed as a percentage of injected dose) was determined, and the SBS clearance was calculated using the method of Ham and Piepsz [4]. Then three ASI method clearances were calculated. First, we assumed that the initial distribution volume (DV) was 20% of body weight (DV20). By dividing the injected activity by this DV20, the initial plasma concentration or intercept of the terminal exponential (A0,20) was calculated. Then, by joining the A0,20 value and the available single sample value at 120 min after tracer injection (P120), the slope (l20) was calculated. Finally, the clearance (CL20) was calculated by using the slope–intercept method, by multiplying the DV20 by l20, or lnðAt =A0;20 Þ  0:2W t where W is body weight. Chantler’s correction for neglecting the first exponential was also performed [5]. These three steps were then repeated by assuming the initial distribution volume to be, respectively, 25% and 30% of body weight, resulting in the clearance values CL25 and CL30. These three different values for the distribution volume (respectively, 20%, 25% and 30% of body weight) were chosen as they represent the usual range of this parameter.

The same approach has been applied retrospectively to two groups of patients chosen from a large slope– intercept clearance database. The first group consisted of 150 randomly selected children in whom the calculated distribution volume DV was between 20% and 35% of body weight. The second group consisted by all children (n = 27) in whom the slope–intercept clearance resulted in a distribution volume of less than 15% or more than 40% of body weight, suggesting the presence of a possible error. In these two patient groups the clearance value by the SBS method and the three ASI clearances were calculated (CL20, CL25 and CL30). For each patient, the maximal difference (D) between the results of the SBS and the three ASI methods, all normalized to 1.73 m2, were calculated (D20, D25 and D30). To compare the maximal differences in the two groups, a t-test of two independent unequal samples was performed without assuming equal variances (SPSS software).

Results Simulation study The error on the injected dose

Normal clearance For a 6-year-old child (with an assumed weight of 21.8 kg and height of 121.2 cm) with a normal clearance (120 ml
min – 1 per 1.73 m2) (Fig. 1), an error between – 30% and + 20% on the injected dose induced a difference of less than 10 ml
min – 1 per 1.73 m2 between the SBS and the ASI clearances. In the case of an underestimation of the injected dose of more than – 30% as well as in the case of an overestimation of the injected dose of more than + 20% the errors obtained by means of SBS diverged from those obtained by the ASI technique. The higher the underestimation and overestimation of the injected dose, the greater was the difference between the two methods. Slightly reduced clearance For a 6-year-old child with a slightly reduced clearance (80 ml
min – 1 per 1.73 m2) (Fig. 1), an error between – 20% and + 40% on the injected dose induced similar errors on both the SBS and the ASI (a difference of less than 10 ml
min – 1 per 1.73 m2). In the case of an underestimation of the injected dose of more than – 20% as well as in the case of an

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Quality control for single-sample technique in GFR De Sadeleer et al. 257

Fig. 1

Error on dose - child of 6 years with CL = 120 ml.min−1 per 1.73 m² Difference SBS - ASI (ml.min−1 per 1.73 m²)

10.00

−60%

−40%

−20%

0.00 0%

20%

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60%

−10.00 −20.00 −30.00 −40.00 120 ml 30% 120 ml 25% 120 ml 20%

−50.00 −60.00 Error on dose (%)

Error on dose - child of 6 years with CL = 80 ml.min−1 per 1.73 m² Difference SBS - ASI (ml.min−1 per 1.73 m²)

10.00

−60%

−40%

−20%

0.00 0%

20%

40%

60%

−10.00 −20.00 −30.00 −40.00 80 ml 30% 80 ml 25% 80 ml 20%

−50.00 −60.00 Error on dose (%)

Difference SBS - ASI (ml.min−1 per 1.73 m²)

Error on dose - child of 6 years with CL = 40 ml.min−1 per 1.73 m²

−60%

10.00 −40%

−20%

0.00 0%

20%

40%

60%

−10.00 −20.00 −30.00 −40.00 40 ml 30% 40 ml 25% 40 ml 20%

−50.00 −60.00 Error on dose (%)

Effect of error on dose for a 6-year-old child. The different clearance values are given.

overestimation of the injected dose of more than + 40% the errors obtained by means of SBS diverged from those obtained by the ASI technique.

Moderately reduced clearance For a 6-year-old child with a moderatly reduced clearance (40 ml
min – 1 per 1.73 m2) (Fig. 1), an error between + 20% and + 60% on the

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injected dose induced comparable errors on both the SBS and the ASI (a difference of less than 10 ml
min – 1 per 1.73 m2). In the case of an overestimation of the plasma activity of less than + 20% as well as for an underestimation of the plasma activity, the errors obtained by means of SBS diverged from those obtained by the ASI technique. Children of other ages Comparable results are also obtained for a 3-year-old child (with an assumed weight of 14.1 kg and height of 96 cm) and for a 10-year-old child (with an assumed weight of 30.6 kg and height of 134.5 cm). Error on the 120 min plasma sample

Normal clearance When an error was present on the 120 min sample, different situations were observed. For a 6-year-old child with a normal clearance (120 ml
min – 1 per 1.73 m2) (Fig. 2), an error between – 10% and + 60% on the plasma activity induced errors of the same magnitude on both the SBS and the ASI (a difference of less than 10 ml
min – 1 per 1.73 m2). In the case of an underestimation of the plasma activity of more than – 10%, the errors obtained by means of SBS diverged from those obtained by the ASI technique. The higher the underestimation of the plasma activity, the greater was the difference between the two methods. Slightly reduced clearance For a 6-year-old child with slightly reduced clearance (80 ml
min – 1 per 1.73 m2) (Fig. 2), an error between – 30% and + 20% on the plasma activity induced almost identical errors on both the SBS and the ASI (a difference of less than 10 ml
min – 1 per 1.73 m2). In the case of an underestimation of the plasma activity of less than – 30% as well as in the case of an overestimation of the plasma activity of more than + 20%, the errors obtained by means of SBS diverged from those obtained by the ASI technique. The higher the underestimation and overestimation of the plasma activity, the greater was the difference between the two methods. Moderately reduced clearance For a 6-year-old child with a moderately reduced clearance (40 ml
min1 per 1.73 m2) (Fig. 2), an underestimation of the plasma activity between – 50% and – 10% induced almost similar errors on both the SBS and the ASI (a difference of less than 10 ml
min – 1 per 1.73 m2). In the case of an underestimation of the plasma activity of less than – 20% as well as for an overestimation of the plasma activity, the errors obtained by means of SBS diverged from those obtained by the ASI technique.

Children of other ages Comparable results are also obtained for a 3-year-old child (with an assumed weight of 14.1 kg and height of 96 cm) and for a 10-year-old child (with an assumed weight of 30.6 kg and height of 134.5 cm). Patient study

In the first group, the mean value of this maximal difference was 8.94 ml
min – 1 per 1.73 m2 (median = 8.82, first quartile = 7.86, third quartile = 9.70, minimum difference = 5.17, and maximum difference = 20.66). In 85% of the patients, the maximal difference was less than 10 ml
min – 1 per 1.73 m2. In the second group, the mean value of this maximal difference was 42.11 ml
min – 1 per 1.73 m2 (median = 29.51, first quartile = 15.73, third quartile = 44.70 minimum difference = 11.15, and maximum difference = 135.8) (Fig. 3). The difference in the maximal differences in the two groups was highly significant (P < 0.01).

Discussion Since its introduction by Tauxe et al. in 1971 [6], the single-sample method for evaluating renal clearance has been the subject of numerous investigations. There is currently a wide consensus concerning the validity of this approach and procedure guidelines recommend its use in adults as well as in children [1–3]. However, some nuclear medicine physicians are still reluctant to use the technique because the opportunity for routine quality control is lacking. In the BNMS guidelines, use of the SBS technique is not even recommended. However, in the same BNMS guidelines, the SBS technique is recommended as a quality control for the two-blood samples technique, which indirectly suggests the acceptance of the value of the SBS technique. In this study, we present a post-test quality control for the SBS technique in children which was based on the following hypothesis. In the absence of an important error, the differences between SBS clearance and three artificially calculated slope–intercept clearances by assuming the distribution volume as 20%, 25% or 30% should be minimal. A large difference between the two methods would indicate the presence of errors. The results of this study partially validate our hypothesis. The results of the simulated study show that an important error creates an important difference between the results obtained by the SBS and ASI techniques. In the presence of a smaller error, however, the difference between the two methods remains less than 10 ml
min – 1 per 1.73 m2, suggesting the low sensitivity of the proposed post-test quality control method.

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Quality control for single-sample technique in GFR De Sadeleer et al. 259

Difference SBS - ASI (ml.min−1 per 1.73 m²)

Fig. 2

Error on 120 minutes blood sample - child of 6 years with CL = 120 ml.min−1 per 1.73 m² 10.00 −60%

−40%

−20%

0.00 0%

20%

40%

60%

− 10.00 − 20.00 − 30.00 − 40.00 120 ml 30% 120 ml 25% 120 ml 20%

− 50.00 − 60.00 Error on sample (%)

Difference SBS - ASI (ml.min−1per 1.73 m²)

Error on 120 minutes blood sample - child of 6 years with CL = 80 ml.min−1 per 1.73 m² 10.00 −60%

−40%

−20%

0.00 0%

20%

40%

60%

− 10.00 − 20.00 − 30.00 − 40.00 80 ml 30% 80 ml 25% 80 ml 20%

− 50.00 − 60.00 Error on sample (%)

Difference SBS - ASI (ml.min−1 per 1.73 m²)

Error on 120 minutes blood sample - child of 6 years with CL = 40 ml.min−1 per 1.73 m² 10.00 −60%

−40%

−20%

0.00 0%

20%

40%

60%

− 10.00 − 20.00 − 30.00 − 40.00 40 ml 30% 40 ml 25% 40 ml 20%

− 50.00 − 60.00 Error on sample (%)

Effect of error on the 120 min plasma sample for a 6-year-old child. The different clearance values are given.

Using patient data, most patients with a possible error according to the slope–intercept method are detected by this post-test quality control method.

In 85% of patients assumed to be free of error, the maximal difference was less than 10 ml
min – 1 per 1.73 m2, whereas in all 26 patients assumed of having

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an error, the difference was higher than 11 ml
min – 1 per 1.73 m2.

Fig. 3

100

Summary

90

A post-test quality control method for the SBS GFR measurement has been presented. A difference greater than 10 ml
min – 1 per 1.73 m2 between the SBS technique and the artifical slope–intercept method should prompt a search for a potential error. However, if a smaller difference is found, the existence of errors cannot be excluded.

80

ml/min/1.73m2

70 60 50

References 40

1

30

2

20 3

10

4

0 1.00

2.00 group 5

Patient study: difference in the maximal differences in the two groups (group I = DV between 20% and 35% of body weight; group II DV < 15% or > 40% of body weight) was highly significant.

6

Blaufox MD, Aurell M, Bubeck B, Fommei E, Piepsz A, Russell C, et al. Report of the Committee on the use of Radionuclides in Nephrourology and Renal Clearance. J Nucl Med 1996; 37:1883–1890. Piepsz A, Colarinha P, Gordon I, Hahn K, Olivier P, Sixt R, van Velzen J. Guidelines for glomerular filtration rate determination in children by the Paediatric Committee of the European Association of Nuclear Medicine. Eur J Nucl Med 2001; 28:BP31–BP36. Fleming JS, Zivanovic MA, Blake GM, Burniston M, Cosgriff PS. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Nucl Med Commun 2004; 28:759–769. Ham HR, Piepsz A. Estimation of glomerular filtration rate in children using a single plasma sample method. J Nucl Med 1991; 32: 1294–1297. Chantler C, Garnett S, Parsons V, Veall N. Glomerular filtration rate measurement in man by the single injection method using Cr-51 EDTA. Clin Sci 1969; 37:169–180. Tauxe WN, Maher FT, Taylor WF. Effective renal plasma flow: estimation from theoretical volumes of distribution of intravenously injected 131-I orthoiodohippurate. Mayo Clin Proc 1971; 46:524–531.

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Original article

Radioiodine whole-body scans, thyroglobulin levels, 99m Tc-MIBI scans and computed tomography: results in patients with lung metastases from differentiated thyroid cancer ¨ zlem N. Ku¨c¸u¨k, Sinan S. Gu¨ltekin, Gu¨lseren Aras and Erkan ˙Ibis¸ O Objectives The correlation between a 131I whole-body scan (WBS), a 99mTc sestamibi (99mTc-MIBI) WBS, a computed tomography (CT) scan and the value of routine follow-up for 131I WBS and thyroglobulin (Tg) levels in patients with lung metastases from differentiated thyroid cancer was assessed. Method Pulmonary metastases were detected in 32 patients out of 583 with differentiated thyroid cancer (DTC) who were admitted to our clinic between 1985 and 2004 (age range, 22–79 years; mean, 58 ± 19 years; 15 women and 17 men). Pulmonary metastases were diagnosed by considering the 131I WBS, increased Tg levels and/or other positive radiological findings. Papillary carcinoma was diagnosed in 15/32 patients and follicular carcinoma in 13/32. A mixed type found in 4/32 patients was classified histopathologically. A total of 3.7–53.65 GBq (100–1450 mCi) 131I was given to each patient. The duration of follow-up ranged from 36 to 240 months. A 131I WBS, the determination of Tg levels and/or a CT scan were carried out in the assessment of a diagnosis and follow-up of patients with lung metastases. A 99mTc-MIBI WBS was performed on 19 patients who were chosen at random from the 583. Results Nineteen of 32 patients had lung metastases before they received the first 131I treatment. Six of the 32 had distant-organ metastases other than in the lungs. Four of these six patients had only lung and bone metastases. Pulmonary metastases were observed on the 131I WBS patients 31/32 (96.8%) whereas no pulmonary metastases, were detected on the CT scans in 3/32 patients. The last diagnostic whole-body scan (DWBS) was normal in 13/32 patients. At the first examination, the Tg levels in 27/32 (84.4%) patients were below 30 ng
ml – 1. At the final examination, 20/32 (62.5%) patients had Tg levels higher than 30 ng
ml – 1, while Tg levels were lower than

Introduction Thyroid cancer accounts for 90% of all endocrine malignancies although it represents less than 1% of all malignancies. Metastatic disease develops in 7–23% of patients with differentiated thyroid carcinoma (DTC) [1,2]. However, distant metastases at the time of initial diagnosis are observed in only 1–4% of patients [3]. The

30 ng
ml – 1 in 12/32 patients. Tg levels decreased in 21/32 and increased in 3/32 patients. The 131I WBS continued to be abnormal in 2/3 patients with increased Tg levels but became normal in one patient whose CT scan still showed macro-nodular lesions. Tg levels did not change significantly in 8/32 patients. The 131I WBS became normal in 5/8 patients, while the CT scans for 4/5 showed micro-nodules. Metastases were detected in 12/19 patients who underwent 99mTc-MIBI whole-body scanning: 18/19 showed metastases on the 131I WBSs and 17/19 on the CT scans. Of the seven patients without a sign of metastasis on the 99mTc-MIBI WBS, one was negative in terms of metastasis on the 131I WBS and one on the CT scan. Fibrosis was observed on the CT scans of 2/32 patients. One patient developed dedifferentiation, as determined by the negative 131I WBS and positive CT scan. Conclusion 131I whole-body scanning and the determination of Tg levels are the most important procedures for the evaluation of lung metastases in differentiated thyroid cancer. Computed tomography is a useful addition to 131 I whole-body scanning. MIBI imaging alone may not be enough to detect lung metastases from differentiated c 2006 thyroid cancer. Nucl Med Commun 27:261–266 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:261–266 Keywords: iodine-131 whole-body scan, Tg level, differentiated thyroid cancer, lung metastasis Department of Nuclear Medicine, Ankara University Faculty of Medicine, Turkey. Correspondence to Dr Nuriye O. Kucuk, Ankara University Faculty of Medicine, Department of Nuclear Medicine, 06100, Cebeci, Ankara, Turkey. Tel: + 90 312 362 01 98; fax: + 90 312 362 08 97; e-mail: [email protected] Received 3 October 2005 Accepted 2 December 2005

lungs are the most frequent distant localization of metastases from DTC [4]. Complete surgical resection of the thyroid (total or near-total thyroidectomy) and post-operative ablation therapy with 131I are routine treatment approaches for patients with DTC. The patients are followed up after

c 2006 Lippincott Williams & Wilkins 0143-3636

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262 Nuclear Medicine Communications 2006, Vol 27 No 3

thyroidectomy and radioiodine therapy by serial 131I whole-body scans (131I WBSs) and serum thyroglobulin (Tg) measurements [5]. 99mTc sestamibi whole-body scans (99mTc-MIBI WBSs) are particularly useful for the evaluation of the patients with negative 131I WBSs and elevated thyroglobulin levels (non-functional disease) in DTC. Furthermore, a 99mTc-MIBI scan has advantages over a 131I scan: (1) it is not necessary to discontinue hormone therapy when performing scintigraphy; (2) 99m Tc-MIBI provides better quality images; and (3) carrying out a scintigraphic examination is easy and time saving. On the basis of these favourable aspects, the use of 99mTc-MIBI whole-body scanning has been suggested as an alternative to 131I whole-body scanning for the evaluation of metastatic disease in DTC patients [6–8].

Table 1 Clinical, pathological and treatment-related characteristics of the 32 patients with lung metastasis from differentiated thyroid carcinoma

This retrospective study investigated the correlation and value of a 131I WBS and 99mTc-MIBI scan, computed tomography (CT) and Tg levels in the assessment of lung metastases in patients with DTC. The use of routine Tg level measurements and 131I WBSs, combined, as a ‘gold standard’ in the verification and follow-up of lung metastases in patients without any distant-organ metastases after successful thyroid remnant ablation were evaluated.

Materials and methods Patients

Five hundred and eighty-three patients with DTC were admitted to the Department of Nuclear Medicine of Ankara University Medical School between 1985 and 2004. All the patients were examined retrospectively and were evaluated according to the results of clinical examination and [99mTc]pertechnetate thyroid scintigraphy. Routine nuclear medicine methods (131I WBS, 201Tl or 99mTc-MIBI scan, thyroid functions tests) and, when necessary, some biochemical tests and radiological studies (e.g., chest X-ray, neck ultrasonography, neck and thorax CT) were performed for the evaluation and follow-up of patients. Of the patients, 32 had lung metastases: histopathologically, papillary carcinoma occurred in 15/32 patients, follicular carcinoma in 13/32 patients and a mixed type in 4/32. Ages at the first diagnosis of 32 patients ranged from 22 to 79 years (mean, 58 ± 19 years; 15 female and 17 male). The duration of follow-up ranged from 36 to 240 months. Clinical, pathological and treatment-related characteristics are presented in Table 1. Surgery

Thirty-one of the 32 patients underwent total or neartotal thyroidectomy. Lymph node dissections at the initial surgery were performed in 12 patients. Nine patients underwent operations twice to ensure that the result was satisfactory.

Characteristic Gender Male Female Median age at diagnosis (years)

Number or value

Percent

17 15

53.12 46.87

58 ± 19

Surgery None Total or near-total Thyroidectomy Lymph-node dissection

1 31

3.12 96.87

12

37.50

Histology Papillary Follicular Mixed type

15 13 4

46.87 40.62 12.50

Lung metastasis at diagnosis Yes No

19 13

59.37 40.62

The algorithm of treatment and follow-up

Those patients with DTC who had undergone total thyroidectomy received a fixed dose of 131I for the ablation of thyroid remnants. A post-treatment 131I WBS was performed 6 days after ablation therapy. Periodic Tg measurements and diagnostic WBS (DWBS) were used for the follow-up of patients after successful ablation. 201 Tl or 99mTc-MIBI was used for the diagnosis and follow-up in cases of negative DWBS and elevated Tg levels. We could not use [18F]fluorodeoxyglucose (18F-FDG) scanning as there is no PET camera in our department. The algorithm of treatment and follow-up used in our department is given in Fig. 1. 131

I therapy and post-treatment

131

I WBS

Patients were each given a high fixed ablative dose for 131 I treatment. Four weeks before 131I therapy, L-thyroxin replacement therapy was withdrawn low iodine diet protocols were applied. TSH levels increased to at least 30 ng
ml – 1. A total of 3.7–53.65 GBq (100–1450 mCi) 131 I was given to each patient. In the presence of persistent functioning lung metastases, radioiodine therapy was repeated periodically (at least 6 months after the therapy). None of these 32 patients underwent 131I whole-body scanning before the first therapy dose of radioiodine. Post-treatment 131I WBSs, planar and spot images were obtained in anterior and posterior projections. It was performed on the sixth post-treatment day using a largefield-of-view gamma camera equipped with a high energy (peak energy centred on 360 keV with a 20% energy window), parallel hole collimator (GE 4000iXC-T/ STARCAM, GE Medical Systems, Milwaukee, Wisconsin). Post-treatment 131I WBS was used only for the purpose of verifying metastatic disease. Each patient was

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Scan results in patients with lung metastases from thyroid cancer Ku¨c¸u¨k et al. 263

Fig. 1 131I

ablation (2.775–4.625 GBq) (75–125 Mci)

PTWBS > 0

PTWBS < 0

3 months on L-T4: TSH, Tg level 6 months off L-T4: DWBS, TSH, Tg Level

Positive DWBS

Patients with normal DWBS were evaluated with periodic follow-up, at first once a year and then every 2 or 3 years within a 10-year period. However, Tg levels of all the patients were obtained every 6 months. In our department, Tg levels were measured by an immunoradiometric method between 1985 and 2000, and by a chemiluminescence method after 2000. Suppressive hormone therapy was resumed after each 131I WBS. All the patients were evaluated regarding positive pulmonary uptake on the DWBS. 99m

Negative DWBS

Tc-MIBI scan

99m

Tg < 30 ng·ml−1

Tg > 30 ng·ml−1

99m

99m

201

201

Tc-MIBI or T1 WBS on L-T4 Negative scan Tg < 10 ng·ml−1

Tc-MIBI or T1 WBS on L-T4

Negative scan Tg > 10 ng·ml−1

FDG scan if available, CT scan of neck/chest

Follow-up: DWBS, Tg (every 1 to 5 years and 6 months, respectively)

Positive scan Tg > 10 ng·ml−1

Negative scan

Positive scan

Surgery Radioiodine 131 I therapy

The algorithm for treatment and follow-up. PTWBS: post-treatment whole-body scan; DWBS: diagnostic whole-body scan; L-T4: L-thyroxine; Tg: thyroglobulin; TSH: thyroid stimulating hormone; PTWBS > 0: Existence of uptake outside the thyroid bed; PTWBS < 0:Abscence of uptake outside the thyroid bed

examined to determine positive uptake on the posttreatment 131I WBS. Patients with increased pulmonary activity were referred to the department of thoracic diseases and evaluated by bronchoscopy in order to confirm the metastasis. Protocol for routine follow-up

Suppressive hormone therapy (L-thyroxin) was started 48–72 h after radioiodine therapy. Diagnostic 131I WBSs and Tg levels were obtained after the first treatment for the first follow-up in 6 months and at both 3 and 6 months, respectively. Thyroid hormone withdrawal and low iodine diet protocols were applied for 4 weeks. The DWBS was performed with a 185 MBq (5 mCi) dose of 131 I. DWBS, planar and spot images were acquired in anterior and posterior projections at the 24 and 72 h (early and late images) using a GE 4000iXC-T/STARCAM gamma camera equipped with a high energy collimator.

Tc-MIBI whole-body scanning was carried out in 19 patients who were chosen at random. We have used 99m Tc-MIBI in our department since 1990 although it was some time before we started to use it as a routine indication. Therefore not all patients in our group underwent 99mTc-MIBI scanning. We also performed 201 Tl WBSs in some patients but did not include them in this study because the number was small. 99mTc-MIBI whole-body scanning was performed on the patients during suppressive hormonal therapy. 99mTc sestamibi (555–740 MBq, 15–20 mCi), CARDIO-SPECT; (MediRadiopharma Ltd, Hungary) was injected intravenously. Images were obtained 20–30 min later. MIBI WBSs and planar and spot images were obtained in anterior and posterior projections with a large-field-of-view gamma camera equipped with a low-energy (peak energy centred on 140 keV with a 15% energy window), high resolution collimator (Siemens ECAM Dual Head Variable Systems; Siemens Medical Solutions, Illinois). The patients were examined for the similar positive pulmonary uptake regions in 99mTc-MIBI and 131I WBS. Computed tomography

CT was performed when pulmonary metastases were detected by positive 131I WBSs and/or increasing Tg levels. In most of the patients, CT was not performed as a follow-up procedure. All patients underwent conventional CT for the examination of metastatic lesions without the use of contrast media in order not to affect the radioiodine therapy. The CT equipment used did not have high resolution properties. The CT images were obtained with 7 mm slice thickness starting from the apex of the lungs. All CT images were obtained with the patient in the supine position. Patients were evaluated in terms of metastatic pulmonary lesions and the lesions were classified according to size (macro-nodule > 10 mm).

Results Nineteen out of 32 patients had lung metastases before the first 131I treatment. Lung metastases were detected in 13 out of 32 patients during the follow-up period. All patients were given radioiodine therapy. Thyroid remnant ablation was successful in the first DWBS in 22/32 patients. Six of 32 patients had distant-organ metastases other than in the

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Nuclear Medicine Communications 2006, Vol 27 No 3

lungs. Four of the six patients had only lung and bone metastases. Pulmonary metastases were observed on the 131 I WBSs of 31/32 (96.8%) patients whereas no pulmonary metastases were detected on the CT scans of 3/32 patients. The last DWBS was negative in 13/32 (40.6%) patients. DWBS was positive in 19/32 patients. At their initial examination, 27/32 (84.4%) patients had Tg levels higher than 30 ng
ml – 1, and 5/32 (15.6%) patients had Tg levels lower than 30 ng
ml – 1. At their final examination, 20/32 (62.5%) patients had Tg levels higher than 30 ng
ml – 1, and 12/32 (37.5%) patients had Tg levels lower than 30 ng
ml – 1. The final Tg levels changed in 7/32 patients. Eight of 32 patients had Tg levels lower than 30 ng
ml – 1 and negative 131I WBS. However, one patient had a Tg value higher than 30 ng
ml – 1 but had a negative 131I WBS. Four of 32 (12.5%) patients had Tg levels less than 30 ng
ml – 1 and positive 131I WBSs. Thirteen of 32 patients had negative 131 I WBSs after a final diagnostic 131I WBS. Tg levels decreased in 21/32 patients and increased in 3/32. 131

I WBSs continued to be abnormal in two of three patients with increased Tg levels but became normal in one patient whose CT still demonstrated macro-nodular lesions. Tg levels did not change significantly in 8/32. 131I WBSs became normal in 5/8 patients, while 4/5 showed micro-nodules on their CT scans. Fibrosis was observed in 2/32 patients on CT. One patient developed dedifferentiation with negative 131I WBS and positive CT.

Metastases were discerned in 12/19 patients who underwent 99mTc-MIBI whole-body scanning; 18/19 showed metastasis on 131I WBS and 17/19 in CT (Table 2). Of the seven patients without any sign of metastasis upon the 99m Tc-MIBI WBS, one was negative in terms of metastasis on the 131I WBS and one after the CT scan. For 3/4 patients who had negative 131I WBSs and Tg levels higher than 30 ng
ml – 1, 99mTc-MIBI whole-body scanning was used as the last follow-up examination. Two of these three patients had positive 99mTc-MIBI scans. In contrast, the patient who had a negative 131I WBS and a Tg level higher than 30 ng
ml – 1 during initial follow-up had a negative MIBI scan.

Discussion The lungs are the most frequent distant localization of metastases from DTC (1–4%) [4]. The results of 131I Table 2 Comparison of findings using different modalities for patients with serum thyroglobulin levels > 30 ng
ml – 1 Finding

131

I whole-body scanning

99m

Tc-MIBI whole-body scanning

Computed tomography

Positive Negative

18 1

12 7

17 2

Percent

94.7

63.1

89.4

WBSs and Tg levels play an important diagnostic role in the evaluation of lung metastasis in differentiated thyroid cancer [9,10]. In our patient group, it was found to be 5%. Lung metastases were observed on the 131I WBSs of all the patients except one, who had a very high Tg level and negative 131I WBS. The measurement of serum Tg levels is the most sensitive and specific marker of DTC patients, but increased Tg concentration alone is not enough when there is a large thyroid remnant. In our retrospective study Tg levels were measured by an immunoradiometric method between 1985 and 2000 and by chemiluminescence after 2000. The cut-off values were 30 ng
ml – 1 in the first period and 5 ng
ml – 1 in the second period. Values above these were considered for the assessment of metastatic disease. Although it is reported that the chemiluminescence method is more sensitive compared to the immunoradiometric method [11] we took the cutoff value as 30 ng
ml – 1 in this retrospective study because both methods were used during the 5 years that analysis was carried out. Initial Tg concentrations determined after total thyroidectomy or near-total thyroidectomy with lung metastases in 32 patients off thyroxin and before 131I therapy or scanning showed that 15.6% (5/32) of patients had initial levels less than 30 ng
ml – 1 and, in contrast, 84.4% (27/ 32) had initial levels higher than 30 ng
ml – 1. Filesi et al. [12] reported that, in their series of patients, 66.7% with metastases had initial Tg levels higher than 60 ng
ml – 1. Ng et al. [7] reported that 46% of patients with thyroid remnants or metastases had initial serum Tg values higher than 30 ng
ml – 1. In our group, 84.4% of patients with lung metastases had initial Tg levels higher than 30 ng
ml – 1. In addition, Filesi et al. reported that metastases were observed in the initial 131I WBS in 47.8% of patients with Tg values of less than 60 ng
ml – 1. In that study 61.3% of patients for whom the initial 131I WBS was negative for metastases had Tg levels higher than 60 ng
ml – 1. In contrast to this finding, Ng et al. [7] found that only 2.7% of subjects with a negative scan had initial serum Tg levels higher than 30 ng
ml – 1. Our retrospective findings suggested that 3.1% of patients had both a negative 131I WBS and a Tg level higher than 30 ng
ml – 1. Such a result might be associated with thyroid remnants and/or other distant metastases. Thyroid remnant ablation was successful in the first DWBS in 22/32 patients. In our study 6/32 patients had another distant-organ metastasis outside the lung. In addition, our series showed that 16.5% of patients who, initially, had a positive 131I scan for metastases had Tg levels less than 30 ng
ml – 1. So, we considered whether the Tg cut-off level should be reduced to less than 30 ng
ml – 1. Ng et al. [7] reported that with 131I ablation

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Scan results in patients with lung metastases from thyroid cancer Ku¨c¸u¨k et al. 265

and long-term thyroxin suppression, Tg levels tended to fall. In our study, final Tg concentrations were determined in order to measure initial values under the same conditions. We observed that 37.5% (12/32) of patients had final Tg levels less than 30 ng
ml – 1 and, conversely, 62.5% (20/32) had final Tg levels higher than 30 ng
ml – 1. Consequently, we aimed to reduce Tg levels to less than 30 ng
ml – 1 in 65.6% (21/32) of the patients. Furthermore, final Tg values became normal in 37.5% (12/32) of the patients (Fig. 2). The combination of routine Tg measurement and I whole-body scanning can be used as a ‘gold standard’ in the diagnosis and follow-up of lung metastases in DTC patients without the detection of any distant-organ metastasis and after successful thyroid remnant ablation. Increased activity in the lung region in the post-treatment 131I WBS is considered to be pulmonary metastasis. These patients were referred to the department of thoracic diseases and bronchoscopic methods were used to confirm the presence of metastatic disease. 131

The patients were divided into two groups according to the Tg level after L-thyroxine replacement therapy had been stopped: (1) those with Tg levels higher than 30 ng
ml – 1 and (2) those with Tg levels lower than 30 ng
ml – 1. It is thought that patients with Tg concentrations higher than 30 ng
ml – 1 represent a high risk group when considering effective treatment of the disease [1,12]. Hence, by dividing patients into the two groups we were able to evaluate how effective routine Tg determination can be in the follow-up of patients as a preparameter before commencing the multi-step, multiparameter evaluation of progression.

Fig. 2

16 000 14 000 12 000 ng.ml−1

10 000 8000 6000 4000 2000 0 1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 Patient number Thyroglobulin levels in 32 patients with lung metastases from differentiated thyroid cancer.

At the beginning of our study we had 27 patients with Tg levels higher than 30 ng
ml – 1 and five patients with Tg less than 30 ng
ml – 1. At the last evaluation there were only 20 patients with Tg levels higher than 30 ng
ml – 1. The last DWBS was negative in 13 of these 20 patients. DWBS was negative in eight and positive in four of 12/32 patients with Tg less than 30 ng
ml – 1 at the final evaluation. For 2/3 patients with Tg progression, the DWBS continued to be positive at the final evaluation. These results show that, at present, Tg measurement and 131 I whole-body scanning can not be evaluated as a ‘gold standard’ in the follow-up of DTC patients with pulmonary metastasis. Nevertheless, depression of serum Tg levels can be considered an obvious sign that the patient benefits from radioiodine therapy although selecting a lower cut-off value will increase the sensitivity and the chance of better correlation. Although it was not easy to evaluate the efficacy of treatment we were able to gather valuable information to help us decide whether, by using Tg measurement and DWBS together, high-dose cumulative radioiodine therapy would result in long-term stability in our patient group. 99m

Tc-MIBI scans have been particularly useful for the evaluation of patients with negative 131I scans and elevated thyroglobulin levels (non-functional disease) in DTC. Furthermore, a 99mTc-MIBI scan has advantages over a 131I scan: (1) it is not necessary to discontinue hormone therapy when performing scintigraphy; (2) MIBI provides better quality images; and (3) undertaking a scintigraphic examination is easy and time saving. On the basis of these favourable aspects, the use of 99mTcMIBI whole-body scanning has been suggested as an alternative to 131I whole-body scanning for the evaluation of the metastatic disease in DTC patients [6–8]. However, there are different opinions about the sensitivity of 99mTc-MIBI whole-body scanning in DTC. Sundram et al. [13] showed that a 99mTc-MIBI WBS had a sensitivity comparable witha 131I WBS. On the other hand, Dadparvar et al. [14] found poor sensitivity (36%) but a high specificity (89%) for 99mTc-MIBI WBS compared with 131I WBS. Miyamoto et al. [8] reported that a 99mTc-MIBI WBS did not discern pulmonary metastases in more patients than did a 131I scan (75% and 85%, respectively). Our findings suggested that, compared with 131I whole-body scanning, 99mTc-MIBI whole-body scanning was less sensitive (94.2% and 63.2%, respectively) in detecting lung metastases in DTC patients. It has been reported that the sensitivity of thoracic CT is about 80% [15]. On the contrary, some studies have shown that CT scan can detect 3 mm peripheral and 6 mm central nodules, though it still fails to discern the diffuse interstitial type of lung metastases in patients with DTC [16]. We found that CT detected lung metastases in 29/32 patients (90.6%) in our series.

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Nevertheless, our findings suggest that, compared with 131 I whole-body scanning, CT was less sensitive in detecting lung metastasis with DTC patients. It may be difficult to interpret data in an area of the neck which has already been submitted to surgery. However, we also found that fibrosis was observed in 2/32 patients in CT. De-differentiation was observed in one patient, as determined by the negative 131I WBS and positive CT, owing to the fact that CT was an additive effect to the 131 I WBS and Tg level. FDG PET scanning may be useful for locating distant metastases, especially when there is no radioiodine uptake. These metastases may be located in mediastinum or other distant areas [17,18]. FDG uptake was also detected more frequently in patients with poorly differentiated thyroid carcinoma, in whom no detectable 131I uptake could be demonstrated. FDG PET can not replace a 131I scan. However, several investigators reported that FDG PET and 131I whole-body scanning play complementary roles in the detection of recurrent metastatic DTC [18]. We were not able to use PET scanning because this modality is not available in our department. In our study, we have chosen to evaluate our retrospective data with the help of previous data instead of statistically assessing various parameters affecting the survival period. Our aim was to determine whether we had a treatment mode which would result in progress in our patients. Shoup et al. [19] analysed patient-related, tumour-related and treatment-related factors and their relation to disease specific survival using statistical tests. They found that at an age of 45 years or more, a site other than lung only or bone only, and symptoms at the time of diagnosis are associated with poorer outcomes. In addition Ronga et al. [4] reported that young age at diagnosis and radioiodine uptake by metastases are the most important factors positively affecting survival time. They also found that radioiodine therapy with high cumulative 131I activity can lead to longer survival time or complete recovery. We found a prolongation of disease-free period and progress in the parameters followed in our patients who accumulated radioiodine and were younger than 45 years at the first diagnosis. Six of the eight patients (from the 32 with lung metastases) who showed progress in the parameters followed (DWBS, Tg level less than 30 ng
ml – 1) were younger than 45 years and needed less cumulative doses than the other two patients. We believe that radioiodine therapy should continue to be given to patients with metastatic disease because it at least slows the course of the disease even though persistent or recurrent cases may not be cured.

has an additive effect to a 131I WBS. Our findings suggested that, compared with 131I whole-body scanning, 99m Tc-MIBI whole-body scanning was less sensitive (94.2% and 63.2%, respectively) in detecting lung metastases in DTC patients. MIBI imaging alone might not be enough to detect these metastases.

References 1

2

3 4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

Conclusion For the detection of lung metastases and follow-up after total thyroidectomy, 131I whole-body scanning and Tg levels were the most important parameters. A CT scan

19

Schlumberger M, Tubiana M, De Vathaire F, Hill C, Gadret P, Travagli JP, et al. Long-term results of treatment of 283 patients with lung and bone metastases from differentiated thyroid carcinoma. J Clin Endocrinol Metab 1986; 63:960–967. Ruegemer JJ, Hay ID, Bergstralh EJ, Ryan JJ, Offord KP, Gorman CA. Distant metastases in differentiated thyroid carcinoma: a multivariate analysis of prognostic variables. J Clin Endocrinol Metab 1988; 67:501–508. Hoie J, Stenwig AE, Kullmann G, Lindegard M. Distant metastases in papillary thyroid cancer. A review of 91 patients. Cancer 1988; 61:1–6. Ronga G, Filesi M, Montesano T, Di Nicola AD, Pace C, Travascio L, et al. Lung metastases from differentiated thyroid carcinoma. Q J Nucl Med Moll Imaging 2004; 48:12–19. Ozata M, Suzuki S, Miyamoto T, Liu RT, Fierro-Renoy F, Degroot L J. Serum thyroglobulin in the follow-up of patients with treated differentiated thyroid cancer. J Clin Endocrinol Metab 1994; 79:98–105. Nemec J, Nyvitova O, Blazek T, Vicek P, Racek P, Novak Z, et al. Positive thyroid cancer scintigraphy using technetium-99m methoxyisobutylisonitrile. Eur J Nucl Med 1996; 23:69–71. Ng DC, Sundram FX, Sin EA. 99mTc-sestamibi and 131I whole-body scintigraphy and initial serum thyroglobulin in the management of differentiated thyroid carcinoma. J Nucl Med 2000; 41:631–635. Miyamoto S, Kasagi K, Misaki T, Alam MS, Konishi J. Evaluation of technetium-99m-MIBI scintigraphy in metastatic differentiated thyroid carcinoma. J Nucl Med 1997; 38:352–356. Mazzaferri EL, Massoll N. Management of papillary and follicular thyroid cancer: new paradigms using recombinant human thyrotropin. Endocr Relat Cancer 2002; 9:227–247. Pacini F, Capezzone M, Elisei R, Ceccarelli C, Taddei D, Pinchera A. Diagnostic 131-iodine whole-body scan may be avoided in thyroid cancer patients who have undetectable stimulated serum Tg levels after initial treatment. J Clin Endocrinol Metab 2002; 87:1499–1501. Morgenthaler NG, Froehlich J, Rendl J, Willnich M, Alonso C, Bergmann A, Reiners C. Technical evaluation of a new immunoradiometric and a new immunoluminometric assay for thyroglobulin. Clin Chem 2002; 48: 1077–1083. Filesi M, Signore A, Ventroni G, Melacrinis FF, Ronga G. Role of initial iodine131 whole-body scan and serum thyroglobulin in differentiated thyroid carcinoma metastases. J Nucl Med 1998; 39:1542–1546. Sundram FX, Goh AS, Ang ES. Role of technetium-99m sestamibi in localization of thyroid cancer metastases. Ann Acad Med Singapore 1993; 22:557–559. Dadparvar S, Chevres A, Tulchinsky M, Krishna-Badrinath L, Khan AS, Sizofski WJ. Clinical utility of technetium-99m methoxyisobutylisonitrile imaging in differentiated thyroid carcinoma: comparison with thallium-201 and iodine-131 scintigraphy and serum thyroglobulin quantitation. Eur J Nucl Med 1995; 22:1330–1338. Lorenzen J, Beese M, Mester J, Brumma K, Beyer W, Clausen M. Chest X ray: routine indication in the follow-up of differentiated thyroid cancer? Nuklearmedizin 1998; 37:208–212. Piekarski JD, Schlumberger M, Leclere J, Couanet D, Masselot J, Parmantier C. Chest computed tomography in patients with micronodular lung metastases of differentiated thyroid carcinoma. Int J Radiat Oncol Biol Phys 1985; 11:1023–1027. Wang W, Macapinlac H, Larson SM, Yeh SD, Akhurst T, Finn RD, Rosai J, Robbins RJ. 18F-2-Fluoro-2-D-glucose positron emission tomography localizes residual thyroid cancer in patients with negative diagnostic I-131 iodine whole body scans and elevated serum thyroglobulin levels. J Clin Endocrinol Metab 1999; 84:2291–2302. Shiga T, Tsukamoto E, Nakada K, Morita K, Morita K, Kato T, et al. Comparison of (18)F-FDG, (131)I-Na, and (201)Tl in diagnosis of recurrent or metastatic thyroid carcinoma. J Nucl Med 2001; 42:414–419. Shoup M, Stojadinovic A, Nissan A, Ghossein RA, Freedman S, Breennan MF, et al. Prognostic indicators of outcomes in patients with distant metastases from differentiated thyroid carcinoma. J Am Coll Surg 2003; 197:191–197.

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Original article

Striatal dopamine transporter density in drug naive patients with attention-deficit/hyperactivity disorder Rolf Larischa, Wolfgang Sitteb, Christina Antkea, Susanne Nikolausa, Matthias Franzb, Wolfgang Tressb and Hans-Wilhelm Mu¨llera Background and aim Dopamine transporters are the target of psychostimulants used for treatment of attention-deficit/ hyperactivity disorder (ADHD). Therefore, the present study aimed to evaluate striatal dopamine transporter density in adult patients with ADHD. Methods Twenty patients (11 female, nine male; mean age 35 ± 7 years) and 20 control subjects (11 female, nine male, mean age 32 ± 8 years) were examined with SPECT using the specific radiotracer 123I-FP-CIT. The ratio of striatal to cortical radioactivity concentration was used for semiquantitative evaluation of dopamine transporter binding potential (V300 ). There was a significant influence of age (P < 0.001) and a trend towards an influence of gender (P = 0.053) on V300 . An ANCOVA with these covariates showed a slightly higher V300 in the patients than in the control subjects (4.24 ± 0.48 vs. 4.03 ± 0.56; P = 0.02).

Introduction The most common psychiatric condition in children affecting attention, impulsivity and motor activity is attention-deficit/hyperactivity disorder (ADHD). The prevalence of this syndrome ranges from 5 to 10% in childhood [1]. Moreover, it appears that in about one third of children symptoms of ADHD fail to vanish in adulthood [2,3]. Although the pathophysiology of ADHD is not understood so far, the central role of dopaminergic neurotransmission is clearly documented. The most convincing evidence comes from the demonstration of the shortterm efficacy of psychostimulants like the dopamine reuptake inhibitor methylphenidate [4]. Therefore, one focus of research using in-vivo imaging methods has been the cerebral dopamine transporter in children [5–7] and in adults [8–11]. Initial evidence pointed towards an increase of striatal dopamine transporter density in ADHD patients as compared to healthy controls [5,8,9]. However, a consecutive study reported no difference between patients and control subjects [10]. Moreover, two papers even reported a decrease of the patients’ dopamine transporter density in striatum [11] and midbrain [7]. In the present study, the largest sample, so far, of wellselected and drug naive adult subjects was examined.

Conclusion This study provides further in-vivo evidence for an involvement of the dopamine transporter in ADHD. However, compared to previous studies, the increase of dopamine transporter density in the patient group is less c 2006 pronounced here. Nucl Med Commun 27:267–270
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:267–270 Keywords: FP-CIT, ADHD, SPECT, in-vivo imaging, human studies a

Clinic of Nuclear Medicine and bDepartment of Psychosomatic Medicine and Psychotherapy, University Hospital Du¨sseldorf, Germany.

Correspondence to Dr Susanne Nikolaus, Clinic of Nuclear Medicine, University Hospital Du¨sseldorf, Moorenstr. 5, 40225 Du¨sseldorf, Germany. Tel: + 0049 0211 811 7048; fax.: + 0049 0211 811 7041; e-mail: [email protected] Received 22 August 2005 Accepted 7 December 2005

Dopamine transporter density was measured using the specific ligand N-o-fluoropropyl-2b-carbomethoxy-3b-(4[123I]iodophenyl)nortropane (123I-FP-CIT) in 20 adult patients with ADHD and compared to 20 age-matched and gender-matched healthy control subjects.

Subjects, materials and methods Twenty consecutive ADHD patients (11 female, nine male; age 35 ± 7 years (mean ± standard deviation)) were recruited from subjects referred to the clinic for psychosomatic medicine and psychotherapy. In order to establish the diagnosis of ADHD according to DSM-IV criteria [12], patients were evaluated by an experienced clinician. Thereby, contemporary information and patients’ recollections of childhood behaviour (school reports, parent interview) were used. The diagnosis and onset of ADHD symptoms in childhood was confirmed psychometrically using the ‘Wender–Utah Rating Scale’ [13]. Persistent relevant problems were assessed using the Brown Add Scales [14] and the Conner’s Adult ADHD Rating Scale [15]. Other current neurological or psychiatric diseases, including substance abuse, chronic suicidal or self-injurious behaviour were exclusion criteria. Moreover, all patients were naive for any pharmaceuticals that act on the central nervous system or for recreational drugs. Twenty healthy control subjects (11 females, nine males; age 32 ± 8 years) were recruited by an internet

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Nuclear Medicine Communications 2006, Vol 27 No 3

advertisement. They were also naive for any substances acting on the central nervous system. Psychiatric disorders were excluded by anamnesis and physical examination. Each subject gave written informed consent to the study. The study protocol was approved by the Ethics Committee of the University of Du ¨sseldorf and by the federal authorities for radiation protection.

Fig. 1

5.5 5.0 4.5 V3″

268

4.0

123

I-FP-CIT was provided by Amersham Health (Braunschweig, Germany). Subjects received 185 MBq of the tracer intravenously in high specific activity ( > 100 TBqmmol – 1) between 8 and 9.30 a.m. after blockade of thyroid iodine uptake by perchlorate. They were asked to physically relax for the subsequent 30 min. Scanning was done exactly 4 h after injection using a double-headed gamma camera (PRISM 2000, Phillips) with high resolution, parallel hole collimation. At this time point, pseudo-equilibrium of the tracer distribution is reached [16]. A total of 120 projections were obtained within 30 min using a 3601 circular rotation of the detectors. The energy window was set to 159 ± 16 keV. Anatomical orientation was ensured by external fiducial markers filled with about 50 kBq 123I and placed close to the acoustic meatus and the lateral orbita of the subjects.

Reconstruction of the 123I-FP-CIT scans was done in 128  128 matrices using filtered back-projection with a low-pass filter. Attenuation correction was performed assuming uniform attenuation equal to water for an ellipse drawn around the brain similar to Chang’s method. The in-plane resolution (full width at half maximum) of the reconstructed single photon emission computed tomography (SPECT) images was 13 mm, the axial 15 mm. The reconstructed images were realigned in 15 slices of 8 mm thickness parallel to the meato-orbital line according to the fiducial markers. The highest activity resulting from striatal tracer accumulation was found in the middle slide. Two standardized, laterally reversed oval regions of interest were placed over the left and right striatum in the middle slice. The circular reference region was placed medially in the occipital cortex on the same slice. The region templates were initially derived from a series of normal FP-CIT scans. The binding potential of the striatal dopamine transporters was calculated analogously to previous studies of postsynaptic receptor density as V300 [17]. For this, the striatal radioactivity concentration was divided by the radioactivity concentration in the reference region.

Results All 40 subjects displayed similar left and right striatal dopamine transporter density (4.14 ± 0.53 vs. 4.12 vs.

3.5 3.0 2.5 15

20

25

30 35 Age (years)

40

45

50

Influence of age and diagnostic group on the dopamine transporter density (V300 ). The filled circles represent the ADHD patients; the open circles were the control subjects. The solid and the dotted line refer to the regression lines for patients and controls, respectively. Dopamine transporter density declined by about 7% per decade. The coefficient r was 0.53 (P < 0.001). The differences between patients and controls were significant (ANCOVA; P < 0.02).

0.53; P = 0.85, t-test). This was also true for the subgroups of patients (4.24 ± 0.48 vs. 4.23 ± 0.49; P = 0.95) and controls (4.04 ± 0.57 vs. 4.01 ± 0.56; P = 0.85), respectively. Therefore, for the subsequent analyses, the mean of both striata was used. Women had higher V300 values than men (4.28 ± 0.55 vs. 3.96 ± 0.45). This difference was borderline significant (P = 0.053, two-sided t-test). The 16 smokers had lower V300 values than the 24 nonsmokers (4.01 ± 0.35 vs. 4.21 ± 0.61). However, this difference was insignificant (P = 0.24). A linear regression showed the influence of age on V300 . Dopamine transporter density declined by about 7% per decade. The coefficient r was 0.53 (P < 0.001). Thus, gender and age but not smoking habits were considered covariates and were included into the final analysis of group effects on V300 using ANCOVA. This test showed a significant influence of the covariate age (P < 0.001) and a significant group effect (patient vs. control; P < 0.02) but no influence of sex (P = 0.18). The ADHD patients had the higher dopamine transporter density (Fig. 1). At the age of 30, the mean group difference was about 7%.

Discussion The main finding of the present study was an increase of striatal dopamine transporter density in adult ADHD patients as compared to healthy controls. Apart from this finding, age but not the smoking habits of the subjects had an effect on striatal dopamine transporters. Women

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Dopamine transporters and ADHD Larisch et al. 269

had higher dopamine transporter densities than men. However, this difference was not significant in the ANCOVA.

However, at 7% the extent of increase was markedly lower in the present study as in the previous ones and only just significant.

The age-dependent decrease of striatal dopamine transporter density is consistent with earlier reports on this subject [18–20]. Moreover, the magnitude of this agedependent decrease of about 7% per decade is in good accordance with the previous data.

The described differences between studies could be due to varieties regarding both patient selection or the choice of radiotracer. All applied radiotracers were tropane derivatives with high sensitivity and specificity for the dopamine transporter. Furthermore, the in-vivo use of these tracers for dopamine transporter imaging was carefully validated beforehand. Thus, it seems unlikely that the choice of radiotracer might result in the observed differences between studies. In all the studies performed, including the present one, patients were selected carefully according to the criteria of the DSM-IV. These criteria were based on specific meaningful self-rating and external rating and careful physical examination. Nevertheless, there seem to be ADHD patients exhibiting pronounced increases in striatal dopamine transporter density, whereas others have similar or even lower values compared to healthy controls. A reason for this might be the different expression of clinical symptoms with, e.g., either more pronounced hyperactivity or attention deficit symptoms. Another explanation could be that the total group of ADHD subjects consists of several subgroups and that only some of them involve the dopamine transporter. Therefore, we suggest that future studies should relate clinical parameters with the dopamine transporter density. This way, firstly the subgroup of ADHD patients with an involvement of dopamine transporters as the biological origin of their disorder may be identified. Secondly, inter-relations between clinical symptoms and the individual dopamine transporter densities might be detected.

A previous study using 123I-FP-CIT reported higher dopamine transporter densities in women than in men [18]. However, another group using 123I-b-CIT found similar values for both sexes [19]. Here, women had higher 123 I-FP-CIT binding than men. However, the differences were only borderline significant and vanished completely when the covariate age was taken into account. It should be emphasized that in the present study any precedent intake of recreational drugs by the subjects was strictly ruled out. The only exception was nicotine. Actually, half of our patients were cigarette smokers. Unlike a previous report [21], we found no differences in dopamine transporter density between smokers and nonsmokers. Thus, a possible interaction between nicotine and the dopamine transporter is not supported by the present study. Striatal dopamine transporter density in ADHD patients as compared with healthy controls has already been examined in studies with small sample sizes. The Harvard group studied six adult ADHD patients using 123 I-altropane and compared them with their database of control subjects. The patients had a 70% higher dopamine transporter density [8]. The Munich group examined ten adult patients with ADHD and ten control subjects using 99mTc-TRODAT-1 and found a 16% higher dopamine transporter density in the patient group [9,22]. These data on adult subjects were replicated in children. Cheon and colleagues examined nine ADHD children between 6 and 12 years using 123I-IPT and compared them with six healthy children. The ADHD subjects displayed a 50% higher striatal 123I-IPT accumulation than the controls [5]. The Yale group used 123I-b-CIT for dopamine transporter imaging in nine adult ADHD patients and nine healthy controls. In contrast to the authors cited above, they reported no significant difference between patients and controls [10]. Moreover, the Brookhaven group even found dopamine transporter densities lowered by 12% in their group of five adult ADHD subjects who were examined with 11C-cocaine PET and compared with nine controls [11]. The Karolinska group reported lower binding potentials for dopamine transporter in the midbrain of 12 ADHD patients as compared to 10 controls [7]. Here, we confirm the increase of dopamine transporter density in ADHD.

Acknowledgements This work was supported by a grant of Amersham Health Ltd (Braunschweig, Germany). The authors acknowledge the technical assistance of Mrs Manuela Beckert, Mrs Doris Buchholz, Mrs Esther Pa¨ch, Mrs Jola Tomkiewicz and Mrs Ute Werner in performing the SPECT studies.

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Szatmari P, Offord DR, Boyle MH. Ontario Child Health Study: prevalence of attention deficit disorder with hyperactivity. J Child Psychol Psychiatry 1989; 30:219–230. Biederman J. Attention-deficit/hyperactivity disorder: a life-span perspective. J Clin Psychiatry 1998; 59(suppl 7):1093. Lie N. Follow-ups of children with attention deficit hyperactivity disorder (ADHD). Review of literature. Acta Psychiatr Scand Suppl 1992; 368:1–40. Spencer T, Biederman J, Wilens T, Harding M, O’Donnell D, Griffin S. Pharmacotherapy of attention-deficit hyperactivity disorder across the life cycle. J Am Acad Child Adolesc Psychiat 1996; 35:409–432. Cheon KA, Ryu YH, Kim YK, Namkoong K, Kim CH, Lee JD. Dopamine transporter density in the basal ganglia assessed with [123I]IPT SPET in children with attention deficit hyperactivity disorder. Eur J Nucl Med Mol Imaging 2003; 30:306–311. Vles JS, Feron FJ, Hendriksen JG, Jolles J, van Kroonenburgh MJ, Weber WE. Methylphenidate down-regulates the dopamine receptor and transporter system in children with attention deficit hyperkinetic disorder (ADHD). Neuropediatrics 2003; 34:77–80.

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Jucaite A, Fernell E, Halldin C, Forssberg H, Farde L. Reduced midbrain dopamine transporter binding in male adolescents with attention-deficit/ hyperactivity disorder: association between striatal dopamine markers and motor hyperactivity. Biol Psychiatry 2005; 57:229–238. Dougherty DD, Bonab AA, Spencer T, Rauch SL, Madras BK, Fischman AJ. Dopamine transporter density in patients with attention deficit hyperactivity disorder. Lancet 1999; 354:2132–2133. Dresel S, Krause J, Krause KH, LaFougere C, Brinkbaumer K, Kung HF, et al. Attention deficit hyperactivity disorder: binding of [99mTc]TRODAT-1 to the dopamine transporter before and after methylphenidate treatment. Eur J Nucl Med 2000; 27:1518–1524. van Dyck CH, Quinlan DM, Cretella LM, Staley JK, Malison RT, Baldwin RM, et al. Unaltered dopamine transporter availability in adult attention deficit hyperactivity disorder. Am J Psychiatry 2002; 159:309–312. Volkow ND, Sanders M, Wang GJ, Newcorn J, Solanto M, Schultz K, et al. Brain dopamine transporters and D2 receptors in never medicated adults with ADHD. J Nucl Med 2002; 43(suppl):16P. American Psychiatric Association. The Diagnostic and Statistical Manual of Mental Disorders, fourth edition (DSM IV), 1994. Ward MF, Wender PH, Reimherr FW. The Wender Utah Rating Scale: an aid in the retrospective diagnosis of childhood attention deficit hyperactivity disorder. Am J Psychiatry 1993; 150:885–890. Brown TE. Differential diagnosis of ADD versus ADHD in adults. In: Nadeau KG (editor): A Comprehensive Guide to Attention Deficit Disorder in Adults. New York: Brunner/Mazel; 1995, pp. 93–108.

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Conners CK, Erhardt D, Sparrow E. Multi-Health Systems. New York: North Tonawanda; 1999. Booij J, Hemelaar JTGM, Speelman JD, de Bruin K, Janssen AGM, van Royen EA. One-day protocol for imaging of the nigrostriatal pathway in Parkinson’s disease by [123I]FP-CIT. J Nucl Med 1999; 40:753–761. Laruelle M. Imaging synaptic neurotransmission with in vivo binding competition techniques: a critical review. J Cereb Blood Flow Metab 2000; 20:423–451. Lavalaye J, Booij J, Reneman L, Habraken JB, van Royen E. Effect of age and gender on dopamine transporter imaging with [123I]FP-CIT SPET in healthy volunteers. Eur J Nucl Med 2000; 27:867–869. van Dyck CH, Seibyl JP, Malison RT, Laruelle M, Wallace E, Zoghbi SS, et al. Age-related decline in striatal dopamine transporter binding with iodine-123beta-CITSPECT. J Nucl Med 1995; 36:1175–1181. Volkow ND, Ding YS, Fowler JS, Wang GJ, Logan J, Gatley SJ, et al. Dopamine transporters decrease with age. J Nucl Med 1996; 37: 554–559. Krause KH, Dresel S, Krause J, Kung HF, Tatsch K, Ackenheil M. Stimulantlike action of nicotine on striatal dopamine transporter in the brain of adults with attention deficit hyperactivity disorder. Int J Neuropsychopharmacol 2002; 5:111–113. Krause KH, Dresel SH, Krause J, Kung HF, Tatsch K. Increased striatal dopamine transporter in adult patients with attention deficit hyperactivity disorder: effects of methylphenidate as measured by single photon emission computed tomography. Neurosci Lett 2000; 285:107–110.

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Original article

Preparation of 99mTc-labelled conjugates of ouabagenin and their biological evaluation in animal models Mita C. Debnatha, Bharat R. Sarkarb, Shantanu Gangulyb and Somenath Banerjeea Background Ouabagenin and its 1,19-acetonide were conjugated with nitrilotriacetic acid (NTA) and diethylenetriaminepentaacetic acid (DTPA) through their respective anhydrides. Methods The reaction mixtures were exhaustively purified by silica gel column chromatography and preparative highperformance liquid chromatography to furnish the ligands in good purity and moderate yield. These ligands were labelled with 99mTc to produce four chelates in 90–95% yield. Of these chelates the 99mTc-oubagenin–NTA conjugate and the corresponding acetonide exhibited appreciable myocardial uptake with respect to that of other vicinal organs in a guinea-pig model. However, all these 99mTc chelates exhibited poor heart-to-blood ratios, which could be attributed to the absence of a 3b sugar residue in this molecule.

digoxigenin derivatives. Nucl Med Commun 27:271–279 2006 Lippincott Williams & Wilkins.


c

Nuclear Medicine Communications 2006, 27:271–279 Keywords: oubain, Na + K + -ATPase, cardiac glycoside, 201Tl + , 99m Tc-ouabagenin–NTA conjugate, 99mTc-ouabagenin–DTPA conjugate a Department of Nuclear Medicine, Indian Institute of Chemical Biology, Kolkata and bRegional Radiation Medicine Center, Thakurpukur Cancer Center and Welfare Home Campus, Kolkata, India.

Correspondence to Dr Mita Chatterje Debnath, Department of Nuclear Medicine, Indian Institute of Chemical Biology, 4, Raja S.C. Mullick Road, Jadavpur, Kolkata 700032, India. Tel: + 0091 33 2473 3491; fax: + 0091 33 2473 5197; e-mail: [email protected] Financial support for this work was received from the Indian Council of Scientific and Industrial Research.

Conclusion The result is in agreement with that previously reported in connection with radioiodinated digoxin and

Introduction The development of 99mTc-based myocardial imaging agents to replace cyclotron-produced 201Tl is an important target of cardiovascular nuclear medicine [1]. 201 Tl images myocardium by targeting myocardial membrane bound Na + ,K + -ATPase, which is of major importance in myocardial function [2]. Several cationic 99m Tc chelates [3,4] and related compounds [5,6] were developed with the objective of targeting myocardial Na + ,K + -ATPase. Although they image myocardium acceptably well neither of these compounds is transported by Na + ,K + -ATPase: rather, their myocardial uptake occurs by a simple passive diffusion method [7]. Therefore they lack the sensitivity [8] of a typical K + analogue in detecting early pathological changes that occur in myocardium. Some non-K + analogues, such as 99mTc-tetrofosmin [9], a neutral lipophilic chelate of technetium, and several neutral and cationic 99mTc-nitrido-dithiocarbamato analogues, have also been developed [10,11]. Although they exhibit preferential myocardial cell retention none of these agents was an ideal technetium analogue of 201Tl + . The concentration of Na + ,K + -ATPase can be estimated by measuring its binding capacity with 3H-oubain [12],

Received 8 August 2005 Accepted 5 December 2005

therefore imaging with ouabain may provide a sensitive method for evaluating myocardial function. 99m

Tc, though, proved to be a useful tracer for macromolecules [13] but labelling of smaller molecules with this tracer usually causes a drastic alteration of ligand bioactivity [14]. Though the above observation is accepted as a general one there are many studies [15] which report that the low molecular weight ligands can retain an appreciable amount of their bioactivity after 99m Tc chelation. Examples of radiometal binding of steroids have also reported [16] where several oestradiene derivatives were modified with an N2S2 chelating moiety. After radiolabelling with 99mTc or 186Re, the synthesized bifunctional ligand showed the expected receptormediated uptake characteristics of the progesterone receptor present in rat uterus. It would be interesting to see whether ouabain, if subjected to similar molecular modification, can produce any useful image of myocardial Na + ,K + -ATPase. We have already radiolabelled ouabagenin–cysteine conjugates with 99mTc [17]. The biological properties of these chelates closely resemble those reported for radioiodinated digoxigenin derivatives [18] and both types of compound showed an appreciable accumulation

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272 Nuclear Medicine Communications 2006, Vol 27 No 3

in the myocardium. Similar observations with other cardiac glycosides radiolabelled with 99mTc have been reported previously from this laboratory [19]. In a continuation of the above pilot studies, in this study we have conjugated ouabagenin and its acetonide with diethylenetriaminepentaacetic acid (DTPA). DTPA conjugates are quite popular in radiometal labelling of macromolecular proteins [20] and merit investigation with other bioactive systems for similar utility. Therefore, radiolabelling of ouabagenin and its derivatives with 99m Tc, through their DTPA conjugates, appears to be a plausible approach. However, when selecting a 99mTc binding tether for conjugation with ouabagenin proper consideration should be given to its small molecular size; it is expected that a low molecular weight tether may be more suitable for this purpose. Whether DTPA will meet this expectation is not known but previous studies from this laboratory have shown that, after 99mTc chelation, the ouabagenin–cysteine conjugate accumulated appreciably in the myocardium, as mentioned earlier. Therefore, in addition to DTPA we have also conjugated ouabagenin with nitrilotriacetic acid (NTA), which is a DTPA analogue of low molecular weight. Both these types of chelate were labelled with 99mTc and used for biodistribution studies in animals. From the comparative biodistribution studies with these chelates it would be interesting to determine which of the above tethers, viz. NTA and DTPA, is more compatible for conjugation with ouabagenin. It is expected that the more compatible tether, after conjugation with ouabagenin and subsequent radiolabelling with 99m Tc, will produce a bifunctional chelate with a better heart-to-non-target ratio. The results are given in the following sections.

Materials and methods Melting points were determined in open capillary tubes and are uncorrected. Ultraviolet spectra of the compounds in methanolic solution were recorded by using a Hitachi U-2000 spectrophotometer. Infrared spectra of the compounds in potassium bromide disks were recorded by a JASCO 700 spectrophotometer. 1H NMR spectra of the compounds in DMSO-d6 or D2O were recorded by using a 400 MHz Bruker VM-400 spectrometer. Chemical shifts (d) were expressed in ppm. Significant signals were tabulated in the order: chemical shift, number of protons, multiplicity (s, singlet; d, doublet; t, triplet; m, multiplet; br, broad) coupling constant and designation. Elemental analyses were performed in the organic chemistry department of Jadavpur University. 99MoO4– was purchased from Bhabha Atomic Research Center and 99mTcO4– was obtained by 2-butanone extraction from a 5 N NaOH solution of 99 MoO4– . Thin-layer chromatography (TLC) experiments

were performed with pre-coated F254 plates (E. Merck, Germany) with the solvents indicated. Preparative reverse-phase high-performance liquid chromatography (HPLC) was performed by using a m-bondapak C18 column (0.78  30 cm) in a linear gradient of 0.1% trifluoroacetic acid (TFA) with increasing concentration of CH3CN. The HPLC system used consisting of a Model 501 pump U6 K injector, 481 variable wavelength UV detector and a Waters Automated Gradient Controller. After purification, the ligands were assessed by analytical reverse-phase HPLC using a Waters m-bondapak C18 analytical column (0.39  30 cm). Analytical HPLC of the 99mTc chelates was performed isocratically on the same column using phosphate buffer (0.01 M) and a CH3CN mix in different proportions as the mobile phase. The column eluates were analysed by a Beckman Model 170 radioisotope detector with integrator. Various countings were performed in a well type gamma counter from Electronic Corporation of India, Model LV 4755. Synthesis of the ligands

The structures of ouabagenin, ouabagenin monoacetonide (compound 1), a starting point for the synthesis, and of compounds 2 to 6 (the ligands) are shown in Fig. 1. Compound 2

Full chemical name: 1b-19-(1-methylethylidene dioxy)3b,11a-(N,N0 -dicarboxymethyl)aminoacetyl-5,14-di;hydroxy-5b,17-card-20(22)-enolide Alternative name: ouabagenin acetonide-3b,11a-bis(iminodiacetate)

Ouabagenin monoacetonide (1, 0.9 g, 1.89 mmol) was added to nitrilotriacetic anhydride (prepared according to Burns’ procedure [21] using nitrilotriacetic acid (6.0 g, 31.2 mmol)), pyridine (90 ml) and acetic anhydride (3.0 ml, 31.2 mmol). The mixture was stirred at room temperature for 48 h during which time the ouabagenin monoacetonide went into solution. The above solution was stirred continuously at 401C for another 48 h, the solvent was evaporated in vacuo, and the semi-solid mass thus obtained was solidified by ether trituration (50 ml  3). Methanol (20 ml) was added to the solid residue and the mixture stirred and filtered. The filtrate was concentrated and any solid (unreacted tri-acid) that appeared at this stage was filtered off and the solution was chromatographed over a column of silica (1.2  30 cm). The column was washed sequentially with CHCl3/MeOH (95:5, 90:10, 85:15 (v/v) mixtures) and then eluted with CHCl3/MeOH 80:20 v/v to furnish 2 (0.75 g) as a glassy solid in an elution volume of 125–270 ml. The products were further purified by reverse-phase HPLC using a m-bondapak C18 preparative column (30  7.8 cm). Solvent A was a 95:5 (v/v) mixture of H2O (containing 0.1% TFA (pH 2.4)) and CH3CN;

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Preparation and evaluation of

99m

Tc-ouabagenin compounds Debnath et al. 273

Fig. 1

O

O O

OR

O

OR OH

O O

HO CH2

CH2 OH

OH RO

RO

OH

OH

1R=H 2 R = CO.CH2 -N 4 R = CO.CH2 -N

CH2CO2H CH2CO2H

3 R = as in 2 6 R = as in 5

CH2-CO2CH3 CH2-CO2CH3

5 R = COCH2-N -CH2 -CH2N -CH2 -CH2 - N

CH2 - CO2 H CH2 - CO2 H

CH2 -CO2H CH2 -CO2H Chemical structures of nitrilotriacetic acid and diethylenetriaminepentaacetic acid conjugates of ouabagenin and its acetonide. The structure on the left is ouabagenin monoacetonide (1). If R = H the structure on the right is ouabagenin. Compounds 2 to 6 are as follows: 2: Ouabagenin acetonide 3b,11a-bis(iminodiacetate) 3: Ouabagenin 3b,11a-bis(iminodiacetate) 4: Ouabagenin acetonide 3b,11a-bis(methyliminodiacetate) 5: Ouabagenin acetonide 3b,11a-bis(diethylenetriaminepentaacetate) 6: Ouabagenin 3b,11a-bis(diethylenetriaminepentaacetate)

solvent B was HPLC grade CH3CN. Initial conditions were set at A = 100% and B = 0%, with a flow rate of 2.4 ml  min – 1. After injection, a 10 min linear gradient to A = 0% and B = 100% was applied, followed by a 3 min hold at this condition. After 400 injections, the fractions (tR = 9.28 min) were pooled, the pH adjusted to 7.0 with NH4OH, and the solution evaporated in vacuo to furnish a white solid, which was further recrystallized from aqueous MeOH to obtain the analytical material (0.4 g, 26%). Analytical data Melting point: 186–1901C UV lmax (nm): 217 (e = 22 387) IR u (cm – 1): 3420–3360 (OH), 3120, 3100 (NH + ), 1730–1720 (C = O of ester and lactone), 1650–1610 (C-O, COO – , 1400, 1215, 1150, 1085, 1030 1 H NMR (DMSO-d6, d): 0.82 (3H, s, 18-H3), 1.16 and 1.3 (3H, ea, s, acetonide CH3 groups), 2.84 (1H, m, 17a-H), 3.52 (12H, s, 2  OCO-CH2-N-(CH2-COOH)2), 4.38 (1H, m, 1a-H), 4.90 (22H, br, s, 21-H2), 5.08–5.32 (2H, m, 3a-H and 11b-H), 5.94 (1H, s, 22-H)

Quantitative analysis calculated for C38H52N2O18: C, 55.39; H, 6.35; N, 3.41. Found: C, 55.16; H, 6.60; N, 3.15 Compound 3

Full chemical name: 3b,11a-di(N,N0 -dicarboxymethyl) aminoacetyl-1b,5,14b,19-tetrahydroxy-5b,17b-card-20(22)enolide Alternative name: ouabagenin-3b,11a-bis(iminodiacetate)

Sulfuric acid (20%, 0.6 ml) was added to a suspension of the monoacetonide derivative 2, as prepared above (0.3 g, 0.36 mmol), in ethanol (20%, 20 ml), and the mixture warmed over a water bath (601C) for 15 min, then cooled to room temperature and shaken with freshly precipitated BaCO3 (prepared from 3 g of BaCl2 and 1.5 g of Na2CO3). The mixture was filtered, the filtrate evaporated, and the residue was dissolved in the minimum volume of methanol. On the dropwise addition of water a very fine powder separated which was further recrystallized from water to furnish pure 3 (0.21 g, 74%).

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274 Nuclear Medicine Communications 2006, Vol 27 No 3

Analytical data Melting point: 298–3001C (decomposes) UV lmax (nm): 206 (e = 16596) IR u (cm – 1): 3420–3380 (OH), 2980–2920 (NH + ), 1740–1720 (C = O of ester and lactone), 1620–1560 (C = O, COO – ) 1 H NMR (D2O, d): 0.96 (3H, s, 18-H3), 2.94 (1H, m, 17a-H), 3.63 (12H, s, 2  OCOCH2-N (CH2-COOH)2), 3.78 and 3.94 (1H, ea, AB doublet J = 12H2, 19-H2), 4.28 (1H, m, 1a-H), 5.04 (2H, s, 21-H2), 5.25 and 5.40 (1H, ea, s, 3a-H and 11b-H), 6.0 (1H, s, 22-H) Quantitative analysis calculated for C35 H48N2O18: C, 53.62; H, 6.16; N, 3.35. Found: C, 53.25; H, 6.29; N, 3.97

Compound 4

Full chemical name: 1b,19(1-methylethylidenedioxy)3b,11a-di(N,N0 -dimethoxy carbonyl methyl)aminoacetyl5,14b-dihydroxy-5b,17-card-20(22)-enolide Alternative name: ouabagenin acetonide-3b,11abis(methyliminodiacetate) Diazomethane generated in ethereal solution from nitrosomethyl urea (1 g) and KOH (40%, 1 ml) was washed with cold water and added portion-wise to 2 (0.2 g, unchromatographed) suspended in ether (10 ml). The clarity of the solution was maintained by addition of methanol. A few drops of acetic acid were added to the reaction mixture, which was washed once with ether (30 ml) and extracted with ethyl acetate (30 ml  3). The combined organic layer was washed successively with saturated bicarbonate solution, 20% KOH solution, saturated brine solution (dried over anhydrous Na2SO4) and evaporated to furnish an oily material (0.16 g) which was chromatographed over a column of silica gel (1  18 cm). The column was washed sequentially with CHCl3, CHCl3/MeOH (97:3, v/v) and then eluted with CHCl3/MeOH (95:5 v/v) to furnish a foamy solid in the elution volume of 15–35 ml, which was purified by recrystallization from CHCl3/MeOH to furnish the pure compound (0.05 g, 20%). Analytical data Melting point: 122–1241C UV lmax (nm): 214 (e = 20 417) IR u (cm – 1): 3480–3340 (OH), 3160–3120 (NH + ), 1750–1720 (C = O of ester and lactone), 1640–1610 (C = C, NH + ), 1430, 1380, 1210, 1180, 1060, 1010 1 H NMR (CDCl3, d): 0.91 (3H, s, 18-H3); 1.19 and 1.34 (3H, ea, s, acetonide CH3 groups), 2.80 (1H, m, 17a-H), 3.64–3.68 (24, m, 2  OCO-CH2-N(CH2-COOCH3)2, 4.24 (1H, m, 1a-H), 4.80 (2H, s, 21-H2), 5.22–5.32 (2H, m, 3a-H and 11b-H), 5.88 (1H, s, 22-H) Quantitative analysis calculated for C42H60N2O18: C, 57.32; H, 6.87; N, 3.18. Found: C, 57.13; H, 7.26; N, 2.84

Compound 5

Full chemical name: 1b-19(1-methylethylidenedioxy) 3b,11a-di(N,N 0 -bis[2(bis[carboxymethyl]amino)ethyl] glycinyl)-5,14b-dihydroxy-5b,17b-card-20(22)-enolide Alternative name: ouabagenin acetonide-3b,11abis(diethylenetriaminepentaacetate)

A solution of ouabagenin monoacetonide (1, 0.45 g, 0.95 mmol) in dimethylformamide (3 ml) was added to a solution of diethylenetriaminepentaacetic anhydride (5.35 g, 14.97 mmol) in pyridine (150 ml). The reaction mixture was stirred for 72 h at 401C. The solvent was evaporated in vacuo and the semisolid mass thus obtained was solidified by ether trituration (50 ml  3). Methanol (20 ml) was added to the solid residue, which was stirred and filtered. The filtrate was concentrated in vacuo to furnish an oily material (0.88 g) which was chromatographed over a column of silica (1.2  24 cm). The column was washed successively with CHCl3/MeOH (90:10 v/v and 80:20 v/v) and then eluted with CHCl3/ MeOH (65:35 v/v) to furnish 5 as glassy solid. The material was further purified by reverse-phase HPLC using m-bondapak C18 column (30  0.78 cm). Solvent A was a 90:10 v/v mix of H2O (containing 0.1% TFA (pH 2.4) and CH3CN; solvent B was HPLC grade CH3CN. Initial conditions were set at A = 100%, B = 0% with a flow rate of 1.8 ml  min – 1. After injection, a 12 min linear gradient to A = 0%, B = 100% was applied followed by a 5 min hold at this condition. After 250 injections, the fractions (tR = 9.59) were pooled, the pH adjusted to 7.0 with NH4OH, and the solution evaporated in vacuo to furnish white solid, which was further recrystallized from aqueous MeOH to obtain the analytical material (0.25 g, 30%). Analytical data Melting point: 212–2141C (decomposes) UV lmax (nm): 2216 (e = 18 356) IR u (cm – 1): 3400–3380 (OH), 3000–2980 (NH + ), 1740–1720 (C = O of ester and lactone), 1560 (C = O, COO – ), 1450, 1310 1 H NMR (D2O, d): 0.822 (3H, s, 18-H3), 0.94 and 1.02 (3H, ea, s, acetonide CH3 groups), 2.92 (1H, m, 17a-H), 3.64–3.89 {(36H, m, 2  O2C-CH2N-(CH2-CO2– )CH2-CH2N(-CH2CO2– )-CH2-CH2N(-CH2CO2– )2}, 5.04 (2H, s, 21-H2), 5.36–5.45(2H, m, 3a-H and 11b-H), 6.0 (1H, s, 22-H) Quantitative analysis calculated for C54H80N6O26: C, 52.77, H, 6.32, N, 6.84. Found: C, 52.59, H, 6.82, N, 6.49

Compound 6

Full chemical name: 3b,11a-di(N,N0 -bis[2-{bis(carboxymethyl)aminoethyl}glycinyl])-1b,5,14b,19-tetrahydroxy5b,17b-card-20(22)-enolide

Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.

Preparation and evaluation of

Alternative name: ouabagenin-3b,11a-bis(diethylenetriamine pentaacetate)

Compound 6 was prepared by treating 5 (0.3 g, 0.60 mmol) with sulfuric acid (20%, 0.6 ml) in ethanol (20%, 15 ml) by a similar procedure to that described for the preparation of 2 and 3. The product was recrystallized from water to furnish pure material (0.2 g, 25%). Analytical data Melting point: 288–2901C (decomposes) UV lmax (nm): 210 (e = 16 580) IR u (cm – 1): 3420–3360 (OH), 2960–2940 (NH + ), 1740–1720 (C = O of ester and lactone), 1610–1580 (C = O, COO – ), 1440, 1380, 1260, 1210 1 H NMR (D2O, d): 0.92 (3H, s, 18-H3), 2.92 (1H, m, 17a-H), 3.69–3.81 {(36H, m, 2 O2C-CH2N-CH2-CO2– ) -CH2-CH2N(-CH2CO2– )-CH2-CH2N(-CH2CO2– )2}, 5.02 (2H, s, 21-H2), 6.02 (1H, s, 22-H) Quantitative analysis calculated for C51H76N6O26 : C, 51.48, H, 6.45, N, 7.06. Found: C, 51.22, H, 6.89, N, 6.77

Radiolabelling

Ligand 2 or 5 was dissolved in an aqueous solution of 99m TcO4– containing 50% ethanol (0.2 ml, 370–740 MBq), whereas ligand 3 or 6 was dissolved in aqueous solution of 99m TcO4– (0.1 ml, 370–555 MBq). Stannous chloride solution (25 ml), prepared by dissolving stannous chloride dihydrate (10 mg) in 6 N HCl (50 ml) and subsequent dilution to the desired volume (10 ml) with nitrogenpurged water, was added to each of the above ligand solutions. The reaction mixture was allowed to stand at room temperature for 30 min prior to use. Radiochemical analysis Thin-layer chromatography

The radiochemical purity of 99mTc chelates of ligands 2, 3, 5 and 6 was determined by a TLC method using pre-coated silica gel strips (10  2.5 cm) as the stationary phase with acetone (system A) and 5% aqueous methanol (system B) as the two developing solvents. The RF values of the 99mTc chelates, 99mTcO4– and 99mTcO2 are, respectively, 0, 1 and 0 in system A; and, respectively, 1, 1 and 0 in system B. A quantitative analysis of the chromatograms was performed by cutting the strips at half the solvent migration distance and counting the radioactivity on each part. High-performance liquid chromatography

Technetium complexes of ligands 2–6 were analysed using a reverse-phase HPLC system with a m-bondapak C18 column (0.39  30 cm). After injecting the pure complex (5–25 ml) the column was eluted isocratically with phosphate buffer (0.01 M, pH 2.4) and acetonitrile in different ratios (1:9, 3:7, 5:5 and 7:3) at the rate of

99m

Tc-ouabagenin compounds Debnath et al. 275

1 ml  min – 1. Results are shown in Table 1. Both analogue and digital data were collected to determine the number, retention time and relative amount of each contributing peak. The chelate solution was left stoppered at room temperature and again analysed by HPLC using the same buffer as above in 5:5 ratio. Biodistribution studies

Biodistribution studies of the chelates were performed on rats (200–300 g), guinea-pigs (400–500 g) and rabbits (1000–1200 g). Animals were anaesthetized with urethane (25%, 0.6 ml  kg – 1 body weight) and the radioactive solution (10 ml, 2960–3700 kBq  kg – 1 body weight) was injected via a previously cannulated femoral vein. At pre-set time interval the animals were killed by intravenous administration of air (2–5 ml). Blood and urine samples were obtained by the puncture of heart and urinary bladder, respectively. Other organs were removed, rinsed with saline and blotted dry to remove residual blood. The results were expressed as percent dose per gram of tissue or percent dose per organ by counting the samples in a gamma counter against suitably diluted aliquots of the injected solution as standard. All animal experiments were carried out in compliance with the relevant national laws relating to the conduct of animal experimentation.

Results Condensation of ouabagenin monoacetonide, 1, with a 16-fold molar excess of the acetylating agent (NTA + Ac2O) for 48 h at 401C gave the desired conjugate, 2, along with a large amount of NTA. Removal of NTA from the product mixture was achieved first by silica gel chromatography, when the desired product, 2, was obtained in 79% purity, mixed with NTA (21%). Preparative HPLC on an RP-18 column furnished the pure conjugate 2 (98%) in 26% yield. The product was further characterized by HPLC, spectroscopic and analytical methods. Deacetonylation of 2 with sulfuric acid in ethanol furnished 3 in 74% yield. In an attempt to develop the 99m Table 1 Retention times (min) of Tc chelates and 99mTcO4– on a reverse-phase C18 HPLC column with different eluting systems 99m

Ratio of phosphate buffer* to acetonitrile 99m

1:9 3:7 5:5 7:3

Tc-2

2.75 3.33 3.86 4.10

99m

Tc chelate

99m

Tc-3

2.44 3.24 3.41 4.00

99m

Tc-5

2.64 3.52 3.77 4.07

99m

TcO4–

Tc-6

2.29 3.17 3.23 3.95

2.10 2.91 3.05 3.59

*

The phosphate buffer was 0.01 M, pH 2.4. 2: Ouabagenin acetonide 3b,11a-bis(iminodiacetate) 3: Ouabagenin 3b,11a-bis(iminodiacetate) 5: Ouabagenin acetonide 3b,11a-bis(diethylenetriaminepentaacetate) 6: Ouabagenin 3b,11a-bis(diethylenetriaminepentaacetate)

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276 Nuclear Medicine Communications 2006, Vol 27 No 3

silica gel chromatographic method for purification, the eluate was esterified with diazomethane to furnish the trimethyl and tetramethyl ester mixture of 2 and NTA, respectively, from which the tetramethyl ester 4 could be isolated easily by silica gel chromatography. Attempts to hydrolyse 4 under mild alkaline conditions resulted in the formation of a mixture from which the desired product 2 could not be isolated. Compound 1, after a similar condensation reaction with DTPA anhydride, furnished 5 which was separated from excess DTPA by silica gel chromatography and reverse-phase preparative HPLC to yield the purified material characterized similarly as in 2(Fig. 2). On deacetonylation, compound 5 gave compound 6 in good yield. 99m

Tc radiolabelling of ligands 2, 3, 5 and 6 was achieved via a stannous chloride procedure in aqueous solution. The radiochemical purities of the above chelates were 92–95% as established by TLC on silica gel plates using two separate solvent systems as developer which can clearly distinguish the chelate from contaminating TcO4– and TcO2. The chelate was subjected to exhaustive HPLC analysis to determine whether more than one radioactive component was present in each of the chelate preparations. For this purpose, phosphate buffer (0.1 M, pH 2.4) and acetonitrile were mixed in different proportions to elute the 99mTc chelate of 2, 3, 5 and 6 along with 99mTcO4– : the latter was used as a reference. Their retention times on the column are recorded in

9.46 (98.8%)

2.48 (1.2%)

2.56 (1.68%)

9.59 (59%)

2.67 (41%)

9.48 (81%)

(b) 2.54 (19%)

(a)

2

5

9.59 (98.32%)

Fig. 2

2

Table 1. Since all the above chelates showed a single component (90–95%) under the above experimental conditions they were considered homogeneous. The chelate solutions was left overnight at room temperature and were again subjected to HPLC analysis. The retention times were the same as previously, indicating their stability. Biodistribution results of the above 99mTc chelates in different animal models are shown in Tables 2, 3, 4 and 5. Blood clearance of all these chelates in all animal models was slow and among the sample data reported, blood showed the highest activity in percent dose per gram at all time points. It also appears that myocardial accumulation of these chelates is greater in guinea-pigs than in rats or rabbits. The heart-to-liver accumulation ratio is also highest in guinea-pigs compared to the other animal models. The above accumulation is highest for 99mTc-2, followed by 99mTc-3, and both have an NTA tether, especially at earlier time points (1.39 and 1.14 at 5 and 15 min) which decreased to a slightly lower value (0.83) at 30 min. However, after 99mTc labelling, compound 2, which was purified by HPLC, showed a better ratio (1.24) at the same time point (30 min). The heart-to-liver accumulation ratio with 99mTc-3 was lower than that obtained with 99mTc-2 starting at 1.0 at 5 min and then having a more or less constant value at 15 and 30 min (0.77), then decreasing further at 60 min (0.55). The corresponding HPLC purified material showed a better uptake ratio than the unpurified material at 30 min (0.91). The DTPA derivatives, 99mTc-5 and 99mTc-6, were inferior to the corresponding NTA derivative in myocardial accumulation and also in myocardial liver accumulation ratio. Therefore the biodistribution experiments were limited to only one time point. All the above radiopharmaceuticals were excreted mainly via the renal route although a small amount of the injected dose cleared via the hepatobiliary pathway. The deacetonylated chelate 99mTc-3 exhibited comparatively higher renal excretion with respect to the acetonide 99m Tc-2. The DTPA derivatives 99mTc-5 and 99mTc-6 showed comparable renal excretion; guinea-pigs were found to be poor renal excretors when compared to the other two animal models in this respect.

Discussion 5

HPLC analysis of ligands 2 and 5: (a) before and (b) after purification by preparative HPLC. The figures on the peaks refer to elution times in minutes with their percentage in the mixture in parenthesis.

Ouabagenin monoacetonide (1) was conjugated with nitrilotriacetic acid (NTA) and diethylenetriaminepentaacetic acid (DTPA) under extremely mild reaction conditions. The resulting ligands, 2 and 5, were deacetonylated to produce two more 99mTc binding ligands, 3 and 6. Radiolabelling of these ligands with 99mTc furnished radiotracers which

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Preparation and evaluation of

Table 2

Biodistribution of

Compound

99m

Tc-ouabagenin compounds Debnath et al. 277

Tc chelates of compounds 2, 3, 5 and 6 expressed in percent dose per gram of tissue* in guinea-pigs

99m

Time after injection (min)

Heart

Blood

Liver

Lung

Spleen

Muscle

Stomach

Mean heart/ liver ratio

99m

Tc-2

5 15 30 30**

0.39 ± 0.06 0.24 ± 0.01 0.15 ± 0.01 0.31 ± 0.05

0.68 ± 0.10 0.37 ± 0.03 0.25 ± 0.02 0.40 ± 0.12

0.28 ± 0.04 0.21 ± 0.05 0.18 ± 0.02 0.25 ± 0.06

0.45 ± 0.07 0.33 ± 0.02 0.21 ± 0.01 0.39 ± 0.06

0.22 ± 0.02 0.16 ± 0.02 0.11 ± 0.01 0.19 ± 0.02

0.13 ± 0.02 0.14 ± 0.01 0.11 ± 0.02 0.19 ± 0.02

0.35 ± 0.05 0.23 ± 0.04 0.16 ± 0.02 0.27 ± 0.06

1.39 1.14 0.83 1.24

99m

Tc-3

5 15 30 30**

0.35 ± 0.06 0.21 ± 0.02 0.24 ± 0.03 0.20 ± 0.06

0.63 ± 0.07 0.48 ± 0.00 0.48 ± 0.03 0.35 ± 0.09

0.34 ± 0.04 0.27 ± 0.01 0.31 ± 0.04 0.22 ± 0.06

0.42 ± 0.06 0.26 ± 0.01 0.35 ± 0.03 0.31 ± 0.08

0.20 ± 0.04 0.14 ± 0.01 0.28 ± 0.05 0.14 ± 0.04

0.13 ± 0.02 0.17 ± 0.01 0.18 ± 0.02 0.10 ± 0.02

0.29 ± 0.04 0.20 ± 0.01 0.62 ± 0.12 0.18 ± 0.05

1.03 0.78 0.77 0.91

99m

Tc-5

30

0.15 ± 0.02

0.26 ± 0.03

0.30 ± 0.01

0.20 ± 0.03

0.10 ± 0.01

0.16 ± 0.03

0.14 ± 0.02

0.5

99m

Tc-6

30

0.16 ± 0.01

0.28 ± 0.01

0.32 ± 0.02

0.19 ± 0.03

0.25 ± 0.06

0.07 ± 0.01

0.18 ± 0.02

0.5

See the footnote to Table 1 and the text for the structures of compounds 2, 3, 5 and 6. Value represents the mean ± SD for eight animals. HPLC purified.

*

**

Table 3

Biodistribution of

Compound

Tc chelates of compounds 2, 3, 5 and 6 expressed in percent dose per organ* in guinea-pigs

99m

Time after injection (min)

Heart

Blood

Liver

Intestine

Urine

Kidney

99m

Tc-2

30 ** 15 30 30**

0.42 ± 0.06 0.33 ± 0.02 0.19 ± 0.01 0.38 ± 0.03

15.05 ± 0.94 11.91 ± 1.70 7.86 ± 1.57 9.97 ± 1.64

2.82 ± 0.29 3.01 ± 0.55 2.14 ± 0.22 2.32 ± 0.34

7.47 ± 1.26 5.09 ± 0.55 3.99 ± 0.88 5.47 ± 1.59

33.93 ± 2.61 21.65 ± 1.03 32.22 ± 1.57 3.93 ± 2.61

5.00 ± 1.04 3.54 ± 0.52 2.68 ± 0.48 3.36 ± 0.28

99m

Tc-3

5 15 30 30**

0.48 ± 0.02 0.26 ± 0.03 0.25 ± 0.04 0.27 ± 0.06

18.18 ± 1.94 12.38 ± 1.0 10.96 ± 1.52 9.45 ± 1.53

4.92 ± 0.70 3.79 ± 0.47 3.55 ± 0.65 2.43 ± 0.44

7.71 ± 0.97 5.54 ± 1.12 5.68 ± 1.86 5.09 ± 0.95

12.38 ± 3.58 23.39 ± 2.37 28.89 ± 2.73 40.60 ± 2.51

5.46 ± 0.56 3.99 ± 0.49 3.46 ± 0.51 3.12 ± 0.27

99m

Tc-5

30

0.18 ± 0.02

7.96 ± 0.50

3.84 ± 0.15

3.46 ± 0.20

40.49 ± 3.89

2.42 ± 0.76

99m

Tc-6

30

0.18 ± 0.01

6.77 ± 0.66

3.94 ± 0.55

4.52 ± 0.41

40.27 ± 0.57

2.47 ± 0.23

See the footnote to Table 1 and the text for the structures of compounds 2, 3, 5 and 6. * Value represents the mean ± SD for eight animals. ** HPLC purified.

Table 4

Biodistribution of

99m

Tc chelates of compounds 2 and 3 in rabbits and rats: percent dose per gram of tissue

Compound

Animal

Time after injection (min)

Heart

Blood

Liver

Lung

Spleen

Muscle

Stomach

99m

Rabbit

15 30 15 30

0.08 ± 0.01 0.07 ± 0.01 0.22 ± 0.03 0.13 ± 0.02

0.19 ± 0.03 0.14 ± 0.02 0.44 ± 0.06 0.32 ± 0.03

0.12 ± 0.01 0.12 ± 0.01 0.43 ± 0.03 0.35 ± 0.01

0.09 ± 0.01 0.09 ± 0.01 0.39 ± 0.04 0.26 ± 0.01

0.09 ± 0.01 0.06 ± 0.02 0.20 ± 0.03 0.15 ± 0.01

0.03 ± 0.01 0.03 ± 0.01 0.19 ± 0.04 0.09 ± 0.01

0.10 ± 0.02 0.07 ± 0.02 0.39 ± 0.08 0.26 ± 0.04

15 30 15 30

0.08 ± 0.01 0.05 ± 0.01 0.20 ± 0.02 0.12 ± 0.02

0.21 ± 0.00 0.11 ± 0.00 0.49 ± 0.04 0.30 ± 0.03

0.14 ± 0.02 0.11 ± 0.01 0.51 ± 0.05 0.38 ± 0.07

0.12 ± 0.01 0.08 ± 0.02 0.32 ± 0.04 0.21 ± 0.03

0.07 ± 0.01 0.06 ± 0.02 0.16 ± 0.03 0.15 ± 0.06

0.04 ± 0.00 0.02 ± 0.00 0.13 ± 0.03 0.06 ± 0.01

0.11 ± 0.02 0.06 ± 0.01 0.49 ± 0.01 0.18 ± 0.02

Tc-2

Rat

99m

Tc-3

Rabbit Rat

See the footnote to Table 1 and the text for the structures of compounds 2 and 3.

could be useful for myocardial Na + ,K + -ATPase imaging because of the well known affinity of the steroidal part of the molecule for the aforementioned enzymes [22]. Analytical and spectroscopic results indicated the presence of two molecules of the tether (either NTA or DTPA) in these conjugates as 3b and 11a monoesters attached to each aglycone moiety. The structure of 99mTc

chelates of ouabagenin–DTPA conjugates can be only speculative because, although 99mTc chelates of DTPA and its various protein conjugates have been shown to be quite useful for several decades, their structures still remain to be elucidated. The structure of the technetium chelate of NTA has been established [23] as Na2[NTATc(IV)(mO)2Tc(IV)NTA]  6H2O where all the carboxylic acid groups are coordinated to technetium. Therefore from this structure it is difficult to propose the structures

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278 Nuclear Medicine Communications 2006, Vol 27 No 3

Table 5

Biodistribution of

99m

Tc chelates of compounds 2 and 3: percent dose per gram of organ

Compound

Animal

Time after injection (min)

Heart

Blood

Liver

Intestine

Urine

Kidney

99m

Rabbit

15 30 15 30

0.19 ± 0.02 0.15 ± 0.04 0.17 ± 0.02 0.09 ± 0.01

14.13 ± 2.94 10.93 ± 1.57 7.88 ± 0.75 5.37 ± 0.92

4.00 ± 0.42 3.36 ± 0.25 3.51 ± 0.50 2.47 ± 0.71

6.95 ± 1.09 6.69 ± 2.08 4.26 ± 0.97 3.96 ± 0.36

32.62 ± 6.90 42.57 ± 1.74 32.71 ± 3.49 38.22 ± 2.04

3.63 ± 1.04 3.00 ± 0.45 3.85 ± 0.53 2.66 ± 0.56

15 30 15 30

0.16 ± 0.02 0.10 ± 0.02 0.18 ± 0.01 0.10 ± 0.00

13.30 ± 1.56 8.71 ± 1.64 8.25 ± 0.85 6.29 ± 1.07

3.75 ± 0.49 2.81 ± 0.58 5.25 ± 0.59 3.56 ± 0.49

8.28 ± 0.99 5.44 ± 1.02 4.26 ± 0.55 4.81 ± 0.49

32.23 ± 1.60 49.65 ± 1.00 28.29 ± 3.38 50.80 ± 2.37

5.65 ± 0.28 3.77 ± 0.36 5.70 ± 0.39 3.41 ± 0.20

Tc-2

Rat 99m

Tc-3

Rabbit Rat

See the footnote to Table 1 and the text for the structures of compounds 2 and 3.

of technetium chelates for conjugates like 2 or 3. In the absence of structural data it is only possible to ensure that the chelate preparations are homogeneous. This was proved by exhaustive HPLC data on these chelates to show that they are essentially pure (90–95%). However, HPLC purified products (100%) showed slightly better biodistribution results in terms of the heart-to-liver accumulation ratio in comparison to unpurified products.

We have used two types of tethers – NTA and DTPA – to conjugate ouabagenin. DTPA is more popular for conjugation with bioactive proteins and such conjugates after radiolabelling with 99mTc are found to be quite useful. However, 99mTc-ouabagenin–DTPA did not furnish the anticipated heart-to-non-target organ ratio. Better results in this respect were obtained with 99m Tc-ouabagenin–NTA conjugates.

Differences in the binding processes of cardiac glycosides and its aglycone with myocardial Na + ,K + -ATPase have been thoroughly investigated [24]. It is observed that irreversible binding with myocardial Na + ,K + -ATPase is only possible with cardiac glycosides (e.g., ouabain) through their sugar residues resulting in rapid disappearance of the agent from blood. The corresponding aglycones (e.g., ouabagenin) also bind with the target enzyme but since the binding assistance from the sugar residue is not available, such binding is reversible and there is appreciable depletion of the substrate from the binding site to blood [25]. This results in a lower heartto-blood ratio for the aglycone in comparison to the parent glycoside. Pharmacologically, the mild and transitory effect of aglycone in comparison to the profound and long lasting therapeutic effect of the cardiac glycoside may be explained on the basis of the above observation.

It is possible that DTPA, with its larger molecular size, changes its molecular nature to such an extent when coupled to ouabagenin, that the expected bioactivity of the aglycone is not observed in such conjugates. NTA on the other hand, because of its smaller molecular size, appears to be a more compatible tether for conjugation to ouabagenin. This is reflected in the biodistribution data when 99mTc-oubagenin–NTA chelates (99mTc-2 and 99m Tc-3) show a better heart-to-liver uptake ratio in comparison with the corresponding DTPA derivatives (99mTc-5 and 99mTc-6).

The above hypothesis offers a satisfactory explanation for the low heart–blood ratio that is observed for the 99mTc chelates described here, for the 99mTc-ouabagenin– cysteine conjugate [17] and also for the radioiodinated digoxigenin derivative [18] discussed earlier. It also indicates that a more favourable biodistribution result could be achieved by attaching a sugar residue to the steroidal moiety in the 3b position. This is already confirmed by the observation that the heart-to-blood ratio of the radioiodinated digoxigenin derivative has been greatly improved [18] while using the corresponding glycosides in biodistribution experiments. Therefore it is anticipated that the unfavourable heart-to-blood ratio which is observed with the radiolabelled ouabagenin is expected to improve appreciably in ouabain derivatives, if it is possible to functionalize and radiolabel the glycoside in accordance with this pilot experiment.

So it can be concluded that ouabagenin and its acetonide can be conjugated with either NTA or DTPA and both the resulting conjugates form a stable chelate with 99mTc. The 99mTc chelate of the NTA conjugate showed biodistribution results that were more promising compared with those of the DTPA conjugate. These and other pilot studies reported from this laboratory indicate that it is possible to functionalize and radiolabel ouabain in a similar manner to furnish a useful agent for myocardial Na + ,K + -ATPase imaging.

Acknowledgement The authors thank Ms Archana Patra for able technical assistance.

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Abstracts

Abstracts of the 34th Annual Meeting of the British Nuclear Medicine Society Manchester International Conference Centre, UK, 27–29 March 2006

CARDIOLOGY I A1 Treadmill exercise and myocardial perfusion imaging: the relationship between target rate and ischaemia A.H. Elmegadmi, C. Randall and N.W. Garvie Royal London Hospital, UK. Aim Treadmill exercise (ETT), which uses ST changes to predict ischaemia, is judged to be unreliable if patients fail to achieve their age-predicted target rate (THR), but nevertheless remains the best physiological stress technique for use with myocardial perfusion imaging (MPI). How important, however, is the THR in the induction of ischaemia, as assessed by MPI? Methodology One hundred and one patients (35 female) referred with suspicion of ischaemia, were stressed by ETT prior to MPI. Beta blockers and nitrates were discontinued beforehand. Thirty failed to attain their THR due to fatigue, breathlessness or other causes. MPI was performed using a 2-day protocol, and images were subjected to attenuation correction. Results Twenty-four patients had ischaemia on MPI. Of these, 11/30 (37%) had a sub-maximal ETT, compared to 13/81 (16%) who achieved or surpassed their THR. ST depression developed in 17 patients, of whom 5 had ischaemia on MPI. Conclusion Surprisingly, on a proportionate basis, ischaemia was twice as common in patients who had performed a sub-maximal ETT, compared to those who fulfilled the criteria for adequate performance, who mostly (84%) had normal subsequent MPI studies. It must be assumed that patients with ischaemia are often unable to stress to completion due to a compromised coronary circulation. The ETT should not be discontinued prior to radiopharmaceutical administration if patients fail to attain their target rate.

Results From planar images, observers identified only 55% of patients not requiring a repeat study and 48% of patients who required a repeat study. There was no significant difference in the results for each of the 5 angles. Comparison of equivalent images at the optimum angle derived from the cine correctly identified whether repeat images would still have overlapping bowel. Conclusion A planar view cannot predict bowel overlap. However, it can show if the patient has waited for sufficient time before the repeat study.

A3 Prognostic significance of ST depression using adenosine stress in the setting of a normal myocardial perfusion study M.M. Pandit, M.C. Prescott, R.S. Lawson, J.M. James and P. Arumugam Department of Nuclear Medicine, Manchester Royal Infirmary, UK. Background Patients who have a normal exercise myocardial perfusion scan are generally thought to have a low future risk of cardiac events (non-fatal MI or cardiac death). Two ultrasoundbased studies suggested that this may not be true if ST depression is associated with vasodilator stress even in the context of normal perfusion [1,2]. Aim We wanted to investigate the prognostic implications of ST depression with vasodilator stress in the presence of a normal MPI within our population group. Methods and Results 1107 patients underwent vasodilator stress for MPI in the year 2001, out of which 33 (0.029%) have been identified as having ST depression with normal perfusion. Case notes of these 33 patients were retrospectively reviewed for cardiac events or revascularization for a mean follow up period of 26 months. Two patients had cardiac events (1 non-fatal MI 25 months later and 1 revascularization at 13 months). The annual rate of adverse cardiac events was 2.8%. Conclusions In our study the event rate was less than that quoted by Klodas [1] and Abbott [2]. It remains higher, however, than the accepted annual event rate of < 1% with a normal stress perfusion study.

A2 Can a planar image predict bowel overlap in tomographic myocardial perfusion scans? S. Russell, W.H. Thomson, J. O’Brien, N.B. Smith and A. Notghi City Hospital, Birmingham, UK.

References 1 Klodas E, et al. Nucl Cardiol 2003; 10:4–8. 2 Abbott BG, et al. J Nucl Cardiol 2003; 10:9–16.

Aim Bowel activity can mask myocardial uptake in tomographic myocardial studies. This is only evident after tomographic reconstruction. Normally the patient waits for a repeat scan. Can a single planar view predict this? Method Tomographic studies of 6 patients who required a repeat and 6 patients who did not were examined. Equivalent planar views at 5 angles around the heart were extracted. Each planar view was filtered with the tomographic filter. Randomized images were assessed by two observers for bowel overlap. A further analysis examined 6 patients with repeat studies. Acquisition data were filtered then viewed in cine mode for the optimum planar angle of the overlap activity. This was compared to the equivalent view in the repeat acquisition.

A4 Pacing and racing the heart: enhancing heart rate in asthmatic patients with cardiac pacemakers G. Gnanasegaran, G. Ross, G. Shabo, A. Taylor, A. Morley and T.O. Nunan Department of Nuclear Medicine, Guy’s and St. Thomas Hospital NHS Trust, London, UK. Objective To assess the role of external pacing myocardial perfusion scintigraphy (MPS) in asthmatic patients with pacemakers. Ideally adenosine should be used as the stressor since there may be aberrant conduction. Stressing asthmatic patients with pacemakers is problematic as vasodilators are contraindicated.

c 2006 Lippincott Williams & Wilkins 0143-3636


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282 Nuclear Medicine Communications 2006, Vol 27 No 3

Methods Six patients with pacemaker and suspected CAD (4 females and 2 males, age range 55–80 years) were stressed through their pacemakers. An electrophysiology technician (trained in handling pacemakers) performs the procedure. The technique involves increasing the heart rate gradually through the patient’s pacemaker using a magnet and the radiopharmaceutical injected when the patient reaches 85% of the expected target heart rate (THR). The heart rate is maintained for at least 2 min post-injection before bringing them to the baseline. The images are acquired 1 h postinjection. No adverse effects were noted. The procedure took 5–7 min. Results Five out of 6 patients had abnormal scans. Four patients had reversible ischaemia (two had widespread ischaemia involving multiple territories) and 1 patient had an equivocal scan (reversible ischaemia/artefact). One patient had a hypotensive response. Conclusion External pacing to achieve the required THR in asthmatic patients with pacemaker is a useful technique. However, liaison with the referring physician and the cardiology team is important.

Methods Case records of 40 patients who underwent MIBI imaging prior to parathyroidectomy between 1997 and 2004 were reviewed. Results At operation 33 had uniglandular and 4 multiglandular disease. One was a carcinoma, 1 multi-endocrine neoplasia and 1 gland was normal. Twenty-three of 40 (50%) had had previous neck exploration. Twenty-five scans were reported as positive, 8 equivocal and 7 negative. The side of the lesion was correct in all positive reports with the quadrant accurately predicted in 100% of single and 80% of the multiple glands. Gland size and weight were significantly larger amongst the MIBI positive group (P = 0.04, Mann–Whitney U). All the equivocal sites corresponded to true abnormal glands at surgery. Only 1 of 7 ectopic glands and none of the intrathyroidal adenomas were detected. In only one of the multiglandular cases did the scan reveal the second gland. Conclusion A positive MIBI scan was still an excellent predictor of lesion site. The surgeons were never misdirected to the wrong side. However, the sensitivity for detection of ectopic, intrathyroidal and multiglandular disease was low.

A5 Chest pain during myocardial stress procedures. Does it imply the presence of ischaemia? A.H. Elmegadmi, Y. Pantic, C. Randall and N.W. Garvie Royal London Hospital, UK.

A7 Role of 99mTc-sestamibi scintigraphy and ultrasound in preoperative localization of parathyroid lesions C.M. Lee, M. Norris, R. Bliss and J.J. Lloyd Royal Victoria Infirmary, Newcastle Upon Tyne, UK.

Purpose Chest pain (CP) is not infrequently described by patients undergoing stress procedures prior to myocardial perfusion imaging (MPI). To what extent is this predictive of ischaemia, as evidenced on the subsequent images, and does this vary for particular stress techniques? Methodology One hundred and forty-four patients who developed CP during stress were evaluated for ischaemia on the basis of subsequent MPI, which was performed using a 2 day tetrofosmin protocol. Patients were allocated according to suitability to a particular standard stress protocol (adenosine, treadmill exercise or dobutamine). Nitrate preparations were discontinued for 2 days beforehand. Results Thirty-one of 144 patients with CP were found to have ischaemia on MPI (21%). Of the individual stress procedures, 17/90 (19%) patients undergoing stress by adenosine infusion were found to have ischaemia. The corresponding figures for those undergoing dobutamine stress, and exercise treadmill stress, were 5/26 (19%) and 9/28 (30%), respectively. The overall positive predictive value of CP for ischaemia was 20%. Conclusions Chest pain during pharmacological myocardial perfusion stress procedures is poorly predictive of ischaemia, and patients and supervising staff can be accordingly reassured. The data suggest, however, that CP developing during treadmill exercise is more linked to ischaemia.

NEUROENDOCRINE I

A6 Usefulness of pre-operative MIBI scanning in patients with primary hyperparathyroidism N.V. Hamilton, G.J. Ellul, G.I. Al-Bahrani, J.M. James, M.C. Prescott and N.R. Parrott Department of Nuclear Medicine, Manchester Royal Infirmary, UK. Aim To assess the accuracy of dual phase MIBI scans in hyperparathyroidism following a change from the Tl–Tc subtraction technique.

Background Scintigraphy and ultrasound are standard modalities for preoperative localization in the management of primary hyperparathyroidism. With an increasing trend towards minimally invasive parathyroidectomy, preoperative localization is becoming particularly important. We introduced localization using 99mTc-sestimibi (MIBI) washout and subtraction technique in 2003. Methods We reviewed the accuracy of the MIBI scan (both washout and subtraction techniques) and ultrasound in 62 patients with primary hyperparathyroidism by reference to findings at surgery and histology. Results Fifty-eight (94%) patients had curative surgery and were included in our analysis. Forty-five (78%) had parathyroid adenoma, 12 (20%) had parathyroid hyperplasia and 1 (2%) had parathyroid carcinoma. The sensitivity and positive predictive value (PPV) for MIBI were 52% and 63%, respectively; the corresponding values for ultrasound were 39% and 71%. However, localization was correct in 20 of the 22 cases where MIBI and ultrasound agreed ( i.e., 91% PPV for localization by both scans). Conclusions MIBI and ultrasound both had low sensitivity; reasons for this will be discussed at the presentation. However, concordant preoperative MIBI and ultrasound findings represents a reliable localization technique (PPV 91%) when used together and is especially useful for directing minimally invasive parathyroidectomy and sparing the patient from bilateral neck exploration. A8 Octreoscan in the UK: a BNMS survey G. Gnanasegaran and S.E.M. Clarke Department of Nuclear Medicine, Guy’s and St. Thomas’ Hospital NHS Trust, London, UK. Background Octreoscan is commonly used in the diagnosis of neuroendrocrine tumours (NETs). The common indications are for the detection, localization, assessment of receptor status, and selection of patients for treatment/follow-up. This study

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34th BNMS meeting: abstracts 283

surveyed the use and imaging protocols in the UK over the last few years. Methods A questionnaire was sent to 241 centres performing radionuclide imaging asking for details of Octreoscan imaging (the key question related to indications, protocols, acquisitions etc). Results Replies were received from 183 centres (76%) (44% by telephone). Of the responding centres, 76 (41%) performed Octreoscan. The main indication was for NETs (GEP/carcinoids). The injected activity ranged from 100 to 270 MBq; 85% used a dual-head gamma camera. Whole-body imaging was acquired in all patients. Seventy-one percent used SPECT regularly and 22% used it occasionally. Twenty-six percent stopped somatostatin analogues and 47% prescribed laxatives before imaging. The image analysis and reporting was by visual analysis in all centres (100%), 3 centres use additional CT registration. Conclusion The main indication for using Octreoscan is for GEP/ carcinoids tumours. The majority of departments follow the standard protocol for imaging and reporting. However, many were not consistent with correct scheduling of imaging and patient preparation. The results confirm that the role of Octreoscan in imaging neuroendocrine tumours is appreciated. These findings indicate that a national guideline to assist departments in developing protocols should be established. A9 Octreotide and MIBG scans in patients with neuroendocrine tumours: correlation with survival and tumour grade A.-M. Quigley, J.R. Buscombe, M.E. Caplin and A.J.W. Hilson Royal Free Hospital, London, UK. Aim To correlate octreotide and MIBG uptake with patient survival and tumour grade in neuroendocrine patients. Method One hundred and forty-nine patients underwent octreotide and MIBG scans (on referral) over a 4 year period, and classified into 1 of 7 groups: both studies negative, both studies positive with identical lesions, different lesions, more lesions on octreotide, positive octreotide with negative MIBG, more lesions on MIBG, positive MIBG with negative octreotide. Survival time or time to death from referral and histological grade were obtained from patient records. Kaplan–Meier survival analysis was performed for each of the 7 groups and also according to grade (Ki-67 score). Results Data were available for 147 patients. No statistical difference was observed for each of the 7 groups (mean survival 31.7–44.7 months). However, if patients’ scans were concordant (both negative or identical lesions) survival was significantly poorer than if scans were discordant (P = 0.028). Higher grade tumour was also associated with significantly poorer survival (P = 0.001). Conclusion As expected, higher grade tumour is associated with poorer survival. The relationship between scan appearance and survival is less clear, but agreement between octreotide and MIBG appearances (both scans negative or identical lesions) was observed to be associated with poorer survival.

A10 An audit to assess the role and importance of nuclear medicine imaging in the management of neuroendocrine tumours V. Ramana, S.V. Biswasb, G. Gnanasegarana, N. Mulhollanda and S.E.M. Clarkea Departments of aNuclear Medicine and bRadiology, Guy’s and St Thomas’ NHS Trust, London, UK. Purpose To review the role of nuclear medicine (NM) imaging in the management of neuroendocrine tumours (NETs).

Method NETs are a heterogeneous entity and share certain biological features. They arise from endocrine and neuroendocrine glands and from endocrine cells found within the gastrointestinal and respiratory tract. Their diagnosis and management is multi-modality and complex. We reviewed all the NM studies, which include 99mTc(V)-DMSA, 131I-MIBG and 111In-pentreotide scintigraphy, performed between 2003 and 2005, the histopathological diagnosis where available, and any radiological imaging. This was performed using data available on electronic patient records and referral letters. Results A total of 104 patients received NM imaging and 177 investigations performed (19%: 99mTc(V)-DMSA; 39%: 131 I-MIBG; and 42%: 111In-pentreotide). These were categorized into those for diagnostic, staging, surveillance and therapeutic purposes; the proportion of scans devoted for the latter were 34%, 11%, 51% and 4%, respectively. Thirty-eight histological diagnoses were documented and the results of the NM investigations correlated well with the above with a disparity seen in only 6 cases. Conclusion The management of NETs is complex and multimodal and, as outlined in the recently published guidelines, integrally involves NM imaging in a variety of capacities to ensure that there is a good correlation with the above and the tissue diagnosis. References 1 Guidelines for the management of gastroenteropancreatic neuroendocrine (including carcinoid) tumours. Gut 2005; 54(suppl 4):l–16. 2 Oberg K. Ann Oncol 2004; 15(suppl 4):293–298. 3 Sherman SI. Lancet 2003; 361:501–511. 4 Lenders JWM, et al. Lancet 2005; 366:665–675. 5 Gregory AK, et al. Endocrine Rev 25:458–511.

PHYSICS A11 Comparison of different methodologies for lesion volume determination in PET M. Hatta, N. Boussiona, K. Carsonb, P. Jarrittb, F. Lamarea, Y. Bizaisa, C. Cheze le Resta and D. Visvikisa a U650 INSERM, Laboratoire du traitement de l’information me´dicale (LaTIM), Brest, France; and bNorthern Ireland Regional Medical Physics Agency, Belfast, UK. Current methodologies for functional volume definition in emission tomography involve the use of a priori CT knowledge or are based on empirical threshold based techniques. The purpose of our study was to compare these methodologies with a new automatic algorithm accounting for noise present in PET images. Images of the IEC phantom containing spheres of variable diameter were simulated, considering different source-to-background (S/B) ratios and statistical noise levels. For thresholding purposes, 50% and 75% of the ROI maximum value as well as 3 times the standard deviation computed on a background ROI were used. A recently developed automatic methodology based on a hidden Markov chain model combined with a fuzzy measure (FHMC) was also tested. The threshold-based techniques were sensitive to the level of noise in the image and were found to perform better with high S/B ratios and for lesion sizes > 2 cm in diameter. The FHMC algorithm, however, was less sensitive to noise levels and performed better throughout the different lesion sizes considered. The performance of empirical threshold-based

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methods for lesion delineation greatly depends on image characteristics. A newly developed automatic algorithm including noise modelling greatly improved the accuracy of volume determination.

A12 Improved triple energy window scatter correction for 131I E. Mora-Ramireza, L. Barndenb and B.F. Huttona a Institute of Nuclear Medicine, UCL, UK; and bQueen Elizabeth Hospital, Adelaide, Australia. An improved method for triple energy window (TEW) scatter correction is proposed for use with 131I, which accounts for the influence of down-scatter from high-energy emissions. Theoretical considerations suggest the use of different weights for the 2 energy windows used for scatter estimation. GATE Monte Carlo (MC) simulation was carried out for 4 source depths in a water phantom for a gamma camera with a highenergy collimator (septal thickness 1.73 mm). Energy spectra and images were generated for a range of energy window widths and positions and used to derive the optimal window settings and choice of weighting factors. Results were compared with conventional TEW. Optimal results were obtained using a 6% window at 318 keV (to avoid the 284 keV photopeak) and a 15% window centred at 475 keV. This higher than expected window avoids the Compton edges for higher emission energies whose spatial distribution does not match the down-scatter in the photopeak. The weights used to correct for scatter in the 20% photopeak window were approximately 2.0 and 0.2, respectively. The modified approach reduced the scatter error to < 2% compared to > 15% for conventional TEW. The proposed method offers potential to improve the quantitative accuracy of 131I SPECT.

A14 The effectiveness of different types of syringe shields for 90Y S.A. Hoopera, W.H. Thomsonb, C.A. Crossleyc, J.H. Pearced and J.M. Jonesd Departments of aNuclear Medicine, Velindre NHS Trust, Cardiff, b Physics and Nuclear Medicine, Sandwell & West Birmingham Hospitals NHS Trust, cRadiation Protection Service, Velindre NHS Trust, Cardiff, and dMedical Physics, University Hospital of Wales, Cardiff, UK. Background Newer therapies involving beta emitting radionuclides require manipulation of high activities e.g. 2500 MBq 90 Y may be handled. Traditionally, Perspex is recommended for shielding because this is thought to reduce Bremsstrahlung. Tungsten is also effective, but heavier. A hybrid syringe shield design for 90Y-Zevalin labelling uses a combination of lead and plastic. Methods Dose rate measurements at 50 cm were obtained for these shield types. The measurements were correlated with Bremsstrahlung spectra measured with a germanium detector. Thermoluminescence dosimetry (TLD) measurements on the shield surface were also carried out. Results For 10 ml syringes, the dose rate at 50 cm for Perspex was more than twice that for tungsten or the Zevalin shield. These results were consistent with the emission spectra and TLD values. The Zevalin shield gave the lowest figure for TLD surface dose (0.33 mSv  GBq – 1  h – 1: cf. tungsten –1 –1 1.7 mSv  GBq  h , Perspex 3.1 mSv  GBq – 1  h – 1 and unshielded 7068 mSv  GBq – 1  h – 1). Conclusions The 10 ml Perspex shield is the least effective in reducing exposure to staff based on TLD, dose rate reading and spectral analysis. Tungsten is effective because it reduces low energy Bremsstrahlung. The Zevalin shield provides the best protection due to incorporated lead and to the wall thickness. However, our results indicate improvements to the Zevalin shield design that we are investigating.

A13 What is an achievable and acceptable level of head misalignment along the axis of rotation direction for gamma camera SPECT? M.P. Avisona, M.C. Barnfieldb, M.T. Burnistonb and S. Brownc Departments of aMedical Physics, Bradford Royal Infirmary, bMedical Physics, St James’s University Hospital, Leeds, and cNuclear Medicine, Southend Hospital NHS Trust, UK.

A15 Radiation protection implications in 68 Ga-DOTATOC PET imaging R. Patel, D. Berehoudougou, M.L. Rahman, D.J. Towey, A. Al-Nahhas, Z. Win and K.S. Nijran Hammersmith Hospitals NHS Trust, London, UK.

We present results demonstrating that several models of modern dual-head gamma cameras fail the performance guideline for head misalignment of less than 2 mm (along the axis of rotation or Z direction) issued in IPEM Report 86. The average head misalignment was 5 mm from which we concluded that either the guideline is too rigorous or that some cameras offer inferior performance. We designed and constructed a phantom that enabled us to assess the effect of head alignment on reconstructed SPECT resolution. Line spread measurements were performed in all 3 dimensions at many positions in the field of view and results confirmed that Z axis misalignment has little effect on X or Y resolution but affects resolution in the Z direction more strongly. Full range head misalignment in the Z direction of 6.6 mm led to a degradation of resolution (FWHM in mm) from 9 mm (SD 0.3 mm) to 11.8 mm (SD 0.3 mm) in the absence of scatter and at a radius of 14 cm. In clinical scanning conditions these differences would not be clinically significant and we conclude that a maximum Z misalignment of 6.5 mm would be permissible without compromising scanning quality. Our phantom has a range of additional uses in evaluating SPECT acquisition and reconstruction performance, examples will be presented.

Aim This study investigates the practical radiation protection issues encountered in 68Ga-DOTATOC PET imaging, including personal and environmental dose rate measurements. Introduction A new imaging technique using 68Ga-DOTATOC for whole-body PET imaging was introduced for patients with neuroendocrine tumours. Thus far, 10 patients have been imaged using the new technique. The high beta (Emax 1.9 MeV) and gamma (1.08 MeV) emission from 68Ga introduces the need to assess the radiation protection implications. Method Patients received 80–150 MBq 68Ga-DOTATOC. Imaging was carried out 30 min post-injection using a dedicated Siemens ART PET scanner. Staff finger doses were measured using thermoluminescence dosimetry (TLD) badges during preparation and injection. Environmental doses were monitored using optically stimulated luminescence (OSL) badges positioned at the operator console. Dose rates near the patient were measured 15 min post-injection at several positions, 30 cm from the head, waist and feet, using an energy-compensated Geiger–Muller tube. Results: Staff finger doses for dispensing and injection were found to be below 0.4 mSv. Environmental dose rate

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measurements were below 0.04 mSv. The maximum dose rate measured near the patient was 15 mSv  h – 1.

QUANTITATIVE METHODS A16 Using whole-body counting to determine neutrophil trafficking and loss in COPD K.R. Szczepuraa, P.R. Rupareliab, D. Biltonb, E.R. Chilversb and A.M. Petersa Departments of aRadiology, University of Cambridge, and bMedicine, University of Cambridge, Addenbrooke’s and Papworth Hospitals, UK. COPD is an inflammatory condition of the airways characterized by airway neutrophilia. Using a whole-body-counter (WBC) we have assessed neutrophil trafficking and loss in COPD using small activities of 111In-labelled neutrophils. Patients were scanned in the WBC 45 min, 1, 2, 4, 7 and 10 days after injection of autologous 111In-labelled ‘pure’ neutrophils and whole-body 111 In retention recorded. Using a single-slit collimator, biodistribution was also recorded from the posterior projection as a count profile from head to toe. SPECT on another occasion with 99m Tc neutrophils was used to validate bio-distribution. Mean retention at 10 days was: controls 89.3% (SEM 0.78) (n = 12), COPD 88.9% (1.00) (n = 10) and bronchiectasis 85% (2.69) (n = 10). Profile data showed that the neutrophils initially localized to the liver/spleen region in all patients and thereafter redistributed to the pelvic and thoracic areas. SPECT (n = 3) demonstrated that 19.6% of the thoracic uptake was in the lungs with the remaining signal in the rib and vertebral bone marrow. Whilst it is known that 111In-labelled neutrophils migrate to the lungs in COPD, mobilization into the airways and subsequent loss from the body is insufficient to quantify by WBC. This technique does provide accurate monitoring of neutrophil trafficking to the liver/spleen and BM compartments, however.

A17 Modelling and correcting for respiratory motion in PET N.S. Vyasa, P. Schleyera, D.L.G. Hillb and P.K. Marsdena a King’s College London School of Medicine at Guy’s, King’s College and St Thomas’ Hospitals; and bCentre for Medical Image Computing, University College London, UK. We have developed a complete simulation of the PET data acquisition and reconstruction process based on the NCAT/SIMSET software, in order to assess the effect of respiratory motion and motion correction techniques on PET data. Regular arrays comprising lesions of different sizes and contrast were incorporated into the NCAT torso phantom and data were reconstructed using filtered backprojection at 16 phases of the respiratory cycle. Affine registrations were performed on the 16 images to create a single summed motion corrected PET frame. The transformations were derived from (i) the PET images themselves and (ii) simulated CT images at the same respiratory phases. Results indicate that respiratory motion both blurs lesions and decreases the accuracy of quantification of PET images. These effects varied with lesion size and SUV. The volume of lesions is seen to increase and the uptake values are seen to decrease in lesions undergoing motion. The smaller and lower contrast lesions show more differences between static, time averaged and motion corrected frames. Preliminary comparisons between corrected and static frames show that

the CT corrected frame is closer to the static frame than the PET corrected one.

A18 Simple scatter correction methods for gamma camera images: Do they work? D.A. Wright and S.T. Chandler Regional Medical Physics Department, Darlington Memorial Hospital, UK. Four simple methods of scatter correction for gamma camera images: Compton window (CW), split window (SW), a reduced width window (15% photopeak window), and a Triple energy window (TEW) were assessed and compared. Point source images of 99mTc were assessed by the FWTM and FWHM of the image profiles. A Williams phantom was also imaged, for which ROIs were used to calculate contrast and signal to noise ratio (S/N) values. Two observers evaluated the visibility of lesions in randomized images of the phantom. All techniques reduced the FWTM value apart from the 15% photopeak method. Contrast improvements were observed for the SW method and less so for the TEW method, particularly at low counts. Both suffered from a decreased S/N. No improvement in lesion detection was observed for any scatter correction technique due to the reduced S/N. Applying a 9-point smooth to the scatter image before subtraction from the primary image marginally improved the visualization of the lesions over the unsmoothed method. These methods of scatter correction were not found to improve lesion detection but the SW method is believed to be the most promising. Further work to investigate ways of reducing the noise would be justified.

A19 Computer simulation of gamma camera images of the kidney A.H. Dawson, J.S. Fleming, S.M.A. Hoffmann, L. Papaspyrou and S. Peel Department of Medical Physics and Bio-engineering, Southampton General Hospital, UK. Background Since the true activity distribution within a patient is never known, the accuracy of measurements from gamma camera images cannot be assessed. If convincing simulated images can be created, then measurements can be compared to the distribution defined for the simulation. Methods Anatomical models were created from CT scans of a patient who had also undergone DMSA imaging. The renal cortex and medulla were modelled separately, because literature suggests that the ratio of DMSA uptake in the cortex and medulla is 22:1. An iterative approach was used to define the activity distribution for the simulation, by comparing the simulation with the patient DMSA scan. Results The real and simulated kidneys had visually similar activity distributions. Analysis of the real and simulated DMSA scans gave values for the relative right kidney function of 85% and 84%, respectively. The optimal simulation had a cortex to medulla activity ratio close to 1:1, suggesting the uptake of DMSA in the cortex and medulla was approximately equal, rather than the majority of uptake being within the cortex. Conclusion A technique for the realistic simulation of static renal images has been devised and validated. This should be useful in the evaluation of image analysis software.

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ONCOLOGY I A20 Phase I/II study of fractionated radioimmunotherapy in relapsed low grade nonHodgkin’s lymphoma M.A. Zivanovica,b, M.C. Baynea, T.M. Illidgea,b Y. Dua,c, V.J. Lewingtona,d and P.W. Johnsona a Southampton University Hospitals NHS Trust, bChristie Hospital NHS Trust, Manchester, cMiddlesex Hospital, London, and dRoyal Marsden Hospital NHS Trust, Sutton, UK. Radiolabelled rituximab is a promising new treatment for ‘low grade’ non-Hodgkin’s lymphoma (NHL). Multiple or fractionated infusions may improve efficacy and are possible because immune reactions are rare, as rituximab is a chimeric antibody. A dose escalation protocol was used to assess the safety and efficacy of fractionated radioimmunotherapy (RIT) in relapsed CD20 positive NHL using two fractions of 131I-labelled rituximab preceded by unlabelled induction courses. All 16 patients in the first 4 cohorts responded with acceptable haematological toxicity. Responses and duration of response was significantly better than that seen following their last combination chemotherapy regime. Nine patients (56%) had a complete response, the median duration of which is 18 months. Eight of these are ongoing. Sequential analyses have shown wide variations in the serum concentration of unlabelled rituximab and the effective half-life of 131I-rituximab between patients and within the same patient as treatment progresses. These results suggest that both the induction rituximab and the 131I-rituximab dose and scheduling need individual tailoring in order to improve biodistribution and optimize therapeutic ratio. Fractionated RIT is safe and efficacious and enables larger whole-body doses to be administered safely than has been achieved previously with nonmyeloablative RIT. A21 The microscopic tumour biodistribution of 131 I-labelled mAb is critical in predicting successful radioimmunotherapy of lymphoma Y. Dua,b, M.J. Glennieb, P.W. Johnsonb and T.M. Illidgeb,c a Institute of Nuclear Medicine, University College London, and Cancer Sciences Divisions, bUniversity of Southampton, and cUniversity of Manchester, UK. Purpose The importance of radiation dosimetry in radioimmunotherapy (RIT) is currently controversial [1,2]. We have investigated tumour dosimetry of radioimmunoconjugates at a cellular level and correlated this with the therapeutic effects in a BCL1 lymphoma tumour model. Methods Groups of tumour inoculated mice were treated with 9.25 or 18.5 MBq 131I-labelled mAbs. Parallel groups of mice were killed for organ dosimetry and biopsies were taken for the determination of the intratumoural biodistribution of mAbs by immunohistochemistry. Results Long-term survival was achieved with 18.5 MBq 131Ianti-MHCII but not with 18.5 MBq 131I-anti-CD45 plus antiidiotype mAb, although conventional organ dosimetry demonstrated that both radioimmunoconjugates delivered similar high doses of radiation to the tumour target organ (the spleen). However, immunohistochemistry revealed marked differences in the microscopic intratumoural biodistribution of these mAbs. The anti-CD45 mAb distributed homogenously within the spleen, whilst the anti-MHCII localized only to the B-cell zones where the tumour cells were located. Thus the different microscopic biodistribution patterns of these mAbs led to

important differences in tumour radiation microdosimetry and therapeutic outcome. Conclusions Radiation microdosimetry appears critical to predicting successful RIT. Conventional dosimetric methods which assume homogenous tumour biodistribution of radiolabelled mAbs can substantially underestimate the radiation dose delivered by RIT. References 1 Britton KE. J Nucl Med 2004; 45:924–925 2 Goldenberg DM, et al. J Nucl Med 2005; 46:383–384

A22 Zevalin: First experience of 18 months of non-research use V.S. Warbeya, A.-M. Quigleya, M.H.L. Nga, R. Hussaina, C. McNamarab, J.R. Buscombea and A.J.W. Hilsona Departments of aNuclear Medicine, and bHaematology, Royal Free Hospital, London, UK. Purpose To review the outcome of patients treated with Zevalin. Method We reviewed the case notes and contacted the referring physicians for information regarding all the patients treated with Zevalin at our institution between December 2003 and July 2005. Results Thirteen patients (6 male, 7 female, age range 42–72 years) referred from 6 different institutions have been treated with Zevalin at our hospital over the past 20 months. Follow-up was available on all 13 patients, 10 are still alive. One patient experienced bronchospasm during intravenous infusion and 1 patient reported nausea and vomiting 4 days post-therapy. Three patients developed severe pancytopenia, 2 requiring blood/platelet transfusions. Six (46%) patients had a complete response to Zevalin, 2 (15%) a partial response, 2 (15%) stable disease and 3 (23%) progressive disease. These latter patients had large volume disease at the time of treatment and have all since died. Five patients remain in complete remission, while the other developed progressive disease at 6 months. One of the partial responders developed progressive disease at 12 months. Two patient’s disease remains stable. One patient is awaiting a decision regarding funding for a second treatment. Conclusion These data are extremely encouraging and in line with previously published response rates.

A23 Radioimmunotherapy in advanced pancreatic cancer: Is it feasible? A. Sultanaa, R. Jayanb, S. Chauhana, S. Shorea, J. Evansc, C. Garveyc, P. Ghaneha, S. Vinjamurib and J.P. Neoptolemosa a Division of Surgery and Oncology, and Departments of bNuclear Medicine and cRadiology, Royal Liverpool University Hospital, UK. Aims To assess the feasibility and safety of radiolabelled antiCEA monoclonal anibody in the treatment of patients with advanced pancreatic cancer. Methods An open, randomized, phase I/II, dose escalation study was devised to evaluate the safety, tolerability, pharmacokinetics, antigenicity and efficacy of 131I-labelled monoclonal antibody when administered either intra-arterially or intravenously in patients with surgically unresectable pancreatic adenocarcinoma. On confirmation of uptake on a diagnostic study, a therapeutic dose of the antibody conjugated with 131I was given 5–7 days later, with imaging for radiotracer localization 1 week following treatment. Results Nineteen patients were enrolled into the study between February 2003 and July 2005. Eighteen (95%) showed

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demonstrable uptake in either the pancreatic cancer site and/or metastases. Nine patients received treatment via the intraarterial route and 9 by the intravenous route (1 patient did not proceed to treatment due to lack of localization of the radiotracer on the dosimetry scan). Post-treatment scans confirmed tumour localization in all patients. Conclusion We have shown the feasibility of using radiolabelled antibodies for the diagnosis and treatment of advanced carcinomas such as pancreatic cancer. Further evaluation of the safety, tolerability and efficacy of this agent is under way. A24 Sodium iodide symporter in metastatic breast carcinoma: Initial experiences with [99mTc]pertechnetate J.E. Prossera, V. MacLarenb, E. Renwicka, M. Boydb, R.J. Mairsb, T.R.J. Evansb, J. McKillopc and G. Gillena. a Department of Nuclear Medicine, Western Infirmary, Glasgow; and b Division of Cancer Science and Molecular Pathology, and cDepartment of Medicine, University of Glasgow, UK. Aim The sodium iodide symporter (NIS) is responsible for concentration of iodide in the thyroid and forms the basis of radioiodine therapy. NIS expression has also been demonstrated in breast tumours and raises the possibility of radionuclide therapy in breast carcinoma [1,2]. [99mTc]pertechnetate behaves similarly to iodide and the aim of this study was to determine if sites of metastatic breast carcinoma could be identified with [99mTc]pertechnetate and to optimize image acquisition. Methods Eighteen patients with metastatic breast carcinoma were split into two groups. Patients in group 1 (n = 9) were administered 600 MBq [99mTc]pertechnetate; patients in group 2 (n = 9) were additionally administered 100–300 mg perchlorate. Images were acquired at 20 min, 4 h and 24 h after injection and were reviewed by a group that was blinded to patient history. Results Physiological uptake of [99mTc]pertechnetate was a problem for patients in group 1. Administration of perchlorate reduced thyroid uptake but bowel activity was still prominent at 24 h. Sites of convincing uptake were seen in 2/9 patients in group 1 and 4/9 patients in group 2. Conclusion Imaging with [99mTc]pertechnetate is feasible in this group but measures must be taken to reduce physiological uptake. References 1 Tazebay UH. Nature Med 2000; 6:871–878. 2 Wapnir I. Clin Can Res 2004; 10:4294–4302. NEUROENDOCRINE II A25 SPECT CT in neuroendocrine tumours R.H. Reid, I. Rachinski and W. Kocha London Health Sciences Centre, Canada. Neuroendocrine tumours, though relatively rare, are a diverse group of pathological entities, including carcinoid tumours, pancreatic neuroendocrine tumours, insulinomas, glucagonomas, VIPomas, medullary thyroid tumours and parathyroid adenomas. All have the ability to produce ectopic hormones and be mulifocal. The accepted clinical investigations for localization are combination of CT, MRI, ultrasound and scanning with 123Ior 131I-MIBG, 111In-Octreoscan and 99mTc-MIBI. The comparison between anatomic imaging and radioisotope imaging is difficult. Image fusion has been tried but is complicated and

requires scanning on two separate modalities. The development of a dedicated single instrument that includes attenuation correction and SPECT CT, has opened up significant opportunity in the assessment of neuroendocrine tumours. These tumours may be small at the boundary of resolution on anatomic imaging but are metabolically active on radioisotope imaging. They may be heterogeneous in the avidity for radioisotope tracers and more precise localization can allow differentiation between lesions that are treatable with single agent or may require combination therapy. More precise localization can be valuable in assessment of patients whose hormone levels are abnormal despite prior surgery. We have performed 120 SPECT CT scans on patients with neuroendocrine pathology. The results in 27 scans changed therapeutic management of patients and reduced operative time required. Six cases are discussed to illustrate the clinical usefulness of SPECT CT.

A26 68Ga-DOTA-Octreotate PET–CT in primary and recurrent neuroendocrine tumours I. Kayani, A.M. Groves, S.M. Gacinovic, P.J. Ell and J. Bomanji Institute of Nuclear Medicine, University College London, UK. Aim To describe our experience with 68Ga-DOTA-Octreotate PET–CT in patients with neuroendocrine tumours. Methods 68Ga-DOTA-Octreotate PET–CT studies from 15 consecutive patients with primary or suspected recurrent metastatic neuroendocrine tumours (8 carcinoid, 2 bronchial carcinoid, 2 pancreatic neuroendocrine tumour, 2 gastrinomas and 1 medullary carcinoid tumour) were reviewed. Patients received 120–200 MBq of 68Ga-DOTA-Octreotate and scans were performed 45 min after injection. Scans were reviewed for normal technical quality and tracer uptake was correlated to sites of primary and metastatic lesions. Results Five of the 15 patients had a primary neuroendocrine tumour, 68Ga-DOTA-Octreotate was true positive in all 5 patients. Eight of 15 patients had proven metastatic disease; in these 8 patients 68Ga-DOTA-Octreotate PET identified metastatic disease in 6. Finally, 3 of 15 patients were proven to be free of recurrent tumour and all 3 patients had negative 68 Ga-DOTA-Octreotate uptake. There were no false positive cases of increased tracer uptake. Conclusion Our initial results suggest that 68Ga-DOTAOctreotate PET–CT is a useful technique for the staging of primary and recurrent neuroendocrine tumours.

A27 18F-FDOPA PET: A superior technique in the detection of phaeochromocytomas? N. Seshadri, J. Wat and K. Balan Department of Nuclear Medicine, Addenbrooke’s Hospital, Cambridge, UK. Aim Accurate localization of phaeochromocytoma is essential for its effective management. Several imaging methods are available to localize this condition but there is emerging evidence on the superiority of 18F-FDOPA PET in the detection of primary and metastatic phaeochromocytoma. The purpose of this investigation was to evaluate the usefulness of 18F-FDOPA PET imaging in patients with phaeochromocytoma where conventional MIBG studies were negative. Materials and methods Five patients, 4 male and 1 female (mean age 47 years) with known or suspected phaeochromocytoma and negative 123I-MIBG scintigraphy underwent PET scanning using 18F-FDOPA. The studies were done within a month of each other.

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Results 18F-FDOPA PET successfully localized the tumor in all five patients. In 1 patient 123I-MIBG showed faint bilateral adrenal uptake but this was not observed on 18F-FDOPA PET. Surgical confirmation was available in 3/5 patients; 2 had benign tumors and 1 had malignant phaeochromocytoma. In the 2 patients without surgical confirmation, a CT scan had shown lesions, but there was no serum elevation of catecholamines. Conclusions 18F-FDOPA PET can detect and localize nonMIBG-avid phaeochromocytomas with high accuracy and it has the potential to become the functional imaging method of choice for this indication in the future.

A28 68Ga-DOTATATE PET imaging in neuroendocrine and neuroectodermal tumours Z. Win, L. Rahman, S. Khan, J. Murrell and A. Al-Nahhas Department of Nuclear Medicine, Hammersmith Hospital, London, UK. Aim To investigate the role of 68Ga-DOTATATE PET imaging in neuroendocrine tumours (NETs), and in malignant neuroectodermal tumours, in comparison with 111In-octreotide and 123 I-MIBG. Methods Seven patients with biochemically active NETs (2 insulinomas, 1 carcinoid, 2 gastrinoma, 1 VIPoma, 1 somatostatinoma) and 2 patients with malignant neuroectodermal tumours (1 paraganglioma, 1 phaeochromocytoma) underwent 68Ga-DOTATATE PET imaging. All patients with NETs underwent imaging with 111In-octreotide; 1 also had imaging with 123I-MIBG. Both patients with paragangliomas underwent imaging with 123I-MIBG; 1 also had imaging with 111 In-octreotide. Results Table 1 lists the results for the three radiopharmaceuticals. Conclusion With the exception of one patient (low administered activity) 68Ga-DOTATATE PET is shown to be very sensitive in detecting NETs and neuroectodermal tumours compared with 123I-MIBG and 111In-octreotide, and provides superior resolution as a PET agent. Furthermore, 68 Ga-DOTATATE PET imaging should be considered in malignant paragangliomas especially if 123I-MIBG imaging is negative, with a view to targeted radionuclide therapy with suitable beta emitters such as 90Y-DOTATATE. Table 1 Tumour Gastrinoma Insulinoma VIPoma Gastrinoma Insulinoma Somatostatinoma Carcinoid

Paraganglioma Phaeochromocytoma

68

Ga-DOTATATE

Positive Positive Negative Negative Negative Negative Negative (low administered activity) Positive Positive

111

In-octreotide Negative Negative Negative Negative Negative Negative Positive

Positive

123

I-MIBG

Negative

Aim To devise a practical, high quality PET–CT scanning protocol. Methods We assessed our 68Ga-DOTATOC PET–CT protocol using the results of 17 patients scanned to date. Patients were injected with 122–229 MBq of 68Ga-DOTATOC following oral hydration and 500 ml of oral CT contrast. At 45 min postinjection, PET whole-body (vertex to upper thigh) 2-D acquisition; 6 min per bed position were acquired. An additional 3-D PET acquisition (10 min single bed position) was performed over an ROI. A whole-body CT dataset was acquired for fusion with PET. Results All 17 patients tolerated the study well. The range of injected activity had no significant impact on the diagnostic quality of the scans. Oral hydration alone was sufficient to prevent excessive renal tracer accumulation and allow excellent discrimination of renal versus adrenal uptake. Conclusion 68Ga-DOTATOC PET–CT is a simple, welltolerated procedure providing high quality diagnostic images. Prior planning and prioritizing of patients and on-site supply of 68 Ga is important to optimize the technical quality of study. A30 Can complex PET research studies be performed on a state of the art PET/CT system? N.A. Benatar, R.M. Dobbin, P.J.Schleyer, J.E. Mackewn and P.K. Marsden King’s College London School of Medicine at Guy’s, King’s College and St Thomas’ Hospitals, London, UK. Modern state of the art PET/CT systems have been primarily designed to deliver ever faster, high quality results in clinical applications. Experience of using the GE Discovery ST system for research applications, however, has thrown up many challenges that have had to be overcome. The new scanners have raised several quantification and PET performance issues. Furthermore, the use of CT for attenuation correction raises radiation dosage issues when scanning volunteers and the separate CT and PET components of the scanner necessitates that the patient must be moved during the scan. The increased scanner length delivers several ‘access’ issues, not least with regard to blood sampling techniques. The lack of positioning aids on the PET component of the new scanners has also required a solution for ensuring the minimization of patient movement between scan acquisitions, particularly on studies of 2 h or more. Over the last 12 months we have developed several techniques and approaches in order to meet these challenges. We will demonstrate that should these techniques be adopted then research applications can be carried out successfully, largely overcoming the limitations imposed by the new scanners. A31 Radiographer reporting: The referrer verdict S.L. Johns and P.M. Kemp Southampton University Hospitals’ NHS Trust, UK.

Negative Negative

TECHNOLOGISTS I

A29 68Ga-DOTATOC PET–CT: A technologist’s perspective G. Heath, P. Blanchard, J. Dickson and I Kayani Institute of Nuclear Medicine, University College London, UK.

Background A survey was conducted of referrers to gather opinion on radiographers reporting nuclear medicine investigations prior to considering the extending role. Eighty-three consultants received questionnaires, and 53 (64%) responded. Results Seventy-four percent supported radiographer reporting, 17% against, 9% if doctor unavailable. Sixty percent would interpret the report as if issued by a doctor, 11% would ask for a medical opinion, and 40% would be more sceptical of a radiographer report. Seventy-seven percent would discuss unexpected results – 42% with a doctor, 24% with the radiographer, 13% with either. With regard to referrer opinion

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of radiographer reporting in radiology and ultrasound, 80% of reports were of a good standard and 88% of radiographers normally reported to the same standard as doctors. Conclusions Most referrers are in favour of radiographer reporting. New practice is likely to raise uncertainty. It is understandable that referrers may question radiographer reports and may require confirmation from a doctor. Where radiographer reporting is established referrers confirm reports are accurate and to a medical standard. This is encouraging and suggests the approach could work in nuclear medicine. Radiographers could reduce demands on overstretched medical staff. Training and audit are essential. This survey, if repeated as part of an audit cycle, could assess whether referrer opinion changes. A32 How should we measure the diagnostic accuracy in reporting? K.G. Holmesa and M.E. Welshb a University of Salford, and bMorecambe Bay Hospitals Trust, UK. In the spring of 1999 the University of the West of England in Bristol began the first postgraduate ‘Technical Reporting’ module for technologists employed in nuclear medicine. In spring 2002 Salford University ran their first ‘Clinical Reporting Course in Nuclear Medicine’. To date, more than 50 students have successfully completed the two study programmes. Previous conference papers have discussed the growing body of evidence that technologists can report nuclear medicine studies with a degree of accuracy similar to medical practitioners. The course content, assessments and philosophy have also been published [1,2]. The diagnostic accuracy is usually characterized by the sensitivity and specificity of a test but can this method be used when there is no accurate ‘gold standard’? Can the use of concordance of reports be used to compile data to set standards? Using evidence from these courses and the analysis of case studies this paper will explore the methods which may be used to assess competence in the reporting process. It will discuss how to assess the accuracy of reports and factors which may cause errors in the process. It evaluates the audit process, and puts forward the rationale for a possible protocol for standard setting in nuclear medicine reporting. References 1 Hogg P, Holmes K. J Diag Radiogr Imaging 2000; 3:77–85. 2 Holmes K, et al. Synergy 2003; March:10–14. A33 Nicer than NICE: A nuclear cardiology service at a teaching hospital G. Ross, G. Gnanasegaran, A. Taylor, A. Robertson, G. Shabo, N. Mulholland, D. Sharp, E. Lee, W. Gibbs, A. Morley and T.O. Nunan Department of Nuclear Medicine, Guys and St Thomas’ Hospital NHS Trust, London, UK. Background and aim The National Institute for Health and Clinical Excellence (NICE) has published the results of its appraisal of myocardial perfusion scintigraphy (MPS). The current average waiting time is 20 weeks and the recommended appropriate upper limits of waiting time are 6 weeks and 1 week for routine and urgent studies, respectively. The aim of this study was to evaluate nuclear cardiology practice at our institution in accordance with the NICE guidelines. Method We reviewed patient files from April 2000 to July 2005 for the number of scans performed annually and the waiting time. We also evaluated factors such as number of staff involved in stress sessions per week.

289

Results 9968 MPS scans were performed during these periods. The waiting times for non-urgent MPS were 5–10 weeks in 2000, 5–11 in 2001, 9–18 in 2002, 6–22 in 2003, 6–19 in early 2004 and in the mid 2005 it was < 1 week for urgent scans and 4–5 weeks for routine requests. This was despite no reduction in referral numbers. The number of staff has remained the same. However, over the last 2 years, 3 technologists have been trained in stress testing. Currently, dedicated secretarial staff/ cardiac nurse are involved in bookings/scheduling. This has resulted in a reduction from 35% to 20% of those patients who did not attend their appointments. Conclusion Overall, our hospital performs urgent MPS in < 1 week and routine requests in < 5 weeks. However, education/ training of both technologists and nurses can bring about the required change. We should emphasize the benefits of MPS in a NICE (nice) way.

TECHNOLOGISTS II A34 Escalating demand for the application of information systems in a clinical environment J.J. Steele and M. Masoomi St. Mary’s Hospital, Portsmouth and Royal Hospital Haslar, Gosport, UK. Aim To provide a system whereby patient appointments, the results and administered radionuclide activities can be recorded for all department radionuclide diagnostic tests and therapies at multiple locations. Method The booking of diagnostic tests and therapies requires prompt implementation. Provision of such services is very demanding and time consuming and may not be achieved by the availability of trained staff over multiple site locations and out of hours. The availability of results to oncology is vital for prompt planning and treatment. The system allows appointments to be booked for predefined slots by designated or nominated members of hospital staff without direct supervision. The system has been developed in Visual Basic for Excel allowing portability and accessibility over the hospital trust network, is built around a calendar-style database, and uses forms to restrict user input. Access to the system has been categorized accordingly. Periodic revision takes place to monitor access and functionality. Time restrictions allow continual access. Conclusion The system is fully automated and additional databases have been developed for data analysis, patient statistics and pollution inventory. It provides instant access for booking appointments, and by allowing patients to be given appointments in a clinic reduces the number of cancellations. The system saves time between referral and appointments and provides prompt access to results for diagnosis and treatment within trust targets. A35 Experience of sentinel node lymphoscintigraphy in penile carcinoma D.E. Hoffland, S.A. Bassingham, A. Irwin, N. Gulliver, P. Hadaway, J. Crawshaw and S.D. Heenan St George’s Healthcare NHS Trust, London, UK. Pre-operative sentinel node lymphoscintigraphy (SNL) and biopsy has been widely accepted in melanoma and other tumour staging. Drawing on our experience with melanoma SNL ( > 1000 cases) we report as the first NHS trust in the UK on setting up and delivering a service for penile carcinoma. Over 30

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patients have undergone this procedure in the last year. We discuss the resources required, administration technique, imaging procedures, and the need to work as a multidisciplinary team in order to provide an efficient and responsive service. We consider factors including ARSAC licence (research at present), patient preparation, information for wards, liaison with theatre and urology teams, availability and QC of gamma probe, training of urology team in use of gamma probe, coordination with ultrasound imaging and fine-needle aspiration (FNA), technologist/radiographer training, experience and resources (requires 2 members of staff), and potential for radiographer reporting. We will present some case studies demonstrating drainage pathways and technical issues that may arise. A36 PET/CT scan with 18F-FDOPA in the diagnosis of congenital hyperinsulinism in children A. Zonoozia, P.A. Beaumontb, G. Plantc, K. Hussaind and J.R.W. Halle a Guildford Diagnostic Imaging, bSt Peter’s Hospital, Chertsey; cNorth Hampshire Hospital, dGreat Ormond Street Hospital, and eFrimley Park Hospital, UK. Background Congenital hyperinsulinism (CHI) is a neuroendocrine disease characterized by profound hypoglycaemia caused by inappropriate insulin secretion from neuroendocrine tumours (NETs) with both diffuse and focal forms. NETs can take up amine precursors and convert them into biological amines. This study evaluates the findings of [18F]fluorodihydroxyphenylalanine (18F-FDOPA) PET/CT scans in children with CHI and also describes the aspects and challenges of carrying out this type of scan in paediatrics. Methods Five patients with CHI were initially enrolled in this study. All patients were injected with 4 MBq  kg – 1 18F-FDOPA and imaging performed 1 h later. The scan was acquired for 10 min in the abdomen area. Results Four of 5 scans showed diffuse abnormality rather than focal uptake in the pancreas. One patient showed abnormality of the pancreas with focal areas of intense activity in an almost nodular pattern throughout the head, body and tail. Conclusion The ability of this noninvasive method is in visualizing NETs and also distinguishing between focal and diffuse forms of CHI in children. Maintaining a stable blood sugar and keeping the child calm and occupied post-injection were 2 of the major challenges. A37 A study to assess the feasibility of introducing radiographer reporting in a nuclear medicine department K. Custis Radiology Department, Princess Royal University Hospital, Bromley, Kent, UK. Purpose To determine whether a radiographer could attain the same diagnostic accuracy as a consultant radiologist in the reporting of nuclear medicine studies; to implement a radiographer reporting protocol; to establish radiographer reporting in nuclear medicine. Methods A senior 1 radiographer and a consultant radiologist independently reported 500 nuclear medicine bone, lung V/Q and renal scans. The radiographer had completed an accredited course in ‘Clinical Nuclear Medicine Double Reporting’. Reports were compared and categorized by both radiologist and radiographer. Cases where agreement was not reached were referred to a second consultant radiologist for decisional scoring.

Results The agreement rate for bone scans was 98.5% with 7.8% having a trivial difference, 3.3% having a sustainable difference and only 1.4% having a non-sustainable (potentially affecting patient management) difference. For renal scans 99.37% totally agreed, 3.67% had trivial differences, 0.2% had a sustainable difference and 0.6% had a non-sustainable difference. Lung V/Q scans had agreement of 98.1%, 4.37% with trivial, 5.0% with sustainable and 1.87% with nonsustainable differences. Conclusion The overall average agreement rate was 98.7% and it is concluded that it is therefore feasible to train radiographers to report nuclear medicine examinations with protocols in place as required by trust governance bodies.

BONE A38 99mTc-depreotide: a potent imaging marker of the activity of bone inflammation N. Papathanasiou, Ph. Rondoyianni, N. Pianou, E. Skoura, E. Vlontzou and I. Datseris Nuclear Medicine Department, ‘‘Evangelismos’’ General Hospital, Athens, Greece. Aim Somatostatin receptors are over-expressed in inflammatory cells (macrophages and activated lymphocytes). The purpose of this study is to assess the utility of 99mTc-depreotide – a somatostatin analogue with high affinity for the receptor’s subtypes 2, 3 and 5 – as a bone inflammation marker. Methods Seventeen patients underwent depreotide scintigraphy, a three-phase bone scan as well as imaging by other modalities (CT, MRI, gallium scan). The patient population consisted of 9 active osteomyelitis cases and 8 various other pathologies (avascular necrosis, fracture, healing osteomyelitis, aseptic loosening, periodontal infection with concomitant bone reaction, septic arthritis and 2 cellulitis cases). Results The depreotide scan was positive in the 9 cases of verified osteomyelitis as well as in the osteonecrosis case (probably due to the co-existing inflammation). Three scans were negative in the cases of the bone fracture, the aseptic loosening and the healing osteomyelitis, indicating the success of therapeutic interventions. In the remaining 4 cases, the depreotide scan, when interpreted along with the bone scan, enabled us to identify the exact site of infection and rule out osteomyelitis. Conclusion The results suggest a potential role of 99mTcdepreotide as a marker of the activity of bone inflammation. A39 Wrist registration scintigraphy: Use by specialist hand surgeons N.J.R. Mulholland, G. Gnanasegaran, V. Raman, S.E.M. Clarke, B. Poulsen and I. Fogelman Guy’s and St Thomas’ NHS Trust, London, UK. Background and aim Wrist registration scintigraphy (WRS) combines functional information with anatomic detail to localize pathology. The objectives of this study are (1) to evaluate the referral base, indication and use of WRS, and (2) to develop an algorithm for its clinical use. Method All WRS performed between May 2003 and July 2005 at Guy’s Hospital were reviewed with referrer and indication noted. Where available, follow-up information was obtained via case notes and orthopaedic lead consultant.

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Results Of the 61 scans performed, 54 were for the investigation of occult wrist pain referred by specialist orthopaedic hand surgeons. Follow-up is ongoing and presently is available in 20/54 cases. WRS altered the management in 13/20. Five arthroscopies were avoided, 8/20 proceeded to different surgery, 3 were discharged and 1 patient was medically managed. Although WRS did not alter management in 7/20 cases, in 4 provisional management was confirmed. In 3/20 cases WRS was non-contributory. Conclusion WRS is a useful tool in the specialist management of occult wrist pain, which can avoid unnecessary arthroscopy and alter the surgery offered to patients. We will present an algorithm for the use of WRS in the management of occult wrist pain.

corticosteroids (80 mg triamcinalone) under fluoroscopy. The mean VAS score was reduced from 7 to 3 at 6 weeks follow-up, a reduction of 42%. Conclusion We conclude that bone SPECT of the SIJs in the post-surgical setting can identify patients with sacroiliitis who may then benefit from local steroid therapy.

A40 Paediatric wrist injury: Scintigraphic evaluation and clinical management H.K. Cheow, P. Set, S. Robinson and K.K. Balan Addenbrookes Hospital, Cambridge, UK.

Background From January 2002 to February 2005, 200 patients were referred by GPs for isotope bone scanning in bone pain where conventional radiography has proved inconclusive. A majority of these patients have had a previous carcinoma. Method Analysis of the clinical records and scans was carried out to determine indication and reports. Result Metastatic disease was present in 23.5% of (47/200) patients. A majority of patients had breast cancer 40.4% (19/47), lung carcinoma 10.6% (5/47), and prostatic carcinoma 8.5% (4/47). Conclusion Referral from a GP is shown to be clinically effective and obviates the need for initial referral to an oncologist. In this model, a patient would see the primary care physician and be referred only if in need of further management. This would result in considerable cost and time savings and reduce the psychological pressures on the patients. This study suggests this should be offered routinely, rather than the informal provision presently in departments. The clinical effectiveness of a primary referral system within this context is demonstrated. This study raises future research opportunities into the nature of bone pain and its relationship to osteoclastic and osteoblastic disease.

Aim To evaluate the role of skeletal scintigraphy in the diagnosis and management of paediatric wrist trauma. Materials and methods All bone scintigrams and plain radiographs performed over 2 years in children under 16 years were reviewed retrospectively. Patients were referred when the immediate and post-injury radiographs (mean 10 days) were considered normal. Results Fifty children were studied, 31 were normal in both studies. Six, however, were treated for a further median period of 9 days on clinical grounds. Four patients with scintigraphic soft tissue injury received no further treatment. Four of 6 patients with suspected fracture on scintigraphy were treated for 20–44 days (median 27 days). Two patients with possible fracture on radiographs and definite fracture on scintigraphy were treated for 17 and 35 days. One of 3 patients with a positive radiograph and normal scintigraphy was treated for 36 days. All 4 patients with positive scintigram and negative radiograph went on to have treatment for 2–47 days (median 35 days). No complications were recorded. Conclusion Bone scintigraphy is a useful non-invasive method for the assessment of paediatric wrist injury in the context of normal plain radiographs. Clinicians generally treat the scintigraphic information seriously. A41 Bone SPECT following spinal surgery P. Ryan, A. Agarwal and A. Hammer Medway Maritime Hospital, Chatham, UK. Background Back pain following spinal surgery is difficult to evaluate with multiple diagnoses possible. This study investigated the importance of sacroiliitis in this setting and the potential role of the bone scan with SPECT of the sacro-iliac joints (SIJs). Methods Patients with low back pain of more than 7/10 on a VAS scale 6 months post-decompression or discetomy without other post-operative complications were investigated with 2phase bone scintigraphy of the lumbar spine and pelvis with SPECT. Results Of 76 patients in total, 32 had lumbar decompression, 24, discetomy, 12 decompression with posterolateral fusion, and 8 dynamic stablilization. Sacroiliitis was diagnosed by identification of high uptake in the SIJs on expert physician judgment using visual inspection of the images aided by quantification. Fifty-six (73%) patients were considered positive for sacroiliitis and 1 or both joints treated with periarticular

A42 A retrospective study of GP referrals for undiagnosed bone pain in patients at risk of metastatic disease S. Khana, T.C. Ohb, J.C. Hilla,c and J. Coffeya a Royal Preston Hospital, bBlackpool Victoria Hospital, and cUniversity of Salford, UK.

CARDIOLOGY II A43 The impact of arrhythmias during gated myocardial perfusion tests: Is there a need to repeat the acquisition? P. Arumugam, K. Herman and A. Bradley Manchester Royal Infirmary, UK. Previous publications have indicated that functional parameters [1] and possibly the summed perfusion images [2] derived from gated myocardial perfusion tests can be erroneous if the patient suffers from an arrhythmia during the acquisition. Currently, we repeat the study with an ungated acquisition when there is data loss due to arrhythmias to ensure the perfusion images are uncorrupted. Clinicians felt that there were no significant differences in the perfusion images between the two studies. The aim of this study was to confirm this impression and reduce the need to repeat scans. The original data from 10 patients were altered to decrease the total counts in various frames to simulate arrhythmia. Several datasets were produced for each patient varying from count losses in a single frame to multiple frames. An experienced observer reviewed the original data and the altered data to look for changes in the reconstructed perfusion images. In most cases no significant changes were observed. The same observer confirmed the study findings in a number of patients where the test had been repeated, due to the loss of data caused by an

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arrhythmia during the acquisition, by comparison of the original gated and un-gated acquisitions.

Table 1

References 1 Nichols K, et al. J Nucl Cardiol 2001; 8:19–30. 2 Germano, et al. Clinical Gated SPECT. Futura Publishing; 1999, p. 102.

IRACSC IRAC IRSC IR FBP FBPSC

A44 Patient motion during acquisition of myocardial perfusion SPECT studies: When is motion correction required? K.C. Cockburn, G.A. Wright, A.C. Tweddel, G. Avery and G. Davies Hull and East Yorkshire Hospitals NHS Trust, UK. Introduction Patient movement during SPECT may create artefacts. It is unclear what magnitude of motion affects clinical interpretation. Aim To investigate the effect of motion on SPECT myocardial perfusion studies. Method Three types of y-direction motion were simulated in one normal patient study and two phantom studies (one with an anterior defect). For type A motion, two projections were moved. Type B simulated a step mid-acquisition. Type C simulated motion on odd-numbered projections. After reconstruction, images were divided into five segments (lateral, anterior, septal, inferior, apical) and scored on a 5-point scale (1 = definitely normal, 2 = probably normal, 3 = equivocal, 4 = probably abnormal, 5 = definitely abnormal). Artefacts were considered present in normal segments with scores = 3. True defects were considered affected if the segment score was < 4. Results No effects were seen for type A motion of up to 100 pixels. Type B motion = 2.4 pixels introduced artefacts. Type C motion = 2.0 pixels introduced artefacts. In all reportable phantom images, true defects were given a score of 5. Conclusion The effect of motion varies with motion type. No adverse effects were seen with motion < 2.0 pixels. Motion on only two frames had no noticeable effects.

A45 A comparison of six reconstruction methods for myocardial perfusion SPECT: An ROC phantom study G.A. Wright, K.C. Cockburn, A.C. Tweddel, G. Avery and G. Davies Hull and East Yorkshire Hospitals NHS Trust, UK. Introduction Iterative reconstruction (IR) with attenuation and scatter correction (AC & SC) may reduce artefacts on SPECT myocardial perfusion scintigraphy (MPS). A variety of reconstruction methods are available and the optimal method has not been established. Aim To compare the diagnostic accuracy of six reconstruction types: filtered back-projection (FBP), FBP with SC (FBPSC), IR, IR with SC (IRSC), IR with AC (IRAC), and IR with AC & SC (IRACSC). Methods Twenty-six SPECT images of a myocardial phantom (with and without simulated perfusion defects) within an anthropomorphic torso phantom were reconstructed using each technique. Hawkeye CT images were used for AC. Images were analysed using Cedars Sinai QPS software to obtain defect scores for anterior, antero-lateral, infero-lateral, inferior, inferoseptal, antero-septal and apical segments. ROC analysis was performed on 182 segments and the area under the curves (AUCs) calculated.

Reconstruction method

AUC ± std error

P(vs. FBP)

0.83 ± 0.04 0.82 ± 0.04 0.82 ± 0.04 0.83 ± 0.04 0.74 ± 0.05 0.72 ± 0.05

0.009 0.061 0.002 0.007 – 0.699

Results Significantly higher AUCs were obtained for IR than FBP (see table). AC or SC did not significantly affect the AUC (Table 1). Conclusion IR is likely to improve the diagnostic accuracy of MPS without the need for attenuation or scatter correction. A46 The correlation of ejection fraction and end systolic volume in gated SPECT: Should we report one or both? R.A. Peace and J.J. Lloyd Regional Medical Physics Department, Royal Victoria Infirmary, Newcastle upon Tyne, UK. Objective Left ventricular ejection fraction (EF) and end systolic volume (ESV) are measures of cardiac function which are frequently reported from gated myocardial perfusion SPECT. However, the correlation of EF and ESV gSPECT measurements and their relative significance for normal and infarcted myocardium has not previously been published. Method Gated SPECT studies with normal perfusion and no known cardiac disease (NORM, n = 127); normal perfusion, but clinical diagnosis of MI (MIND, n = 39); fixed defect consistent with MI and a diagnosis of MI (MIFD, n = 110) were selected. Patients were imaged with a 2 day stress–rest 400 MBq 99mTctetrofosmin protocol. The QGS software package was used to calculate EF and ESV. Results There was a strong correlation between EF and log10(ESV) for NORM, MIND and MIFD study groups (r = 0.89, 0.88 and 0.94). The data for the three groups as a whole was also highly correlated (r = 0.95). The logarithmic relationship demonstrates that EF is more sensitive than ESV to functional changes when function is normal. Although the relationship suggests that ESV is more sensitive in the presence of dysfunction, the advantage is outweighed by increased ESV measurement errors. Conclusion The EF, rather than ESV, should be reported for patients with and without cardiac dysfunction due to MI. A47 Does attenuation correction of gated tomographic cardiac blood pool studies affect left ventricular volumes and ejection fraction values? F. Sundram, A. Notghi and A. Deakin Department of Physics and Nuclear Medicine, City Hospital, Birmingham, UK. Aim We examined the effects of transmission attenuation correction (TAC) on left ventricular volumes (LVV) and ejection fraction (LVEF) values obtained from tomographic cardiac blood pool (TBP) studies. Methods Fourteen patients had TBP studies after erythrocyte labelling and injection of 800 MBq [99mTc]pertechnetate. Emission scans parameters were: 128  128 matrix, pixel size 4.8 mm and 2041 angular range. This was followed by 3601 transmission scans using the BEACON system (employing 133Ba point sources). Iterative reconstruction generated short-axis

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Table 1

Range Mean ± SD

LVEF (non-AC)

LVEF (AC)

LVEDV (non-AC)

LVEDV (AC)

LVESV (non-AC)

LVESV (AC)

20–85% 56 ± 18%

22–74% 52 ± 15%

35–232 ml 126 ± 59 ml

46–251 ml 134 ± 57 ml

8–167 ml 63 ± 47 ml

12–196 ml 72 ± 49 ml

(SA) slices (with and without AC) which were automatically processed by the QBS program (Cedars-Sinai Medical Centre), yielding paired LVV and LVEF values. Minitab (v.14) was used for statistical analysis. Results See Table 1. There was statistically significant correlation between non-AC and AC LVEF (r = 0.92, P < 0.001) and non-AC and AC EDV and ESV values (r = 0.93, P < 0.001). Conclusions In patients in whom photon attenuation may pose difficulties, attenuation corrected LVEF values can be used in clinical practice. A48 Comparison of first-pass and tomographic radionuclide ventriculography for right ventricular ejection fraction values F. Sundram, A. Deakin and A. Notghi Department of Physics and Nuclear Medicine, City Hospital, Birmingham, UK. Background/aim First-pass ventriculography (FPV) is used to calculate right ventricular ejection fraction (RVEF). The technique is dependent on optimal technical factors. Tomographic ventriculography (TV) better delineates the RV and eliminates some technical factors. We compared RVEF obtained by FPV and TV in 12 patients. Methods FPV was performed after erythrocyte labelling. 800 MBq [99mTc]pertechnetate was injected and a 30 s anterior dynamic acquisition commenced. The data were analysed to obtain RVEF. TV was performed immediately after FPV utilizing a dual-headed elliptical step and shoot technique, employing a 2041 angular range. Data were reconstructed using low-pass filter, order 6, cut-off 0.33 cycle/pixel to yield short-axis (SA) slices. The slices were processed automatically by the QBS program to yield RVEF values. Results FPV RVEF range was 17–33% (mean 26 ± 6%). TV RVEF range was 40–72% (mean 52 ± 13%). Linear regression analysis of paired RVEF values yielded a correlation coefficient (r) value of 0.37, P = 0.24 (not statistically significant). Conclusions There is poor agreement between FPV and TV RVEF values with discrepancies between first-pass and tomographic RVEF values. Further validation with perhaps, MRI RVEF values is needed to assess the reliability of either of the two techniques. RADIOPHARMACY A49 Alkoxysilane groups for instant labelling of biomolecules with 18F U. Choudhry, K.E. Martin, S. Biagini and P.J. Blower Biosciences Department and School of Physical Sciences, University of Kent, Canterbury, UK. Current methods for 18F labelling of peptides for PET imaging are time-consuming. More efficient methods are required because of the short half-life (110 min). It has been suggested [1] that single-step nucleophilic fluorination with fluoride ions may be possible using silicon–fluorine chemistry.

This work aims to identify suitable alkyl groups to include in the required alkoxytrialkylsilane, in order to achieve the fastest radiolabelling (nucleophilic displacement of alkoxide by fluoride) and slowest Si–F bond hydrolysis (hence optimal in vivo stability). Four model trialkylfluorosilanes were synthesized and evaluated using HPLC and NMR to determine rates of fluorination and hydrolysis of Si–F bonds: t-butyldimethylfluorosilane, triphenylfluorosilane, t-butyldimethylfluorosilane and dimethylphenylfluorosilane. Rapid Si–F bond formation on reaction of trialkylalkoxysilanes with fluoride was confirmed by 19F and 1H NMR in all cases, showing that rapid fluorination is feasible. However, only t-butyldiphenylfluorosilane was hydrolytically stable for sufficient time to allow imaging. Close to 100% stability was observed over at least 5 h at 451C in water and in phosphate buffered saline. It is therefore feasible that biomolecules containing the t-butyldiphenylalkoxysilane group could be quickly labelled with [18F]fluoride ions, and that the labelled product would possess adequate in vivo kinetic stability for effective imaging after several half-lives. Reference 1 Walsh JC, et al. J Labelled Comp Radiopharm 1999; 42(suppl): S1–S3.

A50 Iodogen labelling method preserves immunoreactivity of high specific activity 131I-labelled rituximab for clinical RIT Y. Dua,b, M. Bayneb, M. Zivanovicb,c, P.W. Johnsonb and T.M. Illidgeb,c3 a Institute of Nuclear Medicine, University College London, bCancer Sciences Division, University of Southampton, and cCancer Studies Division, University of Manchester, UK. Purpose The anti-CD20 rituximab is a good candidate for radioimmunotherapy of B-cell lymphoma. However, compromised immunoreactivity has been reported when rituximab was labelled with only dosimetric level of 131I activities [1]. As part of a phase I/II clinical trial evaluating the safety and therapeutic efficacy of 131I-rituximab in fractionated radioimmunotherapy for B-cell lymphoma, the immunoreactivity of varying specific activities of 131I-labelled rituximab were measured in 70 consecutive radiolabelling procedures. Methods Five mg of rituximab was labelled by the Iodogenbead method with activities of 131I ranging from 300 MBq to 3.0 GBq. The immunoreactivity of radiolabelled rituximab was monitored by the Daudi cell binding assay, whilst the labelling efficiency and radiochemical purity were measured by both ITLC and HPLC. Results Varying specific activities of 131I-labelled rituximab showed no statistically different cell binding capacity, with saturation at 63.3% ± 5.75%. This is comparable to the binding capability of trace amount 125I-labelled rituximab. High labelling efficiency of greater than 95% was consistently achieved with no recognizable degradation of antibody seen with HPLC. Conclusion Over a wide range of 131I specific activity, a high level of 131I labelling efficiency was achieved, with no significant

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change in immunoreactivity. This range covered both clinical diagnostic/dosimetric and therapeutic applications. Reference 1 Schaffland AO, et al. J Nucl Med 2004; 45:1784–1790.

A51 Synthesis of phosphonate conjugates of 188Re-DMSA for treatment of bone metastases U. Choudhry and P.J. Blower University of Kent, Canterbury, UK. Background 186/188Re-bisphosphonate complexes show promise clinically for treatment of bone metastases in prostate cancer. However, target-to-background ratios are lower than in 99mTc bone scans and there is considerable soft-tissue retention. The complexes are inhomogeneous, unstable in vivo and structurally ill-defined, because, while the bisphosphonate group is effective for bone targeting, it is a poor rhenium chelator. Our aim is to improve 186/188Re bone agents by synthesizing bifunctional conjugates in which the rhenium-chelating function is separate from the bisphosphonate group. Methods Because of its excellent in-vivo stability, ease of synthesis and intrinsic bone targeting properties, we chose 188 Re(V)-DMSA as the chelate component. 188Re(V)-DMSA was converted to its bis(anhydride) by treatment with 1-ethyl(3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), followed by reaction with a series of aminophosphates to prepare bisphosphonate conjugates. Alternatively, conjugates could be prepared directly from 188Re(V)-DMSA in water by activation with EDC. The reaction could be controlled to give either the cyclized imide product or the ring-opened form. The products, identified by mass spectometry and HPLC, were stable in serum and bound readily to hydroxyapatite. Conclusion We conclude that the 188Re(V)-DMSA core is an attractive basis for synthesis of stable, well-defined bisphosphonate conjugates from which optimal bone agents may be developed.

A52 Comparison of 99mTc-HYNIC and 99mTcphenylhydrazine in vitro using a gastrin analogue R.C. Kinga, S.J. Mathera, P.J. Blowerb and B. Surfrazb a Cancer Research UK, Department of Nuclear Medicine, St Bartholomew’s Hospital, London, bCancer Research Group, Department of Biosciences, University of Kent, Canterbury, UK. Objectives Hydrazinonicotinamide (HYNIC) has been established as a useful chelator of 99mTc. Using an analogue of gastrin attached to HYNIC or phenylhydrazine, we have undertaken a comparison to determine the importance of a nitrogen atom within the cyclic ring of HYNIC as a complex stabilizing factor. Three different co-ligand systems were explored; tricine (T), tricine/nicotinic acid (T/N), and EDDA/tricine (E/T). Methods For radiolabelling, 10 mg peptide, 50 mg co-ligand, 200–400 MBq 99mTc, 30 mg stannous chloride in ethanol solution was used. Conditions were altered to optimize radiolabelling. The resulting complexes were purified using a standard sep-pak method and stability (in phosphate and plasma) was detected by HPLC. A protein binding assay was performed of the purified complex in plasma. Results The HYNIC derivatives showed higher stability, notably using tricine or tricine/nicotinic acid co-ligands than the phenylhydrazine derivatives. In particular, there was a marked difference in the stability of the derivatives in plasma, for example, using tricine as co-ligand the percentage of 99mTc-

labelled peptide at 5 h was 90.2% for the HYNIC derivative and 31.9% for the phenylhydrazine derivative. There was a greater degree of protein binding of the phenylhydrazine derivatives. Conclusions The stability of the peptide–phenylhydrazine–99mTc complex in vitro is inferior to the peptide– HYNIC–99mTc complex. This further supports the theory that the nitrogen atom of the cyclic ring of HYNIC is involved in the stability of the 99mTc–co-ligand–HYNIC complex.

A53 Solid phase synthesis of peptide radiopharmaceuticals using Fmoc-N-o-(HYNIC-Boc)lysine, a technetium-binding amino acid M.B. Surfraza, R. Kingb, S.J. Matherb, S. Biaginic and P.J. Blowera a School of Biosciences, The University of Kent, Canterbury, bDepartment of Nuclear Medicine, St Bartholomews Hospital, London EC1 7BE; and c School of Physical Sciences, The University of Kent, Canterbury, UK. Labelling of proteins with radionuclides for use in radiopharmaceuticals involves covalently attaching a bifunctional chelator such as HYNIC. If there is more than one lysine/primary amine in the peptide, mixed products can result. To circumvent this problem, a lysine–HYNIC conjugate, Fmoc-No-(HYNIC-Boc)-Lys, was synthesized from NHS-HYNIC-Boc [1] for incorporating into peptides during solid phase peptide synthesis (SPPS) for subsequent labelling with 99mTc. Fmoc-No-(HYNIC-Boc)-Lys can thus be incorporated in place of any amino acid at any chosen location in the sequence, with no further modification once the peptide is cleaved from the resin. Salmon calcitonin was efficiently labelled using this strategy, with HYNIC–lysine in place of Lys-18 [2]. During cleavage of the peptide from the resin with trifluoroacetic acid, the hydrazine functionality can become trifluoroacetylated. This reaction was optimized to yield either the trifluoroacetamide-protected product or the free hydrazine. Optimized conditions for the removal of trifluoroacetyl group were found. Procedures for direct labelling of the trifluoroacetamide by-product with 99mTc are under investigation. We conclude that Fmoc-N-o-(HYNIC-Boc)-Lys is a versatile amino acid for incorporation into peptides during SPPS. Amino acid chelators for radionuclides other than 99mTc are currently being pursued for synthesis of peptide libraries for radiolabelling. References 1 Abrams MJ, et al. J Nucl Med 1990; 31:2022–2028. 2 Greenland WEP, et al. J Med Chem 2003; 46:1751–1757.

NEUROLOGY

A54 Radiological features, clinical significance and utility of DaTSCANs in the management of atypical presentations suspicious for Parkinson’s disease C. Cronin, N. Ellis, D. Lohan, T. Counihan and D. O’Keeffe University College Hospital, Department of Radiology and Neurology, Galway, Ireland. Purpose Parkinson’s disease is diagnosed in more than 1/200 worldwide. However, 20–25% of these are misdiagnosed. As the average population age is increasing, the need for accurate diagnosis and treatment is highlighted. We retrospectively reviewed the radiological features, and in correlation with clinical background, determined the clinical and radiological

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34th BNMS meeting: abstracts 295

utility of DaTSCANs in diagnosis and management of 40 diverse patients with atypical presentations suggestive of PD. Materials and methods Forty patients’ (22 male, 18 female, aged 28–80 years,mean 60.5 years, Hoehn/Yahr stage 1–2) images were reviewed independently by 2 radiologists. Striatial radiotracer uptake was assessed visually, abnormal images graded on a 0–3 scale. Results Number/severity of symptoms and sides involved, correlated with degree of deficit at visual assessment. Twelve of 40 suspicious for essential tremor (ET), 10/40 for drug/toxin induced Parkinson’s (D/TIP), 6/40 for Parkinson-plus syndrome (PPS)and 12/40 for unclassical PD. DaTSCAN supported diagnosis of ET in 5/12, D/TID in 5/10; PPS in 2/6 PPS and IPD in 12/12 suspicious for IPD and 16/28 from other 3 groups. Imaging changed the clinical diagnosis in 16/40, medication in 17/40. No discrepancies were identified at follow-up to date. Conclusion The cost of a DaTSCAN is not inconsiderable (approximately 5–10 months of anti-Parkinson’s medications). However, long-term cost supports scanning, limiting inappropriate investigations/medications. Our experience demonstrates DaTSCANs have a significant role in diagnosis and management of unusual presentations for PD.

A55 A rCBF study of hallucinations in dementia R.T. Staffa, K. Benjaminb, A.D. Murrayc, H.G. Gemmella and C. Wischikb a Department of Bio-Medical Physics, Aberdeen Royal Infirmary, bTauRx Therapeutics Pte Ltd, Institute of Medical Science, University of Aberdeen, and cDepartment of Radiology, University of Aberdeen, UK. Aim Hallucinations are one of the most common noncognitive symptoms seen in patients with dementia. The presence of hallucinations strongly contributes to institutionalization, reduced patient well-being and increased burden on the caregiver. A possible mechanism for the genesis of hallucinations in dementia is that they arise from a lesion in a particular brain region. In this work we aim to test this hypothesis. Methods We imaged 62 patients who were routinely referred for 99mTc-HMPAO brain SPECT imaging as part of their work-up for dementia. From this group 12 had been experiencing visual hallucinations. After reconstruction and correction for attenuation we performed a SPM group comparison between those with and without visual hallucinations using gender, age and cerebellar activity as confounding (nuisance) variables. Results After correcting for multiple comparisons, we found a region in the visual cortex of significantly lower perfusion in the hallucinating group that in the group without hallucinations (P = 0.011 voxel level, P < 0.001 cluster level). Conclusion It has previously been thought that the presence of such non-cognitive symptoms is associated with a greater level of disease severity. Visual hallucinations in dementia are caused by a defect in the visual cortex.

A56 Correlation between visual interpretation and quantification of DaTSCAN SPECT imaging A. Nafati, J.R. Buscombe and A.J.W. Hilson Royal Free Hospital, London, UK. Background DaTSCAN (123I-ioflupane) is used for the differentiation of Parkinson syndrome (PS) from parkinsonism without nigrostriatal degeneration [1].

Objective To compare the visual assessment of DaTSCAN with specific binding ratios (SBRs) as calculated using semiquantitative analysis. Methods The study included 31 patients with parkinsonism (19 male, 12 female, mean age 55.3 ± 15 years). Three nuclear medicine consultants rated the images according to the classification reported by Catafau et al. [1]. SBRs were calculated for the striatum and striatal subregions (head of caudate and putamen) semi-quantitatively from 2 summed consequtive transaxial slices with the most intense striatal binding [2]. Results Approximately 1/3 of the patients were rated as normal and 2/3 as abnormal. Seventy percent of abnormal scans were put in ‘abnormal 2’ grade, 15% in ‘abnormal 1’ and 15% in ‘abnormal 3’. The mean nigrostriatal SBR decreased substantially from 3.20 ± 0.91 in the visually assessed normal scans, to 1.97 ± 0.79 in ‘abnormal 1’, to 1.47 ± 0.51 in ‘abnormal 2’ and to 0.51 ± 0.27 in ‘abnormal 3’ scans with similar decrease in all nigrostriatal subregions. The visual assessment grades showed highly significant negative correlation with SPRs calculated (mean Sprearman’s correlation coefficient 0.815, P < 0.0001). Conclusion Semi-quantitative DaTSCAN imaging showed high significant correlation with the severity of PS as assessed visually. References 1 Catafau AM, et al. Mov Disord 2004; 19:1175–1182. 2 Tatsch K, et al. Eur J Nucl Med Mol Imaging 2002; 29: BP23–BP29.

A57 DaTSCAN: qualitative or quantitative interpretation in Parkinson’s disease G. Shabo, A.G. Kettle and M.J. O’Doherty Guy’s and St Thomas’ Hospital, London, UK. Background This study demonstrates the effects of image reconstruction methods, visual and quantitative analysis on the interpretation of DaTSCAN studies and the impact of these effects on the management of the patients with suspected parkinsonian syndrome. Methods Visual interpretation of 53 consecutive DaTSCANs performed in our department between 2002 and 2004 were re-read by 3 experienced clinicians. The imaging set-up was biased to utilize for each patient the smallest radius of rotation. The datasets qualitatively assessed were from oblique re-orientated transverse slices obtained by iterative reconstruction. The patient projection data were also reconstructed by filtered back-projection with and without Chang attenuation correction. Semi-automated quantification using the Southampton method was applied to the datasets generating specific binding ratio (SBR) values for comparison. Results Kappa values for inter- and intra-observer qualitative interpretation showed moderate to good agreement. Over 80% of the readings agreed with the original reports. Inter- and intraoperator R2 values showed good correlation for manual quantification. SBR values applying Southampton semi-quantification could not confidently segregate between normal and parkinsonian scans and these values for an individual patient varied with reconstruction technique applied. Conclusion Semi-quantification did not change the reports significantly. The qualitative assessment of DaTSCAN images is as good as the quantitative assessment. The semi-quantitative method is a useful aid but not indispensable.

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A58 The 3-D fractal dimension of DaTSCAN images L. Bolt, J.S. Fleming and P.M. Kemp Southampton University Hospitals NHS Trust, UK. Background Fractal analysis was recently applied to DaTSCAN images using 2-D summed images with scaling based on the maximum striatal counts. This study further investigates its potential by (1) using an alternative scaling approach and (2) expanding it to 3 dimensions. Method The analysis was carried out on a ‘slab’ that included all the slices with striatal counts. The scale k was defined as the percentage of the mean non-specific uptake, NSmean, measured from the whole brain except the striatum. The striatal uptake was measured as the number of voxels N(k) with counts above k. N(k) followed the fractal relationship N(k) = constant  k( – FD) (where FD is the fractal dimension) within the range 220–265% of NSmean. This becomes linear on a log–log plot and the slope of the regression line gave FD. A fully automated analysis was performed on 25 ‘normal’ and 30 ‘abnormal’ scans. Results The fractal dimension was able to separate normal from abnormal scans with both sensitivity and specificity above 96%, higher than those achieved previously. Conclusion The 3-D analysis and the use of a scale based on NSmean improve the robustness of the FD. With its high clinical concordance fractal analysis provides a useful adjunct to visual assessment.

A59 Dopamine transporter imaging of tremulous disorders D.J. Hensmana, P.G. Baina and J.W. Frankb Departments of aClinical Neurosciences, Charing Cross Campus, Imperial College London, and bNuclear Medicine, Charing Cross Hospital, London, UK. Objective To evaluate the role of DAT imaging in the diagnosis of tremulous disorders by a movement disorder specialist (MDS). Background SPECT imaging of the dopamine transporter, using 123I-FP-CIT, reveals depletion of the nigrostriatal dopaminergic pathway in parkinsonian syndromes. We report imaging outcomes for patients with various tremulous disorders. Methods Fifty-seven consecutive DAT studies requested by a MDS, performed from 2001 to 2005, were compared to patients’ data. The results were analysed according to scan indication and clinical signs. Results Imaging confirmed diagnosis of Parkinson’s disease (PD) in 19/19 patients; of these, a relationship was found between clinical signs and reduced uptake in the contralateral striatum. DAT imaging was abnormal in 8/12 essential tremor (ET) patients who developed features of parkinsonism and were indistinguishable clinically. DAT imaging was helpful for complex movement disorders. It was abnormal in 0/4 patients with orthostatic tremor, 5/6 atypical Parkinsonism, 3/3 Holmes’ tremor, 4/5 with a differential diagnosis of psychogenic tremor or depression versus parkinsonism, 1 with isolated rest tremor and 4/6 with secondary dystonia. Conclusion DAT imaging is valuable where clinical diagnosis is difficult, particularly in ET with features of parkinsonism. In PD DAT imaging reflects the laterality of signs.

ONCOLOGY II A60 18F-FDG PET–CT in patients with newly diagnosed pre-invasive endobronchial lesions I. Kayania, A.M. Grovesa, P.J. Ella, J. Georgeb and J. Bomanjia a Institute of Nuclear Medicine and bDepartment of Respiratory Medicine, University College London, UK. Background The aim of study was to assess the role of combined 18F-fluorodeoxyglucose (FDG PET/CT) in detecting bronchial carcinoma in patients with newly diagnosed bronchial dysplasia as diagnosed on fluorescent biopsy. Methods Patients underwent 18F-FDG PET/CT following the diagnosis of pre-invasive endobronchial lesions (bronchial dysplasia or carcinoma in situ). The degree of uptake of 18F-FDG at the known pre-invasive lesion and at remote sites in the lung and mediastinum was compared with histology. Results Increased uptake of 18F-FDG was detected (peak SUV range 3.6–11) in 4/15 patients at the site of bronchoscopic lesion. Of these 4 patients, 2 had cancer at the site of dysplastic lesion. In the other 2 patients, with uptake at site of dysplastic lesion, there was no evidence of cancer with histology or followup. One patient had disseminated cancer at the site of known dysplastic site and at remote sites. Finally, in 3 of 15 patients there was no uptake at dysplastic site but 18F-FDG PET/CT detected cancer at remote sites. Conclusion This initial study suggests the probability of FDG PET/CT detectable bronchial carcinoma is high in patients with newly diagnosed pre-invasive endobronchial lesions, and therefore FDG PET/CT would appear to be a useful investigation in this cohort of patients.

A61 A comparison of 11C-choline and 18F-FDG for monitoring response of oesophageal cancer to chemotherapy A. Welch, S. Hammonds, L. Schweiger, S. Suttie and K. Park The John Mallard Scottish PET Centre, School of Medical Sciences, University of Aberdeen, UK. FDG PET has been shown by a number of groups to be an effective technique for monitoring the response of a wide variety of tumours to chemotherapy. However, while FDG is a sensitive tracer for imaging tumours, it is also taken up by a range of other cells, such as macrophages whose levels increase as therapy progresses. Therefore, the question remains as to whether a more specific tracer may be more effective at monitoring response. We have previously shown that 11C-choline uptake correlates with cell proliferation in vitro. In this study we compare dynamic FDG and choline imaging in 17 oesophageal cancer patients undergoing chemotherapy. Analysis is performed using both standardized uptake values (SUVs) and compartmental modelling techniques and a number of methods for scaling imagederived input functions and correcting for metabolites are applied and compared. The results show that, while SUV correlates well with metabolic rate, as measured using compartmental analysis, for FDG, the same is not true for choline. The results also show that the rate constants for choline uptake are very sensitive to the metabolite fractions and do not correlate with FDG rate constants; supporting the hypothesis that choline is measuring a different metabolic pathway to FDG.

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A62 FDG PET for prediction of chemotherapy response in primary breast cancer patients G.M. McDermotta, A. Welcha, R.T. Staffb, F.J. Gilbertc, L. Schweigera, S.I.K. Semplec, A.W. Hutcheond, I.D. Millere, I.C. Smithg and S.D. Heysf a School of Medical Sciences, University of Aberdeen, Departments of b Biomedical Physics & Bioengineering, cRadiology, dOncology, ePathology, f Surgery, Aberdeen Royal Infirmary, and gEli Lilly, Surrey, UK.

GUIDELINES AND SERVICE DELIVERY

FDG PET imaging is emerging as an effective technique for predicting the pathologic response of breast tumours to neoadjuvant chemotherapy at an early stage. However, questions remain as to the optimal method to use for quantifying FDG uptake, the optimal point during therapy at which to acquire the images and the efficacy for a large patient population. In this study we address these questions by comparison of a number of FDG uptake parameters in a group of patients for whom images were acquired before therapy (n = 96), after the first cycle (n = 74), at the midpoint (n = 58) and at the endpoint (n = 66) of anthracycline-based chemotherapy. The results show that (1) FDG PET had no predictive power for response when image contrast was low; (2) there was little change in FDG uptake between the midpoint and end of therapy; and (3) no statistically significant improvement in predictive power after the first cycle. The best results were obtained for a population with a pre-therapy tumour to background ratio greater than 5, using the activity distribution in a background region of interest to define the active tumour volume. With this method a sensitivity of 100% and a specificity of 77% were achieved; although several methods produced similar results.

Objectives To standardize the criteria for requesting radiographs to accompany abnormal bone scans, and to extend the requesting task to clinical scientists. Method Previously, in Sheffield, radiologists assessed abnormal bone scans to decide which radiographs were required to aid scan interpretation. However, these radiologists often lacked formal nuclear medicine training and, as a result, inconsistent and inappropriate radiographs were often requested. Therefore, guidelines for radiograph requesting were formalized by two experienced nuclear medicine radiologists in conjunction with nuclear medicine staff. These guidelines were then employed by both radiologists and clinical scientists and audited over a 3 month period (227 scans) to assess their validity. Results Guidelines were easy to follow in 93% of cases. The reporting radionuclide radiologist deemed the radiographs to be appropriate in 92% of cases. Radiographs aided reporting in 87% of cases. Additional radiographs would only have been useful in 4% of cases. Conclusion In the majority of cases, the new guidelines allowed radiologists and clinical scientists to request appropriate radiographs. Further evaluation of a similar patient group, prior to the introduction of the guidelines, is being undertaken to confirm that these guidelines reduce the number of inappropriate radiographs.

A63 The direct use of combined 18F-FDG PET–CT images in radiotherapy treatment planning for patients with non-small-cell lung cancer K.J. Carsona, V.P. Cosgroveb, A. Zatarib, R. Eakinc, J.C. Clarked, D.P. Stewartc, J. McAleesec, L. Flemingc, A.R. Hounsellb and P.H. Jarritta a NI Regional Medical Physics Agency and dDepartment of Radiology, Royal Victoria Hospital, Belfast, and bNI Regional Medical Physics Agency and cDepartment of Clinical Oncology, Belvoir Park Hospital, Belfast, UK. Aim To investigate delineation of target volumes for radiotherapy of non-small-cell lung cancer (NSCLC) patients using 18 F-FDG PET–CT images. Method Ten patients, who had had a recent diagnostic PET– CT scan and were intended for radical radiotherapy, had treatment planning scans performed on a GE Discovery LS PET–CT scanner. The CT data were used to plan the radiotherapy treatment in the normal way, while the PET data was used for research purposes only. Gross tumour volume has been outlined on the CT and PET images, by oncologists with input from a radiologist. Tumours have also been outlined automatically using thresholding. Results In the majority of patients volumes outlined on the PET and CT images are different. It was noted that most of these patients had had chemotherapy prior to their planning scan. Tumours were not always delineated successfully using thresholds determined from phantom experiments. Conclusion It is important to set consistent guidelines for outlining tumour volumes on PET images. Tumour volumes outlined on PET and CT images have been found to be different. If radiotherapy planning PET–CT scans are being performed it will be important to consider timing as these patients often receive chemotherapy before radiotherapy.

A64 Evaluation of new local guidelines for requesting radiographs to accompany radionuclide bone scans D. Raw, V. Prakash, P. Hillel and E. Lorenz Sheffield Teaching Hospitals NHS Foundation Trust, UK.

A65 Is chest radiography triage in the investigation of suspected pulmonary embolism helpful? A. Denison, M.E. Brooks, A.D. Murray and J. Davidson Department of Nuclear Medicine, Aberdeen Royal Infirmary, UK. Introduction Ventilation perfusion (VQ) imaging is recommended as the initial imaging investigation for suspected pulmonary embolism (PE), if chest radiography (CXR) is normal [1]. Some centres proceed directly to CT pulmonary angiography (CTPA) where CXR is abnormal. There is little consensus on what degree of CXR abnormality should prompt CTPA rather than VQ imaging. We investigated the utility of abnormal CXR in VQ imaging. Methods CXR of all consecutive patients referred for VQ imaging over 2 months were assessed by a radiologist before review of scintigraphy. The radiologist judged whether CTPA would have been the preferred first choice investigation (rather than VQ). The VQ result was compared with the CXR comment and with CTPA (where performed). Results Of the 107 VQ cases, 48 had abnormal CXRs. Of these, 10 were considered sufficiently abnormal that CTPA would have been the preferred investigation. One was high probability (HP) of PE, 6 intermediate probability (IP) and 3 low probability (LP). All 6 IP cases had CTPA, which identified PE in 1 patient. Thirty-eight cases had abnormal CXRs but VQ remained the preferred imaging choice. Twenty-two were LP, 7 normal and 9 IP (of which CTPA identified PE in one). CXR was normal in 59 cases: 13 were HP, 4 IP (none of which had PE on CTPA), 11 LP and 31 normal.

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Conclusion The use of CXR appearances in the triage of patients for VQ or CTPA has limited use. Reference 1 British Thoracic Society guidelines for the management of suspected acute pulmonary embolism. Thorax 2003; 58:470–483.

A66 BTS Guidelines: Is compliance compatible with service provision? J.M. James, G. Al-Bahrani and M.C. Prescott Manchester Royal Infirmary, UK. Aim The 2003 BTS guidelines [1] specify appropriate management of suspected pulmonary embolism. The aim of this audit was to evaluate compliance within Central Manchester Trust. Method Referral details and results of lung scans (V/Q) and CTPAs from September 2005 were evaluated against the relevant elements of the BTS guidelines. Results Of 104 V/Qs 2.9% were follow-ups. Referrals frequently failed to document clinical probability or D-dimer but most were performed in timely fashion. CXR was available in over 90%. 11.5% were normal, 43.3% low, 24% (25) intermediate and 8.6% high probability. 9.6% (10) received a cautious report pending repeat CXR. Twelve of 22 CTPA scans followed V/Q. Two revealed subsegmental emboli (1 intermediate probability V/Q). Of 20 negative CTPAs 10 had intermediate and 1 high probability V/Q. No alternative diagnosis was found in 10; 7 with intermediate probability V/Q. Referral for CTPA as initial modality was generally appropriate. Delay between request and imaging was 0–21 days (75% within 3 days). Including patients given a cautious V/Q report 23 patients were not imaged appropriately according to the guidelines. Conclusion BTS guidelines were not followed by all groups. Stringent application of guidelines would require more resources to prevent delay in diagnosis. Reference 1 British Thoracic Society Standards of Care Committee Pulmonary Embolism Guideline Development Group. Thorax 2003; 58:470–484. A67 Can rapid access to PET be achieved with a mobile service? S. Dizdarevica,b and K.A. Milesa,b a Brighton & Sussex University Hospitals NHS Trust, and bBrighton & Sussex Medical School, UK. Aim The National Institute for Clinical Excellence (NICE) has stated that every cancer network must have a rapid access to FDG PET for lung cancer patients (i.e. within 4 weeks). This paper describes an audit of waiting times following establishment of a mobile service in the Sussex Cancer Network (SCN) serving a population of just over 1 million. Methods In June 2004, a PET service comprising 200 scans annually commenced using a mobile PET–CT visiting fortnightly. The service level was based on the workload and case mix previously referred from SCN to a tertiary centre. The SCN produced clinical guidelines recommending PET primarily for patients with lung cancer, lymphoma and recurrent colorectal cancer. The case mix and waiting times were audited after 6 months. Results Seventy-eight percent of scans were performed for patients with the 3 recommended tumour types indicating effective use of guidelines. 83.3% of the 36 examinations for

lung cancer were performed within 4 weeks with the waiting time increasing exponentially over the audit period. Conclusions Despite effective use of clinical guidelines to control workload, rapid access to PET could not be maintained using a fortnightly mobile service. The results of a re-audit following establishment of a weekly service are awaited.

MULTIMODALITY A68 Sentinel lymph node biopsy and ultrasoundguided fine needle aspiration in penile carcinoma patients: Preliminary findings J. Crawshawa, P. Hadwayb, D. Hofflanda, S. Bassinghama, N. Watkinb, J. Pilcherc, S. Heenana and R. Allana Departments of aNuclear Medicine, bUrology and cUltrasound, St George’s Hospital, London, UK. Aim To assess the utility of sentinel lymph node lymphscintigraphy (SLNL) and ultrasound guided FNA in patients with penile carcinoma. Methods Twenty-nine patients aged 28–81 years with penile carcinoma stage (T1 G2 or greater, clinically N0) underwent SLNL. 11–32 MBq 99mTc-nanocolloid were injected intradermally around the tumour or excision scar. Anterior dynamic series of 10 second frames for 15–20 min, followed by anterior and lateral delayed static images acquired. Ultrasonography of both inguinal regions (26/29 patients) with FNA of suspicious nodes (16/52 basins) was performed before skin marking of sentinel lymph nodes (SLNs). Patients underwent same-day radioguided sentinel lymph node biopsy. Results Lymphatic drainage was demonstrated in all patients (26/29 bilaterally). SLNL identified 84/92 (91%) nodes removed at surgery and 71/76 (93%) blue stained nodes. Fourteen of 92 nodes contained metastases (12 nodal basins, 10 patients); SLNL located 13/14 (93%) and all nodal basins. Of 10/12 involved basins scanned, ultrasound correctly identified metastases in 6/10; FNA missed 4/6 (sensitivity of US 60%; positive predictive value 38%) Conclusions SLNL localization of SLN correlates well with surgery. SLNL in penile carcinoma is comparable with SLNL in other cancers. Ultrasound of impalpable inguinal lymph nodes appears not cost effective. A69 Staging of lung cancer by CT, PET and PET/CT in the same patients R.P. Claussa, R. McAvincheyb and M. Illsleyc a Nuclear Medicine Department, Royal Surrey County Hospital, b Radiology Department, East Surrey Hospital, and cOncology Radiotherapy, Royal Surrey County Hospital, UK. Purpose Since the introduction of PET/CT recently, it is not unusual that patients have to be restaged after initial CT staging, once they have undergone a PET/CT scan. The aim of this investigation was to determine the proportion of lung cancer patients that undergo restaging after PET/CT investigation. Method Twenty-two (12 male/10 female) adult patients with clinically diagnosed lung cancer underwent a PET /CT scan as part of their work-up. The CT scan was reported by a radiologist and the PET scan by a nuclear medicine physician. The PET/CT report was a consensus read between the radiologist and the nuclear medicine physician.

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34th BNMS meeting: abstracts

Result When the CT report was compared to the PET report, 9/22 (41%) patients had their staging changed (7 upgrades and 2 downgrades). When the CT report was compared to the PET/ CT consensus read, 8/22 (36%) patients had their staging changed (6 upgrades and 2 downgrades). There were 4/22 (18%) discrepancies between PET and PET/CT reports, resulting 1 upgrade and 3 downgrades after the PET/CT consensus read. Conclusion The PET/CT consensus read appears the best way to stage lung cancer with 36% of patients restaged after the initial CT investigation.

A70 CT scan and 18F-FDG PET in the restaging of lymphoma R. Challa, R. Jayan, C.N. Ramesh and S. Vinjamuri Department of Nuclear Medicine, Royal Liverpool University Hospital, and BUPA Murrayfield Hospital, Wirral, UK.

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Conclusion The foot and ankle reflects a good example of the strengths of functional anatomical imaging as well as the challenges facing this modality.

POSTER PRESENTATIONS P1 Comparison of radioiodine with radioiodine plus lithium in the treatment of hyperthyroidism K. Ahmeda, G.M.K. Nijherb, A. Bannerjeec, J. Frankd and K. Meerane Departments of aEndocrinology and bNuclear Medicine, Charing Cross Hospital, London, UK. Radioactive iodine (RAI) is a well-established and effective treatment for hyperthyroidism. The use of Li as an adjunct to RAI has been postulated, but information on the subject is limited 1,2. Our aim was to evaluate the efficacy of RAI therapy alone and RAI combined with Li. Forty-one patients with hyperthyroidism were randomly assigned to treatment with RAI (controls) or RAI plus Li and evaluated for changes in thyroid function on the day and weeks 1, 3, 9, 12 and at least 6 months post-treatment. Urinary iodine (UI) excretion measurements from 24-h urine collections of 14 Li vs. 15 non-Li patients were obtained at – 7, 0 and + 7 days from therapy. One of 22 patients treated with RAI plus Li (4.5%) and 2 of 19 patients treated with RAI alone (10.5%) were not cured (euthyroid + hypothyroid) (P = NS). There was no significant difference in UI excretion in the Li vs. non-Li group (P = NS). Lithium does not appear to improve the efficacy of RAI. Lack of a significant difference may be due to the excellent cure rate achieved in both groups. A more definitive conclusion will be reached upon greater patient recruitment and completion of the continuing trial.

Purpose The accurate staging of Hodgkin’s and non-Hodgkin’s lymphoma is important in the management of these patients. PET scanning has been shown to play a useful role in assessing response to therapy, for re-staging and to detect suspected recurrence. We retrospectively analysed the role of CT and PET in the restaging of lymphoma post-chemo/radiotherapy Methodology We reviewed 30 patients (14 male, 16 female) with lymphoma referred for PET scanning. All patients had previous imaging with CT and 2 patients had an additional MRI scan. Twenty-three patients had history of previous chemotherapy, 6 patients had both chemotherapy and radiotherapy and 1 patient received an autograft. Results The age range was from 5 to 87 (average, 45.5 years). 18 patients had Hodgkin’s lymphoma, 12 had non-Hodgkin’s lymphoma. Most patients (n = 29) had a residual mass on CT; 1 patient had normal CT but persistent symptoms. PET scanning showed evidence of active disease in 13 patients and no evidence of active disease in 17. PET scanning was found to be complementary in the evaluation and restaging of these patients. Conclusion CT and PET play a complementary role in restaging of lymphoma post-therapy.

References 1 Bal CS, et al. Thyroid 2002; 12:399–405. 2 Bogazzi F, et al. J Endocrinol Metab 1999; 84:499–503.

A71 The role of SPECT/CT in the evaluation of the painful ankle and foot P.A. Fielding, J.I.S. Rees, L.E. Bartley, P.E. Facey and J. Jones Radiology Service Centre, University Hospital of Wales, Cardiff, UK.

P2 Expression of an anti-cd33 single-chain antibody by Pichia pastoris for radioimmunotherapy of acute myeloid leukaemia L.M. Emberson, A.J. Trivett, P.J. Blower and P.J. Nicholls Department of Biosciences, University of Kent, Canterbury, UK.

Background Combined functional anatomical imaging with hybrid SPECT/CT offers a new opportunity to combine functional and anatomical information in a variety of clinical settings. Pain in the foot or ankle may be due to a wide variety of bony or soft tissue disorders. Bone scintigraphy and labelled white cell scans are sensitive in the detection of bony pathology but due to the complex three dimensional anatomy of the ankle and foot, localization of abnormal activity, and therefore differential diagnosis, may be difficult. Methods We examined the value of SPECT/CT in the ankle and foot in a retrospective analysis of patients scanned in our department between September 2003 and September 2005. Results There were 14 studies in 13 patients. SPECT/CT was felt to have been helpful in all of the cases where it was used. The cases are discussed along with, in many cases, correlative images from volumetric CT and MRI. Optimal positioning of the foot (in dorsi or plantar flexion) was found to depend upon the location of probable abnormality and the clinical question.

CD33 is a cell surface glycoprotein expressed on cells of myelomonocytic lineage, leukaemic cells, but not haematopoietic stem cells, and therefore is a popular target for radioimmunotherapy of acute myeloid leukaemia, e.g. with 213 Bi-labelled anti-CD33 antibodies. However, the short halflife of alpha emitters (213Bi, 211At) is not well-matched to the biokinetics of whole antibodies. Therefore, the methylotrophic yeast Pichia pastoris strain KM71H was used to produce an antiCD33 single chain variable fragment (scFv) in shake flask cultures, for conjugation to a therapeutic radionuclide. Flow cytometry and Biacore analysis demonstrated specific binding of the scFv to CD33, both on the surface of human leukaemic cell lines HL-60 and U937, and in soluble form. A stability assay using flow cytometry after incubation of the anti-CD33-scFv in human serum confirmed that no functionality was lost over 10 days at 371C. Cell proliferation/viability and DNA synthesis inhibition studies, performed with the unlabelled scFv, have shown that it has no effect on HL60 or U937 cells, and will serve as a comparison with the radiolabelled scFv. This

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anti-CD33-scFv has demonstrated specific and high affinity binding to target antigen, and warrants further evaluation as a vehicle for targeted radionuclide therapy. P3 Case report: The value of 90Y-DOTA-lanreotide therapy in the management of neuroendocrine tumours S. Khan, Z. Win and A. Al-Nahhas Department of Nuclear Medicine, Hammersmith Hospital, London, UK.

phantom resulted in a basal artefact of 23%, while re-centred 1801 acquisitions resulted in an anterior artefact of 29%. Without re-centring the latter acquisition resulted in a defect of 32% and an overall shift in counts into the lateral wall of the left ventricle. Reference 1 Liu Y, et al. Nucl Med 2002; 43:1115–1124. P6

P4 The design of a gated SPECT cardiac phantom for assessing ejection fraction estimates in MPI A.R. Wilkinsona, I. Beltona and R. Lawsonb a Cumberland Infirmary, Carlisle, and bManchester Royal Infirmary, UK. A gated myocardial phantom was constructed using latex epicardial and endocardial surfaces to investigate the accuracy of ejection fraction measurements in gated SPECT myocardial perfusion processing software. Water was pumped into the phantom by means of a piston operated by a 24 V motor. A basic electronic circuit was built to supply the trigger to the camera incorporating a Hall effect switch. The phantom had an ESV of 99 ± 1 ml, an EDV of 156.2 ± 1.6 ml and an ejection fraction of 36.6% ± 0.9%. Acquisition data obtained using the phantom was processed on two different MPI software packages: ECToolbox and QGS. Studies performed using a typical number of myocardial counts obtained in clinical studies produced an EDV of 95 ± 2.7 ml, an ESV of 58.8 ± 2.0 ml and an ejection fraction of 37.5% ± 2.6% in ECToolbox. Values obtained processing the same data in quantitative gated SPECT (QGS) were 109 ml for EDV, 61 ml for ESV and 43% for ejection fraction. An independent region of interest analysis from raw short-axis slice data produced estimates of EDV = 175.0 ml, ESV = 109.6 ml and an ejection fraction of 37.4%. It would appear that a volume calculation error exists in two of the most commonly used gated myocardial perfusion software packages, ECToolbox and quantitative gated SPECT. P5 Myocardial acquisition and processing parameters: 3601 vs. 1801 and the effect of orbit type, re-centring and reconstruction algorithms M.P. Avison Department of Medical Physics, Bradford Royal Infirmary, UK. Background Two sources of artefacts in myocardial tomography are well documented: attenuation and breathing. Other artefact(s) typically in the antero-apical or antero-septal regions which have been mentioned in the literature, sometimes termed the insertion artefact are less well understood. 1801 acquisitions of non-central organs have been implicated [1]. In this work, the effect of bringing the organ to centre on all projections before reconstruction is investigated. Additionally the effect of reconstruction algorithm and circular or contoured orbits are examined. Method Contoured 1801, circular 1801 and circular 3601 orbit acquisitions were made of an idealized left ventricle phantom placed in 3 orientations. These were reconstructed with backprojection and iterative algorithms, using unmodified and centred pre-processing, giving a total of 36 acquisition, orientation and processing combinations. Results Levels of non-uniform response of similar magnitudes with different acquisition and processing strategies were shown. Circular 3601 acquisitions for the physiologically orientated

Correlation of lung/heart ratio of post-stress Tc-tetrofosmin in MPI with extent of perfusion abnormalities in patients A.K. Paul, A. Singh, Z. Win, J. Murrell, D. Towey and A. Al-Nahhas Department of Nuclear Medicine, Hammersmith Hospital, London, UK.

99m

Aim To assess the significance of lung/heart ratio (LHR) in post-stress 99mTc-tetrofosmin MPI and its correlation with extent of perfusion abnormalities and severity of CAD. Methods We retrospectively studied 48 patients (M:25, F:23; age range 39–82 years) chosen at random, who underwent stress/rest 99mTc-tetrofosmin MPI. Twenty-one patients had adenosine and 27 had added exercise. Nine-segment analysis showed normal perfusion/minimal ischaemia (22), moderate ischaemia (18) and severe ischaemia (8). From the anterior image of the stress SPECT, 2 equal-sized ROIs were drawn in upper left lung and a myocardial segment with maximum count density. LHR = mean counts/pixel (left lung) / mean count/ pixel (myocardium). Results The mean LHR was 0.30 ± 0.03 in normal perfusion/ minimal ischaemia, 0.33 ± 0.05 in moderate ischaemia, and 0.39 ± 0.03 in severe ischaemia. LHR in severe ischaemia was significantly greater than in normal perfusion scans (P = 0.000) and moderate reversible ischaemia (P < 0.001). There was no statistically significant difference between normal perfusion scans and moderate reversible ischaemia (P > 0.05). There were no significant differences between patients who underwent adenosine or adenosine and exercise in all groups. Conclusion Post-stress LHR using 99mTc-tetrofosmin MPI may provide supportive information regarding the extent of perfusion abnormalities and thus the severity of CAD. P7 Assessing function in 99mTc myocardial perfusion scintigraphy: local normal ranges or published values? D.O. Halla and E.J. Owensb Departments of aMedical Physics and Bioengineering, and bRadiology, United Bristol Healthcare NHS Trust, UK. This study was carried out to determine whether the use of published values rather than a local normal range for end systolic volume (ESV) and left ventricular ejection fraction (LVEF) calculated from gated MPS would have a significant effect on whether patients were considered to have normal or abnormal function. Sixty-nine consecutive gated clinical studies (28 normal perfusion), were included. The functional calculations were considered to be abnormal if they had LVEF more than 2 SD below the mean, or ESV more than 2 SD above the mean. These were also compared with a normal cut-off of 45% for LVEF, used for example by the UK DVLA, and a published normal range of 30–55 ml for ESV. The normal values were LVEF mean 73(SD 7)% , and ESV 28(18) ml. For LVEF, 43/69 were normal for both, 8/69 abnormal for both methods, and 18/69 results differed. For ESV, 37/69 were normal for both, 26/69 abnormal for both, and 6/69 results differed. Using published values instead of local normal range would have changed the result for 18/69 (26%) of LVEF values, and for

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6/69 (9%) of ESV values. Results should be quoted with a local normal range.

Conclusion A threshold method applied to TERNA allowed accurate and reproducible measurement of right and left ventricular end systolic and end diastolic volume.

P8 Myocardial perfusion in diabetics with normal coronary arteriograms J. John, G.A. Wright, K. Cockburn, C. Bourantas, G. Avery, G. Davies and A.C. Tweddel Hull and East Yorkshire Hospitals, UK.

P10 Case report: 99mTc-HMPAO labelled leukocyte imaging using planar scintigraphy and SPECT in a coronary artery mycotic aneurysm J. Crawshawa, N. Beharrya, S. Bassinghama, E.E.J. Smithb and

Background Diabetics have been shown to have subtle abnormalities of left ventricular performance which predate large vessel atheromatous disease. Echo colour Doppler studies suggest these may involve the subendocardium with the underlying pathology demonstrated in animal models that of focal endomyocardial necrosis and perivascular fibrosis. Aim To assess myocardial perfusion and function in longstanding diabetics with arteriographically normal coronary arteries. Methods Twenty patients with normal coronary arteriograms, 12 female, underwent 2 day stress/rest gated SPECT technetium perfusion imaging. Images were displayed and reported (1) visually using a 20 segment model, scaled 0–4 (normal–absent) and (2) using QPS/QGS. Results Good quality images were obtained, and overall left ventricular function was normal in all patients. Summed stress score was > 5 in 8 patients (range 0–9) and summed difference score was > 5 in 4 patients (range 0–8). Visually, 6 patients were normal, 2 had ‘reverse redistribution’, 5 were probably abnormal and 7 definitely abnormal. Perfusion deficits were often small, patchy and involved inferior n = 6, lateral n = 4, septum/apex n = 5, and anterior wall n = 8. Summary Abnormalities of myocardial perfusion may occur in diabetic patients with normal coronary arteriograms, reflecting underlying pathophysiological changes prior to large vessel atheromatous disease.

P9 Cardiac volume determination using tomographic equilibrium radionuclide angiography with threshold edge detection I.P. Clements, B.P. Mullan, J.F. Breen and C.G. McGregor Mayo Clinic, USA. Purpose and methods Right (R) and left (L) ventricular (V) endsystolic and end-diastolic volumes (ml) were assessed by electron beam computed tomography (EBCT) and tomographic equilibrium radionuclide angiography (TERNA) in 17 patients. To generate volumes using TERNA, RV and LV regions of interest in systole and diastole were outlined respectively on the horizontal long-axis and short-axis slices and RV and LV edges detected using a threshold method; slices, pixel number and size generated volumes. In a further 18 patients, the TERNA method was validated by comparison with EBCT and reproducibility was assessed. Results Compared to EBCT, volume (combining end diastolic and end systolic) measured by TERNA was similar using a 45% threshold for RV (178 ± 99 vs 175 ± 84) and a 50% threshold for LV (79 ± 47 vs 76 ± 36). In the validation group, EBCT compared to TERNA volume did not differ for the RV (182 ± 80 vs 180 ± 74) or LV (84 ± 43 vs 87 ± 35) with excellent correlations for the RV (r = 0.92) and LV (r = 0.87). Repeated TERNA measurements were highly correlated with low SEE (RV: r = 0.987, SEE = 12 ml) and (LV: r = 0.997, SEE = 3 ml).

N. Buncec Departments of aNuclear Medicine, bSurgery and cCardiology, St George’s Hospital, London, UK. P11 Quantifying driver exposure from patients who have undergone FDG scans C. Ferrisa, M. Girlingb and J. Lowec a Royal Free Hospital, London, bMount Vernon Hospital, Northwood, and cPaul Strickland Scanner Centre, Mount Vernon Hospital, UK. This study examines the factors affecting the radiation exposure from patients undergoing PET and aims to determine how many journeys a driver can perform and remain under the annual exposure limit (0.3 mSv). The dose rates from 28 patients were measured, for different postures, at distances of either 0.5 or 1 m from the sternal notch following injection of FDG. The affect of voiding on patient dose rate was also analysed. Total driver exposure per patient is given by: exposure = Dr  S  T  L, where Dr is the dose rate from a patient standing 1 m from the detector, S corrects Dr for variation in patient posture and distance, T accounts for any delay in departure using the FDG decay curve and L, the integral of the decay curve, accounts for journey time. By dividing the annual limit by the exposure the number of identical trips can be determined as in Table 1. Additional voiding did not reduce the patients’ dose rate. To reduce driver exposure the distance to the patient must be maximized or the departure delayed.

Table 1 Number of trips

Average patient dose rate = 15 lSv  h – 1 Small car (seated 0.5 m from driver) Large car (seated 1 m from driver)

1 h journey

2 h journey

8

4

24

14

P12 Does respiratory effort have any significant influence of the spatial registration of the PET/CT on lung lesion? H.K. Cheow, A. Winship, S. Rankin, D. Landau and M.J. O’Doherty King’s College School of Medicine at Guy’s, King’s and St Thomas’ Hospital, London, UK. Aim The purpose of this study was to evaluate the role of respiratory effort in the PET and CT spatial registration on the lung lesion. Materials and methods A retrospective review of PET/CT performed in patients with suspected lung carcinoma. PET/CT images were acquired using an end expiratory breath hold protocol during CT acquisition of the thorax and free breathing

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during the PET acquisition. Exclusion criteria included lesions with a standard uptake value < 2.5, lesion < 1 cm in its largest diameter and lesions associated with consolidation. Significant image fusion misregistration was considered if there was more than 75% displacement of the lesion by visual assessment on the fused PET image with the CT image. Results Of the 57 patients studied, 10 cases had significant image misregistration. Five of them had lesions in the upper lobe and the other half in the lower lobe. Sixteen patients were unable to comply with the breathing instruction. Only 1 out of 16 (6.3%) patients had significant image misregistration as compared to 9 out of 41 (30.0%) who complied with breathing instruction. Conclusion This study suggested that there was no significant difference between end expiratory protocol and quiet breathing.

acquired with different noise levels. To simulate respiration the phantom was placed on a moving platform, which allows variation of the frequency and amplitude of movement. Results The threshold required depends on lesion size and image contrast. For lesions with volumes greater than 10 ml it is possible to apply a constant threshold value. If the object is moving the threshold required is lower than when it is static. Conclusion Thresholding is a simple technique to apply, but is dependent on the size, contrast and motion of the tumour.

P13 PET/CT in the evaluation of patients with colon carcinoma. Consideration for surgical resection of solitary metastasis in the lung or liver J.C. Hill, J.C. Coffey and R. Kulshrestha Lancashire Teaching Hospitals, UK.

We have evaluated the GE Discovery ST PET–CT scanner for performing high count-rate brain studies for short-lived tracers such as 15O. Brain activity was simulated using a cylindrical phantom with spheres to present contrast between hot and cold regions. A chest phantom was placed outside the field of view to simulate contribution of random and scattered coincidences from the heart. Activity concentrations for the phantoms were calculated from dynamic brain and cardiac PET studies. Images were acquired using an interlaced 2-D and 3-D acquisition protocol and reconstructed using filtered back-projection using the CT for attenuation correction. Noise equivalent count rates in 3-D acquisitions were greater than those in 2-D up to an equivalent whole-body activity of 7 GBq, with the 3-D peak at 5.1 GBq (2-D peak at B20 GBq). Visual quality of the 3-D images was in all cases superior to the 2-D images up to activities several times those likely to be used. Distortions were not seen in the 2-D images. We conclude that, on the Discovery ST, 3-D 15O brain studies provides better quality images than 2D over all relevant ranges of administered activity. A similar evaluation for the body is ongoing.

Method A review of 8 consecutive patients presenting for PET/ CT was undertaken in order to determine whether they are candidates for surgical resection of solitary metastasis presenting in the liver or lung. All patients were considered by conventional imaging techniques to have a single site of disease prior to PET/CT. Summary Seven out of 8 patients had disease other than that recorded on the conventional staging imaging techniques (MRI and CT). Six patients were upstaged; 1 patient was downstaged. There was agreement with prior imaging in one case. A retrospective review of conventional imaging showed no additional features. Conclusion PET/CT is the preferred modality for demonstration of M1 disease in all patients being considered for solitary metastasis resection. The numbers of patients undergoing complex surgical techniques would be reduced and therefore patients could be targeted to the most appropriate therapy such as chemotherapy. This study suggests that metastatic disease is far more advanced than currently appreciated. All patients should have PET/CT prior to surgical procedures.

P14 Determination of target volumes for external beam radiotherapy using 18F-FDG PET–CT images: Static and dynamic phantom experiments A. Zataria,b, K. Carsonb, A. Hounsella, V. Cosgrovea and P. Jarrittb Medical Physics, aBelvoir Park Hospital, and bRoyal Victoria Hospital, Belfast, UK. Aim If 18F-FDG PET–CT images are to be used in radiotherapy treatment planning, consistent techniques for delineating tumours on the PET images must be developed. One such approach is thresholding. The aim of this study was to determine appropriate thresholds for delineating tumours in non-small-cell lung cancer patients, including consideration of the effects of respiration. Method The threshold required to outline objects of different size and contrast was determined using a PET NEMA IEC body phantom scanned on a GE Discovery LS PET–CT scanner. This phantom contains 6 fillable spheres with volumes 0.5 ml to 26.5 ml. The spheres and background were filled with 18F concentrations giving contrasts from 10:1 to 2:1. Images were

P15 Comparison of 2-D and 3-D image quality for high count rate brain studies on the GE discovery ST PET–CT scanner P.J. Schleyer, J.T. Dunn and P.K. Marsden King’s College London School of Medicine at Guy’s, King’s College and St Thomas’ Hospitals, London, UK.

P16 An interpolation method for reconstruction of data from multilayer PET systems J.E. Mackewn, P.J. Schleyer and P.K. Marsden King’s College London School of Medicine at Guy’s, King’s College and St Thomas’ Hospital, London, UK. Small, high resolution, PET systems are currently in demand for novel pre-clinical molecular imaging techniques. For such systems the ratio of the crystal depth to scanner diameter may be much greater than that of a human system, resulting in a dramatic degradation in spatial resolution from the centre to the edge of the field of view. Depth of interaction techniques, such as the use of multiple concentric crystal rings, can be used to reduce this effect. The sampling patterns of such systems, however, are complex and highly non-uniform compared to that of single ring systems and conventional methods of binning the data into a sinogram prior to image reconstruction can not be used. We describe an interpolation technique to produce sinograms, which can be reconstructed using standard reconstruction algorithms, and have validated this by simulating a small (75.5 mm) diameter PET system comprised of 4 concentric rings of crystals. Simulations were performed using the SimSET Monte Carlo package. We have also examined how the method can be used to overcome the undersampling that usually occurs at the centre of the field of view of most PET scanners.

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34th BNMS meeting: abstracts

P17 Case report: Investigation of the role of positron emission tomography in the diagnosis of aortic graft infection following abdominal aortic aneurysm repair: preliminary case studies Z. Wina, S. Khana, A. Singha, A. Al-Nahhasa and J. Frank Departments of Nuclear Medicine, aHammersmith Hospital, and b Charing Cross Hospital, London, UK. P18 18F-FDG PET in the diagnosis and monitoring of vertebral osteomyelitis: a comparison with MRI Z. Wina, L. O’Flynna, A. Singha, S. Khana, E.J. O’Rourkea, G.S. Cookb, J.S. Friedlandb and A. Al-Nahhasa. Departments of aNuclear Medicine, and bInfectious Diseases, Hammersmith Hospital, London, UK. Aim To demonstrate the role of PET in the diagnosis and monitoring of vertebral osteomyelitis, compared to MRI. Method A 47-year-old woman presented with a 2 month history of fever and rigors. The illness began whilst travelling to India where she developed diarrhoea and fever. On return to the UK she developed right sided hip pain and back pain. Salmonella paratyphi A was grown from blood cultures. Results X-ray of the lumbar spine and 99mTc-HMPAO white cell scan were normal. The MRI scan of the lumbar spine 1 month after presentation showed non-specific signal abnormalities in the L5-S1 disc. An 18F-FDG PET performed at the same time, demonstrated avid tracer uptake in the region of the L5S1 disc in keeping with acute infective disciitis/osteomyelitis. Two repeat MRI scans confirmed vertebral osteomyelitis, but showed persistent signal abnormalities even after several weeks. Two further 18F-FDG PET scans showed decreasing SUV of the lumbar lesion and, finally, resolution of the infection after longterm antimicrobial therapy. Conclusion The singular advantage of 18F-FDG PET over MRI is the ability to monitor quantitatively the response to antimicrobial treatment. The SUV is expected to decrease as antimicrobial therapy takes effect as in our case study. P19 Assessment of a simultaneous dual isotope lung perfusion/ventilation technique using 99mTc-MAA and 81m Kr gas R. Gadd, J. Pattison, J.W. Oxtoby, R. Venkannagari, R. Wheat, C. Eustance, S. Dainty and P.J. Mountford University Hospital of North Staffordshire, Stoke-on-Trent, UK. During simultaneous dual isotope lung perfusion/ventilation imaging downscattered events from the 81mKr gas (ventilation) will be recorded in the 99mTc-MAA (perfusion) energy window, resulting in reduced contrast of mismatched perfusion defects. For the simultaneous dual acquisition technique employed this average downscatter was measured to be less than 20%. The significance of this downscatter was evaluated for 50 patients undergoing simultaneous acquisition. For each patient, counts in the smoothed ventilation image corresponding to 20% downscatter were both added to and subtracted from the perfusion image. In this way three sets of images were generated for each patient study: (1) downscatter corrected ( – 20%), (2) no downscatter correction, and (3) artificially increased downscatter ( + 20%). Four experienced observers blindly reported the images and the results were compared to the clinical report generated with knowledge of all available information. As the scatter in the perfusion image decreased there was an overall upgrading of the probability of PE. The effect of scatter correction is to increase the degree of perceived mismatch, i.e., the number of high/intermediate reports increased. To confirm

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the possible benefit of applying scatter correction a prospective study is required comparing dual acquisition with and without scatter correction with independent perfusion/ventilation image acquisition. P20 Data-driven estimation of the attenuation coefficient in lung for attenuation correction of ventilation/perfusion SPECT scans K. Kacperskia, M. Nunezb and B.F. Huttona a Institute of Nuclear Medicine, UCL, London, UK, and bUniversity School of Medical Technology, Montevideo, Uruguay. The density of lung tissue shows a relatively wide range of variability from patient to patient making it unfeasible to assume a universal value of the linear attenuation coefficient. We propose a method of calculating the attenuation coefficient of the lungs based on the emission data only, avoiding the necessity of a separate transmission acquisition. We make use of the fact that for the ventilation/perfusion scans practically all the activity is deposited in the lungs, and its distribution is fairly uniform. The attenuation coefficient of tissues outside lungs is to a good approximation constant for most patients. The outline of the body can be obtained from the scattered photon projections and lungs delineated from the emission data. Then uniform attenuation coefficients in several predefined lung segments can be calculated from the count ratios of carefully chosen projection subset pairs. The proposed technique was evaluated using both the NCAT phantom and patient CT data for activity levels representative of patient studies. The ratios of projection counts were calculated for a range of assigned attenuation coefficients (0.02–0.06 cm – 1) in the lung. For optimally selected projections an accuracy of better than 10% is predicted, sufficient for use in attenuation correction using iterative reconstruction. P21 Pulmonary vein stenosis after radiofrequency (RF) ablation for atrial fibrillation (AF): a clinical and diagnostic conundrum N.A. Kapse, A.R. Wright, N. Peters and R.T. Dhawan Departments of Radiology and Cardiology, St Mary’s Hospital, London, UK. Recent advances in the understanding of the pathophysiology of AF have led to the development of RF catheter techniques aimed at ablating arrhythmogenic foci in pulmonary veins. While this is a potentially curative technique it can result in pulmonary vein stenosis (PVS) in up to one third of those treated. The clinical presentation in these patients is nonspecific; both dyspnoea and haemoptysis can occur mimicking more common conditions such as asthma, pneumonia and pulmonary embolism (PE). This remains an under-recognized clinical syndrome that may lead to misdiagnosis and inappropriate treatment. We present a series of 4 cases with CT imaging demonstrating the post ablation PVS. One of these patients developed haemoptysis and shortness of breath prompting investigations for PE. A ventilation–perfusion (VQ) scan with a lobar mismatch was recognized as false positive with the subsequent CT pulmonary angiography confirming PVS and excluding arterial thrombus. Perfusion lung scintigraphy predictably demonstrates mismatches in haemodynamically significant PVS, which may erroneously lead to a diagnosis of pulmonary embolism given similar clinical presentation. With increasing use of this ablative technique, we, as imagers, need to be aware of this potential pitfall of VQ scanning which can only be recognized if a high index of suspicion is maintained.

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P22 A fast method for calculation of non-uniformity in nuclear medicine images by Fourier transformation in low count density S. Rasaneh, H. Rajabi and M.R. Farsi Nejad Department of Medical Physics, School of Medical Sciences, Tarbiat Modarres University, Tehran, Iran. Purpose A uniformity test is essentially the only required daily QC procedure in nuclear medicine practice. The present methods for the evaluation of uniformity and nonuniformity require many counts and take considerable time. Thus, the methods are not suitable for busy nuclear medicine departments. In this study we used fast Fourier transformation (FFT) methods to distinguish non-uniformity in low counts. Methods and materials Using the Monte Carlo method, uniform and non-uniform simulation flood images of different matrix sizes and counts were generated. The uniformity of the images was calculated using the present (integral and differential uniformity) and Fourier analysis. We use the ratio of highest frequency amplitude of the noise to the average amplitudes for evaluation non-uniformity. Results A simulation study shows that the minimum requires a count density for the calculation integral, and the differential uniformity is 3000 and 9800 counts per pixel but FFT with 400 counts per pixel could perfectly reflect non-uniformities. The time needed for collecting these value of counts was 5 min. Conclusions In a daily uniformity test (low count density), Fourier transformation of flood images was more sensitive and accurate than current methods. Reference 1 Buvat I. Phys Med Biol 1995; 40:1357–1374.

P23 An audit of soft copy display screens used for the reporting and review of images within a nuclear medicine department D.A. Ibbett Derbyshire Royal Infirmary, UK. The introduction of soft copy reporting has added a new risk in the reporting process for nuclear medicine images. Poor display screen performance can lead to compromised reporting and reduced efficiency. To assess the suitability of existing display screens, a qualitative audit of performance has been carried out. The American Association of Physicists in Medicine Task Group 18 has published an extensive report on medical display screen acceptance testing and QC [1]. To keep the audit simple and achievable, within the limited resources of a nuclear medicine service, a reduced set of tests patterns was used. These were displayed using ImageJ [2], to remove any viewer dependence in the results. Amongst other parameters, monitors were defined as acceptable for medical use if they had uniform luminance, no dead pixels, correctly displayed the bit depth test image and both the 5% and 95% contrast squares were visible in the general QC test pattern. Ten LCD, display screens were assessed for the audit. Two failed to meet the standard set for medical use. Soft copy reporting on display screens introduces a new risk into the reporting process. Suitable QC will ensure that medical images continue to be reported correctly and efficiently. References 1 Samei E. Med Phys 2005; 32:1205–1225. 2 http://rsb.info.nih.gov/ij/

P24 A quantitative dual-modality approach to quality control in SPECT L. Livieratosa, R. Fernandeza, M.D.R. Thomasb and D.L. Baileyc a Nuclear Medicine Department, Guy’s and St Thomas’ Hospitals, London, bJoint Department of Physics, Royal Marsden Hospital, Sutton, UK, and cNuclear Medicine Department, Royal North Shore Hospital, Sydney, Australia. Despite rigorous quality control of individual measures of gamma-camera performance, overall image quality in SPECT and assessment of data processing schemes such as image reconstruction remain restricted in qualitative observations. The recent development of intra-modality image registration algorithms and hybrid PET/CT and SPECT/CT systems offers the potential to incorporate high-resolution structural data in the quality control process in emission tomography. We have previously reported [1] on a dual-modality approach to quality control in SPECT and PET. The methodology compares a high-resolution CT-based data set against the experimentally acquired emission data of a phantom to derive an objective measure of spatial resolution. In the present work we demonstrate the application of this method in order to compare the performance of different SPECT systems and in addition to compare various reconstruction schemes in a quantitative manner. This approach proves to be a useful tool in assessing the combined performance of the data acquisition system and image reconstruction scheme.

Reference 1 Thomas MDR, et al. Phys Med Biol 2005; 50:N187–194.

P25 Role of 99mTc-MAG3 renal scintigraphy in detecting cisplatin-induced renal tubular toxicity T. Zehraa, M. Sohaibb, S. Saeedb, A.N. Khanc and R.A. Jafria a Nuclear Medicine, Oncology and Radiotherapy Institute, and bPakistan Institute of Engineering & Applied Sciences, Islamabad, Pakistan, and c North Manchester General Hospital, UK. Cisplatin, a cytotoxic chemotherapeutic agent, is highly toxic to proximal renal tubules in a dose-related and cumulative manner, its major dose-limiting factor. This study was performed to determine the role of dynamic renal scintigraphy using the tubular agent 99mTc-MAG3 in assessing cisplatin nephrotoxicity. Ten patients, diagnosed cases of solid tumours, destined to receive i.v. cisplatin-based chemotherapy, were included in the study. Dynamic renal scintigraphy was carried out twice using 99m Tc-MAG3: first, as a baseline study, and repeated, at an interval of about 6 weeks of first dose of therapy. The images were analysed qualitatively as well as quantitatively, viz. by renogram generation, calculating time to maximum activity (Tmax), time to half of the maximum activity (T1/2max) from renogram, total renal uptake (TRU) determination at 1.5–2.5 min post-injection of radiopharmaceutical by camera based method, and calculation of effective renal plasma flow (ERPF) by the single-sample method [1] on each occasion. Tmax and T1/2max values showed no significant rise before and after cisplatin dose (P = 0.4 and 0.5, respectively). TRU revealed a fall (15.46 ± 10.36%, P = 0.1), though this was less significant than ERPF, i.e., 19.96 ± 5.42%, P = 0.001). It was concluded that the ERPF calculation using 99mTc-MAG3 and a single plasma sample could be used clinically to assess renal tubular toxicity following i.v. administration of cytotoxic drugs [2].

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34th BNMS meeting: abstracts

References 1 Bubeck B. Semin Nucl Med 1993; 23:73–86. 2 Meijer S. Oncology 1983; 40:170–173. P26 Audit of paediatric DMSA scanning: Are we still scanning too many, too prematurely? A local experience N.R. Jefferson, E.J. Owens and D. Dunlop The Royal United Hospital, Bath, UK. Introduction EANM and BNMS guidelines for performing DMSA scans following proven paediatric urinary tract infection exist. To detect scarring, these recommend DMSA scanning no earlier than 6 months after clinical infection. Premature DMSA scanning may give false positive results [1,2]. In general, clinicians are unhappy to wait this long due to concerns about repeated infections delaying imaging indefinitely. We have adopted a pragmatic approach which is to scan no earlier than 3 months after infection. Despite this, clinicians seem to persist in requesting imaging before 3 months have elapsed. Method We undertook a prospective audit over 8 months of 59 referrals for paediatric DMSA scanning. Our questionnaire noted patient age, PMHx, previous radiological studies, total previous infections, last infective episode, prophylactic antibiotic therapy and result of DMSA scan. Results Four cases showed definite scarring. Two reports were inconclusive, having been performed too soon after clinical infection. Fifty-three other cases showed no scarring. Range of time since the previous infection varied from active infection to 15.5 months, with 12% of scans requested before a three month cut-off period. The majority of scans were requested between 3 and 6 months after infection. Nineteen percent were requested 6 months after infection. Conclusion Clinicians continue to ignore well-established guidelines warning against premature DMSA scanning after paediatric urinary tract infection; potentially necessitating repeat imaging, with the associated additional radiation risk. References 1 EANM Guidelines on 99mTc-DMSA Scintigraphy in Children. 2000. 2 BNMS Guidelines on Renal Cortical Scintigraphy. February 2003. P27 Comparison of geometric mean with posterior view renography in the assessment of renal function J.S. Fleming Southampton University Hospitals NHS Trust, UK. Background Renography is used to assess both absolute and relative right to left renal function (RRF). Quantification is usually carried out using posterior images. Errors in RRF may occur if the kidneys are at different depths. Estimation of depth is possible but this is often not very precise. Geometric mean images from combined anterior and posterior views are much less affected by kidney depth and offer the opportunity of reduced errors. Aim To compare the precision of absolute renal uptake and RRF from geometric mean images with that of posterior views only. Methods Simultaneous anterior and posterior MAG3 renography was performed on 20 adult subjects. Each study was analysed twice by the same observer using both geometric mean and posterior analysis. The coefficient of variation (COV) of the difference between repeat measurements was calculated.

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Results The COV for absolute uptake from the posterior view was 4.6% reducing significantly to 2.3% from the geometric mean view (P = 0.02). The COV for the RRF improved similarly from 1.4% to 0.7% (P = 0.02). Conclusion Geometric mean renography results in more repeatable analysis of both absolute uptake and RRF. The largest cause of this difference is the variability in estimating renal depth with the posterior analysis. P28 Scintigraphic renal function evaluation in postradical cystectomy patients: A randomized study comparing direct and antirefluxive uretero-intenstinal anastomosis in orthotopic ileal neobladder H. Gad, M. Abdel-Latif , A. Musbah and A. Shaban Urology and Nephrology Center, Mansoura University, Egypt. Objectives (1) To assess the reliability of dynamic scintigraphy in renal function evaluation and in the diagnosis of urinary obstruction. (2) To examine the possible benefit of using an antireflux system in patients with orthotopic ileal neobladders. Patients and methods Between January 2002 and March 2004, 60 patients (53 men and 7 women, mean age ± SD, 52.7 ± 7.3 years; range, 31–68) who were candidates for orthotopic neobladders were selected. Preoperative evaluation included IVU, cystoscopic biopsy and radioisotope renography. Renography was done by injecting 111 MBq; of 99mTc-DTPA i.v. and data analysis included, relative split function and differential GFR. The ureters were randomized to either a direct anastomosis into a 5-cm ileal chimney on one side or to be implanted using the antireflux serous-lined extramural tunnel on the contralateral side in the same patient. Regular follow-up included IVU and radioisotope renography every 6 months in cancer-free patients. Results Mean follow up period ± SD was 23 ± 9.6 months, range 6–38 months. Six ureters had developed early anastomotic strictures (1 direct and 5 anti-refluxing), and were treated. Renographic studies were within normal except in cases developed anastomotic strictures where there was reduction in function. Data were comparable in both groups. Mean GFRs were 55.1, 50.7, 49.4, 52.2 and 53.9 ml  min – 1 for direct side and 56.1, 53, 52.4, 53.2 and 50.4 for renal units with anti-refluxing implantation at preoperatively, 6, 12, 18 and 24 months after cystectomy, respectively. Significant deterioration of GFRs was observed due to anastomotic strictures; from 48.6 ± 6.7 ml  min – 1 preoperatively to 31.8 ± 15.9 ml  min – 1 after revision (P = 0.01). Conclusion Dynamic scintigraphic renography is the ‘gold standard’ radiological tool for renal function evaluation and diagnosis of urinary obstruction. Anti-reflux procedures were associated with a relatively higher incidence of anastomotic strictures than direct methods in which significant deterioration in renal function was observed. Long-term follow-up is awaited. P29 Effect of recent BNMS guidelines on paediatric GFR and carboplatin dosing M. Attheya and J.J. Lloydb a Newcastle General Hospital, and bRoyal Victoria Infirmary, Newcastle upon Tyne, UK. Purpose GFR may be measured using a slope–intercept calculation applied to 51Cr-EDTA clearance measurements. The effect of implementing recent BNMS guidelines on GFR measurement and subsequent carboplatin dosing for children was investigated in a retrospective study. The technique at this

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Table 1 Dementia SPM Defect Precuneus Medial temp. Either

Lewy body dementia Whole-brain

Late-onset Alzheimer’s disease Masked-brain

Whole-brain

Masked-brain

Sens

Spec

Sens

Spec

Sens

Spec

Sens

Spec

73% 7% 73%

100% 100% 100%

87% 53% 93%

94% 94% 90%

59% 55% 81%

100% 100% 100%

80% 71% 96%

94% 94% 90%

centre uses 1, 2, 3 and 4 h samples without fast-exponential correction as this was the method used at the time of derivation of the carboplatin dosing formula [1]. Proposed changes at this centre include omission of the 1 h sample and addition of rapid exponential clearance correction. Patients and methods 51Cr-EDTA clearance measurements for 65 children (mean age 8.6 years, range 0.1–17.8 years) were used to calculate GFR and T1/2 using the original and BNMS recommended methods. Carboplatin doses were calculated using T1/2 as favoured by UK Children’s Cancer Study Group [1]. Results: Applying these changes gave mean changes (SD) of T12 = + 5.8% (6.3%), GFR = – 18.9% (6.1%), carboplatin dose = – 7.1% (12.9%). Conclusion Inclusion of a 1 h sample leads to an underestimate of T12 which increases carboplatin doses in children as the preferred dosing formula uses T12 rather than GFR. Changes in dose would be higher if GFR were used.

The aim was to assess the effect of the accuracy of SPM to detect abnormalities in patients with mild dementia by performing the analyses on critical areas of the brain. The data consists of 90 images: 44 late-onset Alzheimer’s disease (70–92 years), 15 Lewy body dementia, LBD, (61–86 years), the latter confirmed by abnormal DATscans, and 31 volunteer controls (40–83 years). Medial temporal lobes and precuneii were identified from the Talairach atlas and used to mask the images. SPM was run for both whole brain and masked patient images: each compared with the group of control images. The specificity of SPM was determined using the leaveone-out method on the control group. Results are given in Table 1. The sensitivity increases for both patient groups when both regions are selectively analysed, however the reduction in specificity must be considered. We believe these important findings warrant a prospective trial to assess the efficacy of HMPAO SPECT imaging with SPM interpretation in clinical practice.

Reference 1 Newell DR, et al. J Clin Oncol 1993; 12:2314–2323. P30 Assessment of error in 51Cr-EDTA plasma clearance estimation associated with the number and timing of blood samples D.N. Strouda, E.A. Hockadaya, N.S. Kennedya and S.A. Ogstonb a Ninewells Hospital and Medical School, and bUniversity of Dundee, Section of Public Health, UK.

P32 201Tl SPECT for assessing chemotherapy response in gliomatous brain tumours R. Jayana, C. Ramesha, P. Warnkeb and S. Vinjamuria Departments of aNuclear Medicine, Royal Liverpool University Hospital, and bNeurosurgery, Walton Neurocentre, Liverpool, UK.

Guidelines adopted by the British Nuclear Medicine Society recommends blood samples for estimation of a patient GFR be derived from 2, 3 or 4 blood samples taken between 2 and 5 h following a single injection of a suitable radiopharmaceutical. The ‘slope–intercept used for the analysis relies on regression parameters derived from these blood samples. In this work we examine sources of error based on the number and timing of plasma samples taken for the 51Cr-ethylenediaminetetraacetic acid (51Cr-EDTA) plasma clearance. At our centre 4 or 5 blood samples are taken after injection. Both a computer simulation and a database of 3147 test results have been used to observe the dispersion of each GFR using the number and timing of 2, 3, 4 and 5 samples. Fitted regression parameters are evaluated with the standard error and the coefficient of correlation. Results show that in some cases a single GFR test can vary by more than 10% and by several millilitres per minute based simply on the number of points used in the calculation and by the timing of blood samples. In conclusion, a means of evaluating the quality of the test results is a necessary component of the GFR study.

Aim To evaluate the role of 201Tl brain SPECT in assessing metabolic response to chemotherapy with procarbazine, cyclophosphamide and vincristine (PCV). Material and methods We assessed the brain scans of 14 patients who were scanned before and after receiving 1 or more cycles of PCV chemotherapy. Visual analysis and semi-quantitative analysis of the thallium scans were performed by calculating tumour to contralateral hemisphere background thallium uptake ratio (Tl index). Results Out of 14 patients (8 males, 6 females), 9 belonged to WHO histological classification grade 2 and 5 to WHO grade 3. In 12 out of 14 patients, a decrease in thallium index ratio was seen. In only 3 of these was the decrease significant (reduction in thallium index > 50%) according to the modified McDonald’s criteria (reduction in thallium index by 56, 76 and 72%). Ten patients were classified as having stable disease by their thallium scan. One patient was shown to have progressive increase in thallium index which reflects increased metabolic activity and likely poor response to chemotherapy. Conclusion 201Tl SPECT appears to have a useful role in assessing metabolic response of brain tumours to PCV chemotherapy.

P31 Improved dementia diagnosis by targeting critical brain regions in SPM statistical analyses of HMPAO brain SPECT S.M.A. Hoffmanna, T. Warda and P.M. Kempb Departments of aMedical Physics & Bioengineering, and bNuclear Medicine, Southampton General Hospital, UK.

P33 Impact of camera and reconstruction algorithm on observer and quantitative assessment of DaTSCAN imaging J.C. Dicksona, M.D. Aldridgea and B.F. Huttonb Institutes of Nuclear Medicine, aMiddlesex Hospital, London, and b University College London, UK.

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34th BNMS meeting: abstracts 307

After several years of successful use, many sites are now intending to continue imaging DaTSCAN using new gamma cameras and reconstruction algorithms. The aim of this paper was to assess the impact of such a change on observer impressions, and quantification, including ‘normal’ ranges. Thirty-one patients consented to be imaged consecutively on a dual-headed and triple-headed gamma camera with parallel hole and fan beam collimators, respectively. Parallel hole data were reconstructed using FBP, and fan beam using OSEM with correction for scatter and attenuation. Data were assessed by 4 observers using a 6point scale and by a fully automatic method of quantification. Observer agreement between systems (k = 0.78) was similar to that between observers on the same system (k = 0.80). There was also a good correlation of the quantitative striatal uptake measure between systems (r2 = 0.89). Existing normal ranges applied after regression analysis (y = 1.37x + 0.2; SEE = 0.33) demonstrated agreement in quantitative scan interpretation in 89% of cases. Agreement in visual interpretation between two very different systems and reconstruction algorithms was good, and once the required conversion of quantitative measures had been performed, this type of VOI analysis gave results adequate for continuing longitudinal studies. P34 Case report: Bone scintigraphy in the evaluation of mandibular condylar hyperplasia A. Anilkumar and R.H. Ganatra Department of Radiology, Queen’s Medical Centre, Nottingham, UK. P35 Case report: Pictorial review of non-skeletal bone scan abnormalities N.R. Jefferson and D. Dunlop Royal United Hospital, Bath, UK. P36 Diagnosis of prosthetic joint infections: Role of Tc-sulesomab K. Iyengar, R. Jayan, C. Ramesh and S. Vinjamuri Department of Nuclear Medicine, Royal Liverpool University Hospital, UK.

99m

Aim To assess the role of 99mTc-labelled anti-granulocyte monoclonal antibody Fab0 fragment (sulesomab) in the diagnosis of prosthetic joint infections. Material and methods A retrospective analysis of 38 patients (18M:20F) over an age range of 54 to 89 years, with suspected prosthetic joint infections. The scintigraphic diagnosis done with 99mTc-sulesomab were compared with the final clinical diagnosis and information collected from routine blood tests, plain radiographs, appropriate microbiology, culture and histology. The final diagnosis was determined by conclusive microbiology, culture and/or histology, intra-operative findings, aspiration, complementary investigations like CT/MRI and long-term clinical follow-up. The findings of 99mTc-sulesomab images were compared with the clinical outcome to arrive at the decision of true positive/false positive/true negative/false negative results. Using the above definitions sensitivity, specificity and diagnostic accuracy of 99mTc-sulesomab for suspected prosthetic joint infection was calculated. Results Outcome classification revealed 10 true positives, 22 true negatives, 5 false positives and 1 false negative. The overall sensitivity was 90.90% and specificity 81.48% with a negative predictive value of 95.65%. Conclusion 99mTc-sulesomab proved to be useful in excluding prosthetic joint infection than confirming it with a high negative predictive value of 95.65%.

P37 Efficacy of 99mTc-sulesomab in the diagnosis of long bone infections K. Iyengar, R. Jayan, C. Ramesh and S. Vinjamuri Department of Nuclear Medicine, Royal Liverpool University Hospital, UK. Aim To assess the role of 99mTc-labelled anti granulocyte monoclonal antibody (sulesomab) in the diagnosis of long bone infections. Material and methods A retrospective analysis of 32 patients (17M:15F) over an age range of 21–86, with suspected long bone infection. The scintigraphic diagnosis with 650 MBq of 99m Tc-sulesomab was compared with the final clinical diagnosis. The final diagnosis was determined by conclusive microbiology, intra-operative findings, aspiration, CT/MRI and long term clinical follow-up. The findings of 99mTc-sulesomab images were compared with the clinical outcome to arrive at the decision of true positive/false positive/true negative/false negative results. Using the above definitions sensitivity, specificity and diagnostic accuracy of 99mTc-sulesomab for suspected appendicular infection was calculated. Results Outcome classification revealed 12 true positives, 19 true negatives, 1 false positive and no false negative results. The overall sensitivity was 100%, specificity of 95%, diagnostic accuracy of 97% with a negative predictive value of 100%. The individual sensitivity and specificity of each category were compared. Conclusion 99mTc-sulesomab seems to have a high efficacy in the diagnosis of long bone infections. With a high negative predictive value, 99mTc-sulesomab scintigraphy excluded orthopaedic infection with greater efficacy.

P38 Case report: branching (atypical) Paget’s disease of bone G. Gnanasegaran, A.E.B. Moore, G.M. Blake, S. Vijayanathan, S.E.M. Clarke and I. Fogelman Guy’s and St Thomas’ Hospital NHS Foundation Trust, Guy’s, King’s and St Thomas’ School of Medicine, London, UK.

P39 Bone mineral density in normal children D.E. Simpson, S. Stevens, V.S. Dontu, V. Lowe, L.J. Archbold, H. Bashir, M.J. O’Doherty, N. Martin and A.J. Coakley East Kent Hospitals NHS Trust, Kent and Canterbury Hospital, UK. Our centre has acquired bone mineral density data from 453 children between the ages of 5 and 17. The protocol involves hip, spine and whole body scanning with a Hologic QDR 4500W, a physical examination including pubertal staging and the completion of dietary questionnaires. These children have been recruited from local schools. In accordance with a preliminary analysis of this data, those children younger than 11 have had their spine and whole-body scans analysed using paediatric algorithms. Those above 11 have used the adult algorithm. The results are given in Table 1 and 2. The physical examination allows these data to be presented as a function of development as well as age.

P40 Facts and artefacts in dual energy X-ray absortiometry G. Gnanasegaran, G.M. Blake, F.M. Crane, D. Dulnoan, S.E.M. Clarke and I. Fogelman Department of Nuclear Medicine, Guy’s and St. Thomas Hospital NHS Trust, London, UK.

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308 Nuclear Medicine Communications 2006, Vol 27 No 3

Table 1

Results for boys

Age range (years)

Number

Spine (L1–L4) Mean (g  cm – 2)

SD (g  cm – 2)

Mean (g  cm – 2)

SD (g  cm – 2)

0.506 0.561 0.569 0.656 0.809 0.900 0.990

0.046 0.053 0.046 0.061 0.076 0.165 0.166

0.602 0.708 0.737 0.782 0.934 0.998 1.194

0.082 0.067 0.076 0.084 0.127 0.170 0.115

<7 7–8 9–10 11–12 13–14 15–16 17

31 38 38 18 19 15 7

Table 2

Results for girls

Age range (years)

<7 7–8 9–10 11–12 13–14 15–16 17

Number

49 44 52 48 36 38 20

Hip (total hip)

Spine (L1–L4)

Hip (total hip)

Mean (g  cm – 2)

SD (g  cm – 2)

Mean (g  cm – 2)

SD (g  cm – 2)

0.515 0.537 0.583 0.736 0.857 0.966 0.999

0.042 0.053 0.068 0.083 0.107 0.075 0.120

0.589 0.645 0.682 0.778 0.874 0.936 0.941

0.058 0.063 0.084 0.098 0.125 0.085 0.143

Osteoporosis is a progressive disorder characterized by low bone mass, leading to increased bone fragility and fracture risk. Bone mineral density (BMD) is measured by dual energy X-ray absortiometry (DXA) scanning. Bone densitometry is clinically established for the diagnosis of osteoporosis and also for determining fracture risk. DXA scans are widely used to diagnose osteoporosis, to assist making decisions in treatment, and follow-up response to therapy. The advantages of DXA include low radiation dose to patients, short scan times and good precision. However, DXA has limitations, including artefacts such as degenerative disease or metallic implants, which may confound BMD results. We present a review of variants and artefacts that affect DXA scans. This pictorial consists of various interesting artefacts that adversely affect the BMD measurements. The artefacts include degenerative disease, patient movement, scoliosis, aortic calcification, CT scan contrast, bone metastases, post implant, post-surgery, navel ring, jeans pocket stud, braces/suspenders and under-wired bra. Familiarity with pitfalls, variants and recognition of artefacts will lead to better interpretation without erroneous results. Factors like patient positioning, scan acquisition mode, quality and data analysis are also crucial for correct interpretation of the findings. Taking a brief history will help in anticipating/recognizing these artefacts and also in careful scrutiny of DXA scan images. P41 Case report: Bone scintigraphy and the pelvis: Pathology and pitfalls I. Hagan and D. Dunlop Royal United Hospital, Bath, UK.

Methods The case notes of 24 patients (18 female, 6 male; age 15–64; mean, 41 years) who had HIG scintigraphy because of inconclusive clinical features of active synovitis were retrospectively reviewed. Prior to scintigraphy 11 patients were on a disease modifying anti-rheumatic drug (DMARD); and 13 patients were on a NSAID. Findings HIG scintigraphy was normal in all 13 patients without DMARDs. The negative scan result correlated well with blood parameters, X-ray, musculoskeletal ultrasound and final clinical diagnoses. No patient developed an erosive arthropathy and none required DMARDs subsequently. In the patients already on DMARDs, synovial HIG uptake was negative in 7 and positive in 4 patients. DMARDs were stopped in 2 patients following negative HIG scintigraphy but were continued in 5 patients because of the clinical improvement. All 4 patients with positive scintigraphy developed joint erosions, recurrent flare-ups and required long-term DMARDs. Conclusion Normal HIG scintigraphy excludes active inflammatory arthritis reliably whereas positive HIG scintigraphy can be associated with active joint inflammation and subsequent erosive arthropathy. P43 Case report: Characteristic appearances of non-malignant areas of increased radiotracer uptake on bone scintigraphy – A review C. Cronin, D. Lohan, N. Gough, E. Delappe and D. O’Keeffe Department of Radiology, University College Hospital, Galway, Ireland. P44 How good is scintimammography: A metaanalysis R. Hussain and J.R. Buscombe Department of Nuclear Medicine, Royal Free Hospital, London, UK. Aim To review clincial trials using scintimammography to determine its utility in both single centre or muti-centre trials. Methods Using PUBMED trials of scintimammography were identified after 1997 when the technique had become mature. The results of 10 trials involving a total of 1489 patients in single centre trials was then compared with a total of 1192 patients in three prospective multicentre trials. The overall sensitivity, specificity and where possible the PPV and NPV were calculated. To prevent re-counting of patients appearing in multiple studies the most recent paper from each study centre was considered. Results The results from single centres were better than those from multi-centre trials with a sensitivty of 87% for single centres compared with 78% in the multicentre trials, specificity was 87% vs. 74% for the multi-centre trials. The PPV was 84% for single sites and 78% for multicentre trials the NPV was likewise 85% vs. 87% for the multicentre trial sites Conclusion As expected, single site research produced better results than multi-centre trials however the accuracy of scintimammography remained acceptable in both types of trial.

P42 How reliable is 99mTc human polyclonal immunoglobulin (HIG) scintigraphy in inflammatory arthropathy? S. Han, J.B. Neilly, F.W. Poon and H.W. Gray Department of Nuclear Medicine, The Royal Infirmary, Glasgow, UK.

P45 Lymphoscintigraphy in lower extremity lymphoedema: standardizing interpretation of lymphoscintigrams A. Singh, W.E. Svensson, D.J. Towey, J. Murrell, K.S. Nijran and A. Al-Nahhas Hammersmith Hospitals NHS Trust, London, UK.

Aim To review the impact of HIG scintigraphy in the differential diagnosis and management of inflammatory arthritis.

Purpose Lymphoscintigraphy is used for diagnosing lymphoedema [1,2], but no effective standards for interpretation have

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34th BNMS meeting: abstracts 309

been established. We attempt to standardize the interpretation of lymphoscintigrams combining qualitative and quantitative assessment. Methods and materials Re-analysis of 99mTc-Nanocoll lymphoscintigrams was performed in 13 patients (7M 6F, age range 18–74) for 26 lower extremities (21 oedematous, 5 nonoedematous). For all patients, images were acquired from 0–120 min, clinical records were reviewed, and visual assessment of lymphoscintigrams was performed using summed lymphatic score (SLS). Percentages of inguino-pelvic node tracer uptake at 120 min and tracer clearance from injection sites were calculated. Statistical analysis was performed to compare data. Results Lymphoedema was established in 16 (11 primary, 5 secondary) and excluded in 5 oedematous extremities. Lymphatic function of 5 non-oedematous extremities was also analysed. For SLS sensitivity, specificity, PPV, NPV and accuracy were 71.4%, 40%, 83.3%, 25% and 80.76%; and for tracer clearance from injection site 61.9%, 40%, 81.25%, 20% and 80.76%, respectively. Correlation of clinical assessment and follow-up with combined qualitative and quantitative assessment of lymphoscintigrams showed improvement in management of patients. Conclusion Our findings suggest that interpretation of lymphoscintigrams maybe standardized using a combination of visual grading and measure of tracer clearance from injection sites. References 1 Szuba A, et al. Vasc Med 1998; 3:145–156. 2 Bull RH, et al. Lancet1993; 341:403–404.

P46 A Dedicated multidisciplinary one stop FNA thyroid clinic: Does it help? S. Dizdarevica,b, R.S. Rathinaezhila, A. Williamsa, C. Zammita, H.K. Mohana, E. Pottera, K.A. Milesa,b and A.M. Petersa,b Brighton & Sussex aUniversity Hospitals NHS Trust, and bMedical School, Sussex, UK. Aim To determine the reduction in insufficient fine-needle aspirates through establishing a multidisciplinary one-stop FNA thyroid clinic. Methods In 2004, the clinic team comprising a nuclear medicine consultant, cytologist and endocrine surgeon saw 93 patients. Preliminary work-up included TFTs, thyroid antibodies, ultrasound and imaging with 99mTc-pertechnetate and MIBI. The rate of inadequate thyroid FNA was compared to that of the previous year. Results The inadequacy thyroid FNA in 2003 was 42% resulting in several outpatient visits for diagnosis. Following establishment of the FNA clinic, this rate fell significantly to 26% (95% CI 17–34.5%, P < 0.02) enabling diagnosis at the first visit in 91% of patients. Sixty-seven percent were discharged, 24% underwent surgery and 9% required follow-up. Mismatched pertechnetate/MIBI uptake was present in 6 of 26 patients undergoing 99mTc-pertechnetate/MIBI thyroid imaging. Two of these 6 had a neoplasm on final histology, one of which had been negative on FNA. All 20 patients with negative MIBI thyroid scan (no mismatch MIBI/pertechnetate) uptake had benign disease. Conclusion Establishment of a dedicated thyroid FNA service resulted in a reduction in the inadequate FNA rate and outpatient visits and is likely to be cost effective. Pertechnetate/MIBI imaging contributed to decision-making.

P47 Son of CuATSM: towards optimized secondgeneration copper complexes for hypoxia imaging R.L. Paula,b, J. Dearlingc, P. Halstedb, J. Ballingerb, J.S. Lewisd, K. Martina, P. McQuaded, P.K. Marsdenb, M.J. O’Dohertyb and P.J. Blowera a University of Kent, Canterbury, UK, bGuy’s, King’s, St Thomas’ School of Medicine, London, UK, cRoyal Free Hospital, London, UK, and d Washington University, St Louis, USA. Tumour hypoxia contributes to failure to respond to radiotherapy in cancer patients. Hypoxia imaging may be of value in radiotherapy planning, both to predict response and to identify hypoxic regions within tumours requiring higher radiation doses. The hypoxia imaging agent Cu-diacetylbis(N4-methylthiosemicarbazone) (CuATSM) is selectively taken up in hypoxic cells. Imaging studies in animals and cancer patients have demonstrated that this behaviour is manifested in vivo. Analogues of CuATSM have been synthesized and in vitro studies show that hypoxia selectivity is a general property of many of them, allowing modification of the pharmacokinetic properties of the complexes while retaining control over hypoxia selectivity. We report that a new complex, CuATSE (Cu-diacetyl-bis(N4-ethylthiosemicarbazone), is hypoxia selective in vitro (EMT6 rat mammary carcinoma cells) but is taken up in cells with less extreme hypoxia (38 mmHg) than CuATSM (1 mmHg, a level much lower than ‘radiobiological hypoxia’). CuATSE also shows higher tumour uptake and lower liver and kidney uptake than CuATSM. This control of pharmacokinetic properties affords a route to development of second-generation hypoxia imaging agents with improved imaging properties. We therefore report the design and implementation of a novel target handling system for semi-automated production of 64Cu (and 60Cu) on an 11 MeV cyclotron, in quantities suitable for clinical studies.

P48 Oesohageal transit in gastro-oesophageal reflux disease (GORD) K. Harding, A. Hawash, N. Smith and R. Spychal Departments of Surgery and Nuclear Medicine, City Hospital, Birmingham, UK. Background Swallowing function after anti-reflux surgery for GORD is an indicator of the quality of surgery. Methods We have examined oesophageal transit (OT) using condensed images in 14 subjects with GORD both before and after anti-reflux surgery. On each visit the fasting subject performed 9 consecutive swallows of 3 liquid, 3 semi-solid (scrambled egg) and 3 solid (toast) meals. They were asked in addition to perform dry swallows every 30 s. Each meal was labelled with 40 MBq 99mTc-DTP. Results Results were generally reproducible between swallows in the same subject with a liquid transit time (TT) under 20 s and a semi-solid TT of 11–42 s. Solid swallows were more variable both before and after surgery: 4 subjects had a solid TT in excess of 100 s, with the remaining subjects being between 13–82 s. Repeat swallows had most variation with solid swallows, which correlates with the variability of clinical reporting of swallowing function in GORD. Conclusion Oesophageal transit measurement in GORD must be reported with respect to both the constituency of the meal swallowed and whether subjects have undergone surgery.

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310 Nuclear Medicine Communications 2006, Vol 27 No 3

P49 Sentinel node biopsy in malignant melanoma P.J. Ryan, F. Sagias, R. Lindley, L. Shaal and O. Khan Medway Maritime Hospital, Kent, UK. Sentinel node biopsy (SNB) for malignant melanoma in the UK remains controversial. This study examined the prognostic value of SNB in a district hospital setting. One hundred and sixteen patients of average age 54 years with malignant melanoma of the limbs and torso, Breslow thickness 0.7 mm to 4 mm, have undergone SNB. Mean follow-up was 34 months. Patients were examined with 4 or 6 intradermal injections of Tc-Nanocoll using a same day procedure (20 MBq) with dynamic and delayed 3 h images, and marking of nodes. Sentinel nodes were excised using probe assisted surgery. Sixteen patients (13.8%) were SNB positive and underwent block dissection, where a further 44 lymph nodes were positive. Intransit metastases were found in 5.1% of patients (SNB negative). Distant metastases developed in 8.6%. Among the SNB positive patients, 6 out of 16 (37.5%) died within 25 months (mean survival 22.35). Ninety-seven percent of SNB negative patients were still alive (mean survival 34 months). There was minimal morbidity with 9% seroma, 2 % wound infection. Our results confirm a far better prognosis for SNB negative compared to SNB positive patients and support the use of SNB to provide prognostic information in routine practice. P50 Comparison of the use of lead shielding and image count truncation for visualization of nodes in sentinel node imaging S. Russell, W.H. Thomson and J. O’Brien City Hospital, Birmingham, UK.

Aim Sentinel nodes have < 1% injected activity and are close to the injection site. The injection site maximum count and scatter can mask the nodes on images. Some departments position lead shields over the injection site. However, it is also possible to alter the image display scaling by directly changing the maximum count value in the image header (image truncation). We have used a phantom to compare these approaches for visualisation of small nodes. Method A high activity source simulating injection and a low activity object simulating nodes were placed at 8 cm depth in a water tank. The distance between the sources was varied from 5 cm to 1 cm. At each position an image was acquired with and without a lead shield over the high activity source. Images without the shield were truncated to display the nodes. Results At all separations the nodes were visible on the lead shielded and truncated images. Count profiles showed no significant differences in the scatter produced with and without the shield. Conclusion Shielding the injection site gives no advantage over image count truncation for node visualisation. Since the placement of the lead shield can be problematic, image truncation is the recommended display method.

P51 Is scatter correction in clinical I-123 MIBG images using a high-sensitivity collimator comparable with images acquired with a medium-energy collimator? Fletcher AM, Motherwell DW, McCurrach G, Small AD, Martin W, Cobbe SM Departments of Nuclear Cardiology and Medical Cardiology, Glasgow Royal Infirmary. Background In addition to the prime photopeak at 159 keV, I-123 also emits high-energy photons at 529 keV. Septal

penetration of these high-energy photons causes degradation of the images when using a low energy high-sensitivity (LEHS) collimator. Myocardial I-123 MIBG uptake is commonly measured using the heart to mediastinal uptake ratio (H/M). The aim of this study was to investigate the variability in the determination of H/M ratios from images obtained using a medium-energy (ME) collimator, a LEHS collimator and a LEHS collimator with scatter correction. Method Images were obtained from 20 patients with clinical evidence of chronic heart failure. Regions of interest were drawn around the myocardium and the mediastinum by two independent observers. Results There was no difference in H/M ratios when using the ME or LEHS with scatter correction compared to the LEHS images (P < 0.001 in both instances). There was no difference in H/M ratios between the ME and LEHS collimator. Additionally there was no significant difference between quantitative uptake measurements between two observers using the LEHS collimator with scatter correction. Conclusion The use of a LEHS collimator combined with scatter correction is adequate for quantitative analysis of clinical I-123 MIBG images.

TECHNOLOGIST POSTERS

T1 Evaluating the impact of leaked 99mTc-DTPA from two lung ventilation systems I.D. Vrettos and P.J. O’Sullivan Royal Berkshire Hospital NHS Trust, Reading, UK.

The aim of this investigation was to estimate the impact of 99mTc-DTPA leaked into the camera room from two lung ventilation systems (Venticis and SmartVent). Routine environmental monitoring had shown that the air conditioning unit could concentrate 99mTc-DTPA after VQ scanning. The activity concentrated within the air conditioning unit was monitored for 6 months using the Venticis system and 8 months using the SmartVent system. The results showed a reduction in the average cps above background per VQ examination of over 30% in the SmartVent over the Venticis results. The amount of concentrated activity was significant enough to change the designation of the room from supervised to controlled on 4 occasions in 6 months with the Venticis system and 2 occasions in 8 months with the SmartVent system. With the 2 systems tested the air conditioning counts were lower on average with the SmartVent system. This fact supported the department’s decision to change to the SmartVent ventilation system. This investigation concludes that daily monitoring of the air conditioning unit is vital for departments using 99mTc-DTPA for lung ventilation studies.

T2 Does dose matter in breast sentinel node imaging? A. Burns, R.F. Bury, H.E. Wilson and K. Horgan Leeds General Infirmary, UK. Background In October 2004 the nuclear medicine department at the Leeds General Infirmary introduced lymphoscintigraphic

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34th BNMS meeting: abstracts 311

localization of sentinel nodes in breast cancer patients. Measurements confirmed that, due to the low volume used, actual injected activity was considerably less than that intended. This did not adversely affect the success of the technique. Introduction Sentinel lymph node biopsy (SLNB) is a reliable method for staging axillary lymph node involvement in breast cancer. Recommended technique requires the intradermal injection of 40 MBq 99mTc-nanocolloid in 0.1 ml on the day prior to surgery. Method The 0.1 ml doses were dispensed accurately using aseptic technique and no extra volume was added to accommodate for the dead space in the syringe. Activity in each syringe and needle was measured before and after injection, and the actual injected dose was recorded. The patients were scanned normally and underwent surgery the following day. Results One hundred and three doses were measured, showing injected activities of between 4 and 47.6 MBq. One hundred patients received less than 90% of the recommended dose, but nodes were successfully located in theatre on every occasion. Conclusion SLN imaging can be carried out successfully using very little injected activity. Measures to ensure that the entire dose enters the patient are unnecessary.

T3 Which drink is it the real thing for myocardial fizziology? B. Hughes, R.S. Lawson and M.C. Prescott Manchester Royal Infirmary, UK. Images from myocardial perfusion studies using 99mTc Myoview and 99mTc-sestamibi are often affected by radioactivity in the gastro-intestinal tract. This can impair the interpretation of the images if it comes too close to the heart. If this happens the routine procedure in our department is to ask the patient to drink some diet Coca-Cola. We have found that the gas released by this drink often helps to move GI activity away from the heart. In order to test whether other drinks might be equally effective, we have performed laboratory investigations to compare the volume of gas produced by 7 different carbonated drinks and by sachets of Carbex. Two sachets of Carbex produced 407 ± 16 ml of gas. The volume of gas available in the carbonated drinks after pouring into a glass ranged from 253 ± 12 ml (Lilt) to 647 ± 18 ml (Sprite). All drinks could give more gas if drunk through a straw straight from the can, ranging from 533 ± 42 ml (Sprite) to 1230 ± 56 ml (Pepsi-Cola). Sprite, Coca-Cola, Pepsi-Cola, carbonated water and soda water gave most gas. Patients may therefore now be able to have a choice of different fizzy drinks to aid their myocardial scan.

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NEWS AND VIEWS March 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)1274 383 174, E-mail [email protected] Nuclear Medicine Communications 2006, 27:313–314

NHS role in the management of radiation emergencies – 6 December 2005 – meeting report

The meeting as a whole was aimed at presenting information on what is expected of the NHS in a variety of radiation emergency situations. Speakers included representatives of the Department of Health, the emergency services, the Health Protection Agency Radiation Protection Division (formerly NRPB), A&E departments and medical physics departments. There were also talks from the Birmingham Regional Resilience Team and the Government Decontamination Service, neither of which I had heard of prior to this meeting. The first talk, by Monika Temple, described the role of the Department of Health (DoH) Emergency Preparedness Division. The division’s responsibility, as its name suggests, is to coordinate the DoH response to an emergency, and the talk was largely a description of work in progress, and ranged over much wider issues than radiation, such as pandemics, isolated cases of scary illnesses, and the need to respond to international incidents such as a tsunami or a Chernobyl. The division is revising the 1998 NHS Emergency Planning Guidance. The 2005 guidance can be found at: http://www.dh.gov.uk/assetRoot/04/ 12/36/04121236.pdf

Dr Rob Carr, a consultant in communicable disease control spoke on command and control arrangements and on arrangements for giving public health advice. The command arrangements consist mainly of acronyms, many of which are in flow chart boxes. The arrangements vary according to whether we have a major incident (serious but local, e.g. a possible bomb), a mass incident (e.g. possibly bird flu), or a catastrophic incident (e.g. an accident at a power station). The chain of command goes through the Strategic Health Authority and down to trusts, and there are gold, silver and bronze command levels. Don’t worry, we won’t get any higher than bronze. There is a parallel structure for advice headed by the Chief Medical Officer and including the Regional Director of Public Health, the Health Protection Agency and Local Health Advisory Teams. Trusts have a duty to prepare their own MAJAX plans and arrange for ‘business continuity’. Leslie Prosser spoke on the role of NRPB, now the HPA-RPD. She described the NAIR scheme (National Arrangements for Incidents involving Radioactivity) and the RPD’s role in preparing for emergencies, including training. She went on to discuss the sort of radioactive sources that might be used by terrorists, including large sources planted in a public place, a poor

man’s dirty bomb made up of smoke detectors, and the possible use of large caesium or strontium sources. Mick White from the Fire Service and Ian Chell from the Department of Health told us about the new approach to dealing with incidents. The emergency services are now probably better equipped with radiation measuring equipment than the average medical physics department, as they have personal alarming dosemeters, contamination and radiation monitors and, in some cases, hand-held gamma spectrometers. One interesting thing to note is that a fireman will accept a dose of up to 100 mSv to save your (salvageable) life; a paramedic will only go up to 1 mSv. The Fire and Rescue Service (FRS) is now extremely well equipped with acronyms for dealing with CBRN and HAZMAT. These include DIM, IRU, USAR and HVP (Detection Identification Monitoring units, Incident Response units, Urban Search and Rescue units and High Volume Pumping modules, respectively: all fire engines to you and me). Neil Higgins (HPA, Oxford) told us about ongoing discussions on iodine prophylaxis – the administration of stable iodine to reduce the uptake of the radioactive variety. The threshold for distribution remains, for the moment, at a thyroid dose of 30 mGy. There is discussion about lowering this to 10 mGy for children.

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314 Nuclear Medicine Communication 2006, Vol 27 No 3

Nick Gent spoke of the decontamination plans and equipment for A&E departments. Essentially, A&E departments are supposed to cope largely on their own, with only advice (rather than personnel) from medical physics. However, the final clean-up of the A&E department following the incident is seen as the duty of medical physics departments. Nick Gent’s highly positive view of NHS arrangements was followed by a highly cynical ‘self-confessed rant’ from Dr Anthony Bleetman, an A&E consultant in Birmingham. His main points were that in an emergency, many people will self-refer to A&E, whatever advice is given, and that he has severe doubts about the NHS decontamination suits (as used for chemicals). In his hospital he advocates the use of a simplified set of clothing for all incidents (not the inflatable suits, but still ‘over-thetop’ for radiation incidents). Karen Goldstone described her experiences in setting up regional monitoring units for incidents. It was clear that there are major differences between trying to do this in a rural community with many small centres of population, and in a large city where everyone has the opportunity to descend on a hospital or monitoring unit. Matt Furber described the work of the Government Decontamination Service. The most important message

is that this is an advisory body. Therefore it will not carry out the actual decontamination or pay for decontamination or confirm that decontamination standards have been achieved. David Temperton presented a rather brief talk, owing to lack of time, on what is expected of a medical physics department – making sure there is a plan and that it is correctly activated, and making sure that A&E is cleaned up after an emergency. Chris Taylor, Leeds Teaching Hospitals Trust Meeting Announcements

Annual Clinical PET CT course – a user’s guide Dates: 8–10 March 2006 Overview of basic science, PET CT imaging and interactive teaching. Contact for information and registration: [email protected] BNMS Spring Meeting Dates: 27–29 March 2006 Venue: Manchester, UK Website: www.bnms.org 2nd European IRPA Congress on Radiation Protection Dates: 15–19 May 2006 Venue: Paris, France Website: www.irpa2006europe.com BNMS Autumn Meeting Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk

EANM Annual Meeting Dates: 30 September to 4 October 2006 Venue: Athens, Greece Website: www.eanm.org 9th World Congress of Nuclear Medicine and Biology Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/congress2006/ index02.htm Education and Training

EANM Learning Courses Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected] EANM distance learning in nuclear cardiology

This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. http://www.eanm.org/eduOnline/ edu_start.php?navId = 332

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The editors wish to express their thanks to all the advisory editors and the reviewers listed below for their contribution to the journal. We realize this takes time and are grateful for the valuable comments that continue to ensure improvements in quality of the papers received. We hope for your continuing support Aboagye, E. Agress, H. Alavi, A. Allen, S. Anselmi, O.E. Avril, N. Badawi, R. Balan, K.K. Ballinger, J. Barlow, M. Barrington, S. Belohlavek, O. Behr, T.M. Blake, G. Bhargava, K. Boerman, O. Bolster, A. Bombardieri, E. Bradley, J. Bradley, K. Bradnam, M. Chung, J-K. Coffrey J. Coleman, R. Cook G. Cooper M. Costa, D. Darcourt, J. Decristoforo, C. Deehan, C. De Bondt, P. De Sutter, J. Dodig, D. Duogall, P. Deckart, H. Duatti, A. Dunn J.

Dutton, J. Ellis, B.L. Ellison, D. Fazio, F. Fleming, J.S. Fettich, J. Fogelman, I. Frank, J.W. Franklyn, J. Frier, M. Gambhir, S.S. Gardner, G. Gillen, G. Glesson, F. Gordon, I. Gray, H. Grosset, D. Gru ¨ning, T. Haberkorn, U. Hain, S.F. Hall, J. Ham, H.R. Harbinson, M. Harmer, C. Hesslewood, S. Hilson, A.J.W. Hojgard, L. Hoefnagel, C.A. Horowitz, N. Houston, A.S. Hutton, B.F. Ilic, S. Israel, O. Jones, T. Keating, D. Kelion, A. Kelman, A.

Kemp, P.M. Kettle, A.G. Kulkarni, P. Lawson, R. Lewington, V. Livierator, L. Meris, R. Mallick, U. Maltby, P. Marton, B. McCready, R. McDougall, I. McEwan, A.J. Menzel, C. Miller, A. Metcalfe, M. Miles, K. Mountford, P. Murray, T. Naidoo, V. Narula, J. Neilly J.B. Neto, S. Nemec, J. Nijran, KS. Nobili, F. Notghi, A. Nunan, T.O. Oyen, W. Palmer, M. Pattesson, J. Piepsz, A. Pons, F. Poon, F-W. Redmond, P. Reed, L. Reid, C.

Robinson, P.S. Royal, H.D. Sambuceti, G. Schillaci, O. Schleyer, P. Sayman, H.B. Schey, S. Schlumberger, M.J. Schwaiger, M. Sharp, P. Siegel, B. Simpson, D. Smith, ML. Somer, E. Staff, R. Spitzweg, C. Szulc, M. Tamaki, N. Tindale, W. Thorley, P. Tuttle, T. Tweddel, A. Underwood, S.R. Van des Wall, H. Vinjamuri, D. Vinjamuri, S. Vita, N. Wegner, E. Wolf, G. Wong, W. Wong, W.L. Wyper, D. Yamashina, S. Zhang, H.

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Editorial

Optimal imaging of patients with ischaemic heart failure Riemer H.J.A. Slarta, Pieter L. Jagera, Dirk J. van Veldhuisenb and Jeroen J. Baxc Nuclear Medicine Communications 2006, 27:317–320 a Department of Nuclear Medicine and Molecular Imaging, bDepartment of Cardiology, University Medical Center Groningen and cDepartment of Cardiology, Leiden University Medical Center, the Netherlands.

Correspondence to Dr Riemer H.J.A. Slart, Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen,

Introduction The increasing number of patients with coronary artery disease and ischaemic left ventricular dysfunction poses a major burden on clinical cardiology. Potential reversibility of chronic left ventricular dysfunction is an important clinical consideration in patients with ischaemic cardiomyopathy who are considered for revascularization [1,2]. Different imaging techniques for the detection of myocardial ischaemia and viability are available, including (gated) single photon emission computed tomography (SPECT), positron emission tomography (PET), magnetic resonance imaging (MRI) and (stress) echocardiography. These imaging techniques are in constant development and innovations are rapidly implemented in the clinical evaluation of patients with ischaemic heart failure. This editorial addresses recent developments in viability imaging. In addition, evaluation of the autonomic nervous system may be important in the assessment of patients with ischaemic cardiomyopathy [3]. In recent years, several PET and SPECT radiotracers have been developed for evaluation of cardiac innervation [4]. The potential value of imaging the autonomic nervous system by nuclear techniques in patients with ischaemic cardiomyopathy is also addressed in this brief point of view.

Viability assessment: PET or (DISA) SPECT? Conventional rest–stress gated myocardial perfusion SPECT is a widely used and well-validated technique for the assessment of myocardial ischaemia and left ventricular function, with a high accuracy for detection of coronary artery disease. The addition of gating to SPECT substantially increases the accuracy of the test and has now become the standard. Attenuation correction also increases the accuracy of myocardial perfusion scintigraphy for the detection of ischaemia and viability, but is less used in clinical practice [5]. The use of nitrates during flow tracer administration has also been postulated by several SPECT studies to improve the assessment of

Hanzeplein 1, P.O. Box 30001, 9700 RB, Groningen, the Netherlands. Tel: + 0031 50 361 3541; fax: + 0031 50 361 1712; e-mail: [email protected]

Received 13 October 2005 Accepted 7 December 2005

myocardial viability. Sciagra et al. [6] reported that nitrate administration significantly enhanced cardiac 99mTcsestamibi uptake, allowing for better identification of viable myocardium with superior prediction of recovery of function after revascularization. Gated myocardial perfusion SPECT also has improved assessment of myocardial viability, by implementing preserved wall thickening and wall motion on gated SPECT as markers of viable myocardium. However, fluorodeoxyglucose (FDG) imaging is still considered the most accurate method for the detection of myocardial viability with a sensitivity of 93% determined from pooled analysis of 20 studies with 598 patients [7]. Dynamic acquisition with PET allows absolute quantification of myocardial metabolism and perfusion, which may be preferred for assessment of myocardial viability, in particular in patients with severely depressed left ventricular function [8]. Whether FDG should be used in combination with perfusion (to assess perfusion–metabolism mismatches) or can be used in isolation (when percentage FDG uptake is used as marker of viability) is a matter of debate and more studies are needed to resolve this issue. Of interest, at present gated FDG PET is possibly an attractive PET approach, because this technique is less time-consuming and has been reported to accurately assess myocardial viability and left ventricular function simultaneously [9,10]. FDG is a tracer for PET imaging, but can also be used in conjunction with conventional gamma cameras using ultra-high energy collimators [11]. A recent study showed an overall good agreement between 99mTc-sestamibi/FDG DISA SPECT and PET for the assessment of myocardial viability in patients with severe left ventricular dysfunction [12]. Quantitative or visual analysis of the DISA SPECT data does not influence the agreement with PET, suggesting that visual assessment may be sufficient for clinical purposes [12].

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Studies using DISA SPECT for the prediction of improvement in regional and global left ventricular function in patients undergoing revascularization are scarce, but recent data indicate good accuracy to predict improvement of function [13]. At present, DISA SPECT is not widely implemented in nuclear departments, mainly due to: 1. The limited data in the literature on the prediction of functional recovery using DISA SPECT for the detection of myocardial viability. However, new data about this issue is in press [14]. Also, the relative merit of the technique in comparison to other modalities is not established yet. 2. The limited availability, costs and reimbursement of FDG. However, with the increasing number of PET cameras, FDG imaging will become accessible. In addition, the wide availability of SPECT systems equipped with 511 keV collimators will also increase implementation of DISA SPECT in clinical practice.

Viability assessment: other techniques Magnetic resonance imaging and echocardiography are other potent diagnostic tools for myocardial viability assessment. Both MRI and echocardiography have been used in combination with dobutamine stress to assess contractile reserve as a marker of myocardial viability. Various studies have indicated that patients with substantial contractile reserve indeed exhibited improvement in LVEF after revascularization and showed a good prognosis [15,16]. More recently, contrast-enhanced MRI has been used for the assessment of myocardial viability [17]. To be more precise, this technique actually allows accurate delineation of scar tissue, rather than assessment of viable myocardium. Animal experiments have demonstrated the excellent agreement between contrast-enhanced MRI and histology in the delineation of scar tissue [18]. The high spatial resolution allows precise delineation of the transmurality of the scar formation, which is currently not possible with other non-invasive imaging techniques. However, MRI and echocardiography are also characterized by several disadvantages. Echocardiography is hampered by poor acoustic windows in a substantial number of patients, and the inter-observer and intraobserver variability is limited; also quantification of results is difficult. Dobutamine stress MRI has been demonstrated to be highly accurate in the prediction of functional recovery [19], but the procedure and the analysis are time-consuming. Also, the use of MRI is limited in the presence of (biventricular) pacemakers and ICDs, which are frequently implanted in patients with heart failure.

Computed tomography (CT) is not generally used for the assessment of myocardial viability. However, a recent report has demonstrated the feasibility of assessment of contrast-enhancement as a marker of scar tissue, in a comparable manner as used with MRI [20]. A novel approach would be to combine the left ventricular anatomy (thinned walls, aneurysms) and fuse these images with PET images (obtained with FDG), as is now possible with PET–CT scanners. Another attractive application of multi-slice CT (MSCT) is the non-invasive visualization of coronary arteries with atherosclerotic lesions and obstructive stenoses [21]. MSCT (16-row or 64-row detector systems) has indeed compared well with invasive angiography, with reported sensitivities of 80– 90% and specificities > 95% [22]. Although the MSCT technique is very useful for exclusion of coronary artery disease (with negative predictive values > 95%), the integration of coronary artery stenosis, left ventricular anatomy (thin walls, aneurysms) and the presence of an occluded, stenotic or non-stenotic coronary artery is important in the decision-making on optimal therapeutic treatment. Currently, no data are available supporting this concept, however.

Mapping of cardiac innervation The autonomic nervous system plays a key role for regulation of cardiac performance, and the clinical relevance of alterations of innervation in the pathophysiology of various cardiac diseases has been increasingly recognized. In ischaemic heart failure, abnormalities of myocardial sympathetic function may contribute to arrhythmogenesis [23], and the denervated area has been demonstrated to correspond well with the area of risk [24]. Importantly, patients with an acute infarction may show preserved left ventricular function after reperfusion, while sympathetic innervation may already be impaired [25]. Nuclear imaging techniques have been introduced which allow global and regional evaluation of the myocardial nervous system [26]. In particular, the guanethidine analogue meta-[123I]iodobenzylguanidine (123I-MIBG) is promising for scintigraphic mapping of presynaptic sympathetic innervation and has been used extensively to determine innervation and denervation in patients with heart failure, and impaired cardiac adrenergic innervation as assessed by MIBG imaging was strongly related to mortality in patients with heart failure [27]. In addition, MIBG imaging can also be used to monitor effect of therapy on innervation [28]; preliminary data in 13 patients with heart failure undergoing MIBG imaging before and 6 months after cardiac resynchronization therapy demonstrated favourable changes in the neurohumoral system [29]. Cardiac innervation can also be assessed with PET tracers. For example, beta-adrenergic receptor density can be measured with PET and (S)-11C-CGP 12388 [30].

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Editorial Slart et al. 319

Myocardial beta-adrenoceptor density 1 month after acute myocardial infarction predicts left ventricular volumes at 6 months, and it has been demonstrated that beta-adrenergic receptor density is reduced after acute infarction, and the severity of reduction predicts later left ventricular dilation [31].

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Also, 11C-hydroxyephedrine (11C-HED) has been used to study innervation and denervation in patients who underwent cardiac transplantation [32]. It is the hope that the relation between regional denervation on the one hand, and ventricular arrhythmias on the other hand, will eventually allow better selection of patients who may need ICD implantation. This is a very timely topic, since currently, based on MADIT II criteria [33], many patients with previous infarction, and depressed left ventricular function will be referred for ICD implantation, and the costs may exceed the resources.

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Future perspectives and conclusions Cardiac imaging techniques play an important role in the non-invasive evaluation of ischaemia and myocardial viability in patients with ischaemic heart failure. Various techniques are available and these help to guide the therapeutic decision-making process. The currently available techniques include SPECT, PET, stress echocardiography and MRI. The newer developments in nuclear cardiology, in particular the use of PET–CT scanners, may further help to optimize non-invasive evaluation of patients with ischaemic cardiomyopathy. In addition, non-invasive assessment of innervation, as possible with PET and SPECT, may further help in risk stratification of patients with heart failure, and may particularly be useful in selection of patients for sophisticated therapies including resynchronization therapy or ICDs.

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28 Somsen GA, van Vlies B, de Milliano PAR, Borm JJJ, van Royen EA, Endert E, et al. Increased myocardial [I-123]-metaiodobenzylguanidine uptake after enalapril treatment in patients with chronic heart failure. Heart 1996; 76:218–222. 29 Erol-Yilmaz A, Verberne HJ, Schrama TA, Hrudova J, de Winter RJ, Eck-Smit BLF, et al. Cardiac resynchronization induces favorable neurohumoral changes. Pace-Pacing and Clinical Electrophysiology 2005; 28:304–310. 30 de Jong RM, Willemsen ATM, Slart RHJA, Blanksma PK, van Waarde A, Cornel JH, et al. Myocardial beta-adrenoceptor downregulation in idiopathic dilated cardiomyopathy measured in vivo with PET using the new

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Original article

Evaluation of left ventricular ejection fraction by the quantitative algorithms QGS, ECTb, LMC and LVGTF using gated myocardial perfusion SPECT: investigation of relative accuracy Magdy Mohamed Khalila,b, Abdelhamid Elgazzara,b and Wafaa Khalilc Aim To compare the quantitative algorithms Emory Cardiac Toolbox (ECTb), quantitative gated SPECT (QGS), layer of maximum counts (LMC), and left ventricular global thickening fraction (LVGTF) using gated myocardial tomography in the calculation of the left ventricular ejection fraction using the regression without truth (RWT) technique. Materials and methods Seventy-four consecutive patients were included in the study (59 males). All patients underwent stress–rest myocardial perfusion SPECT using 99m Tc-tetrofosmin. Analysis of variance (ANOVA), the paired Student’s t-test, the Pearson correlation coefficient and Bland–Altman were used for comparing the methods. The relative accuracy was performed by RWT. Results ANOVA revealed a significant difference among the methods in calculating the ejection fraction. RWT showed that ECTb and QGS outperformed the other two methods. The ECTb was slightly better than QGS, and LMC was slightly better than LVGTF. QGS and ECTb achieved good correlations in end diastolic volume, end systolic volume and ejection fraction measurements. One-way ANOVA demonstrated that QGS was the only software program affected by the category of the perfusion summed stress score (SSS), P = 0.038. The ejection fraction determined by the QGS, ECTb and LVGTF methods correlated significantly with defect size (r = 0.545, P < 0.0001; r = 0.530, P < 0.0001; and r = 0.419, P < 0.0001,

Introduction Gated single photon emission computed tomography (SPECT) imaging has allowed the simultaneous assessment of myocardial perfusion and the calculation of the left ventricular volumes, end diastolic volume (EDV), end systolic volume (ESV) and ejection fraction. Myocardial functional parameters are valuable indicators in stratifying patients to assess the most appropriate treatment strategy. It has been shown that the post-stress ejection fraction is the best predictor of cardiac death [1]. In the last decade, there have been several approaches for measuring the left ventricular volumes and the computation of ejection fraction. Starting from simple methods to

respectively), but the LMC method was not significantly correlated (r = 0.216, P = 0.067). Conclusions There was a considerable variation among the quantitative gated SPECT methods in the evaluation of the ejection fraction. RWT revealed that the ECTb and QGS outperformed the other two methods with respect to the bias and precision of the measurements. Pair-wise correlations of the four methods ranged from mild to good with large agreement limits. Results of RWT provided important information in ranking the quantitative gated c 2006 SPECT methods. Nucl Med Commun 27:321–332
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:321–332 Keywords: Gated Myocardial SPECT, Relative accuracy, Quantitative methods, left ventricular EF a Nuclear Medicine Department, Faculty of Medicine, Kuwait University, bMubarak Al-Kabeer Hospital, Ministry of Health, Kuwait and cBiophysics Department, Faculty of Science, Cairo University, Egypt.

Correspondence to Magdy Mohamed Khalil, Nuclear Medicine Department, Faculty of Medicine, Kuwait University, P.O. Box 24923, Code 13110, Safat, Kuwait. Tel: + 00965 531 2300 ext. 6412; fax: + 00965 533 8936; e-mail: [email protected] This work was presented in part at the 52nd Society of Nuclear Medicine Annual Meeting, June 18–22, 2005, Toronto, Canada. Received 19 October 2005 Accepted 30 December 2005

advanced three-dimensional modelling of the left ventricle, each method has its own advantages and drawbacks. The basic assumptions might be count density sampling, geometric modelling (spherical, cylindrical, ellipsoidal), systolic thickening, or geometric-count-based principles [2–9]. This apparent heterogeneity among the methods may stimulate researchers to look for the most reliable and accurate method. However, in myocardial SPECT imaging, there are many degrading factors that may substantially hinder the gated SPECT methods to accurately estimate the left ventricular volumes and ejection fraction [6,10,11].

c 2006 Lippincott Williams & Wilkins 0143-3636


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322 Nuclear Medicine Communications 2006, Vol 27 No 4

Two methods are commonly seen in nuclear cardiology laboratories: quantitative gated SPECT (QGS, CedarsSinai Medical Center, Los Angeles, California, USA) [2,12] and Emory Cardiac Toolbox (ECTb, Emory University, Atlanta, Georgia, USA) [3,13]. Both methods have gained a clinical acceptance for estimating myocardial functional parameters. Several studies have demonstrated that both methods are significantly correlated with each other and with ‘gold standard’ techniques [2–4,9,14–16]. Another two methods have been reported in the literature: the layer of maximum counts (LMC) and the left ventricular global thickening fraction (LVGTF). The LMC method was reported recently and works in the prolate spheroid coordinate system [5,6]. It avoids the step of contouring the endocardium for volume estimation. Furthermore, the method depends on a reference source for the calculation of the left ventricular ejection fraction [6]. The LVGTF method depends on partial volume phenomenon and rests on systolic count change to calculate the regional myocardial thickening. It estimates the ejection fraction through appropriate geometric assumptions and measurements of the systolic thickening [7,8]. The diversity seen among the methods and the differences observed in comparison studies may confuse the end user with regard to the most appropriate one to use even though each has achieved good correlation with reference techniques and correlated significantly in estimating myocardial functional parameters [2–4,9]. In an attempt to give a measure of precision and accuracy of the estimation methods, a mathematical tool called regression without truth (RWT) was developed to partially resolve the problem [17,18]. In brief, this technique requires a statistical distribution of the ‘gold standard’ data and iteration algorithm to reach the maximum likelihood of the regression parameters that describe the relationship between the estimate of interest, i.e., ejection fraction by the ‘gold standard’ and the estimation methods. The developers of the method derived an expression for the log-likelihood of the model parameters which did not require knowledge of the true ejection fraction (i.e., without the use of a ‘gold standard’). This is analogous to fitting lines without the use of the x axis [17]. The objective of the study was to utilize the new technique (RWT) to compare and rank the four methods (QGS, ECTb, LVGTF and LMC) in the calculation of the left ventricular ejection fraction.

the study (59 males, 15 females, mean age 57 ± 10 years, range 36–80 years). Thirty-four of the patients had had previous myocardial infarction, 33 had diabetes, 19 were smokers, 26 had angina, 37 were hypertensive, six had a coronary artery bypass graft, seven were obese, three had bronchial asthma and one had undergone percutaneous transluminal coronary angioplasty. All patients underwent 99mTc-tetrofosmin gated myocardial perfusion SPECT in a 2-day protocol. A stress test was performed either by a treadmill exercise or pharmacological stress. Thirty-eight patients performed the treadmill exercise using the Bruce protocol and 36 patients underwent pharmacological stress (dipyridamole, 0.57 mg  kg – 1 over a 4-min infusion). Patients were instructed to avoid coffee and other caffeine-containing products for 24 h before the test. A dose of 740–925 MBq (20–25 mCi) tetrofosmin was injected at peak exercise and the test continued for 60 s after injection. A dual head gamma camera (Millenium MG, General Electric Medical systems, Milwaukee, Wisconsin, USA) in cardiac 1011 position equipped with high resolution collimators was used for data acquisition. The acquisition arc was from the right anterior oblique to the left posterior oblique. The projection time was 20 s for a total of 36 projections using the step-and-shoot mode. The number of gates was 8 frames per cardiac cycle using the R-wave trigger and an acceptance window of 50–150% of the mean pre-acquisition heart rate. The energy peak was 140 keV with a 20% window. The acquisition matrix was 64  64 with a zoom factor of 1.33, giving a pixel size of 6.78 mm. Patients with arrhythmia, atrial fibrillation, severe motion artifacts, and those with high extra-cardiac activity were excluded from the study. Processing

Gated SPECT raw data were reconstructed by a conventional filtered back-projection algorithm accompanied by a ramp filter. Projections were pre-filtered using the Butterworth filter (cut-off value was 0.35 cycle/cm for gated data but 0.40 cycle/cm for ungated data, order 7). After data reconstruction, the transverse slices were reoriented into the three orthogonal slices (short, horizontal and vertical long axis slices) for display and interpretation. No scatter or attenuation correction was applied. The functional parameters were calculated from the stress data using a Xeleris workstation (version 1.06, GE medical systems).

Patients

Gated SPECT methods The quantitative gated SPECT and Emory Cardiac Toolbox algorithms

Seventy-four consecutive patients who showed abnormal myocardial perfusion were retrospectively collected for

The QGS and ECTb methods are described fully in the literature [2,3]. Both methods have different basic

Materials and methods

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Evaluation of LVEF by four algorithms Khalil et al. 323

assumptions and geometric approximations. The QGS uses an ellipsoidal model for myocardial sampling whereas the ECTb uses two coordinate systems (cylindrical coordinate for basal and mid-myocardial segments but spherical coordinate for the apex). They achieved good results with ‘gold standard’ techniques in estimating the left ventricular volumes and ejection fractions [2–4,12]. The QGS method uses asymmetric Gaussian profiles fitted to normally drawn profiles from the cavity centre on the myocardial walls [2]. The method utilizes the counts within and among frames in addition to the conservation of mass in outlining the myocardial borders. On the other hand partial volume and Fourier transform are employed by the ECTb to theoretically compute the myocardial endocardial offsets [3,13]. The left ventricular global thickening fraction and the layer of maximum counts algorithms

The LVGTF method was developed to avoid outlining the endocardial boundaries [7] and it is based upon the principle of the partial-volume effect. The method utilizes the systolic count change during cardiac contraction to compute the systolic thickening. Theoretically, myocardial systolic thickening could give an estimate of myocardial ejection fraction. According to this hypothesis in addition to geometric assumptions, a formula was derived to compute the global left ventricular ejection fraction from a measure of regional myocardial thickening. Clinical validation of the LVGTF method demonstrated good correlation with equilibrium gated blood pool [8]. Measurements of the ejection fraction, determined by the LVGTF method, were obtained from the cardiac software ‘Myovation’ designed for data reconstruction and display in a Xeleris Workstation (GE Medical Systems). The LMC method was reported recently and depends on the calculation of the layer of maximum counts to compute the enclosed volume (Vmax) which is then used to calculate the corresponding left ventricular ejection fraction (LVEFmax). The method assumes a linear relationship between the maximum ejection fraction of the maximum counts (EFmax) and the left ventricular ejection fraction using the equation EF ¼ EFmax ð1 þ constantÞ ! 1: Plotting the LVEF computed by a reference method versus the ejection fraction of the maximum counts (EFmax), the regression slope (1 + constant) can be obtained, which is then applied to compute the LVEF [6]. The LMC was not extensively evaluated in patients with perfusion abnormalities. Since the method needs a reference method, we calibrated the EFmax versus that obtained from gated blood pool. For the sake of calibration, nine patients who underwent gated SPECT and gated blood pool within less than

1 month of each other were selected (eight males and one female; mean age 56 ± 11 years). The ejection fraction of the layer of maximum counts (EFmax) was plotted versus the ejection fraction calculated by gated blood pool. By applying the linear regression fit, the slope was calculated and employed in the calculation of the ejection fraction of the study population using Equation 1. The slope obtained was 0.918, with a correlation coefficient of r = 0.907. Recently, we used the same calibration curve to evaluate the LMC method in patients with small hearts [19]. Perfusion quantification

Both stress and rest studies were processed using the 20segment model for perfusion scoring. The segments were automatically scored regarding radiotracer uptake, using a 5-point scoring system (0 = normal, 1 = equivocal, 2 = moderately reduced, 3 = severely reduced and 4 = absent) using the ECTb software package. Scoring results were classified according to the following: normal for scores < 4, mildly abnormal for score 4–8, moderately abnormal for scores 9–13, and severely abnormal for scores > 13 [20]. The score is based on the normal limits generated by a polar map produced from patients with a low likelihood of coronary artery disease. The most suited normal database was a 2-day sestamibi file. The mean defect size (as a percent of the total myocardium) was 25.0 ± 15.4% or as a myocardial mass of 37.6 ± 27.8 g. The summed stress severity score was, on average, 641 ± 443 as calculated by the ECTb [21]. Results of myocardial perfusion scores are given in Table 1. Using the summed stress score (SSS) the distribution of patients according to perfusion abnormalities was two with SSS < 4, eight with SSS = 4–8, 16 with SSS = 9–13, and 48 with SSS > 13. The regression without truth technique

‘Regression without truth’ is a statistical technique proposed to rank the estimation methods for the evaluation of the LVEF [17,18]. The technique works to estimate the relative accuracy and consistency of the methods used without assuming a priori that one of the methods is the ‘gold standard’. In other words, the developers of the method answered the question whether Table 1 Perfusion quantification as calculated by the Emory Cardiac Toolbox software program Perfusion parameters

Mean value

Range

Summed stress score Summed rest score Summed difference score Myocardial mass (ungated) Defect mass Defect mass (as percent of total myocardium) Severity score (stress)

17.0 ± 8.0 11.4 ± 8.0 5.6 ± 5.6 145 ± 34 g 38 ± 28 g 25 ± 15%

0–39 0–32 0–24 80–259 g 5–95 g 5–59%

641 ± 443

0–1422

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324 Nuclear Medicine Communications 2006, Vol 27 No 4

we can plot the estimated EFs versus those values calculated by the reference method without knowing the true values. Traditionally such a process needs a reference method, as commonly seen in the medical literature. RWT succeeded in solving this issue by assuming a parameterized distribution of the ‘gold standard’ (i.e., a truncated normal distribution and a beta distribution). Furthermore, a statistical model that related the true ejection fraction to estimates of its value was assumed [18]. By using the measured data with the postulated assumptions the model parameters and the parameters characterizing the assumed distribution can be estimated. To accomplish this task the maximum likelihood estimator was employed. (A more detailed explanation of the underlying theoretical basis for this method is given in the Appendix.) The idea behind the maximum likelihood parameter estimation is to determine the parameters that maximize the probability (likelihood) of the sample data. From a statistical point of view, the method of maximum likelihood is considered to be more robust and yields estimators with good statistical properties. Here, the assumed statistical model is the regression line with its well known parameters: slope, am, intercept, bm, and a noise level characterized by sm. Two distributions were studied and demonstrated a good performance in simulation studies, a truncated normal distribution (with a varying mean and variance) and beta distribution [17]. The maximum likelihood estimator finds the maximum probability of the parameters that make the measured data by each gated SPECT method fit the model assumed. The ranking of the methods is then performed by calculating the error term (s=a). This term was derived considering the root mean square error (the underlying assumption of regression fitting). For the linear model assumed, the accuracy of the estimation can be achieved approximately by adjusting the measurements using the estimated model parameters am and bm. After this correction is made, the variance can be expressed as s=a. It was shown that the root mean square error increases in accordance with 1=am and therefore the term s=a combines between accuracy and precision in the ranking task [17,18]. Advantages of the ‘regression without truth’ method over other statistical methods

Correlation analysis and linear regression fitting reveal the statistical relationship between two methods regardless of their accuracy and precision in parameter estimation. Bland–Altman analysis plots the mean value versus the differences and if the differences follow a normal distribution within a variation of ± 2 SD, agreement is achieved and both methods could be described as

interchangeable [22]. However, the agreement limits and regression trend are substantially considered to accept agreement in the clinical setting. Nevertheless, all these statistical techniques could not allow for objective determination or even ranking of the estimation methods. RWT treated this point in an interesting way as mentioned above (see Appendix). Because such a statistical technique has no bias or imprecision towards a patient’s physiological state, it could provide a fair ranking of the quantitative gated SPECT methods avoiding some of the limitations that are encountered by ‘gold standard’ methods in the clinical environment. Statistical analysis

Continuous data were expressed as the mean ± standard deviation (SD). Student’s paired t-test was used for comparison of means. The standard error of an estimate (SEE) was calculated to determine the accuracy with which the sample mean estimates the population or ‘true’ mean. Analysis of variance (ANOVA), assuming equality of variance, was used to test the statistical significance of the mean ejection fraction calculated by the four methods: QGS, ECTb, LVGTF and LMC. In addition, we used the Bonferroni correction for multiple comparisons in post-hoc analysis. The Pearson correlation coefficient and regression analysis were used to study the statistical relationship between the gated SPECT methods. A Bland–Altman plot was applied to study the agreement limits and to search for systematic errors and trends [22]. A P value < 0.05 was considered statistically significant. A statistical software package for windows, version 12.0 (SPSS Inc., Chicago, Illinois, USA) was used for data analysis.

Results Results of RWT are given in Table 2. The figure of merit (s=a) was better in ECTb compared to the other methods. This was consistent by the two assumed distributions of the ‘gold standard’ (truncated normal and beta distributions). Table 2 Results of the regression without truth technique for beta and truncated normal distributions Method QGS

ECTb

LVGTF

LMC

Tnorm. dist. Beta dist.

1.0015 0.9510

1.2000 1.1456

0.7483 0.7116

0.5000 0.5000

Tnorm. dist. Beta dist.

– 0.2989 – 0.2322

– 0.3688 – 0.2934

– 0.0065 0.0425

0.0671 0.0816

Tnorm. dist. Beta dist. s/a Tnorm. dist. Beta dist.

0.0463 0.0468

0.0384 0.0372

0.1312 0.1311

0.0863 0.0867

0.04623 0.0492

0.0320 0.0324

0.1753 0.1842

0.1726 0.1734

a

b

s

QGS, quantitative gated SPECT; ECTb, Emory Cardiac Toolbox; LVGTF, left ventricular global thickening fraction; LMC, layer of maximum counts.

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Evaluation of LVEF by four algorithms Khalil et al. 325

However, ECTb was slightly better than QGS. The figure of merit (s=a) was 0.0320 and 0.0324 for ECTb, but 0.04623 and 0.0492 for QGS, corresponding to truncated normal and beta distributions, respectively. The other two methods, LMC and LVGTF, showed a lower performance than ECTb and QGS. The LMC method was slightly better than LVGTF: the error values were 0.1726 and 0.1734 for LMC, but for LVGTF method, the values were 0.1753 and 0.1842 in the truncated normal and beta distributions, respectively. Moreover, the slope values for QGS and ECTb were very close to unity and intercepts were close to zero. This indicated that both methods are correlating well with the ‘gold standard’ rather than the other two methods (Table 2).

ences at higher mean values (r = 0.205, P = 0.080, and r = 0.222, P = 0.058, respectively) (Fig. 2 (a and b)). The results for the ejection fraction as determined by QGS and ECTb also showed a good correlation (y = 0.792x + 3.2%, SEE = 5.6%, r = 0.929, P < 0.0001). For LVGTF and LMC, correlations with ECTb were y = 0.604x + 23.4, SEE = 13.3%, r = 0.627, P < 0.0001

Fig. 1

(a) 350 300

y = 1.002x + 1.7 r = 0.926, P < 0.0001 SEE = 20.3 ml

According to the results showed by the RWT technique, we studied the correlation of the QGS, LMC and LVGTF methods with ECTb which demonstrated the best performance in calculations of the ejection fraction.

QGS EDV (ml)

250 200 150 100

The ejection fraction as determined by the ECTb was significantly higher than that by QGS and LMC (P < 0.0001). However, it was not significantly different from the value determined by the LVGTF method (P = 0.445). The mean ejection fraction determined by QGS was significantly lower than that by the LVGTF method but was not significantly different from the LMC method (P < 0.0001 and P = 0.125, respectively). The ejection fraction by LVGTF was significantly higher than that by LMC (P < 0.0001) (Table 3). Volumes estimated by QGS and ECTb showed good correlations. For EDV, regression analysis results were y = 1.002x + 1.7, SEE = 20.3 ml, r = 0.926, P < 0.0001 but for ESV y = 1.020x – 10.5, SEE = 15.16 ml, r = 0.951, P < 0.0001 (Fig. 1 (a and b)). Bland–Altman plots for volumes demonstrated that both methods had a nonsignificant trend to increase the EDV and ESV differ-

50 0 0

50

100

150 200 250 ECTb EDV (ml)

300

350

(b) 250 y = 1.020x −10.5 r = 0.951, P < 0.0001 SEE = 15.2 ml

200 QGS ESV (ml)

Both revealed a significant difference among the methods in calculating the ejection fraction (P < 0.0001). The mean EDV by QGS was not significantly different from that calculated by ECTb (123.8 ± 53.2 ml vs. 121.9 ± 49.2 ml, P = 0.406). However, the mean ESV by QGS was significantly higher than that by ECTb (71.0 ± 48.8 ml vs. 59.2 ± 45.5 ml, P < 0.0001).

150

100

50

0 0

50

100 150 ECTb ESV (ml)

200

250

Scatter plots of left ventricular volumes as calculated by the quantitative gated SPECT (QGS) and Emory Cardiac Toolbox (ECTb) methods. (a) End diastolic volume (EDV); (b) end systolic volume (ESV). The line of identity is overlaid.

Table 3 Results of end diastolic volume, end systolic volume and ejection fraction as calculated by the four methods QGS, ECTb, LVGTF and LMC

End diastolic volume (ml) End systolic volume (ml) Ejection fraction (%)

QGS

ECTb

LVGTF

LMC

P

123.8 ± 53.2 71.0 ± 48.8 46.4 ± 15.0

121.9 ± 49.2 59.2 ± 45.5 55.7 ± 17.6

–

–

57.1 ± 17.0

45.2 ± 10.8

0.406 < 0.0001 < 0.0001

QGS, quantitative gated SPECT; ECTb, Emory Cardiac Toolbox; LVGTF, left ventricular global thickening fraction; LMC, layer of maximum counts.

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326 Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 2

(a)

Fig. 3

150

(a) 100 90

y = 0.792x + 3.265 r = 0.926, P < 0.0001 SEE = 5.6%

80

100 50 Mean + 2 sd d

0 0

100

200

−50

QGS EF (%)

(ECTb-QGS) EDV (ml)

70 60 50 40 30 20

Mean− 2 sd

10 0 0

−100 −150 100

(b) 100 90 80 70 60 50 40 30 20 10 0

y = 0.351x + 25.6 r = 0.571, P < 0.0001 SEE = 8.9%

LMC EF (%)

(ECTb-QGS) ESV (ml)

(b)

Mean EDV (ml)

10 20 30 40 50 60 70 80 90 100 ECTb EF (%)

50 Mean + 2 sd 0 0

100

0

200 Mean− 2 sd

−50

(c) 100 90

y = 0.604x + 23.4 r = 0.627, P < 0.0001 SEE = 13.3%

80 −100

Bland–Altman plots using the quantitative gated SPECT (QGS) and the Emory Cardiac Toolbox (ECTb) methods for the calculation of (a) end diastolic volume (EDV) and (b) end systolic volume (ESV).

70 LVGTF EF (%)

Mean ESV (ml)

10 20 30 40 50 60 70 80 90 100 ECTb EF (%)

60 50 40 30 20

and y = 0.351x + 25.6, r = 0.571, SEE = 8.9%, P < 0.0001, respectively (Fig. 3 (a–c)), but their correlation results with QGS were y = 0.649x + 26.3, SEE = 14.0%, r = 0.574, P < 0.0001 and y = 0.444x + 24.19, SEE = 8.6%, r = 0.615, P < 0.0001, respectively. Lower but significant correlation was found between LVGTF and LMC (y = 0.520x + 33.6, SEE = 16.14%, r = 0.332, P < 0.0001) (Fig. 3 (c)). Agreement analysis showed that QGS and LMC had a significant trend to increase the ejection fraction difference with ECTb as the mean increased (r = 0.398, P < 0.0001, and r = 0.524, P < 0.0001, respectively). For LVGTF, no significant trend was seen across the whole range of ejection fractions studied with ECTb (r = 0.048, P = 0.684) (Fig. 4 (a–c)). One-way ANOVA revealed that QGS was the only software program influenced by the category of the

10 0 0

10 20 30 40 50 60 70 80 90 100 ECTb EF (%)

Scatter plots of the four algorithms. (a) Quantitative gated SPECT (QGS), (b) layer of maximum counts (LMC) and (c) left ventricular global thickening fraction (LVGTF) with the Emory Cardiac Toolbox (ECTb). Regression lines are demonstrated and the line of identity is overlaid. EF, ejection fraction.

perfusion score (Fig. 5). The significance level was P = 0.038, 0.186, 0.657 and 0.918 for QGS, ECTb, LVGTF and LMC, respectively. Post-hoc analysis for ejection fraction by QGS showed a significantly lower ejection fraction values by patients with (SSS > 13) than those with (SSS = 4–8) (45.3 ± 14.6% vs. 59.8 ± 12.2%, respectively; P = 0.039). The mean ejection fraction for patients with SSS > 13 was not significantly different

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Evaluation of LVEF by four algorithms Khalil et al. 327

Fig. 4

75

70

50

60

25 0 −25

50

Mean + 2 sd

0

50

Mean− 2 sd 100

−50

Mean EF (%)

(ECTb-QGS) EF (%)

(a)

Fig. 5

40 30 20 10

−75

Mean EF (%) 0

(b)

SSS = 4 − 8

75

(ECTb-LMC) EF (%)

50 25 0 0

50

−25

−75

Mean− 2 sd 100

Mean EF (%)

75

Mean+2 sd

25 0 0 −25

50

100

Mean− 2 sd

−50 −75

Discussion We have compared four different methods developed for the calculation of the left ventricular function. Three of them (QGS, ECTb, and LVGTF) are commercially available and one (LMC) is in the preliminary stage of assessment. The comparison was performed in patients with abnormal myocardial perfusion SPECT.

50 (ECTb-LVGTF) EF (%)

SSS > 13

Histogram of the mean ejection fraction by using the four alogrithms – QGS ( ), ECTb ( ), LVGTF ( ) and LMC ( ) – versus the summed stress score. (Abbreviations as in the legend to Fig. 4.)

Mean + 2 sd

−50

(c)

SSS = 9 − 13

Mean EF (%)

Bland–Altman plots of the (a) quantitative gated SPECT (QGS), (b) layer of maximum counts (LMC) and (c) left ventricular global thickening fraction (LVGTF) methods with the Emory Cardiac Toolbox (ECTb) method for the estimation of ejection fraction (EF).

from those with SSS = 9–13 (45.3 ± 14.6% vs. 46.1 ± 15.6%, respectively; P = 0.981). Also, the ejection fraction by SSS = 4–8 and SSS = 9–13 was not significantly different (P = 0.103). The ejection fraction by QGS, ECTb and LVGTF inversely correlated with defect size (r = 0.545, P < 0.0001; r = 0.530, P < 0.0001; and r = 0.419, P < 0.0001, respectively) but the LMC method was not significantly correlated with defect size (r = 0.216, P = 0.067) (Fig. 6 (a–d)).

The study results showed a considerable variation among the four methods in the calculation of the LVEF. Since a ‘gold standard’ modality is not usually available, and just comparing the methods using the traditional statistical tests (Student’s t-test, regression line fitting, correlation coefficient and Bland–Altman plots) is not indicative of the accuracy or the precision of the measurements, a mathematical technique called regression without truth (RWT) was developed to partially resolve this issue [17,18]. Applying this technique to data obtained from patients with wide ranges of perfusion abnormalities and functional parameters calculated post-stress showed that the ECTb and QGS methods had the lowest error in estimating the ejection fraction compared to the LMC and LVGTF methods. Accordingly, we used the ECTb as a reference for evaluating the other methods, keeping in mind that it was not a ‘gold standard’ for the estimation task. There are some imaging confounders that limit the performance of these methods in precisely determining the ventricular volumes and/or estimating the systolic count change [6,8,10,11,23]. In addition, the variability among patients when gated SPECT imaging is performed (for example, patients with small hearts, hypertrophy and severe perfusion defects) may influence the accuracy of the estimation process and the clinical outcome as well

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328 Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 6

y = −0.83x+ 76.0 r = 0.530, SEE = 14.7% P < 0.0001

ECTb EF (%)

80 60 40

100 QGS EF (%)

100

20

60 40 20 0

0 0

10

20

30 40 Extent (%)

50

60

70

0

100

100

80

80

60 40 y = − 0.65x+ 72.8 r = 0.419, SEE = 15.5% P < 0.0001

20 0 0

10

20

30 40 Extent (%)

50

60

70

LMC EF (%)

LVGTF EF (%)

y = − 0.74x + 65.3 r = 0.545, SEE = 12.6% P < 0.0001

80

10

20

30 40 Extent (%)

50

60

70

y = − 0.21x + 50.1 r = 0.216, SEE = 10.6% P < 0.067

60 40 20 0 0

10

20

30 40 Extent (%)

50

60

70

Scatter plots of the ECTb, QGS, LVGTF and LMC methods with defect extent. Correlation results are shown. (Abbreviations as in the legend to Fig. 4.)

[9–11,19]. Various research efforts have demonstrated that the interchangeability of gated SPECT methods is of limited clinical value due to the above mentioned reasons in addition to the intrinsic characteristics of each method. The results presented here support these findings [9,14,16]. Different correlations were found between each pair of the four methods and considerable systematic and random errors were observed. Moreover, the agreement limits were high to the extent that narrowly allowed for interchangeability.

comparison to ECTb [4,16]. This is also in line with our results, as the mean difference between ECTb and QGS was 8.4%. More recently, the ejection fraction calculated by ECTb was not significantly different from that by MRI, but the ejection fraction determined by QGS underestimated the values determined by MRI (53.2 ± 11.5% vs. 62.7 ± 13.7% vs. 60.6 ± 13.9% for QGS, ECTb and MRI, respectively) and, again, the correlation between QGS and MRI was better than between ECTb and MRI [24].

The percentage of patients with previous myocardial infarction in the current study was relatively high (46%) and the majority of patients (B65%) had SSS > 13. The reliability of edge-based techniques may decrease in such cases due to inaccuracy in outlining the hypoperfused myocardial segments. Although ECTb and QGS depend, in part, on edge detection, they showed a better estimation by RWT than did LMC and LVGTF.

QGS significantly underestimated the ESV calculated by ECTb, which can be explained by the lower temporal resolution used (8 frames/cardiac cycle). It was demonstrated that QGS underestimated the ejection fraction, on average, by 3.71% if 8 frames/cycle is used instead of 16 frames/cycle [2]. In a meta-analysis, QGS was reported to underestimate the ejection fraction calculated by contrast left ventriculography, on average by 7.6 ± 17.4% (higher SD due to the heterogeneity of studies included in the analysis) in case 8 frames/cycle was used but the ejection fraction was not significantly different when 16 frames was applied [25].

When QGS and ECTb were compared with magnetic resonance imaging (MRI), QGS showed a better correlation than ECTb, whereas ECTb did not show a significant bias when compared to MRI [4]. This is consistent with our findings as the figure of merit (s/a) measures the degree of biasing from the ‘gold standard’. In the same study and others, QGS provided an underestimation of ejection fraction, on average, by 8% in

Shaefer et al. recently reported that QGS underestimated the ejection fraction by MRI due to the higher temporal sampling by MRI (20 gates/cycle) versus that used in QGS (8 gates/cycle), this underestimation was absent in

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Evaluation of LVEF by four algorithms Khalil et al. 329

ECTb [24]. These findings agreed with the results by RWT in that ECTb could estimate the ejection fraction slightly better than QGS. In patients with large myocardial infarction, QGS was shown to underestimate the ejection fraction calculated by gated blood pool. This was ascribed to volume overestimation when the ventricular volume increased [26]. When QGS was compared with gated blood pool in patients with large perfusion defects, it underestimated the ejection fraction by 4.7 ± 7.3% in addition to wide agreement limits in both 99mTc and 201Tl tracers [27]. In the previous two studies no apparent trend was seen by the difference in ejection fraction as the mean increased in Bland–Altman plots [26,27]. The technique by QGS uses a three-dimensional approach for sampling the myocardium and applying asymmetric Gaussian curves on normally subtended count profiles. Then 65% of the SD is taken to estimate the endocardial and epicardial borders. In the case of very poor count statistics along the count profiles, the standard deviations are combined with those of each of its four spatial neighbouring profiles [2]. More refinement is then applied by anatomical constraint of constant myocardial volume throughout the cardiac cycle. The dependence of QGS on count profiles might explain the sensitivity of the method to perfusion defects as resulted from the difference in ejection fraction by SSS = 4–8 and SSS > 13. In ECTb, the algorithm forces the hypoperfused segment to be a smooth connection between adjacent non-infarcted portions of the wall and because this segment is not thickening, it is pinned to its end diastolic positions [3]. RWT revealed that the LMC and LVGTF methods are beyond ECTb and QGS in the calculation of the ejection fraction. Furthermore, correlation results of the two methods with ECTb were lower than that by QGS. The LVGTF method showed the lowest systematic error with ECTb, as both methods are heavily dependent on the partial-volume effect and the variation of the end diastolic and end systolic pixel counts during systolic contraction [3,7]. Moreover, both methods exhibited a lower bias (small mean differences) in the Bland–Altman plot. However, the agreement limits were large with a 95% confidence interval of ( – 28.6% to 31.3%). The ejection fraction determined by the LMC method correlated fairly well with the ECTb, with a systematic increase in the ejection fraction difference as the mean increased. The LMC method is based on two assumptions. Firstly, that the layer of maximum counts remains at a fixed position in the left ventricular myocardium for the same imaging system during the heart cycle. Secondly, that the volumetric ratio between the left

ventricular myocardium and the cavity at the end diastolic phase is within a narrow physiological range [6]. This ratio could probably be underestimated due to either overestimation of the left ventricular cavity, especially in patients with large defects, or underestimation of the myocardial volume, as it was reported that SPECT imaging underestimates the myocardial wall thickness [28,29]. The weak correlation between LMC and LVGTF might be caused by the differences in the underlying assumptions and the geometric approximations by each method. The LMC method showed good results in patients with small hearts [6,19]. However, one major limitation of the method is that it depends on another reference source for calculating the left ventricular ejection fraction. This may influence the accuracy of the method and its analysis for patients with characteristics differ from those used in the calibration process. Recently, we reported that the LMC method outperformed the other three methods in patients with small hearts with respect to gated blood pool studies [17]. This advises the clinically oriented users to understand the advantages and disadvantages of gated SPECT methods and their differential utility in patient subsets. ANOVA showed that the ejection fraction measured by the ECTb had a non-significant trend to decrease as the category (SSS) of perfusion abnormality increased but it was significant with QGS. For LMC and LVGTF, the ejection fraction values were not influenced by the SSS. Moreover, QGS, ECTb and LVGTF showed significant correlation with defect extent (expressed as the percentage of myocardium). This is consistent with the ischaemic cascade where perfusion abnormalities induce regional dysfunction which may then influence the global function depending on the amount of ischaemia produced. However, this correlation might be statistically confounded due to myocardial stunning [30,31]. A recent report revealed that regional ejection fractions correlated significantly with regional perfusion in the anterior walls (r = 0.71, P = 0.0001), the lateral walls (r = 0.66, P = 0.0001) and inferior walls (r = 0.54, P = 0.0004). However, weaker correlation was found in anteroseptal and inferoseptal walls (r = 0.26 and 0.44 with P = 0.11 and < 0.005, respectively) [32]. These results agreed with our findings, as the global ejection fraction correlated significantly with the defect extent and demonstrated correlation values averaged over the higher and lower regional correlations. However, these observations may need further research due to the small number of patients represented in the SSS ranges of 4–8 and 9–13. Comparing the gated SPECT methods by RWT provided important information and could potentially avoid some limitations of ‘gold standard’ techniques as in MRI, such as breath holding versus no breath holding, inclusion and

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exclusion of papillary muscles and cardiac trabeculae, and inter-observer variation in myocardial tracing. A recent report has indicated that the inclusion and exclusion of the myocardial trabeculae confound the accuracy of MRI in estimating the left ventricular volumes and consequently the computation of the ejection fraction [33]. Based on our results and prior studies in evaluating the gated SPECT methods, it is important to carry out onsite evaluation of the program used routinely in the nuclear medicine clinic. This is not method validation: rather, it allows the determination of the degrees of variation that may occur in clinical practice. Each gated SPECT method postulates assumptions in an attempt to accurately model the left ventricle and hence to precisely extract the functional information from the acquired data. However, the underlying assumptions and the dimensional approximations may not meet the realistic conditions in all situations. Although the correlations between the methods evaluated here ranged from mild to good, using any pair of programs interchangeably is very limited. Furthermore, the agreement limits are large and exceed the serial reproducibility of the test (B5%) [30]. Drawbacks of the ‘regression without truth’ technique

The assumption that the ejection fraction could be represented as unimodal is the subject of argument as patients with normal ejection fractions may have certain peaks that significantly differ from those with abnormal ejection fractions. To avoid the contribution of such limitations, we have selected those patients with abnormal myocardial perfusion to approach the unimodal assumption as far as possible [17,18]. The truncated normal and beta distributions, bounded between 1 and zero, were assumed for the ‘gold standard’. However, distributions that encompass the entire real line may be more challenging as stated by Kupinski et al. [17]. Moreover, the model assumed was a linear relationship between the ‘gold standard’ and the estimated values. This is not always valid as other mathematical relationships could show better representation of the true values. The authors, accordingly, have kept room for further investigations in order to consider other possibilities [17]. Since many nuclear medicine professionals are familiar with investigations using gated SPECT methods with medical standards, accepting the RWT technique may be a little confusing. Nevertheless, the results presented here are in agreement with previous reports.

Conclusion Our study showed considerable variation among the four quantitative gated SPECT methods (ECTb, QGS, LMC and LVGTF) in the evaluation of the left ventricular

ejection fraction. The ‘regression without truth’ technique revealed that ECTb and QGS outperformed the other two methods with respect to the bias and precision of the measurements. However, ECTb was slightly better than QGS, and LMC was slightly better than the LVGTF method. Pair-wise correlations of the four methods ranged from mild to good with large agreement limits. The best correlation was found between QGS and ECTb in the estimation of volumes and ejection fractions. ECTb, QGS and LVGTF correlated significantly with defect extent. Interchangeability of the gated SPECT methods to manage a patient’s therapy or follow-up is not recommended. Results of the RWT technique are consistent with previous reports but further research is warranted to thoroughly utilize the technique in the presence of medically relevant standards.

Acknowledgements We thank Dr Grant Gullberg, E.O. Lawrence Berkeley National Laboratory, Berkeley, USA, and Dr Bing Feng, Department of Radiology, University of Massachusetts, Worcester, for providing us with the software code of the layer of maximum counts method. We acknowledge the technical support given by the technologist of Mubarak Al-Kabeer Hospital, Ministry of Health, Kuwait.

References 1

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Appendix Regression without truth technique assumes that there is a linear relationship between the true ejection fraction and its estimated value ypm by the formula ypm ¼ am yp þ bm þ epm

ð1Þ

where am, bm and epm are the slope, intercept and error terms of the regression line measured by method m. Three important assumptions were given to continue the theoretical derivation of the technique. First, the true ejection fraction measure y does not vary for a given patient across methods and is statistically independent from patient to patient. Second, the regression parameters am and bm are characteristic of the methods and independent of the patient. Third, the error terms epm are statistically independent and normally distributed with zero mean and variance s2m . By using this latter assumption, the probability density function for the noise term for a given patient p and M modalities could be formulated and solved in Equation 1. Then, the probability of the estimated ejection fractions for multiple methods and a specific patient (given the linear model parameters am, bm and sm, and the true ejection fraction) could be written as 
  ypm j am ; bm ; s2m ; yp   M Y 2 1 1  pffiffiffiffiffiffiffiffiffiffiffi exp  ¼ ypm  am yp  bm : 2ps2m 2ps2m m¼1

pr




ð2Þ

The notation y represents the estimated ejection fractions for a given patient p over M methods. By using the conditional probability and marginal probability law, the probability of the estimated ejection fraction for a specific patient across all methods given the linear model parameters becomes 
 ypm j am ; bm; s2m " # Z M X   2 1  ¼ dyp pr yp :S exp  2 ypm  am yp  bm 2sm m¼1

pr




ð3Þ where S¼

M Y

1 pffiffiffiffiffiffiffiffiffiffiffi : 2ps2m m¼1

From the first assumption and considering the total number of patients, Ptot, the likelihood of the linear

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332 Nuclear Medicine Communications 2006, Vol 27 No 4

model parameters can be expressed as Ptot Z Y dyp prðyp Þ L¼ p¼1

"



#) 2 1 1  exp  2  2 ypm  am yp  bm 2sm 2sm m¼1 M X

Taking the log and rewriting the products as sums, we obtain the log likelihood of the probability function l ¼ lnðLÞ ¼Ptot lnðSÞ þ "

Z Ptot X ln dyp prðyp Þ

values determined with the maximum likelihood estimation characterize the relationship between the estimates and the ‘gold standard’ of each modality, the noise in these estimates, and the distribution of the true values for the patient population. Maximum-likelihood estimators produce results by an iterative procedure which is implemented by a code written in the Matlab software package (Mathworks Inc.) and executed in a personal computer. The quasi-Newton optimization method was used to determine the maximum of the likelihood. (This appendix is summarized from the original papers by Kupinski et al. [17] and Hoppin et al. [18].).

p¼1

#) M X 2 1  exp  2 ypm  am bp  bm 2sm m¼1 The scalar quantity l is the maximum likelihood of the model parameters (am, bm and s2m ) that need to be determined for each method. Once again the parameter

The quasi-Newton optimization method was used to determine the maximum of the likelihood. We summarized the theoretical derivation assuming normal distribution; however, beta distribution is similar with changes in the formula representing the probability density function. (This appendix is summarized from the original papers by Kupinski et al. [17] and Hoppin et al. [18].).

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Original article

Prognostic value of normal exercise 99mTc-sestamibi myocardial tomography in patients with angiographic coronary artery disease Min-Fu Yanga, Ke-Fei Doub, Xiu-Jie Liua, Yue-Jin Yangb and Zuo-Xiang Hea Background and aim Previous studies have documented the prognostic value of normal exercise 201Tl myocardial perfusion imaging in patients with angiographic coronary artery disease (CAD). However, data on exercise 99m Tc-sestamibi myocardial single photon emission computed tomography (SPECT) are scant. Accordingly, the purpose of this study was to investigate the prognostic value of normal exercise 99mTc-sestamibi SPECT in patients with angiographic CAD. Methods We retrospectively investigated 90 consecutive patients who had a normal exercise 99mTc-sestamibi myocardial SPECT but angiographic CAD. A group of 69 consecutive patients with both normal exercise 99m Tc-sestamibi myocardial SPECT and coronary arteries were included as control. Results During a mean follow-up of 50 ± 19 months, a total of three hard cardiac events (non-fatal myocardial infarction) and seven soft cardiac events (late revascularization) were observed. The annual hard cardiac event rate between the two groups was not significantly different (0.6% vs. 0.3%, v2 = 0.47, P = NS), nevertheless the annual soft cardiac event rate was higher in patients with angiographic CAD (1.9% vs. 0, v2 = 5.74, P = 0.02). Moreover, the annual hard cardiac events rate in patients with angiographic CAD who were treated medically was also not

Introduction Stress myocardial perfusion imaging is currently the most reliable and non-invasive method for the detection of coronary artery disease (CAD). It also provides important prognostic information in patients with known or suspected CAD [1]. A large number of studies have documented that patients with a normal stress myocardial perfusion imaging with 201Tl or 99mTc-sestamibi had an annual rate of non-fatal myocardial infarction or cardiac death < 1% [2–5]. Several studies have demonstrated that patients with a normal exercise 201Tl myocardial perfusion imaging but angiographic CAD have a low incidence of cardiac events (non-fatal myocardial infarction or cardiac death) during short-term or long-term follow-up [6–8]. Data on exercise 99m Tc-sestamibi myocardial perfusion imaging in such patients are scanty, however. Accordingly, the purpose of

significantly different from that of the control group (0.8% vs. 0.3%, v2 = 0.77, P = NS). Among patients with angiographic CAD, the annual hard cardiac event rate was not statistically different between those treated medically and those who underwent revascularization (0.8% vs. 0, v2 = 0.53, P = NS). Conclusions Our data demonstrate that normal exercise 99m Tc-sestamibi myocardial SPECT despite angiographic CAD suggests a low rate of cardiac death or non-fatal myocardial infarction but a relatively high rate of late revascularization during an intermediate term of c 2006 follow-up. Nucl Med Commun 27:333–338 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:333–338 Keywords: exercise test, radionuclide imaging, coronary artery disease, prognosis Department of aNuclear Medicine and bCardiology, Cardiovascular Institute and Fu Wai Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China. Correspondence to Professor Zuo-Xiang He, Department of Nuclear Medicine, Medical Imaging Center, Fu Wai Hospital, CAMS and PUMC, 167, Bei Li Shi Lu, Beijing 100037, China. Tel: + 0086 10 883 98967; fax: + 0086 10 683 31769; e-mail: [email protected] Received 14 August 2005 Accepted 5 January 2006

this study was to investigate the clinical outcome of patients with a normal exercise 99mTc-sestamibi SPECT but angiographically significant CAD.

Materials and methods Study population

From our inpatient database, we retrospectively reviewed all patients who had had a normal exercise 99mTcsestamibi myocardial SPECT but angiographically significant CAD between January 1993 and March 2001. A control group was composed of the consecutive patients with a normal exercise 99mTc-sestamibi myocardial SPECT and angiographically normal coronary arteries between January 1994 and June 1998. All of the patients were referred for the evaluation of chest pain. Patients were excluded if they had one or more of the following conditions: valvular heart disease, prior Q-wave myocardial infarction, left bundle branch block, previous history

c 2006 Lippincott Williams & Wilkins 0143-3636

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Nuclear Medicine Communications 2006, Vol 27 No 4

of revascularization (percutaneous coronary intervention or coronary artery bypass grafting surgery) or idiopathic cardiomyopathy. Exercise testing

All patients underwent a symptom-limited exercise test on a bicycle ergometer. Anti-anginal medications were not routinely discontinued. Exercise was terminated if one or more of the following conditions occurred: unable to exercise for angina pectoris or dyspnea, systolic blood pressure depressed Z 15 mmHg compared with the baseline, or Z 2 mm ST-segment depression. Positive ECG was defined as Z 1 mm of horizontal or downsloping ST-segment depression or Z 1.5 mm of upsloping, lasting at least 80 ms after the J point. 99mTcsestamibi (740–925 MBq) was intravenously injected at the peak exercise and the patients were ordered to exercise for an additional 1 min. Blood pressure, heart rate and ECG were continuously monitored and recorded at baseline, at each level of exercise, and during the recovery period [9]. Myocardial SPECT imaging 99m

Tc-sestamibi myocardial SPECT imaging was performed 1–1.5 h after the exercise testing. Image acquisition was performed using a single-head or triple-head SPECT equipped with a general purpose, parallel-hole collimator. Projection data were acquired over a 1801 anterior arc from 451 right anterior oblique view to 451 left posterior oblique at 61 intervals and with an acquisition time of 40 s per frame when a single-head SPECT was used, and over a 3601 arc (1201 arc per detector) at 61 interval with an acquisition time of 40 s per frame when a triple-head SPECT was used. The myocardial SPECT imaging was visually analysed by two experienced nuclear medicine physicians who were unaware of the angiographic results. A normal scan was interpreted as no fixed or reversible perfusion defects.

Coronary angiography

Coronary angiography was performed using standard techniques within 90 days (mean ± SD, 20 ± 2 days) of the exercise myocardial SPECT imaging. During that period, none of the patients suffered cardiac events. Catheterization was interpreted by two experienced interventional cardiologists. Angiographic CAD was diagnosed when Z 50% luminal diameter stenosis was present in at least one of the three major coronary arteries or their main branches. Proximal lesions included lesions of the left main coronary artery, the lesions of the left anterior descending (LAD) artery before the first septal or the first diagonal branch, lesions of the left common circumflex (LCx) artery and lesions of the right coronary artery (RCA) before the crux, the remaining coronary artery lesions were considered as distal lesions [8]. Left ventricular ejection fraction (LVEF) was calculated according to the left ventriculography.

Follow-up

All patients, their physicians or relatives were contacted by telephone or letter. The questionnaire specifically addressed the occurrence of sudden and non-sudden cardiac death, non-fatal myocardial infarction and revascularization. Medical records of subsequent hospitalizations were reviewed to confirm events. The period of follow-up was estimated according to the last contact. The follow-up was finished if a patient developed a hard cardiac event (non-fatal myocardial infarction or cardiac death), but continued if soft cardiac event (late revascularization, i.e., > 3 months after the initial myocardial imaging) occurred. Statistical analysis

All variables were reported as mean ± SD or frequencies when appropriate. The Student’s t-test was used to compare mean difference of the continuous variables between two groups. Likewise, the chi-squared test was used to compare categorized variables between the two groups. Survival curves were constructed using the Kaplan–Meier method and were compared with the logrank test. The SPSS 10.0 software package was used for the statistical analysis. A P value of < 0.05 was considered statistically significant.

Results Patient characteristics

There were a total of 90 patients with a normal exercise 99m Tc-sestamibi myocardial SPECT but angiographic CAD and 69 patients with both a normal exercise 99m Tc-sestamibi myocardial SPECT and angiographically normal coronary arteries. Follow-up was completed in 97.5% of the patients (four patients were lost during follow-up, two in each group). Patients with complete follow-up were recruited into the final analysis. Their demographic data are summarized in Table 1. Patients with or without angiographic CAD had similar frequencies of family history of CAD, diabetes mellitus and hyperlipidemia. LVEF was normal (50%) in all patients and not significantly different between the two groups. However, patients with documented CAD were older, and more patients with documented CAD were men and had hypertension and were cigarette smokers. Patients with or without angiographic CAD had similar exercise duration. However, more patients with angiographic CAD had anti-anginal medications at the time of exercise testing. More patients with angiographic CAD experienced chest pain during exercise. The peak heart rate was lower in patients with angiographic CAD. Angiographic data

In the group of patients with angiographic CAD, onevessel disease was documented in 52 (59%) patients, two-

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Outcome of patients with normal exercise myocardial SPECT but angiographic CAD Yang et al. 335

Table 1

Patient’s characteristics

Characteristic

Men Age (years) LVEF (%) Diabetes Hypertension Hypercholesterolaemia Cigarette smoking Family history of CAD Anti-angina medications Exercise grades Total exercise time (min) Peak HR (bpm) % MPHR Chest pain Exercise ECG ( + )

Presence of angio- Absence of angiographic CAD graphic CAD (n = 88) (n = 67)

P value

72 (82) 56 ± 8 75 ± 8 9 (10) 50 (57) 28 (32)

45 (68) 52 ± 9 73 ± 7 4 (6) 27 (40) 26 (39)

NS 0.003 NS NS 0.042 NS

44 (50) 37 (42)

22 (33) 27 (40)

0.032 NS

22 (25)

8 (12)

0.049

3±1 9±3

3±1 9±3

NS NS

132 ± 17 81 ± 12 11 (13) 13 (15)

144 ± 18 86 ± 11 2 (3) 6 (9)

0.0001 0.016 0.034 NS

MPHR = maximal predicted heart rate; NS = not statistically significant; LVEF = left ventricular ejection fraction; CAD, coronary artery disease. Figures in parentheses are presented as percentage.

Table 2 Angiographic data in patients with angiographic coronary artery disease

Number of patients Diseased coronary arteries Left main artery LAD LCx RCA Number of diseased vessels 1 2 3 Location of lesions Proximal Distal Extent of stenosis Z 95% Z 70% 50B70%

Revascularization

Medical therapy

Total

24

64

88

4 19 12 10

2 49 21 27

6 68 33 37

12 7 5

40 15 9

52 22 14

18 6

24 40

42 46

4 22 2

4 37 27

8 59 29

vessel disease in 22 (25%) patients and three-vessel disease in only 14 (16%) patients. Only 9% of those patients had subtotal or total coronary occlusion (Table 2). Follow-up

During 50 ± 19 months of follow-up, most (131/155) of the patients were treated medically. There were three hard cardiac events (all non-fatal myocardial infarctions), two in the group of patients with angiographic CAD, and one in the control group (Table 3). Twenty-four patients with angiographic CAD underwent coronary revascularization (20 for percutaneous coronary intervention, four for coronary artery bypass grafting surgery), seven of them

were late revascularization. None of the patients in the control group suffered from revascularization. The annual hard cardiac events rate was low and not significantly different between the two groups (0.6% vs. 0.3%, w2 = 0.47, P = NS) (Fig. 1). Nevertheless, the annual soft cardiac event rate was higher in patients with angiographic CAD (1.9% vs. 0, w2 = 5.74, P = 0.02) (Fig. 2). Moreover, the annual hard cardiac event rate in patients with angiographic CAD who were treated medically was also not significantly different from that of the control group (0.8% vs. 0.3%, w2 = 0.77, P = NS) (Fig. 3). Among patients with angiographic CAD, the annual hard cardiac event rate was not statistically different between those treated medically and those who underwent revascularization (0.8% vs. 0, w2 = 0.53, P = NS) (Fig. 4).

Discussion To the best of our knowledge, this is the first study to evaluate the prognostic value of normal exercise 99mTcsestamibi scintigraphy in patients with angiographically documented CAD. Our data demonstrate that patients with a normal exercise 99mTc-sestamibi myocardial SPECT despite of angiographic CAD are at low risk for cardiac death or non-fatal myocardial infarction under the crossover with revascularization. This is similar to patients with normal exercise 201Tl myocardial perfusion imaging and angiographic coronary arteries reported in the previous studies. Several investigators have reported outcomes of patients with normal exercise 201Tl myocardial perfusion imaging but angiographic CAD. Brown et al. [6] firstly investigated 75 patients who underwent planar 201Tl scintigraphy for 2 years, and found only one cardiac event (myocardial infarction). It is regretable that the author did not provide information about revascularization during followup. In another study, Fattah et al. [7] investigated 97 patients with normal 201Tl myocardial SPECT but angiographic CAD who were followed for 32 months. Of these patients, 21 underwent revascularization. During the follow-up, an annual event rate of 1.1% was observed (two cardiac deaths and one non-fatal myocardial infarction). Pavin et al. [8] evaluated the long-term prognosis of 69 patients with normal 201Tl scintigraphy and angiographic CAD. Over a mean follow-up period of 8.6 years, most patients (65%) were treated medically and the others underwent revascularization, one fatal and eight non-fatal cases of myocardial infarction were observed. There was no significant difference in the overall incidence of combined major cardiac events (cardiac death and non-fatal myocardial infarction) in these patients compared with those with both normal exercise scintigraphy and angiographically normal coronary arteries (13% vs. 4%, P = NS). However, while the cardiac mortality remained very low (1.4%), the incidence of

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Table 3

Hard cardiac events

Patient

Gender

Age (years)

% MPHR

Exercise ECG

Angiography

Revascularization

Follow-up (months)

Cardiac event

ZM WLC CYL

Male Male Female

41 49 62

81 85 72

(0) (0) (0)

Three-vessel disease RCA Normal

(0) (0) (0)

21 40 53

MI MI MI

MI, myocardial infarction; MPHR, maximal predicted heart rate.

Fig. 3

Fig. 1

1

1

0.95 Hard event-free survival

Hard event-free survival

0.95 Absence of CAD Presence of CAD

0.9 0.85

Log rank
2 = 0.47, P = NS

0.8

Absence of CAD

0.9

Presence of CAD treated medically 0.85 0.8

Log rank 2
= 0.77, P = NS

0.75

0.75

0

20

40 60 Follow-up (months)

80

0

100

20

40 60 Follow-up (months)

80

Hard event-free survival curves of the patients with angiographic coronary artery disease treated medically and the control group.

Fig. 2

Fig. 4

1

1

0.95

0.95

0.9

Hard event-free survival

Soft event-free survival

Hard event-free survival curves of the patients with and without angiographic coronary artery disease.

Absence of CAD Presence of CAD

0.85 Log rank 2
= 5.74, P = 0.02

0.8 0.75

Presence of CAD underwent revascularization

0.9

Presence of CAD treated medically 0.85 0.8 Log rank
2 = 0.53, P = NS

0.75

0

20

40 60 Follow-up (months)

80

100

100

0

20

40 60 Follow-up (months)

80

100

Soft event-free survival curves of the patients with and without angiographic coronary artery disease.

Hard event-free survival curves of the patients with angiographic coronary artery disease treated medically and who underwent revascularization.

non-fatal myocardial infarction in these patients was similar to that with abnormal exercise scintigraphy and angiographical CAD. Therefore, the good long-term

prognosis of these patients is mainly due to the very low cardiac mortality, which is comparable to that in the patients without CAD.

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Outcome of patients with normal exercise myocardial SPECT but angiographic CAD Yang et al. 337

Although many researches clarified that a normal stress myocardial perfusion imaging suggests benign prognosis in patients with suspected or known CAD and adds incremental prognostic value over clinical or angiographic factors [10], some other studies reminded us that determination of outcomes in these patients is complicated and may be influenced by other clinical factors. Patients who underwent pharmacological stress myocardial perfusion imaging, had previous CAD (myocardial infarction or revascularization), or had ischaemic ECG findings during stress testing would experience higher risk of cardiac events than those who underwent exercise stress, had no history of CAD or had no ischaemic ECG findings during stress [11–14]. Moreover, some investigators found the cardiac event rate after a normal stress myocardial perfusion imaging might change with time. Elhendy et al. [15] studied 218 patients who had normal myocardial perfusion assessed by exercise 99mTc-sestamibi SPECT for a follow-up period of 7.4 years, found the annual mortality rate was 0.6% in the first 5 years and 1.8% between the sixth and eighth years, the annual cardiac event rate was 0.7% in the first 5 years and 1.5% between the sixth and eighth years. Hachamovitch et al. [16] performed a large-scale follow-up of 7376 consecutive patients with normal exercise or adenosine myocardial tomographic imaging, and revealed that in patients without previous CAD the level of risk (non-fatal myocardial infarction and cardiac death) was uniform with time, but in patients with known CAD, risk increased with time (e.g., risk in the first year was less than in the second year, hence, a dynamic temporal component of risk was present). Due to its retrospective characters of this study, we can not precisely know to what extent coronary revascularization contributed to the benign outcome of these patients with angiographically significant CAD. In the present study, 27% patients with angiographic CAD received revascularization after initial myocardial perfusion imaging. Like most of other prognostic studies in patients with normal myocardial perfusion imaging, the favourable outcome of this study was obtained under the crossover with revascularization. Furthermore, some studies have demonstrated that anti-anginal medications could significantly reduce the presence, extent and severity of defects on the exercise or pharmacological myocardial perfusion imaging [17–19]. Although 25% of patients with angiographic documented CAD in this study had antianginal medications before exercise testing, the overall annual hard cardiac event rate was very low (0.6%). The effect of anti-anginal medications on the prognostic evaluation by myocardial perfusion imaging needs further clarification [20].

Conclusion Our data demonstrate that a normal exercise 99mTcsestamibi myocardial SPECT despite of angiographic CAD suggests a low rate of cardiac death or non-fatal myocardial infarction but a relatively high rate of late revascularization during intermediate term of follow-up.

References 1

2 3 4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

Beller GA, Zaret BL. Contributions of nuclear cardiology to diagnosis and prognosis of patients with coronary artery disease. Circulation 2000; 101:1465–1478. Brown KA. Prognostic value of thallium-201 myocardial perfusion imaging: a diagnostic tool comes of age. Circulation 1991; 83:363–381. Brown KA, Altland E, Rowen M. Prognostic value of normal technetium-99m sestamibi cardiac imaging. J Nucl Med 1994; 35:554–557. Soman P, Parsons A, Lahiri N, Lahiri A. The prognostic value of a normal Tc-99m sestamibi SPECT study in suspected coronary artery disease. J Nucl Cardiol 1999; 6:252–256. Raiker K, Sinuas AJ, Wackers FJTh, Zaret BL. One year prognosis of patients with normal planar or single-photon emission computed tomographic technetium 99m-labeled sestamibi exercise imaging. J Nucl Cardiol 1994; 1:449–456. Brown KA, Rowen M. Prognostic value of a normal exercise myocardial perfusion imaging study in patients with angiographically significant coronary artery disease. Am J Cardiol 1993; 71:865–867. Fattah AA, Kamal AM, Pancholy S, Ghods M, Russell J, Cassel D. Prognostic implications of normal exercise tomographic thallium images in patients with angiographic evidence of significant coronary artery disease. Am J Cardiol 1994; 74:769–771. Pavin D, Delonca J, Siegenthaler M, Doat M, Rutishauser W, Righetti A. Long-term (10 years) prognostic value of a normal thallium-201 myocardial exercise scintigraphy in patients with coronary artery disease documented by angiography. Eur Heart J 1997; 18:69–77. He ZX, Liu YZ, Shi RF. Clinical evaluation of 99mTc-MIBI myocardial tomography for detecting coronary artery disease. Chin Med Sci J 1992; 7:1–4. Hachamovitch R, Berman DS, Shaw LJ, Kiat H, Cohen I, Cabico JA, et al. Incremental prognostic value of myocardial perfusion single photon emission computed tomography for the prediction of cardiac death: differential stratification for risk of cardiac death and myocardial infarction. Circulation 1998; 97:535–543. Heller GV, Herman SD, Travin MI, Baron JI, Santos-Ocampo C, McClellan JR. Independent prognostic value of intravenous dipyridamole with technetium-99m sestamibi tomographic imaging in predicting cardiac events and cardiac-related hospital admission. J Am Coll Cardiol 1995; 26:1202–1208. Hendel RC, Layden JJ, Leppo JA. Prognostic value of dipyridamole thallium scintigraphy for evaluation of ischemic heart disease. J Am Coll Cardiol 1990; 15:109–116. Klodas E, Miller TD, Christian TF, Hodge DO, Gibbons RJ. Prognostic significance of ischemic electrocardiographic changes during vasodilator stress testing in patients with normal SPECT images. J Nucl Cardiol 2003; 10:4–8. Abbott BG, Afshar M, Berger AK, Wackers FJ. Prognostic significance of ischemic electrocardiographic changes during adenosine infusion in patients with normal myocardial perfusion imaging. J Nucl Cardiol 2003; 10:9–16. Elhendy A, Schinkel A, Bax JJ, van Domburg RT, Poldermans D. Long-term prognosis after a normal exercise stress Tc-99m sestamibi SPECT study. J Nucl Cardiol 2003; 10:261–266. Hachamovitch R, Hayes S, Friedman JD, Cohen I, Shaw LJ, Germano G, et al. Determinants of risk and its temporal variation in patients with normal stress myocardial perfusion scans. J Am Coll Cardiol 2003; 41:1329–1340. Mahmarian JJ, Fenimore NL, Marks GF, Francis MJ, Morales-Ballejo H, Verani MS, et al. Transdermal nitroglycerin patch therapy reduces the extent of exercise-induced myocardial ischemia: results of a double-blind, placebo-controlled trial using quantitative thallium-201 tomography. J Am Coll Cardiol 1994; 24:25–32. Sharir T, Rabinowitz B, Livschitz S, Moalem I, Baron J, Kaplinsky E, et al. Underestimation of extent and severity of coronary artery disease by dipyridamole stress thallium-201 single-photon emission computed

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tomographic myocardial perfusion imaging in patients taking antianginal drugs. J Am Coll Cardiol 1998; 31:1540–1546. 19 Taillefer R, Ahlberg AW, Masood Y, White CM, Lamargese I, Mather JF, et al. Acute beta-blockade reduces the extent and severity of myocardial perfusion defects with dipyridamole Tc-99m sestamibi SPECT imaging. J Am Coll Cardiol 2003; 42:1475–1483.

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Original article

Computed tomography-based attenuation correction in neurological positron emission tomography: evaluation of the effect of the X-ray tube voltage on quantitative analysis Mohammad Reza Ay and Habib Zaidi Background The advent of dual-modality positron emission tomography/computed tomography (PET/CT) imaging has revolutionized the practice of clinical oncology by improving lesion localization and facilitating treatment planning for radiation therapy. In addition, the use of CT images for CT-based attenuation correction (CTAC) allows the overall scanning time to be decreased and a noise-free attenuation map (lmap) to be created. The most common procedure requires a piecewise linear calibration curve acquired under standard imaging conditions to convert the patient’s CT image from low effective CT energy into an attenuation map at 511 keV. Aim To evaluate the effect of the tube voltage on the accuracy of CTAC. Methods As different tube voltages are employed in current PET/CT scanning protocols, depending on the size of the patient and the region under study, the impact of using a single calibration curve on the accuracy of CTAC for images acquired at different tube voltages was investigated through quantitative analysis of the created lmaps, generated attenuation correction factors and reconstructed neurological PET data using anthropomorphic experimental phantom and clinical studies. Results For CT images acquired at 80 and 140 kVp, average relative differences of – 2.9% and 0.7%,

Introduction The advent of combined positron emission tomography/ computed tomography (PET/CT) units is considered as a major advance in medical imaging technology and health care. As the name implies, PET/CT combines the information produced by two sophisticated imaging modalities, the functional information from PET with the anatomical information from CT, into a single procedure [1]. Dual-modality imaging correlates functional and anatomical data to improve disease localization and facilitates treatment planning for radiation oncology or surgery [2]. PET/CT systems offer significant advantages over stand-alone PET, including decreased overall scanning time and increased accuracy in lesion localization. The high-resolution anatomical information from PET/CT improves the differentiation of the physiological (normal) uptake of 18F-fluorodeoxyglucose ([18F]FDG)

respectively, from the images acquired at 120 kVp were observed for the absolute activity concentrations in five regions of the anthropomorphic striatal phantom when CT images were converted using a single calibration curve derived at 120 kVp. Likewise, average relative differences of 1.9% and – 0.6% were observed when CT images were acquired at 120 kVp and CTAC used calibration curves derived at 80 and 140 kVp, respectively. Conclusion The use of a single calibration curve acquired under standard imaging conditions does not affect, to a visible or measurable extent, neurological PET images reconstructed using CTAC when CT images are acquired in different conditions. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:339–346 Nuclear Medicine Communications 2006, 27:339–346 Keywords: attenuation correction, brain imaging, PET/CT, quantification, tube voltage

Division of Nuclear Medicine, Geneva University Hospital, Geneva, Switzerland. Correspondence to Habib Zaidi, Division of Nuclear Medicine, Geneva University Hospital, CH-1211 Geneva 4, Switzerland. Tel: + 41 22 372 7258; fax: + 41 22 372 7169; e-mail: [email protected] Sponsorship: This work was supported by the Swiss National Science Foundation under grant SNSF 3152A0-102143. Received 22 November 2005 Accepted 13 January 2006

and other radiopharmaceuticals from that associated with disease, thereby reducing false-positive errors in comparison with lesion characterization using PET imaging alone. Several physical factors can degrade the image quality and quantitative analysis of PET: the most important is photon attenuation in tissues, which can affect both the visual interpretation and quantitative analysis of PET data [3]. Several transmissionless methods (that do not require a transmission scan) have been devised to correct for attenuation in neurological PET studies [4–7]. One of the advantages of PET/CT is the ability to generate a noise-free attenuation map (mmap) to be used for attenuation correction purposes. With the introduction of hybrid PET/CT systems into the clinical setting, precise conversion from CT numbers derived from low-

c 2006 Lippincott Williams & Wilkins 0143-3636

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340 Nuclear Medicine Communications 2006, Vol 27 No 4

energy polyenergetic X-ray spectra to linear attenuation coefficients at 511 keV has become essential in order to apply accurate CT-based attenuation correction (CTAC) to the PET data. Several CTAC strategies have been developed, including scaling [8], segmentation [9], hybrid segmentation/scaling [10], piecewise linear scaling [11,12] and dual-energy decomposition methods [13]. Most commercially available PET/CT scanners use the bilinear calibration curve method, which is generally calculated at a preset tube voltage (120–140 kVp) and current. Kamel et al. [14] investigated the effect of varying tube current, and showed that a low-current CT is sufficient for CTAC using comparative quantitative analysis of reconstructed clinical PET images. As patient CT images may be acquired at different tube voltages and currents, depending on the patient size and region under study, and considering the fact that the CT number of a particular tissue is tube voltage dependent, it was hypothesized that the use of a single calibration curve calculated at a specific tube voltage for CT images acquired under different scanning conditions might propagate a significant uncertainty during the CTAC procedure. Bai et al. [12] argued that the slope of the bilinear calibration curve for CT numbers higher than 0 Hounsfield units (HU) was tube voltage dependent. Other studies have reported on the relevance of deriving tube voltage-dependent CTAC schemes for PET/CT [15]. These topical developments, combined with the lack of detailed studies investigating the effect of tube voltage on the quantitative analysis of non-clinical PET data, where the ground truth is known, motivated the work presented in this paper. This study was designed to provide answers to the following legitimate questions of the clinician or physicist: ‘what is the magnitude of error of acquiring CT at, for example, 80 kVp when the calibration curve is the manufacturer’s standard of 120 or 140 kVp?’ or, vice versa, ‘what is the magnitude of error of acquiring CT images at specific tube voltages and varying the voltage for the derivation of calibration curves’. The assessment was carried out through quantitative analysis of created mmaps, generated attenuation correction factors (ACFs) and reconstructed neurological PET data using experimental anthropomorphic phantom and clinical studies.

Materials and methods Phantom and clinical studies

Our department is in the process of installing a commercial PET/CT scanner; however, this was not available during the actual study design, which relied on the use of PET and CT data acquired on separate PET and CT scanners. One of the motivations behind the choice of brain imaging is that automated multimodality coregistration algorithms work relatively well (in contrast with whole-body imaging) and can be applied most

successfully to neurological studies, where the skull provides a rigid structure that maintains the geometrical relationship of structures within the brain. An anthropomorphic head phantom (Radiology Support Devices Inc., Long Beach, California, USA), designed specifically for the assessment of quantitative imaging capabilities of the striatum relevant for PET studies of the presynaptic and postsynaptic dopaminergic system, was employed in order to quantitatively assess the effect of using a single calibration curve on the accuracy of CTAC when CT images were acquired at different tube voltages. The phantom has five compartments which can be filled separately: left and right caudate nucleus (LCN and RCN), left and right putamen (LPU and RPU) and the rest of the brain (main chamber). The main chamber itself is embedded in a bone-like structure to provide properties similar to the human head. For an activity ratio of 1 : 8 between the main chamber and small cavities, 2.942 MBq of 18F (in 0.9 ml of 0.9% NaCl) was diluted in distilled water and used to fill the striatum. A total activity of 13.2 MBq diluted in 1.1 ml of NaCl was added to the main chamber. Subsequently, the main chamber was totally filled with distilled water. The fully three-dimensional emission study lasted 25 min, whereas a two-dimensional pre-injection transmission scan (10 min) was acquired using 137Cs single-photon point sources on an ECAT ART PET scanner (CTI/Siemens, Knoxville, Tennessee, USA). This is a whole-body, three-dimensional tomograph having less than one-half the number of bismuth germanate detectors compared with a full ring scanner with the equivalent field of view, upgraded to use collimated point sources of 137Cs, and is capable of producing high-quality, scatter-free data in this continuously rotating partial-ring tomograph. The measured transmission scan was scaled for the difference between 662 and 511 keV energies by normalizing to a slab phantom scan and correcting for scatter and crosssection variation using a log-linear transformation of the attenuation factors. Thereafter, CT data of the same phantom were acquired on an Aquilion CT scanner (Toshiba Medical Systems Corporation, Tokyo, Japan) at 240 mA in order to apply the CTAC procedure to the emission data. This CT scanner has 40 parallel detector rows with 35 840 detector elements, 32 mm detector length along the patient axis and offers 16 slices (0.5 mm thickness) with each 0.5 s gantry revolution. Calibration curves calculated at 80, 120 and 140 kVp were used to create sets of mmaps from images acquired at 80, 120 and 140 kVp. The emission data were corrected for attenuation using the measured transmission method (MTM) as well as the set of mmaps generated using CTAC. Apparent recovery coefficients (ARCs), representing the apparent (observed or partial volume corrected) regional radioactivity concentration to true activity ratio, and absolute activity concentrations were calculated for the five compartments of the striatal phantom as figures of merit

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CT-based attenuation correction in PET Ay and Zaidi 341

for the quantitative analysis of reconstructed neurological PET images. A polyethylene cylindrical phantom (f = 250 ± 0.5 mm), containing 16 cylindrical holes (f = 20 ± 0.5 mm), was made in order to calculate the bilinear calibration curves required for application of the CTAC procedure. Fourteen syringes were filled with a solution of K2HPO4 and water, with concentrations varying between 50 and 900 mg/cm3 to simulate cortical bone with different densities. The prepared syringes, together with two additional syringes containing water and air, were inserted into the polyethylene phantom’s holes (see Fig. 1). Subsequently, the phantom was scanned on a HiSpeed X/iF CT scanner (General Electric Healthcare Technologies, Waukesha, Wisconsin, USA) using three different tube voltages (80, 120 and 140 kVp). This CT scanner uses a Highlight (Y2Gd2O3:Eu) ceramic scintillator and has a 541 mm source to isocentre and 949 mm source to detector distances and 816 detector elements (793 active elements) with a physical dimension of 0.8 mm. The bilinear calibration curves at different tube voltages for both CT scanners were calculated according to the method proposed by Bai et al. [12]. Patient brain CT scans acquired at 120 kVp on the HiSpeed X/iF CT scanner were selected from the database and used for the clinical evaluation of the effect of the tube voltage. Attenuation correction and image reconstruction

The computation of ACFs derived from CTAC involved down-sampling the CT image matrix to 128  128, followed by Gaussian smoothing using a 6 mm kernel, to match the spatial resolution of the PET scanner employed in this study. CT numbers (in HU) were then transformed to linear attenuation coefficients at 511 keV

using the calculated bilinear curve. The created mmaps were forward projected to generate 47 ACF sinograms. The attenuation-corrected projections were reconstructed using the 3DRP reprojection algorithm implemented within ECAT 7.2.1 software (CTI Molecular Imaging Inc., Knoxville, Tennessee, USA) with a maximum acceptance angle corresponding to 17 rings and a span of 7. The default parameters used in clinical routine were applied (ramp filter; cut-off frequency, 0.35 cycles/ pixel). The reconstructed images consist of 47 slices with 128  128 resolution and a voxel size set to 1.72  1.72  3.4 mm3. The acquired CT and preliminary PET images reconstructed using calculated attenuation correction were coregistered using the commercial Hermes multi-modality fusion software (Hermes multi-modality fusion software, Nuclear Diagnostics AB, Stockholm, Sweden) to limit potential artefacts arising from the misalignment of images during the CTAC procedure. The slice thickness of CT images was adjusted during the coregistration to match the thickness of PET images. To increase the accuracy of quantitative analysis, partial volume effect correction of the striatal phantom’s PET images was performed using the geometric transfer matrix-based method [16], where the regions of interest (ROIs) were delineated on the CT images to allow the computation of the corrected estimates without a priori knowledge on any activity level. Briefly, the algorithm directly computes the degradations introduced by the limited spatial resolution of the PET scanner, as well as smoothing introduced during image backprojection, and further modulation during extraction of regional tracer concentration. In practice, these partial volume factors are computed from the simulation of noise-free regional spread function images and sampling with a user-defined set of ROIs.

µ511 keV (cm−1)

Fig. 1

0.2 0.18 0.16 0.14 0.12 0.1 0.08 0.06 0.04 0.02 0 −1000

Results 140 kVp

120 kVp 80 kVp

Toshiba aquilion GE hispeed X/iF

− 500

0

500 1000 1500 2000 CT number (HU) Calculated bilinear calibration curves for conversion of computed tomography (CT) numbers (Hounsfield units, HU) into linear attenuation coefficients at 511 keV at different tube voltages for both Aquilion and HiSpeed X/iF CT scanners. The in-house designed polyethylene cylindrical phantom containing 16 cylindrical holes is shown in the top left corner.

Figure 1 shows the calculated bilinear calibration curves for both CT scanners used in this study at different tube voltages (80, 120 and 140 kVp). The XCOM photon cross-sections database [17] was used to calculate the corresponding linear attenuation coefficients of the inserted solutions at 511 keV. The slopes of the calibration curves for CT numbers greater than 0 HU increase with increasing tube voltage. It is worth noting that these curves have slightly different slopes for different scanners for the same tube voltage. A typical slice of the original clinical brain CT image (512  512 matrix), acquired at 120 kVp, is shown in Fig. 2. Three mmaps (128  128 matrix) were calculated from the same data set using calibration curves calculated at different tube voltages (Fig. 2). It was considered unethical to scan the patients with varying tube voltages owing to the additional radiation dose and the absence of any direct clinical

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Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 2

Original CT

120/80 kVp

120/120 kVp

120/140 kVp

Effect of using different calibration curves during the computed tomography-based attenuation correction (CTAC) procedure. From left to right: original clinical computed tomography (CT) image acquired at 120 kVp; derived mmaps at 511 keV using calibration curves calculated at tube voltages of 80, 120 and 140 kVp.

behaviour is reversed when using a calibration curve derived from a lower tube voltage (80 kVp).

Fig. 3

0.16

(a)

140 kVp 120 kVp 80 kVp

µ511 keV (cm−1)

0.14 0.12 0.1 0.08 0.06 0.04 0.02 0

1

21

41

30

(b)

61 81 Pixel number

101

121

140 kVp 120 kVp 80 kVp

25

ACF

20 15 10 5 0 1

21

41

61

81 101 121 141 161 181 Bin number

(a) Horizontal profiles through the mmaps shown in Fig. 2 and (b) central profiles through generated attenuation correction factor (ACF) sinogram (view 23/47).

benefit to the patients. Figure 3 shows the differences in the calculated mmaps and ACFs by displaying horizontal profiles through the middle of the same slice to demonstrate quantitatively the differences when using different calibration curves. Both mmaps and ACFs are overestimated when using a calibration curve derived from a tube voltage (140 kVp) higher than that used during actual CT scanning (120 kVp) of the patient. The

Figure 4 shows the Radiology Support Devices’ striatal phantom’s mmap obtained through transmission scanning using 137Cs sources, as well as the mmaps calculated by CTAC when CT images were acquired at 120 kVp and scaled using calibration curves derived at different tube voltages (80, 120 and 140 kVp) and when CT images were acquired at different tube voltages (80, 120 and 140 kVp) and scaled using a single calibration curve derived at 120 kVp. The difference between the mmaps and ACFs calculated by the different methods is shown in Fig. 5. A small but noticeable difference is visible on the horizontal profiles of the ACFs as a result of the overestimation/ underestimation of the attenuation coefficients depending on the combination of tube voltages used for acquisition/calibration curve derivation. The created mmaps based on the different methods (Fig. 4) were used for the attenuation correction of the emission data shown in Fig. 6. There is no visually significant difference between the images corrected for attenuation using CTAC with different combinations of tube voltages for CT image acquisition/calibration curve derivation. It should be noted that the illustrated mmaps are for different slices than the striatal images used for evaluation and shown in Fig. 6, where the differences between the mmaps generated using different conditions are small in the central region corresponding to the brain compartment (data not shown). However, the ACFs are created using three-dimensional forward projection of mmaps, and thus the noticeable differences in the bony regions might bias the ACF estimates in the striatal regions. Figure 7 shows the absolute activity concentrations estimated from the reconstructed PET images before and after partial volume correction for five individual compartments of the anthropomorphic striatal phantom. The ARCs for each compartment calculated before and after partial volume correction are also shown in Tables 1 and 2, respectively.

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CT-based attenuation correction in PET Ay and Zaidi 343

Fig. 4

MTM

80/120 kVp

120/80 kVp

120/120 kVp

140/120 kVp

120/140 kVp

Attenuation maps at 511 keV of the anthropomorphic striatal phantom calculated using different methods. From left to right: measured transmission method (MTM) using 137Cs single-photon sources; computed tomography-based attenuation correction (CTAC) method using different combinations of tube voltages for image acquisition and calculation of calibration curves.

Fig. 5

µ511 keV (cm−1)

(a)

0.2 0.18 0.16 0.14 0.12 0.1 0.08 0.06 0.04 0.02 0

MTM 80/120 kVp 120/80 kVp 120/120 kVp 140/120 kVp 120/140 kVp

1

ACF

(b)

5 4.5 4 3.5 3 2.5 2 1.5 1 0.5 0

21

41

61 81 Pixel number

101

121

MTM 80/120 kVp 120/80 kVp 120/120 kVp 140/120 kVp 120/140 kVp

1

21

41

61

81 101 121 141 161 181 Bin number

(a) Horizontal profiles through the mmaps shown in Fig. 4 and (b) central profiles through generated attenuation correction factor (ACF) sinogram (view 23/47). MTM, measured transmission method.

Discussion The advent of dual-modality PET/CT imaging has had a strong impact on the value of diagnostic PET in the localization, evaluation and therapeutic monitoring of head and neck cancer, and may be equally valuable for other localizations that are difficult to pinpoint [18]. The decrease in the examination time as a result of the use of low-noise CT data for attenuation correction is another benefit of combined scanners. PET/CT systems have demonstrated their ability to facilitate attenuation correction using an X-ray-based patient-specific attenua-

tion map that can be produced more rapidly and more accurately than attenuation maps generated with external radionuclide sources [2]. The general feasibility of CTAC has already been proven [10], but some practical technical issues remained to be explored. A high tube current improves CT image quality at the expense of increasing patient dose. It has been reported that effective doses of 8.81 and 18.97 mSv are delivered to the patient for a whole-body scan in highspeed and high-quality mode, respectively [19]. This is in contrast with the relatively low effective doses of 0.15 and 0.08 mSv for thoracic and whole-body transmission scans using positron-emitting 68Ga/68Ge and singlephoton-emitting 137Cs radionuclide sources, respectively [3]. This study was designed to assess the impact of using a single calibration curve on the accuracy of CTAC when CT images were acquired at different tube voltages and, vice versa, that is acquiring CT images at specific tube voltages and varying the voltage for the derivation of calibration curves. The feasibility of using a single calibration curve during practical application of CTAC for CT images acquired at different tube voltages was investigated through quantitative analysis of created mmaps, generated ACFs and reconstructed neurological PET data using experimental phantom and clinical studies. Moreover, the possibility of using low-dose CT for the purpose of attenuation correction was investigated for two commercial scanners to confirm the validity of the results reported in the literature using only clinical data (data not shown) [14]. More recently, a new preprocessing algorithm has been proposed which uses a single ultra-low-dose CT scan for both attenuation map construction and lesion localization [20]. The difference between the slopes of the calibration curves calculated at various tube voltages (Fig. 1) is due to the fact that the probability of photoelectric interaction increases with decreasing tube voltage, particularly in materials with high atomic numbers. Consequently, CT numbers in these regions increase with decreasing tube voltage. As the calculation of calibration curves is based

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344 Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 6

MTM

80/120 kVp

120/80 kVp

120/120 kVp

140/120 kVp

120/140 kVp

Reconstructed positron emission tomography (PET) images of the anthropomorphic striatal phantom corrected for attenuation using different methods. From left to right: measured transmission method (MTM); computed tomography-based attenuation correction (CTAC) method using different combinations of tube voltages for image acquisition and calculation of calibration curves.

on the CT numbers of air, water and cortical bone, the tube voltage dependence of the cortical bone’s CT number is the reason for the difference in the slopes of the calibration curves obtained at different tube voltages. Likewise, the difference between the calibration curves at a particular tube voltage for different scanners can be explained by possible differences in detector calibration procedures, X-ray spectral shape and the reconstruction algorithms used by different scanner manufacturers. As the X-ray tube spectrum is polyenergetic, with the exact energy spectra being determined by physical factors, including the characteristics of tubes, filters, etc., the spectrum for a specific tube voltage may differ slightly between different devices. It should be emphasized that, even for a particular CT scanner and fixed tube voltage; there may be changes in calibration curves obtained at different periods of time. The underestimation of clinical mmaps and ACFs when using a calibration curve derived at 80 kVp is due to the lower slope of the calibration curve in comparison with that obtained at 120 kVp (Figs. 2 and 3). The same behaviour was observed when using the anthropomorphic striatal phantom (Figs. 4 and 5). Generally, the difference between the mmaps and ACFs when using different calibration curves seems to be significant. The use of a calibration curve calculated at a tube voltage higher than that employed during CT scanning tends to overestimate the mmaps and ACFs. This behaviour is reversed when the calibration curve is derived at a tube voltage lower than that used during CT acquisition (Fig. 5). The underestimation of ACFs calculated by MTM, in comparison with CTAC, is the consequence of the underestimation of bone’s linear attenuation coefficients at 511 keV when using low-count, low-resolution transmission scans [4,5]. In contrast, the differences between the reconstructed PET images corrected using CTAC with different combinations of tube voltages for image acquisition/calibration curves are not qualitatively (Fig. 6) or quantitatively (Fig. 7b and Table 2) significant. One possible explanation is that the difference in ACFs varies at different projections, but is small on average. Likewise, the backprojection procedure averages the differences observed in the ACFs in projection space during the

reconstruction process. The noticeable underestimation of the absolute activity concentrations (Fig. 7a) and ARCs (Table 1) for the small brain structures (LPU, RPU, LCN and RCN) within the Radiology Support Devices’ striatal phantom is the result of the partial volume effect [16]. After partial volume correction of the emission images corrected for attenuation using CT images acquired at 80 and 140 kVp, average relative differences of – 2.9% and 0.7%, respectively, from the images acquired at 120 kVp were observed for the absolute activity concentrations in five regions of the anthropomorphic striatal phantom when CT images were converted to mmaps using a single calibration curve derived at 120 kVp. Likewise, average relative differences of 1.9% and – 0.6%, respectively, were observed when CT images acquired at 120 kVp and calibration curves derived at 80 and 140 kVp were used during CTAC. We conclude that the use of a single calibration curve for application of the CTAC procedure to images acquired at different tube voltages does not significantly affect the visual qualitative interpretation and quantitative analysis of neurological PET images. Despite the fact that increasing tube current increases the signal-to-noise ratio and decreases the statistical fluctuations in reconstructed CT images, thus improving image quality, CT numbers and the derived attenuation maps are tube current (mA) independent (data not shown). The statistical fluctuations of CT numbers in the low-current CT images are removed during the downsampling and smoothing procedures inherent to the CTAC procedure. Consistent with the observations reported by Kamel et al. [14], it appears that the tube current used during CT scanning does not affect significantly the quantification of clinical PET images for the purpose of calculating tumour uptake. As discussed above, it may happen that some slight differences are observed at the level of mmaps and ACFs when using different tube currents and a fixed tube voltage. These differences, however, will not induce significant differences during the quantitative analysis of reconstructed neurological PET images.

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CT-based attenuation correction in PET Ay and Zaidi 345

Fig. 7

Activity concentration (Bq/cm3)

(a) 60000

50000

40000

MTM 80/120 kVp 120/80 kVp 120/120 kVp 140/120 kVp 120/140 kVp True value

30000

20000

10000

0 Main chamber

Activity concentration (Bq/cm3)

(b) 60000

50000

40000

LPU

RPU Volume of interest

LCN

RCN

LPU

RPU Volume of interest

LCN

RCN

MTM 80/120 kVp 120/80 kVp 120/120 kVp 140/120 kVp 120/140 kVp True value

30000

20000

10000

0 Main chamber

Comparison between the true and calculated absolute activity concentrations in different brain structures of the striatal phantom when using the different attenuation correction methods before (a) and after (b) partial volume correction. MTM, measured transmission method; LCN, left caudate nucleus; LPU, left putamen; RCN, right caudate nucleus; RPU, right putamen.

As this study was carried out using separate PET and CT systems for the reasons mentioned in ‘Materials and methods’, it was limited to the use of a neurological research brain phantom and clinical brain images, rather than an anthropomorphic whole-body phantom and whole-body clinical images, which might impose a far greater challenge to the accuracy of attenuation correction due to the much larger attenuating volume, larger bony structures and more complex juxtapositions of media with different attenuating properties, e.g. lung/ soft tissue/bone in the thorax. It is hard to predict whether the answers will be equivalent or whether the same conclusions will be reached in the clinically challenging situations mentioned above. Further investi-

gation using whole-body data is guaranteed when the inline PET/CT system becomes fully operational in our department. Given the increasing use of CT contrast media and the severe challenge that such media present for accurate attenuation correction in PET/CT, this paper does not consider the effects that might be observed in the presence of contrast media or other non-human tissue (metallic implants etc.), which are addressed in a separate paper [21].

Conclusion The impact of the tube voltage (kVp) on the accuracy of CTAC in neurological PET studies was investigated in

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346 Nuclear Medicine Communications 2006, Vol 27 No 4

Table 1 Apparent recovery coefficients for different volumes of interest (VOIs) corresponding to different structures within the anthropomorphic brain phantom before partial volume correction (a/b kVp denotes computed tomography image acquired at a kVp scaled using a calibration curve calculated at b kVp) VOI

Volume (cm3)

MTM (662 keV)

CTAC (80/120 kVp)

CTAC (120/80 kVp)

CTAC (120/120 kVp)

CTAC (140/120 kVp)

CTAC (120/140 kVp)

Main chamber Left putamen (LPU) Right putamen (RPU) Left caudate nucleus (LCN) Right caudate nucleus (RCN)

1290 6 6 4.9 4.9

101.74 70.01 71.00 68.69 65.08

111.87 72.78 75.90 71.79 70.10

105.00 69.30 72.33 68.73 67.14

107.79 70.67 73.73 69.95 68.32

106.76 70.14 73.18 69.49 67.87

108.62 71.10 74.16 70.33 68.69

CTAC, computed tomography-based attenuation correction; MTM, measured transmission method

Table 2 Apparent recovery coefficients for different volumes of interest (VOIs) corresponding to different structures within the anthropomorphic brain phantom after partial volume correction (a/b kVp denotes computed tomography image acquired at a kVp scaled using a calibration curve calculated at b kVp) VOI

Volume (cm3)

MTM (662 keV)

CTAC (80/120 kVp)

CTAC (120/80 kVp)

CTAC (120/120 kVp)

CTAC (140/120 kVp)

CTAC (120/140 kVp)

Main chamber Left putamen (LPU) Right putamen (RPU) Left caudate nucleus (LCN) Right caudate nucleus (RCN)

1290 6 6 4.9 4.9

103.36 94.86 96.97 100.32 90.79

113.83 98.26 103.44 104.47 97.68

106.76 93.62 98.66 100.20 93.73

109.63 95.45 100.52 101.90 95.31

108.58 94.74 99.79 101.24 94.70

110.49 96.02 101.11 102.42 95.80

CTAC, computed tomography-based attenuation correction; MTM, measured transmission method.

detail using experimental phantom and clinical studies. It was concluded that the application of a single calibration curve derived under standard scanning conditions during the CTAC procedure to images acquired at different tube voltages does not affect significantly the visual qualitative interpretation and quantitative analysis of neurological PET images. The same behaviour was observed when calibration curves were derived at different tube voltages and used for the conversion of CT images acquired at a fixed tube voltage. These results may contribute to alleviate the quality assurance procedures required for daily operation of PET/CT scanners in a clinical environment.

8

9

10 11

12

13

14

Acknowledgements The authors would like to thank Dr O.G. Rousset for providing the PVC software and F. Schoenahl, T. Ruest and N. Andreini for their valuable support.

15

16

References 1 2

3 4 5

6

7

Townsend DW, Beyer T, Blodgett TM. PET/CT scanners: a hardware approach to image fusion. Semin Nucl Med 2003; 33:193–204. Hasegawa BH, Zaidi H. Dual-modality imaging: more than the sum of its components. In: Zaidi H, editor. Quantitative analysis in nuclear medicine imaging. New York: Springer; 2005. pp. 35–81. Zaidi H, Hasegawa B. Determination of the attenuation map in emission tomography. J Nucl Med 2003; 44:291–315. Weinzapfel BT, Hutchins GD. Automated PET attenuation correction model for functional brain imaging. J Nucl Med 2001; 42:483–491. Zaidi H, Montandon ML, Slosman DO. Magnetic resonance imaging-guided attenuation and scatter corrections in three-dimensional brain positron emission tomography. Med Phys 2003; 30:937–948. Zaidi H, Montandon ML, Slosman DO. Attenuation compensation in cerebral 3D PET: effect of the attenuation map on absolute and relative quantitation. Eur J Nucl Med Mol Imaging 2004; 31:52–63. Montandon M-L, Zaidi H. Atlas-guided non-uniform attenuation correction in cerebral 3D PET imaging. Neuroimage 2005; 25:278–286.

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Beyer T, Kinahan PE, Townsend DW, Sashin D. The use of X-ray CT for attenuation correction of PET data. Proc IEEE Nucl Sci Symp Med Imag Conf 1994; 4:1573–1577. Kinahan PE, Hasegawa B, Beyer T. X-ray-based attenuation correction for positron emission tomography/computed tomography scanners. Semin Nucl Med 2003; 34:166–179. Kinahan PE, Townsend DW, Beyer T, Sashin D. Attenuation correction for a combined 3D PET/CT scanner. Med Phys 1998; 25:2046–2053. Burger C, Goerres G, Schoenes S, Buck A, Lonn AH, Von Schulthess GK. PET attenuation coefficients from CT images: experimental evaluation of the transformation of CT into PET 511-keV attenuation coefficients. Eur J Nucl Med Mol Imaging 2002; 29:922–927. Bai C, Shao L, Da Silva AJ, Zhao Z. A generalized model for the conversion from CT numbers to linear attenuation coefficients. IEEE Trans Nucl Sci 2003; 50:1510–1515. Guy MJ, Castellano-Smith IA, Flower MA, Flux GD, Ott RJ, Visvikis D. DETECT-dual energy transmission estimation CT for improved attenuation correction in SPECT and PET. IEEE Trans Nucl Sci 1998; 45:1261–1267. Kamel E, Hany TF, Burger C, Treyer V, Lonn AHR, Von Schulthess GK, et al. CT vs 68Ge attenuation correction in a combined PET/CT system: evaluation of the effect of lowering the CT tube current. Eur J Nucl Med 2002; 29:346–350. Rappoport V, Carney JPJ, Townsend DW. CT tube-voltage dependent attenuation correction scheme for PET/CT scanners. Proc IEEE Nucl Sci Symp Med Imag Conf 2004; 6:3853–3857. Rousset OG, Ma Y, Evans AC. Correction for partial volume effects in PET: principle and validation. J Nucl Med 1998; 39:904–911. Berger MJ, Hubbell JH, Seltzer SM, Chang J, Coursey JS, Sukumar R, et al. XCOM: photon cross sections database. NBSIR 87-3597, 1998; http:// physics.nist.gov/PhysRefData/Xcom/Text/XCOM.html Branstetter BF IV, Blodgett TM, Zimmer LA, Snyderman CH, Johnson JT, Raman S, et al. Head and neck malignancy: is PET/CT more accurate than PET or CT alone? Radiology 2005; 235:580–586. Wu TH, Huang YH, Lee JJS, Wang SY, Wang SC, Su CT, et al. Radiation exposure during transmission measurements: comparison between CT- and germanium-based techniques with a current PET scanner. Eur J Nucl Med Mol Imaging 2004; 31:38–43. Li J, Hsieh J, Colsher J, Stearns C, Lonn AHR. Towards single ultra-low dose CT scan for both attenuation map creation and localization in PET-CT application. Proc IEEE Nucl Sci Symp Med Imag Conf 2004; 4: 2396–2398. Ay MR, Zaidi H. Assessment of errors caused by x-ray scatter and use of contrast medium when using CT-based attenuation correction in PET. Eur J Nucl Med Mol Imaging 2006; 33: in press.

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Original article

Different methods for the detection of small changes in uptake between single-photon emission computed tomography (SPECT) examinations: 99mTc-sestamibi in chemotherapy for breast tumours Hans Jacobssona, Brigitte Wilczeka, Jonas Berghb, Eva von Schoultzb, Elina Erikssonc and Stig A. Larssond Aim This study was undertaken to evaluate different methods for the detection of small changes in uptake between single-photon emission computed tomography (SPECT) examinations in the same individual. No standard exists for making digital evaluations at single-photon examinations. For this purpose, we employed a patient cohort from a previous study assessing the response to neoadjuvant chemotherapy for breast cancer using 99m Tc-hexakis-2-methoxyisobutylisonitrile (99mTc-sestamibi). Methods The tumour uptake in 29 women with locally advanced breast cancer was examined using 99m Tc-sestamibi and SPECT before neoadjuvant chemotherapy and, on average, 19 days after one chemotherapy cycle. The histology of the finally resected tumour confirmed a therapeutic response. Different assessments of the uptake, various levels of background activity subtraction and different reference tissues for relative activity calculations were used. The tumour uptake and activity of the reference tissues were also related to the administered activity. Results Different definitions of tumour activity had little influence. Relating the tumour uptake to a large portion of the abdomen, as well as visual evaluation, showed a therapeutic response. Comparison with the administered

Introduction In order to ensure objectivity in medical imaging, approaches such as blind assessments, consensus evaluations and semi-quantitative scoring systems are used for the evaluation of clinical examinations and scientific studies. Since nuclear medicine examinations have become digitized, this has also been used for the evaluation of such examinations. Digital data are easily subjected to further processing. The general perception is that digital evaluation is more objective and accurate than visual assessment, although there is little support for this in the literature. In addition, no effort has been made to determine how to perform digital evaluations of single-photon examinations. There are no established standards for this purpose, and different methods are used ad hoc for assessments and comparisons. This is

activity showed that the apparent responses were due to an increased activity of the reference tissues. Referring the tumour uptake to the administered activity truly depicted a therapeutic response. Conclusions A critical attitude is necessary when making digital evaluations at SPECT. Digital data may seem more relevant than they really are. Relative comparisons may be unreliable. It may be necessary to develop standardized methods for this purpose. Nucl Med Commun 27:347–352

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:347–352 Keywords: breast cancer, mammoscintigraphy, neoadjuvant chemotherapy, single-photon emission computed tomography (SPECT), therapy evaluation Departments of aRadiology, bOncology (Radiumhemmet), cPathology and d Hospital Physics, Karolinska University Hospital, Stockholm, Sweden. Correspondence to Dr Hans Jacobsson, Department of Radiology, Karolinska University Hospital, SE-171 76 Stockholm, Sweden. Tel: + 46 8 517 73581; fax: + 46 8 517 74939; e-mail: [email protected] Sponsorship: Expenses for the statistical analyses were covered by a grant from DuPont Pharma, the supplier of 99mTc-sestamibi. Received 17 November 2005 Accepted 10 January 2006

in contrast with positron emission tomography (PET), where the calculation of standardized uptake values (SUVs) is an established technique. The aim of this study was to evaluate different methods for the detection of small changes in tumour uptake between two consecutive single-photon emission computed tomography (SPECT) examinations in the same patient. For this purpose, we employed a patient cohort from a previous study assessing the response to neoadjuvant (preoperative) chemotherapy for breast cancer using 99m Tc-hexakis-2-methoxyisobutylisonitrile (99mTc-sestamibi) [1]. Standard therapy for locally advanced breast cancer includes neoadjuvant chemotherapy. The response to such therapy is assessed by clinical examination and mammography. These methods have shortcomings,

c 2006 Lippincott Williams & Wilkins 0143-3636

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348 Nuclear Medicine Communications 2006, Vol 27 No 4

including a low sensitivity [2] due to the delay until tumour shrinkage and difficulties in discriminating between remaining fibrotic tissue and residual tumour [3–7]. In our previous study, examination using 99mTc-sestamibi after completion of neoadjuvant chemotherapy demonstrated a response, whereas, in another patient group examined after one therapy cycle, the effect was very weak and not significant in the restricted evaluation [1]. This made the latter patient group suitable for the current study. A comparison was made of the tumour 99mTc-sestamibi uptake in examinations before and after the first chemotherapy cycle. Different calculations of the tumour uptake, various levels of background activity subtraction and different reference tissues for the assessment of the relative tumour activity were used. Comparisons of the absolute uptake and visual evaluation were also made.

Patients and methods Patients

Twenty-nine women (mean age, 49 years; range, 31–66 years) with newly diagnosed large breast cancer (15 right and 14 left), who were scheduled for neoadjuvant chemotherapy, were examined. The study was approved by the local ethics and radiation safety committees. The diagnosis was confirmed by fine-needle aspiration cytology at least 1 week before baseline scintigraphy. The second examination was performed an average of 19 days after the first chemotherapy course, on the day before the second course or on the same day but prior to the initiation of chemotherapy. Thirty patients were initially enrolled in the study. At the subsequent analysis, one patient differed markedly from the others by showing a very strong therapeutic response between the two examinations, thus giving rise to a skewness outside ± 1.5 in several of the statistical evaluations. In order to assess small changes between the examinations, which was the aim of this study, this patient was excluded to allow for the use of parametrical statistical methods. Neoadjuvant chemotherapy

Chemotherapy was given as either standard FEC courses (5-fluorouracil, 600 mg/m2 body surface area; epirubicin, 60 mg/m2; cyclophosphamide, 600 mg/m2) every third week or dose-escalated and toxicity-modulated FEC courses (5-fluorouracil, 300–600 mg/m2; epirubicin, 38– 120 mg/m2; cyclophosphamide, 450–1800 mg/m2) with intended cycle intervals of 3 weeks [8]. Scintigraphic examination

Identical SPECT examinations were performed after the administration of approximately 500 MBq of 99mTcsestamibi (Cardiolite, DuPont Pharma, Billerica, Massachusetts, USA). The syringe activity was measured with a radioisotope calibrator (CRC-120, Capintec, Ramsey,

New Jersey, USA) before the injection. From this, 3.9% was subtracted, and the remaining activity was considered to be administered to the patient. In a previous study, this fraction of 99mTc-sestamibi was trapped in the same type of syringe [9]. The examination was performed using a triple-headed gamma camera (Triad XLT, Trionix Inc., Twinsburg, Ohio, USA) equipped with low-energy, ultrahigh-resolution, parallel-hole collimators, and operating on a 128  128 matrix. No maintenance or changes in the camera performance were carried out between examinations in the same patient. With the patient supine and wearing a non-metallic brassie`re, a 30 min acquisition in 90 angular steps over 3601 was initiated 5 min after intravenous administration of the tracer. The data were corrected on-line with regard to energy, linearity and uniformity. The projections were prefiltered with a twodimensional Hamming filter (cut-off frequency, 1.40 cycles/cm) and transverse sections were reconstructed using a ramp filter and filtered backprojection. Correction was made for photon attenuation [10]. The projection data for each section were weighted together from five different consecutive rows according to a kernel of 1, 2, 5, 2 and 1 before reconstruction. Mean tumour activity and reference tissue activity

A tumour uptake was always identified. For each study, all transverse sections with a visible tumour uptake were summed into one image. In this, a region of interest (ROI) was drawn around the tumour with a small margin, representing the ‘total tumour’ activity (Ttotal). In the same image and breast, an identical ROI was positioned outside the tumour at the same depth from the chest wall, representing the ‘total background’ activity (Btotal). In the same image, the entire cross-section of the trunk was also enclosed by an ROI (‘cross-section’ reference activity). A ‘lung’ reference region was defined by a tumour-sized ROI positioned in the centre of the right lung, using the same number of sections as in calculating the ‘total tumour’ activity. As the ‘cross-section’ reference region is heterogeneous, in contrast with these other regions, the same number and similarly positioned sections were used when calculating this activity for the two examinations in the individual patient. After normalization to the number of sections, the ‘mean tumour’ (Tmean), ‘background’ (Bmean), ‘lung’ and ‘cross-section’ activities were expressed as the count rate/pixel. The decrease in activity between the two examinations in each individual was visually assessed before the digital evaluations. This was performed by an experienced nuclear medicine physician (HJ), blind to the order of the examinations, from rotating maximum intensity projection (MIP) images of both examinations displayed simultaneously. The evaluation was performed according to a semi-quantitative scale from 0 to 3, where 0 represents no change or increasing activity and 3 a marked decrease in activity.

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Digital evaluations in SPECT Jacobsson et al. 349

Maximum tumour activity

For each examination, the transverse section with the highest tumour uptake was selected by applying a colour scale with discrete steps. Using the standard software, the pixel count with the highest count rate in the tumour ROI was designated the ‘maximum tumour’ activity. Differences in relative tumour uptake between examinations

Background correction was performed by subtracting 33%, 50% and 100% of the ‘background’ activity, respectively, from the ‘mean tumour’ activity (Tmean – 0.33Bmean, Tmean – 0.50Bmean and Tmean – Bmean, respectively). For each examination, ratios were then calculated between these background-corrected ‘mean tumour’ activities, on the one hand, and the ‘background’, ‘lung’ and ‘crosssection’ reference activities, respectively, on the other. For the evaluation of the therapeutic response, the ratio for the baseline study was subtracted from the ratio for the second study. Ratios were also calculated between the ‘maximum tumour’ activity without correction for background activity, on the one hand, and the ‘breast background’, ‘lung’ and ‘cross-section’ reference activities, respectively, on the other. Identical evaluations as above were then carried out. Differences in tumour uptake of the administered activity between the examinations

For this purpose, the ‘total tumour’ activity (Ttotal) and the ‘total background’ activity (Btotal) were used, and thus without correction for the number of sections and pixels. Background correction was performed by subtracting 33%, 50% and 100% of the ‘total background’ activity, respectively, from the ‘total tumour’ activity (Ttotal – 0.33Btotal, Ttotal – 0.50Btotal and Ttotal – Btotal, respectively). These values were thereafter divided by the administered activity (counts/MBq). For the evaluation of the therapeutic response, the ratio for the baseline examination was subtracted from the ratio for the second examination. Ratios were calculated between the ‘maximum tumour’ activity without correction for background activity and the administered activity. Thereafter, identical evaluations as above were carried out. Histopathological evaluation

Semi-quantitative assessments of the response to chemotherapy were made from haematoxylin–eosin-stained sections of the finally resected tumour by an experienced pathologist (EE) according to the following: 3, total disappearance of the tumour cells and replacement by fibrosis or less than 10% remaining tumour cells with nonviable morphology; 2, 50–90% of the tumour cells replaced by fibrosis and moderate degenerative changes

in the remaining tumour cells; 1, less than 50% of the tumour cells replaced by fibrosis and slight degenerative changes in the remaining tumour cells; 0, all tumour mass left and with morphologically viable tumour cells. Statistical methods

The differences in the relative tumour activity (‘the ratio change’) and the differences in the tumour uptake of the administered activity between the examinations were summarized using means and standard deviations. The population mean change was compared with the null hypothesis value m = 0. The decrease in activity between the two examinations, assessed visually and by the pathologist, was analysed using a significance test for a single proportion. The proportion of patients with a response of 1 or more was calculated, and the population mean for the proportion was compared with the null hypothesis value p = 50%. The associations between the differences in the relative tumour activity (‘the ratio change’) between the examinations, the differences in tumour uptake of the administered activity between the examinations and the visual assessment of the uptake reduction between the examinations, on the one hand, and the pathologist’s assessment, on the other, were calculated by Spearman rank-order correlation analysis.

Results The correlation analysis of the therapeutic response evaluated by histological analysis and all the different assessments of the scintigraphic change between the two examinations revealed a very weak relationship or no relationship. Spearman’s rank-order correlation coefficient ranged from – 0.25 to 0.14. The P values ranged from 0.19 to 0.98 (findings not illustrated). The pathologist’s assessment of the finally resected tumour showed a significant therapeutic response in terms of the entire patient group. In addition, a therapy effect between the examinations was found in the visual evaluation of the SPECT examinations (Table 1). The differences in the relative tumour uptake between the two examinations are shown in Table 2. The use of normal breast tissue and lung tissue as reference showed a therapy effect only when comparing the maximum tumour activity with breast tissue; however, the use of ‘cross-section’ as reference always showed an effect. This was the case irrespective of the amount of breast background activity that was subtracted from the tumour uptake. There was a small decrease in the breast background and lung activity in relation to the administered activity

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(6–8%), whereas the cross-section activity increased considerably (28%) between the examinations (Table 3). The differences in tumour uptake from the administered activity (absolute uptake) between the two examinations are shown in Table 4. At all evaluations, there was a significant difference between the examinations.

Discussion Several groups have reported that examination with 99m Tc-sestamibi after completion of neoadjuvant chemotherapy for breast cancer shows a response [11–14]. Histological analysis of the resected tumour confirmed that most patients had responded to therapy, although the lack of correlation between the decrease in activity between the two examinations and the histopathological evaluation of the therapeutic response cannot be explained. A restricted agreement between the decrease in 99mTc uptake and the histopathological response after successful neoadjuvant chemotherapy of breast cancer has also been described previously [15]. By tradition, histopathology is considered to be the ‘gold standard’, and nuclear medicine and histopathology measure different properties. This issue, however, is beyond the scope of this study. Tracer uptake reflects metabolic Effect of neoadjuvant chemotherapy in 29 women with breast cancer. Pathologist’s assessment of the therapeutic response at examination of the finally resected specimen and visual evaluation of the decrease in tumour uptake between single-photon emission computed tomography (SPECT) examinations with 99mTc-hexakis-2-methoxyisobutylisonitrile (99mTcsestamibi) before and after the first therapy cycle. The therapy effects were assessed using a semi-quantitative scale (0 – 3). Population means for the proportion of patients with a decrease of 1 or more are shown Table 1

Proportion of patients with a therapeutic response of Z 1 at histological assessment Proportion of patients with an activity reduction of Z 1 at second examination (visual assessment)

Mean (%)

P value

77

< 0.004

70

< 0.05

status. It is related to blood flow and mitochondrial transmembrane electronegativity, and inversely related to necrosis or fibrosis [16,17]. P-glycoprotein was not analysed. This is involved in multi-drug resistance and enhances tracer efflux after rapid uptake [18]. All examinations were performed soon after tracer administration. The lateral prone (planar) view is used in clinical mammoscintigraphy for the detection of tumours. For the evaluation of a known tumour, SPECT also depicts possible reference tissues other than the surrounding normal breast. In addition, SPECT is better for quantification. Our routine Chang correction for attenuation is not ideal for the chest, but allows for intraindividual comparisons to be made [10]. The myocardium was found to be insufficiently thick to serve as a reproducible reference. The bone marrow cannot be used because of possible chemotherapy effects. The liver activity is not stable as the tracer is excreted by the biliary system. The lung is large and reproducible. It was used for the same purpose in a similar study [14]. The ‘cross-section’ includes several tissues, thereby representing a ‘global’ reference. Subtraction between the ratios, as performed previously, is considered to be better than dividing the ratios by one another [1]. Although large differences in uptake between scintigraphic examinations can be detected by the naked eye, small changes may have to be established by digital evaluations. In this methodological study, the single individual who lay outside the normal distribution was excluded, thereby making the use of parametrical statistical methods possible. The application of such methods allows the detection of very small differences between observations, which was the aim of this study. As the tumour could not be outlined exactly in the added sections, a certain portion of surrounding tissue was always included, necessitating the subtraction of back-

Table 2 Differences in tumour uptake versus various reference tissues at single-photon emission computed tomography (SPECT) examinations with 99mTc-hexakis-2-methoxyisobutylisonitrile (99mTc-sestamibi) before and after one cycle of neoadjuvant chemotherapy in 29 women with breast cancer. Various fractions of breast background activity were subtracted from the tumour uptake when calculating the mean tumour uptake Differences in activity ratio

Mean tumour uptake

Mean activity ratio Background activity subtraction (%)

Baseline examination

Second examination

Mean

± SD

P value

Breast background

33 50 100 33 50 100 33 50 100 0 0 0

1.70 1.54 1.04 0.60 0.53 0.33 0.13 0.11 0.07 5.89 1.97 0.38

1.54 1.37 0.87 0.57 0.50 0.29 0.12 0.10 0.06 5.16 1.79 0.34

– 0.17 – 0.17 – 0.17 – 0.03 – 0.03 – 0.04 – 0.01 – 0.01 – 0.01 – 0.73 – 0.17 – 0.04

0.50 0.50 0.50 0.19 0.18 0.16 0.03 0.03 0.02 1.86 0.62 0.08

0.08 0.08 0.08 0.45 0.39 0.21 < 0.05 < 0.05 < 0.05 < 0.05 0.14 < 0.05

Lung

Cross-section

Maximum tumour uptake

Change between examinations

Reference tissue

Breast background Lung Cross-section

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Digital evaluations in SPECT Jacobsson et al. 351

ground activity. When enclosing an ‘ideal’ spherical tumour in a cylinder with the same diameter and height, one-third of the cylinder volume is outside the sphere. As the configuration of some tumours may deviate from a sphere, larger fractions of background activity (50% and 100%) were also subtracted. In the evaluation of SPECT studies, calculations of some type of ‘mean’ activity of the target uptake are usually made. Assessment of the ‘maximum’ activity is uncommon, but may be compared with the calculation of SUVmax in PET. Relating the uptake to the administered activity is associated with several uncertainties and is not usual at SPECT. However, as performed in the present study and used for comparison in the same patient this evaluation method is considered relevant. Few reports have described or evaluated different methods for the quantification and comparison of findings at single-photon examinations, even though such studies are used for therapy evaluation as well as scientific purposes. Serial bone scintigrams are usually scrutinized by the naked eye in the work-up of metastases [19]. One study has evaluated different digital methods for the assessment of the uptake of 99m Tc-sestamibi in order to distinguish benign from malignant breast lesions when using planar scintigraphy [20]. In a few studies using planar and SPECT examinations to evaluate neoadjuvant chemotherapy in breast cancer with 99mTc-sestamibi, digital and visual evaluations were performed, but these were not compared [13,14,21]. In a SPECT study using 99mTc-sestamibi in head and neck cancer, the choice of reference region for the calculation of relative tumour uptake was studied [22]. One study assessed average versus maximal count Uptake of the administered activity in different reference tissues at single-photon emission computed tomography (SPECT) examinations with 99mTc-hexakis-2-methoxyisobutylisonitrile (99mTc-sestamibi) before and after one cycle of neoadjuvant chemotherapy in 29 women with breast cancer (counts/pixel/ MBq) Table 3

Breast background Lung Cross-section

Baseline examination

Second examination

0.16 0.40 15.7

0.15 0.37 20.1

ROIs at SPECT of indeterminate lung nodules using 99m Tc-depreotide [23]. Quantifications at single-photon examinations are usually performed as comparisons with reference tissue, assuming that this remains stable between examinations. Using normal breast and lung as reference tissues was not adequate to detect a therapy effect, but reference to the ‘cross-section’ showed an effect. Calculating the uptake relative to the administered activity revealed a very small difference in the breast background activity and lung uptake between the examinations, whereas a considerable increase in the ‘cross-section’ activity explained the apparent therapy effect detected in this comparison. The reason for this cannot be explained, but it is presumed that chemotherapy affects many tissues and organs in different ways. This illustrates that relative comparisons may be unreliable and that controlled conditions are necessary for this purpose. On relating the tumour uptake to the administered activity, there was a significant decrease between the examinations. This evaluation was more robust than relative comparisons, and may be compared with the calculation of SUVs at PET. The distribution of a radiopharmaceutical in the body, however, represents a zero-sum game, why the effects of chemotherapy on large indifferent organs or tissues may indirectly affect the uptake of a target organ by a general redistribution of activity. Visual assessment of the change in uptake between examinations is always possible. This is completely subjective, but adequate for many aspects of medical imaging. The simultaneous display of MIP images from both studies provides optimal conditions for comparison. The exact quality assessed in the visual evaluation cannot be stated. Although it is an adequate measure of the therapeutic response, assessment must be made by relative comparisons within the image, why this evaluation may be similar to comparisons with the ‘crosssection’ reference. In many respects, digital information is robust and objective. Digital data are also easily subjected to further processing and their applicability for image analysis is

Differences in tumour uptake of administered activity at single-photon emission computed tomography (SPECT) examinations with 99mTc-hexakis-2-methoxyisobutylisonitrile (99mTc-sestamibi) before and after one cycle of neoadjuvant chemotherapy in 29 women with breast cancer. Various fractions of background activity were subtracted from the tumour uptake when calculating the total tumour activity Table 4

Tumour uptake of administered activity (counts/MBq)

Total tumour uptake

Maximum tumour uptake

Change between examinations

Background activity subtraction (%)

Baseline examination

Second examination

Mean

± SD

P value

33 50 100 0

448 398 249 0.73

305 267 158 0.62

– 144 – 130 – 91 – 0.11

180 168 138 0.20

< 0.001 < 0.001 < 0.01 < 0.01

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352 Nuclear Medicine Communications 2006, Vol 27 No 4

obvious. Our findings, however, indicate that a critical attitude is necessary when making such evaluations. Digital data are easily produced in various and more or less controlled ways. Digits seem ‘seductive’ when compared with analogous information and, depending on how they are acquired, may appear to be more relevant than they really are. The choice and definition of the ROIs from which digital information is usually extracted represent crucial steps. Our findings indicate that relative evaluations of tumour uptake in chemotherapy may be misleading. Consequently, for the evaluation of tumour therapy at single-photon examinations, methods standardized with regard to the radiopharmaceutical and possibly also with regard to the therapeutic regimen may have to be developed. Another important conclusion was that an effect of neoadjuvant chemotherapy for breast cancer could be detected after one therapy course provided that an adequate evaluation was performed. This was in contrast with our previous analysis of this patient cohort based on relative comparisons. Thus, examination with 99mTcsestamibi, instead of the more expensive PET examinations, may be useful for the evaluation of chemotherapy for tumours, provided that adequate methods are applied.

6

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10 11

12

13

14

15

16

Acknowledgement The professional statistical analysis by Elisabeth Berg, BSc (Medical Statistics Unit, Karolinska Institute, Stockholm, Sweden) is greatly appreciated.

17

18

References 1

2 3

4

5

Wilczek B, von Schoultz E, Bergh J, Eriksson E, Larsson SA, Jacobsson H. Early assessment of neoadjuvant chemotherapy by FEC-courses of locally advanced breast cancer using 99mTc-MIBI. Acta Radiol 2003; 44:284–287. Bonadonna G, Valagussa P, Zucali R, Salvadori B. Primary chemotherapy in surgically resectable breast cancer. CA Cancer J Clin 1995; 45:227–234. Cocconi G, Di Blasio B, Alberti G, Bisagni G, Botti E, Peracchia G. Problems in evaluating the response of primary breast cancer to systemic therapy. Breast Cancer Res Treat 1984; 4:309–313. Herrada J, Iyer RB, Atkinson EN, Sneige N, Buzdar AU, Hortobagyi GN. Relative value of physical examination, mammography, and breast sonography in evaluating the size of the primary tumor and regional lymph node metastases in women receiving neoadjuvant chemotherapy for locally advanced breast carcinoma. Clin Cancer Res 1997; 3:1565–1569. Moscovic EC, Mansi JL, King DM, Murch CR, Smith IE. Mammography in the assessment of response to medical treatment of large primary breast tumor. Clin Radiol 1993; 47:339–344.

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Segel MC, Paulus DD, Hortobagyi GN. Advanced primary breast cancer: assessment at mammography of response to chemotherapy. Radiology 1988; 169:49–54. Yang WT, Lam WW, Cheung H, Suen M, King WW, Metreweli C. Sonographic, magnetic resonance imaging, and mammographic assessments of preoperative size of breast cancer. J Ultrasound Med 1997; 16:791–797. Bergh J, Wiklund T, Erikstein B, Lidbrink E, Lindman H, Malmstro¨m P, et al. Tailored fluorouracil, epirubicin, and cyclophosphamide compared with marrow-supported high-dose chemotherapy as adjuvant treatment for high-risk breast cancer: a randomised trial. Lancet 2000; 356:1384–1391. Wilczek B, Svensson L, Danielsson R, Celebiouglu F, Larsson SA, Jacobsson H. 99mTc-exametazime as a breast tumor-seeking agent: comparison with 99mTc-sestamibi. J Nucl Med 2004; 45:2040–2044. Chang L-T. A method for attenuation correction in radionuclide computed tomography. IEEE Trans Nucl Sci 1978; 25:638–643. Cwikla JB, Buscombe JR, Barlow RV, Kelleher SM, Parbhoo SP, Crow J, et al. The effect of chemotherapy on the uptake of technetium-99m sestamibi in breast cancer. Eur J Nucl Med 1997; 24:1175–1178. Maini CL, Tofani A, Sciuto R, Semprebene A, Cavaliere R, Mottolese M, et al. Technetium-99m-MIBI scintigraphy in the assessment of neoadjuvant chemotherapy in breast carcinoma. J Nucl Med 1997; 38:1546–1551. Mankoff DA, Dunnwald LK, Gralow JR, Ellis GK, Drucker MJ, Livingston RB. Monitoring the response of patients with locally advanced breast carcinoma to neoadjuvant chemotherapy using [technetium 99m]-sestamibi scintimammography. Cancer 1999; 85:2410–2423. Tiling R, Linke R, Untch M, Richter A, Fieber S, Brinkba¨umer K, et al. 18 F-FDG PET and 99mTc-sestamibi scintimammography for monitoring breast cancer response to neoadjuvant chemotherapy: a comparative study. Eur J Nucl Med 2001; 28:711–720. Cwikla JB, Kolasinska AD, Buscombe JR, Parbhoo SP, Jones AL, Hilson AJ. How does chemotherapy for breast cancer affect the uptake of Tc-99m MIBI in-vivo [Abstract]? J Nucl Med 1999; 40(Suppl):233. Khalkhali I, Cutrone J, Mena I, Diggles L, Venegas R, Vargas H, et al. Technetium-99m-sestamibi scintimammography of breast lesions: clinical and pathological follow-up. J Nucl Med 1995; 36:1784–1789. Piwnica-Worms D, Kronauge JF, Chiu ML. Uptake and retention of hexakis (2-methoxyisobutyl isonitrile) technetium(I) in cultured chick myocardial cells. Mitochondrial and plasma membrane potential dependence. Circulation 1990; 82:1826–1838. Del Vecchio S, Ciarmiello A, Pace L, Potena MI, Carriero MV, Mainolfi C, et al. Fractional retention of technetium-99m-sestamibi as an index of P-glycoprotein expression in untreated breast cancer patients. J Nucl Med 1997; 38:1348–1351. Cook GJ, Fogelman I. The role of nuclear medicine in monitoring treatment in skeletal malignancy. Semin Nucl Med 2001; 31:206–211. Dunnwald LK, Hartnett SD, Mankoff DA. Utility and reproducibility of semiquantitative analysis of sestamibi breast images. J Nucl Med Technol 1997; 25:106–109. Tiling R, KeXler M, Untch M, Sommer H, Linke R, Brinkba¨umer K, et al. Breast cancer: monitoring response to neoadjuvant chemotherapy using Tc-99m sestamibi scintimammography. Onkologie 2003; 26:27–31. Seok JW, Kim IJ, Kim YK, Kim JY, Wang SG. Which background is more useful and optimal for detection of head and neck cancer with Tc-99m MIBI SPECT [Abstract]? Eur J Nucl Med Mol Imaging 2002; 29(Suppl):239. Montilla JL, Bridwell RS. Average vs. maximal count region of interest quantitative analysis of lung nodules using 99mTc depreotide. Is this nodule malignant [Abstract]? J Nucl Med 2002; 43(Suppl):300.

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Original article

Cell death induced by 131I in a differentiated thyroid carcinoma cell line in vitro: Necrosis or apoptosis? Kathrin Marx, Detlef Moka, Klaus Schoma¨cker, Thomas Fischer, Beate Gabruk-Szostak, Carsten Kobe, Markus Dietlein and Harald Schicha Aim The apoptotic and necrotic dose–response of thyroid carcinoma cells following irradiation with 131I was evaluated. Methods In our in-vitro model, cells of well-differentiated papillary thyroid carcinoma (B-CPAP) were incubated with increasing activity concentrations of 131I for 2 days. Changes in cell viability and the extents of necrosis and apoptosis were evaluated both immediately and 2 days after irradiation. Results Viability of B-CPAP cells diminished with increasing 131I activity concentration. No apoptosis was detectable immediately after irradiation. Two days after irradiation significant apoptosis was found. The lowest 131 I activity concentration at which apoptosis was detectable corresponds to about 1 MBq
ml – 1. At higher activity concentrations a larger percentage of cells became apoptotic but the proportion decreased again at activity concentrations > 10 MBq
ml – 1. Likewise, necrosis was minimal at low activity concentrations and showed an exponential increase with rising 131I activity concentrations ( > 5–10 MBq
ml – 1). Necrosis was already detectable immediately after irradiation and

Introduction Radioiodine therapy (RIT) with 131I is widely used in the treatment of benign thyroid diseases and in cases of well-differentiated thyroid carcinoma for eliminating thyroid (carcinoma) cells [1–6]. In general, two different types of cell death are known: apoptosis and necrosis. The term apoptosis is used to describe a programmed physiological cell suicide induced by different environmental stimuli [7,8]. In contrast to apoptosis, necrosis is a mechanism of cell death resulting from overwhelming cellular injury produced by high levels of toxic agents such as radiation [8]. One major distinction between apoptosis and necrosis in vivo, which is of clinical relevance in RIT, is that complete elimination of apoptotic cells by phagocytes prevents an inflammatory response. Necrosis, on the other, is characterized by an early loss of cell membrane integrity, resulting in leakage of cytoplasmic contents and the induction of an inflammatory response with injury to the surrounding tissue [9–12].

was the predominant form of cell death at high activity concentrations. Conclusion The data suggest that the nature of the cytotoxic effect of 131I and whether it leads to apoptotic or necrotic cell death is dose-dependent. High 131I doses seem to produce mainly necrotic phenomena, whereas at low 131I activity concentrations apoptotic phenomena prevail. The predominance of delayed apoptosis could explain why radioiodine therapy at lower doses is often linked to delayed onset and possible continuation of thyroid volume reduction over some months and even c 2006 up to a year. Nucl Med Commun 27:353–358 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:353–358 Keywords: necrosis, apoptosis, iodine-131, thyroid carcinoma Department of Nuclear Medicine, University of Cologne, Germany. Correspondence to Dr Kathrin T. Marx, Department of Nuclear Medicine, University of Cologne, Kerpenerstrasse 62, 50937 Cologne, Germany. Tel: + 0049 221 478 4050; fax: + 0049 221 478 6777; e-mail: [email protected] Received 13 September 2005 Accepted 25 November 2005

Due to the high thyroid absorbed doses delivered ( > 200–400 Gy), high-activity RIT is generally associated with rapid elimination of thyroidal cells, that is within days, if not hours, of treatment. In conjunction with this, in about 5–20% of cases, high-activity RIT produces inflammatory side-effects such as a painful local inflammation of the thyroid tissue (radiothyroiditis) or sialadenitis. On the other hand, administration of lower 131I doses ( < 200 Gy) produces fewer of these inflammatory reactions [13–16]. In contrast to high-activity RIT, loweractivity RIT is associated with delayed onset of therapeutic response and possible continuation of thyroid volume reduction over some months to a year [17,18]. These observations raise the question of whether 131I at lower doses is more likely to induce apoptosis whereas higher doses might be more likely to lead to direct, necrotic cell death [19,20]. In our experimental design we therefore investigated cells of well-differentiated papillary thyroid carcinoma exposed to different activity concentrations of 131I and changes in cell viability,

c 2006 Lippincott Williams & Wilkins 0143-3636

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Nuclear Medicine Communications 2006, Vol 27 No 4

apoptosis and necrosis were evaluated at different times after irradiation.

changes in cell viability in relation to 131I activity concentration, as compared to the control group (both corrected for background levels).

Materials and methods Cell line

Photometric detection of necrosis and apoptosis

The cell line B-CPAP of well-differentiated papillary thyroid carcinoma obtained from the German Collection of Microorganisms and Cell Cultures (DSMZ, Braunschweig, Germany) was grown as an adherent monolayer in RPMI 1640 medium supplemented with 10% fetal bovine serum (GIBCO BRL, Grand Island, New York, USA) [21]. The cells were cultured at 371C in an incubator with a humidified atmosphere containing 5% CO2. The culture medium was exchanged every 3–4 days. At a doubling time of about 30 h, confluent cultures were split every 4–6 days using trypsin/EDTA.

The extents of necrosis and apoptosis in cultures of B-CPAP cells were determined by photometric quantification of cytoplasmic histone-associated DNA fragments (mononucleosomes and oligonucleosomes) using the CellDeathDetectionELISAplus (Roche Diagnostics, Penzberg, Germany). This assay discriminates between necrotic and apoptotic cells on the basis of the localization of DNA fragments. As a characteristic of necrosis, rupture of the plasma membrane liberates DNA fragments which are detectable and floating freely in the cell supernatant. In apoptosis, cell membrane integrity is typically maintained so that induced DNA fragments are only detectable after induction of cell lysis. In the CellDeathDetectionELISAplus assay, aliquots were transferred immediately after irradiation and 2 days later (for detection of apoptosis in both cases after cell lysis) from each well of the incubation plate to an ELISA multititre plate. They were then incubated with specific peroxidase-conjugated monoclonal antibodies directed against DNA and histones. After removing the unbound antibodies by washing the cells, peroxidase substrate was added until sufficient colour for photometric analysis had developed. The sample absorbance in the multititre plate was measured with an ELISA reader. The results were expressed as enrichment factors, relative to the control group, of DNA fragments (from necrosis or apoptosis) released as a function of 131I activity concentration. Corrections were made for background (incubation buffer + peroxidase substrate).

Irradiation with

131

I

4

Aliquots of 10 cells/well were plated in multititre plates in a volume of 100 ml/well of activity-free medium and were pre-cultured before irradiation for 1 day. Irradiation was started by adding 100 ml/well of different 131I solutions in final activity concentrations of 0.1–185 MBq
ml – 1 medium (prepared from soluble sodium 131I and RPMI medium). In general, each test group of the different 131I activity concentrations included eight wells. Untreated, non-irradiated control cultures were included in all experiments. All plates were incubated for 2 days with 131 I. After irradiation, changes in cell viability, necrosis and apoptosis were evaluated immediately. For additional investigation of longer-term effects of 131I, medium was replaced after irradiation in some test groups with nonradioactive medium and incubation was continued for 2 days before evaluation. The longer-term assays were done for detection of apoptosis that did not immediately follow irradiation but required a longer activation time [22]. At a cell doubling time of about 30 h, induction of the apoptotic process in up to three cell cycles (2 days of irradiation + 2 days of continued incubation) could be detected. Cell viability assay

To evaluate the cytotoxic effect of 131I the WST-1 assay (Roche Diagnostics, Penzberg, Germany) was used. Upon incubation with viable cells, the tetrazolium ring of WST (4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]1,3-benzene disulfonate) is cleaved by mitochondrial dehydrogenases to yield a purple formazan product [23– 25]. Briefly, aliquots of the WST-1 solution were added to each well for substrate reaction either immediately or 2 days after irradiation. The absorbance was measured at the respective wavelength with an ELISA plate reader (ELISA reader ‘SpectraIII’, Crailsheim, Germany). The photometric absorbance is correlated with the activity of mitochondrial dehydrogenases and therefore closely reflects the number of metabolically active, viable cells in the culture. The results were expressed as relative

Calculation of absorbed dose

The 131I dose applied to the cells was calculated using a modification of the Marinelli formula. Assuming that the activity is distributed throughout the soluble milieu in each well and given the long range (0.5–2.2 mm) of the 131 I beta particles, the cell monolayer will be continuously and uniformly irradiated even without any intracellular 131I uptake. Cells were exposed for 2 days to 131I present in the culture medium. After 2 days more than 10% of the 131I activity had decayed and may have led to radiation damage; the remaining activity is eliminated on medium exchange. The formula used was adjusted for the irradiation time of 2 days.

Results Cell viability (WST-1 assay)

At medium doses (activity concentrations from 1 to 10 MBq
ml – 1) a proliferation increase of up to 70% (compared to controls) was detected (Fig. 1(a)). After 2 days of continued activity-free incubation this viabilitystimulating effect was no longer detected, but a significant reduction in viable cells was found (Fig. 1(b)).

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Cell death after

Fig. 1

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I Marx et al. 355

Fig. 2

(a) 2.0

(a) 4.0

1.8

3.5

1.6

3.0

1.4 2.5

1.2 1.0

2.0

0.8

1.5

0.6

1.0

0.4 0.5

0.2 0.0 0.01

0.1 (0.4 Gy) 1 (4.3 Gy) 10 (43 Gy) 100 (432 Gy) 1000

0.0 0. 01

Activity concentration (MBq·ml )/calculated dose (Gy)

(b) 2.0

(b) 4.0

1.8

3.5

1.6

3.0

1.4

0.1 (0.4 Gy)

1 (4.3 Gy) 10 (43 Gy) 100 (432 Gy) 1000

Activity concentration (MBq.ml−1)/calculated dose (Gy)

−1

2.5

1.2 2.0

1.0 0.8

1.5

0.6

1.0

0.4

0.5

0.2 0.0 0.01

0.0 0.01 0.1 (0.4 Gy) 1 (4.3 Gy) 10 (43 Gy) 100 (432 Gy) 1000

0.1 (0.4 Gy) 1 (4.3 Gy) 10 (43 Gy) 100 (400 Gy) 1000 Activity concentration (MBq.ml−1)/calculated dose (Gy)

Activity concentration (MBq·ml−1)/calculated dose (Gy)

(a) Cell viability dose–response immediately after 2 days of irradiation with 131I. (b) Cell viability dose–response after 2 days continued incubation in activity-free medium (following 2 days of irradiation with 131 I). Both panels: Cell viability relative to control (enrichment factors, control = 1).

At higher doses (activity concentrations > 10–100 MBq
ml – 1) a reduction in viability of up to 50% was detected immediately after incubation with 131I (Fig. 1(a)), while continued incubation for 2 days led to a further 40% decrease of cell viability in some samples (Fig. 1(b)). Necrosis and apoptosis (CDD-ELISAplus)

Necrosis was seen very rarely at low activity concentrations ( < 1 MBq
ml – 1). An exponential increase of necrosis of 150–350% was detected with rising 131I activity concentrations > 1MBq
ml – 1 (Fig. 2(a and b)). At medium doses (activity concentrations from 1 to 10 MBq
ml – 1) necrosis was moderately increased (150– 180%) after continued incubation for 2 days (Fig. 2(b)). Corresponding to the reduction of cell proliferation (Fig. 1(a and b)), higher activity concentrations ( > 10–100 MBq
ml – 1) produced significant necrosis

(a) Necrotic dose–response immediately after 2 days of irradiation with 131 I. (b) Necrotic dose–response after 2 days continued incubation in activity-free medium (following 2 days of irradiation with 131I). Both panels: Necrosis relative to control (enrichment factors, control = 1).

immediately after irradiation and this was then the predominant form of cell death. Apoptosis was not detected immediately after irradiation (Fig. 3(a)). At 2 days after irradiation B-CPAP cells showed a significant enrichment of DNA fragments from apoptosis, peaking at about 5 MBq
ml – 1 (Fig. 3(b)). The lower activity concentration at which the apoptotic process was detected corresponds to about 1 MBq
ml – 1. At higher doses a higher percentage of cells underwent apoptosis (increase of about + 180%), while at activity concentrations > 10 MBq
ml – 1 where necrosis was predominant the percentage of apoptotic cells fell (Fig. 3(b)).

Discussion To clarify the mechanism of action of RIT of thyroid carcinoma, the relative contributions of apoptosis and necrosis of the cytotoxic effect of 131I on a thyroid cancer

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was only temporary and no longer detected in evaluations after continued incubation, when cell viability decreased by 50%. How far the observed increase in proliferation might reflect a malignant cellular transformation/degeneration induced by the lower acitivity concentrations remains speculative, according to our data.

Fig. 3

(a) 2.0 1.8 1.6 1.4 1.2 1.0 0.8 0.6 0.4 0.2 0.0 0 1(4.3 Gy)

2

3 (13 Gy) 4

5 (22 Gy)

6

7 (30 Gy)

Activity concentration (MBq·ml−1)/calculated dose (Gy)

(b) 2.0 1.8 1.6 1.4 1.2 1.0 0.8 0.6 0.4 0.2 0.0 0.01

0.1 (0.4 Gy) 1 (4.3 Gy) 10 (43 Gy) 100 (432 Gy) 1000 Activity concentration (MBq·ml−1)/calculated dose (Gy)

(a) Apoptotic dose–response immediately after 2 days of irradiation with 131I. These experiments were made at the lower dose range only, therefore a linear x-axis scale was chosen. (b) Apoptotic dose– response after 2 days continued incubation in activity-free medium (following 2 days of irradiation with 131I). Both panels: Apoptosis relative to control (enrichment factors, control = 1).

cell line in vitro was determined. Our data showed that irradiation by 131I generally had a cytotoxic effect on B-CPAP cells. Nevertheless, at low and medium activity concentrations an increase in cell viability ( = mitochondrial dehydrogenases activity) was found immediately after irradiation. This effect disappeared after an activityfree continued incubation for a further 2 days. This radiation-induced rise in cell viability might be explained by the phenomenon of ‘hormesis’, first described by Feinendegen and colleagues [24–29]. On the other hand, an early cellular response to radiation damage (by DNA repair process or cellular changes in early apoptosis) might increase the overall activity of mitochondrial dehydrogenases and therefore only simulate an increase in the number of viable cells. The latter possibility is supported by the observation that the foregoing effect

At higher activity concentrations above a threshold of about 10 MBq/ml – 1 cell viability was significantly reduced immediately after irradiation and this fall was even more marked after incubation in activity-free medium. By our dose calculation an activity concentration of 10 MBq/ml – 1 corresponds to a radiation dose of about 40–50 Gy, which is comparable to doses achieved in external radiation applied for treatment of thyroid carcinoma (about 50–60 Gy) [30]. The cytotoxic effect of 131I at an absorbed dose of 43 Gy resulted primarily from necrosis. Increasing the absorbed dose further, to 430 Gy, sharply increased the amount of necrosis (to 300– 400% and 200–300%, respectively) of that in unirradiated control cells immediately and 2 days after the end of incubation in 131I-containing medium. Irradiation with 131I at these doses seems to exacerbate already severe cellular damage leading to direct, necrotic cell death. 131I interacts with cell material indirectly through ionization and the formation of radical species (secondary radiation effects). Further cytoplasmic, membraneous and nuclear damage at activity concentrations >10 MBq/ml – 1 appears to be lethal to B-CPAP cells. At low activity concentrations ( < 1 MBq/ml – 1, corresponding to about 5 Gy) necrosis was seen very rarely, and it is probable that the cellular damage induced by 131I is not severe enough to lead to direct, necrotic cell death at these doses. With regard to apoptosis, in general, only a moderate increase of apoptosis was observed after exposure to 131I. Directly after irradiation no significant increase in apoptosis could be detected. Increased apoptosis was measured at the chosen ‘2 day point’ after continued incubation following 131I irradiation suggesting that apoptosis required a longer ‘activation time’ before it could be detected. Apoptosis is not a synchronous process, so that cells at different stages of apoptotic death will coexist in the same cell section [31]. The invitro assay CDDplus used in our experiment can only detect intermediate stages of apoptosis, i.e., DNA fragmentation, but not the early or late stages, such as loss of membrane asymmetry and phagocytosis. Hence, our measurements might underestimate the total number of cells undergoing apoptosis in response to irradiation damage. Moreover, the apoptotic cells are removed rapidly by phagocytes in vivo [9]. In contrast, those apoptotic cells lose their cell membrane integrity (‘secondary necrosis’) under in-vitro conditions and will hence simulate the ELISA-detected characteristics of necrotic cells. It is therefore difficult to be sure whether some of the necrotic cells detected by ELISA, especially

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Cell death after

after continued incubation, were not actually apoptotic cells simulating ‘secondary necrosis’ after cell lysis in vitro. Furthermore, a high baseline rate of apoptosis is characteristically seen in papillary thyroid carcinoma under ‘physiological’ conditions [32]. With such a high ‘background level’ of apoptosis (control group), a further strong increase in apoptosis could scarcely be caused by 131I. Our data showed an apparent increase in apoptosis 2 days after continued incubation following 131I irradiation, suggesting that in B-CPAP cells the apoptotic process is more likely be a consequence of secondary effects which require a longer activation time of up to several cell cycles, as in ‘delayed interphase cell death’ or ‘delayed mitotic cell death’ [20]. This may be related to the mutated form of apoptosis-associated oncoprotein p53 found in B-CPAP cells [21]. In contrast to wild-type p53, which plays an important role in the induction of ‘early apoptosis’, mutated p53 is incapable of inducing p53-associated apoptosis [33,34]. Apoptosis was significantly increased, compared to controls, 2 days after irradiation at medium doses (activity concentrations 1–10 MBq), peaking at about 5 MBq
ml – 1, which corresponds to a thyroid dose of about 20 Gy by our calculation. Cell death caused by medium 131I doses could be attributed to an activation of the apoptotic process, possibly delayed after several cell cycles or a longer process of unsuccessful or improper repair of cellular damage. With medium doses a moderate increase of necrosis (150–180%) was measured after continuing incubation for 2 days. As noted, a portion of the cells identified as necrotic may actually have been lysed apoptotic cells.

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Fig. 4

3.0

3--30 Gy

(‘Stunning dose range’)

2.5

300 Gy (Ablation) Necrosis

2.0 1.5 Apoptosis 1.0 0.5 0 0.01

Cell viability

N

*

0.1 (0.4 Gy) 1 (4 Gy) 10 (40 Gy) 100 (400 Gy)

1000

Activity concentration (MBq.ml−1)/calculated dose (Gy)

Overview: Changes in cell viability, necrosis and apoptosis after 2 days continued incubation in activity-free medium (following 2 days of irradiation). Cell viability diminishes with rising 131I activity concentration. At medium activity concentrations (1–10 MBq
ml – 1) apoptosis is predominant, while necrosis is only moderate. At higher activity concentrations ( > 10 MBq
ml – 1), apoptosis decreases and necrosis is predominant. A cytotoxic radiation effect (apoptosis > necrosis) is already observed, even within the stunning dose range (3–30 Gy) discussed above [35–39]. *‘Early’ cell viability directly after irradiation; phenomenon of ‘hormesis’? [26–29]. Enrichment factors (control = 1), ‘late effects’ 2 days after irradiation.

necrosis (Fig. 4) [35–39]. This observation should be considered in further discussions on the stunning phenomenon.

References 1

Conclusion In conclusion, our data show that irradiation with 131I has a generally cytotoxic effect on B-CPAP cell cultures. The mechanism of cytotoxicity, that is, whether primarily necrosis or apoptosis, depends on the absorbed dose of 131 I, with apoptosis predominating at lower absorbed doses and necrosis at high absorbed doses. Thus, the use of high doses of 131I appears to favour necrotic phenomena and an inflammatory reaction in the surrounding tissue, as observed in about 5–20% of highactivity RIT cases [14]. The use of lower 131I doses is biologically characterized by apoptotic phenomena which are less serious as they are not accompanied by inflammatory reactions. Predominance of apoptosis could explain why RIT at lower activities is often associated with a delayed onset and a possible continuation of thyroid volume reduction over some months and even up to a year [17]. Interestingly, the low-dose range of 3–30 Gy (1–10 MBq
ml – 1), previously described as the ‘stunning dose range’ already produced cytotoxic cell damage, resulting in apoptosis and to some extent

Greig WR. The treatment of thyrotoxicosis with radioactive iodine. Scot Med J 1966; 11:307–314. 2 Dietlein M, Dressler J, Farahati J, Schicha H, Schober O. Procedure guidelines for radioiodine therapy of differentiated thyroid cancer (version 2). Nuklearmedizin 2004; 43:115–120. 3 Horst W, Rosler H, Schneider C, Labhart A. 306 cases of toxic adenoma: clinical aspects, findings in radioiodine diagnostics, radiochromatography and histology; results of 131-I and surgical treatment. J Nucl Med 1967; 8:515–528. 4 Schicha H, Dietlein M. Radioiodine therapy in differentiated thyroid gland carcinoma. Zentralbl Chir 1997; 122:266–273. 5 Wieler H, Bartenstein P, Becker HP, Bell E, Decker P, Jakob R, et al. Guideline for therapy of malignant thyroid tumors: pleading for an actualisation. Nuklearmedizin 2004; 43:121–123. 6 Haq MS, McCready RV, Harmer CL. Treatment of advanced differentiated thyroid carcinoma with high activity radioiodine therapy. Nucl Med Commun 2004; 25:799–805. 7 Kerr JF, Winterford CM, Harmon BV. Apoptosis. Its significance in cancer and cancer therapy. Cancer 1994; 73:2013–2026. 8 Tomei LD, Cope RO. Current Communications in Cell and Molecular Biology. New York: Cold Spring Harbor; 1991; 3:410. 9 Savill JS, Wyllie AH, Henson JE, Walport MJ, Henson PM, Haslett C. Macrophage phagocytosis of aging neutrophils in inflammation. Programmed cell death in the neutrophil leads to its recognition by macrophages. J Clin Invest 1989; 83:865–875. 10 Steller H. Mechanisms and genes of cellular suicide. Science 1995; 267:1445–1449. 11 Van Furth R, Van Zwet TL. Immunocytochemical detection of 5-bromo-2deoxyuridine incorporation in individual cells. J Immunol Methods 1988; 108:45–51.

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12 Wyllie AH, Kerr JF, Currie AR. Cell death: the significance of apoptosis. Int Rev Cytol 1980; 68:251–306. 13 Albrecht HH, Creutzig H. Funktionsszintigraphie der Speicheldru¨sen nach hochdosierter Radiojodtherapie. Fortschr Ro¨ntgenstr 1977; 125: 541–551. 14 Alexander C, Bader JB, Schaefer A, Finke C, Kirsch CM. Intermediate and long-term side effects of high-dose radioiodine therapy for thyroid carcinoma. J Nucl Med 1998; 39:1551–1554. 15 Allweiss P, Braunstein GD, Katz A, Waxman A. Sialadenitis following I-131 therapy for thyroid carcinoma: concise communication. J Nucl Med 1984; 25:755–758. 16 Spiegel W, Reiners C, Bo¨rner W. Einschra¨nkungen der Speicheldru¨senfunktion nach hochdosierter Radiojodtherapie. Nuklearmediziner 1986; 3:159–166. 17 Dederichs B, Otte R, Klink JE, Schicha H. Volume reduction of the thyroid after radioiodine therapy in patients with autonomous goiter and Basedow’s goiter. Nuklearmedizin 1996; 35:164–169. 18 Erdogan MF, Kucuk NO, Anil C, Aras S, Ozer D, Aras G, Kamel N. Effect of radioiodine therapy on thyroid nodule size and function in patients with toxic adenomas. Nucl Med Commun 2004; 25:1083–1087. 19 Lennon SV, Martin SJ, Cotter TG. Dose-dependent induction of apoptosis in human tumour cell lines by widely diverging stimuli. Cell Prolif 1991; 24:203–214. 20 Radford IR, Murphy TK, Radley JM, Ellis SL. Radiation response of mouse lymphoid and myeloid cell lines. Part II. Apoptotic death is shown by all lines examined. Int J Radiat Biol 1994; 65:217–227. 21 Fabien N, Fusco A, Santoro M, Barbier Y, Dubois PM, Paulin C. Description of a human papillary thyroid carcinoma cell line. Morphologic study and expression of tumoral markers. Cancer 1994; 73:2206–2212. 22 Menz R, Andres R, Larsson B, Ozsahin M, Trott K, Crompton NE. Biological dosimetry: the potential use of radiation-induced apoptosis in human T-lymphocytes. Radiat Environ Biophys 1997; 36:175–181. 23 Liu SQ, Saijo K, Todoroki T, Ohno T. Induction of human autologous cytotoxic T lymphocytes on formalin-fixed and paraffin-embedded tumour sections. Nature Med 1995; 1:267–271. 24 Shirahata S, Watanabe J, Teruya K, Yano T, Osada K, Ohashi H, et al. E1A and ras oncogenes synergistically enhance recombinant protein production under control of the cytomegalovirus promoter in BHK-21 cells. Biosci Biotechnol Biochem 1995; 59:345–347. 25 Teruya K, Yano T, Shirahata S, Watanabe J, Osada K, Ohashi H, et al. Ras amplification in BHK-21 cells produces a host cell line for further rapid

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Original article

Radioactive iodine treatment in medullary thyroid carcinoma Murat Faik Erdogan, Alptekin Gursoy, Gurbuz Erdogan and Nuri Kamel Background Elevated levels of basal and stimulated calcitonin are commonly seen in hereditary and sporadic medullary thyroid cancer (MTC) following total thyroidectomy. The cause of these high levels can be residual thyroid tissue, possibly with C-cell hyperplasia, and/or residual micro-MTC foci. MTC does not have the ability to concentrate radioactive iodine. However, radioactive iodine trapped by thyroid follicular cells may affect the neighbouring parafollicular cells. Aim To investigate the effect of radioactive iodine treatment as adjuvant therapy to surgery in seven patients with persistent elevation of basal and stimulated calcitonin levels.

calcitonin levels were decreased significantly in only one. At follow-up, of the three patients who showed no decrease in basal and stimulated calcitonin levels, two developed further regional lymph node and distant metastases. Conclusions In patients with persistently elevated basal and stimulated calcitonin levels, radioactive iodine treatment may be the therapy of choice for C-cell hyperplasia and/or micro-MTC after optimal thyroid surgery, especially if the disease has not spread beyond c 2006 the thyroid gland. Nucl Med Commun 27:359–362 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:359–362

Methods Pentagastrin testing was performed in each case immediately before surgery and at intervals of 6 months over a maximum period of 5 years (range, 44–60 months) after surgery. Results A significant decrease in basal and stimulated calcitonin levels was observed in three patients whose disease was localized to the thyroid gland at the final visit. In the remaining four patients, who initially had lymph node involvement at surgery, basal and stimulated

Introduction Medullary thyroid cancer (MTC) is a form of neuroendocrine neoplasia in which the tumour originates from the parafollicular cells (known as C-cells) of the thyroid gland. MTC can be hereditary or sporadic [1,2]. Surgery is the main treatment strategy, as the tumours do not accumulate radioactive iodine and do not respond well to external beam radiation or conventional cytotoxic chemotherapy [2]. However, even in thyroidectomies performed by very experienced surgeons, diseased tissue may be left behind. This tissue may contain C-cell hyperplasia (CCH) and/or residual microscopic foci of MTC, especially in the context of hereditary cases. Such residual tissue frequently causes elevated basal and stimulated calcitonin levels, and may lead to tumour recurrence [3,4]. This is especially true in patients with hereditary MTC with multifocal or bilateral diffuse CCH [5], as these patients have ret proto-oncogene mutations, meaning that, theoretically, residual C-cells can form a new MTC focus. Calcitonin is a very useful tumour marker for the diagnosis of MTC and for the assessment of the response

Keywords: calcitonin, medullary thyroid cancer, radioactive iodine, treatment Department of Endocrinology and Metabolism, School of Medicine, Ankara University, Ankara, Turkey. Correspondence to Alptekin Gursoy, Department of Endocrinology and Metabolism, School of Medicine, Ankara University, Ek Bina, M1 06100, Samanpazari, Ankara, Turkey. Tel: + 90-312-3094505; fax: + 90-312-3094505; e-mail: [email protected] Received 28 August 2005 Accepted 16 January 2006

to treatment. The serum calcitonin level is related to tumour load, and increases or decreases in serum calcitonin reflect the efficacy of treatment [6]. Serum basal calcitonin levels below 10 pg/ml and stimulated calcitonin levels below 30 pg/ml, measured by radioimmunoassay, are generally accepted to be cure criteria in the treatment of MTC [7,8]. In the normal thyroid gland, parafollicular cells are scattered amongst thyroid follicles, but these cells are not easily identified by light microscopy [1,9]. Any inflammation or necrosis that destroys thyroid follicular cells may also affect C-cells, as these cells are located very close to normal thyrocytes. Radioactive iodine treatment causes severe destruction, necrosis and fibrosis of thyroid follicle cells [10]. Normal and malignant C-cells do not concentrate radioactive iodine. The 131 I isotope used for radioactive iodine therapy becomes trapped in thyroid follicular cells and affects tissues in the surrounding 2 mm by b-ray emission [11–13]. The main aim of this study was to observe the effects of radioactive iodine as adjunctive therapy in patients who

c 2006 Lippincott Williams & Wilkins 0143-3636

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360 Nuclear Medicine Communications 2006, Vol 27 No 4

had undergone surgery for MTC and had persistently elevated calcitonin levels.

Methods Patients

The study involved patients with hereditary or sporadic MTC who were followed up in our department and fulfilled the following criteria: histopathological diagnosis of MTC; basal and/or stimulated serum calcitonin level above 30 pg/ml (measured by radioimmunoassay) at least 6 months after MTC surgery; no distant metastasis, as confirmed by extensive diagnostic imaging, including cervical ultrasonography and cervical, thoracic and abdominal computed tomography. Seven patients (five women and two men; mean age, 41 ± 8.1 years; age range, 29–55 years) met these criteria. Informed consent was obtained from each participant. The patients’ clinical and demographic characteristics are presented in Table 1.

slow intravenous injection over 3 min at a dose of 0.5 mg/kg. Blood was drawn from an antecubital vein via a lightly heparinized indwelling cannula. Serum calcitonin levels were measured using a Diagnostics Systems Laboratories (DSL) (Webster, Texas, USA) calcitonin radioimmunoassay kit. The sensitivity was 3.5 pg/ml. The intra-assay and inter-assay coefficients of variation were 5.3% and 7.1%, respectively, at 75 pg/ml. DNA analysis for ret proto-oncogene mutations

Pentagastrin test and measurement of serum calcitonin

In each case, genomic DNA was extracted from lymphocytes in a sample of peripheral blood collected in an ethylenediaminetetraacetic acid (EDTA) tube. Analysis was performed with the Heliosis DNA extraction module (Metis Biotechnology Ltd., Ankara, Turkey) according to the manufacturer’s protocol. Polymerase chain reaction (PCR) was used to amplify exons 10, 11, 13, 14, 15 and 16 of the ret proto-oncogene. This was performed using 50 ml reaction mixtures (one for each primer set for each exon) containing 75 mM Tris/HCl, pH 8.8, 200 mM (NH4)2SO4, 0.1% Tween-20, 1.5 mM MgCl2, 50 mM of each deoxynucleoside triphosphate (dNTP), 50 pmol of each of the two primers, 1.25 U of Taq DNA polymerase (DNAmp Ltd., Farnborough, UK) and 50 ng of genomic DNA. The PCR machine (an automated thermocycler; MJR, Watertown, Massachusetts, USA) was programmed for 3 min of denaturation at 941C, followed by 30 cycles of 30 s at 941C, different settings for the amplification of each exon (1 min at 681C for exon 10, 651C for exons 11 and 15, 621C for exons 13 and 16, 701C for exon 14) and 5 min of annealing at 721C.

Pentagastrin testing was performed in each case immediately before and 6 months after radioactive iodine therapy. Patients were followed up at intervals of 6 months over a maximum period of 5 years (range, 44–60 months). Pentagastrin testing was repeated at each visit. Blood samples were collected for serum calcitonin evaluation at 0 (baseline), 2, 5, 7 and 10 min after pentagastrin injection. The injection of pentagastrin and blood collection for serum calcitonin were performed with the patient in the supine position. Each patient fasted overnight (including abstinence from caffeine and tobacco) prior to testing, and pentagastrin (Peptavlon; Zeneca Pharma, Cergy, France) was administered as a

The amplification products were purified using a PCR Product Purification System (Metis Biotechnology Ltd.), followed by analysis by electrophoresis in a 2% agarose gel. Each purified product was sequenced by the cycle sequencing method using a dye terminator cycle sequencing kit (Amersham Pharmacia, Piscataway, New Jersey, USA), according to the manufacturer’s protocol, and the OpenGene automated DNA sequencing system (Visible Genetics, Toronto, Ontario, Canada). The sequences were compared with ret gene proto-oncogene data obtained from the GenBank database (Accession no. AJ243297).

Surgical procedures and histopathological examination

All patients had undergone standard surgical treatment for MTC, including total thyroidectomy and central lymph node dissection. Four had also undergone both central and laterocervical node dissection. Each thyroidectomy specimen was examined using standard histopathological techniques. The entire specimen was embedded in a paraffin block, and sections were prepared with haematoxylin and eosin staining and immunohistochemical staining.

Table 1 Clinical and demographic characteristics of the seven patients in this study Patients age (years)/sex 29/female 40/male 55/female 39/female 45/female 44/male 36/female

Type of medullary Mutation detected thyroid carcinoma MEN2A MEN2A MEN2A Sporadic Sporadic Sporadic Sporadic

Codon 634 Codon 634 Codon 634 – – – –

Extent of disease Localized LNI Localized Localized LNI LNI LNI

LNI, local lymph node involvement; MEN2A, multiple endocrine neoplasia type 2A.

Results As shown in Table 1, four patients had sporadic MTC and three had hereditary MTC. The last three patients were from three different multiple endocrine neoplasia type 2A (MEN2A) families. All the MEN2A patients had a cysteine to arginine mutation at codon 634 of exon 11 of the ret proto-oncogene. At initial diagnosis with MTC, three of the seven patients had disease restricted to the thyroid gland, and the other four had cervical lymph node involvement. None of the patients had distant metastasis at the time of diagnosis.

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Radioactive iodine treatment in medullary thyroid carcinoma Erdogan et al. 361

Table 2 Patients

Patients’ clinical and laboratory characteristics before Extent of disease Recurrence after 131 before 131I I

1 2 3 4 5 6 7

Localized LNI Localized Localized LNI LNI LNI

No No No No No Yes Yes

RAI (mCi)

131

I treatment and at the final visit

Calcitonin levels before

100 100 100 100 100 150 200

131

I

Calcitonin levels at final visit

B

S

B

S

61 21 17 925 1309 1815 1708

1859 288 101 1815 3214 4736 3610

45 9 9 8 874 591 2026

297 21 14 24 2025 6500 4206

Follow-up (months) 47 46 45 60 48 44 52

LNI, local lymph node involvement; RAI, radioactive iodine; B, basal calcitonin; S, stimulated calcitonin.

All seven patients received one dose of 131I treatment in the range 100–200 mCi. None experienced side-effects related to radioactive iodine treatment. The three individuals with disease restricted to the thyroid gland showed a significant decrease in basal and stimulated serum calcitonin levels after radioactive iodine therapy. Two had serum basal calcitonin levels of less than 10 pg/ml and stimulated calcitonin levels of less than 30 pg/ml (generally accepted cure criteria in the treatment of MTC). Only one of the four patients who underwent lymph node dissection showed a significant decrease in basal and stimulated serum calcitonin levels. None of the patients who initially had cervical lymph node involvement attained cure criteria. The results of the pentagastrin stimulation tests before radioactive iodine treatment and at the final visit are presented in Table 2. The three patients who showed no decline in serum basal and stimulated calcitonin levels at the final visit had different findings and outcomes during follow-up. One individual (patient 7) developed mediastinal, nasopharyngeal and paranasal sinus metastases. Another (patient 6) underwent radical neck dissection in which a lymph node metastasis was revealed. The third patient with fine needle aspiration-proven lymph node metastasis refused further surgical intervention. During the pentagastrin stimulation tests, all patients experienced adverse effects, including substernal discomfort, nausea, metallic taste in the mouth, abdominal cramping, flushing, warmth, tachycardia and headache, after the injection of pentagastrin. No serious complications occurred during the tests.

Discussion Normal and hyperplastic parafollicular C-cells do not have the ability to trap iodine. MTC, in contrast with differentiated thyroid carcinoma, fails to concentrate iodine and thus is not routinely treated with radioactive iodine [14]. However, in certain studies, it has been observed that radioactive iodine treatment can lead to a decrease in elevated post-operative calcitonin levels, which are thought to be due to residual tumour, even in metastatic MTC cases [15–18].

However, other studies have reported that radioactive iodine treatment has no value in the therapy of MTC cases. In two separate studies performed on sporadic and hereditary MTC cases with residual biochemical MTC, it was shown that radioactive iodine treatment following surgery did not cause a significant decrease in calcitonin levels, change the survival rates or decrease the tumour recurrence [19,20]. The underlying mechanism of 131I treatment in MTC cases responding to radioactive iodine is not yet understood. Follicular carcinoma foci can be found in primary MTC tumours and MTC metastases. As a result, 131 I treatment may be useful in these mixed-type tumours [15–18]. Indeed, MTC cases showing follicular histopathology and behaviour, and trapping radioactive iodine in their metastases, have been reported; these types of mixed tumour have been named ‘pseudomedullary carcinoma’ [21,22]. Another more probable mechanism is that follicular cells, remaining in the thyroid bed after total thyroidectomy, may trap a sufficient amount of isotope and thus destroy adjacent non-iodine-trapping cancer cells, especially if they are located within the short range of the b-radiation emitted by radioactive iodine. According to calculations, a 1 mm piece of residual tissue in the thyroid gland is exposed to a radiation dose of approximately 340 Gy by 131I treatment. This dose is much higher than the external radiotherapy dose, although it is limited by the penetration of b-rays, which is only around 2 mm [10]. When the histopathological distribution of C-cells amongst follicular cells is considered, although, theoretically, this penetration is not effective in larger tumours, it can be effective in benign or malignant CCH or micromedullary carcinoma foci. It is known that the 131I radiation dose used for differentiated thyroid cancer is not harmful to vital tissues, such as the trachea, oesophagus and spinal cord [23]. In reports supporting the concept that b-rays affect parafollicular cells following 131I application, neonatal rat thyroid C-cells have been found to be very sensitive to radioactive iodine; the gland tissue showed a dramatic decrease in the number of C-cells and calcitonin levels [11–13]. In two different reports, basal and stimulated calcitonin levels were significantly decreased in patients with Graves’ disease or toxic nodular goiter who were

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treated with 131I [24,25]. The destruction of the vascular support of parafollicular cells by the inflammation and fibrosis caused by radioactive iodine may have a role in this effect [24].

3

4 5

In this study, radioactive iodine treatment was given to seven MTC patients (three with disease localized to the thyroid gland and four with lymph node involvement) with elevated post-operative basal and stimulated calcitonin levels and no evidence of distant metastases. Basal and stimulated calcitonin levels were measured before surgery and at intervals of 6 months over a maximum period of 5 years after radioactive iodine treatment. Three patients whose disease was localized to the thyroid gland showed a significant decrease in basal and stimulated calcitonin levels after radioactive iodine treatment. In two of these patients, during the MTC follow-up period, basal and stimulated calcitonin levels decreased to the desired values. Only one of the remaining four patients with lymph node involvement initially showed a significant decrease in basal and stimulated calcitonin levels. None of the patients with lymph node involvement initially attained cure criteria. At follow-up of the three patients with no decline in basal and stimulated calcitonin levels after radioactive iodine treatment, two developed regional lymph node and/or distant metastases. In conclusion, radioactive iodine treatment may possibly be regarded as adjuvant therapy to surgery in patients with persistently elevated basal and stimulated calcitonin levels, especially for residual CCH and/or micro-MTC foci left in the thyroid bed after thyroid surgery. Theoretically, this approach may be more important in hereditary cases, which are almost always multifocal with concomitant diffuse CCH. In patients with lymph node involvement in whom sufficient lymph node dissection has been performed, but with continuing high levels of basal and stimulated calcitonin post-operatively, radioactive iodine treatment could be applied, assuming that the high levels originate from the thyroid bed. If there is a lack of response after radioactive iodine treatment, an extensive search for lymph node and distant metastases should be undertaken. In theory, decreased calcitonin levels following radioactive iodine treatment can provide a survival advantage. Clearly, further longitudinal data are required to clarify the possible role of radioactive iodine treatment in MTC patients.

References 1 2

Hazard JB. The C cells (parafollicular cells) of the thyroid gland and medullary thyroid carcinoma. A review. Am J Pathol 1977; 88:213–250. Lairmore TC, Wells SA Jr. Medullary carcinoma of the thyroid: current diagnosis and management. Semin Surg Oncol 1991; 7:92–99.

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Becker KL, Silva OL, Snider RH, Moore CF, Geelhoed GW. The surgical implications of hypercalcitonemia. Surg Gynecol Obstet 1982; 154: 897–908. Raue F. Postsurgical follow-up and management. Recent Results Cancer Res 1992; 125:197–211. Wolfe HJ, Melvin KE, Cervi-Skinner SJ, Saadi AA, Juliar JF, Jackson CE, et al. C-cell hyperplasia preceding medullary thyroid carcinoma. N Engl J Med 1973; 289:437–441. Tashjian AH Jr, Wolfe HJ, Voelkel EF. Human calcitonin. Immunologic assay, cytologic localization and studies on medullary thyroid carcinoma. Am J Med 1974; 56:840–849. Guilloteau D, Perdrisot R, Calmettes C, Baulieu JL, Lecomte P, Kaphan G, et al. Diagnosis of medullary carcinoma of the thyroid (MCT) by calcitonin assay using monoclonal antibodies: criteria for the pentagastrin stimulation test in hereditary MCT. J Clin Endocrinol Metab 1990; 71:1064–1067. Barbot N, Calmettes C, Schuffenecker I, Saint-Andre JP, Franc B, Rohmer V, et al. Pentagastrin stimulation test and early diagnosis of medullary thyroid carcinoma using an immunoradiometric assay of calcitonin: comparison with genetic screening in hereditary medullary thyroid carcinoma. J Clin Endocrinol Metab 1994; 78:114–120. Perry A, Molberg K, Albores-Saavedra J. Physiologic versus neoplastic C-cell hyperplasia of the thyroid: separation of distinct histologic and biologic entities. Cancer 1996; 77:750–756. Thurston V, Williams ED. The effect of radiation on thyroid C cells. Acta Endocrinol (Copenhagen) 1982; 99:72–78. Shah KH, Oslapas R, Calandra DB, Prinz RA, Ernst K, Hofmann C, et al. Effects of radiation on parafollicular C cells of the thyroid gland. Surgery 1983; 94:989–994. Ott RA, Hofmann C, Oslapas R, Nayyar R, Paloyan E. Radioiodine sensitivity of parafollicular C cells in aged Long-Evans rats. Surgery 1987; 102: 1043–1048. Feinstein RE, Gimeno EJ, el-Salhy M, Wilander E, Walinder G. Evidence of C-cell destruction in the thyroid gland of mice exposed to high 131I doses. Acta Radiol Oncol 1986; 25:199–202. Ljungberg O. Medullary carcinoma of the human thyroid gland. Autoradiographic localization of radioiodine. Pathol Microbiol Scand 1966; 68:476–480. Hellman DE, Kartchner M, Van Antwerp JD, Salmon SE, Patton DD, O’Mara R. Radioiodine in the treatment of medullary carcinoma of the thyroid. J Clin Endocrinol Metab 1979; 48:451–455. Deftos LJ, Stein MF. Radioiodine as an adjunct to the surgical treatment of medullary thyroid carcinoma. J Clin Endocrinol Metab 1980; 50: 967–968. Nusynowitz ML, Pollard E, Benedetto AR, Lecklitner ML, Ware RW. Treatment of medullary carcinoma of the thyroid with I-131. J Nucl Med 1982; 23:143–146. Michael BE, Forouhar FA, Spencer RP. Medullary thyroid carcinoma with radioiodide transport. Effects of iodine-131 therapy and lithium administration. Clin Nucl Med 1985; 10:274–279. Saad MF, Guido JJ, Samaan NA. Radioactive iodine in the treatment of medullary carcinoma of the thyroid. J Clin Endocrinol Metab 1983; 57: 124–128. Nieuwenhuijzen Kruseman AC, Bussemaker JK, Frolich M. Radioiodine in the treatment of hereditary medullary carcinoma of the thyroid. J Clin Endocrinol Metab 1984; 59:491–494. Hales M, Rosenau W, Okerlund MD, Galante M. Carcinoma of the thyroid with a mixed medullary and follicular pattern: morphologic, immunohistochemical, and clinical laboratory studies. Cancer 1982; 50:1352–1359. Spencer RP, Garg V, Raisz LG, Karimeddini MK, Penikas V, Yamase H, et al. Radioiodide uptake and turnover in a pseudo-medullary thyroid carcinoma. J Nucl Med 1982; 23:1006–1010. Snyder WS, Fisher HL Jr, Ford MR, Warner GG. Estimates of absorbed fractions for monoenergetic photon sources uniformly distributed in various organs of a heterogeneous phantom. J Nucl Med Suppl 1969; 3:47–52. Body JJ, Demeester-Mirkine N, Corvilain J. Calcitonin deficiency after radioactive iodine treatment. Ann Intern Med 1988; 109:590–591. Bayraktar M, Gedik O, Akalin S, Usman A, Adalar N, Telatar F. The effect of radioactive iodine treatment on thyroid C cells. Clin Endocrinol (Oxford) 1990; 33:625–630.

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Original article

Effect of a 188Re-SSS lipiodol/131I-lipiodol mixture, 188 Re-SSS lipiodol alone or 131I-lipiodol alone on the survival of rats with hepatocellular carcinoma Elienne Garina, Herve´ Rakotonirinab,c, Florence Lejeunea, Benoit Denizotb,c, Jerome Rouxc, Nicolas Noiretd, Habiba Mesbahe, Jean-Yues Herrya, Patrick Bourgueta and Jean-Jacques Lejeuneb Background and aim It has been shown that the use of a cocktail of isotopes of different ranges of action leads to an increase in the effectiveness of metabolic radiotherapy. The purpose of the present study was to compare with a control group the effectiveness of three different treatments in rats bearing hepatocellular carcinoma (HCC), using (1) a mixture of lipiodol labelled with both 131I and 188 Re, (2) lipiodol labelled with 131I alone and (3) lipiodol labelled with 188Re alone. Material and methods Four groups were made up, each containing 14 rats with the N1-S1 tumour cell line. Group 1 received a mixture composed of 22 MBq of 188Re-SSS lipiodol and 7 MBq 131I-lipiodol. Group 2 received 14 MBq 131 I-lipiodol. Group 3 received 44 MBq of 188Re-SSS lipiodol and group 4 acted as the control. The survival of the various groups was compared by a non-parametric test of log-rank, after a follow-up of 60, 180 and 273 days.

Conclusions In this study, 131I-lipiodol is the most effective treatment in HCC-bearing rats, because this is the only method that leads to a prolonged improvement of survival. These results cannot necessarily be extrapolated to humans because of the relatively small size and unifocal nature of the lesions in this study. It appears necessary to carry out a study in humans with larger tumours in order to compare these three treatments, particularly with a view to replacing 131I-labelled lipiodol by 188Re-labelled lipiodol. However, this study clearly demonstrated that, for small tumours, as in an adjuvant setting for exemple, 131 I-labelled lipiodol should be a better option than 188 Re-labelled lipiodol. Nucl Med Commun 27:363–369
c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:363–369 Keywords: metabolic radiotherapy, study

131

I,

188

Re, liver neoplasm, animal

Results Compared with the controls, the rats treated with a mixture of 188Re-SSS lipiodol and 131I-lipiodol show an increase in survival, but only from day 60 onwards (P = 0.05 at day 60 and 0.13 at days 180 and 273). For the rats treated with 131I-lipiodol, there was a highly significant increase in survival compared with the controls at day 60, day 180 and day 273 (P = 0.03, 0.04 and 0.04, respectively). There is no significant increase in survival for the rats treated with 188Re-SSS lipiodol, irrespective of the follow-up duration (P = 0.53 at day 60, 0.48 at day 180, and 0.59 at day 273).

a UPRES EA 3890/Service de Me´decine Nucle´aire, Centre Euge`ne Marquis, Rennes, bINSERM U646, Angers, cAnimalerie, Universite´ d’Angers, dENSCR UMR CNRS 6052, Rennes-Beaulieu and eDe´partement d’informatique me´dicale, Centre Euge`ne Marquis, Rennes, France.

Introduction

131

Hepatocellular carcinoma (HCC) is one of the most frequent cancers, being ranked fifth in importance worldwide with 437 000 new cases per year [1]. The prognosis for this cancer is extremely poor, and a curative treatment (liver transplantation, resection, ablative procedure like alcoholization or radio frequency) can only be carried out in less than 25% of cases. For patients who are not candidates for a radical procedure and who do not present with an excessively advanced form, a palliative treatment can be proposed, mainly by lipiodol-chemoembolization or arterial metabolic radiotherapy. At present,

Correspondence to Dr Etienne Garin, Centre Euge`ne Marquis, rue de la Bataille Flandres Dunkerque, CS 44229, 35042 Rennes Cedex, France. Tel: + 0033 2 99 25 30 75; fax: + 0033 2 99 25 32 60; e-mail: [email protected] This study received financial support from the League Against Cancer (Ille et Vilaine and Morbihan, France). Received 4 November 2005 Accepted 19 December 2005

I-labelled lipiodol is the most widely used therapeutic agent in arterial metabolic radiotherapy of HCC [2]. However, new methods of radiolabelling lipiodol with lipophilic complexes of 188Re have recently been described [3–8] which could be used to replace 131Ilipiodol because of the less severe radiation protection constraints and the lower production cost. However, the therapeutic effectiveness of a treatment by I-lipiodol remains a matter of debate. Indeed, except in the particular case of patients with portal thrombosis [9], there is no evidence that treatment by 131I-lipiodol

131

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364 Nuclear Medicine Communications 2006, Vol 27 No 4

leads to an improvement of survival. Moreover, studies comparing lipiodol labelled with 131I or 188Re against lipiodol chemoembolization have failed to reveal a significant difference in effectiveness between these two treatments [10,11]. These results emphasize the need to optimize the therapeutic effectiveness of intra-arterial metabolic radiotherapy using radiolabelled lipiodol. The use of a cocktail of isotopes of different paths of action [12,13] is one of various techniques already evoked to improve the therapeutic effectiveness of metabolic radiotherapy, and is an approach that could be adopted using lipiodol radiolabelled with a mixture of 131I and 188Re. The aim of this study was to compare, with a control group, the effectiveness of three different treatments on the HCC-bearing rat, involving (1) a mixture of lipiodol labelled with both 131I and 188Re, (2) lipiodol labelled with 131I alone, and (3) lipiodol labelled with 188Re alone.

Materials and methods Experiments were carried out on 56 Sprague–Dawley female rats with hepatoma weighing 227 ± 14 g, in compliance with the French regulations in force (Law 0189.4 of 24 January 1990). Tumour model Cell line

The well-established rat hepatocarcinoma cell line N1-S1 (ATCC, Maryland, USA) was used for tumour emplacement. We cultivated this cell line in a DMEM culture medium (Dulbecco’s Modified Eagle Medium, Cambrex Biosciences, Verviers, France) supplemented by 20% horse serum (Cambrex Biosciences, Verviers, France), 5% fetal calf serum (Cambrex Biosciences, Verviers, France) and 1% antibiotics (Antibiotic Antimycotic Solution (100  ) stabilized, Sigma-Aldrich, Lyon, France). After 3 days, we obtained 25  106 to 30  106 cells from an initial population of 0.8  106 cells in these cultures. Tumour inoculation

Anaesthesia was induced by intra-peritoneal injection of 0.1 ml xylazine (Rompun 2%; Bayer, Puteau, France) and ketamine (Ketalar 1000; Vetokinol, Lure, France). A subxiphoidian laparotomy was carried out from 1.5 to 2 cm long, and the left hepatic lobe was exposed and placed on a sterile compress. Then, 6  106 cells in 0.15 ml DMEM were injected under the hepatic capsule in 30 s with a 27G1/2 needle (MicrolanceTM 3; BD, New Jersey, USA). A gentle compress was then applied for 15 s with absorbent gauze composed of cotton and polyamide (B Braun, Boulogne, France) in order to avoid bleeding and reflux of the cells. The incision was closed again by two planes.

Products and activities used Products 188

Re-super six sulfur lipiodol (188Re-SSS lipiodol) was prepared as described previously [7,8]. This product consists of a lipophilic complex of 188Re solubilized in lipiodol (the complex is 188Re-(PhCS2)(PhCS3)2, abbreviated as 188Re-SSS complex). 188Re was eluted from a 188 W/188Re generator (Oak Ridge National Laboratory, Tennessee, USA). The 131I-lipiodol (Lipiocis) was obtained from Schering CISbio International (Gif sur Yvette, France). Activities used

With the animal model used here, it was not possible to place a permanent catheter or catheterize the hepatic artery twice. Indeed, the hepatic artery of the rat is too small and the injection technique (see below) rules out access via the gastroduodenal artery. For this reason, we could not carry out personalized pre-therapeutic dosimetry with an injection of a scout dose of radiolabelled lipiodol (or an angiographic study) in order to determine the level of tumour uptake and effective half-life required for a dosimetric study for each animal. Therefore, we adopted a pragmatic approach using the MIRD formalism to calculate the standard dose to inject in each batch of rats, assuming that the tumour is a sphere with a homogeneous distribution of radiolabelled lipiodol, and that the tumour forms the only source. Finally, we assumed that the tumour uptakes of 131 I-lipiodol and 188Re-SSS lipiodol were identical. In fact, even if no comparative studies have been carried out, the results obtained with 131I-lipiodol and 188Re-SSS lipiodol, as well as with lipiodol labelled by other lipophilic complexes of 188Re, all yield similar biodistributions, i.e., a predominant tumour uptake with a high tumour/non-tumour liver uptake ratio, considerable hepatic uptake, weak pulmonary uptake and an absence of uptake on other organs [3,6,8,14]. When using 131I-labelled lipiodol, the activity that is usually administered is calculated to deliver a dose at the tumour higher than 100–120 Gy [15–18]. We thus decided to deliver a dose of 120 Gy to the tumour. The delivered dose, D (in mGy) was calculated by the formula D ¼ AS where S (in mGyMBq1s1) is defined for a given radioelement and a given diameter of source; and A, the cumulated activity (in MBq), is given by the expression A ¼ 1:44Teff A0 where Teff is the effective half-life, and A0 is the initial activity in the sphere.

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Effect of

In the rat, the effective tumour half-life is 15 h for 188Relabelled lipiodol [19] and 6 days for 131I-labelled lipiodol [14]. On the other hand, the A0 activity is less precisely known, with reported activities of 10% [20], 13% [19] and 34% [3] of the injected activity in the tumour or 0.72% of the injected activity per millilitre volume [21]. Due to those considerations we used 20% as the percentage uptake of the radiolabelled lipiodol for a tumour of 1 cm diameter.

188

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I-lipiodol on HCC in rats Garin et al. 365

during the injection. The radiolabelled lipiodol was injected slowly, the needle was rinsed with 0.1 ml physiological salt solution, and then the proximal part of the gastroduodenal artery (upstream of the puncture point) was tied off. The thread around the coeliac artery was withdrawn in order to restore hepatic arterial flow.

From these various considerations, the activity of 188Re that should be injected to obtain a dose of 120 Gy at the level of a tumour of 1 cm is 44 MBq, and the corresponding activity of 131I is 14 MBq.

Four groups of 14 rats bearing hepatoma were made up. Group 1 was treated with a mixture of 188Re-SSS lipiodol and 131I-lipiodol (with 22 and 7 MBq of the respective radioelements). Group 2 was treated with 131I-lipiodol alone (14 MBq). Group 3 was treated by 188Re-SSS lipiodol alone (44 MBq). Finally, group 4, representing the control, received 0.1 ml physiological salt solution.

The activity of 188Re-SSS lipiodol used here (44 MBq) is very close to the activity of 90Y used by Lin et al. (37 MBq) in a study on HCC-bearing rats. These authors compared the effectiveness of 90Y-labelled microspheres injected directly into the tumour or into the hepatic artery [22].

After the injection, the rats were put back into cages, and their survival was studied for up to 273 days after the treatment. Surviving rats were killed 273 days after the treatment. Autopsy was performed to check the tumour status.

Thus, in our study, we injected 44 MBq of 188Re into the rats treated with 188Re-SSS lipiodol alone, and 14 MBq of 131 I into the rats treated with 131I-lipiodol alone. For the rats treated with the cocktail 188Re-SSS lipiodol/131Ilipiodol, we injected 50% of each dose – that is, 22 MBq of 188Re-SSS lipiodol and 7 MBq of 131I-lipiodol. In each case, the volume injected was 0.1 ml. Treatment

Tumour growth in the liver and the absence of extrahepatic extension (peritoneal carcinomatosis in particular) were monitored surgically at the time of administering the treatment, 14 days after inoculation. Only rats with proved macroscopic hepatoma and no extra-hepatic tumour dissemination were included in this study. The tumour was then measured. The volume, V, of the tumour was calculated using the formula p V ¼ hl e 6 where h is the height, l is the length and e is the thickness. The injection of 0.1 ml of radiolabelled lipiodol (or physiological salt solution for the control group) via the hepatic artery was carried out, after having checked the tumour status, in the same operation, as described previously [3,19,23]. Briefly, the gastroduodenal artery was identified and carefully dissected until the coeliac and the hepatic arteries were exposed. The distal part of the gastroduodenal artery was ligated, and a thread was placed around the coeliac artery to stop arterial flow temporarily (to avoid back-flow during the injection). The gastroduodenal artery was punctured upstream of the distal ligature using a 27G butterfly needle (Valu-Set; BD, New Jersey, USA). The gastroduodenal artery was ligated onto this needle in order to avoid any reflux

Statistics

The weight of the tumours between the various groups of rats was analysed by applying the Bonferroni test for comparison of the means (SPSS software). Survival curves were established at 60, 180 and 273 days after the treatment. We used the non-parametric test of log-rank test to compare survival between the different treated groups and the control group, as well as between the treated groups themselves, at days 60, 180 and 273. P r 0.05 was considered statistically significant.

Results By using this tumour model it was possible to obtain tumours weighing 2.54 ± 1.69 g 14 days after inoculation of tumour cells. The tumour induction rate was 82% (69 rats not bearing hepatoma were necessary to obtain 56 rats bearing hepatoma after inoculation of tumour cells). The tumour weight was 2.34 ± 1.36 g in the 188Re-SSS lipiodol/131I-lipiodol group, 2.73 ± 1.98 g in the 131Ilipiodol group, 2.53 ± 1.53 g in the 188Re-SSS lipiodol group and 2.56 ± 1.89 g in the control group. A comparative analysis of the mean weights of the tumours did not reveal any significant difference in weight between the various groups (P = 0.95). The sizes and weights of the tumours, and survival of the animals of the different groups are presented in Table 1. The cause of death was tumour progression, except for (1) the seven rats with early post-operative death (no evidence of tumour progression at the macroscopic postmortem examination: one in group 1, two in group 2, three in group 3, and one in group 4) and (2) the 22 rats killed 273 days after the treatment (no visual evidence of disease, even though a tumour was present initially).

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366 Nuclear Medicine Communications 2006, Vol 27 No 4

Group 1

Group 2

Group 3

Group 4

Size (mm)

Weight (g)

Survival (days)

10  14  22 11  13  14 14  20  25 15  15  20 12  16  20 15  16  18 14  15  20 15  20  20 17  17  26 10  15  15 10  10  15 10  12  18 20  22  25 16  16  23

1.16 1.05 2.87 2.35 2.01 2.26 2.19 3.14 3.93 1.17 0.78 1.13 5.75 3.08

273 77 273 17 134 273 123 3 273 24 273 273 12 164

12  15  18 11  14  24 6  7  10 9  12  16 14  15  20 17  17  20 14  15  20 20  25  30 11  14  18 16  20  28 14  17  25 15  20  28 8  10  20 11  22  30

1.69 1.93 0.21 0.91 2.19 3.02 2.19 7.85 1.45 4.69 3.11 4.39 0.83 3.81

273 273 273 1 273 23 50 143 273 273 273 273 1 63

12  20  25 8  10  12 10  10  15 11  17  20 10  14  18 10  12  16 14  20  28 20  22  25 14  14  18 14  15  28 18  20  20 15  24  26 8  15  20 11  16  24

3.14 0.51 0.78 1.95 1.13 1.01 4.11 5.75 1.84 3.07 3.76 4.91 1.25 2.21

1 273 273 237 273 17 273 41 11 273 11 5 2 91

20  28  28 10  12  20 8  10  12 10  12  13 10  15  15 13  15  22 9  10  14 14  16  18 12  18  20 20  25  28 15  15  30 15  18  24 12  15  20 16  24  26

3.34 1.25 0.62 0.81 1.17 2.24 0.73 2.11 2.26 7.33 3.53 3.39 1.88 5.22

14 1 17 273 44 33 273 13 273 25 58 31 105 18

Group 1 = 188Re-SSS lipiodol + 131I-lipiodol; group 2 = 131I-lipiodol; group 3 = 188Re-SSS lipiodol; group 4 = controls.

For the group of rats treated with the 188Re-SSS lipiodol/ 131 I-lipiodol mixture, there was an increase in survival compared with the reference group, but this is at the limit of significance (P = 0.05) at day 60. On the other hand, there was no significant difference in survival for this group, compared to the reference group, with a longer follow-up of 180 and 273 days (P = 0.13 in both cases).

120 Survival rate (%)

Group

Fig. 1

50 25 00 0

10

20

30 40 Survival (days)

50

60

Survival curves over 60 days for rats treated by the 131I-lipiodol/188ReSSS lipiodol mixture (group 1, grey line; P = 0.05), by 131I-lipiodol (group 2, red line; P = 0.03) or by 188Re-SSS lipiodol (group 3, green line; P = 0.53), as well as for the control group (group 4, blue line). Statistical significance is given in comparison to the control groups.

Fig. 2

1.00 Survival rate (%)

Table 1 Size and weight of the tumours, and survival for the different treatment groups

0.75 0.50 0.25 0.00 0

50

100 150 200 Survival (days)

250

300

Survival curves over 273 days for rats treated by the 131I-lipiodol/188ReSSS lipiodol mixture (group 1, grey line; P = 0.13), by 131I-lipiodol (group 2, red line; P = 0.04), or by 188Re-SSS lipiodol (group 3, green line; P = 0.59), as well as for the control group (group 4, blue line). Statistical significance is given in comparison to the control groups.

For the group of rats treated with 131I-lipiodol, there is a highly significant improvement of survival compared with the reference group, not only at day 60 but also at days 180 and 273 (P = 0.03 at day 60 and 0.04 at days 180 and 273). For the group of rats treated with 188Re-SSS lipiodol alone, there was no significant improvement of survival, compared to the reference group, whatever the follow-up period (P = 0.53 at day 60, P = 0.48 at day 180 and P = 0.59 at day 273). The survival curves established over 60 and 273 days are presented in Figs 1 and 2. Finally, we analysed the survival over 273 days by excluding from the study those seven animals that died at an early stage (in the 5 days following the treatment). In such cases, we might suspect post-operation complications to be responsible for the deaths as there was no

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Effect of

Re- and

131

I-lipiodol on HCC in rats Garin et al. 367

required to obtain an optimal probability of effectiveness is between 0.5 and 12 mm, while O’Donoghue et al. [12] have estimated that it is between 2.5 and 5 mm. Lesions smaller than 0.5–2.5 mm remain insensitive to this radioelement [12,13,30]. For 188Re, this optimal size is 26 mm [12]. Some authors have thus proposed combining radioelements having different ranges of action, in the case of tumour lesions of different sizes, with the aim of optimizing the effectiveness of the treatment [12,13].

Fig. 3

1.00 Survival rate (%)

188

0.75 0.50 0.25 0.00 0

50

100 150 200 Survival (days)

250

300

Survival curves over 273 days after removing early deaths ( r day 5) for rats treated by the 131I-lipiodol/188Re-SSS lipiodol mixture (group 1, grey line; P = 0.11), by 131I-lipiodol (group 2, red line; P = 0.01) or by 188 Re-SSS lipiodol (group 3, green line; P = 0.26), as well as for the control group (group 4, blue line). Statistical significance is given in comparison to the control groups.

evidence of tumour progression at the macroscopic postmortem examination of the liver. Under these conditions, there was also a very significant improvement of survival for the rats treated by 131I-lipiodol compared to the control (P = 0.01) and an absence of significant improvement in survival for the rats treated by the 188Re-SSS lipiodol/131I-lipiodol mixture or by 188ReSSS lipiodol alone (P = 0.011 and 0.026, respectively). The survival established over 273 days by excluding the early deaths is presented in Fig. 3.

Discussion Various techniques have been tested to increase the therapeutic effectiveness of radiolabelled lipiodol. The association with chemotherapy has yielded encouraging results [24,25]. The use of an emulsion gives contradictory results according to whether it is stabilized or not [26,27]. The concomitant administration of a vasoconstrictor agent has also been evoked because, in certain cases, it can lead to a preferential redistribution of arterial flow towards the tumour [28,29]. The simultaneous use of two radioelements of different ranges of action to improve the effectiveness of metabolic radiotherapy has also been mentioned [12,13,30]. This approach could be currently applied by using 131I-labelled lipiodol along with 188Re-labelled lipiodol, but no studies have so far tested this option. Theoretically, it could also be applied by using 131I-labelled lipiodol along with 90Ylabelled lipiodol but as 90Y-labelled lipiodol is not available yet this hypothesis was not tested in this study. Various studies have indeed pointed out that the sensitivity of a lesion to a given radioelement depends directly on the size of the lesion [12,13,30]. For 131I, Amin et al. [30] have estimated that the lesion size

This idea has recently been applied in vitro by the concomitant use of lipiodol labelled with 131I and 125I on a hepatocarcinoma cell line [31]. In fact, the 131I-lipiodol + 125I-lipiodol ‘cocktail’ has a greater cytotoxic effect than either 131I-lipiodol alone or 125I-lipiodol alone. The approach has also been tested in vivo on rats using analogues of somatostatin labelled with 90Y and 177Lu [32]. In this study, rats with neuroendocrinal tumours of different sizes (0.1 and 8 cm2) were treated either with a somatostatin analogue labelled with 90Y (278 MBq) or the same somatostatin analogue labelled with 177Lu (555 MBq) or a mixture of both (with 185 and 278 MBq, respectively). The survival of the rats was 57, 50 and 145 days, respectively, for the treatments with 90Y, 177Lu and the 90Y + 177Lu mixture. The large study on HCC-bearing rats presented here aims to assess the therapeutic effectiveness of a 188ReSSS lipiodol/131I-lipiodol mixture, as well as 188Re-SSS lipiodol and 131I-lipiodol in comparison with a control group. To our knowledge, it is the only study carried out to date on this subject. To evaluate the effectiveness of these various treatments, we carried out a survival study with a long follow-up time, rather than a morphological analysis of the tumour response (by computed tomography or ultrasonography) as proposed in the study by Lin et al. [22]. Indeed, when an early evaluation of the tumour response needs to be obtained, only a morphological assessment is possible. On the other hand, when an early evaluation is not essential, as in the case of this study on the animal, an evaluation of survival is preferable. Moreover, an early morphological evaluation can give erroneous results in two situations. Firstly, in the case of a real tumour response, but with replacement of the tumour by cicatricial tissue, the morphological evaluation can erroneously be an absence of response. Secondly, in the case of real tumour response, but with a secondary escape related to the selection of cellular strains resistant to the treatment, the early morphological response can be converted into an absence of response with a longer follow-up. This study highlights an improvement of the therapeutic effectiveness compared with the control group, which appears only at day 60 just above the level of significance when using the 188Re-SSS lipiodol/131I-lipiodol mixture. Indeed, at day 180 and day 273, there is no longer any

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368 Nuclear Medicine Communications 2006, Vol 27 No 4

significant difference in survival between this group and the control group. This result stresses the importance of carrying out a survival study with a prolonged follow-up. In addition, we did not detect any significant improvement in survival for the rats treated by 188Re-SSS lipiodol alone, whereas 131I-lipiodol was the most effective treatment, with a highly significant improvement in survival even with a prolonged follow-up. Certain factors could explain these disappointing results concerning the use of the 188Re-SSS lipiodol/131I-lipiodol mixture or the 188Re-SSS lipiodol alone. Firstly, this study did not find any significant improvement of survival for the group of rats treated by 188Re-SSS lipiodol alone compared to the control group. This result is probably explained by the relatively small size of the tumours (on average, 2.54 g) in the rats studied, which corresponds to a sphere of 1.68 cm diameter compared with the optimal lesion diameter of 26 mm for 188Re [12]. It would be practically impossible to obtain larger tumours with the tumour model used here because we observed the appearance of haemorrhagic ascites and a peritoneal carcinomatosis in the majority of rats having a tumour diameter larger than 2–2.5 cm (these rats were excluded from the study). This lack of effectiveness of 188Re-SSS lipiodol, alone, may be sufficient to explain the absence of improvement of the therapeutic effectiveness with the 188Re-SSS lipiodol/131I-lipiodol mixture. Secondly, the idea of improving the therapeutic effectiveness by using a cocktail of isotopes of different ranges of action, rather than the isolated use of one of the isotopes, was based on a mathematical modelling of homogeneous tumours and applied in the case of tumours of different sizes. In our study, we put forward a slightly different hypothesis: we assume that the use of a cocktail of isotopes of different ranges can improve the therapeutic effectiveness on very heterogeneous tumours. The isotopes with short paths are mainly useful for treating uptake zones, while the isotope with longer paths are for treating zones with weak uptake (paucicellular or poorly vascularized zones). With the tumour model used here, we first checked whether the tumours obtained were of a highly heterogeneous character [8]. Thus, it may be impossible to improve the therapeutic effectiveness for very heterogeneous tumours using a cocktail of radioelements. It is also possible that our assumption is valid, but that we used tumours of excessively small size. Thirdly, the presence of prolonged survivals in the control group (three rats with initial evidence of macroscopic hepatoma killed at 273 days, presenting with a macro-

scopically full response) raises a question about the aggressiveness of the tumours obtained and thus whether the tumours are equally aggressive in the various groups. Such a factor could have skewed our results. However, this phenomenon appears inevitable with hepatocellular carcinoma, considering that such observations have also been noted with patients [33]. On the other hand, we checked that there was no significant difference in tumour size between the various groups, since this factor could also have had an influence on our results. Indeed, we showed previously with this tumour model that the tumour uptake of the radiolabelled lipiodol decreases significantly with increasing tumour weight [23]. This study clearly shows that treatment by 131I-lipiodol is the only really effective treatment, particularly in comparison with 188Re-SSS lipiodol. However, the results obtained with 188Re-SSS lipiodol cannot necessarily be extrapolated to others where 188Re labelling (e.g. 188ReHDD and 188Re-NDEDC lipiodol) has been used; preclinical studies comparing the different compounds should be necessary to select the best one. Furthermore, the results we obtained in rats with relatively small HCCs lead us to consider whether it would be appropriate to substitute 131I-labelled lipiodol by 188Re-labelled lipiodol in humans. Indeed, a treatment by radiolabelled lipiodol is most often proposed in patients presenting with multifocal lesions in association with large and small tumours for which surgery cannot be considered. In the light of the present results, clinical studies in humans, comparing treatments by 188Re- or 131 I-labelled lipiodol, are necessary before considering whether the former could be an alternative to the latter.

Conclusion This study shows that, in HCC-bearing rats, treatment by 131 I-lipiodol is more effective compared with the 188ReSSS lipiodol/131I-lipiodol mixture or 188Re-SSS lipiodol alone, because it is the only treatment that makes it possible to obtain a prolonged improvement of survival. This study also shows that treatment by 188Re-SSS lipiodol alone is ineffective with the tumour model used here, which is built up of a single, small tumour. Taking into account the limitations related to the use of this tumour model, these results cannot necessarily be extrapolated to patients, who often have larger tumours. Thus, it would appear necessary to carry out a study in humans to compare these three treatments. In particular, a comparison should be made between the effectiveness of 131I-labelled and 188Re-labelled lipiodol with a view to replacing the former treatment by the latter. However, this study clearly demonstrated that, for small tumours, 131I-labelled lipiodol is more effective than

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Effect of

188

Re-labelled lipiodol. So in such clinical situations, and particularly in an adjuvant setting, 131I-labelled lipiodol should be a better option than 188Re-labelled lipiodol.

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Yoo HS, Lee JT, Kim KW, Kim BS, Choi HJ, Lee KS, et al. Nodular hepatocellular carcinoma. Treatment with subsegmental intraarterial injection of iodine 131-labeled iodized oil. Cancer 1991; 68:1878–1884. Maini CL, Scelsa MG, Fiumara C, Tofany A, Sciuto R, Tipaldi L, et al. Superselective intra-arterial radiometabolic therapy with I-131 Lipiodol in hepatocellular carcinoma. Clin Nucl Med 1996; 21:221–226. Wang SJ, Lin WY, Chen MN, Hsieh BT, Shen LH, Tsai ZT, et al. Biodistribution of rhenium-188 lipiodol infused via the hepatic artery of rats with hepatic tumours. Eur J Nucl Med 1996; 23:13–17. Tsai C, Kusumoto Y, Harada R, Shima M, Nakata K, Kono K, et al. Effect of intra-hepatic arterial infusion of 131I-labelled lipiodol on hepatocellular carcinoma of rat. Ann Acad Med 1986; 15:521–524. Madsen MT, Park CH, Thakur ML. Dosimetry of iodine 131 ethiodol in the treatment of hepatoma. J Nucl Med 1988; 29:1038–1044. Brans B, Bacher K, Vandevyver V, Vanlangenhove P, Smeets P, Thierens H, et al. Intra-arterial radionuclide therapy for liver tumours: effect of selectivity of catheterization and 131I-lipiodol delivery on tumour uptake and response. Nucl Med Commun 2003; 24:391–396. Lin WY, Tsai SC, Hsieh JF, Wang SJ. Effects of 90Y-microspheres on liver tumours: comparison of intra-tumoural injection method and intra-arterial injection method. J Nucl Med 2000; 41:1892–1897. Garin E, Denizot B, Roux J, Noiret N, Lepareur N, Moreau M, et al. Description and technical pitfalls of a hepatoma model and of intra-arterial injection of radiolabelled lipiodol in the rat. Laboratory Animals 2005; 39:314–320. Brans B, Van Laere K, Gemmel F, Defreyne L, Vanlangenhove P, Troisi R, et al. Combined iodine-131 lipiodol therapy with low-dose cisplatin as radiosensitiser: preliminary study results in hepatocellular carcinoma. Eur J Nucl Med 2002; 29:928–932. Raoul JL, Boucher E, Rolland Y, Garin E. Association of CDDP and intra-arterial injection of 131I-lipiodol (lip 131I) in the treatment of hepatocellular carcinoma (HCC): results of a phase II trial. J Clin Oncol 2002; 21:563. De Baere T, Zhang X, Aubert B, Harry G, Lagrange C, Ropers J, et al. Quantification of tumour uptake of iodized oils and emulsions of iodized oils: experimental study. Radiology 1996; 201:731–735. Garin E, Denizot B, Roux J, Noiret N, Lepareur N, Moreau M, et al. Effect of stabilized lipiodol emulsion on experimentally induced hepatocellular carcinoma in rats. J Vasc Intervent Radiol 2005; 16:841–848. Burton MA, Gray BN, Self GW, Heggie JC, Towensend PS. Manipulation of experimental rat and rabbit liver tumour blood flow with angiotensin II. Cancer Res 1985; 45:5390–5393. Burton MA, Gray BN, Coletti A. Effect of angiotensin II on blood flow in the transplanted sheep squamous cell carcinoma. Eur J Cancer Clin Oncol 1988; 24:1373–1376. Amin AE, Wheldon TE, O’Donoghue JA, Barret A. Radiobiological modeling of combined targeted 131I therapy and total body irradiation for treatment of disseminated tumours of differing radiosensitivity. Int J Radiat Oncol Biol Phys 1993; 27:323–330. Towu E, Boxer G, Begent R, Zweit J, Spitz L, Hobbs K, Winslet M. In-vitro uptake of radioactive lipiodol I-131 and I-125 by hepatoblastoma: implications for targeted radiotherapy. Pediatr Surg Int 2001; 17:609–613. De Jeong Jeong M, Bernard HF, Breeman WAP, Bijster M, van den Berg A, van Gameren A, Krenning EP. Combined use of 90Y- and 177Lu-labelled somatostatin analogues for radionucleide therapy. Eur J Nucl Med 2002; 29(suppl 1):S119. Kraczynski J, Hansson G, Remotti H, Wallerstedt S. Spontaneous regression of HCC. Histopathology 1998; 32:147–150.

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Original article

Preparation and evaluation of 99mTc-EDDA/HYNIC-[Lys3]bombesin for imaging gastrin-releasing peptide receptor-positive tumours Guillermina Ferro-Floresa, Consuelo Arteaga de Murphyb, Jeanette Rodrı´guez-Corte´sb,c, Martha Pedraza-Lo´pezb and Marı´a Teresa Ramı´rez-Iglesiasb Background Bombesin is a peptide that was initially isolated from frog skin and which belongs to a large group of neuropeptides with many biological functions. The human equivalent is gastrin-releasing peptide (GRP), whose receptors are over-expressed in a variety of malignant tumours.

studies demonstrated a high stability in serum and cysteine solutions, specific cell receptor binding and rapid internalization. Biodistribution data showed a rapid blood clearance, with predominantly renal excretion and specific binding towards GRP receptor-positive tissues such as pancreas and PC-3 tumours.

Aim To prepare a HYNIC-[Lys3]-bombesin analogue that could be easily labelled with 99mTc from lyophilized kit formulations and to evaluate its potential as an imaging agent for GRP receptor-positive tumours.

Conclusion 99mTc-EDDA/HYNIC-[Lys3]-bombesin obtained from lyophilized kit formulations has promising characteristics for the diagnosis of malignant tumours that over-express the GRP receptor. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:371–376 

Methods HYNIC was conjugated to the e-amino group of Lys3 residue at the N-terminal region of bombesin via succinimidyl-N-Boc-HYNIC at pH 9.0. 99mTc labelling was performed by addition of sodium pertechnetate solution and 0.2 M phosphate buffer pH 7.0 to a lyophilized formulation. Stability studies were carried out by reversed phase HPLC and ITLC-SG analyses in serum and cysteine solutions. In-vitro internalization was tested using human prostate cancer PC-3 cells with blocked and non-blocked receptors. Biodistribution and tumour uptake were determined in PC-3 tumour-bearing nude mice.

Nuclear Medicine Communications 2006, 27:371–376 Keywords: bombesin, imaging

99m

Tc labelled HYNIC-bombesin, GRP receptor

a Instituto Nacional de Investigaciones Nucleares, Ocoyoacac, Mexico, bInstituto Nacional de Ciencias Me´dicas y Nutricio´n, Salvador Zubira´n, Mexico and c Universidad Auto´noma del Estado de Mexico.

Correspondence to Dr Guillermina Ferro-Flores, Departamento de Materiales Radiactivos, Instituto Nacional de Investigaciones Nucleares, Km. 36.5 Carretera Me´xico-Toluca, Ocoyoacac, Estado de Me´xico, C.P. 52045, Mexico. Tel: + 0052 55 532 97200; fax: + 0052 55 532 97306; e-mail: [email protected]

Results 99mTc-EDDA/HYNIC-[Lys3]-bombesin was obtained with radiochemical purities > 93% and high specific activity (B0.1 GBq
nmol – 1). Results of in-vitro

This study was supported by the CONACyT-Mexico (SALUD-2004-01-003) and the International Atomic Energy Agency (IAEA).

Introduction

use in nuclear medicine for the detection of malignant tumours and for staging breast and prostate cancers and their lymph nodes [1–8].

The small peptide bombesin (14 amino acids), which was isolated from frog skin, belongs to a large group of neuropeptides with many biological functions. The human equivalent is gastrin-releasing peptide (GRP) whose receptors (GRP-r) are over-expressed in the tumour cell membrane. The strong, specific, bombesin–GRP-r binding is the basis for labelling bombesin with radionuclides [1–5]. 99mTc-diaminedithiol-[Lys3]-bombesin (99mTcDADT-[Lys3]-bombesin), [DTPA1,Lys3(99mTc-DADT),99m Tc(I)-2-picolylamine-N,N-diacetic Tyr4]-bombesin, acid-5-aminovaleric acid-bombesin (99mTc(I)-PADA-AVAbombesin), 99mTc-Cys-6-amino-n-hexanoic acid-bombesin and 64Cu-DOTA-[Lys3]-bombesin have been prepared for

Received 13 December 2005 Accepted 6 January 2006

The availability of simple, efficient and reproducible radiolabelling procedures is essential in the development of radiopharmaceuticals for routine clinical use. Although the technology of 99mTc-HYNIC involves an indirect labelling procedure where the hydrazinonicotinamide (HYNIC) bifunctional chelator is conjugated to the peptide and ethylenediamine-N,N0 -diacetic acid (EDDA) used as co-ligand to complete the technetium coordination sphere, it can be obtained easily from instant freezedried kit formulations [9,10].

c 2006 Lippincott Williams & Wilkins 0143-3636 

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372 Nuclear Medicine Communications 2006, Vol 27 No 4

The aim of this study was to prepare the HYNIC-[Lys3]bombesin analogue that could be labelled with 99mTc from lyophilized kit formulations and to evaluate its in-vitro and in-vivo binding to human prostate cancer PC-3 cells.

pack C-18 column (10 mm) at a flow rate of 1 ml
min – 1 using 0.1% TFA/water (solvent A) and 0.1% TFA/CH3CN (solvent B). The gradient started at 100% solvent A for 3 min, changed to 50% solvent A over 10 min, held constant for 10 min, changed to 30% solvent A over 3 min and finally back to 100% solvent A over 4 min. In this system the retention times for HYNIC, [Lys3]-bombesin and HYNIC-[Lys3]-bombesin were 8, 11 and 11.4 min, respectively. According to the conjugation yield (average 60%), the collected sample containing HYNIC-[Lys3]bombesin was dried under vacuum and reconstituted with 0.6 ml of 10% ethanol (1 mg
ml – 1).

Materials and methods Preparation of HYNIC-[Lys3]-bombesin

[Lys3]-bombesin and succinimidyl-N-Boc-HYNIC were supplied by Bachem-USA and ABX-Germany, respectively. The peptide was prepared at a concentration of 1.0 mg/0.5 ml in 0.1 M HEPES (N-(2-hydroxyethyl)piperazine-N0 -(2-ethanesulfonic acid)) buffer, pH 9.0, to which 60 ml of a fresh 15 mg
ml – 1 solution of succinimidyl-NBoc-HYNIC in dry dimethylformamide (DMF) was added with agitation. The final HYNIC-to-peptide molar ratio was 5 : 1. The mixture was reacted at room temperature for 60 min and purified by solid phase extraction. Boc-HYNIC-(Lys3)bombesin was loaded on the preconditioned Sep-Pak C-18 cartridge followed by 5 ml of 20% CH3CN to elute Boc-HYNIC. The conjugate peptide was eluted with 1 ml of 50% CH3CN, deprotected by addition of 300 ml of trifluoroacetic acid (TFA) and dried under vacuum for 30 min (Fig. 1). The sample was reconstituted with 0.8 ml of 20% CH3CN and purified by reverse phase HPLC (Waters, 1 ml loop) with a UV–photodiode array in-line detector and a m-Bonda-

HYNIC-[Lys3]-bombesin lyophilized formulations

The procedure was carried out under aseptic conditions in a facility certified for good medical practice: 0.4 mg of HYNIC-[Lys3]-bombesin in 0.4 ml of 10% ethanol was added to a solution of EDDA–tricine–mannitol which was previously prepared by mixing 0.4 g of EDDA, 0.8 g of tricine (N-tris[hydroxymethyl]methylglycine) and 2 g of mannitol in 39 ml of water for injection under gentle heating and stirring. Finally, 0.8 ml of a freshly prepared anhydrous stannous chloride solution (1 mg
ml – 1 in 0.012 M HCl) was added under a nitrogen atmosphere. The mixture was sterilized by membrane filtration (Millipore, 0.22 mm) and 1.0 ml aliquots were dispensed into 40 pre-sterilized serum vials and lyophilized for 24 h.

Fig. 1 OH O O H

O NH

N

Tc

N H N H HO N OH NH O O N

NH2 NH N

HN O

NH2

HN O

O

O O

H N OO

HN

H HN N

N H

O

NH2

O

O

1. Succinimidil-N-Boc-HYNIC, pH = 9

O

HN NH O N H

H N

O

NH2

O NH

N H

H N O

O

HN

O

N

O

H N

NH2

H2N

99mTcO −, 4

N H

H N O

N H

NH

N H

O

O NH H N O

N H

O

HN

O N H

H N O

N

O N H

N H

HN O

NH2

NH2

HN O

O

NH

O HN O

NH

NH O

O S

H2N

Overall reaction scheme for the preparation of

O

N

O S

O

HN

O O

92˚C, 15 min HN

O

O

H N

O

EDDA/tricine/SnCl2

O NH

HN

O

NH2

O HN O

O

O

H N

H HN N

N H O

NH HN

O N H

H2N

O

HN

O

2. TFA O

O

H N

O

O H2N

O

99m

H2N

S

H2N

Tc-EDDA/HYNIC-[Lys3]-bombesin.

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Preparation of

Additionally, 100 ml of a 0.2 M buffer phosphate pH 7 solution was prepared under aseptic conditions, sterilized by membrane filtration (Millipore, 0.22 mm), 5.0 ml aliquots were dispensed into pre-sterilized serum vials and stored at 41C. All vials were tested as sterile and apyrogenic preparations by conventional pharmaceutical procedures. Labelling HYNIC-[Lys3]-bombesin with

99m

Tc

Radiolabelling was carried out by adding 1 ml of 0.2 M phosphate buffer pH 7.0 to the freeze-dried kit formulation then, immediately, 740–1110 MBq (1 ml) of [99mTc]pertechnetate followed by incubation at 921C for 15 min. Evaluation of radiochemical purity

Analyses of radiochemical purity were performed by instant thin-layer chromatography on silica gel (ITLCSG, Gelman Sciences), solid phase extraction (Sep-Pak C-18 cartridges) and reverse phase high-performance liquid chromatography (HPLC). ITLC-SG analysis was accomplished using three different mobile phases: 2-butanone to determine the amount of free 99mTcO4 (RF = 1), 0.1 M sodium citrate pH 5 to determine the 99mTc co-ligand and 99mTcO4– (RF = 1), and methanol/ 1 M ammonium acetate (1 : 1 v/v) for 99m Tc-colloid (RF = 0). RF values of the radiolabelled peptide in each system were 0.0, 0.0 and 0.7–1.0, respectively. The Sep-Pak cartridges were preconditioned with 5 ml of ethanol followed by 5 ml of 1 mM HCl and 5 ml of air. An aliquot of 0.1 ml of the labelled peptide was loaded on the preconditioned Sep-Pak cartridge followed by 5 ml of 1 mM HCl to elute free 99mTcO4– . The radiolabelled peptide was eluted with 3 ml of ethanol/saline (1 : 1) and the hydrolyzed-reduced 99mTc or 99mTc-colloid remained in the cartridge. HPLC analyses were carried out with a Waters instrument running Millenium software with both radioactivity and UV–photodiode array in-line detectors. A YMC ODSAQ S5 column (5 mm, 4.6  250 mm) at a flow rate of 1 ml
min – 1 using the gradient system described above. In this system retention times for free 99mTcO4– , 99mTc co-ligand and 99mTc-EDDA/HYNIC-[Lys3]-bombesin were 3 min, 3.5–4.5 min and 12 min, respectively. Serum stability

A volume of 50 ml of the labelled peptide solution (0.5 mg/50 ml). was incubated at 371C with 1 ml of fresh human serum. Radiochemical stability was determined taking samples of 10 ml at different times from 5 min to 24 h for analysis by radio-HPLC. Recovery of radioactivity was routinely determined.

99m

Tc-EDDA/HYNIC-[Lys3]-bombesin Ferro-Flores et al. 373

Cysteine challenge 99m

Tc-EDDA/HYNIC-[Lys3]-bombesin was tested for instability toward cysteine. A fresh cysteine solution was prepared (10 mg
ml – 1 in 0.1 M PBS, pH 7.0) and diluted to different concentrations. Then, 12 ml of each cysteine solution was mixed with 90 ml of the 2.2 mM of the labelled peptide solutions. The molar ratios of cysteine to peptide were between 5 : 1 and 500 : 1. Each test tube was incubated at 371C and radiochemical purity determined 1 h later by ITLC.

Protein binding

Ultrafiltration (Ultrafree-PFL 30,000 NMWL, Millipore Co.). was used to estimate the percent of labelled peptides bound to serum proteins. A volume of 10 ml of the labelled peptide solution (0.5 mg/50 ml) was incubated at 371C with 1 ml of fresh human serum. Samples at different times up to 24 h were ultrafiltered and the percent of the activity in the filter and the eluant determined. Cell lines

The human prostate cancer cell PC-3 line was originally obtained from ATCC (USA). The cells were routinely grown at 371C, in a 5% CO2 atmosphere and 100% humidity in RPMI medium supplemented with 10% newborn calf serum and antibiotics (100 mg
ml – 1 streptomycin). Internalization assay and non-specific binding

PC-3 cells (1  106) supplied with fresh medium were incubated in 6-well plates with about 200 000 cpm of 99m Tc-EDDA/HYNIC-[Lys3]-bombesin (0.2 nmol total peptide) for 2 h at 41C. After the pre-incubation cells were washed 3 times with ice-cold culture medium. Warmed culture medium was added to the plates which were incubated in triplicate at 371C for 2, 4, 6 and 24 h in order to allow internalization. Cell-surface bound radioligand was removed by two steps of 5 min acid wash (50 mM glycine HCl/100 mM NaCl, pH 2.8) at room temperature. Cells were solubilized by incubation with 1 N NaOH at 371C to determined internalized radioligand. Results were expressed as percentage of total activity internalized. The non-specific binding was determined in parallel but in the presence of 10 mM Tyr4-bombesin (Bachem-USA) (blocked receptor cells). Animal model

Tumour uptake and biodistribution studies in mice were carried out according to the rules and regulations of the Official Mexican Norm 062-ZOO-1999. Athymic male mice (20–22 g) were kept in sterile cages with beds of sterile wood shavings, constant temperature, humidity, noise and 12 : 12 light periods. Water and feed (standard PMI 5001 feed) were given ad libitum.

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374 Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 2

(a) 80.00

60.00 50.00

Biodistribution

Imaging

The nude mice with the implanted tumours were scanned with a gamma camera with a pinhole collimator 2 h after the radiopharmaceutical was injected in the tail vein.

40.00 30.00 20.00 10.00 0.00 0.00

5.00

10.00

15.00 20.00 Minutes

25.00

30.00

12.146

(b) 2.00 1.50 1.00 0.50 0.00 − 0.50

10.532

mV

Athymic mice with induced tumours were used for biodistribution studies. 99mTc-EDDA/HYNIC-[Lys3]bombesin, 1.11 MBq (30 mCi) in 0.04 ml was injected in a tail vein. The mice (n = 4) were killed 2 h postinjection. Whole heart, spleen, pancreas and kidneys and samples of tumour, lung, liver, blood, intestines and muscle were rinsed in saline, blotted with paper and placed into pre-weighed plastic test tubes. The activity was determined in a well-type scintillation detector (Canberra) along with six 0.5 ml aliquots of the diluted standard representing 100% of the injected activity. Mean activities were used to obtain the percentage of injected activity per gram of tissue (% IA/g). For non-specific uptake into the tumour or receptor-positive organs a biodistribution study was done in parallel but with animals injected with 50 mg of Tyr4-bombesin 30 min previously (blocked receptor group, n = 4).

Radioactivity

70.00

mV

Prostate tumours were induced by subcutaneous injection of PC-3 cells (1  106) resuspended in 0.2 ml of phosphate-buffered saline, into the backs of 6- to 7week-old nude mice. The sites of injection were observed at regular intervals for the appearance of tumour formation and progression.

4.035

Tumour induction in athymic mice

− 1.00 5.00

10.00

15.00 20.00 Minutes

25.00

30.00

Reverse phase HPLC radiochromatograms of 99mTc-EDDA/HYNIC[Lys3]-bombesin (a) after labelling, and (b) 24 h after incubation in fresh human serum.

Pathology studies

Tumours taken from the mice were formalin-fixed and paraffin-embedded. In order to determine their microscopic characteristics the tumour sections were stained with haematoxylin&eosin and viewed under a light microscope (Zeiss). Statistics

Specific uptake was determined by comparison of blocked and unblocked animals or cells using the Student t-test.

Results The results obtained by TLC, SepPak and HPLC analyses showed a mean radiochemical purity for 99mTcEDDA/HYNIC-[Lys3]-bombesin of 95 ± 2% (n > 30) without post-labelling purification (Fig. 2(a)). The average specific activity was B0.1 GBq
nmol – 1. After 24 h in human serum the radiochemical purity remained > 90% (Fig. 2(b)). Protein binding was 29 ± 1.4% and 34 ± 2.1% at 1 and 24 h, respectively. After incubation with a 1 : 500 cysteine:peptide molar ratio, both ITLC and HPLC analyses showed that the

radioactivity dissociated from 99mTc-EDDA/HYNIC[Lys3]-bombesin is minimal and less than 9% (Fig. 3). Results of the in-vitro assays showed a rapid internalization (8% at 2 h and 11.5% at 4 h) and a specific receptor binding of 99mTc-EDDA/HYNIC-[Lys3]-bombesin to PC-3 cells since there were significant differences in the percentage of uptake between blocked and unblocked cells at different times (P < 0.05) (Fig. 4). The tissue distribution of radioactivity 2 h after 99mTcEDDA/HYNIC-[Lys3]-bombesin (unblocked and blocked animals) exhibited a rapid clearance from the blood and most tissues with predominantly renal excretion (Table 1). The highest non-specific uptake was found in kidneys. A significant uptake of radioactivity was observed in the pancreas which expresses GRP receptors. The tumour also exhibited specific uptake of radioactivity. The specificity was confirmed by the receptor blocking study in which the previous injection of Tyr4bombesin diminished the activity in pancreas and in PC-3 tumours. Reduction uptake percentages were 70% in the

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Preparation of

Tc-EDDA/HYNIC-[Lys3]-bombesin Ferro-Flores et al. 375

Table 1 Biodistribution in mice with induced PC-3 tumours 2 h after administration of 99mTc-EDDA/HYNIC-[Lys3]-bombesin

Fig. 3

10

Tissue

9

Blood Heart Lung Liver Spleen Pancreas Kidney Intestine Muscle Tumour

8 7 % Dissociation

99m

6 5 4

Unblocked

Blocked

0.08 ± 0.03 0.05 ± 0.02 0.10 ± 0.04 0.17 ± 0.04 0.08 ± 0.04 1.29 ± 0.31* 4.70 ± 1.20 0.17 ± 0.10 0.05 ± 0.03 0.30 ± 0.11*

0.11 ± 0.06 0.07 ± 0.03 0.11 ± 0.05 0.16 ± 0.04 0.10 ± 0.05 0.34 ± 0.16* 5.20 ± 1.8 0.26 ± 0.11 0.04 ± 0.02 0.09 ± 0.03*

* Statistical significant difference (P < 0.05) between blocked and unblocked animals. Results are expressed as a percentage of the injected activity per gram of tissue (%IA/g)(mean ± SD, n = 4). Blocked animals received an additional gastrin-releasing peptide receptor blocking dose of Tyr4-bombesin in order to determine the non-specific uptake of radioactivity.

3 2 1 0 0

50 100 150 200 250 300 350 400 450 500 Cysteine:peptide (molar ratio)

Dissociation of 99mTc from 99mTc-EDDA/HYNIC-[Lys3]-bombesin following incubation with cysteine at different molar ratios (mean ± SD, n = 3).

Fig. 4

26 Unblocked Blocked

Internalization (% of total activity)

24 22

internal viscera, highlighted the 99mTc-EDDA/HYNICbombesin uptake in tumour PC-3 cells (Fig. 5). The pathology studies confirmed that the cell line showed highly undifferentiated cells corresponding to human prostate cancer. The tumours, 0.2–2.3 g (n = 24) were white and of elastic consistency. Microscopically, the cell characteristics corresponded to human prostate carcinoma metastatic to soft tissues.

Discussion

Time dependent internalization of 99mTc-EDDA/HYNIC-[Lys3]bombesin in unblocked and blocked PC-3 cells expressed as percent of total activity (mean ± SD, n = 3). Blocked cells were incubated with an additional GRP receptor blocking dose of Tyr4-bombesin in order to determine the non-specific binding of radioactivity.

As Baidoo et al. reported [1] modification of Lys3 in bombesin had no effect on functional activity since the active portion of the peptide is the C-terminal eight amino acids. The peptide could be modified without side-chain protection, since the E-amino group of Lys is the only moiety susceptible to modification under the reaction conditions (pH = 9). However, molecular mechanics and quantum-mechanical calculations have shown that for [Lys3]-bombesin, the only site available to introduce HYNIC as the Tc chelator was Lys3 even if reaction conditions are carried out at pH 7. In fact, the conjugation via HATU (O-(7-azabenzotriazolyl)-1,1,3, 3-tetramethyluronium hexafluorophosphate) produces high HYNIC-Lys3-bombesin conjugate yields, obtaining a very high thermodynamic stability without interference in the stereospecificity of the C-terminal eight residues which are believed to contain the domain responsible for receptor recognition [11].

tumour (0.30% IA/g vs. 0.09% IA/g, P < 0.05) and 73% in the pancreas (1.29% IA/g vs. 0.34% IA/g, P < 0.05). The uptake in non-targeted tissues was not significantly reduced by the blocking dose. Tumour-to-blood, tumour-to-muscle and pancreas-to-blood ratios were 3.75, 7.5 and 16, respectively. In-vivo images showed a clear tumour uptake and a dissection process to eliminate

Conjugation of HYNIC to bombesin for the preparation of 99m Tc-EDDA/HYNIC-[Lys3]-bombesin modifies the lipophilic and pharmacokinetic properties of bombesin, producing a radiopharmaceutical with low hepatobiliary clearance and predominantly renal excretion. This is an important result because most of the 99mTc-labelled bombesin analogues have a tendency to accumulate in the liver and intestine as a result of their high lipophilicity and hepatobiliary clearance [1–5]. The high

20 18 16 14 12 10 8 6 4 2 2

4

6

24

Time (h)

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376 Nuclear Medicine Communications 2006, Vol 27 No 4

bioconjugate toward GRP receptors. The uptake into the GRP receptor-positive PC-3 was significantly lower as in the pancreas, but the tumour uptake was highly specific as it could be reduced in the ‘blocked’ experiment.

Fig. 5

(a)

Kidneys and pancreas Tumour

Bladder

As Scopinaro et al. [6,7] reported, technetium-labelled bombesin (99mTc-Leu13-bombesin1) is able to detect breast, lung and prostate cancer and invasion of pelvic nodes, even when this bombesin derivative shown hepatobiliary excretion. After preliminary human studies, the authors have concluded that the use of 99mTcbombesin may play an important role in diagnosing and staging prostate cancer [7]. Then, an important advantage of the 99mTc-EDDA/HYNIC-[Lys3]-bombesin reported in this work is the feasibility to be obtained from instant freeze-dried kit formulations for possible routine clinical use in breast and prostate cancer detection with minimal accumulation in the abdominal area. In conclusion, 99mTc-EDDA/HYNIC-[Lys3]-bombesin obtained from lyophilized kit formulations has promising characteristics for diagnosis of tumours that over-express the GRP receptor.

(b)

References 1

Tumour

Bladder

Uptake of 99mTc-EDDA/HYNIC-[Lys3]-bombesin in tumour PC-3 cells in athymic mouse. (a) Whole mouse and (b) mouse with dissection of internal viscera to highlight tumour uptake.

accumulation of radioactivity might interfere during the detection of bombesin/GRP receptor-positive cancers and their metastases in the abdominal areas. The uptake of radioactivity in the pancreas demonstrated the ability of the radiopharmaceutical to target in-vivo GRP receptor-bearing cells. Furthermore, the receptor blocking studies confirmed the specificity of the

Baidoo KE, Lin KS, Zhan Y, Finley P, Scheffel U, Wagner HN. Design, synthesis, and initial evaluation of high-affinity technetium bombesin analogues. Bioconjugate Chem 1998; 9:218–225. 2 La Bella R, Garcia-Garayoa E, Langer M, Bla¨uenstein P, Beck-Sickinger AG, Schubiger PA. In vitro and in vivo evaluation of a 99mTc(I)-labeled bombesin analogue for imaging of gastrin releasing peptide receptor-positive tumors. Nucl Med Biol 2002; 29:553–560. 3 Varvarigou AD, Scopinaro F, Leondiadis L, Corleto V, Schillaci O, De Vicentis G, et al. Synthesis, chemical, radiochemical and radiobiological evaluation of a new 99m Tc-labelled bombesin-like peptide. Cancer Biother Radiopharm 2002; 17:317–326. 4 Smith JC, Gali H, Sieckman GL, Hayes DL, Owen NK, Mazuru DG, et al. Radiochemical investigations of 177Lu-DOTA-8-Aoc-BBN[7–14]NH2: an in vitro/in vivo assessment of the targeting ability of this new radiopharmaceutical for PC-3 human prostate cancer cells. Nucl Med Biol 2003; 29:101–109. 5 Chen X, Park R, Hou Y, Tohme M, Shahinian AH, Bading JR, et al. microPET and autoradiographic imaging of GRP receptor expression with 64Cu-DOTA[Lys3]Bombesin in human prostate adenocarcinoma xenografts. J Nucl Med 2004; 45:1390–1397. 6 Scopinaro F, Varvarigou AD, Ussof W, De Vicentis G, Sourlingas TG, Evangelatos GP, et al. Technetium labelled bombesin-like peptide: preliminary report on breast cancer uptake in patients. Cancer Biother Radiopharm 2002; 17:327–335. 7 Scopinaro F, De Vicentis G, Varvarigou AD, Laurenti C, Iori F, Remediani S, et al. 99mTc-bombesin detects prostate cancer and invasion of pelvic lymph nodes. Eur J Nucl Med Mol Imaging 2003; 30:1378–1382. 8 Lin KS, Luu A, Baidoo KE, Hashemzadeh-Gargari H, Chen MK, Brenneman K, et al. A new high affinity technetium-99m-bombesin analogue with low abdominal accumulation. Bioconjugate Chem 2005; 16:43–50. 9 Von-Guggenberg E, Mikolajczak R, Janota B, Riccabona G, Decristoforo C. Radiopharmaceutical development of a freeze-dried kit formulation for the preparation of [99mTc-EDDA-HYNIC-D-Phe1, Tyr3]-octreotide, a somatostatin analog for tumor diagnosis. J Pharm Sci 2004; 93:2497–2506. 10 Gonza´lez-Va´zquez A, Ferro-Flores G, Arteaga de Murphy C, Gutie´rrez-Garcı´a Z. Biokinetics and dosimetry in patients of 99mTc-EDDA/ HYNIC-Tyr3-Octreotide obtained from lyophilized kits. Appl Rad Isot 2005; in press. 11 Ferro-Flores G, Arteaga de Murphy C, Ramı´rez F de M, Pedraza-Lo´pez M, Rodrı´guez-Corte´s J, Mele´ndez-Alafort L. Preparation and evaluation of third generation technetium radiopharmaceuticals. International Symposium on Trends in Radiopharmaceuticals. IAEA publication IAEA-CN-130. Vienna: International Atomic Energy Agency; 2005, pp. 10–11.

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Original article

Hu¨rthle cell carcinoma: a clinicopathological study of thirteen cases ¨ zlem Ku¨c¸u¨k, Hu¨lya Kulak, Emel Tokmak, Pınar Tarı, Erkan ˙Ibis¸ N. O and Gu¨lseren Aras Background Hu¨rthle cell carcinoma (HCC) of the thyroid is a variant of follicular cancer which has been considered by many as a more aggressive disease than the usual well-differentiated carcinoma of the thyroid. Aim To investigate the clinico-pathologic characteristics, treatment and outcome of Hu¨rthle cell carcinoma. Material and methods During a 7-year period, 13 patients (seven male, six female; mean age at diagnosis 48.4 ± 13.2 years) with HCC were treated and monitored at the Ankara University. The measured diameter of the tumours varied from 1 to 6 cm in diameter with pathological examination. Three of the HCC had extrathyroid invasion, five had intrathyroid invasion, and five were encapsulated. One of the patients had a history of low-dose external radiation to the head and neck in childhood. Treatment consisted of a total thyroidectomy in 12 patients, and a near total thyroidectomy in one patient. At surgery, lymph node metastases were present in three patients and lymph node dissection were performed in these patients. Distant metastases were detected in only one patient (lung metastasis).

being followed up. After a median follow-up period of 85 months, there was no recorded mortality due to the disease and 12/13 of the patients were categorized as disease free (criteria for ablation were a negative 131I whole-body scan and very low serum thyroglobulin levels). Conclusion We did not find higher incidences of local recurrences, distant metastases or mortality rates compared to well differentiated thyroid carcinomas. HCC of the thyroid and well differentiated thyroid carcinomas have similar biological behaviour. Their treatment should be similar, including total or near-total thyroidectomy plus modified cervical node dissection when there is lymph node involvement. Radioactive iodine therapy and suppressive laevothyroxin therapy c 2006 should follow. Nucl Med Commun 27:377–379 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:377–379 Keywords: Hurthle cell carcinoma, radioiodine therapy, response distant metastases Faculty of Medicine, Department of Nuclear Medicine, Ankara University, Turkey.

Results All patients had radioiodine ablation therapy for residual thyroid tissue. Twelve of the 13 patients were ablated with a single dose of 131I (3.7–5.5 GBq). A second dose of radioiodine therapy was required in only one patient who had lung metastases and this patient is still

¨ zlem Ku¨c¸u¨k, Ankara University, Faculty of Medicine, Correspondence to Dr N. O Department of Nuclear Medicine, Ankara, Turkey. Tel: + 90 312 595 6445; fax: + 90 312 362 0897; e-mail: okucuk@ medicine.ankara.edu.tr

Introduction

131

Hu ¨rthle cell carcinoma (HCC) is reported to represent only 3% of differentiated thyroid cancers [1]. According to WHO histological classification, HCCs of the thyroid are generally assigned to the group of folliculary carcinomas. The frequency is reported only in about 3–6% with large patients groups [2–6]. The tumour is predominantly seen in women (the male/female ratio is 2/7) and during the fourth and seventh decades. The 5-year survival rate is approximately 85% and the relapse rate is 50% [7]. When there are distant metastases the 5-year survival rate is 60% [8–10]. HCC is usually multifocal and the first choice of therapy is total or near-total thyroidectomy [10,11]. Although the

Received 7 November 2005 Accepted 4 January 2006

I uptake is less than 10%, 131I therapy is commonly performed to ablate thyroid remnants [12–15]. This present study is a retrospective analysis to investigate the clinicopathologic characteristics, treatment and outcome of HCC.

Patients, material and methods During a 7-year period (1997 through 2004), 510 patients with differentiated thyroid carcinoma were treated and monitored at Ankara University. Thirteen patients (seven female and six male; mean age at diagnosis 48.4 ± 13.2 years, range 29–84 years) with Hu ¨rthle cell tumours were treated.

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378 Nuclear Medicine Communications 2006, Vol 27 No 4

A non-Hu ¨rthle cell thyroid carcinoma was found within the thyroid gland in four (30%) of the patients: one was papillary and three were follicular carcinomas. The tumours measured 1–6 cm in diameter. Three of the Hu ¨rthle cell carcinomas had extrathyroid invasion, five had intrathyroid invasion, and five were encapsulated (i.e., they had intracapsular invasion only). One of the patients had a history of low-dose external radiation to the head and neck in childhood. Laboratory tests

Serum thyroglobulin (Tg) and anti-Tg concentrations were determined by chemoluminescence using a commercially available kit (IMMULITE 2000 Tg kit/L2KTY2; normal range, 1.6–59.9 ng
ml – 1, IMMULITE 2000 AntiTg Ab kit/L2KTG; normal range, 0–40 IU
ml – 1). Surgical treatment

Treatment consisted of a total thyroidectomy in 12 patients, and a near total thyroidectomy in one patient. At surgery, lymph node metastases were present in three patients and lymph node dissection were performed in these patients. Distant metastasis was detected at diagnosis only in one patient (lung metastases). Radioiodine ablation

The patients were informed about the different therapeutic strategies and the need for more than one course of radioiodine treatment to eliminate thyroid remnants. All of the patients in our retrospective study had radioiodine ablation therapy for residual thyroid tissue. After primary treatment, all patients received L-thyroxine in suppressive doses. Scintigraphic techniques

Patients were followed up for 3, 6 and 12 months after treatment, and then on a yearly basis. A 131I whole-body scan was performed 6 days after the initial radioiodine treatment. T4 therapy was withdrawn 30 days before scan. A subsequent whole-body scan was performed 6 months after radioiodine treatment and repeated therapy was given if the scan showed positive uptake or high serum Tg levels. In each patient successful ablation was documented by 131I whole-body scintigraphy 6 months after the last therapeutic dose. An absence of pathological uptake of 131I in the neck and a Tg level of less than 1 ng
ml – 1 were the criteria for ablation.

Results All of the patients had radioiodine ablation therapy for residual thyroid tissue. Twelve of the patients were ablated with a single dose of 131I (3.7–5.5 GBq). In all patients, serum thyroglobulin concentrations were within the normal range (1.6–59.9 ng
ml – 1). A second dose of radioiodine was required only in one patient, because of lung metastases, and this patient is still being followed

up. After a median follow-up period of 85 months, there was no recorded mortality due to the disease and 12 of the patients were categorized as disease-free using the criteria for ablation outlined above.

Discussion Hu ¨rthle cell carcinomas are uncommon and may be more aggressive than other differentiated thyroid cancers. The relapse rate is high (50%) and the 5-year survival rate is 60% in patients with distant metastases [1–7]. In this series, it is also observed to be 3% in all differentiated thyroid carcinomas, in the fourth to fifth decade. This result is similar to other studies. The aggressiveness of Hu ¨rthle cell carcinoma is reported to be higher than other types of differentiated thyroid cancer. A recurrence rate of 36.4% has been reported [1] although we did not find this. Three of our patients had exthrathyroidal invasion, five had intrathyroidal invasion and five were encapsulated (they had only intracapsular invasion) and all were ablated with radioiodine except one patient with pulmonary metastases. Carcangy´u et al. [16] have shown that local recurrence of HCC correlated with the extent of surgery, with recurrence rate for nodulectomy, thyroid lobectomy and total thyroidectomy of 75%, 40% and 15%, respectively. Kushchayeva et al. [13] investigated the possible factors of aggressiveness of differentiated thyroid cancer and concluded that initial metastasis and initial surgical treatment influenced the prognosis. In our series, 12/13 patients had total thyroidectomy and one patient had near-total thyroidectomy, lymph node metastases were observed in three patients (23%) and lymph node dissection was performed in these patients. The low rate of recurrence in our patients may be due to the extent of surgery performed. Chen et al. also reported the low recurrence rate and explained it with the extent of surgery [15]. The mortality rate at 5 years has been reported as 80% [9]. Pulmonary metastases are most common but also occur in bone, liver and the central nervous system [4,16]. We also observed pulmonary metastasis at the initial diagnosis in our series in one patient. In our series the rate was low, which might be due to the small number of patients. The size of tumours is another factor that influences the prognosis of HCC. Several authors have reported that the risk of malignancy was higher when the tumour size was up to 2 cm. Thompson et al. [17] showed the risk of malignancy was high and is up to 2 cm in diameter, and Azadian et al. [18] up to 4.6 cm. On the other hand Carcagny´u et al. [16] evaluated 153 patients with HCC and showed that tumours of less than 1 cm were benign and more than 10 cm were malignant [16]. Chen et al. [15] reported that the size of a Hu ¨rthle cell tumour was a predictor of malignancy. In our series, the tumours were

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Hu¨rthle cell carcinoma Ku¨c¸u¨k et al. 379

1–6 cm in diameter. Distant metastasis was observed in one patient with tumours greater than 4 cm. All patients except one with pulmonary metastasis were categorized as disease-free during the follow-up period of 85 months. Our results were also comparable with other studies. Although HCC cells take up radioiodine, the amount is less than 10% of the uptake into normal tissue. 131I treatment is given to ablate residual tissue and to identify persistent or recurrent disease. Hu ¨rthle cell carcinomas generally secrete Tg and serum anti-Tg levels may be used to follow up these patients. We gave 131I therapy to ablate remnant tissue and we used Tg levels to follow up recurrence. Twelve of 13 patients were ablated with first 131 I treatment and their Tg levels did not increase. They were accepted as ‘disease-free’. 131I therapy was given twice in only one patient. His Tg level did not change and there was no recorded mortality due to the disease during 85 months.

References 1

2 3

4

5 6

7

8 9

10

Conclusion We did not find higher incidences of local recurrences, distant metastases or mortality rates compared to well differentiated thyroid carcinomas [1]. Hu ¨rthle cell carcinomas of the thyroid and well differentiated thyroid carcinomas have similar biological behaviour. Their treatment should be similar, including total or near-total thyroidectomy plus modified cervical node dissection when there is lymph node involvement. Radioactive iodine therapy and suppressive laevothyroxine therapy should follow. When treated aggressively, Hu ¨rthle cell carcinoma of the thyroid, an oncocytic variant of follicular carcinoma, has a favourable outcome, similar to that of well differentiated thyroid carcinomas.

11

12 13 14 15

16 17 18

Hundahl SC, Fleming ID, Fremgen AM, Mench HR. A national cancer database report on 53,856 cases of thyroid carcinoma treated in the U.S., 1985–1995. Cancer 1998; 82:2638–2648. Cooper DS, Schneyer CR. Follicular and Hurthle cell carcinoma of the thyroid. Endocrinol Metab Clin North Am 1990; 19:577–591. Hamann A, Gratz KF, Soudah B, Fritsch RS, Georgi A, Hundeshagen H. The clinical course of oxyphilic carcinoma of the thyroid. Nuklearmedizin 1992; 31:230–238. Har-el G, Hadar T, Segal K, Levy R, Side J. Hurthle cell carcinoma of the thyroid gland. A tumor of moderate malignancy. Cancer 1986; 57: 1613–1617. Tollefsen HR, Shah JP, Huvos AG. Hurthle cell carcinoma of the thyroid. Am J Surg 1975; 130:390–394. Watson RG, Brennan MD, Goellner JR, van Herden JA, McConahey WM, Taylor WF. Invasive Hu¨rthle cell carcinoma of the thyroid: natural history and management. Mayo Clin Proc 1984; 59:851–855. McDonald MP, Sanders LE, Silverman ML, Chan HS, Buyske J. Hurthle cell carcinoma of the thyroid gland: prognostic factors and results of surgical treatment. Surgery 1996; 120:1000–1005. Janser JC, Solis C, Rodier JF, Ghnassia JP. Oncocytic cancers of the thyroid. Hu¨rthle cell cancers. Chirurgie 1996; 121:28–36. Ruegemer JJ, Hay ID, Bergstrahl E J, Ryan JJ, Offord Kp, Gormon CA. Distant metastases in differentiated thyroid carcinoma. J Clin Endocrinol Metab 1988; 67:501–508. Sanders L, Silverman M. Follicular and Hurthle cell carcinoma: predicting outcome and directing therapy. Surgery 1998; 124:967. Arganini M, Behar R, Wu T-C, Straus F 2nd, McCormick M, De Groot L J, Kaplan EL. Hurthle cell tumors: a twenty-five-year experience. Surgery 1986; 199:1108. Sugino K, Ito K, Mimura T, Komeyama K, Iwasaki H, Ito K. Hurthle cell tumor of the thyroid: analysis of 188 cases. World J Surg 2001; 25:1160. Kushchayeva Y, Duh QY, Kebemew E, Clark OH. Prognostic indications for Hu¨rthle cell cancer. World J Surg 2004; 28:1266–1270. Yutan E, Clark OH. Hurthle cell carcinoma. Curr Treat Options Oncol 2001; 2:331–335. Chen H, Nicol TL, Zeiger MA, Dooley WC, Ladenson PW, Cooper DS, et al. Hurthle cell neoplasms of the thyroid: are there factors predictive of malignancy. Ovid Chen Ann Surg 1998; 227:542–546. Carcangy´u ML, Bianchi S, Savino D, Vaynick IM, Rasai J. Follicular Hurthle cell tumors of the thyroid. Cancer 1991; 68:1944–1953. Thompson NW, Dunn EL, Batsakis JG, Nishiyama RH. Hurthle cell lesions of the thyroid gland. Surg Gynecol Obstet 1974; 139:555–560. Azadian A, Rosen IB, Walfish PG, Asa SL. Management considerations in Hurthle cell carcinoma. Surgery 1995; 118:711–715.

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Original article 123

I-FP-CIT in progressive supranuclear palsy and in Parkinson’s disease: a SPECT semiquantitative study

Luca Filippia, Carlo Mannia, Mariangela Pierantozzib, Livia Brusab, Roberta Danielia, Paolo Stanzioneb and Orazio Schillacia Background and aim It is still debated whether or not 123 I-FP-CIT single photon emission computerized tomography (SPECT) is able to differentiate between Parkinson’s disease and progressive supranuclear palsy (PSP). Our aim was to use SPECT semiquantitative analysis to assess the capacity of 123I-FP-CIT to characterize Parkinson’s disease versus PSP. Patients and methods Twenty-one Parkinson’s disease patients, 15 disease duration- and age-matched PSP patients and 20 age-matched healthy controls were included in this study. SPECT imaging was always performed at 4 h post-injection. The ratios of striatal (S) to non-specific occipital (O) binding for the entire striatum (S/O), caudate nuclei (C/O), putamina (P/O) were calculated in both the basal ganglia. The asymmetric index (AI) for the whole striatum was also calculated for Parkinson’s disease and PSP.

compared to Parkinson’s disease group. The asymmetric index (AI) was significantly higher (P < 0.001) in Parkinson’s disease than in PSP (AI: 23.6% ± 15.07% vs. 9.66% ± 5.83), but with an overlap between the two groups. Conclusion Our results confirm that 123I-FP-CIT SPECT is clinically useful for detecting nigrostriatal degeneration both in Parkinson’s disease and PSP. Moreover, in our series, semiquantitative analysis using 123I-FP-CIT SPECT allowed Parkinson’s disease and PSP to be discriminated because PSP patients presented a more severe and symmetric dopamine transporter loss, and the results for S/O were more accurate. Nucl Med Commun 27:381–386

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:381–386 Keywords: dopamine transporter imaging, progressive supranuclear palsy

123

I-FP-CIT, Parkinson’s disease,

Results Compared to healthy controls, S/O, C/O and P/O were significantly reduced (P < 0.001) both in Parkinson’s disease ( – 46%, – 43%, – 49%, contralaterally to the most affected side; – 41%, – 37%, – 41%, ipsilaterally) and in PSP ( – 58%, – 57%, – 59%, contralaterally; – 58%, – 57%, – 59%, ipsilaterally). S/O, C/O and P/O ratio values were significantly (P < 0.001) lower in PSP patients when

Departments of aBiopathology and Diagnostic Imaging and bNeurological Sciences, University ‘‘Tor Vergata’’, Rome, Italy.

Introduction

Since in both Parkinson’s disease and atypical parkinsonian syndromes the loss of dopaminergic cells is associated with a depletion of dopamine transporters (DATs), several SPECT tracers (i.e., 2b-carboxymethoxy3b-(4-[123I]iodophenyl)tropane (123I-b-CIT), N-o-fluoropropyl-2b-carboxymethoxy-3b-(4-[123I]iodophenyl)nortropane (123I-FP-CIT), and 99mTc-TRODAT-1) have been introduced as sensitive and accurate tools for the diagnosis of Parkinson’s disease at the early stage [5–7]. In particular, 123 I-b-CIT has been recently applied to discriminate between Parkinson’s disease and PSP [8,9]. However, 123 I-b-CIT is characterized by relatively slow kinetics so that images are commonly acquired 20 h after injection.

Nuclear imaging with both positron emission tomography (PET) and single photon emission computerized tomography (SPECT) tracers are considered to be accurate and sensitive in the diagnosis of Parkinson’s disease [1,2]. However, several pathological studies [3,4] have demonstrated that the clinical accuracy in the diagnosis of Parkinson’s disease diagnosis is less than 80%, thus up to 10–20% of patients in whom Parkinson’s disease is suspected present a different diagnosis. Atypical parkinsonian syndromes such as multiple system atrophy, progressive supranuclear palsy (PSP) and corticobasal degeneration account for most cases erroneously classified as Parkinson’s disease. In fact, discrimination between Parkinson’s disease and PSP on the basis of clinical features may be difficult, especially in the early stages of disease, when most of the characteristic signs of PSP have not been manifested.

Correspondence to Dr Orazio Schillaci, viale Mazzini 121, 00195 Rome, Italy. Tel: + 0039 06 2090 2419; fax: + 0039 06 3735 8644; e-mail: [email protected] Received 21 October 2005 Accepted 12 December 2005

123

I-FP-CIT shows faster kinetics than 123I-b-CIT allowing SPECT to be performed 3–4 h after injection so that a single-day study protocol is feasible. Moreover, it is commercially available and routinely used in many clinical

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382

Nuclear Medicine Communications 2006, Vol 27 No 4

centres. However, only two studies [10,11] have evaluated the scintigraphic pattern of 123I-FP-CIT distribution in Parkinson’s disease and PSP patients. With regard to the possible usefulness of 123I-FP-CIT in discriminating Parkinson’s disease versus PSP, though, these papers lead to divergent conclusions [10,11]. Therefore, the aim of this study was to evaluate if 123IFP-CIT with semiquantitative analysis is able to distinguish Parkinson’s disease from PSP.

Patients, materials and methods Parkinson’s disease patients

Twenty-one patients (aged 51–75 years; mean, 64.3 ± 7.7 years; nine females and 12 males) affected by a rigid akinetic form of idiopathic Parkinson’s disease, diagnosed according to the United Kingdom Parkinson’s Disease Society Brain Bank (UKPDSBB) criteria were consecutively recruited from the movement disorder outpatient clinic. Exclusion criteria were the presence of cerebral lesions in the computerized tomography and metabolic disorders. All patients underwent an accurate neurological examination; the stage of Parkinson’s disease, assessed using the Hoehn & Yahr staging scale ranged between 2 and 3. The mean duration disease from the first symptoms onset to SPECT imaging was 32 ± 22.2 months. All Parkinson’s disease subjects showed a Mini Mental Scale Evaluation Z 24. Eleven patients presented bilateral motor signs with slight one-side prevalence; 10 subjects were affected to a similar extent bilaterally. Eight patients were drug free; the other 13 were taking L-DOPA. Patients with progressive supranuclear palsy

We enrolled 15 patients (aged 50–70 years; mean, 63.5 ± 5.6 years; six females and nine males), according to diagnostic criteria for probable disease by the NINDSSPSP International workshop [12]. Motor disability related to parkinsonism ranged from 2.5 to 4 on the Hoehn & Yahr scale. The mean duration disease from the first symptoms onset to SPECT imaging was 32 ± 13.8 months. Six out of 15 patients presented a Mini Mental Scale Evaluation < 24. Nine patients were free from antiparkinsonian drugs, six subjects were taking L-DOPA.

Imaging was performed 4 h after the administration of 185 MBq of 123I-FP-CIT (DATSCAN, Amersham Health, UK) using a variable angle dual-head gamma camera (Millennium VG, GE Medical System) equipped with high resolution, low energy, parallel hole collimators. The energy window was set at 159 keV ± 10%. SPECT was acquired in a 128  128 matrix, obtaining multiple view over 3601 at 40 s acquisition time per projection with an angular step of 31. Images were reconstructed using Butterworth filtered back-projections (cut-off 0.5 and order 10); transverse, sagittal, coronal cantho-meatal oriented slices were generated. Chang’s correction method was used to compensate for photon attenuation using a coefficient, m, of 0.11 cm – 1 [13]. Qualitative analysis was performed independently by three well-experienced nuclear medicine physicians (L.F., C.M. and O.S.) who were blind to patients’ clinical data. For semiquantitative analysis, to calculate the ratio of specific to non-specific uptake, the three adjacent transaxial slices with the highest tracer uptake were summed. A standard region of interest (ROI) template, constructed as previously described by Booij et al. [14], including fixed ROIs for both caudate nuclei, putamina and occipital cortices, was placed on the summed images. Small variations of individual brain required movement of the ROIs within the template, without changing their size or shape, for optimal positioning. Therefore, exactly the same ROIs were used in all patients for both images. The ratios of specific to non-specific binding for caudate nuclei (C/O) and putamina (P/O) was then calculated as 123 I-FP-CIT binding = (ROI – O)/O, in which ROI represents the mean counts in the region of interest (putamen or caudate nucleus) and O in the occipital cortex. The average striatal ROI activity was then calculated for the entire striatum (S/O), left and right striatum, as the size-weighted average of right and left putamen and caudate activity [15]. The specific to non-specific uptake ratios of 123I-FP-CIT were expressed as mean ± standard deviation (SD) and also as percentages of the ratios found in healthy volunteers.

Controls

Twenty age-matched healthy control subjects (aged 51–74 years; mean, 63.8 ± 7 years; 11 females and nine males) with no current or past history of neuropsychiatric diseases were also evaluated. SPECT imaging

All subjects received perchlorate (1000 mg) 30 min before scanning to block thyroid uptake of free radioactive iodide.

The asymmetric index (AI) for the whole striatum was also calculated for Parkinson’s disease and PSP patients from the formula AI = [(Sips – Scont)/Sav]  100, where Sips is the ipsilateral S to the side of dominant symptoms, Scont is the contralateral S, and Sav is the average ipsilateral and contralateral S. In the case of bilateral disease without a predominant side and in the control group, the right S was arbitrarily assumed as the contralateral.

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123

I-FP-CIT in supranuclear palsy and Parkinson’s disease Filippi et al. 383

Statistical analysis

A two-tailed unpaired Student’s t-test was used to analyse differences in DAT activity between Parkinson’s disease patients versus healthy controls: the significance was established at the P < 0.05 level.

Results No significant differences in age (P = 0.72) and in disease duration (P = 0.99) were found between Parkinson’s disease and PSP patients. Qualitative analysis 123

In all the subjects of the control group, I-FP-CIT uptake was symmetrically distributed and highly concentrated in the striatal region. All the 21 Parkinson’s disease patients showed bilateral asymmetric reduction of tracer uptake, more severe in the putamen contralateral to the side of dominant symptoms. Visual analysis revealed severe and symmetric DAT loss in all PSP subjects (Fig. 1). Semiquantitative analysis

Table 1 summarizes the absolute ratios of striatal to nonspecific binding in healthy subjects, in Parkinson’s disease and in PSP patients. When compared to controls, the Parkinson’s disease group showed significant (P < 0.001) reduction in the S/O, C/O and P/O ratios both contralaterally ( – 46%, – 43%, – 49%, respectively) and ipsilaterally ( – 41%, – 37%, – 41%, respectively) to the dominant symptomatic side. Moreover, in Parkinson’s disease patients DAT loss was significantly (P < 0.001) greater in putamina than in caudate nuclei. There was also a significant (P < 0.001) reduction in the S/O, C/O and P/O ratios ( – 58%, – 57%, – 59%, contralaterally; – 58%, – 57%, – 59%, ipsilaterally) in

the PSP patients compared with healthy subjects. S/O, C/O and P/O ratio values were significantly (P < 0.001) reduced in PSP patients as compared to the Parkinson’s disease group. In particular, minimal overlap in striatal to non-specific binding values was found between Parkinson’s disease and PSP patients because all but one PSP subject showed an S/O value at least 1 SD lower than the mean value measured for Parkinson’s disease (Fig. 2). Table 1

Overview of the results

Striatal to non-specific binding ratio values S/O Healthy controls Parkinson’s disease Progressive supranuclear S/O contralateral Healthy controls Parkinson’s disease Progressive supranuclear C/O contralateral Healthy controls Parkinson’s disease Progressive supranuclear P/O contralateral Healthy controls Parkinson’s disease Progressive supranuclear S/O ipsilateral Healthy controls Parkinson’s disease Progressive supranuclear C/O ipsilateral Healthy controls Parkinson’s disease Progressive supranuclear P/O ipsilateral Healthy controls Parkinson’s disease Progressive supranuclear Asymmetric index Parkinson’s disease Progressive supranuclear

Mean ± SD

Range

palsy

2.85 ± 0.44 1.66 ± 0.15 1.22 ± 0.16

2.35–3.35 1.42–1.94 1.01–1.52

palsy

2.91 ± 0.38 1.57 ± 0.15 1.23 ± 0.17

2.33–3.64 1.37–2.00 0.98–1.57

palsy

3.14 ± 0.41 1.79 ± 0.17 1.36 ± 0.21

2.45–3.79 1.54–2.21 1.11–1.84

palsy

2.78 ± 0.40 1.40 ± 0.13 1.12 ± 0.17

2.26–3.66 1.18–1.58 0.94–1.40

palsy

2.93 ± 0.40 1.73 ± 0.17 1.22 ± 0.15

2.30–3.67 1.47–2.10 1.03–1.51

palsy

3.20 ± 0.48 2.01 ± 0.26 1.37 ± 0.18

2.48–4.49 1.70–2.60 1.18–1.71

palsy

2.73 ± 0.38 1.60 ± 0.15 1.12 ± 0.17

2.1–3.60 1.34–1.84 0.86–1.40

palsy

23.60 ± 15.07 9.66 ± 5.83

4.20–51.20 0–22.90

S/O: striatal to non-specific uptake ratio; C/O: caudate nucleus to non-specific uptake ratio; P/O: putamen to non-specific uptake ratio. P value < 0.001 in all cases.

Fig. 1

The scintigraphic pattern of dopamine transporters activity imaged by 123I-FP-CIT SPECT in a healthy subject (a), in a patient with Parkinson’s disease (b) and in a subject with progressive supranuclear paralysis (c).

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384 Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 2

2.5

S/O PD S/O PSP

S/O value

2

1.5

1

0.5

0 No. patients The 123I-FP-CIT striatal to non-specific occipital binding ratio values in Parkinson’s disease (S/O PD) and in progressive supranuclear palsy (S/O PSP): note the minimal overlap between the two groups.

Fig. 3

70

AI PD AI PSP

60

AI % value

50 40 30 20 10 0 No. patients The asymmetric index (AI) calculated in Parkinson’s disease (AI PD) and in progressive supranuclear palsy (AI PSP): a consistent overlap was found between the two groups.

A significantly (P < 0.001) more symmetric reduction of 123 I-FP-CIT binding was found in PSP (AI: 9.66% ± 5.83%) than in the Parkinson’s disease group (AI: 23.6% ± 15.07%); in spite of a consistent overlap between the two groups (Fig. 3).

Discussion Our study indicates that the differentiation between Parkinson’s disease versus PSP can be feasible using 123 I-FP-CIT SPECT with semiquantitative analysis. We found that 123I-FP-CIT uptake is significantly reduced

both in PSP and Parkinson’s disease as compared to controls; nevertheless, for PSP in our series we found a significantly more severe and symmetric DAT loss. We evaluated the distribution of 123I-FP-CIT in PSP and Parkinson’s disease patients both by qualitative and semiquantitative methods. Visual analysis showed bilateral reduction of tracer uptake both in Parkinson’s disease and PSP. Although Parkinson’s disease patients showed a more asymmetric scintigraphic pattern of DAT loss than PSP subjects, it was not possible to discriminate Parkinson’s disease and PSP by qualitative examination alone. In contrast, semiquantitative assessment enabled a precise calculation of the differences in 123I-FP-CIT uptake between Parkinson’s disease and PSP. In particular, the degree of 123I-FP-CIT binding reduction (S/O, – 57%) found in our PSP series is similar to the data reported in other previously published studies performed with PET tracers showing 50% to 60% reduction in caudate nucleus and putamen binding ratio values [16]. However, relatively few experiences have been performed using PET tracers in PSP patients; in particular, Brooks and co-workers [16] reported that 18F-DOPA uptake was more symmetrically reduced in PSP than in Parkinson’s disease patients. It is of note that 18F-DOPA uptake depends on its conversion by the amino acid decarboxylase which might be upregulated in the early phase of the disease, so that this PET radioligand is not considered the most adequate tool to detect dopaminergic damage in the early phase of Parkinson’s disease. Since the introduction of 123I-b-CIT , the assessment of DAT loss has become feasible by the more diffuse SPECT technology. The only two studies [8,9] specifically performed on the possible discrimination between Parkinson’s disease and PSP by 123I-b-CIT SPECT present discordant findings. In fact, although Pirker et al. [8] found that asymmetry in tracer uptake seemed to be less pronounced in PSP than in Parkinson’s disease, this difference did not reach statistical significance. On the contrary, in a small series of PSP patients (n = 5). Messa et al. [9] reported that the analysis of 123I-b-CIT binding in discrete striatal areas allowed to discriminate PSP from Parkinson’s disease patients, since in Parkinson’s disease the head of caudate nucleus is relatively spared from dopaminergic degeneration. It has been shown that 123I-FP-CIT is clinically useful in the diagnosis of Parkinson’s disease, especially regarding the differential diagnosis between Parkinson’s disease and essential tremor [17,18]; nevertheless, there are relatively few data on its possible usefulness to discriminate Parkinson’s disease versus PSP. This is of particular importance considering the long average delay in the diagnosis of PSP after the onset of the first symptoms. In

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I-FP-CIT in supranuclear palsy and Parkinson’s disease Filippi et al. 385

fact, several published papers have reported that PSP is diagnosed at the first clinical examination in only 50% of cases, whereas the remaining 50% of PSP patients are correctly diagnosed 3.5 years or more after the onset of symptoms [19–22]. The difficulties in the early clinical diagnosis of PSP may be partly ascribed to the variable clinical presentations of the disorder and to the fact that the most typical sign (i.e., supranuclear gaze palsy) frequently occurs months or years after the first symptoms. In our series, we performed DAT imaging, on average, 3 years after the onset of clinical symptoms, although, of course, it would be clinically useful to reach the correct diagnosis at an early phase of the disease. Nevertheless, our results suggest that semiquantitative 123 I-FP-CIT SPECT might be of great value for correctly categorizing PSP at an early phase. Recently, Antonini et al. investigated the role of 123I-FP-CIT in differentiating Parkinson’s disease and atypical parkinsonian syndromes. These authors found that this radiopharmaceutical was able to discriminate Parkinson’s disease from PSP, while no significant difference was found between Parkinson’s disease and multiple system atrophy [10]. However, although Parkinson’s disease and PSP subjects presented similar age (62 ± 13 years vs. 64 ± 8 years, respectively) and disease duration (5 ± 4 years vs. 4 ± 3, respectively), the differences between the two groups were not examined statistically and the AI, which may be useful for characterizing the scintigraphic pattern of dopaminergic degeneration, was not calculated [10]. We have carefully matched PSP and Parkinson’s disease patients for age and disease duration. We think that the accuracy in comparing Parkinson’s disease and PSP depends strictly on the characteristics of the study population; in particular, the two groups of patients (i.e., Parkinson’s disease and PSP) should be matched for age to avoid confounding factors due to age-dependent striatum atrophy. To discriminate Parkinson’s disease and PSP, we mainly considered two indices: the S/O and the AI. The calculation of the AI resulted of limited clinical usefulness in our series: although Parkinson’s disease patients showed a relatively more asymmetric reduction of 123IFP-CIT binding than PSP subjects, there was a consistent overlap between the two groups. In particular when the mean + 1 standard deviation of the AI value calculated in PSP (i.e., 9.66 + 5.83) is considered as a cut-off for differential diagnosis, a misclassification occurs in 10/21 (47.6%) of Parkinson’s disease patients. In contrast, by using the S/O index it was possible to obtain an accurate separation between Parkinson’s disease and PSP patients since the overlap between the two groups was minimal. Therefore, the S/O index resulted of utmost value in discriminating Parkinson’s disease versus PSP, suggesting that the severe reduction of tracer binding in PSP might reflect a more profound dopaminergic impairment.

In our series, we used the clinical criteria as the ‘gold standard’. However, the standard clinical criteria have some limitations; in fact, several neuropathological studies have shown that more than 18% of patients thought to be affected by idiopathic Parkinson’s disease according to clinical assessment present other diagnoses at post-mortem examination [23]. In particular, it has been reported that PSP represents 4% of cases of parkinsonism [24]. To better characterize the striatal dopaminergic activity in parkinsonism, besides the DAT tracers several ligands binding post-synaptic D2 receptors have been also introduced in nuclear medicine practice [25,26]. In particular, 123 I-IBZM has been successfully used for the scintigraphic assessment of the post-synaptic dopamine cells loss both in Parkinson’s disease and in atypical parkinsonian syndromes [27,28]. PSP patients showed reduced binding of S-( – )N-[(1-ethyl-2-pyrrolidinyl)methyl]-2-hydroxy-3-[123I]iodo6-methoxybenzamide (123I-IBZM), with frequent overlap with controls. Regarding the possible use of 123I-IBZM in discriminating Parkinson’s disease versus PSP, Buck et al. [29] reported that D2 receptor scintigraphy may be clinically useful to detect the D2 receptor loss in early PSP, contrary to early Parkinson’s disease which often shows slightly increased or unchanged D2 receptor density. In a recent paper [11], Plotkin et al. studied a small cohort (n = 8) of PSP patients both by 123I-FP-CIT and 123 I-IBZM SPECT in order to evaluate the accuracy of these two combined imaging modalities. In this situation, 123 I-FP-CIT was not able to differentiate patients having PSP from those with Parkinson’s disease. The discrepancy between our findings and those obtained by Plotkin’s group might be explained considering the different characteristics of patient population; in particular, in our series both Parkinson’s disease and PSP patients were studied at a relatively early stage when the differential diagnosis is of great clinical usefulness. Moreover, in the previously cited paper [11] the authors found significant DAT loss in 7/8 PSP patients, while a reduction of D2 receptors was found in only 6/8 subjects. These findings suggest that DAT imaging might have higher sensitivity in PSP diagnosis than D2 scintigraphy as previously reported by Kim et al. [28]. However, 123IIBZM was characterized by a substantially lower ability to diagnose atypical parkinsonian syndromes than 123I-FPCIT [11]. So, a normal finding at 123I-IBZM examination cannot exclude an atypical parkinsonian syndrome and 123 I-FP-CIT SPECT should be performed to detect DAT loss particularly at an early phase of PSP. Hence, further studies with larger series are needed to more fully assess the value of combined DAT and D2 receptor imaging in the diagnosis of PSP. Our findings confirm that 123I-FP-CIT represents a useful tool to differentiate patients with nigro-striatal degeneration (i.e., Parkinson’s disease and PSP) from

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healthy controls and indicate that the discrimination between Parkinson’s disease and PSP might be feasible by using 123I-FP-CIT SPECT with semiquantitative analysis. In particular, our data indicate that the S/O value is more useful than the AI to this end. However, accurate clinical examination of patients with suspected parkinsonism is an essential prerequisite, because the clinical impressions and the results of DAT imaging should be integrated to obtain the correct diagnosis. In early PSP, certain clinical signs might be not be clearly recognized, so our results strongly suggest that DAT imaging with 123I-FP-CIT might be useful for supporting a clinical evaluation by providing functional information on the dopaminergic neuropathological damage.

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Bohnen NI, Frey KA. The role of positron emission tomography imaging in movement disorders. Neuroimaging Clin N Am 2003; 4:791–803. 2 Seibyl JP. Imaging studies in movement disorders. Semin Nucl Med 2003; 23:105–113. 3 Rajput A, Rodzilsky B, Rajput A. Accuracy of clinical diagnosis of Parkinsonism – a prospective study. Can J Neurol Sci 1991; 18:275–278. 4 Hughes AJ, Daniel SE, Kilford L, Lees AJ. Accuracy of clinical diagnosis of idiopathic Parkinson’s disease: a clinico-pathological study of 100 cases. J Neurol Neurosurg Psychiatry 1992; 55:1142–1146. 5 Marek KL, Seibyl JP, Zoghbi SS, Zea-Ponce Y, Baldwin RM, Fussel B, et al. [123I]-b-CIT/ SPECT imaging demonstrates bilateral loss of dopamine transporters in hemi-Parkinson’s disease. Neurology 1996, 46:231–237. 6 Filippi L, Manni C, Pierantozzi M, Brusa L, Danieli R, Stanzione P, et al. 123IFP-CIT semiquantitative SPECT detects pre-clinical bilateral dopaminergic deficit in early Parkinson’s disease with unilateral symptoms. Nucl Med Commun 2005; 26:421–426. 7 Mozley PD, Schneider JS, Acton PD, Plossl K, Stern MB, Siderowf A, et al. Binding of [99mTc]TRODAT-1 to dopamine transporters in patients with Parkinson’s disease and in healthy volunteers. J Nucl Med 2000; 4:584–589. 8 Pirker W, Asenbaum S, Bencists G, Prayer D, Gerschlager W, Deecke L, et al. [123I]-b-CIT SPECT in multiple system atrophy, progressive supranuclear palsy, and corticobasal degeneration. Mov Disord 2000; 15:1158–1167. 9 Messa C, Volonte` MA, Fazio F, Zito F, Carpinelli A, d’Amico A, et al. Differential distribution of striatal [123I]b-CIT in Parkinson’s disease and progressive supranuclear palsy, evaluated with single photon emission tomography. Eur J Nucl Med 1998; 25:1270–1276. 10 Antonini A, Benti R, De Notaris R, Tesei S, Zechinelli A, Sacilotto G, et al. 123 I-Isoflupane/SPECT binding to striatal dopamine transporters (DAT) uptake in patients with Parkinson’s disease, multiple system atrophy, and progressive supranuclear palsy. Neurol Sci 2003; 24:149–150. 11 Plotkin M, Amthauer H, Klaffe S, Kuhn A, Ludemann L, Arnold G, et al. Combined 123I-FP-CIT and 123I-IBZM SPECT for the diagnosis of parkinsonian syndromes: study on 72 patients. J Neural Transm 2005; 112:677–692.

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Original article

Colloid scintigraphy in non-alcoholic steatohepatitis: A conventional diagnostic method for an emerging disease Deniz Gu¨ney Dumana, Fuat Dedeb, Hakan Akına, Feyza S¸enb, ¸ ig˘dem C ¸ elikelc and Nurdan To¨zu¨na Halil Turgut Turog˘lub, C Aim Non-alcoholic steatohepatitis (NASH) is a progressive liver disease characterized by diffuse fatty infiltration and Kupffer cell dysfunction which contributes to its pathogenesis. Since the liver biopsy, which is considered the ‘gold standard’ in diagnosing NASH, has some limitations other imaging methods have been explored as alternatives. Colloid scintigraphy is a good method reflecting Kupffer cell activity and we found it worthwhile to evaluate this technique in NASH. We aimed to present the common scintigraphic features and their clinicopathologic correlations in NASH. Methods Twenty-two new patients (11 female, mean age 43.7 ± 10.8) with biopsy-proven NASH underwent colloid liver scintigraphy. The dynamic, static and SPECT images were performed after intravenous injection of 185 MBq 99m Tc tin colloid. Hepatic perfusion, blood pool clearance time, colloid shift to spleen and bone marrow were assessed and liver right/left lobe ratio was calculated. Results The values calculated on static and tomographic (SPECT) images showed good correlation. Liver right/left lobe ratio was altered in all patients. Blood pool clearance time was prolonged in seven (32%) but hepatic perfusion

Introduction Non-alcoholic fatty liver disease (NAFLD) is the general term which encompasses a clinical spectrum varying from simple hepatosteatosis and non-alcoholic steatohepatitis (NASH) to cirrhosis and hepatocellular carcinoma [1]. Simple steatosis is presumably benign and non-progressive, whereas NASH may progress to liver fibrosis [2]. NASH is a common cause of chronic liver disease and is gaining importance due to its potential to progress to liver failure. It constitutes at least 50% of patients in the transplantation lists. Among obese patients, the prevalence of NASH was reported as 20–25% [3]. Despite the fact that fatty infiltration of liver has been recognized for many years, NASH is a relatively new entity which was first introduced by Ludwig in 1980 [4]. The liver has two major cell populations: (1) parenchymal cells, performing metabolic functions, which account for about 85%; and (2) Kupffer cells, responsible for phagocytosis of the foreign particles, and which account for the other 15% [5]. Images of either cell population

was normal in all patients. Colloid shift to the spleen was observed in 55% of patients using SPECT analysis. No correlation between scintigraphy parameters and histological or biochemical findings were observed. Conclusion Altered liver right/left lobe ratio was the universal finding in all our NASH patients. Other common scintigraphic features of NASH include colloid shift to spleen and prolonged blood pool clearance time. Liver scintigraphy might be a promising non-invasive tool in the follow-up of NASH patients in therapeutic trials. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:387–393 Nuclear Medicine Communications 2006, 27:387–393 Keywords: non-alcoholic steatohepatitis, colloid scintigraphy, right/left lobe ratio, splenic shift, Kupffer cell function Departments of aGastroenterology, bNuclear Medicine and cPathology, Marmara University Hospital, Istanbul, Turkey. Correspondence to Dr Deniz Gu¨ney Duman, Ahmet Refik Sok., Ceylan Apt. No: 19/5, 34730 C ¸ iftehavuzlar, Kady´ko¨y, Istanbul, Turkey. Tel: + 90 5322 470566; fax: + 90 2163 267073; e-mail: [email protected] Received 13 October 2005 Accepted 22 December 2005

obtained by scintigraphy may be assumed to demonstrate similar liver morphology since Kupffer cells are scattered homogeneously amongst the parenchymal cells throughout the liver. In fact, most diseases affect both the hepatocytes and adjacent Kupffer cells more or less to a similar extent. Colloid scintigraphy has long been used in the diagnosis and evaluation of numerous liver diseases such as alcoholic hepatitis and cirrhosis. However, its use has somehow declined with the improvement of other complementary imaging modalities like ultrasonography, computed tomography and magnetic resonance. Nevertheless, liver scintigraphy using single photon emission computed tomography (SPECT) provides valuable anatomical and functional information on diseases causing diffuse liver involvement i.e., alcoholic hepatitis, sarcoidosis, amyloidosis, leukaemia, lymphoma, haemachromatosis and cirrhosis [2,3]. Recent data suggest that dysfunctioning Kupffer cells of the reticuloendothelial system (RES) contribute to the pathogenesis of NASH [6]. As Kupffer cells remove

c 2006 Lippincott Williams & Wilkins 0143-3636

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388 Nuclear Medicine Communications 2006, Vol 27 No 4

colloids from the circulating blood by phagocytosis, they are imaged using radioactive colloids. It may be hypothesized that hepatic dysfunction in patients with NASH can be evaluated by determining the radiocolloid uptake into Kupffer cells. However, our search in the PUBMED database for publications through August 2005 related to liver scintigraphy findings and NASH did not yield any information. The aim of this study was to identify the liver colloid scintigraphy findings in patients with biopsy-proven NASH and to assess their correlations with the clinicopathological variables.

Patients and methods Patients with a recent diagnosis of NASH (diagnosis within past 6 months) based on the clinical, biochemical, histological and ultrasonographic findings were included in the study. The diagnosis of NASH was based on the presence of all of the following four criteria: 1. persistently abnormal liver alanine aminotransferase (ALT) levels; 2. a daily alcohol intake of less than 20 g; 3. appropriate exclusion of other causes of chronic liver disease, such as chronic viral hepatitis, drug-induced hepatitis, haemochromatosis, Wilson’s disease, primary biliary cirrhosis, a1 antitrypsin deficiency, autoimmune hepatitis, jejuno-ileal bypass, hyperalimentation; 4. suggestive features in the liver biopsy, including macrovesicular steatosis ( > 10% of hepatocytes) and lobular inflammation together with ballooning degeneration, Mallory hyaline fibrosis or sinusoidal fibrosis. The study protocol was approved by the local ethics committee and informed consent was obtained from each patient. Laboratory tests

Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyl transferase (GGT), alkaline phosphatase (AP), total bilirubin, ferritin, iron, total iron binding capacity, lipid profile (HDL cholesterol, LDL cholesterol, triglyceride, total cholesterol) were determined by standard methods. Body mass index (BMI) was calculated as weight (in kilograms) divided by height (in metres) squared. Liver biopsies were assessed by an experienced pathologist blinded to the clinical, laboratory and scintigraphic data of the patients. The percentage of hepatocytes involved in macrovesicular steatosis was calculated based on the average number of fatty vesicles divided by the

number of hepatocyte nuclei. Hepatic inflammation was quantified by the number of inflammatory foci, counted throughout the liver biopsy with  20 ocular, divided by the number of zones visualized. Ballooning degeneration, Mallory’s hyaline and lipogranulomas found in biopsies were also recorded for each case. Liver–spleen scintigraphy

Liver–spleen scintigraphy was performed according to the Procedure Guidelines for Hepatic and Splenic Imaging 3.0, published by the Society of Nuclear Medicine. Dynamic imaging at the anterior projection was added for evaluating the hepatic blood flow and blood pool activity. A dynamic study (64  64 matrix, 1 frame/s for 120 s) was performed immediately after the intravenous bolus injection of 185 MBq of 99mTc tin colloid. Anterior and posterior static scintigraphy (256  256 matrix, 1 million counts) and SPECT (64 images over 3601 on a 64  64 matrix) studies were carried out 20 min after the injection. The patients were examined in the supine position and images were acquired using single-headed, circular, gamma camera. In order to evaluate hepatic biodistribution and biokinetics of radiocolloid, hepatic perfusion time, the blood pool clearance time were determined and liver right/left lobe, liver/spleen and liver/bone marrow radiocolloid uptake ratios were calculated. Hepatic perfusion time, the time difference between the visualization of the aorta and liver activity in the dynamic images (Fig. 1), was accepted normal when it was longer than 6 s. Blood pool clearance time: The time–activity curve was generated from the region of interest (ROI) over the cardiac blood pool activity and time to 50% of the peak activity was defined as a parameter of blood pool clearance (Fig. 2). Normal blood pool clearance time was accepted as less than 10 s [7]. Liver right/left lobe uptake ratio (right/left lobe ratio) was calculated using mean radioactivity counts obtained from ROIs on each lobe which were outlined on both anterior static and transaxial SPECT images (Fig. 3). Normal right/left lobe ratio was accepted as greater than 2.5 [7]. Liver/spleen uptake ratio (liver/spleen ratio) was calculated using mean radioactivity counts obtained from ROIs on liver and spleen which were outlined on both posterior static and transaxial SPECT images (Fig. 3). A liver/ spleen ratio of greater than 1.4 was accepted normal [7]. Liver/bone marrow uptake ratio (liver/bone marrow ratio) was calculated using mean radioactivity counts obtained

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Scintigraphy in non-alcoholic steatohepatitis Duman et al. 389

Fig. 1

1 frame/s

Dynamic anterior images showed the normal hepatic perfusion.

Fig. 2

5 Max roi1

kcounts/s

4

3 1

/2 max

2

T- 1/2 max

1

0 0

10

s

20

30

Blood pool clearance time was calculated from the time–activity curve over the cardiac region.

from ROIs on liver and L4 vertebra which were outlined on posterior static images (Fig. 3).

determined by the Pearson correlation test. A P value of < 0.05 was considered significant.

Statistics

Results were expressed as mean ± SD. Continuous variables were analysed by unpaired Student’s t-tests or the Wilcoxon rank sum test. Categorical variables were compared using the chi-squared test or Fisher’s exact test when appropriate. Correlation between scintigraphic and histological variables and serum liver function tests were

Results A total of 22 patients (11 female; mean age, 43.7 ± 10.8 years) with biopsy proven NASH were included. The demographic and laboratory data are summarized in Table 1. Detailed pathological findings of the patients are given in Table 2.

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390 Nuclear Medicine Communications 2006, Vol 27 No 4

Fig. 3

(a)

(b) roi0 Rt

roi1 Lt

Lt

Rt

(c)

(d) roi1 roi0

L

S roi0 L

roi1 S

roi2 BM Liver right/left lobe uptake ratio was calculated from anterior planar (a) and transaxial SPECT images (b), liver/spleen uptake ratio was calculated from posterior planar (c) and transaxial SPECT images (d) and liver/bone marrow uptake ratio was calculated from the posterior planar image (c). Rt, right; Lt, left; S, spleen; L, liver; BM, bone marrow.

Table 1

Summary of patients’ characteristics (n = 22)

Characteristic Age (years) Gender (male/female) Body mass index (kg/m2) Alanine aminotransferase (IU/L) Aspartate aminotransferase (IU/L) (10–37) Gamma-glutamyl transferase (IU/L) (7–49) Alkaline phosphatase (UI/L) (0–270) Total cholesterol (mg/dL) Triglycerides (mg/dL)

Liver pathology findings of patients with non-alcoholic steatohepatitis (n = 22)

Table 2

Mean ± SD (range) 43.7 ± 10.8 (23–60) 11/11 29.9 ± 3.3 (22.6–36.4) 83.8 ± 26.3 (42–146) 49.8 ± 15.8 (24–93) 65.3 ± 27.6 (29–134) 203.1 ± 99.7 (81–431) 218.9 ± 41.8 (142–307) 216.27 ± 172.5 (66–714)

Histology Necroinflammatory grade 1 2 3 Fibrosis stage 0 1 2 3 Ballooning degeneration Mallory bodies Lipogranulomas

Number of patients (%) 11 (50%) 10 (45%) 1 (5%) 5 13 3 1 18 11 8

(23%) (59%) (13%) (5%) (82%) (50%) (36%)

Colloid scintigraphy findings

All patients had diffuse colloid uptake on both static and SPECT images. The mean hepatic perfusion time was 7.27 ± 0.98 s (6–9 s) and, overall, no abnormality was seen in any patient in terms of hepatic perfusion. The blood pool clearance time was 9.27 ± 3.13 s (5–16 s). It was longer than 10 s in seven (32%) patients. When the patients with longer( > 10 s) and shorter( < 10 s) blood pool clearance times were compared, there was no difference in terms of their histological amount of steatosis, inflammation or serological liver function tests (ALT, AST, GGT, AP).

Static liver/spleen ratio was 2.55 ± 1.99 (0.67–10.93). Similarly SPECT liver/spleen ratio was 1.76 ± 1.31 (0.45– 6.67). There was a perfect correlation between static and SPECT liver/spleen ratios (r = 0.923, P = 0.000) (Fig. 4). We found colloid shift to spleen in three (14%) patients by static analysis, 12 (55%) patients by SPECT analysis. Strikingly, right/left lobe ratio was altered in all patients in favour of the left lobe using both static and SPECT images: for static images right/left lobe ratio was

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Scintigraphy in non-alcoholic steatohepatitis Duman et al. 391

blood pool activity, right/left lobe ratio, liver/spleen ratio, liver/bone marrow ratio) and histological findings of hepatic steatosis or inflammation. A negative correlation between age and blood pool clearance time was observed (r = 0.457, P = 0.033). Apart from this there was no correlation between the clinico-demographic findings (i.e., age, sex, BMI, serum hepatic function tests) and the scintigraphic indices.

Fig. 4

SPECT liver/spleen ratio

6

4

Discussion The current study evaluated the colloid scintigraphy findings in biopsy-proven NASH patients. We found that the right/left lobe ratio was altered in all subjects both by static and SPECT measurements. The colloid shift to the spleen and prolonged blood pool clearance time were the other abnormalities encountered in NASH patients.

2

2

4 6 8 Static liver/spleen ratio

10

Correlation between liver/spleen ratios obtained by SPECT and static images in NASH patients (linear regression). (r = 0.923, P = 0.000)

SPECT right to left lobe ratio

Fig. 5

1.6

1.4

1.2

1.0 1.6 1.2 1.4 Static right to left lobe ratio

1.8

Correlation between right/left lobe ratios obtained by SPECT and static images in NASH patients (linear regression). (r = 0.653, P = 0.001)

1.3 ± 0.25 (0.84–1.78) and SPECT images revealed an average right/left lobe ratio of 1.34 ± 0.21 (1.07–1.72). There was a good correlation between static and SPECT liver images regarding right/left lobe ratios (r = 0.653, P = 0.001) (Fig. 5). Liver/bone marrow ratio was 30.35 ± 7.58 (12.75–46). There was no correlation between the scintigraphy parameters (hepatic perfusion time, clearance rate of

The experience with colloid scintigraphy gained in alcoholic hepatitis can be extrapolated to NAFLD, since liver histology findings in NASH resemble those of alcoholic liver disease. An earlier study showed that scintigraphy had a sensitivity of 94% in detecting liver cirrhosis in biopsy-proven chronic alcoholic liver disease while it declined to 33% in detecting the hepatosteatosis in these patients [8]. Another study for the assessment of steatosis in diffuse alcoholic liver disease revealed sensitivities of 66% and 89% with static imaging and SPECT, respectively [9]. Nevertheless, there are no data on the scintigraphic features of NASH. Liver biopsy is considered as the ‘gold standard’ for the diagnosis of NASH, though the uneven distribution of fat throughout the liver is subject to sampling error [10]. However, performing a liver biopsy for each case is not feasible because patients with this very common disease are often asymptomatic [2]. Therefore, non-invasive imaging modalities for diagnosis and follow-up of these patients are strongly needed. Recently, our group has shown that computed tomography is valuable for predicting the amount of hepatic steatosis in NASH [11]. We believe colloid scintigraphy may be another promising non-invasive diagnostic tool in NASH because of the scintigraphic features defined among NASH patients in the present study. NASH patients are often incidentally detected during routine screening by the finding of abnormal liver function tests. The physical findings are not helpful in most cases and occasionally a slightly enlarged liver is the only abnormality. Ultrasonographic studies may reveal coarsened echo-texture or bright liver, which are not specific for NASH. A diagnosis of NAFLD can be confirmed by liver biopsy after excluding other causes of elevated liver transaminase levels in patients who do not consume significant amount of alcohol. Type 2 diabetes, obesity and dyslipidaemia are the components

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of the metabolic syndrome which is frequently associated with NASH. In an autopsy study, NASH was found in 18.5% of obese patients and 2.7% of lean patients [3]. Given the high frequency of NASH, it might be useful if the usual scintigraphic findings of the disease were defined in order to (1) list the disease features and (2) prevent confusion with the findings of scintigraphic examinations carried out in these patients for other indications. Approximately 85% of the colloid is trapped in the Kupffer cells in liver and the remainder goes mostly to spleen and bone marrow. A decreased colloid uptake by the Kupffer cells reflects hepatic dysfunction which may be due to decreased blood flow, shunting or replacement of liver with fatty or fibrotic tissue [7]. Similarly, a prolonged colloid clearance time may be due to either a reduction in hepatic blood flow or a reduction in effective hepatic mass in the absence of cardiac failure [12]. We found prolonged blood pool clearance time in 32% of NASH patients. Since we did not find any difference in terms of histological hepatosteatosis, hepatic inflammation or biochemical liver function tests between the patients with longer ( > 10 s) and normal blood pool clearance time, we may speculate that abnormal RES function rather than decreased hepatic perfusion accounts for prolonged blood pool clearance time in NASH patients. In fact, none of our patients had liver cirrhosis. We have found altered right/left lobe ratios in favour of left lobe in all our NASH patients. The liver has the capacity to regenerate itself when exposed to a toxic injury and most of this regeneration takes place in the left lobe [13]. As the cirrhosis advances, atrophy of the right lobe with compensatory hypertrophy of the left lobe may develop. Although reversal of the right/left lobe ratio suggests the presence of liver cirrhosis [7], none of our patients had histological and clinical evidence of cirrhosis. Furthermore, all our patients had normal hepatic perfusion time which was a scintigraphic finding supporting the fact that cirrhosis was not present in our patients. Hence, reversal of right/left lobe ratio appeared as a sign of chronic liver pathology in NASH patients rather than cirrhosis, in our study. Eventually, we believe, NASH should be added to the list of the diseases which may lead to reversal of right/left lobe ratio. An explanation for the reversed right/left lobe ratio in NASH may be the higher prevalence of small intestinal bacterial overgrowth compared with control subjects [14]. Continuous exposure to the toxic injury (intestinal bacterial overgrowth) in the left liver lobe may account for the triggered and increased colloid uptake in left lobe relative to the right liver lobe. Increased spleen and bone marrow uptakes may be associated with cirrhosis, diabetes, malignancy, melanoma,

mucopolysaccharidosis or hepatitis [12]. Decreased hepatic sequestration due to increased intrahepatic resistance will lead to increased amount of colloid perfusing these organs, so that liver/spleen and liver/ bone marrow ratios decrease. Using SPECT images we found colloid shift to spleen as reflected by a decreased liver/spleen ratio in 55% of patients. Not surprisingly, by using SPECT images rather than the static images we have detected more patients with a colloid shift to the spleen because the former technique is capable of generating higher quality images [15]. Thus, although the static and SPECT techniques for liver and spleen images appear to correlate well in our study, SPECT may be more sensitive in reflecting the pathology in NASH. The data on liver/bone marrow ratio in the literature is scarce; our mean value for NASH patients was 30.35 ± 7.58. The colloid shift to bone marrow, similar to the splenic shift, is an important sign of impaired liver function. It is not possible, however, to further discuss the clinical impact of our results because the normal limits of this parameter has not yet been well established. An explanation for the failure of demonstrating any correlation between the scintigraphic parameters and the clinico-pathological findings may be the differing levels of Kupffer cell dysfunction in those patients. Theoretically, Kupffer cell capacity may not directly relate to the amount of the hepatosteatosis, hepatic inflammation or fibrosis. Therefore, colloid scintigraphy may be a useful adjunct in diagnosing NASH and its follow-up by exploring the Kupffer cell function since liver biopsy can only assess the architecture of the tissue but not its functions. However, further studies in larger series are warranted. In conclusion, a wide variety of scintigraphic manifestations may be observed in NASH patients. Altered right/ left lobe ratio, splenic shift of the tracer, and prolonged blood pool clearance time are the abnormalities shown from colloid studies in our NASH population. The good correlation between static and SPECT images implies that liver scintigraphy with either technique might be of use in therapeutic trials as a non-invasive tool in the follow-up of NASH patients.

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2

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Bugianesi E, Leone N, Vanni E, Marchesini G, Brunello F, Carucci P, et al. Expanding the natural history of nonalcoholic steatohepatitis: from cryptogenic cirrhosis to hepatocellular carcinoma. Gastroenterology 2002; 123:134–140. Dam-Larsen S, Franzmann M, Andersen IB, Christoffersen P, Jensen LB, Sorensen TI, et al. Long term prognosis of fatty liver: risk of chronic liver disease and death. Gut 2004; 53:750–755. Neuschwander-Tetri BA, Caldwell SH. Nonalcoholic steatohepatitis: Summary of an AASLD single topic conference. Hepatology 2003; 37:1202–1219. Ludwig J, Viggiano RT, McGill DB. Nonalcoholic steatohepatitis: Mayo Clinic experiences with a hitherto unnamed disease. Mayo Clin Proc 1980; 55:342–348.

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Oppenheim BE, Wellman HN, Hoffer PB. Liver imaging. In: Gottschalk A, Hoffer PB, Potchen EJ (editors): Diagnostic Nuclear Medicine. Baltimore: Williams and Wilkins; 1988, pp. 538–567. 6 Diehl AM. Nonalcoholic steatosis and steatohepatitis IV. Nonalcoholic fatty liver disease abnormalities in macrophage function and cytokines. Am J Physiol Gastrointest Liver Physiol 2002; 282:G1–G5. 7 Arkles LB. Diffuse disease of the liver and spleen. In: Murray IPC, Ell PJ, Strauss HW (editors): Nuclear Medicine in Clinical Diagnosis and Treatment. Edinburgh: Churchill Livingstone; 1994, pp. 55–62. 8 Meek DR, Mills PR, Gray HW, Duncan JG, Russell RI, McKillop JH. A comparison of computed tomography, ultrasound and scintigraphy in the diagnosis of alcoholic liver disease. Br J Radiol 1984; 57:23–27. 9 Delcourt E, Vanhaeverbeek M, Binon JP, Brasseur P, Calay R, Baudoux M, et al. Emission tomography for assessment of diffuse alcoholic liver disease. J Nucl Med 1992; 33:1337–1344. 10 Brunt EM. Nonalcoholic steatohepatitis: definition and pathology. Semin Liver Dis 2001; 21:3–16.

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Duman DG, C ¸ elikel C ¸ , Tu¨ney D, ˙Imeryu¨z N, Avsar ¸ E, To¨zu¨n N. Computed tomography in nonalcoholic fatty liver disease: a useful tool for hepatosteatosis assessment? Dig Dis Sci 2006; 51:346–351. Harbert JC. The liver. In: Harbert JC, Eckelman WC, Neumann RD (editors): Nuclear Medicine Diagnosis and Therapy. New York: Thieme Medical Publishers; 1996, pp. 651–683. Sodee DB, Velchic MG, Noto RB, Reilley J, Alavi A. Gastrointestinal system. In: Early PJ, Sodee DB (editors): Principles and Practice of Nuclear Medicine. St Louis: Mosby; 1995, pp. 476–534. Wigg AJ, Roberts-Thomson IC, Dymock RB, McCarthy PJ, Grose RH, Cummins AG. The role of small intestinal bacterial, intestinal permeability, endotoxaemia, and tumour necrosis factor a in the pathogenesis of non-alcoholic steatohepatitis. Gut 2001; 48: 206–211. Oppenheim BE, Wellman HN, Hoffer PB. Liver imaging. In: Gottschalk A, Hoffer PB, Potchen EJ (editors): Diagnostic Nuclear Medicine. Baltimore: Williams and Wilkins; 1988, pp. 538–567.

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Original article

Normal values of [99mTc]pertechnetate uptake and excretion fraction by major salivary glands Dalton A. Anjos, Elba C.S.C. Etchebehere, Allan O. Santos, Mariana C.L. Lima, Celso D. Ramos, Raquel B. Paula and Edwaldo E. Camargo The assessment of the functional status of the salivary glands has been used in the scintigraphic evaluation of xerostomia. Several quantitative methods derived from standard dynamic scintigraphy have been suggested. However, the indices proposed are quite variable and unlikely to be useful in clinical practice. The objectives of this study were to obtain reference values of major salivary glands uptake and excretion fraction in healthy subjects and to obtain normal ratios of 99m Tc-pertechnetate uptake by the major salivary glands in comparison to the thyroid gland uptake. The standardization of these values has the purpose of making this evaluation faster and more objective. Fifty volunteers without clinical evidence of xerostomia or thyroid disease underwent static salivary glands scintigraphy with 99mTc-pertechnetate. Static images were obtained at 20 minutes and then at 3 minutes after oral stimulation with lemon juice. Percent uptake, excretion fraction and salivary gland to thyroid ratio rates were calculated for the parotid and the submandibular glands. The mean of the uptake values at 20 minutes for the right and left parotid glands were respectively 0.31% and 0.26%, and for the submandibular glands 0.15%. The excretion fraction of the tracer after the lemon juice stimulation was 70% for the parotids glands, 50% for the right and 49% for

Introduction The salivary glands are structures located in the oral cavity and are responsible for the production and secretion of saliva. The dysfunction of the salivary glands is the main determinant of xerostomia, which can cause alteration of eating habits, difficulty of speech, taste impairment, intolerance to dental prostheses and increased susceptibility to decay and opportunistic infections of the oral cavity [1,2]. Oesophagitis has also been associated with reduction of the buffering capacity of the saliva [3]. Therefore, salivary gland dysfunction can have a serious impact upon the quality of life. Xerostomia has many causes. An adverse drug-related effect is the most common cause. More than 500 medications have been associated with dry mouth complaints. The list includes antihistamines, beta block-

the left submandibular glands. The mean ± SD salivary gland to thyroid count ratio was 0.79 ± 0.45 for the right parotid, 0.78 ± 0.5 for the left parotid, 0.67 ± 0.33 and 0.66 ± 0.34 for the right and left submandibular glands, respectively. Salivary glands scintigraphy with uptake and excretion fraction calculation is an easy to perform, non-invasive and objective method to investigate salivary glands function. These findings help the nuclear physician to interpret salivary gland scintigraphy more objectively, even in patients with thyroid gland dysfunction in whom 99m Tc-pertechnetate thyroid uptake may be abnormal c 2006 Lippincott Nucl Med Commun 27:395–403
Williams & Wilkins. Nuclear Medicine Communications 2006, 27:395–403 Keywords: Salivary glands, Excretion fraction

99m

Tc-pertechnetate, Quantitive analysis,

Division of Nuclear Medicine, Department of Radiology, Campinas State University (UNICAMP), Campinas, Brazil Correspondence to Dr Dalton A. Anjos, Servic¸o de Medicina Nuclear, Hospital das Clı´nicas da UNICAMP, Av. Zeferino Vaz S/N, Cx. Postal 6142, Campinas, SP, CEP 13081-970, Brazil. Tel: + 0055 + 19 3788 7801; fax: + 0055 + 19 3788 7821; e-mail: [email protected] Received XXXX Revised XXXX Accepted XXXX

ers, omeprazole, atropines, benzodiazepines, antipsychotics and tricyclic antidepressants [4]. Radiotherapy, chemotherapy, chronic graft-versus-host disease, Sjo¨gren’s syndrome, sarcoidosis, amyloidosis, HIV, Epstein–Barr virus and hepatitis C virus infections, diabetes mellitus, chronic renal failure, primary biliary cirrhosis, chronic pancreatitis, cystic fibrosis and salivary gland agenesis are other possible aetiologies [5].

Xerostomia is also a common problem among the elderly. Prescribed and over-the-counter medications, Sjo¨gren’s syndrome and previous radiotherapy are the primary causes of hyposalivation in the elderly population [6]. Smoking, chewing habits and menstrual status are also known to change salivary function [4,7].

c 2006 Lippincott Williams & Wilkins 0143-3636


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396 Nuclear Medicine Communications 2006, Vol 27 No 4

The evaluation of the salivary glands can be performed by computerized tomography and sialography. However, the effectiveness of these methods is questioned because in most cases structural abnormalities may not be found [7].

functional capacity of the salivary glands will provide an objective evaluation.

In contrast, salivary gland scintigraphy is a valuable resource because it provides a functional evaluation of the salivary glands. It is a non-invasive study that can be performed easily and quickly, it has a low dosimetry, does not interfere with the normal physiology and is well tolerated by patients [1,8–10].

1. To perform a visual analysis of the blood flow, uptake and excretion of major salivary glands in normal individuals. 2. To perform a semiquantitative analysis of the uptake and excretion of the salivary glands in normal individuals. 3. To compare the visual and semiquantitative analyses. 4. To establish the normal ranges for salivary glands uptake and excretion.

[99mTc]pertechnetate is trapped by the lachrymal and salivary glands, as well as the thyroid gland and gastric mucous membrane [11]. Salivary gland scintigraphy can provide quantitative data about glandular function, as percent uptake, concentration and excretion fraction [12–14]. The main indication of salivary gland scintigraphy is in the evaluation of salivary function in patients with xerostomia. The scintigraphic demonstration of a potentially functioning glandular tissue is decisive in the indication of a therapeutic intervention [15,16]. In spite of the lack of specificity of the scintigraphic results, it is useful in the determination of the type and severity of salivary dysfunction [1,16,17]. Qualitative analysis of salivary gland scintigraphy is usually performed by visual analysis comparing the salivary gland uptake to thyroid gland uptake [9]. This is a subjective evaluation. Other qualitative criteria have been proposed, but are still subjective and do not distinguish the different functional phases (accumulation, concentration and excretion) [1].

The objectives of this study were:

Patients, materials and methods Population

The group under investigation consisted of 50 normal volunteers, with ages ranging from 21 to 54 years (30 women and 20 men) who matched the inclusion criteria of (1) the absence of pathologies in the salivary, lachrymal and thyroid glands and (2) a normal physical examination of the oral cavity and the neck. Exclusion criteria were: 1. 2. 3. 4. 5. 6. 7.

8. Several quantitative and semiquantitative methods have been proposed to assist the nuclear medicine physician in the qualitative assessment of salivary glands scintigraphy. Most papers use indices derived from time–activity curves [17–20]. However, almost all of these indices have a high degree of variability [18] and do not distinguish between normal volunteers and xerostomic populations [21–23]. Quantitative indices are necessary to salivary scintigraphy interpretation as thyroid uptake is to thyroid scintigraphy. Those indices need to be simple and easy to obtain. However, quantitative analysis is complementary to qualitative analysis, and neither can be interpreted without the other. The main objective of obtaining quantitative scores is the differentiation of the normal from the mildly abnormal, where visual analysis has limitations and becomes subjective. Standardization of the normal values for the

9.

Oral discomfort or xerostomic symptoms. Eye discomfort or dry-eye symptoms. History of previous or ongoing cervical radiotherapy. History of previous cervical surgery. Use of drugs that could interfere with the function of the salivary glands. History of thyroid disorders. Use of medications or substances that might interfere with the uptake of [99mTc]pertechnetate into the thyroid. Abnormal values of free thyroxine (0.74–2.1 ng%) or hypersensitive TSH (0.38–6.15 mUI  ml – 1). A positive pregnancy test.

All individuals gave written, informed consent to participate in the study. Salivary gland scintigraphy

In preparation for the study, the volunteers were placed on a low iodine diet and conventional thyroid scintigraphy preparation for 15 days [24]. The volunteers were positioned in the supine position, with the neck slightly extended under the detector of a computerized scintillation camera with a low energy, high resolution, parallel hole collimator. Each participant was positioned so that the major salivary glands and the thyroid were in the centre of the field of view. Sequential images were acquired at 2 s per frame for 80 s and at 15 s per frame for 20 min, in the anterior position of

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Pertechnetate uptake and excretion fraction of salivary glands Anjos et al. 397

the head and neck, immediately after the intravenous injection of 555 MBq (15 mCi) of [99mTc]pertechnetate. At the end of the dynamic acquisition, a static image of the same area was acquired in the same position with 500 000 counts. The volunteers were then orally stimulated with lemon juice and another image of the same area was acquired for the same time interval as the baseline image. Images of the syringe (before and after the injection), the site of radiotracer injection and the thyroid were also recorded for calculation of the percent uptake of the glands. The static approach was chosen because dynamic salivary gland scintigraphy is prone to artifacts caused by a subject’s movement during stimulation by lemon juice. Visual analysis

Two nuclear medicine physicians blindly performed the visual analysis. The image reading was subjective and grey-scale colour bars were not used. If the second nuclear medicine physician’s reading conflicted with the

first physician’s, then a third independent nuclear medicine physician interpreted the study, and this helped to establish the final score by means of majority opinion. All volunteers had the four major salivary glands. The blood flow images were classified as normal, mildly, moderately or markedly increased or decreased. Normal blood flow to the salivary glands was defined as equal to the carotid arteries blood flow 2 s after their peak activity. Salivary gland blood flow was considered as increased or decreased when higher or lower than the peak carotid artery activity, respectively. The thyroid gland uptake was calculated and used as a reference for evaluation of the salivary gland function. Normal salivary gland uptake was defined by comparison with the thyroid uptake (which was used as reference) and the background activity. Salivary gland uptake was graded to be normal (when equal to the thyroid gland uptake), mildly, moderately or markedly increased or decreased (Fig. 1).

Fig. 1

Dynamic images at 100 s per frame show normal [99mTc]pertechnetate uptake by the major salivary glands and by the thyroid gland. Note the presence of unstimulated saliva in the oral cavity after the third image. The normalcy criteria used in this example is the same described above.

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398 Nuclear Medicine Communications 2006, Vol 27 No 4

Salivary gland uptake was considered to be markedly decreased when equal to the background activity (shoulder) and four times lower than the thyroid uptake; as moderately decreased when two times higher than the background activity (shoulder) and three times lower than the thyroid uptake; as mildly decreased when three times higher than the background radiation (shoulder) and two times lower than the thyroid uptake.

Salivary gland uptake was considered to be markedly increased when the uptake was four times higher than the thyroid uptake; as moderately and mildly increased when the uptake was three times and twice as high as the thyroid uptake, respectively. Salivary gland uptake intensities of four, five and six times the background activity were considered as mildly, moderately or markedly increased, respectively.

Fig. 2

Salivary gland excretion was evaluated by comparison of radiotracer concentration before and after lemon juice stimulation (Fig. 2). The excretion was classified as normal, mildly, moderately or markedly increased or decreased. Normal excretion was determined as the activity in the salivary glands after lemon juice stimulation equal to the background activity. When salivary gland activity after stimulation was of similar intensity to the unstimulated salivary gland activity, excretion was considered as markedly decreased. Salivary gland activity after stimulation greater than two and three times the background radiation were classified as mildly and moderately decreased excretion, respectively. Semi-quantitative analysis Uptake into the salivary glands

Regions of interest (ROIs) were drawn over the parotid (RP, LP) and submandibular glands (RS, LS), the thyroid gland (Th) and the background activity (BG). These ROIs were drawn on the images obtained 20 min after injection of the radiotracer. Background ROIs were drawn over the supraclavicular regions (Fig. 3). These ROIs were also drawn on the images obtained after the lemon juice stimulation. Images of the syringe before (SCb) and after (SCa) the injection were also used for calculation of the absolute uptake. The uptake values of the major salivary glands and the thyroid gland were calculated as a percent of the injected dose of [99mTc]pertechnetate. All counts were decay corrected for 99mTc. Uptake into the right parotid gland was calculated as RP counts  BG counts : SCb  SCa Similarly, uptake into the left parotid gland was calculated as LP counts  BG counts : SCb  SCa Images of the cervical area before (A) and after (B) lemon juice stimulation. Note the normal washout of the tracer from the parotid and submandibular glands, and a small amount of unstimulated saliva in the oral cavity.

Uptake into the right submandibular gland was RS counts  BG counts : SCb  SCa

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Pertechnetate uptake and excretion fraction of salivary glands Anjos et al. 399

counts were subtracted from counts obtained for each gland:

Fig. 3

EF% ¼ Left parotid

Right parotid

SG counts before LJS  BG counts after LJS : SG counts before LJS

Results Visual analysis

Right submandibular

Left submandibular

Thyroid Background

Background

The regions of interest drawn around the parotid glands, the submandibular glands and the thyroid gland.

Uptake into the left submandibular gland was LS counts  BG counts : SCb  SCa Uptake into the thyroid gland was Th counts  BG counts : SCb  SCa Uptake ratio of salivary glands to thyroid gland

The same ROIs drawn in the 20 min images were used for calculation of the salivary glands to thyroid gland ratio (SG/Th) using the equation SG SG counts  BG counts ¼ : Th Th counts  BG counts

The blood flow to the salivary glands was classified as normal in 40 individuals. Seven volunteers presented a mild reduction of the flow to the parotid glands, two showed mild reduction to the submandibular glands and one presented marked reduction of the flow bilaterally to the parotid and submandibular glands (Table 1). Accumulation of the radiotracer in the salivary glands was interpreted as normal in 37 of the 50 volunteers. Accumulation of the tracer in the submandibular glands was classified as mildly reduced in four individuals, moderately reduced in three volunteers and markedly reduced in one volunteer. Accumulation of the tracer in the parotid glands was classified as mildly reduced in two individuals. Three volunteers presented reduced accumulation of the tracer in the parotid and submandibular glands (one with mild, one with moderate and one with marked reduction). In all patients in whom the accumulation was reduced, unstimulated salivation could be documented during the dynamic study by the accumulation of saliva in the oral cavity. The excretion fraction followed the same pattern as that observed for the accumulation. Therefore, the excretion of the radiotracer by the submandibular glands was classified as mildly reduced in four individuals, moderately reduced in three and as markedly reduced in one. The excretion of the tracer by the parotid gland was classified as mildly reduced in two individuals. Three volunteers presented reduced excretion by the parotid and submandibular glands (one with mild, one with moderate and one with marked reduction).

Excretion fraction of the salivary glands

The images obtained before and after stimulation with lemon juice stimulation (LJS) were used to calculate the excretion fraction. ROIs were drawn over the right parotid gland (RP), left parotid gland (LP), right submandibular gland (RS), left submandibular gland (LS), thyroid gland (Th) and background activity (BG) in both images (Fig. 3). The excretion fractions (EF%) for each major salivary gland (SG) were calculated by using the equation below and expressed as a percentage. The background (BG)

Table 1 Blood flow, uptake and ejection fraction visual analyses for all 200 salivary glands Blood flow

Normal Mildly reduced Moderately reduced Markedly reduced

Uptake

Ejection fraction

Parotid

Submandibular

Parotid

Submandibular

Parotid Submandibular

84 14

94 4

90 6

78 10

90 6

78 10

0

0

2

8

2

8

2

2

2

4

2

4

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The uptake ratios of the salivary glands to thyroid were equal to 1.0 in 23 of the 50 volunteers. In 23 individuals, the salivary glands uptake was lower than the thyroid gland uptake, and in four the salivary glands uptake was higher (Table 2). Semi-quantitative analysis

Means and standard deviations were calculated for all data. The confidence interval (CI) for the mean with a 95% CI for normal distribution (Table 1) was also calculated for all data. The Shapiro–Wilks W test was used for evaluation of the distribution pattern of the data. The excretion fractions of the right and left submandibular glands, and of the right submandibular/thyroid uptake ratio followed a normal distribution pattern (P > 0.05). Other variables did not follow a normal distribution pattern (Table 3). Thyroid gland uptake values ranged from 0.56 to 0.80% in 95% of the population studied. The normal uptake for 95% of the population ranged from 0.26 to 0.35% for the right parotid gland, 0.22 to 0.31% for the left parotid gland and 0.12 to 0.17% for the right and left submandibular glands. For the excretion fraction the values ranged from 67 to 74.4% for the right parotid gland, 65.5 to 74.5% for the left parotid gland, 45.3 to 54.4% for the right submanTable 2

Salivary gland (SG) to thyroid (Th) uptake ratios

Salivary gland to thyroid uptake ratio

Number of individuals

SG = Th (ratio = 1) SG > Th ( > 1) SG < Th ( < 1)

Table 3

23 23 4

dibular gland and 44.3 to 53.9% for the left submandibular gland. The salivary glands to thyroid gland ratios ranged from 0.66 to 0.93 for the right parotid gland, 0.64 to 0.93 for the left parotid gland, 0.58 to 0.77 for the right submandibular gland and 0.57 to 0.76 for the left submandibular gland. Comparison between visual and semiquantitative analysis

Normal volunteers were divided in two groups: those who had visually normal (n = 37) uptake into the salivary glands and those who had visually abnormal (n = 37) uptake. Difference between mean uptake values were then detected by using t-tests. When the test for normality failed, the Mann–Whitney rank sum test was performed. The test for normality failed only for the comparison of left parotid uptake. There were no significant differences between the visually normal and visually abnormal groups for the parotids and the right submandibular uptake values (P = 0.996 for the right parotid; P = 0.5 for the left parotid; P = 0.182 for the right submandibular). However, the difference between the mean left submandibular uptake values was statistically significant (P = 0.041) when visually reduced volunteers were compared to the visually normal group. Individuals with salivary glands to thyroid ratio visually equal to 1.0 (n = 23) were compared with the 23 volunteers with salivary glands uptake lower than the thyroid on visual analysis. The group with four volunteers in whom the salivary gland uptake was considered greater than thyroid, was not included. The Mann–Whitney rank sum test was performed. The difference in median values between the two groups was statistically significant for all the major salivary glands (P < 0.001 for the right and left parotids; P = 0.007 for the right submandibular; P = 0.003 for the left submandibular). The t-test showed

Mean, standard deviation and confidence intervals for the salivary glands and the thyroid Mean

Age Uptake (%) Right parotid Left parotid Right submandibular Left submandibular Thyroid Ejection fraction (%) Right parotid Left parotid Right submandibular Left submandibular Ratio Right parotid to thyroid Left parotid to thyroid Right submandibular to thyroid Left submandibular to thyroid

32 years 0.3056 0.2620 0.1462 0.1462 0.6766

Standard deviation 9.83 0.16600 0.16100 0.08413 0.08420 0.42100

Confidence interval

P values

(29.2263–34.8137)

< 0.0001

(0.258–0.353) (0.216–0.308) (0.122–0.170) (0.122–0.170) (0.557–0.796)

0.0024 0.0004 0.0154 0.0004 0.0021

70.72 70.03 49.85 49.13

12.99 15.96 16.08 16.88

(67.033–74.416) (65.491–74.561) (45.282–54.424) (44.333–53.925)

< 0.0001 0.0001 0.0978 0.5588

0.79 0.78 0.67 0.66

0.45 0.50 0.33 0.34

(0.66101–0.92019) (0.64232–0.92528) (0.57633–0.76571) (0.56751–0.76169)

< 0.0001 < 0.0001 0.1971 0.0264

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Pertechnetate uptake and excretion fraction of salivary glands Anjos et al. 401

no statistical difference between the mean thyroid uptake of both groups (P = 0.075). Excretion fraction values were also compared between the visually normal group (n = 37) and the visually abnormal (n = 13). The Mann–Whitney rank sum test showed no statistical differences when right and left parotids excretion fraction median values were compared (P = 0.877 for the right parotid; P = 0.894 for the left parotid). The t-test also showed no statistical difference for the right submandibular excretion fraction values (P = 0.109). However, there was a statistically significant difference between the left submandibular excretion fraction mean values of the two groups (P = 0.009).

Discussion Salivary gland dysfunction is detected by scintigraphy. There are clear differences between the normal population and patients with marked xerostomia [17,23]. However, the differentiation between normal individuals and patients with mild xerostomia is difficult in the initial stages of the disease. There is need for reliable, objective, quantitative methods of evaluation of the functional status of the salivary glands. Therefore, the determination of the normal values of [99mTc]pertechnetate uptake and excretion fraction for the salivary glands is essential for a reproducible and objective evaluation. Qualitative analysis of the blood flow to the salivary glands showed that, even in healthy individuals, the arrival of the radiotracer to the salivary glands can be mildly reduced in a small number of normal individuals (20% in the population studied). This could be due to attenuation of the submandibular glands by mandibles of different thicknesses among individuals. The uptake of [99mTc]pertechnetate by the salivary glands can be mildly reduced in 26% of the individuals, probably due to excretion of unstimulated saliva. This phenomenon can easily be recognized by analysing the dynamic phase of the study with a demonstration of early accumulation of the tracer in the oral cavity. The qualitative analysis was fundamental for the recognition of early unstimulated salivation. This phenomenon has been considered as a cause of the variability of the quantitative scintigraphic salivary indices in studies with normal volunteers [18]. Attempts to quantify early unstimulated salivation have been unsuccessful because of the persistent swallowing of radioactive saliva during the study. A possible explanation for this phenomenon is the previous knowledge by the volunteers that, at a certain moment during the study, lemon juice would be given orally. The perception that oral stimulation is imminent can cause anxiety and, consequently, early stimulation of salivation, without lemon juice. The visual detection of

this phenomenon should, therefore, be included as part of the interpretation of the salivary gland scintigraphy in the daily routine of nuclear medicine laboratories. Unstimulated salivation was more intense and more frequent in the submandibular glands than in the parotid glands. Hermann et al. [18] and Malpani et al. [25] also described the same finding. This was proven in the current study, not only by simply looking at the scintigraphic images, but also by calculating [99mTc]pertechnetate uptake, using the same method as described for thyroid gland uptake [26]. The parotid glands uptake values were approximately twice as high as the mean values of the submandibular glands. Loutfi et al. [27] found parotid glands uptake values 1.5–2 times higher than the submandibular glands uptake. There is no explanation for this phenomenon yet. It may be related to the fact that the predominant function of the parotid glands is digestive secretion, whereas the predominant functions of the submandibular glands are lubrication and protection. The mean uptake values obtained in the present study are comparable to those of Stephen et al. [28] and Vigh et al. [29], but lower than those obtained by Bohuslavizki et al. [11] and Klutmann et al. [14]. However, the methodology used by those authors was very different and based on time–activity curves. Attempts of developing semiquantitative methods for uptake and excretion fraction analysis deriving from time–activity curves were published recently by Loutfi et al. [27], who also found higher values of excretion fraction for the parotid glands in comparison to the submandibular glands. The [99mTc]pertechnetate excretion fraction stimulated with lemon juice also proved to be a powerful tool in the functional evaluation of the salivary glands. It was already known that saliva excretion is the first to be impaired in patients with xerostomia, followed by the gland’s accumulation capacity. However, so far there are no reports attempting to establish a normal range for the excretion fraction using salivary glands static scintigraphy. Empirically, the value 50% has been considered as normal for parotid and submandibular glands. The present study was, therefore, the first to establish the normal salivary glands excretion values, using a simple methodology and with potential applicability in clinical practice. The excretion fractions of the parotid glands were usually higher than those of the submandibular glands. The values obtained in the present study are comparable to those obtained by Bohuslavizki et al. [11] and by Aung et al. [17] despite the different methodologies.

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Kohn et al. [1] observed that 31 of 33 normal individuals presented salivary glands uptake 50% higher than the thyroid gland uptake. The relationship between the salivary glands uptake and the thyroid gland uptake has also been empirically assumed to be equal. The present study revealed that the salivary glands uptake is generally lower than that of the thyroid gland. This finding can be very useful because patients with advanced Sjo¨gren’s syndrome may have reduced thyroid gland uptake as well [30]. Comparisons of visual and semiquantitative analysis showed that the salivary gland to thyroid ratio was the only parameter that followed the visual impression for the four major salivary glands, independently of the thyroid uptake. However, the four volunteers in whom salivary gland uptake was higher than thyroid uptake were excluded from these comparisons. Left submandibular gland uptake and excretion fraction also followed the visual interpretation. On the other hand, uptake and excretion fraction of the other major glands were not statistically different between visually normal and visually abnormal volunteers.

Semiquantitative indexes obtained in the present study showed a wide variability and only a few had a normal distribution. This variability has also been experienced by other authors and was attributed to unstimulated salivation [18]. We suppose that the previous knowledge that lemon juice would be given orally during the study is related to early unstimulated salivation and it could be avoided if patients were not previously informed. Variability of the results makes us believe that the semiquantitative scores obtained may not be useful for distinguishing between normal and early-stages xerostomic populations. However, further work is needed using the indexes obtained to evaluate xerostomic patients. Visual and semiquantitative analysis comparisons showed that the salivary gland to thyroid ratio was the only score that followed the visual impression for the four major salivary glands, independently of the thyroid uptake. This finding should be kept in mind when performing visual analysis of salivary gland scintigraphy.

References Study limitations

Some causes of salivary dysfunction, such as smoking, chewing habits and menstrual status were not controlled. This can explain why some supposed normal individuals showed reduced blood flow, uptake and ejection fractions. This choice was made to obtain a control group closer to the normal population.

1

2

3 4

5

The fact that some individuals showed unstimulated salivation may explain why most of the indices did not follow normal distribution. These individuals were not excluded because it would create an artificial population, and semiquantitative data derived from them would not be useful for clinical applications.

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Conclusions To the best of our knowledge, this is the first study to perform visual and semiquantitative analysis of major salivary glands in a large group of normal individuals using static salivary gland scintigraphy. Even in healthy volunteers, visual impairments of the blood flow, uptake and excretion fraction can be visually detected. We believe that known and unknown factors that were not controlled could have interfered with the results [4,7]. Salivary gland scintigraphy is a simple, non-invasive and safe method to evaluate salivary gland function. The quantitative indexes derived from ROIs drawn on the static images are simple and easy to obtain.

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Kohn WG, Ship JA, Atkinson JC, Patton LL, Fox PC. Salivary gland scintigraphy: a grading scale and correlation with major salivary gland flow rates. J Oral Pathol Med 1992; 21:70–74. Garg AK, Kirsh ER. Xerostomia: recognition and management of hypofunction of salivary glands. Compend Contin Educ Dent 1995; 16:576–584. Sarosiek J, McCallum RW. Mechanisms of oesophageal mucosal defense. Bailliere’s Best Pract Res Clin Gastroenterol 2000; 14:701–717. Porter SR, Scully C, Hegarty AM. An update of the etiology and management of xerostomia. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2004; 97:28–46. Pajukoski H, Meurman JH, Halonen P, Sulkava R. Prevalence of subjective dry mouth and burning mouth in hospitalized elderly patients and outpatients in relation to saliva, medication, and systemic diseases. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2001; 92:641–649. Astor FC, Hanft KL, Ciocon JO. Xerostomia: a prevalent condition in the elderly. Ear Nose Throat J 1999; 78:476–479. Streckfus CF, Baur U, Brown LJ, Bacal C, Metter J, Nick T. Effects of estrogen status and aging on salivary flow rates in healthy Caucasian women. Gerontology 1998; 44:32–39. de Rossi G, Calcagni ML. Salivary gland disorders. In: Murray IPC, ELL PJ (eds): Nuclear Medicine in Clinical Diagnosis and Treatment. London: Churchill Livingstone; 1994, pp. 373–376. Maurer AH. Scintigraphic evaluation. In: Gore RM, Levine MS, Laaaufer I (eds): Textbook of Gastrointestinal Radiology, vol. 1. Philadelphia: W.B. Saunders; 1994, p. 316. Mishkin FS. Radionuclide salivary gland imaging. Semin Nucl Med 1981; 11:258–265. Bohuslavizki KH, Brenner H, Klutmann S, Tinnemeyer S, Werner JA, Mester J, et al. Implementation and indications for quantitative sialoscintigraphy. Laryngorhinootologie 1997; 76:614–624. Hays MT, Berman M. Pertechnetate distribution in man after intravenous infusion: a compartmental model. J Nucl Med 1977; 18:898–904. de Rossi G, Focacci C. A computer-assisted method for semi-quantitative assessment of salivary gland diseases. Eur J Nucl Med 1980; 5:499–503. Klutmann S, Bohuslavizki KH, Kroger S, Bleckmann C, Brenner W, Mester J, et al. Quantitative salivary gland scintigraphy. J Nucl Med Technol 1999; 27:20–26. Pilbrow WJ, Brownless SM, Cawood JI, Dynes A, Hughes JD, Stockadale HR. Salivary gland scintigraphy – a suitable substitute for sialography? Br J Radiol 1990; 63:190–196.

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Baum JB, Fox PC, Neumann RD. The salivary glands. In: Harbert JC, Eckelman WC, Neumann RD (eds): Nuclear Medicine: Diagnosis and Therapy. New York: Thieme Medical Publishers; 1996, pp. 439–444. Aung W, Murata Y, Ishida R, Takahashi Y, Okada N, Shibuya H. Study of salivary gland scintigraphy and determination of the clinical stage of Sjo¨gren’s syndrome. J Nucl Med 2001; 42:38–43. Hermann GA, Vivino FB, Shnier D, Krumm RP, Mayrin V, Shore JB. Variability of quantitative scintigraphic salivary indices in normal subjects. J Nucl Med 1998; 39:1260–1263. Umehara I, Yamada I, Murata Y, Takahashi Y, Okada N, Shibuya H. Quantitative evaluation of salivary gland scintigraphy in Sjo¨gren’s syndrome. J Nucl Med 1999; 40:64–69. Demangeat R, Didon-Poncelet A, Cherfan J, Demangeat J. Stimulated salivary pertechnetate clearance revisited. Clin Nucl Med 2000; 25:888–894. Adams BK, Al Attia HM, Parkar S. Salivary gland scintigraphy in Sjo¨gren’s syndrome: are quantitative indices the answer? Nucl Med Commun 2003; 24:1011–1016. Booker J, Howarth D, Taylor L, Voutnis D, Sutherland D. Appropriate utilization of semi-quantitative analysis in salivary gland scintigraphy. Nucl Med Commun 2004; 25:1203–110. Hermann GA, Vivino FB, Shnier D, Krumm RP, Mayrin V. Diagnostic accuracy of salivary gland scintigraphic indices in xerostomic populations. Clin Nucl Med 1999; 24:167–172.

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Correspondence Nuclear Medicine Communications 2006, 27:405

Early clinical experience and impact of [18F]fluorodeoxyglucose positron emission tomography Ali T. Akpinar Uludag University Faculty of Medicine, Bursa, Turkey. Correspondence to Ali T. Akpinar, Nuclear Medicine Department, Uludag University Faculty of Medicine, Gorukle Campus, 16059 Bursa, Turkey. Tel: þ 90 224 4428400 (ext 1787); fax: þ 90 224 4429212; e-mail: [email protected]

I read with interest the article by Gutte et al. [1] in the November 2005 issue of Nuclear Medicine Communications. The authors describe the influence and impact of [18F]fluorodeoxyglucose positron emission tomography ([18F]FDG PET) on patient management, and the referring physicians’ satisfaction. As stated by the authors, their overall results seem to be in accordance with similar studies published previously. However, I have two concerns with this article. First, as far as I understand, this was a prospective study and was built upon questionnaires sent with each FDG PET report. I believe, in addition to these questionnaires, it would have been of value to obtain questionnaires completed by referring physicians reflecting their in-

tended management plan before the PET investigation, as previously performed in a similar prospective study [2]. Such an approach could have increased the objectivity of the results by minimizing the effects of responder bias. Second, no details were provided about the timing and indications of PET examinations. As stated by the authors, the alteration of patient management depends on the timing of the PET scan. In fact, the authors only declared that, in their study, the PET scan was rather late in patient work-up. If they had clarified at which stage (i.e. diagnosis, staging, therapy monitoring, restaging) of patient management the PET scans had been performed, and had analysed separately, the article would have been more clinically valuable. As the number of detailed studies dealing with the impact of PET on patient management increases, the clinical situations in which PET scan is most helpful will become clearer.

References 1 2

Gutte H, Hojgaard L, Kjaer A. Early clinical experience and impact of 18F-FDG PET. Nucl Med Commun 2005; 26:989–994. Hillner BE, Tunuguntla R, Fratkin M. Clinical decisions associated with positron emission tomography in a prospective cohort of patients with suspected or known cancer at one United States center. J Clin Oncol 2004; 22:4147–4156.

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Correspondence

Correspondence Nuclear Medicine Communications 2006, 27:407–408

Controversies in the estimation of glomerular filtration rate David Simpson

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Correspondence to Dr David Simpson. e-mail: [email protected] 5

The British Nuclear Medicine Society (BNMS) guidelines on the measurement of glomerular filtration rate (GFR) include a list of controversies [1]. Further correspondence has uncovered some more [2]. I would like to add two more issues to the standardization ‘oto do listo’. Paediatric normal ranges

The BNMS recommends an age-matched normal range from 20 to 75 which differs by a few percent from a subsequent large normal study [3]. From ages 2–20 both BNMS and EANM guidelines [4,5] recommend the same normal range (95% confidence interval 80.6–129.4 ml
min1 per 1.73 m2). From ages 0–2, however, the normal data indicated by both sets of guidelines is constructed using Chantler’s correction rather than the Brochner-Mortensen correction recommended by the BNMS guidelines. This results in a step change in the 95% normal range (71.9–151.1 ml
min1 per 1.73 m2 for a patient 1.99 years old). Whilst individual data points can be converted between the two correction methods, this cannot readily be done for distributions. Perhaps a collaborative effort between EANM and BNMS could fill this gap. Application to oncology

BNMS guidelines rightly point out the importance of using GFR non-normalized for body surface area for use in the Calvert formula for carboplatin dosing [6]. Other chemotherapy dosing schemes are not clear-cut. Some recommend a percentage dose reduction from a dose calculated using some measure of body size. This would seem to make a normalized GFR appropriate [7]. It is often difficult for a practising pharmacist to obtain consistent guidance on how to adjust doses for renal impairment [8,9]. Perhaps BNMS could work with pharmacy bodies to bring standardization to this important area.

References 1

2

Fleming JS, Zivanovic MA, Blake GM, Burniston M, Cosgriff PS. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Nucl Med Commun 2004; 25:759–769. Piepsz A, Ham R, De Sadeleer C. Correspondence (plus reply from Fleming et al.) Nucl Med Commun 2005; 26:175–178.

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Grewal GS, Blake GM. Reference Data for 51Cr-EDTA measurements of the glomerular filtration rate derived from live kidney donors. Nucl Med Commun 2005; 26:61–65. Piepsz A, Colarinha P, Gordon I, Hahn K, Olivier P, Sixt R, van Velzen J. Paediatric Committee of the European Association of Nuclear Medicine. Guidelines for glomerular filtration rate determination in children. Eur J Nucl Med 2001; 28:BP31–BP36. Piepsz A, Pintelon H, Ham HR. Estimation of normal chromium-51 ethylene diamine tetra-acetic acid clearance in children. Eur J Nucl Med 1994; 21:12–16. Calvert AH, Newell DR, Grumbrell LA, O’Reilly S, Burnell M, Boxall FE, et al. Carboplatin dosage: prospective evaluation of a simple formula based on renal function. J Clin Oncol 1989; 7:1748–1756. http://www.nlcn.nhs.uk/downloads.php?filename ¼ 405renal.pdf (Last checked 1 Nov 2005). Vidal L, Shavit M, Fraser A, Paul M, Leibovici L. Systematic comparison of four sources of drug information regarding adjustment of dose for renal function. BMJ 2005; 331:263–266. BNF, Martindale, AHFS Drug Information. Drug prescribing in renal failure. Correspondence. BMJ 2005; 331:292–294.

Reply John S. Fleminga and Maureen A. Zivanovicb, Glen M. Blakec, Maria Burnistond, and Philip S. Cosgriffe a Southampton University Hospitals NHS Trust bChristie Hospital NHS Trust, Manchester cGuy’s Hospital, London dSt James’ University Hospital, Leeds and e United Lincolnshire Hospitals NHS Trust, UK.

Correspondence to Professor John Fleming. e-mail: [email protected]

We would like to thank Dr Simpson for his comments on the controversies noted in the BNMS GFR guidelines [1]. It is very useful to have challenges to the guidelines as they provide the motivation to improve them. The aim of the guidelines was to present what we felt was the best approach in the light of current knowledge. It is indeed our hope that improved versions will be created in the future as evidence of better methodology appears. Paediatric normal ranges

Dr Simpson rightly points out that existing normal range data for children provided by Piepsz et al. [2] were calculated by the slope–intercept technique using the simplistic Chantler method to correct for the ‘single exponential assumption’ (SEA). This will result in a systematic overestimate of age-dependent normal values compared to the use of the Brochner-Mortensen correction recommended in the BNMS Guidelines. This has recently been studied by Blake et al. [3], who evaluated GFR in children assumed to be normal by virtue of their DMSA scan. When adjusted for the difference in SEA correction technique, they found their results were in good agreement with the normal range given by Piepsz et al. [2]. As the numbers of patients in the study by

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408 Nuclear Medicine Communications 2006, Vol 27 No 4

Piepsz et al. were larger than in the study by Blake et al., it was recommended that the Piepsz normal range, corrected for this systematic difference in technique, be used. In children between 2 and 17 years of age, this gave a mean and standard deviation GFR of 107 and 17 ml
min1 per 1.73 m2, respectively. The modification of the Piepsz figures to calculate new upper and lower limits of the normal range assumed that the population distribution function is not skewed. This assumption may not be correct and its adoption is therefore not entirely satisfactory. However, it seems a reasonable approach at present until better data are available. The adjusted Piepsz equation can be approximated as follows:
GFR ¼ 107  62  0:985100 a where the GFR is measured in ml
min1 per 1.73 m2 and a is the age in years, valid from 0 to 17 years. The 95% confidence limits on this data are 734 ml
min1 per 1.73 m2. Strictly, this leaves a small age gap between 17 and 20 years, as the adult normal range was constructed using data from patients in the 20–75 year age group [1]. There is thus a small step in mean GFR, from 107 ml
min1 per 1.73 m2 at age 17 to 109 ml
min1 per 1.73 m2 at age 20. Extrapolation between these values would seem reasonable. The difference between the 95% confidence limits at either end of the age gap (734 ml
min1 per 1.73 m2 below age 17 years; 725 ml
min1 per 1.73 m2 above 20 years) is somewhat larger than that for the mean values. Again, extrapolation seems reasonable until more recent evidence can be collated. Recent work from Grewal et al. [4] has described the normal range in adult subjects. Both means and standard deviations differ slightly from those quoted in the guidelines. This is not surprising as the methodology was not exactly the same. However, the encouraging conclusion from the work of Grewal et al. and Blake et al. is that these differences are small, provided the methodology is consistent and aimed at producing absolute values of clearance. Nevertheless, we agree with Dr Simpson on the need to update and improve the confidence in the normal values and subsequent revisions of the guidelines should indeed pool results based on this new evidence. In particular, we encourage centres that acquire GFR data on children who have normal DMSA scans to help build a more robust normal range in this population by making their data available. There is also the question of whether the normal ranges for DTPA and EDTA are different [1]. More normal data on both children and adults is required, particularly for DTPA, to help answer this question. Application in oncology

Dr Simpson has very helpfully pointed out a recent review of chemotherapy dosing schemes that include a description of how dose should be adjusted to account for differences in the patient’s renal function [5]. The authors found that the advice given was often unclear.

For example, there were differences between information sources on basic issues such as whether or not a particular drug needed dose adjustment for renal function. There was also considerable variability in the description of renal function, ranging from undefined general terms such as ‘renal impairment’ to numerical GFR values. All references to GFR in the review are given in units of millilitres per minute and therefore should in theory only refer to absolute GFR. However, there was no mention of the difference between absolute GFR (in millilitres per minute) and GFR normalized for body surface area (expressed in millilitres per minute per 1.73 m2), nor any discussion of the method of calculation. There is thus some doubt as to whether all references to ‘GFR’ refer to absolute values. The numerical difference between GFR and BSA-normalized GFR will be significant in children and differences in SEA correction technique are also known to have a large effect on GFR values obtained [6]. The BNMS could certainly comment on methodological issues to help clear up some of this confusion. It would therefore seem valuable to initiate some dialogue between the BNMS and the relevant pharmaceutical bodies. We feel the following conclusions on technique can be deduced from current evidence: K

GFR is the best overall index of the level of kidney function [7].

K

For each drug the advice must be clear about whether GFR or BSA corrected GFR is to be used. This will depend on the source of data describing the dependence of dose on renal function.

K

For blood sample measurements, it is necessary to use a method that produces unbiased estimates of GFR. This can be achieved by following the BNMS guidelines technique, as this methodology can be traced back to full blood sampling evaluation of 51Cr-EDTA clearance and hence to inulin infusion.

References 1

2

3

4

5

6 7

Fleming JS, Zivanovic MA, Blake GM, Burniston M, Cosgriff PS. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Nucl Med Commun 2004; 24:759–769. Piepsz A, Pintelon H, Ham HR. Estimation of normal chromium-51 ethylene diamine tetra-acetic acid clearance in children. Eur J Nucl Med 1994; 21: 12–16. Blake GM, Gardiner N, Gnanassegaran G, Dizdarevic S. Reference ranges for 51 Cr-EDTA measurements of glomerular filtration rate in children. Nucl Med Commun 2005; 26:983–987. Grewal GS, Blake GM. Reference data for 51Cr-EDTA measurements of the glomerular filtration rate derived from live kidney donors. Nucl Med Commun 2005; 26:61–65. Vidal L, Shavit M, Fraser A, Paul M, Leibovici L. Systematic comparison of four sources of drug information regarding adjustment of dose for renal function. BMJ 2005; 331:263–266. Cosgriff PS, Fleming JS, Jarritt PH, Skyrpniuk J, Bailey D, Whalley D, et al. UK audit of GFR measurements. Nucl Med Commun 2002; 23:286. Manjunath G, Sarnak MJ, Levey AS. Estimating the glomerular filtration rate. Do’s and don’ts for assessing kidney function. Post Grad Med 2001; 110:55–62.

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NEWS AND VIEWS APRIL 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 206 6930, E-mail: [email protected]

Nuclear Medicine Communications, 2006, 27, 409–410 A change of direction for the society

At the start of January 2006, BNMS embarked on a wide ranging review of its purpose, aims and programmes. The ball was set in play by John Hall (council member) and perhaps falling membership was the initial spur. A strategic review meeting was organised, mediated by Community Mentors Ltd. Starting out from a core aim of promoting nuclear medicine to the benefit of patients and a set of values, council reviewed every facet of the society: The society’s profile The society’s structure K The forum it provides for the nuclear medicine community K Education and training K Its status as a reference source for official bodies K Its role in standards setting K Its role in promoting research and development. K K

Under each of these seven headings, the existing resources of the society were examined. An action plan was sketched out for the review of each of these functions. Over the ensuing months, these plans will be completed and implemented, so steering the society into a new direction more fit for the challenges ahead. The general feeling at this first step was that the society needs to raise its profile, and

change its structure, separating in some way the development of policy from the implementation of it. The forum did not come under such radical scrutiny because the annual meeting is reviewed in depth through many council meetings during the year, although there was a feeling that it would help the aims of the society if we could reach out more to members of the medical community beyond the nuclear medicine sphere. As far as the society’s standing as a reference source to official bodies goes, we have recently made advances in this direction thanks to the efforts of our current president, Andrew Hilson, and these should form a good foundation on which to build. Our role in standards setting was seen as perhaps one of our stronger points historically, with the clinical guidelines, organisational audit and gamma camera questionnaire, but the review will look into perhaps a wider range of such activities. On research and development, we recognise that, in an ideal world, we would have done more, although effective ideas on how to promote R&D were generally regarded as difficult to achieve financially. No doubt the emphasis in each of these functions will modify as the action plans develop, but this article seeks only to give a summary, a snapshot in time, in January 2006.

Meeting Announcements 2nd European IRPA Congress on Radiation Protection

Dates: 15–19 May 2006 Venue: Paris, France Website: www.irpa2006europe.com BNMS Autumn Meeting

Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk EANM Annual Meeting

Dates: 30 September to 4 October 2006 Venue: Athens, Greece Website: www.eanm.org 9th World Congress of Nuclear Medicine and Biology

Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/congress 2006/index02.htm Education and Training

EANM Learning Courses Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM Executive Secretariat on Tel.: + 43 1 2128030, fax: + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected]

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410 Nuclear Medicine Communications 2006, Vol 27 No 4

EANM Distance Learning in Nuclear Cardiology This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology

studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as

well as the most common clinical indications. http://www.eanm.org/eduOnline/ edu_start.php?navId = 332

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Erratum 411

Erratum The paper entitled ‘Efficiency comparison between 99mTc-tetrofosmin and 99mTc-sestamibi myocardial perfusion studies’ [1] contained four errors. In the abstract the number of repeated scans for 99mTcsestamibi for rest and stress studies was ‘‘16.4%’’ and not ‘‘19.7%’’, and the P value ‘‘0.001’’ instead of ‘‘0.1’’. In Table 2 the re-scan rate for 99mTc-sestamibi in the rest and stress study should be ‘‘16.4 (101/614)’’ and not

‘‘19.7 (101/514)’’, and the P value ‘‘0.001’’ instead of ‘‘0.01’’.

Reference 1

Ravizzini GC, Hanson MW, Shaw LK, Wong TZ, Hagge RJ, Pagnanelli RA, et al. Efficiency comparison between 99mTc-tetrofosmin and 99mTcsestamibi myocardial perfusion studies. Nucl Med Commun (2002); 23:203–208.

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Editorial

Is it time to incorporate quantitative functional imaging data, FDG PET in particular, into the response evaluation criteria in solid tumours? Sandip Basu and Narendra Nair Nuclear Medicine Communications 2006, 27:413–416

Tel: + 91 22 241 49428; fax: + 91 22 241 57098; e-mail: [email protected]

Radiation Medicine Centre (BARC), Tata Memorial Centre Annexe, Jerbai Wadia Road, Parel, Bombay 400 012, India.

Radiological assessment of ‘response-to-treatment’ in clinical trials in oncology by the WHO criteria has traditionally been based on the percentage change of the product of the longest diameter and that perpendicular to it (summed over all measurable lesions), from its baseline value. This was developed by the UICC and the WHO and published in the WHO handbook in 1979 [1]. With time, the WHO criteria faced criticisms; in 1992, the Southwest Oncology Group, in cooperation with the National Cancer Institute suggested a new set of guidelines [2] due to ‘uncertainties in clinical trials objectives, limitations in the resolution of imaging methods, and demands for greater rigor in response and endpoint definitions’. In 2000, a collaborative effort undertaken to unify and give more specificity and objectivity to response assessment criteria by the representatives of various research groups published the revised guidelines [3] in the Journal of National Cancer Institute, recommending one-dimensional measurements of the sum of the longest diameters of tumours instead of bi-dimensional tumour measurements by the WHO criteria. These guidelines are popularly known as RECIST (response evaluation criteria in solid tumours). The RECIST advocates computed tomography (CT) and magnetic resonance imaging (MRI) as the methods to measure target lesions selected for response assessment and spiral CT as the choice for tumours of the chest, abdomen and pelvis. The measurable lesion here has been defined as a lesion that can be accurately measured in at least one dimension with longest diameter > 20 mm using conventional techniques or > 10 mm with a spiral CT scan [3]. Non-measurable lesions include those with smaller dimensions than above, bony metastases, leptomeningeal disease, ascitis, pleural or pericardial effusions, inflammatory breast cancer, lymphangitis carcinomatosa and cystic or necrotic lesions. A major paradigm shift in oncological management is growing interest in appropriate early response assessment due to availability of a number of aggressive but potentially toxic treatment modalities. With accurate early response assessment, timely therapy modification can be accomplished and significant treatment-related

morbidity could be avoided. Reduction in tumour volume is only a late feature of effective therapy and several courses of cytotoxic therapy are usually needed before it can be determined whether the treatment is effective. This means that using size reduction as a criterion for early response may cause an unacceptable number of patients to be falsely labelled as poor responders, even if these cases have achieved an early complete metabolic response with the installed therapy. At the same time patients with non-responding tumours will be treated without benefit over a long period of time. As the functional changes are likely to precede anatomic response, positron emission tomography (PET) can play a major role in detecting early (non-) response. At times, the size change measurement by RECIST can also be misleading in the clinically inactive enlarging lesions following therapy like non-seminomatous germ cell tumours. It might be also the case when a responding tumour becomes more cystic, where there is an apparent increase in size. There is now convincing scientific evidence that 18F-fluorodeoxyglucose (18F-FDG) PET is sensitive and specific for determining therapy response in a wide variety of malignancies, viz. neoadjuvant chemoradiotherapy in head and neck squamous cell carcinoma [4,5] and oesophageal cancer [6,7], neoadjuvant chemotherapy locally advanced breast cancer (LABC) [8] and locally advanced non-small cell lung carcinoma having inoperable disease [9], response to preoperative radiation and 5-fluorouracil-based chemotherapy in colorectal cancer [10,11] and provides important prognostic information. Probably one of the most studied malignancies addressing this is lymphoma [12–15], where the performance of FDG PET in assessing chemoradiotherapy response was significantly better compared to CT so also its predictive value in prognostification or future prediction of relapse. 18F-FDG PET imaging has produced attractive results for monitoring treatment with imatinib mesylate, a relatively specific and powerful inhibitor of ABL tyrosine kinases (e.g., bcr-abl, c-abl), c-KIT and platelet-derived growth factor receptor and a marked decrease in tumour metabolic activity observed as early as 24 h after starting imatinib is now regarded as the sensitive early indication of treatment response in

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414 Nuclear Medicine Communications 2006, Vol 27 No 5

gastrointestinal stromal tumours (GISTs). Response to RECIST criteria by CT can take 4–6 months leading to an initial underestimation of the response rate [16,17]. These results can be extrapolated in assessing the efficacy of novel therapies (many of which are cytostatic rather than cytoreductive), where FDG PET may have important application as a phase II/III trial end point to accelerate evaluation and approval could facilitate drug development as an early surrogate of clinical benefit. According to the RECIST, only patients with measurable disease at baseline should be included in protocols. The measurable lesion here has been defined as one that can be accurately measured in at least one dimension with the longest diameter > 20 mm using conventional techniques or > 10 mm with a spiral CT scan [3]. The present dedicated PET systems with a spatial resolution of around 5–7 mm can be comfortably incorporated in the current definition. It is not infrequent to find in PET images extremely tiny but intensely hypermetabolic metastatic lesions, which may be more representative and preferred over a larger metabolically indolent one while selecting the ‘target’ lesions. With its increasing use, this ‘metabolic resolution’ in FDG PET can be foreseen as a distinct strength in ‘target lesion’ selection to overcome its so-called shortcoming of lower spatial resolution. The uptake of FDG has a definite edge over morphological imaging modalities in the evaluation of the ‘non-measurable’ lesions as there is the facility to carry out a whole-body evaluation in a single go. Imaging with meta-iodobenzylguanidine (MIBG), critical and well recognized for years in the management of neural crest tumours has not been included in the present guidelines. With the wide availability of 123IMIBG, excellent quality single photon emission computed tomography (SPECT) imaging is now possible in neural crest tumours, which makes it crucial for assessing treatment response in this group of tumours. Although there is an increasing number of studies indicating the potential usefulness and clinical application of FDG PET for assessment and prediction of treatment response, there are several issues on which consensus needs to be built: What is the appropriate timing of response measurement: which therapy and when to measure? Which is the optimal and clinically most appropriate approach for quantification of FDG uptake? Which thresholds are to be used to define response as complete metabolic response (CMR), Partial metabolic response (PMR), stable metabolic disease (SMD) and progressive metabolic disease (PMD)? How should it be used judiciously for varying uptake by different tumours or in heterogeneous tumours? So far, the European Organization for Research and Treatment

of Cancer (EORTC) PET Study Group recommendations have been the only firm step in this direction [18]. The EORTC PET study group held a consensus meeting in February 1998 and subsequently in March 1999 to make initial recommendations for a common measurement standard and criteria for reporting alterations in FDG uptake (based upon SUV) to assess clinical and subclinical response.

Selection of appropriate timing of posttreatment PET This has been a point of debate as confounding transient ‘metabolic flare’ has been observed in responding tumours following radiotherapy or hormonal therapy and, to a lesser extent, with chemotherapy. The increase in FDG uptake accompanying radiotherapy during the initial 2–3 weeks following therapy is prominent in epithelial surfaces and is related to inflammatory infiltrate of neutrophils, lymphocytes, and macrophages; the ‘flare’ with hormonal therapy is specific to the partial agonist effect of tamoxifen. This ‘metabolic flare’ is usually short-lasting in chemotherapy: the EORTC PET group suggested a time gap of 1–2 weeks between completion of the chemotherapy and the FDG PET, although they do not make any specific recommendation regarding the timing of follow-up FDG PET in case of radiotherapy. False negative 18F-FDG PET, which has been attributed to vascular damage, altered GLUT-I or hexokinase activity, regardless of tumour viability, has also been reported within 1 month of the post-radiotherapy period [19,20]. In general, at least an interval of 4 weeks (preferably 6–8 weeks) following radiotherapy is suggested to circumvent the misinterpretation. The minimum interval for re-evaluation after starting therapy, according to RECIST, is 4 weeks, where again, FDG PET can be comfortably applied.

Quantitative PET: The optimum method A number of methods have been used to assess tumour FDG uptake which can be broadly categorized into two groups: (1) visual interpretation and semiquantitative indices and (2) metabolic rates for 18F-FDG (MRglu) using kinetic modelling. The optimal trade-off between accuracy and simplicity is yet to be defined [4] in quantitative PET. While the standardized uptake value (SUV) is most commonly used in oncological studies, it is not clear whether this represents the best one. Common sources of error include a variable uptake period (time between injection and imaging), paravenous 18F-FDG injection, residual activity in the syringe and noncorrection of decay of injected activity while calculating SUV. Though good correlation with coefficients of 0.91 [21] and 0.84 [22] have been reported, correction of SUV by body surface area (SUVBSA) reduces the dependency on body weight and is proposed to be the minimum standard by the EORTC PET study group. Compared to

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Imaging in response evaluation of solid tumours Basu and Nair 415

SUV, MRglu, either by non-linear regression (NLR) analysis with a two-tissue compartment model and simplified tracer kinetic approaches, e.g. Patlak–Gjedde analysis (preferred by the EORTC group), is quantitative in nature and has less dependence on uptake time but here evaluable tumour lesions are limited to the specific bed position and additional scans are required for tumour staging. Taking all these into consideration, in a busy clinical setting, at present the SUV appears to be the most practical index for the purpose. To alleviate the problem of wide variability of FDG uptake in different malignancies it is recommended that the percent change in FDG uptake is used rather than the absolute values, which is important in the response stratification. In the common scenario, tumours are often heterogeneous with areas of necrosis interspersed; it is recommended that pretreatment regions of interest (ROIs) be drawn on the high 18F-FDG uptake region and posttreatment ROIs should be drawn as close as possible to these. The EORTC 1999 criteria stratified the tumour response into complete metabolic response, partial metabolic response, stable metabolic disease and progressive metabolic disease, based on the percent change in FDG uptake and percent visible change in the extent of 18F-FDG tumour uptake. These values and the cutoffs, however, require further prospective validation in larger (multicentre) studies. These should be dealt with by a task force that critically reviews and compares the various quantitative models and cut-off values. It is also important to recognize and keep abreast of the information concerning false positive accumulation of FDG (observed in a wide range of anatomical variants and physiological processes as well as in several non-malignant pathologies) to obviate diagnostic errors. The literature describing these continues to grow.

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PET-CT/SPECT-CT is becoming more and more available and unfortunately little thought is given as to how to integrate combined metabolic/anatomic information into response evaluation. There is a growing need to introduce a combined functional–anatomical assessment of tumour characteristics. Many newer tumour-specific PET tracers, currently under active investigation, have the potential to circumvent many of the shortcomings of FDG and more accurately measure the tumour clonogen density. Further studies are required before commenting on their clinical applicability. It is time for the oncology fraternity to take up a collaborative effort as a matter of urgency to integrate functional imaging (PET in particular) and thereby set up appropriate and rational guidelines for assessing treatment response.

References 1

WHO Handbook for Reporting Results of Cancer Treatment. WHO Publication No. 48. Geneva: World Health Organization, 1979.

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Green S, Weiss GR. Southwest Oncology Group standard response criteria, endpoint definitions and toxicity criteria. Invest New Drugs 1992; 10:239–253. Therasse P, Arbuck SG, Eisenhauer EA, Wanders J, Kaplan RS, Rubinstein L, et al. New guidelines to evaluate the response to treatment in solid tumors. J Natl Cancer Inst 2000; 92:205–216. Allal AS, Dulguerov P, Allaoua M, Haenggeli CA, El-Ghazi A, Lehmann W, Slosman DO. Standardized uptake value of 2-[(18)F] fluoro-2-deoxy-Dglucose in predicting outcome in head and neck carcinomas treated by radiotherapy with or without chemotherapy. J Clin Oncol 2002; 20:1398–1404. Dalsaso TA, Lowe VJ, Dunphy FR, Martin DS, Boyd JH, Stack BC. FDG PET and CT in evaluation of chemotherapy in advanced head and neck cancer. Clin Positron Imaging 2000; 3:1–5. Downey RJ, Akhurst T, Ilson D, Ginsberg R, Bains MS, Gonen M, et al. Whole body 18FDG PET and the response of esophageal cancer to induction therapy: results of a prospective trial. J Clin Oncol 2003; 21:428–432. Kato H, Kuwano H, Nakajima M, Miyazaki T, Yoshikawa M, Masuda N, et al. Usefulness of positron emission tomography for assessing the response to neoadjuvant chemoradiotherapy in patients with esophageal cancer. Am J Surg 2002; 184:279–283. Wahl RL, Zasadny K, Helvie M, Hutchins GD, Weber B, Cody R. Metabolic monitoring of breast cancer chemohormonotherapy using positron emission tomography:initial evaluation. J Clin Oncol 1993; 11: 2101–2111. Patz EF, Connolly J, Herndon J. Prognostic value of thoracic FDG PET imaging after treatment for non-small cell lung cancer. Am J Roentgenol 2000; 174:769–774. Guillem JG, Puig-La Calle Jr J, Akhurst T, Tickoo S, Ruo L, Minsky BD, et al. Prospective assessment of primary rectal cancer response to preoperative radiation and chemotherapy using 18-fluorodeoxyglucose positron emission tomography. Dis Colon Rectum 2000; 43:18–24. Haberkorn U, Strauss LG, Dimitrakopoulou A, Engenhart R, Oberdorfer F, Ostertag H, et al. PET studies for fluorodeoxyglucose metabolism in patients with recurrent colorectal tumors receiving radiotherapy. J Nucl Med 1991; 32:1485–1490. Romer W, Hanauske AR, Ziegler S, Thodtmann R, Weber W, Fuchs C, et al. Positron emission tomography in non-Hodgkin’s lymphoma: assessment of chemotherapy with fluorodeoxyglucose. Blood 1998; 91:4464–4471. Jerusalem G, Beguin Y, Fassotte MF, Najjar F, Paulus P, Rigo P, Fillet G. Whole-body positron emission tomography using 18F-fluorodeoxyglucose for post treatment evaluation in Hodgkin’s disease and non-Hodgkin’s lymphoma has higher diagnostic and prognostic value than classical computed tomography scanning. Blood 1999; 94:429–433. Mikhaeel NG, Timothy AR, O’Doherty MJ, Hain S, Maisey MN. 18-FDG-PET as a prognostic indicator in the treatment of aggressive non-Hodgkin’s lymphoma – comparison with CT. Leuk Lymphoma 2000; 39:543–553. Spaepen K, Stroobants S, Dupont P, Van Steenweghen S, Thomas J, Vandenberghe P, et al. Prognostic value of positron emission tomography (PET) with fluorine-18 fluorodeoxyglucose ([18F]FDG) after firstline chemotherapy in non-Hodgkin’s lymphoma; is [18F]FDG-PET a valid alternative to conventional diagnostic methods? J Clin Oncol 2001; 19:414–419. Van den Abbeele AD, Badawi RD. Use of positron emission tomography in oncology and its potential role to assess response to imatinib mesylate therapy in gastrointestinal stromal tumors (GISTs). Eur J Cancer 2002; 38(suppl 5):S60–S65. Stroobants S, Goeminne J, Seegers M, Dimitrijevic S, Dupont P, Nuyts J. 18FDG-Positron emission tomography for the early prediction of response in advanced soft tissue sarcoma treated with imatinib mesylate (Glivec). Eur J Cancer 2003; 39:2012–2020. Young H, Baum R, Cremerius U, Herholz K, Hoekstra O, Lammertsma AA, et al. Measurement of clinical and subclinical tumor response using [18F]fluorodeoxyglucose and positron emission tomography: review and 1999 EORTC recommendations. European Organization for Research and Treatment of Cancer (EORTC) PET Study Group. Eur J Cancer 1999; 35:1773–1782. Greven KM, Williams DW, Keyes Jr JW, McGuirt WF, Watson Jr NE, Randall ME, et al. Positron emission tomography of patients with head and neck carcinoma before and after high dose irradiation. Cancer 1994; 74:1355–1359. Rege S, Safa AA, Chaiken L, Hoh C, Juillard G, Withers HR. Positron emission tomography: an independent indicator of radiocurability in head and neck carcinomas. Am J Clin Oncol 2000; 23:164–169.

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21 Minn H, Leskinen-Kallio S, Lindholm P, Bergman J, Ruotsalainen U, Teras M, Haaparanta M. [18F]-fluorodeoxyglucose uptake in tumours: kinetic vs steady-state methods with reference to plasma insulin. J Comput Assist Tomogr 1993; 17:115–123.

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Original article

A new computer-based decision-support system for the interpretation of bone scans May Sadika, David Jakobssonb, Fredrik Olofssonb, Mattias Ohlssonb,c, Madis Suurkulaa and Lars Edenbrandta,b Objective To develop a completely automated method, based on image processing techniques and artificial neural networks, for the interpretation of bone scans regarding the presence or absence of metastases.

sensitivity of 90%. A false positive classification of metastases was made in 18 of the 69 patients not classified as having metastases by the experienced physician, resulting in a specificity of 74%.

Methods A total of 200 patients, all of whom had the diagnosis of breast or prostate cancer and had undergone bone scintigraphy, were studied retrospectively. Whole-body images, anterior and posterior, were obtained after injection of 99mTc-methylene diphosphonate. The study material was randomly divided into a training group and a test group, with 100 patients in each group. The training group was used in the process of developing the image analysis techniques and to train the artificial neural networks. The test group was used to evaluate the automated method. The image processing techniques included algorithms for segmentation of the head, chest, spine, pelvis and bladder, automatic thresholding and detection of hot spots. Fourteen features from each examination were used as input to artificial neural networks trained to classify the images. The interpretations by an experienced physician were used as the ‘gold standard’.

Conclusion A completely automated method can be used to detect metastases in bone scans. Future developments in this field may lead to clinically valuable decision-support c 2006 Lippincott tools. Nucl Med Commun 27:417–423 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:417–423 Keywords: computer-assisted diagnosis, radionuclide imaging, bone metastases, image processing, neural networks a

Departments of Clinical Physiology, Sahlgrenska University Hospital, Go¨teborg, Department of Clinical Physiology, Malmo¨ University Hospital and cDepartment of Theoretical Physics, Lund University, Sweden. b

Correspondence to May Sadik, Department of Clinical Physiology, Sahlgrenska University Hospital, SE-413 45 Go¨teborg, Sweden. Tel: + 0046 313 42141 6; fax: + 0046 314 11735; e-mail: [email protected] This study was supported by grants from the Swedish Medical Research Council.

Results The automated method correctly identified 28 of the 31 patients with metastases in the test group, i.e., a

Introduction Bone scintigraphy is a highly sensitive method for demonstrating bone metastases, and is an effective diagnostic tool for whole-body examinations. Patients with advanced prostate or breast cancer frequently suffer from bone metastases, and the extent of the disease significantly affects the overall survival and treatment of the patient [1–3]. The interpretation of bone scans is, however, a difficult pattern-recognition task and long experience is required. Non-neoplastic diseases can also reveal abnormalities in the images and a number of differential diagnosis and error sources should be considered [4]. Clinical decision-support systems can improve clinical practice in different medical fields, as has been shown in a recent systematic review of randomized controlled trials [5]. Artificial neural networks is an example of a technique that has been successfully applied to analysis of diagnostic images [6–10]. Lindahl et al. reported significantly improved classifications in myocardial perfu-

Received 4 January 2006 Revised 27 January 2006 Accepted 27 January 2006

sion scintigraphy by the use of a decision-support system [10]. These authors showed that even a very experienced physician can benefit from a computer-based decisionsupport system, expressed as decreased intra-observer and inter-observer variability. This type of decision support will not replace physicians, but will assist them by proposing an interpretation of the studies. Computer-assisted systems have also been applied to the analysis of bone scans. Semi-automated image segmentation programs that present a quantification of the extent of metastases have been developed [11,12]. The results presented by the programs have shown high correlation with corresponding visual or manually drawn region-ofinterest analysis. This type of program may be of value in clinical trials in order to improve the analysis, but they are probably too time-consuming for clinical use. The measuring time for the method presented by Noguchi and co-workers was 16 min per examination, on average [12]. The method presented by Erdi and co-workers

c 2006 Lippincott Williams & Wilkins 0143-3636

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418 Nuclear Medicine Communications 2006, Vol 27 No 5

requires the user to insert a seed point into each metastatic region on the image before the system can demarcate a region of interest. This is time-consuming, however, because patients with bone metastases usually have multiple disease sites [11]. Yin and Chiu developed a computer-aided diagnosis system for bone scans in order to provide physicians with second-reader information [13]. Their system showed a high detection rate, but the mean number of false positive detections was 37 in an abnormal image and 46 in a normal image. Their system performed better for hands and legs and worse for the head and vertebrae regions, where metastatic bone disease is mostly located. Decision-support systems with lower rates of false positive detections and completely automated analysis will most likely be required before physicians can adopt this type of technique in their clinical practice. The purpose of this study was to develop a completely automated method, based on image processing techniques and artificial neural networks, for the interpretation of bone scans regarding the presence or absence of metastases.

Materials and methods Patients

We retrospectively selected 200 consecutive patients with the diagnosis of breast or prostate cancer, who had undergone bone scintigraphy due to suspected metastatic disease during the periods January to June 1999 and January to February 2000 (Table 1). Patients examined with our dual detector gamma camera and with a complete set of technically sufficient images were included. Images with artefacts were excluded from the study (five patients). The total material was randomly divided into a training group and a test group, with 100 patients in each group. The training group was used in the process of developing the image analysis techniques and to train the artificial neural networks. The test group was used to evaluate the method. Bone scintigraphy

Bone scans were obtained approximately 3 h after intravenous injection of 600 MBq 99mTc-methylene diphosphonate (MDP) (Amersham, UK). Whole-body images, anterior and posterior (scan speed 10 cm
min – 1, matrix 256  1024), were recorded digitally with a lowenergy, high-resolution collimator (Maxxus; General Table 1

Study population

Number of patients Female (%) Mean age, years (range) Prevalence of bone metastases (%)

Training group

Test group

100 31 68 (40 – 88) 36

100 32 69 (36 – 89) 31

Electric, Milwaukie, USA) and stored on a computer system (Maxxus, Star Cam RMX, Milwaukie, USA). Energy discrimination was provided by a 15% window centred on the 140 keV of 99mTc. The ‘gold standard’

The ‘gold standard’ classification of the patients regarding the presence or absence of bone metastases was based on the clinical reports. Clinical information such as medical condition, the localization of bone pain and previous history of trauma were available to the reporting physicians. All reports and images were re-examined by a trained technologist and an experienced physician. They estimated the probability of bone metastases on an analogue scale from 0 to 1 on the basis of the clinical reports as well as the images. In order to present the network with the binary classification ‘bone metastases’ or ‘no bone metastases’, 0.5 was chosen as a cut-off value. All patients with values below 0.5 were considered as normal and values equal to or above 0.5 as pathological. This estimation was used as the gold standard. Decision-support system

The method for interpretation of bone scans consists of image processing techniques and artificial neural networks. The program is fed with the anterior and posterior images in digital format and no manual steps are required. The first steps are image segmentation, hot spot detection and feature extraction. The resulting image features are used as input to artificial neural networks, applied as classifiers for the detection of metastases. Image segmentation

Customized image processing algorithms for segmentation, which covers the entire body, were developed. The goal was to automatically delineate the spine (including the sternum in the anterior image), the ribs, the region of the kidneys, the forehead of the anterior image, the head for the posterior image and a region containing the pelvis and legs. The position of the neck and upper pelvic region are detected by calculating the vertical profiles of the image. This is achieved by thresholding the bones from the background into a binary image and summarizing the binary pixel values of each row. The vertical profile can be plotted as a curve or histogram, and has two characteristic dips in the positions of the neck and the upper pelvic region, which can be located automatically by the algorithms. The next step is to segment the spine and the ribs. By using a combination of image filtering and histogram analysis, the spine of the posterior image between the two lines of the neck and the upper pelvic region can be located. The algorithms use the spinal region to extract the ribs by using filtering, edge detection and morphological operations. The spinal region is also applied to the

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Decision-support system for the interpretation of bone scans Sadik et al. 419

anterior image and modified to fit the sternum. Finally, the lower half of the head in the anterior image and triangular regions below the ribs were delineated and hot spots in these regions were not considered in the neural network classifications. Detection of hot spots

The hot spots were located and segmented by a thresholding algorithm. The algorithm uses an optimized threshold value for each region after image segmentation. All hot spots under six pixels in size were excluded. A hot spot corresponding to the bladder, often visible in bone scans, was detected based on its size and position and was excluded from the interpretation. Symmetrical hot spots, likely to represent normal higher bone turnover or arthrosis, were detected based on distances to the spine, size and intensity. The remaining hot spots were used in the automatic interpretation of the images.

one input layer, one hidden layer and one output layer. There were 14 nodes in the input layer, one for each of the image features. The hidden layer contained 10 nodes. The output node encoded whether the image was classified as showing metastases or not. Each multi-layer perceptron was trained using gradient descent applied to a cross-entropy error function. To avoid over-training, a weight elimination [18] regularization term was utilized. The output of the neural network ensemble was computed as the mean of the output of the individual members of the ensemble. In this study, an ensemble size of 100 multi-layer perceptrons was used. All model selections, e.g., obtaining an optimal regularization parameter or finding the best architecture, were performed using a 5-fold cross-validation scheme on the training group. The neural network presented an output value between 0 and 1 for each test case. A threshold in this interval was used, above which all values were regarded as consistent with bone metastases.

Feature extraction

Fourteen features were used to describe the anterior and posterior images. The features were the result of the image processing part and the input to the artificial neural networks used for classification of the images. The features were: 1. Number of hot spots in the anterior image. 2. Number of hot spots in the posterior image. 3. Hot spot coverage of the anterior image, the area of the hot spots divided by the area of the body. 4. Hot spot coverage of the posterior image, the area of the hot spots divided by the area of the body. 5. Hot spot distribution of the anterior image, the number of regions where hot spots are present. 6. Hot spot distribution of the posterior image, the number of regions where hot spots are present. 7. Coefficient of variation (CV) for counts/pixel of the spine in the anterior image. 8. CV for counts/pixel of the spine in the posterior image. 9. CV for counts/pixel of the ribs in the anterior image. 10. CV for counts/pixel of the ribs in the posterior image. 11. Maximum divided by mean for counts/pixel of the spine in the anterior image. 12. Maximum divided by mean for counts/pixel of the spine in the posterior image. 13. Maximum divided by mean for counts/pixel of the ribs in the anterior image. 14. Maximum divided by mean for counts/pixel of the ribs in the posterior image. Artificial neural networks

Artificial neural networks were used as classifiers for the detection of metastases. A more general description of neural networks can be found elsewhere [14–16]. Each classifier consisted of an ensemble of single artificial neural networks. The individual members of the ensemble were standard multi-layer perceptrons [17] with

The study was approved by the Research Ethics Committee at Gothenburg University.

Results Thirty-one of the 100 patients in the test group had metastases, based on the probability estimate by the experienced physician (Fig. 1). In 24 of these 31 patients, the diagnosis was given with high confidence, expressed as a probability estimate of more than 95%. A probability estimate of less than 5% was given in 39 patients. Thirtyseven cases were given intermediate probability estimates, i.e., approximately two thirds of the cases were clear-cut cases and one-third of the cases were more difficult. The decision-support system made correct classifications in 28 of the 31 patients with metastases in the test group, i.e., a sensitivity of 90%. Figure 2 shows an example of one of these 28 patients. All of the 24 patients given a probability estimate of more than 95% were correctly classified by the system. The three patients with a false negative classification had probability estimates of 0.50, 0.50 and 0.68, respectively (Fig. 3). A false positive classification of metastases was made in 18 of the 69 patients not classified as having metastases by the experienced physician, which resulted in a specificity of 74% (Table 2). Six of these 18 cases had probability estimates of less than 5%. The decisionsupport system had difficulty in recognizing symmetrical hot spots (five patients) or classified the bladder as metastases (one patient) in these six patients. Most of the false classifications by the decision-support system were in the group of patients with intermediate probability values. Fifteen of these 37 patients were false positive or false negative. In contrast, only six of the 63

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420 Nuclear Medicine Communications 2006, Vol 27 No 5

cases with low ( < 5%) or high ( > 95%) probability estimates were falsely classified by the decision-support system.

Discussion Main findings

We have demonstrated that a completely automated method based on image processing techniques and artificial neural networks can be used to classify bone

Neural network output

Fig. 1

1.0 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0.0

0

50 Probability estimates

100

Probability estimates used as the ‘gold standard’ and neural network output for the test group (n = 100). The solid line represents the cut-off value, above which all values were regarded as consistent with bone metastases. The broken line represents the gold standard threshold used to discriminate between the presence or absence of metastases. False positive cases are in the upper left quadrant and false negative cases are in the lower right corner.

scans regarding the presence or absence of metastases. Our results show a high detection rate with a sensitivity of 90% in the test group. All patients given a probability estimate of more than 95% were correctly classified by the system. Yin and Chiu [13] presented a system for computer-aided diagnosis with sensitivity similar to that of ours (91.5%). Their system reported a total of 1252 false positive lesions in 27 normal images, i.e., an average of 46 false positive marks per normal image. Their system was used to provide warning marks to direct the physician’s attention to these locations, and not to provide a second opinion regarding the interpretation of the complete image as in our system. Their system performed better in the extremities than in the axial areas, where metastatic bone disease is most often located. Our system was able to correctly identify 74% of the images without metastases and 85% of the images with a probability estimate of less than 5%. The problem of low specificity seems to be much less with our system than with that of Yin and Chiu. Our system made false positive classifications in cases with high and symmetric radiotracer uptake, accumulating in the joints or vertebrae regions, due to arthrosis and spondylosis (Fig. 4). The network also had difficulty in finding symmetry in patients with severe scoliosis or subjects not lying straight, i.e., at different distances from the detectors. One problem with bone scintigraphy is its inability to discriminate between metastatic disease and/ or fracture, and complementary examinations such as standard radiography, computed tomography scanning,

Fig. 2

Patient with multiple hot spots correctly classified by neural networks. Red indicates metastases, green shows symmetrical hot spots, and the bladder is in yellow.

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Decision-support system for the interpretation of bone scans Sadik et al. 421

Fig. 3

A false negative classification made by artificial neural network in a patient given an intermediate probability estimate of having metastases. The interpretation program does flag for suspicious hot spots (arrows) though.

Table 2

False positive cases in the test group

Causes of misclassifications Symmetrical hot spots Fracture or/and metastases Spondylosis Sternectomi Uribag misclassified as metastases Urine catheter Bladder classified as metastasis Total false positive cases

Number of patients 7 4 3 1 1 1 1 18

magnetic resonance imaging or biopsy must occasionally be used to give the correct diagnosis. Decision support systems such as ours will have a problem in classifying such images correctly. Clinical implications

To our knowledge, our decision-support system is the first completely automated method for clinical interpretation of bone scans. Previous methods have been semi-automated and used for quantification of the extent of bone metastases [11,12], or used to indicate

locations on the images in order to direct the physician’s attention towards these locations – and not for an interpretation of the examination [13]. Our purpose has been to develop a method that would provide the physician with a second opinion for the bone scans and not to decide treatment strategy, which of course must be based on additional information. Currently, interpretation of bone scans is a subjective task based on the experience of the physician. A decision-support system of high quality could be a valuable tool to improve the quality of the medical reports in terms of reduced inter-observer and intra-observer variability, and to support less experienced physicians. The aim is not to replace the physician with a computer but to take advantage of the technique. Other applications for the automated decision-support system could be to highlight suspicious uptake for technologists during the acquisition of the images so they could consider obtaining additional views. A previous study has shown that even a very experienced physician will benefit from the advice of artificial neural networks in the classification of myocardial perfusion images [10]. The automated

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422 Nuclear Medicine Communications 2006, Vol 27 No 5

Fig. 4

Artificial neural network misclassifying the bladder as metastasis in the anterior image and not recognizing the symmetry in the shoulders due to severe scoliosis.

method presented in this study is fast, easy to use and does not require any manual steps. The current study is the first step towards an automated decision-support system for the interpretation of bone scans. Further refinements are needed before applying it in the clinical setting.

field may lead to clinically valuable decision-support tools.

References 1

Study limitations

2

The gold standard used in this study was based on the probability of bone metastases estimated by an experienced physician. The differences between our gold standard and the absolute truth regarding presence of metastasis must be recognized. Approximately one third of the cases were not clear cut, but were more or less difficult cases, expressed as probability estimates between 5 and 95%. It is important to include difficult cases in the training and evaluation of the neural network, i.e., cases that can be found in the clinical setting in which the decision-support system is intended to be used. The accuracy of the gold standard is important, however, both in the development phase and in the evaluation. Consensus classifications based on more than one experienced physician or independent gold standard examinations of the patients might be of value. However, in this phase of the development of an automated method, we felt that this type of validation was good enough. With this gold standard method, we could concentrate on including a relatively large patient group, which is important for the training of neural networks.

3 4

5

6 7

8

9

10

11

12

Conclusion A completely automated method can be used to detect metastases in bone scans. Future developments in this

13

Maffioli L, Florimonte L, Pagani L, Butti I, Roca I. Current role of bone scan with phosphonates in the follow-up of breast cancer. Eur J Nucl Med Mol Imaging 2004; 31:143–148. Coleman R, Heidenreich A, Bell R. Managing metastatic bone disease: three case studies. Semin Oncol 2004; 31:83–86. Coleman RE. Metastatic bone disease: clinical features, pathophysiology and treatment strategies. Cancer Treat Rev 2001; 27:165–176. Bombardieri E, Aktolun C, Baum RP, Bishof-Delaloye A, Buscombe J, Chatal JF, et al. Bone scintigraphy procedures guidelines for tumour imaging. Eur J Nucl Med Mol Imaging 2003; 30:107–114. Kawamoto K, Houlihan CA, Balas EA, Lobach DF. Improving clinical practice using clinical decision support systems: a systemic review of trials to identify features critical to success. BMJ 2005; 330:765–768. Mango LJ. Computer-assisted cervical cancer screening using neural networks. Cancer Letts 1994; 77:155–162. Freer TW, Ulissey MJ. Screening mammography with computer-aided detection: prospective study of 12,860 patients in a community breast center. Radiology 2001; 220:781–786. ¨ hlin H, Edenbrandt L. Neural networks – a Olsson S-E, Ohlsson M, O diagnostic tool in acute myocardial infarction with concomitant left bundle branch block. Clin Physiol Func Imag 2002; 22:295–299. Nieminen P, Hakama M, Viikki M, Tarkkanen J, Anttila A. Prospective and randomised public-health trial on neural network-assisted screening for cervical cancer in Finland: results of the first year. Int J Cancer 2003; 103:422–426. Lindahl D, Lanke J, Lundin A, Palmer J, Edenbrandt L. Improved classifications of myocardial bull’s-eye scintigrams with computer-based decision support systems. J Nucl Med 1999; 40:96–101. Erdi YE, Humm JL, Imbriaco M, Yeung H, Larson SM. Quantitative bone metastases analysis based on image segmentation. J Nucl Med 1997; 38:1401–1406. Noguchi M, Kikuchi H, Ishibashi M, Noda S. Percentage of positive area of bone metastasis is an independent predictor of disease death in advanced prostate cancer. Br J Cancer 2003; 88:195–201. Yin T-K, Chiu N-T. A computer-aided diagnosis for locating abnormalities in bone scintigraphy by a fuzzy system with a three-step minimization approach. IEEE Trans Med Imag 2004; 23:639–654.

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Decision-support system for the interpretation of bone scans Sadik et al. 423

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Cross SS, Harrison RF, Kennedy LR. Introduction to neural networks. Lancet 1995; 346:1075–1079. Baxt WG. Application of artificial neural networks to clinical medicine. Lancet 1995; 346:1135–1138. Dybowski R, Gant V. Artificial neural networks in pathology and medical laboratories. Lancet 1995; 346:1203–1207.

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Rumelhart DE, McClelland JL (editors). Parallel Distributed Processing, vols 1 and 2. Cambridge, Massachusetts: MIT Press; 1986. 18 Hanson SJ, Pratt LY. Comparing biases for minimal network construction with backpropagation. In: Touretzky DS (editor): Advances in Neural Information Processing Systems. San Meteo, California: Morgan Kaufmann; 1988, pp. 177–185.

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Original article

Left ventricular ejection fraction and volumes on rest gated 201 Tl perfusion SPECT: Comparison with two-dimensional echocardiography Chetan D. Patela, Murali R. Nadiga, Sumodh Kurienb, Sukanta Baraia, Rajeev Narangb and Arun Malhotraa Background Rest gated 201Tl images are considered to be of poor count statistics due to lower energy and low photon flux of 201Tl in addition to increased attenuation and low dose that can be administered. We compared the left ventricular ejection fraction (LVEF), end diastolic (EDV) and end systolic volume (ESV) obtained on 4 h gated rest 201Tl myocardial perfusion single photon emission computed tomography (SPECT) with those obtained by two-dimensional echocardiography (2-D ECHO) in patients with known or suspected coronary artery disease (CAD). Methods Eighty-two consecutive patients who underwent gated 201Tl stress–rest myocardial perfusion SPECT and 2-D ECHO were studied. The gated thallium images were processed with Siemens e-soft autocardiac processor and LVEF, EDV and ESV were evaluated using Emory Cardiac Toolbox. The same parameters were also assessed on the 2-D ECHO using the modified Simpson method for comparison. Results Out of 82 rest gated images, one study was excluded because of poor count statistics. In 81 (99%) patients there was good linear correlation with 2-D ECHO values and rest gated 201Tl SPECT images for EDV, ESV

Introduction Gated myocardial perfusion single photon emission tomography (gSPECT) has been used extensively for assessing left ventricular function in patients with coronary artery disease (CAD). Most of these studies have been done using 99mTc-based myocardial perfusion tracers [1–4]. The low-energy photons of 201Tl are susceptible to more scatter and attenuation within the body especially in patients with larger body habitus. Further, the low energy and lower photon flux from 201 Tl is considered to result in poor quality images [5,6]. However, several studies have described the feasibility of 201Tl gSPECT to assess left ventricular function [7,8]. The left ventricular functional parameters, such as ejection fraction (LVEF), end diastolic volume (EDV) and end systolic volume (ESV), derived from 99mTclabelled tracers reflect the function at a resting state as even the post-stress images are usually acquired about 30–45 min after peak stress. The thallium post-stress images are acquired within about 10 min and depict the left ventricular functional parameters that are different

and LVEF. Pearson’s correlation co-efficient (r value) for EDV, ESV and LVEF between the two methods was 0.78, 0.79 and 0.88, respectively. A Bland–Altman plot showed close agreement with LVEF but not for EDV and ESV. Conclusion These results suggest that the 4 h rest gated Tl study gives a reliable value for the LVEF compared to 2-D ECHO and can be used in routine clinical practice. c 2006 Lippincott Williams Nucl Med Commun 27:425–429 & Wilkins. 201

Nuclear Medicine Communications 2006, 27:425–429 Keywords: rest gated 201Tl SPECT, left ventricular ejection fraction and volumes, two-dimensional echocardiography Departments of aNuclear Medicine and bCardiology, All India Institute of Medical Sciences, New Delhi, India. Correspondence to Dr Chetan D. Patel, B-54, South Extension Part-I, New Delhi 110049, India. Tel: + 91-11-26594761; fax: + 91-11-26588641; e-mail: [email protected] Received 18 November 2005 Revised 06 February 2006 Accepted 13 February 2006

from the rest images at 4 h. Therefore it may also be difficult to compare gSPECT parameters like LVEF, EDV and ESV in the immediate post-stress state with other modalities like two-dimensional echocardiography (2-D ECHO) and radionuclide ventriculography which are usually done in the resting state. Gating of the 4-h redistribution images provides the same parameters at rest. However, the washout of thallium on the 4 h rest images is believed to further deteriorate the quality when compared to those of the immediate post-stress 201Tl images and 99mTc-based compounds. The aim of this study was to examine gated 4 h rest 201Tl images, which constitute the worst-case scenario among all gated images and compare the LVEF, EDV and ESV with 2-D ECHO derived values. Patients, materials and methods

Eighty-two consecutive patients (57 men and 25 women) of known or suspected CAD referred for stress–rest gated 201 Tl SPECT imaging were studied. The patients’ mean age was 63 ± 14 years (range 32–68 years). All patients

c 2006 Lippincott Williams & Wilkins 0143-3636

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426 Nuclear Medicine Communications 2006, Vol 27 No 5

had 2-D ECHO within a week of the gSPECT study. Forty patients had a history of myocardial infarction, 20 patients had previous revascularization (six had coronary artery bypass grafting (CABG) and 14 had percutaneous coronary angioplasty (PTCA)).

using the Emory Cardiac Tool Box (ECTB) application (version 1.1). The global left ventricular functional parameters for rest-gated studies were obtained using the automated algorithm in ECTB to calculate the LVEF, EDV and ESV.

Informed consent was obtained from all the patients recruited in the study group.

Two-dimensional echocardiography

Exclusion criteria were acute myocardial infarction or unstable angina occurring less than 2 weeks before the study and PTCA or CABG performed within 30 days of the study or any change in the clinical status between acquisition of the gSPECT and 2-D ECHO study. Patients with significant arrhythmia that compromised the gating technique were also excluded. Gated

201

Tl SPECT

Gated 201Tl myocardial perfusion SPECT was performed on a dual-head gamma camera system (Siemens E-CAM, Siemens Medical Systems, Germany). All patients fasted overnight and underwent stress–rest gated 201Tl myocardial perfusion SPECT. Sixty patients had treadmill exercise and 22 were subjected to pharmacological stress using dobutamine. At peak stress, 111–130 MBq (3–3.5 mCi) 201Tl, which is within the acceptable limits in India, was administered intravenously. Gated 201T1 post-stress SPECT imaging was initiated within 10 min after tracer administration using a 30% window centred over the 68–80 keV photopeak and a 20% window over the 167 keV peak. Gating was performed with eight frames per cardiac cycle, using a 100% beat acceptance window. All studies were acquired with a low-energy, all-purpose collimator. Studies were acquired using stepand-shoot mode with the heads at an angle of 901 to each other and an elliptical orbit. Sixty-four projections (32 steps, 2.81 per step) of 25 s per projection were acquired over 1801, from 451 RAO (right anterior oblique position) to 1351 LPO (left posterior oblique position). Rest gated 201 T1 SPECT studies were performed 4 h later with the same acquisition parameters. SPECT image processing

SPECT non-gated projection images were reviewed in cine mode in all patients to assess patient movement and any source of potential attenuation artifacts. The raw images (both gated and nongated datasets) were then pre-filtered with a Butterworth filter (order = 5.0, critical frequency = 0.45 cycle/cm), as determined previously by cardiac phantom studies and also the default vendor provided setting. The resulting transaxial image slices were then reoriented to generate short axis, vertical long axis and horizontal long axis images, using vendor provided software (Seimens, Autocardiac). These were analysed

All the patients underwent 2-D ECHO within a week of the 201Tl gSPECT study for the assessment of the same left ventricular functional parameters. The 2-D ECHO was performed at rest with standard parasternal and apical views. Left ventricular volumes and LVEFs were computed by the modified Simpson’s method. Two nuclear medicine physicians independently examined the gated 4 h rest SPECT 201Tl data for quality of images and evaluation of the LVEF, EDV and ESV from the acceptable rest gated images. One cardiologist performed the 2-D ECHO blinded to the results of the gSPECT.

Results In one patient, evaluation was not possible because of poor count statistics in the rest images. In the remaining 81 (99%) patients 2-D ECHO and gSPECT derived values were as given in Table 1. Pearson’s correlation coefficient (r value) for EDV, ESV and LVEF between 2-D ECHO values and gated SPECT images were 0.78, 0.79 and 0.88, respectively, with the linear regression equation to calculate LVEF by 2-D ECHO (y) from the value obtained by gSPECT (x) being y = 0.49x + 21. The Bland–Altman plot showed close agreement between LVEFs calculated by the two methods. However, good agreement was not observed for EDV and ESV (Figs 1–3).

Discussion Gated myocardial perfusion SPECT is unique in that it allows assessment of left ventricular perfusion and function simultaneously. In this study we tried to validate the LVEF, EDV and ESV derived from 4 h rest gated 201Tl SPECT by comparing the same parameters on 2-D ECHO. We did not take into consideration the post-stress left ventricular functional parameters because the intention of the study was to evaluate the rest gated 201Tl images as they are considered to be of poor count statistics and evaluation of functional parameters is difficult. In our study we found poor quality images in only one patient. Even though we included consecutive Mean ± SD values derived by two-dimensional echocardiography (2-D ECHO) and rest gated 201Tl SPECT

Table 1

Technique

EDV (ml)

ESV (ml)

LVEF (%)

2-D ECHO Rest gated SPECT

116 ± 33 109 ± 84

60 ± 35 61 ± 74

52 ± 14 60 ± 14

EDV, end diastolic volume; ESV, end systolic volume; LVEF, left ventricular ejection fraction.

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Comparison of LVEF and volumes on rest gated

Fig. 1

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Tl with 2-D ECHO Patel et al. 427

Fig. 3

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Average Results of a comparison of end diastolic volumes (EDVs) derived by two-dimensional echocardiography (2-D ECHO) and by rest gated 201 Tl SPECT displayed as a Bland–Altman plot. Difference = difference between EDV measured by the two methods. (Mean ± SD = 6.95 ± 58.5.) Average = mean of EDV measured by the two methods.

− 0.4 0.1

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Results of a comparison of ejection fractions (EFs) derived by twodimensional echocardiography (2-D ECHO) and by rest gated 201Tl SPECT displayed as a Bland–Altman plot. Difference = difference between EF measured by the two methods. (Mean ± SD = 0.09 ± 0.14.) Average = mean of EF measured by the two methods.

of patients in our study ranged from 54 to 97 kg with a mean of 64.5 kg. We did not correlate the perfusion abnormalities of each patient with the left ventricular functional parameters, because the idea behind the study was to validate the rest gated LVEF with 2-D ECHO in the entire spectrum of perfusion abnormalities. In our study, the LVEF values determined by gSPECT 201Tl were higher compared to the 2-D ECHO values especially in the normal range. This is usually seen in patients with small hearts and is well reported [9–11].

Fig. 2

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0

Although echocardiography is an operator-dependent modality, we wanted to compare LVEF and volumes derived from gSPECT with 2-D ECHO, which is widely used in routine practice in the management of these patients.

− 100

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Average Results of a comparison of end systolic volumes (ESVs) derived by twodimensional echocardiography (2-D ECHO) and by rest gated 201Tl SPECT displayed as a Bland–Altman plot. Difference = difference between ESV measured by the two methods. (Mean ± SD = 2.04 ± 49.63.) Average = mean of ESV measured by the two methods.

patients referred to myocardial perfusion studies without selective inclusion of patients with low body weight; the average body weight in our population is less compared to a Western population and this could be one possible reason for relatively better quality of images. The weight

Wright et al. [12] assessed the limitations of quantitative gSPECT myocardial perfusion imaging with 201Tl and compared LVEF from the 201Tl gSPECT with radionuclide ventriculography. These authors concluded that LVEF values derived from gSPECT studies were inadequate for use in clinical practice. However, these authors assessed gSPECT imaging using lower activities of 201Tl (mean injected activity of 62 ± 7 MBq) as compared to the dose administered in our study. Moralidis et al. [13] compared the left ventricular function derived from gSPECT with radionuclide ventriculography and suggested that the left ventricular function may be misinterpreted in a significant proportion of patients if the calculation of LVEF is based on 201Tl gSPECT.

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428 Nuclear Medicine Communications 2006, Vol 27 No 5

Several studies have shown the feasibility of calculating LVEF on gated thallium images. Manoury et al. [8] determined if myocardial gated SPECT with 201Tl was possible and reliable to quantify left ventricular function. The authors found good correlation between post-stress gated 201Tl and rest gated 99mTc sestamibi (r = 0.93 for LVEF, 0.92 for EDV and 0.96 for ESV) and concluded that myocardial gated SPECT quantification of left ventricular function with 201Tl was possible and as reliable as gated SPECT with 99mTc sestamibi. Matsuo et al. [14] assessed both the post-stress and rest LVEF, comparing the decrease in post-stress LVEF with the degree of ischaemia for prognostication of patients with CAD, although the authors did not validate the stress and rest LVEF on the gated 201Tl study with any other modality. Toba et al. [15] compared LVEF, EDV and ESV calculated by the QGS program from 201Tl SPECT study at rest, and estimated its accuracy by comparing it with echocardiography. Although 72.0% of 201Tl ECG-gated SPECT images were fair or poor in image quality, there were good correlations between the values obtained by the QGS program and echocardiography (LVEDV: r = 0.82; LVESV: r = 0.88, LVEF: r = 0.89). Cwajg et al. [16] in their study concluded that quantitative gated tomography, using either 201Tl or 99mTc tracers, has a good correlation ( Z 0.68) with echocardiography for the assessment of left ventricular volumes and ejection fraction. BacherStier et al. [17] also obtained good correlation for LVEF between post-stress and rest LVEF as assessed by gSPECT and echocardiography (r = 0.76 and 0.86, respectively). However, in all these studies the authors have only used Pearson’s correlation coefficient (the r value) for comparing the two modalities. In our study, we have also found good correlation with all the three parameters although the Bland–Altman plot showed good agreement only with respect to the LVEF. The Bland– Altman plot for continuous variables gives better information about agreement between the values derived by two different modalities than just the correlation coefficient. Our results are comparable with the study by Itti et al. [18] who correlated rest and rest–redistribution LVEFs from gSPECT with radionuclide ventriculography. They obtained even higher correlation coefficients (r = 0.92) on 4 h a rest–redistribution study, with good agreement on the Bland–Altman plot. In our study the correlation between 2-D ECHO and gSPECT is good to excellent for EDV, ESV and LVEF in gated 4 h rest studies when compared to 2-D ECHO, but the Bland–Altman plot shows good agreement with the LVEF parameter only and not with EDV and ESV. There appears to be a systematic error between the measurements of EDV and ESV by the two methods. Since this error is much higher at higher values of EDV and ESV, it

may be due to low count statistics. This affect is manifested when individual values of EDV and ESV are considered. However, it is likely to affect the estimation of both EDV and ESV to an equal extent and therefore the LVEF value may not be affected. This study suggests that gated 4 h rest 201Tl images can be used to quantify LVEF reliably, although the EDV and ESV values cannot be assessed accurately.

Conclusion Our results suggest that the gated 4 h rest 201Tl images can be used to obtain values of left ventricular ejection fraction. Further, the LVEF values derived from rest gated SPECT can be compared with post-stress LVEF to determine whether there is any stress-induced left ventricular dysfunction, which is a valuable tool for prognostication of CAD patients.

References 1

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Travin MI, Heller GV, Johnson LL, Katten D, Ahlberg AW, Isasi CR, et al. The prognostic value of ECG-gated SPECT imaging in patients undergoing stress Tc-99m sestamibi myocardial perfusion imaging. J Nucl Cardiol 2004; 11:253–262. Ban K, Nakajima T, Iseki H, Abe S, Handa S, Suzuki Y. Evaluation of global and regional left ventricular function obtained by quantitative gated SPECT using 99mTc tetrofosmin for left ventricular dysfunction. Intern Med 2000; 39:612–617. Yoshioka J, Hasegawa S, Yamaguchi H, Tokita N, Paul AK, Xiuli M, et al. Left ventricular volumes and ejection fraction calculated from quantitative electrocardiographic-gated 99mTc-tetrofosmin myocardial SPECT. J Nucl Med 1999; 40:1693–1698. Yang KT, Chen HD. Evaluation of global and regional left ventricular function using technetium-99m sestamibi ECG-gated single-photon emission tomography. Eur J Nucl Med 1998; 25:515–521. DePuey EG, Parmett S, Ghesani M, Rozanski A, Nichols K, Salensky H. Comparison of Tc-99m sestamibi and Tl-201 gated perfusion SPECT. Nucl Cardiol 1999; 6:278–285. Hyun IY, Kwan J, Park KS, Lee WH. Reproducibility of Tl-201 and Tc-99m Sestamibi gated myocardial perfusion SPECT measurement of myocardial function. J Nucl Cardiol 2001; 8:182–187. Manrique A, Faraggi M, Eltchaninoff H, Cribier A, Le Guludec D, Vera P. Evaluation of the ejection fraction tomoscintigraphy synchronised with an electrocardiogram. Arch Mal Coeur Vaiss 2001; 94:118–123. Manoury C, Chen CC, Chua KA, Thomson CJ. Quantification of left ventricular function with thallium-201 and technetium-99m sestamibi myocardial gated SPECT. J Nucl Med 1997; 38:958–961. Nakajima K, Higuchi T, Taki J, Kawano M, Tonami N. Accuracy of ventricular volume and ejection fraction measured by gated myocardial SPECT: comparison of 4 software programs. J Nucl Med 2001; 42:1571–1578. Ababneh AA, Sciacca RR, Kim B, Bergmann SR. Normal limits for left ventricular ejection fraction and volumes estimated with gated myocardial perfusion imaging in patients with normal exercise test results: influence of tracer, gender, and acquisition camera. J Nucl Cardiol 2000; 7: 661–668. Hambye AS, Vervaet A, Dobbeleir A. Variability of left ventricular ejection fraction and volumes with quantitative gated SPECT: influence of algorithm, pixel size and reconstruction parameters in small and normal-sized hearts. Eur J Nucl Med Mol Imaging 2004; 31:1606–1613. Wright GA, McDade M, Keeble W, Martin W, Hutton I. Quantitative gated SPECT myocardial perfusion imaging with 201Tl: an assessment of the limitations. Nucl Med Commun 2000; 21:1147–1151. Moralidis E, Spyridonidis T, Arsos G, Apostolopoulos D, Karatzas N, Vassilakos P, Karakatsanis K. 201Tl gated single photon emission computed tomographic myocardial perfusion imaging in the assessment of global and regional left ventricular function. Would it be favoured over equilibrium radionuclide angiography? Nucl Med Commun 2004; 25:665–673. Matsuo S, Matsumoto T, Nakae I, Koh T, Masuda D, Takada M, et al. Prognostic value of ECG-gated thallium-201 single-photon emission

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tomography in patients with coronary artery disease. Ann Nucl Med 2004; 18:617–622. Toba M, Ishida Y, Fukuchi K, Fukushima K, Katafuchi T, Hayashida K, Oka H, et al. Assessment of left ventricular function by 201Tl ECG-gated myocardial SPECT. Kaku Igaku 1999; 36:23–30. Cwajg E, Cwajg J, He ZX, Hwang WS, Keng F, Nagwh SF, Verani MS. Gated myocardial perfusion tomography for the assessment of left ventricular function and volumes: comparison with echocardiography. J Nucl Med 1999; 49:1857–1865.

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Bacher-Stier C, Muller S, Pachinger O, Strolz S, Erles H, Moncayo R, Venger M. Thallium-201 gated single photon emission tomography for the assessment of left ventricular ejection fraction and regional wall motion abnormalities in comparison with two-dimensional echocardiography. Eur J Nucl Med 1999; 26:1533–1540. 18 Itti E, Rosso J, Damien P, Auffret M, Thirion JP, Meignan M. Assessment of ejection fraction with Tl-201 gated SPECT in myocardial infarction: Precision in a restredistribution study and accuracy versus planar angiography. J Nucl Cardiol 2001; 8:31–39.

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Original article

Comparison of gastric emptying scintigraphy based on the geometric mean of the gastric proportion of the abdominal radioactivity or on the geometric mean of the intragastric radioactivity Pierre-Yves Salauna, Sole`ne Querelloua, Jean-Michel Nguyenb, Caroline Bodet-Milinc, Thomas Carlierc, Alexandre Turzoa, Yves Bizaisa and Olivier Couturierc Purpose Using gastric emptying scintigraphy the gastric retention rate is commonly calculated within a gastric region of interest (intragastric method). This technique may have significant limitations when left oblique anterior views are acquired, due in part to attenuation resulting from intragastric redistribution. To minimize these drawbacks, it was proposed to express the intragastric content as a percentage of the abdominal radioactivity (abdominal method). Our goal was to compare these two methods when anterior–posterior scanning is used. Methods Antero-posterior scintigraphic data of 272 consecutive patients were analysed by both methods. Retention rates were obtained by both observation and calculation by power exponential fit. Gastric emptying parameters (half-emptying time of solids (T50,S) and liquids (T50,L), lag phase (Tlag) time and real emptying time (TRE)), and quality of fit were also computed and compared. Results For solids, the intragastric method resulted in weakly higher experimental retention rates, whereas retention rates were quite similar for liquids. Differences between experimental and calculated retention rates were smaller for abdominal method, for both liquids and solids. As a result, values for the quality of fit were higher for the

Introduction Gastric scintigraphic procedures are time consuming, and many centres use to perform gastric emptying tests on a single-detector gamma camera, whereas dual-detector gamma cameras are reserved for whole-body scans or tomographic applications. For the determination of gastric emptying by using scintigraphy, the most common procedure involves antero-posterior acquisitions, followed by radioactivity measurements inside a gastric region of interest (intragastric method), physical decay corrections and computation of the geometric mean of each set of antero-posterior data [1,2]. Indeed, using the geometric mean as a tissue attenuation correction factor is essential because the distribution of the meal changes during the

abdominal method. Significant differences were observed only for calculated T50,S (122 ± 46 min vs. 124 ± 48 min, mean difference 2 ± 2 min, P < 0.00001) and TRE (163 ± 64 min vs. 168 ± 68 min, mean difference 4.5 ± 3.8 min, P < 0.05), respectively, for the abdominal and the intragastric methods. However, the Bland–Altman statistical method revealed good agreements ( < 5% outliers). Conclusion Intragastric and abdominal methods can be used indifferently to treat antero-posterior data of gastric c 2006 scintigraphy. Nucl Med Commun 27:431–437
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:431–437 Keywords: gastric emptying, scintigraphy, intragastric method, abdominal method Departments of aNuclear Medicine, University of Brest, bBiostatistics and c Nuclear Medicine, University of Nantes, France. Correspondence to Dr O. Couturier, Department of Nuclear Medicine, Hospital Hoˆtel Dieu, University of Nantes, Place A. Ricordeau, 44093 Nantes, France. Tel: + 0033 2 4008 4136; fax: + 0033 2 4008 4218; e-mail: [email protected] Received 30 December 2005 Revised 13 February 2006 Accepted 13 February 2006

gastric emptying process; first posterior (i.e. in the fundus) after ingestion, the meal moves thereafter anteriorly toward the antrum. As a result, at the beginning of the scintigraphic test, the attenuation on anterior views is emphasized whereas it is less important at the end of the test. This procedure is still considered to be the ‘gold standard’, using either a single-detector or a dual-head gamma camera [1,2]. Left anterior oblique scanning (LAO) is a possible alternative with a single detector, on which attenuation is nearly constant. This method requires the correct repositioning of the subject throughout the whole procedure, to avoid its major drawback, which is the lack of spatial reproducibility between collimator and stomach. However, even with

c 2006 Lippincott Williams & Wilkins 0143-3636


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skin markers, it can be difficult to seat the patient in the same position at each time point, particularly when a patient has undergone a lengthy gastric test [3,4]. Furthermore, good repositioning is important if a second gastric test needs to be performed in the attempt to evaluate the effectiveness of prokinetic agents. These spatial changes may cause technical errors, which can be responsible for poor estimations of gastric emptying or a gastric emptying pattern that is too irregular for accurate curve fitting. In a study with 45 subjects, Lien et al. [5] proposed an original approach to minimize the technical drawbacks of this left anterior oblique approach, by expressing the intragastric counts as a proportion of the total abdominal radioactivity. These authors concluded that this new abdominal method provided some advantages over the classic intragastric method, especially in smoothing the curves, thus increasing the quality of mathematical fits and the determination of gastric emptying parameters. In the present study, we aimed to compare the intragastric and the abdominal methods by using (1) a geometric mean of antero-posterior acquisitions, which is supposed to be the reference method, and (2) a group of 272 consecutive patients with diversified pathologies.

Patients, materials and methods Two hundred and seventy-two consecutive patients (147 women, mean age 44.12 years ± 14.63 and 125 men, mean age 45.69 years ± 15.31) were retrospectively included in this study from January 2001 to September 2005. These patients underwent a gastric emptying scintigraphy for idiopathic dyspepsia (n = 103), dyspepsia in a context of gastroesophageal reflux disease before surgery (n = 70) or after anti-reflux surgery (n = 17), for clinical symptoms of gastroparesis and/or worsening of glycaemia control (n = 55 patients with diabetes mellitus), or prior to heart–lung transplantation (n = 27 patients suffering from cystic fibrosis).

After the meal, the subjects were seated in the upright position. Four markers with about 10 MBq (0.3 mCi) of 99m Tc each, were fixed on the skin as anatomical reference points, two on the upper anterior thorax and the others. A large-field-of-view gamma camera (DSX, SMVi, Buck, France) with a medium-energy collimator was used. The first static acquisition was performed 10 min after the beginning of the meal in an anterior view and was followed 1 min later by a posterior view [1,7–9]. The duration of each acquisition was 1 min and each was obtained using 99mTc and 111In photopeak energy channels (windows ± 10%) on a 64  64 matrix. The interval between sets of anterior–posterior acquisitions was 20 min. Between each set of acquisitions, the patient was allowed to walk or remain seated. Drinking or eating was prohibited. The acquisitions were stopped when the gastric retention rate of solids was no more than one third of the initial maximum activity, in order to ensure that the experimental point of half-emptying was really acquired. Thus, computed gastric emptying parameters (see below) had a physiological meaning and were not simple mathematical estimates. Data analysis

The zero time (T0) was considered the time of the first acquisition, i.e., 10 min after the beginning of the meal. Regions of interest (ROIs) were drawn on a summed Fig. 1

Skin markers Gastric ROI

Test meal and data acquisition

All examinations were performed in the morning after an overnight fast. For the purposes of reproducibility, all patients ingested the same test meal [6], which was calibrated at 370 kcal and contained a well-cooked omelette made with an egg, in which the albumin was labelled with 37 MBq (1 mCi) of 99mTc sulfur colloid (TCK1 Cis Bio, Gif sur Yvette, France), a slice of ham, two slices of buttered toast (10 g of butter), orange juice (100 ml) and a glass of water (100 ml) labelled with 7.4 MBq (200 mCi) of 111In-DTPA (Mallinckrodt, Le Petten, the Netherlands). The glass of water was given at the end of the meal. Six minutes were allotted for the ingestion of the test meal.

Abdominal ROI

Gastric and abdominal regions of interest (ROIs) used in both methods. For the intragastric method, the rate of gastric emptying is assessed by calculating the geometric means of intragastric counts. For the abdominal method, the geometric means of intragastric counts are divided by the sum of the geometric means of intragastric counts plus abdominal counts, i.e., the geometric means of the total abdominal counts.

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Comparison of gastric emptying scintigraphy Salaun et al. 433

111

In image, on the anterior view and on the posterior view, and thereafter were applied on 111In and 99mTc sets (Fig. 1). For the intragastric method, ROIs were drawn around the total stomach, and for the abdominal method, ROIs were also drawn around the abdominal area. Intragastric and abdominal radioactivities were measured inside these ROIs, excluding radioactive skin markers. After correction for Compton downscatter of 111In in the 99m Tc photopeak window, time–activity curves (TACs) were corrected for 99mTc physical decay. Intragastric method

Geometric means of absolute intragastric counts were determined for each set of anterior–posterior 111In and 99m Tc data. Percentages of meal retention (experimental retention rates) were calculated in relation to the maximum gastric activity.

Statistical analysis

Comparison between the intragastric and abdominal methods was carried out using experimental and computed data of corrected TACs, i.e., the retention rates (RR) at different times (from 0 to 180 min) and using the computed parameters (T50,S, T50,L, Tlag and TRE) and the values for the quality of fit. Univariate analyses were performed using paired t-tests. The agreement between the computed parameters obtained by both methods were assessed using the Bland–Altman method [12], in which the differences between two corresponding values were plotted against their mean. All statistical analyses were performed using Graphpad Prism 4.0b 2004 (Graphpad Software, USA). The level of statistical significance was set at 5%. Results were expressed as mean ± standard deviation.

Results Abdominal method

Geometric means of intragastric counts and of abdominal counts were determined for each set of anterior–posterior 111 In and 99mTc data. The proportion of abdominal radioactivity in the stomach was expressed as geometric means of intragastric counts divided by geometric means of total abdominal counts (i.e., intragastric plus abdominal counts). Percentages of intragastric meal retention (experimental retention rates) were calculated in relation to the maximum gastric proportion of intra-abdominal radioactivity. The corrected TACs of both the intragastric method and the abdominal method were fitted using the power exponential function described by Elashoff et al. [10], in which y(t) is the percentage of radioactivity remaining in the stomach at time t (computed retention rates): a

yðtÞ ¼ 2ðt=T50 Þ : The Elashoff parameters a and half-emptying time of solids (T50,S) and of liquids (T50,L) were determined by a non-linear least-squares fitting algorithm (Excel, Microsoft, USA). The biphasic nature of the gastric emptying of solids was estimated by a lag phase time (Tlag) and by a constant emptying time, called the time of real emptying (TRE), using a maximum slope tangent method. We described this approach in a previous paper [11]. The quality of fit (QF) was assessed by a variance ratio and defined as ! s2res QF ¼ 1  s2data where s2res is the residual sum of squares and s2data is the variance of the experimental data.

The intragastric and abdominal methods were applied to the whole group of patients. Because the experimental half-emptying point was systematically acquired for each patient, the calculation of the gastric emptying parameters was always possible by both methods. Experimental and computed retention rates

Table 1 shows the experimental retention rates (RRs from 0 to 180 min, i.e., RR0 to RR180) obtained with the two methods. For the solid component, the intragastric method resulted in higher experimental retention rates than did the abdominal method, except for RR0. However, differences were statistically significant only for RR0, RR160 and RR180 (P < 0.001). At baseline (t = 0), the abdominal method resulted in weakly higher RR0 with a very small standard deviation, than did the intragastric method. For the liquid component, both methods gave almost the same results, except for RR20, RR40 and RR60 (P < 0.001). Table 2 reveals that the differences between the experimental and the computed retention rate values Table 1 Comparison of experimental retention rates obtained using the intragastric method and the abdominal method Experimental retention rates

RR0 RR20 RR40 RR60 RR80 RR100 RR120 RR140 RR160 RR180

Solids

Liquids

Intragastric method

Abdominal method

Intragastric method

Abdominal method

98 ± 4* 96 ± 5 90 ± 10 80 ± 15 68 ± 19 57 ± 21 48 ± 21 39 ± 21 33 ± 21* 27 ± 24*

99 ± 2 96 ± 5 90 ± 10 79 ± 16 68 ± 19 57 ± 21 46 ± 22 37 ± 22 29 ± 22 18 ± 23

100 69 ± 15* 53 ± 16* 43 ± 15* 36 ± 15 30 ± 15 26 ± 14 21 ± 13 17 ± 13 12 ± 14

100 68 ± 15 52 ± 15 42 ± 15 36 ± 15 30 ± 15 25 ± 14 21 ± 13 17 ± 14 12 ± 14

* P < 0.001 for bilateral paired t-test. RR = retention rate (%) at different acquisition times (from 0 to 180 min). All results are expressed as mean ± SD. Values in italic type are significant differences.

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for the abdominal method were sometimes equal to and most often smaller than those obtained with the intragastric method. Although these differences were observed for almost all the mean values and all the standard deviation values, they were statistically significant only for RR0 to RR60 (P < 0.05) for the solids, and for RR20, RR60, RR100, RR120 and RR140 (P < 0.00001) for the liquids. Values for the quality of fit

The abdominal method resulted in a significantly higher quality of fit for both the solid corrected TAC and the liquid corrected TAC, than did the intragastric method

Table 2 Comparison of the difference in retention rates between the experimental and computed values using the two methods of region of interest Times of retention rates (min)

0 20 40 60 80 100 120 140 160 180

Differences for solids

Differences for liquids

Intragastric method

Abdominal method

Intragastric method

Abdominal method

1.4 ± 3.5* 2.2 ± 2.4* 2.4 ± 2.5** 2.5 ± 3.3** 2.6 ± 3.8 2.9 ± 4.9 3.1 ± 6.3 3.0 ± 7.0 2.8 ± 7.5 2.0 ± 7

0.7 ± 2.2 1.7 ± 2 2.0 ± 2.1 2.2 ± 2.8 2.3 ± 3.2 2.6 ± 4.1 2.8 ± 5 2.6 ± 5.2 2.4 ± 5.7 1.5 ± 4.5

0 5.5 ± 9.5* 4.2 ± 8.8 4.1 ± 7.2* 3.5 ± 7.7 3.2 ± 6* 2.9 ± 5.6* 2.7 ± 5.3* 1.9 ± 4 1.3 ± 2.8

0 4.6 ± 6.9 3.5 ± 6.2 3.3 ± 5.5 2.7 ± 4.9 2.7 ± 4.9 2.5 ± 4.7 2.3 ± 4.6 1.9 ± 4.1 1.3 ± 3.0

*

P < 0.00001 and P < 0.05 for bilateral paired t-test. RR = retention rate (%) at different acquisition times (from 0 to 180 min). All results are expressed as mean ± SD. Values in italic type are significant differences.

**

(for the solid QFs: 0.90 ± 0.12 vs. 0.85 ± 0.19, P < 0.00001; for the liquid QFs: 0.91 ± 0.19 vs. 0.89 ± 0.28, P < 0.00001, for the abdominal and the intragastric method respectively) (Table 3). Comparison of gastric emptying parameters using intragastric and abdominal methods

Table 3 shows the results of the comparison for the calculated gastric emptying parameters obtained after fitting of experimental data from abdominal and intragastric methods. Weak, but statistically significant differences were observed for calculated values of T50,S and TRE (for T50,S, 122 ± 46 min vs. 124 ± 48 min, P < 0.00001; for TRE, 163 ± 64 min vs. 168 ± 68 min, P < 0.05, respectively, for the abdominal and the intragastric methods). For liquids, statistical differences were observed only for the unitless shape coefficient a, which has no clear physiological meaning. However, there was no significant difference between the two methods in evaluating the T50,L. Using the Bland–Altman method, the differences between two corresponding values were plotted against their mean. Table 4 displays the mean values of all physiological parameters (i.e., T50,S, TREL, Tlag and T50,L which are expressed in minutes, thus easily interpretable), their mean difference and their 95% confidence interval. The 95% confidence interval was met by all the four parameters with less than 5% of outliers (Fig. 2). In addition, removing these outliers did not modify the results of the bilateral paired t-tests.

Discussion Comparison of computed gastric emptying parameters obtained after fitting the experimental data Table 3

Parameter

Solids Intragastric method

Liquids Abdominal method

Intragastric method

Abdominal method

124.29 ± 47.59* 122.29 ± 45.57 55.33 ± 43.85 54.51 ± 45.85 T50 (min) 0.76 ± 0.25 a 2.09 ± 0.63 2.08 ± 0.66 0.78 ± 0.26* 39.93 ± 28.70 39.03 ± 28.52 Tlag (min) TRE (min) 167.58 ± 68.08** 163.07 ± 64.25 0.90 ± 0.12 0.89 ± 0.28** 0.91 ± 0.19 Quality factor 0.85 ± 0.19* *

P < 0.00001 and P < 0.05 for bilateral paired t-test. T50 = half emptying time; a shape coefficient; Tlag = time of lag phase; TRE time of real emptying. All results are expressed as mean ± SD. **

Table 4

The Bland–Altman method

Gastric emptying parameter (min)

Intragastric method Mean

T50,S Tlag TRE T50,L

Using an LAO acquisition procedure, Lien et al. [5] have proposed an alternative method for plotting the emptying curve, based on the expression of the intragastric counts as a proportion of the total abdominal radioactivity. They concluded that this new abdominal method offered substantial advantages over the conventional intragastric method, related to smaller differences between TAC experimental values and power exponential fitted values, resulting in smoothing the emptying curve with a better goodness of fit. The authors hypothesized that using an LAO procedure, the classical intragastric method neglected the post-gastric radioactivity. Thus, the intragastric redistribution, i.e., when solids move from the more

*

124.29 39.93 167.58 55.33

Abdominal method *

± 1.96  SD

Mean

93.28 56.25 133.44 85.95

122.29 39.03 163.07 54.51

Difference: intragastric abdominal

± 1.96  SD

Mean*

± 1.96  SD

89.32 55.90 125.93 89.87

2 0.90 4.51 0.82

3.96 0.35 7.51 – 3.92

* Shown are the means of differences between the calculated values (minutes) derived from the intragastric and the abdominal methods. To size up the differences, the 95% confidence intervals of each mean value were also given.

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Comparison of gastric emptying scintigraphy Salaun et al. 435

Fig. 2

Half-emptying time of solids (T50,S)

(a) 30

m + 1.96 SD

10 m

0

m − 1.96 SD

−10

Difference (min)

Difference (min)

20

(b)

−20 −30

Time of the lag phase (Tlag)

40

(d)

20

m + 1.96 SD

10 m

0 −10

m − 1.96 SD

− 20

Difference (min)

Difference (min)

m m − 1.96 SD

100

200 300 Mean (min)

400

500

Half-emptying time of liquids (T50,L)

40 30

30

20

m + 1.96 SD

10

m

0 −10

m − 1.96 SD

− 20 − 30

−30 − 40

m + 1.96 SD

0

25 50 75 100 125 150 175 200 225 250 275 300 325 Mean (min) (c)

Times of real emptying (TRE)

90 80 70 60 50 40 30 20 10 0 −10 −20 −30 −40 −50

− 40 0

25

50

75 100 125 150 175 200 Mean (min)

− 50

0

50

100

150 200 Mean (min)

250

300

350

Comparison of (a) the half-emptying time for solids (T50,S), (b) times of real emptying (TRE), (c) time of the lag phase (Tlag) and (d) the half-emptying time for liquids (T50,L) obtained with the intragastric and the abdominal methods using the Bland–Altman method. The differences between two corresponding values were plotted against their mean. The means of differences and their 95% confidence intervals were drawn.

posterior fundus to the more anterior antrum, could account for observed retention rates > 100% in the early periods of the gastric emptying process. In contrast, with the abdominal method, the authors reported that the early rise in the peak in lag phase disappeared. This was mainly due to an inherent characteristic of this new abdominal method, which is a 100% retention rate at T0, obviously related to the fact that soon after the ingestion of the test meal, there is no intestinal radioactivity. In our experience, using geometric means of anterior–posterior views, the classical intragastric method also leads to encounter retention rates of > 100% in the lag phase period. Furthermore, in the present study, retention rates > 100% were also observed with the abdominal method, although the abdominal method rates at T0 were closer to 100% than those obtained with the intragastric method. In contrast to the assumptions concerning the LAO scanning method, in our study this phenomenon could not result from an intragastric redistribution of the test meal, because antero-posterior geometric means take into account both anterior and posterior distributions of the

test meal at each acquisition time. This early rise of retention rates may rather be attributed to self-absorption of a meal [13]. Thus, the TAC maximum value is not necessarily the value at T0, resulting in curves that are not normalized by the value at T0 but by the maximum value of each set of data [14].

Using this normalization by the maximum value and because we ceased the acquisitions only when the retention rate of solids was no more than one third of the maximum gastric activity, we did not encounter any difficulty in fitting the TAC obtained from the intragastric method or from the abdominal method. This acquisition procedure ensured that the half-emptying time was acquired experimentally, so it allowed the computation of gastric emptying parameters that have a physiological meaning and are not simply mathematical extrapolations. Thus, in contrast to the study by Lien et al., both the intragastric method and the abdominal method were applied to all the experimental data from

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Nuclear Medicine Communications 2006, Vol 27 No 5

the 272 gastric scintigraphy scans, with 100% success in calculating the gastric emptying parameter for both methods. Another reason why fitting was easy to perform was that Excel software (Microsoft, USA) was used. Before data are fitted with reference to the particular shape of each TAC, this spreadsheet allows the user to predefine the only two parameters of the power exponential function (i.e., the half-emptying time T50 and the shape coefficient a). Empirically approaching the fit parameters reduces the number of iterations required to solve the mathematical system, and sometimes allows a convergence towards a solution, which would not always be possible otherwise.

In our study, the power exponential function described by Elashoff et al. [10] was applied to both liquid and solid meal components, even if both components exhibited different shapes. This power exponential function permits the characterization of the biphasic emptying of solids [15], allowing the quantification of the half-emptying time (T50,S), the duration of the first phase of the emptying process, i.e., the time of the lag phase (Tlag) [16] and the duration of the second phase of constant emptying [17], which is called the time of real emptying (TRE) in our previous publications [11,14]. Because liquid emptying does not exhibit a lag phase [18], liquid TAC has been conventionally fitted by a simple exponential function [19]. However, when its shape coefficient is equal to 1, the power exponential function becomes a mono-exponential function, and thus can be used for the mathematical expression of the emptying rate of both liquids and solids. Fitting the TACs by this function provided an excellent quality of fit, with values close to 1. However, in agreement with the results obtained by Lien et al., we observed that the goodness of fit was significantly better with the abdominal method than with the intragastric method. The superior quality of fits with the abdominal method is also illustrated in Table 2, showing weaker differences between the experimental and computed abdominal method retention rates, than the differences observed with the intragastric method. A possible explanation was the more pronounced fluctuation of the intragastric method TAC than the abdominal method TAC. Although radioactive skin markers can theoretically minimize this drawback, errors in repositioning or movements of the subjects in front of the gamma camera are probably inevitable during a long period of scanning and can affect the results of the intragastric method which are closely dependent on the intragastric counts. From this point of view, the abdominal method has a substantial advantage over the intragastric method, because both the intragastric counts and the abdominal counts change in the same range, thus minimizing the variation of intragastric counts as expressed as a proportion of the total abdominal counts.

Fitting the TAC with a power exponential function allows the use of the estimated gastric emptying parameters to compare the intragastric method and the abdominal method. Using paired t-tests, Lien et al. did not find any significant difference for the T50,S in the whole group of patients, as well as in the two sub-groups of patients with normal or with delayed gastric emptying. However, calculation of this parameter was not always possible, thus minimizing the power of these statistical tests. Furthermore, the lack of statistical difference with paired t-tests, did not mean that the two methods agreed, but rather that the differences could be equally distributed on both sides of the average difference. For this reason, we performed Bland–Altman statistical tests to assess the level of agreement between the two methods, by comparing the computed parameters having a clinical relevance, i.e., T50,S, T50,L, Tlag and TRE, which are all expressed as times, and then provide an easy understanding of the gastric emptying processes. This statistical method revealed good agreements between the computed parameters, i.e., the 95% confidence interval was met by all parameters with less than 5% of outliers. Because good agreement does not mean identical results, it was not surprising that significant differences were observed for T50,S and TRE. On average, T50,S and TRE obtained for the intragastric method were, respectively, 2 ± 3.96 min (mean ± 1.96  SD) and 4.51 ± 7.51 min higher than the abdominal method T50,S and TRE, in which the differences were too weak to compromise the validity of the results of one or the other method in clinical practice. By contrast, the estimates of Tlag and T50,L were similar with both methods (no significant difference and good agreement).

Conclusion The abdominal method is a reliable and efficient method for plotting the emptying curve. Its principal advantage is to smooth the emptying curve, allowing a better fit by the mathematical function. However, when the half-emptying time is acquired experimentally, fitting is easy to perform and always possible with both methods. In this case, the two methods are in agreement and give similar results for liquids and quite similar results for solids, with only very small differences that are acceptable in clinical practice. When the half-emptying point is not acquired experimentally, fitting the curve has no physiological meaning, because estimates of the gastric emptying parameter represent only mathematical extrapolations. Thus, in the case of severe gastroparesis, we recommend that computed parameters are not used, but, rather, the experimental retention rates at 120 or 180 min and are used to describe the particular shape of the emptying curve (with a slow, linear decrease). We conclude that, when fitting is possible, either the intragastric or the abdominal method can be used to treat antero-posterior data of gastric scintigraphy.

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Maurer A, Knight L, Charkes N, Vitti R, Krevsky B, Fisher R, et al. Comparison of left anterior oblique and geometric mean gastric emptying. J Nucl Med 1991; 32:2176–2180. Ford PV, Kennedy RL, Vogel JM. Comparison of left oblique anterior, anterior and geometric mean methods for determining gastric emptying times. J Nucl Med 1992; 33:127–130. Fahey F, Ziessman H, Collen M, Eggli DF. Left anterior oblique projection and peak-to-scatter ratio for attenuation compensation of gastric emptying studies. J Nucl Med 1989; 30:233–239. Meyer J, Van Deventer G, Graham L, Thomson J, Thomasson D. Errors and corrections with scintigraphic measurements of gastric emptying of solid foods. J Nucl Med 1983; 24:197–203. Lien H-C, Chang C-S, Chen G-H, Kao C-H, Tsai S-C, Wang S-J, et al. Gastric emptying rate assessment based on the proportion of intraabdominal radioactivity in the stomach. J Nucl Med 1999; 40:1106–1110. Lartigue S, Bizais Y, Bruley-des-Varannes S, Murat A, Pouliquen B, Galmiche JP. Inter- and intra-subject variability of solid and liquid gastric emptying parameters. A scintigraphy study in healthy subjects and diabetic patients. Dig Dis Sci 1994; 39:109–115. Goo R, Moore J, Greenberg E, Alazraki N. Circadian variation in gastric emptying of meals in humans. Gastroenterology 1987; 93:515–518. Tothill P, McLoughlin G, Heading R. Techniques and errors in scintigraphic measurements of gastric emptying. J Nucl Med 1978; 19:256–261. Tothill P, McLoughlin G, Holt S, Heading R. The effect of posture on errors in gastric emptying measurements. Phys Med Biol 1980; 25:1071–1077.

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Original article

A study of the possibility of curing Graves’ disease based on the desired reduction of thyroid mass (volume) as a consequence of 131I therapy: a speculative paper Antonio C. Trainoa, Fabio Di Martinoa, Mariano Grossoc, Fabio Monzanid, Angela Dardanod, Nadia Caracciod, Giuliano Marianic and Mauro Lazzerib Purpose The possibility of predicting the final volume of Graves’ disease thyroids submitted to 131I therapy could allow the physician to decide what activity to administer based on the desired volume reduction instead of on a fixed value of the thyroid radiation absorbed dose. In this paper the relationship between maximum uptake of 131I, fractional reduction of thyroid volume and outcome of Graves’ disease is discussed. Methods The results are based on ultrasonography thyroid volume measurements before administration of therapy and at the moment of recovery from Graves’ disease (thyroid stimulating hormone > 0.3 lIU
ml – 1 in the absence of anti-thyroid drug therapy) and on measurements of 131I uptake in 40 patients. It is shown that the possibility of curing Graves’ disease may be individually related to the final volume of the patient’s thyroid. An equation is presented to calculate the ‘optimal’ final thyroid volume.

The corresponding thyroid median absorbed doses are, respectively, 353 Gy and 320 Gy (non-parametric Wilcoxon test, P < 0.02). Conclusion A method to evaluate individually the ‘optimal’ final thyroid mass is presented and discussed. The method based on ‘volume reduction’ could probably reduce the activity and the thyroid absorbed dose compared to the method based on ‘empirical’ calculations, thus allowing the administration of 131I therapy to be optimized. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:439–446  Nuclear Medicine Communications 2006, 27:439–446 Keywords: Graves’ disease,

131

I therapy, dosimetry, thyroid, thyroid volume

a Sezione di Fisica Medica, bU.O. Fisica Sanitaria, Azienda OspedalieroUniversitaria Pisana, cCentro Regionale di Medicina Nucleare, Azienda Ospedaliero-Universitaria Pisana e Universita` di Pisa and dDipartimento di Medicina Interna, Universita` di Pisa, Italy.

Results A comparison between the traditional method, based on absorbed dose, and the final method, based on volume, has been carried out retrospectively. In the first case a median activity of 529 MBq has been administered; in the second, a median activity of 394 MBq (nonparametric Wilcoxon test, P < 0.05) should be administered.

Correspondence to Dr A. Claudio Traino, Sezione di Fisica Medica, U.O. Fisica Sanitaria, Azienda Ospedaliero-Universitaria Pisana Via Roma n. 67, 56125 Pisa, Italy. Tel: + 0039 50 992219; fax: + 0039 50 992513; e-mail: [email protected]

Introduction

Recently, an empirical relationship between post-therapeutic thyroid volume and therapy outcome in Graves’ disease patients has been reported by a number of authors [9–12]. This relationship suggests the possibility of deciding the optimal 131I activity (A0) to administer to the patient based on the desired therapy-induced reduction in thyroid volume.

The use of 131I in the therapy for Graves’ disease has a long history, dating back to the early 1940s. It has largely replaced surgery and is nowadays commonly used because it is easy to perform and has proved to be effective in the definitive treatment of hyperthyroidism. Nevertheless, there is no consensus regarding the most appropriate dosimetric approach for radioiodine therapy. Traditionally, the amount of 131I to be used is based on one of two different modalities: either (1) the administration of a fixed activity of radioiodine, A0, (usually 555 MBq) or (2) the administration of an activity that is calculated individually and on an optimal fixed value of the target absorbed dose, DT (in Gy). Many papers have been published in order to decide which of these two modalities is better [1–4]. Other papers have been published to establish an optimum value of therapeutic activity [5], or target absorbed dose [6–8]. The results presented are in poor agreement.

Received 22 December 2005 Revised 14 February 2006 Accepted 14 February 2006

To date, the foregoing results have not been applied clinically, mainly because any prediction of the reduction in thyroid volume as a result of radioiodine therapy of Graves’ disease has been considered an intractable problem. This is because the reduction in thyroid volume depends on a number of patient-specific parameters that, generally, are not evaluable. A mathematical model of thyroid mass reduction after administration of 131I therapy to patients with Graves’ disease has recently been published [13]. This model

c 2006 Lippincott Williams & Wilkins 0143-3636 

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440 Nuclear Medicine Communications 2006, Vol 27 No 5

relates the therapeutic radioiodine activity, A0, to the desired reduction of the thyroid mass, allowing the optimal therapeutic activity, A0, to be decided predictively; that is, after administration of a diagnostic activity (0.37–1.85 MBq). Starting from this, the present paper describes and discusses a completely new approach: the possibility of curing Graves’ disease by choosing, individually, a therapeutic 131I activity based on therapy-induced reduction of thyroid volume instead of on the target absorbed dose.

Remembering that the reduction of thyroid mass due to 131 I therapy has been observed and reported by many authors, it follows from the second hypothesis and Equation 2 that for each patient U0 UG ¼ m0 mG

ð3Þ

where mG is the thyroid mass (volume) at the moment of cure of Graves’ disease and UG is a parameter related to the uptake by the gland at the same moment. Note that if UG is known, Equation 3 allows mG to be evaluated. Thus the first problem is the determination of an ‘optimal’ value for UG.

Methods and materials This study is based on the following hypotheses. Firstly, that in a Graves’ disease thyroid the 131I activity is uniformly distributed throughout the gland (i.e., the uptake can be considered almost uniform). This means that thyroid iodine trapping, organification and function is uniform throughout the gland. Secondly, that there is a linear correlation between the uptake concentration, U0/ m0, before therapy and the uptake concentration U1/m1 at the end of therapy. From the first hypothesis it follows that the thyroid functional tissue can be described as a set of identical traps uniformly distributed throughout the gland [14]. These traps are able to concentrate iodine from the blood. Each of them is assumed to take up the same fraction of radioiodine from the blood, thus the radioiodine uptake depends on the number of traps in the gland. For each patient the number of traps is linearly related to the mass (volume) of the thyroid, i.e., the mass (volume) of the gland is individually linearly related to the uptake. This does not mean that patients with the same thyroid mass (volume) have the same number of traps (it depends on thyroid uptake), but that, by reducing the mass (volume) of the gland by a fraction, f, the thyroid uptake is reduced by an amount related to the fraction f. The second hypothesis can be expressed by the equation U1 U0 ¼j m1 m0

ð1Þ

where m0 is the basal mass of the gland, U0 is the basal maximum thyroid uptake, m1 is the mass of the gland 6 months to 1 year after administration of therapy, and U1 is the maximum thyroid uptake 6 months to 1 year after administration of therapy (i.e., at the end of 131I therapy). j is a coefficient which takes into account the adjustment of uptake per unit mass due to radioiodine therapy. Note that Equation 1 can be written as U1 U0 ¼ m1 m0 where U1* = U1/j.

ð2Þ

From March 2002 to April 2003, 106 patients with Graves’ disease were treated with 131I in our nuclear medicine department. Forty-six of these patients were enrolled in this study. All patients included had proven thyrotoxicosis from Graves’ disease, as confirmed by suppressed basal thyroid-stimulating hormone (TSH), increased serum triiodothyronine (FT3) and thyroxine (FT4), and the presence of TSH receptor antibodies and/or evidence of ophthalmopathy. Twenty-four hours after administration of 131I (1.85 MBq) all the patients underwent 131I scintigraphy, before receiving the therapeutic activity, to confirm the diagnosis of Graves’ disease. Apart from possible refusal of radioiodine therapy by a patient, exclusion criteria included age younger than 18 years, suspicion of pregnancy, and the presence of any suspicious thyroid nodule on ultrasound examination. All the patients were advised to stay on a generic, lowiodine diet for at least 2 weeks prior to treatment and anti-thyroid drug therapy was withdrawn prior to radioiodine administration (average, 7.5 days; range, 5–30 days). Anti-thyroid drug treatment was restarted (for 30 days) at least 7 days following administration of 131I therapy. All these 46 patients (34 women and 12 men; average age 54 ± 18 years, range 27–82 years) were followed monthly by assessing serum levels of TSH, FT4 and FT3. When they were considered as cured (TSH > 0.3 mmIU
ml – 1 in the absence of anti-thyroid drug therapy) the thyroid volume was evaluated by ultrasonography to calculate mG and then, by Equation 3, UG. Eleven patients (five from the initial 106 patients plus six from the group of 46 patients) with Graves’ disease (eight women and three men; average age of 51 ± 13 years; range, 26–72 years) received a second course of 131I therapy for persistent hyperthyroidism 10–12 months after the first treatment. These patients were followed to validate the second hypothesis and, consequently, Equation 1. The value of the ratio U0/m0 calculated for each patient before the first course of therapy was compared to the value of the ratio U1/m1 calculated before administration of the second course. The coefficient j has been

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Thyroid mass reduction-based method for

131

I Graves’ disease therapy Traino et al. 441

131 Table 1 Thyroid mass m and maximum uptake U before the first (m0, U0) and the second (m1, U1) I treatment of 11 patients submitted twice to radionuclide therapy for Graves’ disease

Patient number

m0

U0

m1

U1

U0/m0

U1/m1

1 2 3 4 5 6 7 8 9 10 11

54 12 15 22 30 40 70 36 33 50 38

0.53 0.92 0.51 0.68 0.47 0.73 0.68 1.00 0.93 0.65 0.56

25 10 10 16 13 21 21 19 12 24 13

0.70 0.75 0.41 0.50 0.33 0.70 0.64 0.77 0.64 0.53 0.42

0.010 0.077 0.034 0.031 0.016 0.018 0.010 0.028 0.028 0.013 0.015

0.028 0.075 0.041 0.031 0.025 0.033 0.030 0.041 0.053 0.022 0.032

evaluated. The results are summarized in Table 1. The other 40 patients became hypothyroid within 1 year of therapy administration. The basal thyroid mass, m0, was calculated by ultrasonography volume measurements performed before radioiodine administration (the thyroid density can be considered as 1 g
ml – 1), the time to maximum uptake, Tmax, the half-life of radioiodine in the gland, Teff, and the maximum activity in thyroid, Am, were calculated for each patient after therapy administration. The choice of the therapeutic 131I activity, A0, administered was based on thyroid volume, maximum uptake (evaluated by two measurements performed 4 and 24 h after administration of the tracer) and the clinical situation of the patient, and mG, was evaluated by ultrasonography volume measurement performed at the moment of disease cure (Table 2). Thyroid volume measurements were done using an AU-4 echograph (ESAOTE; Genova, Italy), equipped with an array 7.5-Hz transducer. All measurements were done by the same nuclear medicine specialist. The ellipsoid model has been used for each thyroid lobe and isthmus volume calculation. The thyroid absorbed dose, DT, was calculated using the equation [15] Dnp U0 A0 DT ¼ m0 ( "
 #) Tmax 1 A 0 Teff 1=2 þ  1  1  2g ð4Þ gA0 2 ln 2 where Dnp = 0.032 mGy
g
MBq – 1
s – 1 is the equilibrium dose constant for non-penetrating radiations [16], U0 = Am/A0 is the maximum radioiodine thyroid uptake and g = 2.2  10 – 6 is a one-dimensional constant [14,15]. Equation 4 allows calculation of the radiation dose to the thyroid by taking into account the change of thyroid mass (volume) during the clearance phase of 131I in the gland.

Results In Table 1 the U/m values calculated before the first (U0/m0) and the second (U1/m1) course of 131I therapy for

11 patients with Graves’ disease who underwent a second treatment are reported. In Fig. 1 the scattergram of U0/m0 vs. U1/m1 with error bars is shown. The error bars represent the maximum uncertainty (about 20%) in U/ m. From the represented data (for 11 patients) the uptake concentrations, U/m, before and after therapy seem to be linearly related (R = 0.91), and an adjustment factor j = 1.2 due to radioiodine therapy (hypothesis 2) has been calculated. Thus, Equation 1 and, consequently, Equations 2 and 3 can be used. The thyroid absorbed dose DT (in Gy) and the UG values calculated for 40 patients with Graves’ disease are shown in Table 2. UG values were calculated by Equation 3, where mG is the thyroid mass evaluated by thyroid volume measurement at the moment of Graves disease cure (TSH > 0.3 mIU
ml – 1 in the absence of anti-thyroid drug therapy). Note that all these 40 patients were cured; that is, they become hypothyroid as a consequence of 131I therapy within 1 year. The median value of DT is 353 Gy (range, 110–1311 Gy); the median value of UG is 0.34 (range, 0.24–0.65). The frequency distribution of UG is shown in Fig. 2. Considering an optimal final value of UG = 0.24 (this is the lower UG value for the 40 Graves’ diseased patients shown in Table 2), from Equation 3 it follows that m0 m0 ¼ 0:24 ð5Þ mG;cal ¼ mfin ¼ UG U0 U0 where mfin is the final mass of the thyroid. Equation 5 allows a retrospective re-calculation of the optimal final thyroid mass (volume) mG,cal (in g) for each of the 40 patients. The comparison between mG and mG,cal for each of 40 patients is shown in Fig. 3(C). Note that mG > mG,cal because UG = 0.24 is the lowest value of UG calculated by Equation 3 for the 40 Graves’ disease patients considered. Based on the hypothesis that the outcome of Graves’ disease is related to the reduction of the thyroid mass (volume) due to 131I, mG,cal can be considered the optimal value of the final thyroid mass. This means that further thyroid mass reduction can not be considered as useful for

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442

Nuclear Medicine Communications 2006, Vol 27 No 5

Table 2

Measured data for 40 patients treated with

Patient number 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40

131

I for Graves’ disease

m0(g)

A0(MBq)

Am(MBq)

Teff(h)

Tmax(h)

DT(Gy)

mG(g)

UG

23 18 40 16 35 23 24 32 21 26 19 16 30 18 23 14 39 29 13 30 16 56 9 64 12 15 18 35 28 19 14 16 23 51 23 14 49 27 29 22

555 1036 370 339 740 555 222 666 555 666 185 444 555 370 555 503 503 370 185 666 370 555 296 925 740 258 740 222 407 444 370 370 650 555 555 185 740 470 555 555

322 991 337 331 604 326 134 590 549 659 151 357 478 269 533 407 355 370 94 595 288 505 137 628 555 215 562 171 407 342 208 295 443 480 511 150 740 315 383 549

122 105 122 141 122 122 122 98 141 109 122 118 122 90 116 139 122 122 126 127 131 94 139 122 122 122 122 122 149 122 122 122 80 108 122 122 107 122 122 169

24 24 24 4 24 24 24 19 28 9 24 18 24 22 12 25 24 24 23 21 20 2 10 24 24 24 24 24 4 24 24 24 20 20 24 24 4 24 24 4

342 1311 196 535 445 346 126 359 757 544 178 503 389 256 520 809 219 297 167 523 448 156 386 272 1194 326 806 110 406 427 347 430 304 200 542 240 317 278 322 863

14 7 15 4 13 13 17 12 8 9 6 13 14 6 14 6 16 11 7 16 6 39 7 23 5 9 7 12 8 7 11 12 9 21 8 9 26 18 13 11

0.35 0.37 0.34 0.24 0.30 0.33 0.43 0.33 0.38 0.34 0.26 0.65 0.40 0.24 0.58 0.35 0.29 0.38 0.27 0.48 0.29 0.63 0.36 0.24 0.31 0.50 0.30 0.26 0.29 0.28 0.44 0.60 0.27 0.36 0.32 0.52 0.53 0.45 0.31 0.49

m0 is the basal mass of the gland; A0 is the activity of radioiodine administered; Am is maximum activity of the thyroid; Teff is half-life of radioiodine in the gland; Tmax is the time for maximum uptake; DT is target absorbed dose; mG is the thyroid mass (volume); and UG is a parameter related to the uptake of the gland. The patients were followed for 1 year after administration of therapy. They all became hypothyroid. The value of DT (Gy) has been calculated by using Equation 4 and UG has been calculated by using Equation 3.

Note that the mG values shown in Table 2 do not represent the final masses (volumes) of the thyroid due to 131I therapy. Values of mG (in g) are only the thyroid mass values measured at the moment of recovery from Graves’ disease. The final thyroid mass values of the 40 patients considered are less than mG. This means that probably too much activity has been administered.

U1/m1

Fig. 1

0.1 0.09 0.08 0.07 0.06 0.05 0.04 0.03 0.02 0.01 0

The final mass of the thyroid due to evaluated using the equation [13] 0

0.01 0.02 0.03 0.04 0.05 0.06 0.07 0.08 0.09 0.1 U0/m0

Graphical representation of the relationship j between maximum uptake per unit mass, U/m, before (U0/m0) and after (U1/m1) 131I therapy of Graves’ disease (Equation 1). The error bars represent the maximum uncertainty of U/m (20%). The linear correlation coefficient r = 0.91; j = 1.2 ± 0.1.

the outcome of radioiodine therapy (mG,cal represents the lower ‘cure’ thyroid mass value (i.e., the optimal final thyroid mass value)).

131

mfin ¼ m0 expðDT Þ

I therapy can be ð6Þ

where a = 0.0038 Gy – 1 is a constant typical of thyroid tissue. From Equations 5 and 6 it follows that
 1 0:24 DT ¼ DT;G ¼  ln ð7Þ a U0 where DT,G is the thyroid dose needed to obtain a final thyroid mass mG,cal, that is, the ‘optimal’ thyroid finalmass value (see Equation 5).

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Thyroid mass reduction-based method for

131

I Graves’ disease therapy Traino et al. 443

Fig. 2

12

11

Number of Patients

10

9

8

7

6 4

3

3 2

2

2

2

1

−0 .6 5 60 0.

−0 .6 0 56 0.

56 0. 0. 51 −

51 0. 0. 47 −

−0 .4 7 0.

42

−0 .4 2 38 0.

3

−0 .3 8 33 0.

.3 −0 29 0.

0.

24

−0 .2 9

0

UG Frequency distribution of UG for 40 Graves’ disease patients. UG has been calculated by using Equation 3.

DT,G has been retrospectively calculated by Equation 7 for the 40 Graves’ disease patients. The results are shown in Fig. 3(B). The median value of DT,G is 320 Gy (range, 173–376 Gy). The corresponding 131I administering activity, A0,G, calculated by Equation 4, has a median value of 394 MBq (range, 138–1092 MBq), whereas the median value of the effectively administered activity, A0, is 529 MBq (range, 185–1036 MBq) (Fig. 3(A)). There is a significant difference between A0 and A0,G (P < 0.05, non-parametric Wilcoxon test) and between DT and DT,G (P < 0.02, non-parametric Wilcoxon test).

Discussion Although 131I therapy of Graves’ disease is well established for definitive treatment of Graves’ hyperthyroidism, the best method for determining the quantity of radioiodine administered remains highly controversial. Traditionally, the choice of the amount of 131I therapy is based on one of two different approaches: (1) a fixed amount of 131I activity (in MBq); and (2) an amount of 131 I activity (MBq) based on a fixed value of the target absorbed dose (in Gy). These two different approaches are not equivalent. This is shown in Fig. 4. There is no correlation between A0 (MBq) and DT (in Gy) (r = 0.56). The approach based on the administration of a fixed activity is only empirical; it is a very inaccurate way to choose the therapeutic activity of 131I [3]. It does not take into account the differences in radioiodine kinetics (U, Teff, Tmax) and in basal mass (m0) between patients. It is very difficult to explain, theoretically, why this approach seems to work well. It probably does so only for high administered activities (about 555 MBq), thus it does not optimize therapy.

On the other hand it is also difficult to explain the meaning of one fixed ‘optimal’ target absorbed dose value. Many studies have been carried out and many papers published trying to decide an ‘optimal’ absorbed dose value. Peters et al. [2] report a success rate of about 40–50% in patients who received a target dose of 100 Gy (with considerable differences due to the basal mass of the thyroid) and a success rate of 80% in patients who received a target dose of 200 Gy. In another paper from the same group [17] the authors demonstrate a strong correlation between the outcome of therapy and the radiation dose absorbed by the thyroid. The rate is 11% for 50 Gy, 50% for 100 Gy, 67% for 150 Gy, 80% for 200 Gy, 84% for 250 Gy, 90% for 300 Gy and 93% for 400 Gy. Reinhardt et al. [6] report a success rate of 73% for 150 Gy, increasing to 92% for 300 Gy. Bajnok et al. [7] report a success rate of 72% 6 months after therapy administration for a thyroid absorbed dose ranging from 70 to 100 Gy. They relate the success of the therapy to the size of the gland. In their study Howarth et al. [8] randomly divide the patients into two groups, one receiving 60 Gy and the other receiving 90 Gy. They report a positive response of 41% of patients of the second group (90 Gy) and of 39% of patients of the first group (60 Gy) 6 months after 131I administration. They conclude that no significant advantage in response rate is gained by using a dose of 90 Gy instead of 60 Gy. More recently, an empirical relationship between posttherapeutic thyroid volume (mass) and therapy outcome in Graves’ disease patients has been reported in the literature. Gomez-Arnaiz et al. [12] report a relation between the thyroid volume 3 and 6 months after therapy and final thyroid function outcome. Haase et al. [11] divide the patients into three groups depending on the

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444 Nuclear Medicine Communications 2006, Vol 27 No 5

Fig. 3

(A)

1200 A0 (MBq)

A0(DTG) (MBq)

1000

Activity (MBq)

800

600

400

200

0 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 Patient number

(B)

1400 DT (Gy)

DTG (Gy)

Thyroid absorbed dose(Gy)

1200

1000

800

600

400

200

0 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 Patient number

(C)

45 40

mG (g)

mGcal (DTG Gy)

Thyroid cure mass (g)

35 30 25 20 15 10 5 0 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 Patient number

Comparison between the effectively administered activity A0 (in MBq) and the activity A0(DTG) (in MBq) which corresponds to the optimal dose value DTG(in Gy) (A); comparison between the dose effectively absorbed by thyroid DT (in Gy) and the optimal dose value DTG (in Gy) (B); comparison between the measured cure mass of thyroid mG (in g) and the calculated (by using Equation 5) cure mass of the gland mG,cal (in g) (C). Note that DTG (Gy) has been calculated by using Equation 7 (i.e., it is the radiation dose which reduces the thyroid mass to the value mG,cal).

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Thyroid mass reduction-based method for

D T (Gy)

I Graves’ disease therapy Traino et al. 445

depends linearly on the reduction of the thyroid mass. This result can probably be used (Table 1 and Fig. 1) even if the data presented involve only 11 patients.

Fig. 4

1400 1200 1000 800 600 400 200 0

131

Conclusion

0

200

400

600 800 A 0 (MBq)

1000

1200

Relationship between administered activity A0 (in MBq) and thyroid absorbed dose DT (in Gy) for 40 Graves’ disease patients. R = 0.56.

basal thyroid volume (mass) and adjust the dose calculation to the patient’s thyroid volume. They report a therapeutic success associated with different target doses in each of the three groups (150 Gy for thyroid volumes < 15 ml; 220 Gy for thyroid volumes in the range 15–25 ml; 260 Gy for thyroid volumes > 25 ml) and they find a significant correlation between post-therapeutic thyroid volume (mass) and clinical outcome. They conclude that an adjustment of the target doses based on thyroid basal volumes will lead to an appropriate reduction of thyroid volumes. This means that the ‘optimal’ absorbed dose value is individual (i.e., it depends on the characteristics of the patient and on the ‘appropriate’ reduction of his thyroid volume). Note that all those results are based only on statistical considerations, without any theoretical model and for this reason the debate on the optimization of 131I therapy of thyroid cancer has a very weak foundation. In this paper the relationship between 131I maximum uptake, fractional reduction of thyroid volume and outcome of Graves’ disease is discussed and a simple method is presented to calculate the ‘optimal’ final thyroid volume (Equation 5) and the corresponding thyroid dose value DT,G (Equation 7). This method is based on a theoretical model, described by Equations 1, 2 and 3.

The possibility of administering 131I activity based on the reduction of thyroid mass instead of a fixed thyroid absorbed-dose value could present a new chance to cure Graves’ disease. In this paper a method to evaluate, individually, the ‘optimal’ final thyroid mass is described (Equation 3) and discussed. It is not more difficult to apply this method than it is to apply the methods usually employed to calculate the activity based on a fixed value of thyroid absorbed dose, DT. This is because the calculation of DT,G only requires a knowledge of the basal thyroid maximum uptake, U0 (Equation 7). From DT,G (in Gy) an ‘optimal’ activity A0,G (in MBq) can be calculated from Equation 4. This equation depends on parameters measurable after the administration of a diagnostic (0.37–1.85 MBq) amount of activity. It is shown that the method based on ‘volume reduction’ could probably reduce the activity and the thyroid absorbed dose with respect to methods based on ‘empirical’ calculations, allowing the administration of therapy to be optimized. This is only a speculative paper and the method presented needs further carefully performed prospective studies before it can be accepted and eventually adopted. On the other hand, it suggests a possible new way to cure Graves’ disease, based on a theoretical model, overcoming the present debate on the best modality for administering radioiodine therapy.

References 1

2

3

4

Some simplified hypotheses must be discussed. The first is the uniform uptake of the Graves’ disease gland. This is a hypothesis which can be accepted for Graves’ disease thyroids. The second important hypothesis is that the uptake concentration U0/m0 is linearly related to the final uptake concentration U1/m1 as a consequence of radioiodine therapy. This is an important hypothesis because it implies Equations 1 and 3, and, consequently, Equation 5. It means that the reduction of the thyroid uptake

5

6

7

8

Catargi B, Leprat F, Guyot M, Valli M, Ducassou D, Tabarin A. Optimized radioiodine therapy of Graves’ disease: analysis of the delivered dose and of other possible factors affecting outcome. Eur J Endocrinol 1999; 141: 117–121. Peters H, Fischer C, Bogner U, Reiners C, Schleusener H. Treatment of Graves’ hyperthyroidism with radioiodine: results of a prospective randomized study. Thyroid 1997; 7:247–251. Jo¨nsson H, Mattsson S. Excess radiation absorbed doses from nonoptimised radioiodine treatment of hyperthyroidism. Radiat Prot Dosim 2004; 108:107–114. Jo¨nsson H, Mattsson S. Current methods for absorbed dose planning for radioiodine treatment of hyperthyroidism in Sweden. Phys Med 2004; 20:99–103. Jarlov AE, Hegedu¨s L, Kristensen LO, Nygaard B, Hansen JM. Is calculation of the dose in radioiodine therapy of hyperthyroidism worth while? Clin Endocrinol (Oxford) 1995; 43:325–329. Reinhardt MJ, Brink I, Joe AY, von Mallek D, Ezziddin S, Palmedo H, et al. Radioiodine therapy in Graves’ disease based on tissue-absorbed dose calculations: effect of pre-treatment thyroid volume on clinical outcome. Eur J Nucl Med 2002; 29:1118–1124. Bajnok L, Mezosi E, Nagy E, Szabo J, Sztojka I, Varga J, et al. Calculation of the radioiodine dose for the treatment of Graves’ hyperthyroidism: is more than seven-thousand rad target dose necessary? Thyroid 1999; 9: 865–869. Howarth D, Epstein M, Lan L, Tan P, Booker J. Determination of the optimal minimum radioiodine dose in patients with Graves’ disease: a clinical outcome study. Eur J Nucl Med 2001; 28:1489–1495.

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446 Nuclear Medicine Communications 2006, Vol 27 No 5

9

Muratami Y, Takamatsu J, Sakane S, Kuma K, Ohshawa N. Changes in thyroid volume in response to radioactive iodine for Graves’ hyperthyroidism correlated with activity of thyroid stimulating antibody and treatment outcome. J Clin Endocrinol Metab 1996; 81:3257–3260. 10 Chiovato L, Fiore E, Vitti P, Rocchi R, Rago T, Dokic D, et al. Outcome of thyroid function in Graves’ patients treated with radioiodine: role of thyroidstimulating and thyrotropin-blocking antibodies and of radioiodine-induced thyroid damage. J Clin Endocrinol Metab 1998; 83:40–46. 11 Haase A, Bahre M, Lauer I, Meller B, Richter E. Radioiodine therapy in Graves’ hyperthyroidism: determination of individual optimum target dose. Exp Clin Endocrinol Diabetes 2000; 108:133–137. 12 Gomez-Arnaiz N, Andia E, Guma A, Abos R, Soler J, Gomez JM. Ultrasonographic thyroid volume as a reliable prognostic index of radioiodine-131 treatment outcome in Graves’ disease hyperthyroidism. Horm Metab Res 2003; 35:492–497.

13

Traino AC, Di Martino F, Grosso M, Monzani F, Dardano A, Caraccio N, et al. A predictive mathematical model for calculation of the final mass of Graves’ disease thyroids treated with 131I. Phys Med Biol 2005; 50:2181–2191. 14 Di Martino F, Traino AC, Brill AB, Stabin MG, Lazzeri M. A theoretical model for prescription of the patient-specific therapeutic activity for radioiodine therapy of Graves’ disease. Phys Med Biol 2002; 47:1493–1499. 15 Traino AC, Di Martino F, Lazzeri M. A dosimetric approach to patient specific radioiodine treatment of Graves’ disease with incorporation of treatment induced changes in thyroid mass. Med Phys 2004; 31:2121–2127. 16 Stabin MG. Internal radiation dosimetry. In: Hankin RE, et al. (editors): Nuclear Medicine, Volume I. St. Louis: Mosby; 1996, pp. 316–333. 17 Peters H, Fischer C, Bogner U, Reiners C, Schleusener H. Radioiodine therapy of Graves’ hyperthyroidism: standard vs. calculated 131-iodine activity. Results from a prospective, randomized, multicentre study. Eur J Clin Invest 1995; 25:186–193.

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Original article

Effects of gender and age on the quantitative parameters of [99mTc]pertechnetate salivary gland scintigraphy in normal subjects Fatih FIrat, Tevfik F. C ¸ ermik, Ali SarIkaya and Sakir Berkarda Aim To assess the effect of gender and age on [99mTc]pertechnetate salivary gland scintigraphy (SGS) in healthy subjects. Methods The study population consisted of 93 normal subjects (46 women, 47 men; age range 20–59 years). The subjects were categorized into eight (four female and four male) subgroups according to age decades. Dynamic SGS was performed after intravenous injection of 370 MBq [99mTc]pertechnetate. Anterior salivary gland images were acquired for 30 min. On the basis of the time–activity curves, three functional parameters were calculated for the parotid and submandibular salivary glands: (1) the first minute uptake ratio, (2) the maximum uptake ratio, and (3) the maximum secretion percentage. Results For women, all parotid and submandibular functional parameters had a peak in the fourth decade group. The comparison of four age groups in the female subjects showed significant differences, except for the third versus the fifth decades, at least for one parameter. The number of significant differences was highest in the comparison between the second versus the fourth decades. Among men, the highest values for all parotid

Introduction [99mTc]pertechnetate salivary gland scintigraphy (SGS) was introduced by Bo¨rner et al. more than 40 years ago [1]. Since then, it has been used in the diagnosis of a variety of salivary gland disorders as well as to determine the effects of some systemic disorders or therapies on the salivary glands, including Sjo¨gren’s disease, radiation therapy of head and neck tumours, radioiodine therapy of thyroid cancers, Bell’s palsy, end-stage renal disease, reflux oesophagitis, and primary salivary glands tumours [2–11]. [99mTc]pertechnetate SGS is an easy, safe and noninvasive method that can evaluate the major functions of salivary glands. However, no standard methods for the assessment of quantitative SGS have been established so far, and there are limited data published regarding the functional disturbance in a normal population and the role of age and gender. In particular, there is a need for further clarification regarding age and gender differences in healthy subjects. The aim of the present study was to determine useful quantitative parameters for the evalua-

and submandibular parameters were in the second decade, except for the first minute uptake ratio in the submandibular gland. The number of parameters that were significantly different among the four age groups in men was lower than in women. The first minute uptake ratio of the submandibular gland had the most significant differences among the groups of male subjects. Conclusion Age and gender differences have a significant effect on salivary gland functions which is more apparent in women than in men. Nucl Med Commun 27:447–453

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:447–453 Keywords: salivary gland scintigraphy, [99mTc]pertechnetate, normal subjects Hospital of the University of Trakya, Turkey. Correspondence to Dr Tevfik F. C ¸ ermik, Gu¨ilapog˘lu Yerles¸kesi 22030 Edirne, TURKEY Tel: + 001 215 662 3021; fax: + 001 215 349 5843; e-mail: [email protected] Received 13 January 2006 Accepted 20 February 2006

tion of salivary gland functions and to assess the effects of age and gender by using [99mTc]pertechnetate SGS in healthy subjects.

Subjects, materials and methods Subjects

The study population consisted of 93 normal subjects (46 women and 47 men; age range, 20–59 years). The subjects were divided into eight subgroups according to age decades: groups M2, M3, M4, M5, F2, F3, F4 and F5, as shown in Table 1. Subjects were excluded if they had a history of radiation therapy or surgery for head and neck tumours, or had a connective tissue or other systemic disease, or were taking any medication. The local ethics committee approved this investigation and each subject gave informed consent prior to participation in the study. Imaging protocol

Subjects fasted for a minimum of 4 h and then dynamic salivary gland scintigraphy was performed after the intravenous bolus injection of 370 MBq [99mTc]pertechnetate. A single-head gamma camera with a parallel hole,

c 2006 Lippincott Williams & Wilkins 0143-3636

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448 Nuclear Medicine Communications 2006, Vol 27 No 5

Table 1

The number of cases, age intervals and mean ± SD of the groups

N Age range (years) Mean ± SD (years)

Group F2

Group F3

Group F4

Group F5

Group M2

Group M3

Group M4

Group M5

11 20–29

12 30–39

12 40–49

11 50–59

12 20–29

13 30–39

11 40–49

11 50–59

26.1 ± 2.8

33.9 ± 2.1

43.5 ± 1.9

51.7 ± 2.8

23.3 ± 3.6

36.2 ± 2.6

44.6 ± 2.2

56.3 ± 3.2

F, female; M, male.

low energy, high resolution collimator and data analysis system (Orbiter; Siemens Inc., Erlangen, Germany) was used. The duration of the scan was 30 min, anterior salivary gland images were acquired sequentially, one frame every 20 s. Images were obtained in a 128  128 matrix with a zoom factor of 1.55. The energy window was 20% around the 140 keV photopeak of 99mTc. The head of each subject was fixed in a slightly extended supine position during imaging, using a hemi-cylindrical plastic holder. Salivary gland secretion was stimulated with 2 ml oral concentrated lemon juice (the sialogogue) instilled with a syringe at 20 min.

Fig. 1

B

RP

LP

RSM

LSM

Semi-quantitative analysis

Circular region of interests (ROIs) were drawn manually around both the parotid and submandibular glands and the temporal area to form the background on a total of 90 frames of summation images of dynamic scintigraphy. A time–activity curve for each salivary gland was obtained (Figs 1 and 2). On the basis of the time–activity curves, the following functional parameters were calculated for each salivary gland: (1) the first minute uptake ratio (FUR), which is c1/c2, where c1 is the mean counts in ROIs of salivary glands at the first minute after injection, and c2 is the mean counts in ROIs of background at first minute after injection; (2) the maximum uptake ratio (MUR), which is c3/c4, where c3 is the highest mean counts in ROIs of salivary glands after injection and c4 is the mean counts in ROIs of background synchronized with the highest mean counts in ROIs of the glands; and (3) the maximum secretion percentage (MSP), which is MUR c5 100; MUR where c5 is the lowest mean counts in ROIs of salivary glands after stimulation with the sialogogue. In addition, the percentage participation to the total function for these parameters of parotid and submandibular glands was calculated. In most cases, the uptake of the frame that showed the highest uptake was used for the calculation of the MUR before the sialogogue was applied. If there was a continuously increased uptake during the 20 min period, the frame just before the sialogogue was applied was accepted as the MUR value.

Thy Regions of interest on the summation image of dynamic scintigraphy. RP, right parotid; LP, left parotid; RSM, right submandibular gland; LSM, left submandibular gland; B, background; Thy, thyroid gland.

Fig. 2

Counts

M

S

L MU

F

LU

FU

0 1 min

20 min

30 Time

Schematic presentation of the time–activity curve for glandular activity in normal pattern on salivary gland scintigraphy. F = vascular perfusion peak at 1 min, M = maximum activity point before sialogogue stimulation, S = sialogogue stimulation point at 20 min and L = lowest activity point after sialogogue stimulation. FU, MU and LU = counts at F, M and L, respectively.

Data analysis

Conventional methods were used to generate descriptive statistics. A comparison of the scintigraphic ratios taken from the eight groups of subjects was carried out by using

the Kruskal–Wallis ANOVA and the Mann–Whitney U-test. P values less than 0.05 were considered to represent a significant difference between the groups.

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Effects of gender and age on salivary gland scintigraphy Firat et al. 449

Table 2 Mean ± SD, range values of trapping, uptake and secretion functions of age groups of female and male subjects, and participation rate to total functions of parotid and submandibular gland Mean ± SD (Min – Max) P-FUR P-MUR P-MSP %SP P-FUR %SP P-MUR %SP P-MSP SM-FUR SM-MUR SM-MSP %SP SM-FUR %SP SM-MUR %SP SM-MSP

Group F2

Group F3

1.74 ± 0.47 (1.18–2.96) 2.08 ± 0.50 (1.50–3.44) 45 ± 10 (26–64) 46 63 65 3.33 ± 0.87 (1.65–5.21) 1.99 ± 0.53 (1.40–3.65) 24 ± 12 (9–62) 54 37 35

1.63 ± 0.26 (1.27–2.11) 2.08 ± 0.49 (1.26–2.71) 53 ± 9 (38–69) 44 64 64 3.99 ± 1.47 (2.41–7.15) 2.28 ± 0.50 (1.41–3.67) 30 ± 9 (15–47) 56 36 36

Group F4

Group F5

Group M2

Group M3

Group M4

Group M5

2.07 ± 0.57 1.75 ± 0.37 2.00 ± 0.46 1.79 ± 0.43 1.91 ± 0.43 1.88 ± 0.33 (1.17–3.36) (1.24–2.44) (1.43–3.11) (1.20–2.89) (1.11–3.06) (1.41–2.51) 2.51 ± 0.61 2.03 ± 0.42 2.75 ± 0.93 2.27 ± 0.52 2.09 ± 0.48 1.83 ± 0.43 (1.33–3.42) (0.91–2.79) (1.44–4.60) (1.43–3.42) (1.42–3.58) (1.09–2.56) 53 ± 13 (33–72) 55 ± 10 (38–71) 55 ± 10 (30–72) 51 ± 11 (31–69) 52 ± 14 (24–68) 48 ± 11 (32–71) 45 46 42 44 48 51 59 67 63 57 66 62 57 66 62 58 63 58 4.46 ± 2.03 4.52 ± 1.94 4.08 ± 1.72 3.66 ± 1.05 4.26 ± 1.66 3.94 ± 1.22 (2.42–11.36) (1.51–8.24) (2.14–7.54) (1.96–5.45) (1.79–8.51) (2.61–6.50) 3.11 ± 1.09 2.18 ± 0.52 2.37 ± 0.81 2.78 ± 0.96 2.21 ± 0.38 2.45 ± 1.00 (1.53–4.83) (1.30–3.16) (1.35–3.78) (1.44–4.44) (1.53–2.89) (1.08–4.11) 40 ± 14 (19–64) 28 ± 9 (12–45) 34 ± 11 (19–55) 37 ± 12 (20–66) 30 ± 12 (12–48) 35 ± 14 (18–57) 55 54 58 56 52 49 51 33 37 43 44 38 43 44 38 42 37 42

P, parotid; SM, submandibular gland; FUR, first minute uptake ratio; MUR, maximum uptake ratio; MSP, maximum secretion percentage; %SP, split function.

Results Values for the FUR, MUR and MSP which were calculated from parotid and submandibular glands and their percentages of participation to the total function in female and male subjects according to the age decade groups are given in Table 2. In male and female subjects, the P values of the comparisons between the groups separately, and between each age decade group with regard to gender, are given in Table 3. Among the four age decade groups for women, except for parotid MSP, the highest values of FUR, MUR and MSP for both parotid and submandibular glands were observed in group 4F (Table 2). The comparisons of these four groups with each other showed significant differences for at least one parameter, except for the comparison of F3 versus F5. The highest number of significant parameters was found in the comparison of F2 versus F4 and F3 versus F4 (Table 4). In women, all parameters had peak values in the F4 group (Figs 3 and 4). In men, the highest values for all parameters were in group M2, except for FUR of the submandibular gland (Table 2). The numbers of significant parameters among the four age decades were lower among male groups than the female groups separately and the FUR parameter of the submandibular gland had the most significant differences (Table 4). Even though the highest functional parameter values were observed in the M2 group, it seems that age has less effect on salivary gland function in the male groups (Figs 5 and 6). In both genders, at all age decades, a high standard deviation (SD) and a wide range of functional parameter values were demonstrated in FUR, MUR and MSP. While the split function of the parotid gland to the total function in female groups was 44–46% for FUR, 59–67%

Table 3 P values obtained by statistical comparison between female and male age groups Parameter and gland

M–F

M2–F2

M3–F3

M4–F4

M5–F5

P-FUR P-MUR P-MSP

NS NS NS

NS NS 0.001

NS NS NS

NS NS NS

NS NS 0.02

SM-FUR SM-MUR SM-MSP

NS NS 0.03

0.003 NS 0.003

NS 0.02 0.02

0.006 0.001 0.01

NS NS 0.03

NS, not significant. Other abbreviations as in the footnote to Table 2.

Table 4 P values obtained by statistical comparison between age groups of the female subjects Parameter and gland

F2–F3

F2–F4

F2–F5

F3–F4

F3–F5

F4–F5

P-FUR P-MUR P-MSP

NS NS 0.01

0.03 0.01 0.04

NS 0.008 0.002

0.001 NS NS

NS NS NS

0.022 NS NS

SM-FUR SM-MUR SM-MSP

NS 0.04 NS

0.007 0.001 0.001

NS NS NS

0.005 0.001 0.004

NS NS NS

0.001 0.001 0.001

NS, not significant. Other abbreviations as in the footnote to Table 2.

for MUR and 57–66% for MSP, these ratios for male groups were 42–51%, 57–66% and 58–63%, respectively. While the FUR value was found to be higher in seven of eight groups for the submandibular gland, values of MUR and MSP were higher in all eight groups for the parotid gland (Table 2). When all 51 comparisons were evaluated, more significant differences were found in parameters of the submandibular gland than of the parotid. While 11 significant differences in 51 comparisons (22%) were calculated in

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Nuclear Medicine Communications 2006, Vol 27 No 5

Fig. 3

Female

Uptake ratios

6.0 5.5

P-FUR

5.0

P-MUR

4.5

SM-FUR

4.0

SM-MUR

3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 F2

F3

F4

F5

Age groups Distribution of perfusion and uptake function values of female age groups. P, parotid; SM: submandibulary gland; FUR, first minute uptake ratio; MUR, maximum uptake ratio.

Secretion %

Fig. 4

60 55 50 45 40 35 30 25 20 15 10 5 0

Female P-MSP SM-MSP

F2

F3

Age groups

F4

F5

Distribution of secretion ratios after sialogogue administration of female age groups. P, parotid; SM, submandibular gland; MSP, maximum secretion percentage.

the parotid, 26 (51%) significant differences were found in the submandibular gland (Tables 3, 4 and 5).

tional loss in salivary glands during the early stages increases the success of treatment or may lead to reevaluation of treatment modalities.

Discussion

The measurement of saliva production using objective techniques is based on the collection of saliva secretion from the gland orifice. However, this technique is difficult and requires special training. In contrast, SGS is a non-invasive method which enables the qualitative and quantitative evaluation of bilateral parotids and submandibular glands. Therefore, for more than 30 years it has been used to evaluate the functional course of diseases that affect the salivary glands.

Due to its antimicrobial effect, saliva is necessary not only for digestion and speech functions, but also for protection and maintenance of the oral mucous membrane and for dental health. Some systemic diseases such as Sjo¨gren’s disease, local salivary gland diseases, radiation therapy to the head and neck area, and radioiodine therapy may permanently damage the functions of salivary glands [2–7,9,10,12–15]. Therefore, the determination of func-

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Effects of gender and age on salivary gland scintigraphy Firat et al. 451

Uptake ratios

Fig. 5

6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0

Male P-FUR P-MUR SM-FUR SM-MUR

M2

M3

M4

M5

Age groups Distribution of perfusion and uptake function values of male age groups. P, parotid; SM, submandibular gland; FUR, First minute uptake ratio; MUR, maximum uptake ratio.

Secretion %

Fig. 6

Male

60 55 50 45 40 35 30 25 20 15 10 5 0

P-MSP SM-MSP

M2

M3

M4

M5

Age groups Distribution of secretion ratios after sialogogue administration of male age groups. P, parotid; SM, submandibular gland; MSP, maximum secretion percentage.

The correlation between 99mTc uptake into salivary glands and saliva secretion has been shown. Helman et al. established that 99mTc substitutes for Cl – in the Na + /K + /Cl – salivary co-transport system and therefore may serve as a measure of saliva secretion by using semiquantitative methods [16]. In semi-quantitative SGS studies, the calculation of three functions (perfusion, uptake and secretion) has been widely recognized. However, there is still controversy about the selection of the appropriate parameters to use in the calculation of these functions and many complex or simple parameters have been suggested [3,13,17–22]. In this study, three parameters calculated by simple methods were used to assess these three main salivary gland functions.

Table 5 P values obtained by statistical comparison between age groups of the male subjects Parameter and gland P-FUR P-MUR P-MSP SM-FUR SM-MUR SM-MSP

M2–M3

M2–M4

M2–M5

M3–M4

M3–M5

M4–M5

NS NS NS

NS NS NS

NS NS 0.02

NS NS NS

NS NS NS

NS NS NS

0.01 NS NS

0.002 NS NS

0.001 NS NS

NS 0.006 0.04

0.001 NS NS

0.04 NS NS

NS, not significant. Other abbreviations as in the footnote to Table 2.

The reason for selecting these simple methods was that they are used routinely. An imaging period of 30 min was chosen to complete the scintigraphic imaging because a

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452 Nuclear Medicine Communications 2006, Vol 27 No 5

longer time can be uncomfortable for the patient. An oral radioactivity index was not used in our study because it is not possible to differentiate which salivary gland produces the results obtained for an oral index. The effect of ageing on the production and secretion of saliva has been calculated by a saliva collection method for the first time and different results have been obtained in the studies. In 1982, Parvinen and Lamas [23] reported that age did not have an effect on stimulated flow rate and pH. Similarly, Tylenda et al. reported that rest and post-stimulation major salivary gland secretions did not decrease according to age [24]. In another study, where the cases were divided by 3-year time periods, unstimulated and stimulated flow rates of the parotid and submandibular gland were not affected by age and it was suggested that a decreased function obtained in elderly subjects may not be considered as physiological [25]. Jones and Ship [26] reported that flow rates from major salivary glands were independent of age, gender and race. However, there are also studies which report that ageing has a negative influence on salivary gland secretions. Pedersen et al. [27] found that salivary flow rates were significantly reduced in subjects aged over 70 years when compared to subjects who were younger than 40 years. Percival et al. [28] divided 116 healthy individuals into 20year time periods and in a comparison between the four age groups the secretion rates of unstimulated whole saliva were shown to decrease by age. However, there is no significant difference between the age groups for flow rates of stimulated parotid saliva. Further, in their study population, a significant decrease in flow rate was calculated for women rather than for men. As mentioned above, although studies have been published which describe the extensive use of quantitative 99mTc SGS, the results of functional parameters of healthy control subjects are still needed [18,29,30]. In a study in which post-menopausal and pre-menopausal women, as well as men older than 50 years of age and men younger than 50 years of age were compared, there was no difference between the groups in trapping, uptake and secretion functions [29]. In our study population, there was a significant difference only in the submandibular MSP when cases were divided according to gender and age. Despite significant differences in some values of the MSP and FUR for the parotid and the MUR of the submandibular gland in the comparison of the same age decade groups between women and men, we could not make a final decision about these findings because of the small number of subjects. In order to detect the functional differences between genders we suggest that the same age groups should be compared. However, this issue needs to be confirmed by further studies. In previously published studies with normal control subjects, the confusing findings are the variability of salivary gland uptake and secretion [31]. This variability does not

allow a clear-cut differentiation in the results between normal glands and abnormal glands to be obtained. The total uptake index has been developed to reduce this effect [18]. However, to calculate this index, blood samples need to be taken at three different times and use of the index is limited by the relatively complex calculation required. The same study showed that uptake into the submandibular gland was higher than into the parotid glands. This finding was supported in our study by a submandibular FUR value in seven age groups being higher than the parotid FUR, and by a higher participation percentage compared to the total function. In contrast, in the parotid, the MUR and MSP values and split function to the total function of these parameters were found to be high in all age groups in our study. The reason for this finding may be the existence of unstimulated spontaneous secretion in the submandibular gland. In our study group, little spontaneous secretion and reaction to sialogogue stimulation at high rates in the parotid were observed. Herman et al. reported different secretion behaviour between the parotid and submandibular gland and higher uptake and secretion rates in the parotid, similar to our study [30]. We believe it is important to obtain significant differences for each of the three parameters in both the parotid and submandibular glands between female age groups. Therefore, a comparison between the scintigraphic parameters of patients with Sjo¨gren’s disease, for example, with those of normal women in same age group may help to provide more accurate results and may enhance the value of semi-quantitative analysis. In contrast, in our study population, the effect of age on functions was limited in male subjects. However, it should be still considered in the evaluation of FUR values of the submandibular gland indicating differences between age groups. In the present study, the results of the calculated significant differences between both age and gender groups for the submandibular gland parameters may be important, given that Sjo¨gren’s disease affects the submandibular much more than the parotid [32,33]. Because of this, it is suggested that participants of the same age groups should be chosen for comparisons between normal subjects and patients in whom Sjo¨gren’s disease is suspected. One limitation of this study is that it did not include healthy subjects over 60 years of age but as salivary gland diseases, such as Sjo¨gren’s disease, begin during middle age the importance of this limitation might be reduced. The variability of normal salivary gland functions is also an important limitation for all parameters that we used. However, by using age groups divided into decades for the comparison, along with gender distinction, may help decrease this limitation.

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Effects of gender and age on salivary gland scintigraphy Firat et al. 453

Conclusion According to the results of data presented, it was clearly demonstrated that the age and gender differences have an effect on salivary gland function which was more evident in the submandibular gland functions of women. We also showed that this can be demonstrated by 99mTc SGS. It is suggested that a control group should be the same gender and in the same age decade as a patient group for accurate assessment when quantitative scintigraphic parameters are used although visual analysis is also important in patients who are being investigated for salivary gland functions.

Acknowledgements The authors would like to thank Serhan C ¸ ´yldavil and Engin Aytekin, technologists at the Department of Nuclear Medicine.

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References Bo¨rner W, Grunberg H, Moll E. Scintigraphic description of salivary glands of the head with technetium-99m. Med Welt 1965; 42:2378–2380. 2 Saito T, Fukuda H, Horikawa M, Ohmori K, Shindoh M, Amemiya A. Salivary gland scintigraphy with 99mTc-pertechnetate in Sjogren’s syndrome: relationship to clinicopathologic features of salivary and lacrimal glands. J Oral Pathol Med 1997; 26:46–50. 3 Bohuslavizki KH, Brenner W, Wolf H, Sippel C, Tonshoff G, Tinnemeyer S, et al. Value of quantitative salivary gland scintigraphy in the early stage of Sjogren’s syndrome. Nucl Med Commun 1995; 16:917–922. 4 Arrago JP, Rain JD, Brocheriou C, Rocher F. Scintigraphy of the salivary glands in Sjogren’s syndrome. J Clin Pathol 1987; 40:1463–1467. 5 Hughes PM, Carson K, Hill J, Hastings D. Scintigraphic evaluation of sialadenitis. Br J Radiol 1994; 67:328–331. 6 Liem IH, Olmos RA, Balm AJ, Keus RB, van Tinteren H, Takes RP, et al. Evidence for early and persistent impairment of salivary gland excretion after irradiation of head and neck tumours. Eur J Nucl Med 1996; 23: 1485–1490. 7 Bohuslavizki KH, Brenner W, Lassmann S, Tinnemeyer S, Tonshoff G, Sippel C, et al. Quantitative salivary gland scintigraphy in the diagnosis of parenchymal damage after treatment with radioiodine. Nucl Med Commun 1996; 17:681–686. 8 Chen SD, Kao CH, Chang CS, Chen GH. Salivary function in patients with reflux esophagitis: effect of cisapride. J Nucl Med 1998; 39:1449–1452. 9 Kao CH, Hsieh JF, Tsai SC, Ho YJ, Chang HR. Decreased salivary function in patients with end-stage renal disease requiring hemodialysis. Am J Kidney Dis 2000; 36:1110–1114. 10 Kaya M, Cermik TF, Ustun F, Sen S, Berkarda S. Salivary function in patients with chronic renal failure undergoing hemodialysis. Ann Nucl Med 2002; 16:117–120. 11 Yamashita T, Ino C, Tomoda K, Kumazawa T. Prognostic determination and submandibular function in Bell’s palsy. Dynamic testing with technetium Tc 99m. Arch Otolaryngol 1985; 111:244–248. 12 Umehara I, Yamada I, Murata Y, Takahashi Y, Okada N, Shibuya H. Quantitative evaluation of salivary gland scintigraphy in Sjogren’s syndrome. J Nucl Med 1999; 40:64–69.

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Hermann GA, Vivino FB, Shnier D, Krumm RP, Mayrin V. Diagnostic accuracy of salivary scintigraphic indices in xerostomic populations. Clin Nucl Med 1999; 24:167–172. Bohuslavizki KH, Brenner W, Klutmann S, Hubner RH, Lassmann S, Feyerabend B, et al. Radioprotection of salivary glands by amifostine in high-dose radioiodine therapy. J Nucl Med 1998; 39:1237–1242. Manthorpe R. New criteria for diagnosing Sjogren’s syndrome: a step forward? Scand J Rheumatol 2001; 115(suppl):14–20; discussion 20–22. Helman J, Turner RJ, Fox PC, Baum BJ. 99mTc-pertechnetate uptake in parotid acinar cells by the Na + /K + /Cl – co-transport system. J Clin Invest 1987; 79:1310–1313. Demangeat R, Didon-Poncelet A, Cherfan J, Demangeat JL. Stimulated salivary pertechnetate clearance revisited: correlation with dynamic scintigraphic indices in Sicca syndrome. Clin Nucl Med 2000; 25:888–894. Vigh L, Carlsen O, Hartling OJ. Uptake index and stimulated salivary gland response in 99Tcm-pertechnetate salivary gland scintigraphy in normal subjects. Nucl Med Commun 1997; 18:363–366. Stephen KW, Robertson JW, Harden RM. Quantitative aspects of pertechnetate concentration in human parotid and submandibular salivary glands. Br J Radiol 1976; 49:1028–1032. Mishkin FS. Radionuclide salivary gland imaging. Semin Nucl Med 1981; 11:258–265. Parret J, Peyrin JO. Radioisotopic investigations in salivary pathology. Clin Nucl Med 1979; 4:250–261. De Rossi G, Focacci C. A computer-assisted method for semi-quantitative assessment of salivary gland diseases. Eur J Nucl Med 1980; 5:499–503. Parvinen T, Larmas M. Age dependency of stimulated salivary flow rate, pH, and lactobacillus and yeast concentrations. J Dent Res 1982; 61:1052–1055. Tylenda CA, Ship JA, Fox PC, Baum BJ. Evaluation of submandibular salivary flow rate in different age groups. J Dent Res 1988; 67:1225–1228. Ship JA, Nolan NE, Puckett SA. Longitudinal analysis of parotid and submandibular salivary flow rates in healthy, different-aged adults. J Gerontol A Biol Sci Med Sci 1995; 50:M285–M289. Jones RE, Ship JA. Major salivary gland flow rates in young and old, generally healthy African Americans and whites. J Natl Med Assoc 1995; 87:131–135. Pedersen W, Schubert M, Izutsu K, Mersai T, Truelove E. Age-dependent decreases in human submandibular gland flow rates as measured under resting and post-stimulation conditions. J Dent Res 1985; 64: 822–825. Percival RS, Challacombe SJ, Marsh PD. Flow rates of resting whole and stimulated parotid saliva in relation to age and gender. J Dent Res 1994; 73:1416–1420. Chuang FJ, Lin CY, Sun SS. Evaluation of the effects of menopause and aging on salivary function by quantitative salivary scintigraphy. Mid Taiwan J Med 2003; 8:32–36. Hermann GA, Vivino FB, Shnier D, Krumm RP, Mayrin V, Shore JB. Variability of quantitative scintigraphic salivary indices in normal subjects. J Nucl Med 1998; 39:1260–1263. Bohuslaviski KH, Brenner W, Tinnemeyer S, Wolf H, Sippel C, Tonshoff G, et al. Normal data base for quantitative salivary gland scintigraphy. Radiol Oncol 1995; 29:297–305. Sugihara T, Yoshimura Y. Scintigraphic evaluation of the salivary glands in patients with Sjogren’s syndrome. Int J Oral Maxillofac Surg 1988; 17:71–75. Hakansson U, Jacobsson L, Lilja B, Manthorpe R, Henriksson V. Salivary gland scintigraphy in subjects with and without symptoms of dry mouth and/ or eyes, and in patients with primary Sjogren’s syndrome. Scand J Rheumatol 1994; 23:326–333.

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Original article

The effect on radiochemical purity of modifications to the method of preparation and dilution of 99mTc-sestamibi Maggie Coopera, Kate Dustanb and Laure Rotureauc Background 99mTc-sestamibi is a useful radiopharmaceutical for myocardial perfusion imaging, parathyroid imaging and breast tumour imaging. However, the preparation is time consuming and the limit of 3 ml on the volume of liquid that can be added to the Cardiolite kit vial means that it is often difficult to draw up small doses for patient studies. Aim To modify the method of preparation of 99mTcsestamibi in order to reduce the preparation time and to give a preparation which is more convenient for withdrawing patient doses. Method A modified kit was prepared by reconstituting a Cardiolite kit vial with 3 ml Sodium Chloride Injection (0.9%) BP, sub-dividing it into two separate nitrogen-filled vials before adding sodium pertechnetate and boiling for radiolabelling. 99mTc-sestamibi was also prepared according to the manufacturer’s recommended method and diluted with sodium chloride injection after preparation. Radiochemical purity was assessed by thin-layer chromatography and high-performance liquid chromatography. Results 99mTc-sestamibi prepared according to the manufacturer’s recommended method had high

Introduction

radiochemical purity (96.9% ± 1.1%) and retained > 90% radiochemical purity over 8 h following dilution. However, 99m Tc-sestamibi prepared by the modified method gave variable and inconsistent results. Conclusion The modified method of preparation was not robust enough to give reproducibly high radiochemical purity. However, dilution of 99mTc-sestamibi prepared according to the manufacturer’s recommended method was satisfactory. This study highlights problems with the analysis of 99mTc-sestamibi and the limitations of modifying the method of preparation. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:455–460 Nuclear Medicine Communications 2006, 27:455–460 Keywords: sestamibi, preparation, radiochemical purity Departments of aPharmacy, bNuclear Medicine, St. Bartholomew’s Hospital, London, UK and cParis XI University, France. Correspondence to Dr M. Cooper, Department of Pharmacy, St Bartholomew’s Hospital, London ECIA 7BE, UK. Tel: + 44 (0)20 7601 7153; fax: + 44 (0)20 7601 7146; e-mail: [email protected] Received 30 January 2006 Accepted 24 February 2006

Tc-sestamibi has been an important radiopharmaceutical for myocardial perfusion imaging for a number of years and, more recently, the licence for the product has been extended to include breast and parathyroid imaging. The radiochemical purity (RCP) of the radiopharmaceutical is obviously important to allow unambiguous interpretation of scans and accurate diagnosis. However, a recent study suggested that this may be more important when considering the interpretation of parathyroid images [1].

that the kit should be cooled for 15 min. This makes the preparation time-consuming compared to many other radiopharmaceutical kits, which require just the addition of sodium pertechnetate, shaking and incubation at room temperature for a few minutes. Since the kit is relatively expensive, many radiopharmacies, like our own, wish to prepare one 99mTc-sestamibi preparation using a single Cardiolite vial for use in more than one institution and therefore need to divide the product after the boiling and cooling phase. The time taken to do this can be constrictive and can interrupt the flow of work.

For the radiopharmacist, 99mTc-sestamibi (Cardiolite) is often considered to be a somewhat time-consuming kit to prepare. The 99mTc-sestamibi complex consists of one atom of 99mTc in a + 1 oxidation state surrounded by six molecules of 2-methoxyisobutylisonitrile (MIBI). In order to obtain the complex with 99mTc in this low oxidation state, it is necessary to boil the kit for 10 min at 1001C in a water bath to reduce the 99mTc to oxidation state + 1. The manufacturer’s instructions then state

A further complication in the preparation of 99mTcsestamibi is that (according to the manufacturer’s instructions) the total volume in the vial should be 1–3 ml. Thus, if the maximum permitted activity of 11.1 GBq is added to the kit vial then the concentration will be at least 3.7 GBq
ml – 1. This high concentration can make it difficult to draw up small doses (e.g., 185– 250 MBq) for cardiac imaging since the volume may be as little as 0.05 ml if the kit is used undiluted.

99m

c 2006 Lippincott Williams & Wilkins 0143-3636

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Nuclear Medicine Communications 2006, Vol 27 No 5

In view of the complications involved in preparing 99mTcsestamibi, we sought to modify the method of preparation to save time and to give a more suitable concentration for drawing up small doses of the radiopharmaceutical. However, we were concerned that any modification to the method should not cause a loss in RCP since this is vital for correct interpretation of scans. We also wanted to ensure that our current practice of preparing the kit and then diluting it was not detrimental to the RCP of the preparation.

and 2 ml to another nitrogen-filled vial (kit B). To kit A was added 4 GBq of sodium [99mTc]pertechnetate to give a final volume of 4 ml and to kit B was added 8 GBq of sodium [99mTc]pertechnetate to give a final volume of 8 ml. (These activities and volumes mimic the typical requirements of the nuclear medicine departments that our radiopharmacy supplies.) Following the addition of sodium [99mTc]pertechnetate the vials were heated for 10 min (n = 12) or 15 min (n = 8) at 1001C in a water bath and then cooled for 15 min prior to analysis.

Materials and methods

Analysis by thin-layer chromatography

Kit preparation

The RCP of the 99mTc-sestamibi was assessed by thinlayer chromatography (TLC) over a period of up to 8 h from the time of preparation as described below. The radioactivity in each section of the chromatography strips was measured in a dose calibrator (Capintec CRC-15R) or using a gamma counter (1282 Compugamma CS, LKB Wallac) as appropriate to the radioactivity on the strip using a cut-and-count method. Analysis was carried out in triplicate for each time point assayed. Two TLC methods were used, the manufacturer’s recommended method [2] and a two-strip method [3]. The second method was introduced because results using the manufacturer’s recommended method were very variable for 99mTcsestamibi prepared using our modified method; we sought to identify the nature of the impurities present by using the second TLC method.

Cardiolite kits were provided by Bristol Myers Squibb, sodium [99mTc]pertechnetate was obtained by elution of a 99Mo/99mTc Drytec generator (Amersham), the eluate was usually from the first elution of a new generator since this is the most common situation when preparing 99m Tc-sestamibi for routine use within our radiopharmacy. The eluate obtained was used within 2 h of elution. Nitrogen-filled 10 ml glass vials (product code N46) were also purchased from Amersham. 99mTc-sestamibi was either prepared according to the manufacturer’s instructions [2] (‘standard method’) followed by dilution, or by using the modified method described below. Standard method

Cardiolite kits were reconstituted with up to 11.1 GBq of sodium [99mTc]pertechnetate diluted to 3 ml with Sodium Chloride Injection (0.9%) BP. The vials were heated immediately for 10 min at 1001C in a water bath. Following heating, the 99mTc-sestamibi was cooled for 15 min at room temperature. If necessary the 99m Tc-sestamibi was then divided for use in two separate institutions. 99mTc-sestamibi was aseptically transferred to a nitrogen-filled vial and diluted with sodium chloride injection to give a final concentration of 750–1300 MBq
ml – 1 depending on clinical need. This standard method is routinely used in our radiopharmacy. Kit dilution

Following reconstitution of a Cardiolite kit with 11.1 GBq of sodium [99mTc]pertechnetate, boiling for 10 min and cooling for 15 min as above, 99mTc-sestamibi was diluted to either 1500 MBq
ml – 1 (high-activity kit) or 500 MBq
ml – 1 (low-activity kit) by transferring 2 ml to a nitrogen-filled vial and diluting to 5 ml with sodium chloride injection (high-activity kit) or by transferring 0.7 ml to a nitrogen-filled vial and diluting to 5 ml with sodium chloride injection (low-activity kit). 99mTcsestamibi was prepared in this way on three separate occasions. Modified method

Cardiolite kits were reconstituted with 3 ml of sodium chloride injection then 1 ml of the reconstituted cold sestamibi was transferred to a nitrogen-filled vial (kit A)

Method A

This is the manufacturer’s recommended method. One drop of ethanol was applied 1.5 cm from the bottom of a pre-cut 2.5  7.5 cm Baker-Flex aluminium oxide plate #1 B-F (J.T. Baker Inc.), using a 25-gauge needle, immediately followed by one drop of 99mTc-sestamibi. The spot was allowed to dry and then the TLC strip was developed to a distance of 5 cm from the spot with ethanol as mobile phase. The strip was then cut at a distance of 4 cm from the bottom and the activity in each half measured. 99mTc-sestamibi migrates to the top section of the strip and impurities remain in the bottom section. Method B

The method devised by Proulx et al. was used, [3]. This is a two-strip method using ITLC-SG as solid phase. One drop of 99mTc-sestamibi was applied to each of two ITLC-SG strips (Gelman), 1.5  9 cm, 1.5 cm from the bottom. The strips were run in 0.9% sodium chloride in water (sodium pertechnetate and other hydrophilic impurities migrates to the solvent front) and acetone (colloidal impurities remain at the baseline). The strips were developed to a distance of 6.5 cm from the spot and cut at a distance of 4.75 cm from the bottom of the strip (this is midway between the origin and the solvent front). The activity in each section was counted as before.

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Effect of preparation method on radiochemical purity of

Analysis by high-performance liquid chromatography

Reverse-phase high-performance liquid chromatography (HPLC) analysis was carried out on a Beckman 128 solvent deliver module with in-line UV and radiometric detection using a C18 column (Jupiter 5u C18 300A, 150  4.6 mm, 5 mm, Phenomenex) as stationary phase and a mobile phase of 45% methanol: 35% 50 mM ammonium sulfate: 20% acetonitrile running at 1 ml
min – 1 [4]. In this system 99mTc impurities elute early at 3.5–5.0 min and the peak corresponding to 99mTcsestamibi elutes at 8.2 min.

Results Standard preparation 99m

Tc-sestamibi prepared according to the manufacturer’s instructions and then diluted to 750– 1300 MBq
ml – 1 consistently showed high RCP after preparation (96.9% ± 1.1% (mean ± SD, n = 10)) when analysed by three different operators using the manufacturer’s recommended TLC method.

Kit dilution 99m

Tc-sestamibi prepared according to the manufacturer’s instructions and then diluted to either 500 MBq
ml – 1 or 1500 MBq
ml – 1 showed initially high RCP (94.9% ± 1.4 (n = 6) and 95.8% ± 2.3 (n = 6) respectively). 99mTc-sestamibi was analysed using the manufacturer’s recommended method on all occasions (Baker-Flex aluminium oxide plates run in ethanol). The high activity kit (1500 MBq
ml – 1) had acceptable RCP over the whole test period ( > 90%), showing a gradual loss in RCP over the 8 h of the test (Table 1). The RCP of the low activity kit (500 MBq
ml – 1) was consistently

99m

Tc-MIBI Cooper et al. 457

lower than for the high activity kit (except at the 8-h time point) but RCP was > 90% 8 h after dilution. Modified method Boiling for 10 min

There was a large amount of inter-assay and intra-assay variability in the measured RCP of 99mTc-sestamibi prepared with this modified method; that is, between results for products prepared in the same way but analysed on different occasions and between duplicate assays run on the same sample. This variability occurred using both of the TLC methods described. Purities as high as 98.9% were obtained on one occasion but more often < 90% RCP was observed. The results were extremely variable and showed no logical pattern either between 99mTc-sestamibi prepared on different occasions or between results for a given preparation either between duplicate samples or over the 8-h test period (Fig. 1). Using the two-strip TLC method [3], the main impurity identified was immobile impurities that remained in the lower section of the ITLC-SG strip run in acetone, but on some occasions, high levels of mobile impurities were 99m Table 1 Radiochemical purity (RCP) of Tc-sestamibi kits prepared according to the manufacturer’s recommended method then diluted to either 500 MBq
ml – 1 or 1500 MBq
ml – 1 (mean RCP ± SD, n = 6)

Time after preparation (h) 0 1 2 4 8

Low-activity kit (500 MBq
ml – 1)

High-activity kit (1500 MBq
ml – 1)

94.9 ± 1.4 94.5 ± 4.8 92.9 ± 1.9 89.9 ± 5.3 93.7 ± 2.2

95.8 ± 2.3 96.5 ± 3.3 94.1 ± 3.3 91.9 ± 2.1 91.8 ± 1.5

Fig. 1

% Radiochemical purity by TLC

120 100 80 60 40 20 0

0

1

2

3

4 5 6 Time after preparation (h)

7

8

9

Scatter graph of the radiochemical purity obtained for 99mTc-sestamibi prepared using the modified method (10 min boiling time) following analysis by manufacturer’s recommended thin-layer chromatography method. The graph shows the variability in results obtained for 18 different assays.

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Nuclear Medicine Communications 2006, Vol 27 No 5

also identified and there was a lack of consistency in the results. We also noted inter-operator and intra-operator variability in the results and it was not clear whether the low RCP often observed was due to operator error, the quality control method used or was, in fact, due to real impurities in the preparation. In order to assess this, we prepared 99m Tc-sestamibi using both the manufacturer’s method and our modified method and analysed the preparations using both TLC systems with two different operators. Our results show that using the manufacturer’s recommended method of preparation, the product had > 90% RCP at all test points whichever operator carried out the assay for both TLC systems (Table 2). However, when the modified method was used the results were very variable suggesting that the quality of the preparation was 99m Table 2 Radiochemical purity (RCP) of Tc-sestamibi prepared according to manufacturer’s recommended method using two different quality control systems

Method

Operator 1

Operator 2

Manufacturer’s method Two-strip method

97.1 ± 0.9 93.8 ± 1.0

97.7 ± 1.0 94.8 ± 1.1

The RCP was > 90% for the three preparations tested for both operators.

Fig. 2

590 000 Standard kit Kit A Kit B

490 000

cpm

Increasing the heating time did not resolve this problem of variability in the RCP observed for these kits. A series of reproducible results were obtained when one experienced operator both prepared the 99mTc-sestamibi and carried out the analysis using the manufacturer’s recommended method (Table 3). This operator was able to achieve a RCP of 95.3% ± 2% (n = 6) for kits containing 4 GBq in 4 ml and of 94.3% ± 2.1% (n = 6) for kits containing 8& GBq in 8 ml. However, on occasions when either the operator preparing the 99mTcsestamibi or the operator carrying out the TLC was changed, the RCP observed fell below 90% regardless of whether the operator was experienced or not. As for the method when the kits were boiled for 10 min, variability was observed between duplicate samples, between different test points and between different operators.

Tc-sestamibi is an important radiopharmaceutical for myocardial perfusion imaging as well as for breast and parathyroid imaging. High RCP is essential for reliable and accurate diagnosis. Various studies have looked into modifying the method of preparation in order to save time [4,5], by preparing 99mTc-sestamibi in a microwave oven or using an instant hot water machine, and have reported RCP > 90% for the product. Other studies have looked at sub-dividing the kit prior to radiolabelling in order to save money [6–8]. Our results clearly show that when the product is prepared according to the manufacturer’s recommended method a high RCP is obtained. In our hands, the RCP was well above the recommended

190 000

90 000

5

Boiling for 15 min

99m

290 000

0

Further evidence showing that impurities were present in the product was obtained by HPLC analysis. 99mTcsestamibi prepared according to the manufacturer’s recommended method showed the expected main peak at 8.2 min with some low level impurities eluting between 3.5 and 5.0 min constituting < 5% of the total radioactivity. However, 99mTc-sestamibi prepared using the modified method showed high levels of impurities eluting between 3.5 and 5.0 min (Fig. 2).

Discussion

390 000

−10 000

the cause of the variable results obtained rather than problems with the TLC method or with operator technique.

10 Time (min)

15

High-performance liquid chromatography radiochromatogram of (1) standard preparation: 99mTc-sestamibi prepared according to the manufacturer’s recommended method; (2) kit A: 99mTc-sestamibi prepared using the modified method of reconstituting with sodium chloride injection, fractionating the kit and then adding 4 GBq of sodium [99mTc]pertechnetate to a final volume of 4 ml and boiling for 10 min; and (3) kit B: 99mTc-sestamibi prepared as for kit A except by adding 8 GBq of sodium [99mTc]pertechnetate to a final volume of 8 ml.

99m Table 3 A series of six results of radiochemical purity of Tcsestamibi prepared using the modified method, boiling for 15 min, where the radiochemical purity was consistently > 90%

Time after preparation (h) 1 2 4 6 8

Kit containing 4 GBq in 4 ml

Kit containing 8 GBq in 8 ml

95.0 ± 2.0 94.8 ± 2.1 94.9 ± 2.3 94.7 ± 2.2 95.3 ± 2.1

93.6 ± 2.7 93.9 ± 2.5 93.2 ± 2.8 93.5 ± 2.5 94.3 ± 2.1

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Effect of preparation method on radiochemical purity of

90% limit at 96.9% even when the product was diluted after preparation. Our results show that dilution of the product following preparation in the standard way is not detrimental to the quality of our routine 99mTc-sestamibi preparations. There was a slight loss in RCP with time but over the 8 h of our study the RCP remained > 90%. We observed a high degree of inter-assay and intra-assay variability in the RCP of 99mTc-sestamibi prepared using our modified method either with boiling times of 10 or 15 min. Several studies have shown similar problems with fractionated kits. Varelis et al. [9] noted variability in RCP of 99mTc-sestamibi. They noted elevated amounts of a polar radiochemical impurity when they prepared 99mTcsestamibi using a wet column generator. They noted that the wet column generator eluate contains sodium nitrate, which is an oxidizing agent. Similarly, Hung et al. [10] found that the RCP was higher using a dry column generator than a wet column generator and they suggest that this may be due to traces of hydrogen peroxide and free radicals in the eluate from the wet column generator. Both sodium nitrate and hydrogen peroxide oxidize stannous ions present in freeze-dried kits and this can result in failure to reduce technetium to the correct oxidation state for the desired complex to form. It may be that this causes high levels of sodium pertechnetate to be present in the preparation or may give rise to other intermediate oxidation state complex impurities in the product. It is possible that these types of intermediate oxidation state complex impurities are being formed using our modified method. Thomson et al. [11] noted that the manufacturer’s recommended TLC method often overestimates the RCP because some of these impurities comigrate with 99mTc-sestamibi on the TLC strip: they found the presence of two lipophilic impurities, in addition to the more hydrophilic impurities that we noted, which probably co-migrate with 99mTc-sestamibi on TLC. These could give rise to the variable results that we found when analysing the kits by TLC. We do not believe that the impurities noted were simply [99mTc]pertechnetate because they did not consistently run to the top of the ITLC-SG strip eluted with 0.9% sodium chloride, and although the retention time on HPLC was similar to that for [99mTc]pertechnetate, this short retention time would also be expected for other hydrophilic impurities. We would speculate that these types of impurities are more likely to form in fractionated kits than when 99mTcsestamibi is prepared according to the manufacturer’s instructions due to oxidation of the relatively low levels of stannous ion present in the kit allowing only partial reduction of the technetium. We would speculate that

99m

Tc-MIBI Cooper et al. 459

this gives rise to other technetium isonitrile complexes. This idea is supported by several other studies. Decristoforo and Riccabona [8] fractionated Cardiolite kits in a manner very similar to our modified method. They dissolved the freeze-dried kit in 3 ml of low dissolved oxygen saline, split the kit into 3  1 ml fractions and froze the fractionated kits. They then reconstituted the thawed fractionated kits with sodium pertechnetate. They found that when they used different generators the RCP sometimes fell below the 90% limit. They suggest that higher levels of dissolved oxygen in the generator eluate may oxidize the stannous ions in the kit. Similarly, Baker [6] fractionated kits and found that the RCP fell below 90% when > 3 GBq of sodium pertechnetate was added to the kit. They modified the fractionated kits by adding extra stannous ion and improved the RCP to 97.4% ± 0.9% (n = 7). We did not use low dissolved oxygen saline in the preparation of our kits nor did we add additional stannous ion because we wanted to make the method simple, quick and in line with the procedures used for routine production in our radiopharmacy. Had we done so, it is likely that we would have seen a similar increase in RCP. However, such an additional modification would be time consuming and the aim of the study was to reduce the time required to produce 99mTc-sestamibi. Various studies have assessed different methods for analysing RCP. In these studies, artificially low RCP was usually achieved by mixing 99mTc-sestamibi, prepared according to the manufacturer’s recommended method, with sodium pertechnetate and/or reduced hydrolysed technetium [12–15]. However, by creating artificially low RCP in this way, the studies may have ignored the possibility of the presence of other intermediate oxidation state complex impurities in kits where the method of preparation has been modified and so they fail to assess whether these alternative analysis systems are able to identify these types of impurities. As Thomson et al. [11] point out, HPLC is a much more reliable way of identifying these types of impurities in the preparation. We found that in one operator’s hands, 99mTc-sestamibi prepared using our modified method with a boiling time of 15 min showed RCP consistently above the 90% limit, but, in view of the variability seen from other operators, we question whether these results represent the real level of impurities in the kits. We were not able to carry out HPLC on most of our samples due to limited availability of HPLC equipment so were unable to confirm the TLC results in every case. It seems likely that when the robustness of the formulation is impaired as a result of changes in the labelling procedure, then subtle variations in operator technique can either result in real changes in the quality of the preparations, perhaps through the introduction of variable amounts of air or errors in the RCP analysis itself. In any case, this

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460 Nuclear Medicine Communications 2006, Vol 27 No 5

modified method was not sufficiently robust to produce reproducible results with different operators. In view of this, we would not recommend using this modified method.

for kind provision of Cardiolite kit vials used in this study and Vadim Savov, Research Institute of Oncology, Minsk, Belarus, for his technical assistance.

Conclusion

References

99m

Tc-sestamibi prepared according to the manufacturer’s instructions with up to 11.1 GBq sodium [99mTc]pertechnetate consistently gives a product with high radiochemical purity. The product can be diluted prior to use to give a more convenient concentration and is stable for up to 8 h following dilution.

1

2 3 4

99m

Tc-sestamibi, prepared using a modified method involving fractionating a cold kit, reconstituting with sodium pertechnetate and boiling for 10 or 15 min at 1001C in a water bath gives variable radiochemical purity and the method is not robust enough to be recommended for routine use.

5 6

7 8

Our studies have highlighted problems with the analysis of 99mTc-sestamibi in that it is possible to miss 99mTc impurities present if TLC methods other than the manufacturer’s recommended method are used for analysis especially when these methods have not been thoroughly validated. Finally, we would recommend following the manufacturer’s recommended method for the preparation of 99m Tc-sestamibi.

Acknowledgements The authors wish to acknowledge the support and use of facilities of the Cancer Research UK Nuclear Medicine Research Laboratory. We also thank Bristol-Myers Squibb

9

10

11 12

13 14

15

Karam M, Dansereau RN, Dolce CJ, Feustel PJ, Robinson LW. Increasing the radiochemical purity of 99mTc sestamibi commercial preparations results in improved sensitivity of dual-phase planar parathyroid scintigraphy. Nucl Med Commun 2005; 26:1093–1098. Cardiolite package insert, Bristol-Myers Squibb S.r.l, Sermoneta, Italy, May 2004. Proulx A, Ballinger JR, Gulenchyn KY. Routine determination of radiochemical purity of 99mTc-MIBI. Appl Radiat Isot 1989; 40:95–97. Hung JC, Wilson ME, Brown ML, Gibbons RJ. Rapid preparation and quality control method for technetium-99m-2-methoxy isonitrile (technetium-99m-sestamibi). J Nucl Med 1991; 32:2162–2168. Wilson ME, Hung JC, Gibbons RJ. An alternative method for rapid preparation of 99Tcm-sestamibi. Nucl Med Commun 1993; 14:544–549. Baker RJ. Properties of Tc-99m radiopharmaceuticals prepared from fractionated cold kits utilizing a stannous ion augmentation technique [Abstract]. J Nucl Med 1996; 37:A325. Chowdhury S, Hung JC. Reconstitution and fractionation of Cardiolite kit into multi-dose cold kits for cost saving. J Nucl Med 1995; 36:A326. Decristoforo C, Riccabona G. Problems with fractionated cold kits. J Nucl Med 1996; 37:1912–1913. Varelis P, Parkes SL, Poot MT. The influence of generator eluate on the radiochemical purity of 99Tcm-sestamibi prepared using fractionated Cardiolite kits. Nucl Med Commun 1998; 19:615–623. Hung JC, Herold TJ, Gibbons RJ. Optimal conditions of 99mTc eluate for the radiolabeling of 99mTc-sestamibi. Nucl Med Biol 1996; 23:599–603. Thomson N, Lai L, Blower PJ. 99mTc sestamibi: what is the value of TLC quality control? Nucl Med Commun 2005; 26:75. Hung JC, Wilson ME, Gebhard MW, Gibbons RJ. Comparison of four alternative radiochemical purity testing methods for 99Tcm-sestamibi. Nucl Med Commun 1995; 16:99–104. Reily RM, So M, Polihronis J, Houle S. Rapid quality control of 99Tcmsestamibi. Nucl Med Commun 1992; 13:664–666. Patel M, Sadek S, Jahan S, Owunwanne A. A miniaturized rapid paper chromatographic procedure for quality control of technetium-99m sestamibi. Eur J Nucl Med 1995; 22:1416–1419. Luebke AL, Wilary DM, Mahoney DW, Hung JC. Evaluation of an alternative radiochemical purity testing method for technetium-99m-sestamibi. J Nucl Med Technol 2000; 28:259–263.

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Novel preparation and characterization of a trastuzumab– streptavidin conjugate for pre-targeted radionuclide therapy James E.S. Hainswortha, Peter Harrisonb and Stephen J. Mathera Introduction This study describes a novel and convenient route for the preparation of a trastuzumab–streptavidin conjugate such as might be used in a pre-targeting system and its in-vitro and in-vivo evaluation. Methods Trastuzumab was irradiated with UV light in the presence of stannous ions to reduce a number of the disulfide bridges to free thiol groups. A range of irradiation times were studied in order to quantify the number of thiols produced and to optimize the reduction process. The conjugate was then prepared by reaction with succinimidyl 4-(N-maleimidomethy cyclohexane)-1-carboxylate (SMCC)-linked streptavidin.

Her-2-expressing xenografts in nude mice was similar to that of labelled antibody. Conclusion Our results demonstrate a new, simple and effective method for the successful synthesis of antibody–streptavidin conjugates which could also be applied to many other heterodimeric protein conjugation c 2006 reactions. Nucl Med Commun 27:461–471
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:461–471 Keywords: pre-targeting, photoreduction, trastuzumab, monoclonal antibody

Results Initial conjugation reactions in phosphate buffer were inefficient, producing low conjugate yields, but conjugation reactions in triethanolamine-based buffer showed greatly increased conjugation yields. A high purity product (B100%) was obtained following purification by gel-filtration HPLC as determined by subsequent size exclusion HPLC analysis. The conjugate was shown to possess an essentially identical immunoreactivity to that of the native, unconjugated antibody and an unaltered biotin binding stoichiometry. Shedding and internalization by Her-2-expressing cells were low and the uptake in vivo by

a Cancer Research UK, Nuclear Medicine Group, St Bartholomew’s Hospital, London and bBioventix Ltd, Farnham, UK.

Introduction

the blood, it may be necessary to inject a clearing agent between the first and last reagents in order to reduce the amount of circulating antibody conjugate.

Conventional radioimmunotherapy requires the systemic administration of a radiolabelled antibody, in the hope that a sufficient proportion of the radioactivity will target and irradiate the tumour cells. However, while this approach has proven to be successful in a minority of tumours, most notably lymphoma, the amount of radioactivity that can be administered is limited by radiation to non-target tissues which either accumulate the labelled antibody (liver, kidney) or are irradiated by circulating antibody as it slowly clears from the blood (bone marrow). In order to increase radiation doses to the tumour while reducing the systemic radiation burden, pre-targeting approaches have been developed [1]. These involve the pre-administration of a tumour-specific antibody conjugate to establish secondary binding sites on the tumour which can then be more efficiently targeted by low molecular weight radiolabelled effector molecules. In order to prevent these effector molecules interacting with molecules of antibody conjugate still circulating in

Correspondence to Dr Stephen J. Mather, Department of Nuclear Medicine, St Bartholomew’s Hospital, London EC1A 7BE, UK. Tel: + 44 (0)207 601 7153; fax: + 44 (0)207 601 7143; e-mail: [email protected] Financial support for this study was provided by Cancer Research UK and the UK EPSRC. Received 19 January 2006 Revised 26 February 2006 Accepted 3 March 2006

Several research groups have used streptavidin-conjugated antibodies as part of the streptavidin–biotin pretargeting strategy [2] and have shown significant benefit over conventional radioimmunotherapy techniques [3]. However, the development of a pre-targeting approach is highly complex, requiring the preparation of a number of pharmaceutical targeting reagents and optimization of the doses and timings of administration of each. Any development which is able to simplify any of the stages in the procedure is therefore welcome. This publication is concerned with the design of the streptavidin-conjugated antibody used in the first stage in the process. It describes the development of a novel approach for conjugate preparation and its application for a novel receptor target which has previously not been employed using a pre-targeting approach, Her-2.

c 2006 Lippincott Williams & Wilkins 0143-3636


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462

Nuclear Medicine Communications 2006, Vol 27 No 5

Fig. 1

Streptavidin

Streptavidin

LY S

LY S

UV

O

NH Sn2+

S

SS

S-

SH SH

S

O

S

N

S S-

S-

O

S-S S-S S-S S

S

S-

S

S-

S S-

O

S-

S

S-

S

S-

S S-

O

N O

O

S H

S-S S-S S-S S S-

S

O O

N

O O

S H

S H

S-S S-S S-S S-S S-S

Na+O−

S H

Sn2+

SH

Sn2+

Sn2+

STAGE 1

STAGE 2

STAGE 3

Scheme showing the reactions involved in the trastuzumab–streptavidin conjugation. The antibody solution is irradiated by UV in the presence of stannous ions in order to reduce the disulfide bridges. Streptavidin is derivatized via lysine residues with SMCC. The maleimide group of SMCC is then reacted with the reduced antibody solution to form a heteroconjugate.

The HER2 (c-erbB2) proto-oncogene encodes a transmembrane receptor protein of 185 kDa, which is structurally related to the epidermal growth factor receptor. HER2/neu protein is expressed at low levels (2  105 to 5  105 molecules per cell) in a wide variety of normal tissues such as breast epithelia, endometrium, lung, gastrointestinal tract, kidney and central nervous system but is overexpressed at much higher levels (up to 2  106 molecules per cell) in 20–25% of primary breast cancers [4–6]. At this high level of expression, the kinase downstream of the receptor becomes constitutively activated probably due to auto-activation of p185HER2/ neu within the cell membrane [7]. This, in turn, produces mitogenic cell signalling which increases cell proliferation. Aberrant receptor activation resulting from gene amplification and overexpression has been correlated with reduced survival and reduced time to relapse in breast cancer patients and HER2 was identified as a potential drug target [8]. The humanized monoclonal anti-HER2 trastuzumab was the first targeted anti-cancer therapeutic agent based on genomic research, being approved by the FDA in 1998. Heterodimeric protein conjugates such as antibody– streptavidin have been conventionally prepared using

traditional protein chemistry techniques involving the use of bifunctional linkers. More recently, emphasis has been placed on the production of fusion proteins using increasingly available recombinant DNA techniques because these have the advantage of improved homogeneity and simple scale-up. Development and optimization of the procedures for fusion protein expression can, however, be time consuming and aberrant folding of domains may result in non-functional constructs. Such approaches are, therefore, not ideal for the preparation of a number of experimental conjugates on a small scale such as may be required in early pre-clinical development of a pre-targeting system. Many chemical conjugation approaches are also quite complex, however, requiring multiple steps that may involve purification and concentration of intermediates, resulting in limited control of the process and poor overall yields. The approach described herein comprises a three-step procedure as shown in Fig. 1 requiring only one intermediate purification and provides a relatively simple procedure that can be employed for the small-scale preparation of such conjugates for pre-clinical characterization.

Experimental procedures Chemicals and reagents

All reagents were purchased from Sigma–Aldrich unless otherwise indicated. Sodium [125I]iodide was purchased from ICN (Basingstoke, Hampshire, UK). The TSMZ

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Preparation of a trastuzumab–streptavidin conjugate Hainsworth et al. 463

buffer contained 80.2 mM triethanolamine, 98.9 mM sodium chloride, 1 mM magnesium chloride and 0.1 mM zinc chloride, pH corrected to pH 7.3 with concentrated sodium hydroxide.

remains at the origin while iodide migrates with an RF = 1.

High-pressure liquid chromatography

The number of free thiols per antibody was determined using two separate thiol quantification assays. In the first, Ellman’s reagent, 5,50 -dithiobis(2-nitrobenzoic acid) (DTNB) [11] was used, while in the second, 5-iodoacetamidofluorescein (5-IAF) [12] was used.

High-performance liquid chromatography (HPLC) analysis was performed using a Beckman 126 solvent-module pump system with a Beckman 168 diode array detector and a sodium iodide flow-through radiochemical detector attached to a Raytest gamma radioactivity monitor and a Bio-SEC-S3000 300  7.8 mm internal diameter column (Phenomenex, Cheshire, UK). Data collection was performed with a PC running System Gold Nouveau (Beckman, High Wycombe, UK). A mobile phase system comprising 0.1 M phosphate buffer with 2 mM ethylenediaminetetraacetic acid (pH 7.4) was used at a flow rate of 1 ml  min – 1. Preparative HPLC was performed using ¨ KTA Explorer connected to a Frac 950 fraction an A collector (Amersham Biosciences, Buckinghamshire, UK) and a Phenomenex HiLoad 16/60 Superdex 200 60-cm column. The mobile phase used was 0.1 M phosphate buffered saline (PBS) (pH 7.4) with a flow rate of 0.5 ml  min – 1. Photoactivation of the antibody

Reduction of the disulfide bridges in the antibody to free thiols in order to allow reaction with the maleimidederivatized streptavidin was performed by photoactivation as described by Stalteri and Mather [9]. A typical irradiation was performed as follows. A Medronate II kit (Amersham Biosciences, UK) was reconstituted with 2 ml of PBS or TSMZ and the entire volume was added to 2 ml of 10 mg  ml – 1 trastuzumab in 0.1 M PBS (at pH 7) or TSMZ in a sterile 10 ml nitrogen-filled borosilicate glass vial (Incstar Ltd, Wokingham, Berkshire, UK). The vial containing the antibody solution was placed centrally in the lamp unit of a Rayonet RMR 3000 photochemical reactor (Southern New England UV Company) and irradiation of the antibody was performed for predefined time periods. Radiolabelling of proteins

The trastuzumab–streptavidin (TSA) conjugate and trastuzumab were both radiolabelled with sodium [125I]iodide using the iodogen method [10]. Twenty micrograms of the protein was diluted to 100 ml with 0.1 M phosphate buffer pH 7. The solution was pipetted into an Iodogen tube followed by 1 ml (6 MBq) of sodium iodide. After a 10-min incubation at room temperature, the labelling mixture was removed and purified on a PD10 size exclusion column eluted with 0.5% bovine serum albumin (BSA) in PBS. The radiochemical purity of the labelled proteins was checked by instant thin-layer chromatography (ITLC) developed in 0.9% sodium chloride before use. In this system labelled protein

Thiol assays using Ellman’s reagent and 5-iodoacetamidofluorescein

Ellman’s assay

At the end of the irradiation (0, 5, 10, 20 min), 250 ml of a 4 mg  ml – 1 solution of DTNB was added to 250 ml (1.25 mg) of the antibody solution. The solutions were briefly mixed and then placed on a rotating mixer at 371C for 20 min after which the antibody was separated from any free DTNB or stannous ions by gel filtration using a PD-10 column (Amersham Biosciences, UK). The absorbance of the antibody solutions were measured at 412 nm on a Beckman DU 600 general purpose UV/visible spectrophotometer against 0.1 M PBS at pH 7. A standard curve was constructed by performing the same assay (without gel filtration) on a range of cysteine concentrations up to 200 mM in 0.1 M PBS. The concentration of thiol groups in the antibody solutions was determined by reference to the standard curve and the number of thiols per antibody molecule calculated by dividing the thiol concentration by the antibody concentration measured by absorbance at 280 nm, assuming a 1 mg  ml – 1 solution to have an absorbance of 1.4. Assay using 5-iodoacetamidofluorescein

A similar procedure was followed for the 5-IAF assay with 250 ml of an 8 mM solution of 5-IAF being added to 250 ml (1.25 mg) of antibody. The fluorescence of a 200 ml sample from each fraction collected from the PD-10 column was measured in a 96-well plate in a SpectraMAX Gemini microplate reader: a dual-monochromator spectrofluorometer system (Molecular Devices) together with blank solutions and a series of 5-IAF solutions (0–200 mM) to provide a standard curve. The data capture and analysis software SoftMax Pro 4.3 was used on an Apple Macintosh computer. A 5-IAF standard curve was constructed using a range of 5-IAF concentrations up to 200 mM. Conjugation of reduced trastuzumab with streptavidin-SMCC

Depending on the scale of the reaction, the required volume of a 20 mg  ml – 1 solution of streptavidin was mixed with a four-fold molar excess of sulfo-SMCC (4 mg  ml – 1) incubated at room temperature for 10 min and then added dropwise with continuous stirring to a two-fold molar excess of photoactivated antibody relative to the streptavidin. After incubation at room temperature for 2 h remaining free thiol groups were re-oxidized by

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464 Nuclear Medicine Communications 2006, Vol 27 No 5

adding a freshly prepared sodium tetrathionate solution to the antibody-conjugation mixture at a final concentration of 5 mM and incubating for 10 min. Identification of conjugation products

The product of the antibody–streptavidin conjugation was characterized by size exclusion HPLC and mass spectrometry. Size exclusion HPLC

HPLC analysis was performed using the chromatographic system described above after calibrating the column with five molecular weight standards ranging from 1350 Da to 670 kDa. A standard curve was generated by plotting retention time against log of molecular mass and the apparent mass of the conjugation products determined by reference to the standard curve. Tryptic digest and mass spectrometry

Ten microlitres of sample at 5 mg  ml – 1 were diluted with 15 ml of sample buffer and 15 ml samples separated for 90 min at 90 V on a vertical Pierce Precise 8–16% 10well native acrylamide gel (Perbio; Tattenhall, Cheshire, UK) in a BioRad Mini Trans Blot tank. After Coomassie staining, bands were cut out from the acrylamide gel using a scalpel and thoroughly dispersed in a 180-ml sample of 25 mM ammonium bicarbonate solution. The mixture was centrifuged and washed twice in ammonium bicarbonate and once with 180 ml aliquot of acetonitrile. After centrifugation and removal of the supernatant 200 ng trypsin in 500 ml ammonium bicarbonate solution was used to rehydrate the gel fragments for 4 h at 371C. The centrifuged gel fragments were dissolved in formic acid with sonication for 15 min before being dried in a SpeedVac. The dried residue was reconstituted in 5 ml of 50% ACN (acetonitrile)/0.1% TFA (trifluoacetic acid) (v/v) and 0.3 ml loaded on to the target plate of a 4700 Proteomics Analyser (Applied Biosystems) together with a 0.3 ml mixture of 3 mg  ml – 1 a-cyano matrix and the internal standard angiotensin I (25 fmol). Mass spectrometry (MS) was run in reflector positive mode. The collated MS and/or MS/MS data was used to search the NCBI database using Protein Prospector with a mass accuracy of 20 ppm. Conjugate binding of labelled DOTA–biotin conjugates 111

In-labelled biotin [13] was incubated with a series of dilutions of the conjugate ranging from 0.2 pM to 2 mM corresponding to biotin:streptavidin ratios of 10 – 5 to 100 : 1. The reaction mixtures were incubated for 30 min at 371C and samples were taken and analysed using Sephadex G-50 spin columns (Amersham Biosciences, UK) to determine the degree of streptavidin binding. Cell binding studies

The immunoreactive fraction of the radiolabelled antibody and antibody conjugate were measured using a

modification of the method described by Lindmo et al. [14]. Cells were harvested from two confluent tissue culture flasks of SKBR3 cells, re-suspended in 5 ml of cold 1% BSA/0.1 M PBS pH 7.4 and the concentration measured using a haemocytometer and light microscope. Five serial 1 : 2 dilutions of 500 ml of cells were prepared in duplicate and incubated with a 250 ml aliquot of the radiolabelled conjugate at a concentration of 50 ng  ml – 1. Non-specific binding was measured by co-incubation of the radiotracers together with 500 mg of unlabelled antibody. After incubation at room temperature on a rotating mixer for 2 h the tubes were centrifuged and the cell pellets washed twice with cold 1% BSA/0.1 M PBS before counting in a 1282 Compugamma gamma counter (LKB; Wallac, Finland). The total radioactive counts added to each tube were divided by the cell-bound counts after subtraction of the non-specifically bound counts and plotted against the reciprocal of cell concentration for each sample. After linear regression analysis using Origin software, the immunoreactive fraction was obtained from the reciprocal of the intercept on the y-axis. The extent and rate of shedding or internalization of the antibody and antibody conjugate were also studied with SKBR3 cells. Cells were grown to confluency in six-well plates and washed with nutrient media. A volume of 150 ml of 125I-labelled conjugate (250 ng  ml – 1) was added together with 1.2 ml of media to each well either in the absence or presence of 150 ml of unlabelled trastuzumab (400 mg  ml – 1). The plates were incubated at 371C for various time points after which triplicate wells were analysed in various ways. (1) To assess shedding, the plates were maintained at 371C for a further 24 h and at various time points the supernatant was removed, the cells washed twice with ice-cold media and cells were lysed with 1 ml of 0.5 M NaOH for 15 min and counted to determine the amount of bound radioactivity at each time point. (2) To determine the rate and extent of antibody dissociation the cell supernatant was removed and replaced either with fresh media or media containing a 1500-fold excess of cold trastuzumab. The plates were then maintained at 371C for a further 48 h and at various time points the supernatant was removed, the cells washed twice with ice-cold media and cells lysed and counted. (3) To assess antibody internalization, the supernatant was removed, the cells washed twice with ice-cold media and the solutions combined and counted to determine (by subtraction from the total counts added) the total amount of cell-associated antibody. The cells were then washed twice with 1 ml of 0.05 M glycine-HCl solution at pH 2.8–1.8 and 1 ml of 0.5% BSA/PBS and the washings combined and counted to determine the amount of radioactivity membrane bound. The cells were finally lysed and counted to determine the amount of internalized radioactivity at each time point.

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Preparation of a trastuzumab–streptavidin conjugate Hainsworth et al. 465

Fig. 3

35 30 25 20 15 10 5 0 −5

MAb, Tin, UV (Fluorimeter) MAb, Tin, UV (Spectrophotometer) MAb, Tin, no UV (Fluorimeter) MAb, Tin, no UV (Spectrophotometer) MAb, No Tin, UV (Fluorimeter) MAb, No Tin, UV (Spectrophotometer)

0

5 10 15 UV irradiation time (min)

mAU

Thiols/Ab

Fig. 2

20

Thiol concentration assays using either 5,50 -dithiobis(2-nitrobenzoic acid) (Ellman’s reagent) or 5-iodoacetamidofluorescein (a thiol-specific fluorescent label). Both techniques were used after irradiation of antibody in solution under either reducing or control conditions. The concentration of the thiol-reactive probes was measured and the number of thiol groups per antibody produced was calculated. MAb, monoclonal antibody.

Biodistribution studies

All animal studies were performed in compliance with the UK Animals (Scientific Procedures) Act 1986 and the Code of Practice for the Housing and Care of Animals used in Scientific Procedures (Home Office, UK). The biodistribution of 125I-labelled TSA was compared with that of 125I-labelled trastuzumab in nude mice bearing human breast cancer xenografts. Female nude mice were injected subcutaneously with 500 000 MDA-MB-435S cells and small tumours were allowed to develop. Once these reached a maximum diameter of 4 mm in any direction the mice were killed, the tumours resected and cut into smaller tumours for subcutaneous implantation in recipient mice after which they were allowed to develop until a few millimetres in diameter. Eight such mice were injected with 80 kBq of radiolabelled TSA and a further eight with 80 kBq radiolabelled trastuzumab. After a delay of 24 and 48 h, four mice were killed from each group and samples of blood, muscle, femur and the entire kidneys, liver, gastrointestinal tract, stomach, spleen and tumour resected, weighed and counted in a gamma counter together with samples of a dilution of the injected material in order to allow calculation of the proportion of the injected dose in each sample.

Results Photoactivation of trastuzumab

Free thiol groups were generated in trastuzumab for subsequent conjugation to streptavidin as shown in Fig. 1 by irradiation in a UV reactor in the presence of stannous ions. In order to control the number of thiol groups produced, the effect of different radiation times was studied initially. To measure the number of thiol groups produced under these irradiation conditions, two assays were compared, the well established colorimetric Ellman’s assay and a less commonly used fluorescent assay

120000 110000 100000 90000 80000 70000 60000 50000 40000 30000 20000 10000 0 −10000

Trastuzumab Trastuzumab post-UV-irradiation Trastuzumab post-UV-irradiation plus air cooling

0

5

10

15 Time (min)

20

25

30

Size exclusion HPLC chromatogram analysis of a sample of trastuzumab photoactivated by UV in the presence of stannous ions. Air cooling the irradiation reactor during reaction was found to prevent the formation of aggregated antibody (seen in front of the main antibody peak).

based on the use of 5-iodoacetamidofluorescein. Similar results were obtained with both assays as shown in Fig. 2. Irradiation for 10 min resulted in a partial reduction with the generation of six to eight thiols per antibody molecule while exposure for 20 min resulted in almost complete reduction with the production of around 30 thiols per antibody. Irradiation in the absence of tin produced no reduction as did treatment with stannous ions in the absence of UV irradiation. An irradiation time of 10 min was chosen for routine application. Antibody irradiated for this period was analysed by size exclusion HPLC whereupon a small degree of antibody aggregation can be seen, as shown in Fig. 3. It was thought likely that this was caused by heating during the irradiation process since the vials are palpably warm to the touch on removal from the reactor. To overcome this problem a system of forced air ventilation through the reactor during irradiation was implemented. No aggregation of antibody irradiated under these conditions was observed, as shown in the figure. Conjugation of reduced trastuzumab to streptavidin

The reduced antibody was mixed with streptavidin that had been pre-treated with the heterofunctional crosslinking reagent sulfo-SMCC in order to generate maleimido functionalities thereon. Various molar ratios of trastuzumab to streptavidin were studied initially on a small (100–200 mg) scale. Reactions were initially performed in PBS, but under these conditions only low conjugation yields were obtained. Size exclusion HPLC analysis of a typical reaction mixture using an antibody: streptavidin ratio of 1 : 2 is shown in Fig. 4. In this example only about 8% of the protein elutes with retention times shorter than that of the unconjugated antibody. Extensive investigation failed to identify the cause of these low conjugation yields but changing the

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466 Nuclear Medicine Communications 2006, Vol 27 No 5

Fig. 4

Fig. 5 PBS buffered conjugation reaction

1.2

Trastuzumab-streptavidin conjugate (not to scale) Trastuzumab

AU (@ 280 nm)

1.0

mAU

2.4 TSMZ buffered conjugation reaction 2:1 MAb: streptavidin reaction (TSMZ) 2.2 2.0 1.8 1.6 1.4 Streptavidin-trastuzumab 1.2 Trastuzumab 1.0 0.8 Multiple streptavidin 0.6 derivatised trastuzumab 0.4 0.2 0.0 −0.2 0 5 10 15 20 25 Retention time (min)

0.8 0.6 0.4 0.2 0.0 0

30

Size exclusion HPLC analyses of conjugation reactions at antibody : streptavidin ratios of 1 : 2 performed in phosphate buffered saline (PBS) (red) and TSMZ buffer (black) solutions showing much greater yields when TSMZ is used. Conjugation at an antibody : streptavidin ratio of 2 : 1 (blue) results in a reduction of the number of multi-derivatized antibody molecules. (See the text for the composition of TSMZ buffer.)

buffer system from PBS to one comprised of triethanolamine, sodium chloride, magnesium chloride and zinc chloride (TSMZ) did result in much higher yields. Using reaction conditions otherwise identical, changing the buffer from PBS to TSMZ improved the yields four-fold. An example is again shown in Fig. 4 in which approximately 40% of the area under the curve was represented by newly synthesized species. However, the downside of this increased yield is the increased heterogeneity of the conjugate product. Additional peaks are present on the HPLC analysis caused by the presence of antibody molecules derivatized with more than one molecule of streptavidin. In order to overcome this problem, reactions employing a molar excess of antibody were studied. The results of size exclusion HPLC analysis of such a reaction (antibody : streptavidin, 2 : 1) are also shown in Fig. 4. It can be seen that, although the conjugation yield falls to 20%, the proportion of conjugate molecules eluting ahead of the main conjugate peak is greatly reduced. Following this small-scale optimization, the reaction was scaled up and conjugations were performed using 30 mg of trastuzumab and 4.5 mg of streptavidin with similar yields. In order to purify the TSA we explored the use of two purification strategies: the iminobiotin column proposed by Hylarides et al. [15] and semi-preparative size exclusion chromatography. Both methods produced very similar results but since the iminobiotin chromatography requires exposure of the preparation to pH 11 and the size exclusion chromatography is performed at neutral pH, we decided to employ the latter approach as

5

10

15 20 Time (min)

25

30

Size exclusion HPLC analysis of the purified trastuzumab–streptavidin conjugate product compared with that of the unconjugated antibody.

described above. The antibody–streptavidin conjugate obtained from this purification was then submitted to analytical size exclusion HPLC with the result shown in Fig. 5. It can be seen that the compound elutes with a retention time some minutes shorter than that of the unconjugated antibody. The symmetrical profile of the peak and the absence of earlier eluting impurities indicate the high purity of the conjugate product. Characterization of the trastuzumab–streptavidin conjugate

Two analytical techniques were employed to characterize the identity of the conjugate. Calibration of the size exclusion column was performed using protein standards of known molecular weight and the graph of retention time (RT) versus log10 molecular weight was found to be linear over the range 44–670 kDa and could be assigned the equation log (MWt) = (RT – 37.8)/ – 4.32. Since the conjugate elutes with a retention time of 14.6 min its calculated molecular weight is 238 kDa which corresponds very well with the combined molecular weight of one molecule of trastuzumab (185 kDa) and one molecule of streptavidin (50 kDa) of 235 kDa. Since a very similar mass could be achieved by conjugation of consitutent chains of the antibody alone (for example addition of one heavy chain) further analysis was performed to confirm the identity of the constituent proteins in the conjugate. After further purification on a ‘native’ polyacrylamide gel the conjugate was digested with trypsin and a ‘fingerprint’ of digestion products obtained, which, after analysis by mass spectroscopy were compared to those in the NCBI database [16]. This analysis showed the presence of sequences from Fab and heavy-chain fragments of trastuzumab plus those from streptavidin-conjugated Fab, confirming that the conjugate does indeed consist of one molecule of streptavidin conjugated to one molecule of trastuzumab.

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Preparation of a trastuzumab–streptavidin conjugate Hainsworth et al. 467

The ability of the conjugate to capture biotin was assessed by mixing a range of concentrations of the protein with a known concentration of 111In-DOTAbiotin and separating the bound from free using size exclusion spin columns. The results show a sigmoidal pattern of binding as seen in Fig. 7. With a large molar excess of streptavidin up to 99% of the 111In-DOTAlysine-biotin (DLB) was protein bound. At a DLB : streptavidin ratio of 4 : 1 (the expected stoichiometric ratio of biotin binding) approximately 98% of the 111In-DOTAbiotin is protein bound indicating that conjugated streptavidin has fully retained its biotin-binding functionality after conjugation. Shedding and internalization studies on the radiolabelled trastuzumab–streptavidin conjugate

To function well as a pre-targeting vehicle it is essential that the conjugate remains bound to the surface of the cell for as long as possible and is neither shed nor internalized. The extent to which these events occur was investigated with Her-2-expressing SKBR3 cells in vitro. Kinetic binding studies at 371C showed that binding equilibrium was reached 3 h after addition of labelled TSA but thereafter the level of binding remained unchanged for at least 24 h showing no evidence of shedding of antibody receptor complexes into the supernatant (Fig. 8). Cells were also first incubated with radiolabelled antibody for 2 h after which the medium containing unbound antibody was removed and replaced either with fresh medium or medium containing an excess of unlabelled trastuzumab antibody. The amount of antibody that remained bound to the cells was monitored over a period of 48 h. There was a slow, exponential release of the conjugate from the cells with 50% dissociation at 24 h and 75% at 48 h. The rate of

35 Total counts/counts bound

The conjugate has two binding functionalities which must be fully retained if the pre-targeting is to be effective; that is, the ability to bind to the target epitope on the tumour and to subsequently administered radiolabelled biotin. To determine whether the immunoreactivity of the antibody portion of the conjugate had been compromised the binding of radiolabelled TSA to Her-2-expressing SKBR3 cells was compared with that of radiolabelled native trastuzumab. This was performed according to the method described by Lindmo et al. [14]. A Burke–Lineweaver plot of the binding data generated by this assay allows the immunoreactive fraction (i.e., the fraction of antibody bound to a theoretically infinite excess of antigen) to be determined from the intercept through the y-axis. The results are shown in Fig. 6. The calculated immunoreactive fraction for the conjugate was 76% compared with 72% for the antibody itself indicating that the antibody retains its epitope-binding functionality after site-selective conjugation to streptavidin.

Fig. 6

I 125 −trastuzumab-streptavidin conjugate I 125 −trastuzumab

30 25 20 15 10 5 0 0

2

4

6

8 10 Dilution factor

12

14

16

18

Immunoreactivity assay of the labelled conjugate and native trastuzumab on SKBR3 cells. The immunoreactive fraction is obtained from the reciprocal of the intercepts of both lines with the y-axis after linear correlation. The conjugate intercept is 1.32 and the trastuzumab intercept is 1.39 corresponding to immunoreactive fractions of 76% and 72%, respectively.

Fig. 7

100

Percentage111 In-DLB bound

Binding assays on the trastuzumab–streptavidin conjugate

80 Corresponds to a 1:4 ratio of T5A:biotin

60 40 20

1E-7

1E-6

1E-5

1E-4

1E-3

Molar ratio of TSA to

0.01

0.1

111

In-DLB

Binding curve indicating the percentage of 111In-labelled biotin bound to the conjugate at a range of molar ratios. Close to 100% binding is observed at a biotin : TSA molar ratios of 4 : 1 indicating that the conjugation process does not influence the biotin-binding capacity. (TSA, trastuzumab–streptavidin; DLB, DOTA-lysine-biotin)

dissociation was, as expected, greatly increased in the presence of competing antibody. The extent to which the dissociated radioactivity was protein bound was measured using TCA precipitation. In the first 24 h, about 8% of the radioactivity was not protein bound. The extent to which the antibody conjugate may internalize after binding to the receptor was assessed by incubating the cells with the radiolabelled conjugate for up to 2 h and then removing surface-bound antibody with an acid wash and measuring the amount of radioactivity

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468 Nuclear Medicine Communications 2006, Vol 27 No 5

Fig. 8

120

A B C

Activity cell bound (%)

100

125 Table 1 Biodistribution of I-trastuzumab and 125I-TSA in nude mice bearing MDA-MB-435S xenografts expressed as either the percentage of injected dose per gram of tissue or per organ plus (standard deviation)

60

40

125

I-trastuzumab

24 h Blood Muscle Bone Tumour Organ Kidney Liver Gut Stomach Spleen

80

125

Tissue

48 h

24 h

I-TSA 48 h

14.4 1.2 1.2 5.7

(3.6) (0.3) (0.8) (1.0)

12.5 1.2 1.8 17.2

(4.1) (0.6) (1.4) (27.5)

7.5 0.7 1.7 6.8

(2.5) (0.1) (1.4) (7.4)

5.3 0.4 0.7 12.5

(2.8) (0.1) (0.7) (13.8)

3.7 4.8 1.7 1.3 5.4

(0.3) (3.5) (0.4) (0.6) (1.3)

3.9 3.8 1.3 2.5 5.6

(3.4) (0.9) (0.4) (3.2) (6.0)

3.1 3.6 1.0 1.7 3.2

(0.9) (0.7) (0.2) (0.9) (0.9)

4.0 2.9 0.7 1.0 2.7

(2.2) (0.7) (0.3) (0.2) (1.3)

TSA, trastuzumab–streptavidin conjugate.

20

0

0

10

20

30 Time (h)

40

50

60

Dissociation of radiolabelled conjugate from SKBR3 cells (A) under equilibrium binding conditions (B) after replacement of non-bound TSA with fresh media (C) after replacement of non-bound TSA with fresh media containing a 1500 excess of cold antibody.

otherwise there are no major differences between the amount of radioactivity in any other normal tissues or organ. The percentage of injected activity retained by the tumours is also similar for each tracer although the variation, especially at 48 h, is large.

Discussion

that remain associated with the cells. For this assay acidwash solutions with a pH range from 2.8 to 1.8 were studied. Following acid wash at pH 2.8 about 30% of the radioactivity remained associated with the cells, but washing with pH 2.3 or 1.8 reduced this to 8–10%. These results combined with those shown in Fig. 8 indicate a low but significant level of antibody internalization under the in-vitro conditions employed. In-vivo targeting

A preliminary study to assess the ability of the conjugate to target the Her-2 receptor in vivo was performed. The ability of a number of Her-2-expressing cell lines including SKBR3, BT474, MDA-MB-361 and MDA-MB435S to grow as solid tumours in nude mice was assessed following sub-cutaneous injection of between 500 000 and 1 million cells. The most efficient take was seen with MDA-MB-435S cells. ‘Parent’ tumours were first established in a small number of animals and these were then transplanted into a second series for the biodistribution studies. Groups of tumour-bearing mice were killed 24 and 48 h after the intravenous injection of tracer amounts of either radiolabelled TSA or radiolabelled trastuzumab and the tumours and samples of tissues and major organs were removed and counted in a gamma counter. The percentage of injected dose per organ or gram of tissue was calculated and the results are shown in Table 1. It can be seen that the blood clearance of the conjugate is considerably faster than that of the antibody but

Trastuzumab has been demonstrated as one of the few humanized monoclonal antibodies that has clinical value and potential [17]. Trastuzumab pre-targeting could combine the antibody-dependent cell cytotoxicity activity of trastuzumab with an added radiation dose to tumours in the same way that Zevalin provides an additive effect to rituximab [18]. The aim of this work was therefore to synthesize and evaluate a trastuzumab–streptavidin conjugate for the intended purpose of pre-targeting HER2-receptor positive tumours. Numerous approaches have been taken to the preparation of heteroconjugates of proteins such as antibody–streptavidin. Typically, this has involved crosslinking the proteins via lysine amino groups using a suitable bifunctional linker [19]. The major disadvantage of this approach is the random substitution pattern brought about by the widespread distribution of lysine residues throughout the antibody sequence. Site-specific conjugation approaches via hinge or engineered thiols [20,21] or carbohydrate side chains [22] have the advantage that they preserve the immunoreactivity of the resulting conjugate since the conjugation is directed to a site distant from the complementary determining regions of the antibody. Others have used conventional chemical reduction strategies in order to generate reactive free thiols from the disulfide bridges in the hinge region of the immunoglobulin. We have previously shown that the use of photoactivation has advantages in simplifying the number of steps required to perform reduction-mediated antibody conjugation [23].

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Preparation of a trastuzumab–streptavidin conjugate Hainsworth et al. 469

We therefore decided to pursue this route for the preparation of the trastuzumab–streptavidin conjugate for pre-targeting. The results of this study confirm our previous findings that photoactivation requires the presence of stannous ions to complete the disulfide reduction and that varying the time of irradiation allows the extent of reduction to be controlled [9]. Since the aim was to produce a conjugate with 1 : 1 stoichiometry, only a small number of free thiols are required and excessive exposure of the protein to UV irradiation is likely to produce generalized damage to its structure and function. In order to control the extent of reduction, an assay which measures the number of thiol groups produced is required. The wellknown Ellman’s assay [24], based on the production of a coloured thionitrobenzene–antibody adduct, has been widely used for this purpose but its disadvantage in the current situation is that the presence of tin results in a high background against which small numbers of thiol groups cannot be accurately measured. In order to overcome this problem the antibody adduct can be purified from low molecular weight contaminants by size exclusion chromatography before colorimetric measurement but this represents an undesirable additional step and may produce artefactual results. For this reason, a method was adapted from the paper by Muddukrishna et al. who used the more thiol-specific 5-iodofluorescein [12]. This paper describes a convenient HPLC-based assay using on-line UV detection of the antibodyconjugated fluorescein. Unfortunately, we were unable to reproduce this assay in our laboratory due to the fact that the analytes were retained by our HPLC columns, although we were able to adapt the procedure to an opencolumn gel-filtration procedure with subsequent analysis of fractions off-line. This also allowed us to use fluorescence detection, which is more sensitive than UV absorbance. In practice, however, the results obtained by the two assays were extremely similar as shown in Fig. 2. UV irradiation for 10 min resulted in the generation of six to eight thiol groups. Although this is probably more than the number absolutely required, it was felt that some excess was desirable provided that the integrity of the antibody was not affected. Size exclusion HPLC analysis of the partially reduced antibody showed no fragmentation but there was initially some evidence of aggregation (Fig. 3). However, this could be prevented by air cooling of the environment during the irradiation period. These results confirm that photoreduction is a highly controllable and reproducible technique for the production of free thiol groups in monoclonal antibodies. As a starting point for the subsequent conjugation we used the publications of Karacay et al. [25] and Hylarides et al. [15] who used similar conditions to prepare streptavidin conjugates based on chemically reduced

antibodies. The initial yields we obtained in a phosphate buffer system were, at around 10%, unexplainedly lower than the 37% achieved by Hylarides and the 20–25% by Karacay. However, an alternative triethanolamine buffer system, TSMZ, for reasons not yet elucidated, produced much higher yields of around 40%. We explored the effect of varying the streptavidin to antibody ratio and analysed the resulting mixtures by size exclusion chromatography. An excess of streptavidin resulted in higher yields but a relatively heterogeneous mixture of products as shown in Fig. 4. An excess of antibody, on the other hand, gave lower yields but a more homogeneous product. Since our primary aim was to prepare a well-characterized product for pre-clinical optimization, we felt that it was preferable to sacrifice yield to achieve higher quality and we therefore chose a streptavidin : antibody ratio of 1 : 2 for subsequent scale-up. This was performed at a scale of 30 mg of trastuzumab which produced a yield very similar to that obtained at the 100 mg scale. Subsequent analysis of the purified material showed it to be pure by analytical size exclusion HPLC with a retention time expected of a protein with a molecular weight of 238 kDa. This is almost the same as the 235 kDa expected of a 1 : 1 conjugate of trastuzumab and streptavidin. Mass spectroscopic analysis of a tryptic digest confirmed that it did indeed have the desired composition and was not simply a combination of reduced antibody chains. The ability of the conjugate to bind to the Her-2 receptor was unchanged to that of the native antibody and its ability to capture biotin was also identical to that of native streptavidin. It can be concluded, therefore that the TSA possessed all the attributes of the ideal antibody–streptavidin conjugate, being both chemically pure and functionally fully active. A preliminary biological evaluation of the TSA was undertaken to assess its likely utility as a pre-targeting reagent. Initially, in-vitro cell binding studies were performed to determine the extent to which the TSA is internalized by or shed from Her-2-expressing tumour cells. These results show that if equilibrium binding conditions are maintained the antigen does not appear to be shed following receptor binding, if the equilibrium is altered by removal of unbound antibody then the antibody itself will, as expected, slowly dissociate from the receptor. Under the conditions employed, 75% of the antibody dissociated over a 2-day period but it is unlikely that this accurately reflects the more complex situation in vivo. The rate of internalization of the conjugate was investigated using the well-established acid-wash approach which strips surface-bound antibody by interfering with the ionic interactions involved in receptor binding. Initial studies using an acid wash at pH 2.8 indicated a relatively high level of antibody internalization (around 50% of that initially bound) but this was at

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470 Nuclear Medicine Communications 2006, Vol 27 No 5

odds with the published data which suggest only a low level of internalization. The effect of decreasing the pH of the acid wash further, initially to pH 2.3 and then to pH 1.8, was therefore studied. Under these conditions the level of apparent internalization was reduced to around 20%. However, microscopic analysis of the cells after acid wash did indicate some loss of integrity, especially at pH 1.8, which might suggest that the lower level of cell retention was in part due to leaching of cell contents and not removal of cell-surface activity. However, two other findings also indicate the extent of internalization if the TSA is relatively low. The first is that, by addition of a large excess of cold antibody, it is possible to displace B90% of the activity associated with the cell. If a significant proportion of TSA were internalized, this would not be the case. The second is that when a TCA precipitation on the cell supernatant following extended incubation with labelled TSA was performed, > 90% of the radioactivity was found to be still protein bound. If a significant level of internalization had occurred, then higher levels of free iodide in the supernatant would be expected following catabolism in the lysosomal compartment and subsequent externalization from the cell [26]. The final characterization step to be performed was an assessment of the ability of the TSA to target Her-2 receptors in vivo. The tumorigenicity of a number of Her2-positive cell lines was studied and the most successful results were obtained with MDA-MB-435S cells. The biodistribution of 125I-TSA was therefore compared with that of 125I-trastuzumab in a small group of nude mice bearing transplanted MDA-MB-435S tumours. The results show that the TSA cleared from the blood significantly faster than the antibody but that otherwise their biodistribution in all other tissues sampled was generally similar although with a trend towards lower retention by the TSA, as would be expected in the face of more rapidly declining blood levels. Tumour uptake of the two tracers was also similar although the level of variation was high perhaps because of variation in size and blood supply.

Conclusion Photoreduction appears to be a very useful route for the preparation of immunoconjugates based on linkage through reduced disulfide bridges. In particular it has been shown to be suitable for the preparation of a trastuzumab–antibody conjugate which possess all the attributes required of a pre-targeting agent for radioimmunotherapy. Preliminary biological studies indicate that the Her-2 receptor is a potential target for such an approach, although, as is the case in all multi-step methodology, optimization of each step including a more extensive in-vivo assessment of Her-2 pre-targeting is required.

Acknowledgements We gratefully acknowledge the use of the facilities of the Barts Foundation for Research and the technical assistance of Maria Stalteri, David Ellison Sandra Peak and the CR-UK mass spectroscopy unit of Dr Nick Totty.

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Goodwin DA, Meares CF, McCall MJ, McTigue M, Chaovapong W. Pretargeted immunoscintigraphy of murine tumors with indium-111-labeled bifunctional haptens. J Nucl Med 1988; 29:226–234. Boerman OC, van Schaijk FG, Oyen WJ, Corstens FH. Pretargeted radioimmunotherapy of cancer: progress step by step. J Nucl Med 2003; 44:400–411. Paganelli G, Malcovati M, Fazio F. Monoclonal antibody pretargetting techniques for tumour localization: the avidin–biotin system. International Workshop on Techniques for Amplification of Tumour Targetting. Nucl Med Commun 1991; 12:211–234. Konecny G, Pauletti G, Pegram M, Untch M, Dandekar S, Aguilar Z, et al. Quantitative association between HER-2/neu and steroid hormone receptors in hormone receptor-positive primary breast cancer. J Natl Cancer Inst 2003; 95:142–153. Lemoine NR, Staddon S, Dickson C, Barnes DM, Gullick WJ. Absence of activating transmembrane mutations in the c-erbB-2 proto-oncogene in human breast cancer. Oncogene 1990; 5:237–239. Natali PG, Nicotra MR, Bigotti A, Venturo I, Slamon DJ, Fendly BM, et al. Expression of the p185 encoded by HER2 oncogene in normal and transformed human tissues. Int J Cancer 1990; 45:457–461. Kraus MH, Popescu NC, Amsbaugh SC, King CR. Overexpression of the EGF receptor-related proto-oncogene erbB-2 in human mammary tumor cell lines by different molecular mechanisms. Embo J 1987; 6:605–610. Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 1987; 235:177–182. Stalteri MA, Mather SJ. Technetium-99m labelling of the anti-tumour antibody PR1A3 by photoactivation. Eur J Nucl Med 1996; 23:178–187. Salacinski PR, McLean C, Sykes JE, Clement-Jones VV, Lowry PJ. Iodination of proteins, glycoproteins, and peptides using a solid-phase oxidizing agent, 1,3,4,6-tetrachloro-3 alpha,6 alpha-diphenyl glycoluril (Iodogen). Anal Biochem 1981; 117:136–146. Ellman GL. Tissue sulfhydryl groups. Arch Biochem Biophys 1959; 82: 70–77. Muddukrishna SN, Chen A, Qi P, Smolenski MA. Quantitation of reduced disulfide groups in monoclonal antibodies using 5iodoacetamidofluorescein: a novel size exclusion-HPLC technique. Appl Radiat Isot 1995; 46:1015–1026. Hainsworth J, Harrison P, Mather SJ. Preparation and characterization of a DOTA–lysine–biotin conjugate as an effector molecule for pretargeted radionuclide therapy. Bioconjug Chem 2005; 16:1468–1474. Lindmo T, Boven E, Cuttitta F, Fedorko J, Bunn Jr PA. Determination of the immunoreactive fraction of radiolabeled monoclonal antibodies by linear extrapolation to binding at infinite antigen excess. J Immunol Methods 1984; 72:77–89. Hylarides MD, Mallett RW, Meyer DL. A robust method for the preparation and purification of antibody/streptavidin conjugates. Bioconjug Chem 2001; 12:421–427. Bienvenut WV, Deon C, Pasquarello C, Campbell JM, Sanchez JC, Vestal ML, et al. Matrix-assisted laser desorption/ionization–tandem mass spectrometry with high resolution and sensitivity for identification and characterization of proteins. Proteomics 2002; 2:868–876. Glennie MJ, Johnson PW. Clinical trials of antibody therapy. Immunol Today 2000; 21:403–410. Witzig TE, Gordon LI, Cabanillas F, Czuczman MS, Emmanouilides C, Joyce R, et al. Randomized controlled trial of yttrium-90-labeled ibritumomab tiuxetan radioimmunotherapy versus rituximab immunotherapy for patients with relapsed or refractory low-grade, follicular, or transformed B-cell non-Hodgkin’s lymphoma. J Clin Oncol 2002; 20:2453–2463. http://www.piercenet.com Stimmel JB, Merrill BM, Kuyper LF, Moxham CP, Hutchins JT, Fling ME, et al. Site-specific conjugation on serine right-arrow cysteine variant monoclonal antibodies. J Biol Chem 2000; 275:30445–30450. Natarajan A, Xiong CY, Albrecht H, DeNardo GL, DeNardo SJ. Characterization of site-specific ScFv PEGylation for tumor-targeting pharmaceuticals. Bioconjug Chem 2005; 16:113–121.

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Karacay H, Sharkey RM, Govindan SV, McBride WJ, Goldenberg DM, Hansen HJ, et al. Development of a streptavidin–anti-carcinoembryonic antigen antibody, radiolabeled biotin pretargeting method for radioimmunotherapy of colorectal cancer. Reagent development. Bioconjug Chem 1997; 8:585–594. Reist CJ, Garg PK, Alston KL, Bigner DD, Zalutsky MR. Radioiodination of internalizing monoclonal antibodies using N-succinimidyl-5-iodo-3pyridinecarboxylate. Cancer Res 1996; 56:4970–4977.

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Correspondence

Correspondence Nuclear Medicine Communications 2006, 27:473

Preparation of 99mTc sestamibi for parathyroid imaging Alistair M. Millara and Tom Murrayb Royal Infirmary of Edinburgh, UK bWestern Infirmary, Glasgow, UK.

a

Correspondence to Dr Alistair M. Millar, Radiopharmacy, Royal Infirmary of Edinburgh, Little France Crescent, Edinburgh EH16 4SA, UK. Tel: þ 44 131 242 2929; fax: þ 44 131 242 2931; e-mail: [email protected]

We have serious concerns about the 99mTc sestamibi preparation described in the paper, ‘Increasing the radiochemical purity of 99mTc sestamibi commercial preparations results in improved sensitivity of dual-phase planar parathyroid scintigraphy’ [1]. (1) The authors wrote, ‘The standard preparation was calculated to have a technetium (99mTc and 99Tc) to MIBI ratio of 1 mg technetium per 1 mg MIBI. In the modified preparation, the technetium (99mTc and 99Tc) to MIBI ratio was 0.34 mg technetium per 1 mg MIBI.’ Our calculations show that 18.5 GBq of 99mTc contains 0.1 mg 99mTc. Assuming that the generator had remained uneluted for 24 h, the 99Tc to 99mTc ratio would be 2.6. There would therefore be 0.26 mg 99Tc, making a total of 0.36 mg technetium. This is in reasonable agreement with the ratio quoted for the modified preparation, but insufficient information is provided on how the two ratios were achieved. (2) The authors wrote, ‘In the standard preparation a sufficient quantity of stannous chloride dihydrate was added to adjust the total to 63 mg.’ This is less than the 75 mg stannous chloride dihydrate that a vial of Cardiolite contains. Also, we consider that adding stannous chloride to a kit makes it non-standard.

Baker-Flex aluminium oxide plates, not instant thin-layer chromatography, which are silica plates. Also, the thinlayer chromatographic method specified by Bristol-Myers Squibb is capable of only limited impurity detection. We consider that the addition of stannous chloride to a kit has great potential to upset the radiochemical purity of the final product. A much more sophisticated analytical technique, such as that specified in the British, European and United States pharmacopoeias, is therefore required to confirm that the radiochemical purity is satisfactory. (4) Our main criticism is that the Cardiolite kits were reconstituted with 18.5 GBq 99mTc. In the Cardiolite instructions, a maximum reconstitution activity of 11.1 GBq is specified in the package leaflet (May 2004) supplied with the product marketed in the UK, and a maximum activity of 5.55 GBq is specified in the package leaflet (May 2003) on the Bristol-Myers Squibb USA website. To exceed the manufacturer’s recommendations and then criticize the quality and performance of the resulting product is unreasonable. The authors offer no explanation of why they chose to exceed the manufacturer’s recommended maximum activity. Accommodating the additional activity by increasing the amount of stannous chloride in the kit seems to be a complicated method of achieving a satisfactory product, when reducing the activity added to the kit would probably have had the same effect. In the introduction to the paper, the authors quote a seminal study in which 99mTc sestamibi with a radiochemical purity greater than 95% was used [2]. The authors of this study did not specify the activity at which they reconstituted the sestamibi kits, but perhaps they achieved high radiochemical purity by complying with the manufacturer’s recommendations.

References 1

(3) The authors wrote, ‘All 99mTc sestamibi preparations were evaluated for radiochemical purity by instant thinlayer chromatography, which was performed in accordance with the procedure recommended in the package labelling.’ The package leaflet specifies the use of

2

Karam M, Dansereau RN, Dolce CJ, Feustel PJ, Robinson LW. Increasing the radiochemical purity of 99mTc sestamibi commercial preparations results in improved sensitivity of dual-phase planar parathyroid scintigraphy. Nucl Med Commun 2005; 26:1093–1098. Taillefer R, Boucher Y, Potvin C, Lambert R. Detection and localization of parathyroid adenomas in patients with hyperparathyroidism using a single radionuclide imaging procedure with technetium-99m-sestamibi (doublephase study). J Nucl Med 1992; 33:1801–1807.

c 2006 Lippincott Williams & Wilkins 0143-3636


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NEWS AND VIEWS May 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up-to-date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 206 6930, E-mail: [email protected]

Nuclear Medicine Communications, 2006, 27:475–476

The Nuclear Medicine Community podcast

Late last year, an experiment in the communication of nuclear medicine ideas – a nuclear medicine podcast service – was launched by Rob Williams. At the time of writing, there have been four podcasts, covering topics from SPECT reconstruction algorithms to DNA therapy, the topics and their treatment designed to appeal to and interest all disciplines. Podcasting is a term coined just a few years ago that refers to an automated form of downloading audio files, generally in mp3 format. The nomenclature is slightly confusing, in that neither podcasting nor listening to podcasts requires an iPod or other portable player, and no over-the-air broadcasting is required. The name association came about simply because Apple Computer’s iPod was the best-selling portable digital audio player when podcasting began and was used by early practitioners. It is perfectly possible to listen to podcasts by using a PC, downloading the files and running software that can read them, such as Windows media player, but doing this would negate the major advantage; playing the files away from the workplace. Another major advantage of podcasting is the ability to subscribe to various services then allow the software to automatically

download the files and alert you, obviating the need to keep checking for updates. You might decide, for example, to subscribe to some of the BBC radio podcasts (these at present are mostly limited to the spoken word because of music copyright issues), the nuclear medicine service and perhaps one relating to your own particular hobby. All these can be automatically delivered to your portable mp3 player for you to listen to at your leisure. Subscription does not necessarily imply a financial cost, most podcasting services are free. So is this just a gimmick reserved for the seriously geeky or could it really become a useful part of the glue that holds together our nuclear medicine community? It certainly has a number of advantages over the more traditional hard copy magazines or journals in that it minimises costs and maximises the speed of delivery. It is quite possible to make a podcast with just a little technical knowledge, coupled with time and energy. Once it has been recorded, it can be made available to subscribers almost instantly, unlike the lead time of several months that is standard with magazines and journals. This makes it an ideal method to distribute news while it still really is news. In his first podcast in November, Rob Williams was able to pay tribute to Hal Anger who died on 31 October. A major

advantage of podcasting is that it enables us to use technology to fit in with lives that are ever more frantic. Podcasts can act as nuclear medicine updates for people ‘on the run’ – you can listen to them while on the train, in the car, or even at the gym. Another major advantage arises from the relaxed ‘magazine style’ of most podcasts, so that interviews with leaders in the field can exist alongside tips and tricks for practical nuclear medicine practitioners. The disadvantages of podcasting are those it shares with the information explosion brought about by the development of the Internet. The fact that anyone can make and distribute a podcast means that there is no regulation over content, no editorial control and it places the responsibility back with the listener in deciding if what they are hearing is factually correct or whether opinions expressed are biased. Ultimately, the future of nuclear medicine podcasting will depend on whether its subscribers perceive it as a useful service, and this in turn will depend on whether the quality and diversity of speakers and topics can be maintained. There have been a few teething problems with the availability of the podcasts but they are certainly worth a listen (you can find them available at: http://feeds. feedburner.com/TheNuclearMedicine AndMolecularMedicinePodcast) and

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476 Nuclear Medicine Communications 2006, Vol 27 No 5

please consider responding to Rob Williams’ plea for contributions. The podcast really only can be as good as its contributors! News of members – Ken Herman

We are very sorry to report the death of Ken Herman on 15 January 2006. Ken was the Chief Technologist in the Nuclear Medicine Department of Manchester Royal Infirmary, where he had worked since 1972. The team of people in medical physics and nuclear medicine owes much of its team spirit to Ken and his leadership. Ken was trained as a chemist and was a valuable source of expertise in the world of physics and pharmacy that he joined. In those early days, his practical approach was invaluable: given a problem, he would quickly reach for the bottles on the shelves and cook up a new recipe. He played a special part in the synthesis of 123IHippuran for imaging the kidneys, and thereby made a significant contribution to the department’s research and development. Ken worked hard for the patients of the north-west region, carrying out diagnostic and therapeutic procedures, teaching others, and helping to improve patient services. During 33 years, his devotion to the work in nuclear medicine was a splendid example to the staff who worked with him. For many years, his leadership was a key factor in the success of the department and its service. He was popular and well respected and his keen sense of humour

was much enjoyed by his colleagues. In recent years, he had not enjoyed the best of health, but he was always cheerful and maintained a positive and encouraging outlook on life. He was well-known in the world of nuclear medicine, not only amongst practising professionals but also amongst members of the pharmaceutical and equipment industry with whom he maintained cordial relations. He will be greatly missed and long remembered by his many friends and colleagues.

Venue: Seoul, South Korea Website: www.wfnmb.org/ congress2006/index02.htm

Meeting Announcements 2nd European IRPA Congress on Radiation Protection

Education and Training

Dates: 15–19 May 2006 Venue: Paris, France Website: www.irpa2006europe.com 5th European Symposium on Paediatric Nuclear Medicine

Dates: 25–28 May 2006 Venue: Girona, Spain Website: http://5esopnm. thesaurus.net/

International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM)

Dates: 13–15 November 2006 Venue: Vienna, Austria Website: www-pub.iaea.org/MTCD/ Meetings/Announcements. asp?ConfID = 146

EANM Learning Courses Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on Tel: + 43 1 2128030, fax: + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected]

BNMS Autumn Meeting

Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk EANM Annual Meeting

Dates: 30 September to 4 October 2006 Venue: Athens, Greece Website: www.eanm.org 9th World Congress of Nuclear Medicine and Biology

Dates: 22–27 October 2006

EANM distance learning in nuclear cardiology

This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. http://www.eanm.org/eduOnline/ edu_start.php?navId = 332

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Editorial

Current practice in pancreatic imaging: The role of nuclear medicine Aviral Singh, Adil Al-Nahhas and Zarni Win Nuclear Medicine Communications 2006, 27:477–480 Keywords: Pancreatic imaging, neuroendocrine tumours, pancreatic carcinoma

Correspondence to Dr Adil Al-Nahhas, Department of Nuclear Medicine, Hammersmith Hospital, Du Cane Road, London W12 0HS, UK. Tel: + 0044 208 383 4923; fax: + 0044 208 383 1700; e-mail: [email protected]

Department of Nuclear Medicine, Hammersmith Hospital, London, UK.

Introduction The pancreas is a dual function organ with both exocrine and endocrine cell types. The vast bulk of the pancreas is composed of exocrine tissue, which secretes digestive enzymes. Embedded within the pancreatic exocrine tissue are islets of Langerhans, the endocrine component of the pancreas, which produce hormones such as insulin, glucagon, somatostatin and pancreatic polypeptides. Diseases of the pancreas include hereditary conditions, inflammation (acute and chronic pancreatitis) and pancreatic masses and tumours. The aetiology of the latter is diverse and includes malignant as well as benign lesions. Common benign lesions are inflammatory masses (chronic pancreatitis) and cystic lesions, while common malignancies include adenocarcinoma of the exocrine cells, neuroendocrine tumours (NETs) of the islet cells and pancreatic metastases. Pancreatic adenocarcinoma is one of the foremost causes of death from all cancers and occurs more commonly than NETs, which are rare and differ in histology, hormone production, and biological and clinical behaviour.

Conventional radiology: Current role and limitations A variety of radiological investigation are available for imaging the pancreas. These include plain abdominal film, ultrasound (US), computed tomography (CT), magnetic resonance imaging (MRI), angiography, endoscopic retrograde cholangio-pancreatography (ERCP), percutaneous pancreatography and barium studies. In practical terms US, CT, MRI and ERCP are the primary diagnostic tools.

detectable, small central solid pancreatic masses, particularly small NETs, may still be difficult to identify. US scanning is useful but is highly user dependent and visualization of the entire pancreatic gland may be limited by overlying bowel gas. Endoscopic US can overcome this problem but is relatively invasive. MRI is useful in the assessment of small islet cell tumours [1] while magnetic resonance cholangio-pancreatography (MRCP) provides an accurate non-invasive method to assess the biliary tree and pancreatic duct. However, adjacent biliary stents and surgical clips may degrade image resolution, and presently the relatively limited resolution does not allow accurate visualization of peripheral biliary or pancreatic ductal branches. Angiography has largely been superseded by helical CT, but remains useful in the detection of small islet cell tumours by selective arterial catheterization and portal venous sampling. It remains a very invasive procedure, however. Other invasive procedures include ERCP, which permits the assessment of the biliary and pancreatic ducts. In summary, the major limitations of conventional radiology are the low sensitivity for preoperative localization of neuroendocrine tumours, the difficulty in assessing response to therapy and the invasive nature of some procedures such as ERCP and angiography. There was therefore a need for the development of non-invasive nuclear medicine procedures using radiopharmaceuticals that can provide alternative ‘physiological’ imaging.

Pancreatic endocrine scintigraphy Multiphase intravenous contrast-enhanced CT is generally considered the most reliable technique for assessing the pancreas and the peripancreatic tissue. Ultra-fast helical CT scanners now provide highly detailed images of the pancreas and major blood vessels in a single breath hold. Although pancreatic adenocarcinoma is readily

Pancreatic scintigraphy dates back to early 1960s, using gamma camera75Seselenomethionine imaging, followed by digital-data-processing techniques using 75Se and 198 Au [2]. These techniques fell out of favour due to the long half-life of 75Se (120 days), conveying a large radiation dose to the patient, and its tendency to

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478 Nuclear Medicine Communications 2006, Vol 27 No 6

accumulate in peripancreatic tissue, resulting in poor image resolution. Interest in radionuclide imaging of the pancreas waned over the following years due to unavailability of suitable radiopharmaceuticals that could target specific tissues or receptors of endocrine origin, or be able to detect metabolic activity in tumours. In addition, planar gamma camera imaging suffered from high background noise resulting in poor resolution with subsequent reduction in the ability to detect smaller lesions. The major breakthrough in imaging neuroendocrine tumours, including those of the pancreas, was the development of somatostatin receptor imaging (SRI). Of the various somatostatin analogues developed so far, octreotide, with affinity to human somatostatin receptor 2 (hSSTR2) and hSSTR5, was the most widely used in clinical endocrinology and later in SRI as 111In-DTPA-DPhe1-octreotide (111In-octreotide). Its success was augmented by developments of gamma camera hardware including the availability of multi-headed cameras and the use of better crystals. The wider use of single photon emission tomography (SPECT) resulted in dramatic improvement in image resolution. Krenning et al. performed SRI in over a thousand patients with NETs, and demonstrated sensitivity of 100% for gastrinomas (12/12) and glucagonomas (3/3), 96% for carcinoids (69/72) and 69% for insulinomas (14/23). The low sensitivity for insulinoma was attributed to the existence of hSSTR subclass [3]. However, Schillaci et al. studied 14 patients with biochemically proven insulinomas and reported a sensitivity of 87.5% using SPECT, which was far superior to that of CT (31.3%) and MRI (43.8%) [4]. Chiti et al. compared SRI to CT and US in 131 patients with NETs of whom 39 had pancreatic lesions including 14 carcinoids, eight gastrinomas, three insulinomas, two glucagonoma and 12 non-secretory tumours. The overall sensitivity for detecting primary NET with SRI, CT and US was 62%, 43% and 36% respectively. For detection of liver metastasis, sensitivity with SRI, CT and US were 90%, 78% and 88% respectively. Interestingly, eight previously unknown malignant pancreatic NETs were also detected with SRI [5]. The MAURITIUS trial involved the use of another SRI radiopharmaceutical, 111In-DOTA-lanreotide (111In-lanreotide), which has a high binding affinity to hSSTR2-5, and weak affinity to hSSTR1. Its value, in a limited number of pancreatic NETs, was compared to that of 111 In-octreotide. Both radiopharmaceuticals had a sensitivity of 100% for insulinomas, but 111In-octreotide had a higher sensitivity of 100% for gastrinomas compared to 75% for 111In-lanreotide [6].

The obvious physical and technical advantages of using a technetium-labelled somatostatin analogue led to the development of 99mTc-depreotide, which has a high receptor-binding affinity to hSSTR2, 3 and 5. In comparative studies, however, it failed to supersede the sensitivity of 111In-octreotide for detecting neuroendocrine tumours [7].

PET imaging of neuroendocrine tumours In comparison to single photon imaging, PET provides better spatial resolution (around 5 mm) and allows quantification of tracer uptake. It can be used to stage disease and monitor response to therapy. Both techniques have the added advantage of detecting unsuspected lesions. PET imaging of somatostatin receptor positive tumours was first described by Hoffman et al. [8] in carcinoid tumours and by Kowalski et al. [9] in neuroendocrine tumours using Gallium-68 labelled with a DOTAconjugated analogue of somatostatin (DOTATOC). The PET tracer 68Ga (with a half-life of 68 min) is available commercially as a 68Ge/68Ga generator, negating the need for a cyclotron, and can be used efficiently to label DOTATOC as well as other DOTA peptides such as DOTATATE and DOTANOC. Higher tumour to nontumour contrast and higher tumour detection rates were achieved with 68Ga-DOTATOC compared with 111 In-octreotide scintigraphy [8,9]. High uptake in pancreatic NETs was clearly demonstrated [10]. In our own clinical experience, 68Ga-DOTATATE PET scans were superior to 111In-octreotide in detecting pancreatic NETs (unpublished data). Neuroendocrine cells characteristically take up decarboxylate amine precursors such as L-DOPA and 5-hydroxytryptophan (5-HTP), and store their biogenic amines (dopamine and serotonin) [11]. L-DOPA and 5-HTP have been labelled with 11C and 18F and the radiolabelled compounds have been used for PET imaging of functional NETs. Most of these studies have reported an advantage over anatomical imaging in detecting primary as well as metastatic lesions [12]. However, difficulty in localizing pancreatic lesions may be encountered, particularly in the head of the pancreas, due to increased adjacent renal activity. Pre-medication with carbidopa was shown to overcome this limitation by significantly reducing background activity and hence improving image quality [13]. Hyperinsulinism is another neuroendocrine disease resulting from a focal adenoma or diffuse abnormality of which the former may benefit from surgery. Based on immunohistochemical analyses, Ribeiro et al. showed that 18 F-DOPA PET was able to distinguish between the two forms [11]. However, due to the high physiological accumulation of tracer in the kidneys, co-registration of

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Editorial Singh et al. 479

PET images with MRI was advised for optimal surgical planning. The localization provided by 18F-DOPA was as precise as that obtained by pancreatic venous sampling. However, the latter is invasive, technically difficult and requires anaesthesia and cessation of all medications for 5 days prior to the procedure. PET radiopharmaceuticals targeting somatostatin receptors continue to develop and include 110mIn-DTPA-DPhe1-octreotide, 86Y-DOTA0-D-Phe1-Tyr3-octreotide and the 18F-labelled carbohydrate analogue of octreotide. The cholinergic PET radioligand 4-[18F]fluorobenzyltrozamicol has been suggested for assessing the functioning betacell mass in the pancreas through visualization of uptake in sympathetic neuroreceptors. Although in its early stages, this approach may provide an in-vivo method of determining the insulin-producing beta-cell mass and may play a role in diabetes research.

Pancreatic exocrine scintigraphy The use of labelled amino acids such as 11C-tryptophan and 11C-methionine in imaging the exocrine pancreas dates back to the late 1970s. The radiopharmaceutical was reported to accumulate in normal pancreatic tissue while the neoplasms were seen as a defect of radiotracer uptake. When visible, the pancreas was easily located and distinguishable from the intestinal image but differentiation between carcinoma and pancreatitis was impossible [14]. Using a gamma camera to image 11C, a positron emitter, was a clear disadvantage resulting in lower resolution compared to modern day PET scanners. Receptor imaging of vasoactive intestinal peptide (VIP) receptors and vascular endothelial growth factor (VEGF) receptors were also attempted. Receptor expression is noted in adenocarcinomas as well as in NETs, but its use in the latter has been largely superseded by SRI. The diagnostic value of 123I-VIP was investigated by Raederer et al. in 60 patients suffering from primary pancreatic adenocarcinoma. The sensitivity for primary tumours confined to the pancreas was 90%, but that for locoregional and liver metastasis was 32%, reducing the overall sensitivity to 58% [15]. However, these results were challenged by other studies showing extremely low sensitivity in patients with adenocarcinoma of the colon and pancreas [16]. This was attributed to a higher number of VIP receptors in normal tissues compared to neoplastic tissues. Imaging with 123I-vascular endothelial growth factor (123IVEGF) has been developed for the localization and diagnosis of a variety of human solid tumours. In-vitro specific binding to pancreatic tumour cells has been demonstrated [17], but this still remains a research tool.

Functional imaging of exocrine pancreatic tumours was revolutionized with the development of PET scanners. Several radiopharmaceuticals have been used for this purpose such as 18F-FDG, 11C-methionine, 11C-acetate, 11 C-hydroxytryptophan and 18F-DOPA. However, the most practical and widely used radiopharmaceutical is 18 F-FDG. The use of 18F-FDG to evaluate pancreatic malignancies is based on the tendency of malignant cells to overexpress glucose transporters and, thus, to have a high uptake of the glucose analogue FDG. Sensitivity figures ranging between 85 and 100% and specificity between 67 and 99% have been reported and 18F-FDG PET has been shown to significantly add to the diagnostic accuracy of ERCP and CT in the assessment of pancreatic malignancies [18–20]. The main drawback affecting specificity of 18F-FDG PET is the difficulty in differentiating malignant from benign lesions. Delbeke et al. studied 65 patients with pancreatic tumours and found that quantitative analysis of 18F-FDG PET using a cut-off SUV of 3.0 to be helpful in differentiating malignant from benign pancreatic lesions. 18 F-FDG PET had a sensitivity and specificity of 92% and 85% compared to 65% and 61% for CT [18]. Despite its reduced specificity, 18F-FDG PET performs better than CT in this respect. Higashi et al. retrospectively investigated 231 pancreatic cancer patients of whom 53 (23%) had equivocal results on conventional imaging with its inability to differentiate malignant from benign lesions. They were referred for 18F-FDG PET, which showed sensitivity, specificity and accuracy of 65%, 93% and 81%, respectively [19]. Other methods of differentiating malignant from benign pancreatic lesions include scanning at 2 h and observing the pattern of uptake. Pre-operative staging of pancreatic cancer with 18F-FDG PET is recommended. Nishyama et al. investigated the value of 18F-FDG PET in detecting distant metastases in 42 patients with metastatic pancreatic cancer to liver, lung, bone, peritoneum and lymph nodes [20]. 18F-FDG PET had sensitivity, specificity and accuracy of 82%, 98% and 97% as compared to 64%, 98% and 94% for CT. Others found 18F-FDG PET helpful for staging in 40% of cases, with additional metastases or unsuspected lesions detected in 38% cases [19]. Studies concerning follow-up of post-treatment patients are not extensive, due to the poor prognosis of pancreatic cancer. Maisey et al. have suggested that the absence of 18 F-FDG uptake at 1 month following chemotherapy in patients with pancreatic carcinoma is an indicator of improved overall survival [21].

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Interest in PET radiopharmaceuticals that are used to delineate the normal pancreas was resumed with the use of 11C-methionine to detect impaired exocrine function. Likewise, 11C-acetate has been shown to provide good anatomical delineation of the pancreas. Loss of functional pancreatic tissue such as in chronic pancreatic masses and adenocarcinoma showed markedly reduced avidity for 11 C-acetate. However, only a moderate reduction in 11Cacetate avidity was observed in cases of uncomplicated acute pancreatitis. Hence, 11C-acetate has been suggested to complement the metabolic activities revealed by 18F-FDG in distinguishing between acute inflammation and neoplasms of the pancreas [22]. Various novel pharmaceuticals are being investigated for pancreatic imaging. A laboratory study compared phenylalanine derivatives 123I-AIPA and 123I-IPA with 18F-FDG PET, in pancreatic adenocarcinoma, and proposed them as potential pancreatic cancer imaging agents with comparatively lower uptake in pancreatic inflammation [23]. The recent availability of PET–CT scanners has the combined value of imaging anatomy and function and recent data suggest clinical and economical advantage that will enhance future management of pancreatic cancer. In summary, the last two decades have witnessed a significant development in the use of nuclear medicine techniques to assess pancreatic tumours. The introduction of receptor and metabolic imaging has added function to the anatomical information perceived by cross-sectional and interventional radiology, resulting in better patient management. The success of SRI in the assessment of NETs is evident by its incorporation into clinical practice and the use of PET with 68Ga-DOTA radiopharmaceuticals is bound to expand in the foreseeable future.

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The use of F-FDG PET in adenocarcinoma of the pancreas continues to flourish and its value in detecting unsuspected lesions makes it useful in pre-surgical staging. Its specificity continues to improve with the development of a variety of techniques to distinguish adenocarcinoma from pancreatitis. Further progress is expected with the wider use of hybrid PET–CT scanners and further development of optimal PET and receptor radiopharmaceuticals.

19

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References 1 2

Outwater EK, Gordon SJ. Imaging of the pancreas and biliary ducts with MR. Radiology 1994; 192:19–21. Blanquet PC, Beck CR, Fleury J, Palais CJ. Pancreas scanning with 75-selenomethionne and 198Au using digital-data-processing techniques. J Nucl Med 1968; 9:486–488.

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Krenning EP, Kwekkeboom DJ, Bakker WH, Breeman WA, Kooij PP, Oei HY, et al. Somatostatin receptor scintigraphy with [111In-DTPA-D-Phe1]- and [123I-Tyr3]-octreotide: the Rotterdam experience with more than 1000 patients. Eur J Nucl Med 1993; 20:716–731. Schillaci O, Rita M, Scopinaro F. 111In-pentetreotide scintigraphy in the detection of insulinomas: Importance of SPECT imaging. J Nucl Med 2000; 41:459–462. Chiti A, Fanti S, Savelli G, Romeo A, Bellanora B, Rodari M, et al. Comparison of somatostatin receptor imaging, computed tomography and ultrasound in the clinical management of neuroendocrine gastro-enteropancreatic tumours. Eur J Nucl Med 1998; 25:1396–1403. Virgolini I, Traub T, Leimer M, et al. New trends in peptide receptor radioligands. Q J Nucl Med 2001; 45:153–159. Lebtahi R, Cloirec JL, Houzard C, et al. Detection of neuroendocrine tumours: 99mTc-P829 scintigraphy compared with 111In-pentetreotide scintigraphy. J Nucl Med 2002; 43:889–895. Hofmann M, Maecke H, Borner R, et al. Biokinetics and imaging with the somatostatin receptor PET radioligand (68)Ga-DOTATOC: preliminary data. Eur J Nucl Med 2001; 28:1751–1757. Kowalski J, Henze M, Schuhmacher J, et al. Evaluation of positron emission tomography imaging using [68Ga]-DOTA-D Phe(1)-Tyr(3)-Octreotide in comparison to [111In]-DTPAOC SPECT. First results in patients with neuroendocrine tumors. Mol Imaging Biol 2003; 5:42–48. Henze M, Schuhmacher J, Dimitrakopoulou-Strauss A, et al. Exceptional increase in somatostatin receptor expression in pancreatic neuroendocrine tumour, visualized with 86Ga-Dotatoc PET. Eur J Nucl Med Mol Imaging 2004; 31:466. Ribeiro MJ, Lonlay PD, Delzescaux T, et al. Characterisation of hyperinsulinism in infancy assessed with PET and 18F-fluoro-L-DOPA. J Nucl Med 2005; 46:560–566. Orlefors H, Sundin A, Garske U, et al. Whole-body 11C-5hydroxytryptophan positron emission tomography as a universal imaging technique for neuroendocrine tumours: comparison with somatostatin receptor scintigraphy and computed tomography. J Clin Endocrinol Metab 2005; 90:3392–3400. Orlefors H, Sundin A, Lu L, et al. Carbidopa pretreatment improves image interpretation and visualisation of carcinoid tumours with 11C-5hydroxytryptophan positron emission tomography. Eur J Nucl Med Mol Imaging 2006; 33:60–65. Syrota A, Comar D, Cerf M, et al. 11C-methionine pancreatic scanning with positron emission computed tomography. J Nucl Med 1979; 20: 778–781. Raederer M, Kurtaran A, Qiong Y, et al. Iodine-123-vasoactive intestinal peptide receptor scanning in patients with pancreatic cancer. J Nucl Med 1998; 39:1570–1575. Hessenius C, Bader M, Meinhold H, et al. Vasoactive intestinal peptide receptor scintigraphy in patients with pancreatic adenocarcinomas or neuroendocrine tumours. Eur J Nucl Med 2000; 27:1684–1693. Li S, Peck-Radosavljevic M, Kienast O, et al. Iodine-123-vascular endothelial growth factor-165 (123I-VEGF165) Biodistribution, safety and radiation dosimetry in patients with pancreatic carcinoma. Q J Nucl Med Mol Imaging 2004; 48:198–206. Delbeke D, Rose DM, Chapman WC, et al. Optimal interpretation of FDG PET in the diagnosis, staging and management of pancreatic carcinoma. J Nucl Med 1999; 40:1784–1791. Higashi T, Saga T, Nakamoto Y, et al. Diagnosis of pancreatic cancer using fluorine-18 deoxyglucose positron emission tomography (FDG PET) – Usefulness and limitations in ‘clinical reality’. Ann Nucl Med 2003; 17:261–279. Nishyama Y, Yamamoto Y, Yokoe K, et al. Contribution of whole body FDG-PET to the detection of distant metastasis in pancreatic cancer. Ann Nucl Med 2005; 19:491–497. Maisey NR, Webb A, Flux GD, Padhani A, Cunningham DC, Ott RJ, Norman A, et al. FDG-PET in the prediction of survival of patients with cancer of the pancreas: a pilot study. Br J Cancer 2000; 83: 281–283. Shreve PD, Gross MD. Imaging of the pancreas and related diseases with PET carbon-11-acetate. J Nucl Med 1997; 38:1305–1310. Hellwig D, Menges M, Schneider G, Moellers MO, Romeike BF, Menger MD, et al. Radioiodinated phenylalanine derivatives to image pancreatic cancer: a comparative study with [18F]fluoro-2-deoxy-D-glucose in human pancreatic carcinoma xenografts and in inflammation models. Nucl Med Biol 2005; 32:137–145.

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Original article

FDG PET for the assessment of myometrial infiltration in clinical stage I uterine corpus cancer Tatsuo Torizukaa, Fumitoshi Nakamuraa, Munetaka Takekumab, Toshihiko Kannoa, Tomomi Ogusuc, Etsuji Yoshikawac, Hiroyuki Okadac, Makoto Maedab and Yasuomi Ouchia Objective For surgical planning of uterine corpus cancer, prior knowledge of the depth of myometrial invasion is important. Curative tumour resection is possible in superficial invasion (stages IA and IB), while post-surgical chemotherapy or radiation therapy is required in deep invasion (stage IC). We evaluated the value of positron emission tomography with 2-[18F]fluoro-2-deoxy-D-glucose (FDG PET) for estimating the myometrial invasion in uterine corpus cancer. Methods We studied 22 patients with clinical stage I uterine corpus cancer, who underwent FDG PET prior to surgery. Standardized uptake value (SUV; tracer activity per injected dose normalized to body weight) was calculated on the PET image. PET findings were compared with magnetic resonance imaging (MRI) and the surgical staging. Results The surgical stage was IA in five, IB in 11 and IC in six patients. SUVs in deep invasion (15.69 ± 4.73, 8.83–21.84) were significantly higher than those in superficial invasion (9.09 ± 3.29, 2.68–15.41) (P < 0.005). Using 12.0 as a cut-off value of SUV for the differentiation of these two groups, PET results were correct in 19 patients but were incorrect in three patients. Although both PET and MRI provided correct staging in 14 patients, only MRI

Introduction Uterine corpus cancer is the common gynaecological malignancy and the fourth most common cancer in women. In Europe more than one in 20 female cancers are of the endometrium [1] and over 40 000 new cases are reported annually in the United States [2]. Surgery is the treatment of choice in patients with non-invasive or locally advanced corpus cancer [3]. In 1988, the International Federation of Gynecology and Obstetrics (FIGO) adopted a surgico-pathological system for the staging of corpus cancer [4]. According to data from the FIGO, approximately 75% of patients are surgical stage I at presentation and only 3% are surgical stage IV [5]. Curative tumour resection is possible in stage IA (tumour limited to endometrium) and stage IB (invasion of less than 50% of the myometrial thickness), while dissection of para-aortic lymph node and post-surgical chemotherapy or radiation therapy are required in stage IC (invasion of

overestimated the myometrial invasion in four patients with stage IB and showed inconclusive findings in one patient with stage IC. Four of these five patients were post-menopausal. Conclusions The cut-off value of SUV ( = 12.0) may be a useful index for the differentiation of superficial invasion and deep invasion. FDG PET may be feasible for predicting the myometrial infiltration of uterine corpus cancer, especially when uterine atrophy makes it difficult at MRI in post-menopausal patients. Nucl Med Commun 27:481–487

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:481–487 Keywords: FDG PET, uterine corpus cancer, diagnosis, myometrial infiltration a Positron Medical Center, bDepartment of Gynecology and Obstetrics, Hamamatsu Medical Center, Japan and cCentral Research Laboratory, Hamamatsu Photonics K.K., Japan.

Correspondence to Dr Tatsuo Torizuka, Positron Medical Center, Hamamatsu Medical Center, 5000, Hirakuchi, Hamamatsu, 434-0041, Japan. Tel: + 0081 53 585 0366; fax: + 0081 53 585 0367; e-mail: [email protected] Received 19 November 2005 Accepted 13 February 2006

greater than 50% of the myometrial thickness). Lymph node metastases are unlikely unless deep myometrial invasion is present [6–8]. Thus, surgical planning, which may include the dissection of para-aortic lymph nodes, may be altered by a prior knowledge of the depth of myometrial invasion. The diagnostic performance of magnetic resonance imaging (MRI) and X-ray computed tomography (CT) in assessing myometrial invasion of corpus cancer has been extensively evaluated. Results from studies [9,10] in which MRI was compared with CT, suggest MRI is more accurate than CT. On MRI, the degree of myometrial invasion has been evaluated on the basis of the appearance of the junction zone and myometrial thickness [11,12]. Preservation of the junction zone usually indicates superficial invasion whereas focal disruption or absence of the zone is associated with deep

c 2006 Lippincott Williams & Wilkins 0143-3636

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invasion. However, the junction zone is rarely observed in post-menopausal women, in whom the majority of uterine corpus cancer is most frequently observed. Positron emission tomography (PET) with 2-[18F]fluoro2-deoxy-D-glucose (FDG) has emerged as an extremely useful technique in clinical oncology [13–15]. However, there have been only a few studies for estimating the diagnostic value of PET for patients with uterine corpus cancer. In preoperative studies, Horowitz et al. [16] showed that FDG PET was highly specific in the prediction of pelvic and para-aortic lymph node metastasis but the use of PET had a limitation for detecting microscopic metastases. In post-operative studies [17,18], FDG PET appears useful for the accurate localization of suspected recurrences and for the detection of asymptomatic recurrent disease and may afford important information in the management of patients. To our knowledge, there is no study that describes the clinical application of FDG PET in preoperative assessment of myometrial infiltration of uterine corpus cancer. This retrospective study was designed to evaluate the value of FDG PET in predicting the degree of myometrial invasion in patients with clinical stage I uterine corpus cancer.

starting 60 min after intravenous administration of FDG (400–500 MBq) [20]. The acquisition time was 6 min per bed position. A transmission scan for segmented attenuation correction was then acquired with a 68Ge external ring source for 3 min per bed position. To minimize the accumulation of FDG activity in the urinary bladder, the patients were asked to void just before the emission scan. Transaxial, coronal and sagittal images were reconstructed by means of the ordered subsets expectation maximization method (OSEM) with eight subsets and three iterations. The average reconstructed x–y spatial resolution was about 4.0 mm full width half maximum (FWHM) in-plane. Magnetic resonance imaging

MRI was acquired on a 1.5-T magnetic resonance system (Magnetom Symphony, Siemens, Erlangen, Germany) using a pelvic phased-array coil. T1 weighted spin-echo images (TR/TE = 550/15) were obtained in the transaxial plane. T2 weighted fast spin-echo images (TR/effective TE = 4000/80–100) were obtained in the transaxial and sagittal planes. Using 0.1 mmol
kg – 1 of gadopentetate dimeglumine (Magnevist, Schering, Osaka, Japan), transaxial, coronal and sagittal T1 weighted spin-echo images (TR/TE = 550/15) were acquired. Image analysis

Methods Patient population

Whole-body FDG PET scans were performed on 22 patients with clinical stage I uterine corpus cancer for preoperative assessment. Lesions in all patients were diagnosed on the basis of findings at dilatation and curettage and physical examination. These 22 patients underwent magnetic resonance examination of the pelvis 12–20 days after the dilatation and curettage and received PET examination 1–13 days after MRI. The diagnosis was confirmed at surgery 4–15 days after PET examination. The surgical procedures performed included a total abdominal hysterectomy, bilateral salpingo-oophorectomy and bilateral pelvic lymphadenectomy. When indicated, omentectomy was performed. The size of each primary lesion was recorded as the largest and smallest diameters at surgical fixed specimens. All patients provided written informed consent for participation in the study, which was approved by our institutional review board. PET imaging

All patients fasted for at least 5 h before PET studies. Serum glucose levels measured at the time of FDG injections were < 150 mg
dl – 1 in all patients. A wholebody PET scanner was used (model SHR22000, Hamamatsu Photonics, K.K., Hamamatsu, Japan) [19]. The SHR22000 scanner permits simultaneous acquisition of 63 transverse planes each of thickness 3.6 mm and encompassing an axial field of view of 23.0 cm. Wholebody emission scan was obtained with five bed positions,

PET images were interpreted visually and analysed with knowledge of clinical information. Increased FDG uptake relative to the background was considered to be positive for primary tumour. For semiquantitative analysis, the standardized uptake value (SUV = tracer activity/injected dose normalized to body weight) was calculated in each patient. A computerized semi-automated algorithm was employed to eliminate interobserver discrepancy [21]. This method helps to define the maximal uptake in a small square region of interest (ROI) (1.2  1.2 cm; 3  3 pixels) placed within the whole tumour. The highest SUV in one pixel was used to minimize the partial volume effect. The findings of PET imaging were compared with those of MRIs, and related to histopathological findings at surgery. Assessment with MRI for the depth of myometrial invasion was based on the established criteria [12]. Stage IA was diagnosed if the junction zone or endometrial– myometrial interface was intact, and there was no area of abnormal signal intensity in the myometrium. Stage IB was diagnosed if the magnetic resonance image showed a mass that invaded, at most, 50% of the myometrial wall thickness and disrupted the junction zone or the endometrial–myometrial interface. Stage IC was diagnosed if the magnetic resonance image showed a mass that invaded more than 50% of the myometrial wall thickness and disrupted the junction zone or the endometrial–myometrial interface. Magnetic resonance images were interpreted by experienced radiologists.

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FDG PET for uterine corpus cancer Torizuka et al. 483

Statistical analysis

Student’s unpaired t-test was used to compare SUV with the FIGO surgical staging and histological grade. Pearson’s correlation coefficient was used to correlate between SUV and tumour size. A two-sided P-value of < 0.05 was considered significant.

Results The findings of PET and magnetic resonance images and surgical histopathology in our series of 22 women with clinical stage I uterine corpus cancer are presented in Table 1. Based on the surgical FIGO staging system [4], the depth of myometrial invasion of primary tumour was evaluated. Five patients had stage IA (tumour limited to endometrium), 11 patients had stage IB (invasion of less than 50% of the thickness of the endometrium) and six patients had stage IC (invasion of greater than 50% of the thickness of the endometrium). The histology of primary tumour was endometrial adenocarcinoma in 17 patients, carcinosarcoma (malignant mixed epithelial and mesenchymal tumour) in four patients and clear-cell carcinoma in one patient. Of the 17 patients with endometrial adenocarcinoma, tumour histologic grade included highly differentiated adenocarcinoma (G1) in three patients, moderately differentiated adenocarcinoma (G2) in 11 patients and poorly differentiated adenocarcinoma (G3) in three patients. The mean age of the patients was 62 years (range, 40–76 years). Seven patients were premenopausal and 15 patients were post-menopausal. Primary tumour was visualized on PET images in all patients. Tumour SUVs in stage IA, stage IB and stage IC

Table 1

According to the depth of myometrial invasion, our 22 patients were divided into two groups: superficial invasion (stages IA and IB, n = 16) and deep invasion (stage IC, n = 6). Using 12.0 as a cut-off value for the differentiation of these two groups, PET imaging was correct in 19 of the 22 patients but was incorrect in two patients with stage IB (numbers 6 and 7) and one patient with stage IC (number 22) (Fig. 1). Although both PET and magnetic resonance images were correct in 14 patients, only MRI overestimated the depth of myometrial invasion in four patients with stage IB (Fig. 3) and showed inconclusive finding in one patient with stage IC (Fig. 4). Four of these five patients were post-menopausal. There was no patient in whom both PET and magnetic resonance findings were incorrect for the estimation of myometrial invasion. Overall, for the diagnosis of deep invasion, the sensitivity, specificity, accuracy, positive predictive value and negative predictive value were 83.3%, 87.5%, 86.4%, 71.4% and 93.3%, respectively, for PET imaging while those values were 83.3%, 75.0%, 77.3%, 55.6% and 92.3%, respectively, for MRI.

Patients and tumour characteristics and imaging findings

Patient number

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

were 7.62 ± 2.99 (2.68–10.32), 9.75 ± 3.33 (2.83–15.41) and 15.69 ± 4.73 (8.83–21.84), respectively. SUVs in stage IC were significantly higher than those in stage IA and stage IB (P < 0.01), while no significant difference of SUVs was observed between stage IA and stage IB (Fig. 1). The relation between tumour SUV and the largest diameter in the surgical specimen is shown in Fig. 2. There was a moderate but significant correlation between these two parameters (P < 0.05).

Age (years)

49 40 63 69 51 72 57 73 42 53 51 62 67 62 66 74 72 50 69 76 65 72

FIGO stage

IA IA IA IA IA IB IB IB IB IB IB IB IB IB IB IB IC IC** IC IC IC IC

PET SUV

10.32 9.14 8.90 7.04 2.68 15.41 13.54 11.30 10.99 10.13 9.82 9.82 8.10 8.39 6.97 2.83 21.84 19.88 16.37 13.33 13.86 8.83

Magnetic resonance stage IA IA IA IA IA IB IB IB IB IC IC IB IC IB IB IIA IC IC IB/IC IC IC IC

Tumour characteristics Histology

Size (cm)

Grade*

AdenoCA AdenoCA AdenoCA AdenoCA Carcinosarcoma AdenoCA AdenoCA Carcinosarcoma AdenoCA AdenoCA AdenoCA AdenoCA AdenoCA AdenoCA AdenoCA Clear-cell CA AdenoCA AdenoCA AdenoCA Carcinosarcoma AdenoCA Carcinosarcoma

2.0  1.0 2.0  1.0 2.5  1.2 2.0  1.5 2.0  1.0 3.2  1.8 2.5  2.5 5.0  2.0 3.0  1.3 4.5  2.0 6.0  4.0 3.3  2.0 3.0  3.0 2.0  1.5 0.8  0.4 2.5  1.9 5.5  3.0 6.6  5.2 3.0  3.0 13.0  5.0 10.0  8.0 5.0  4.0

1 1 3 2 n/a 2 2 n/a 2 1 2 3 2 2 2 n/a 3 2 2 n/a 2 n/a

FIGO, International Federation of Gynecology and Obstetrics. AdenoCA = endometrial adenocarcinoma; carcinosarcoma = malignant mixed epithelial and mesenchymal tumour; clear-cell CA = clear-cell carcinoma. * 1 = highly differentiated; 2 = moderately differentiate; 3 = poorly differentiated; n/a = not available. **Final diagnosis was stage IIIC with para-iliac nodal metastasis.

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Fig. 1

Fig. 2

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15 r = 0.453 P < 0.05

10 Size (cm)

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SUV Correlation between tumour SUV and the largest diameter in the surgical specimen. There is a moderate but significant correlation between these two parameters (P < 0.05).

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Fig. 3

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IB Stage

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Comparison of tumour standardized uptake value (SUV) between stage IA, stage IB and stage IC in 22 patients with uterine corpus cancer. SUVs in stage IC are significantly higher than those in stage IA and stage IB (P < 0.01).

SUV = 9.82

BL

Tumour SUVs and histological grade were compared in the 17 patients with endometrial adenocarcinoma (Fig. 5). Mean SUV was 9.86 ± 0.63 (9.14–10.32) in three patients with highly differentiated adenocarcinoma (G1), 11.85 ± 4.27 (6.94–19.88) in 11 patients with moderately differentiated adenocarcinoma (G2) and 13.52 ± 7.22 (8.90–21.84) in three patients with poorly differentiated adenocarcinoma (G3). There was no significant difference of SUVs between these three groups, although the number of patients was small in each group. In pathological examination of resected lymph nodes, only one patient with deep myometrial invasion (number 18) had one positive node in left para-iliac region. Metastatic involvement was only 2.5 mm microscopically. Before surgery, MRI showed an enlarged lymph node (2 cm) while PET finding was negative, probably because of the microscopic metastatic disease. The final diagnosis

BL

A 51-year-old woman with endometrial carcinoma (patient number 11). (a) T2 weighted sagittal magnetic resonance image shows endometrial tumour (arrow) of the uterine body. A junction zone, a hypo-intense rim between endometrium and myometrium, is present along the posterior wall of the uterine body and interrupted along the anterior wall (short arrows). Endometrial tumour appears to infiltrate the myometrium to greater than 50% of its thickness. (b) Sagittal PET image shows increased FDG uptake (SUV = 9.82), corresponding to the endometrial tumour. At surgery, histology was endometrial adenocarcinoma, grade 2, infiltrating less than 50% of the myometrial wall thickness (stage IB). BL = urinary bladder.

was FIGO stage IIIC endometrial cancer in this patient. In the other 21 patients, there was no nodal and distant metastasis in surgical histology and clinical findings. Thus, post-surgical chemotherapy or radiation therapy was performed for the six patients with deep myometrial

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FDG PET for uterine corpus cancer Torizuka et al. 485

Fig. 4

Fig. 5

(a)

25

(b)

SUV = 16.37 20

BL BL

SUV

A 69-year-old woman with endometrial carcinoma (patient number 19). (a) T2 weighted sagittal magnetic resonance image shows ill-defined iso-intense endometrial tumour (arrow). A junction zone is intact along the anterior wall of the uterine body and absent along the posterior wall (short arrows). The depth of myometrial invasion is inconclusive in the posterior wall because of the atrophic myometrium. (b) Sagittal PET image shows intense FDG uptake (SUV = 16.37), corresponding to the endometrial tumour. At surgery, histology was endometrial adenocarcinoma, grade 2, extending across more than 50% of the myometrial wall thickness (stage IC). BL = urinary bladder.

15

10

5

invasion (numbers 17–22), including one patient with stage IIIC.

Discussion Uterine corpus cancer is the most common gynaecological malignancy in the female pelvis. It is known that patients with greater than 50% myometrial invasion will have a sixfold to seven-fold higher prevalence of pelvic or paraaortic lymph node metastases and of advanced surgical stage when compared with patients with less than 50% myometrial invasion [22–24]. The reported 5-year survival rate in patients with deep myometrial invasion or with poorly differentiated endometrial cancer is 58%, compared with 89% for patients with highly differentiated endometrial cancer [25]. Therefore, more aggressive treatment, including post-surgical chemotherapy or radiation therapy should be considered for patients with deep myometrial invasion. This study demonstrates the feasibility of using preoperative FDG PET for estimating the depth of myometrial invasion in patients with uterine corpus cancer. Tumour SUVs in deep invasion were significantly higher as compared to those in superficial invasion (P < 0.005) (Fig. 1). Using an SUV of 12.0 as a cut-off value for the differentiation of these two groups, our PET results were correct in 19 patients and incorrect in only three patients. Although magnetic resonance results were correct in 17 of the 22 patients, only magnetic resonance images revealed false or inconclusive findings in five patients, including four post-menopausal patients. Therefore, preoperative assessment of FDG PET may

0 G1

G2 Grade

G3

Comparison of tumour SUV with histological grade in 17 patients with endometrial adenocarcinoma; G1: highly differentiated (n = 3), G2: moderately differentiated (n = 11), G3: poorly differentiated (n = 3). There is no significant difference of SUV between these three groups.

be effective especially in such post-menopausal patients. In most institutions, patients with greater than 50% myometrial invasion are considered for further surgical staging, which includes pelvic and para-aortic lymphadenectomy, and the use of adjuvant chemotherapy or radiation therapy [26]. When tumour SUV is higher than 12.0 and magnetic resonance findings suggest deep myometrial invasion, patients need referral to a gynaecological oncologist. Figure 2 shows a moderate but significant correlation between SUV and tumour size in uterine corpus cancer (P < 0.05). This finding is consistent with the previous reports of recurrent rectal cancer and oesophageal cancer [27,28]. The study of oesophageal cancer [28] demonstrated that FDG uptake was significantly correlated with the depth of tumour invasion as well as the size of tumour and that tumour with high FDG uptake carried a poorer prognosis as compared to low FDG uptake. Higashi et al. [29] showed that FDG uptake is significantly related to

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486 Nuclear Medicine Communications 2006, Vol 27 No 6

cell proliferation of non-small cell lung cancer. They recently demonstrated that FDG uptake can predict intratumoural lymphatic vessel invasion and lymph node metastasis in patients with lung cancer and is an important factor in the planning of appropriate surgical treatment [30]. In our data, FDG uptake by corpus cancer was helpful for predicting the depth of myometrial invasion. One might assume that our patients with superficial invasion showed lower FDG uptake probably because partial volume effects affected smaller tumours in this group. It is known that recovery of radioactivity concentration in structures less than twice the spatial resolution will be decreased. For the PET scanner we used, the spatial resolution is 4.0 mm. Only one lesion (number 15, SUV = 6.97) displayed a diameter less than 8 mm, measured at surgical fixed specimen in this study. Therefore, it appears unlikely that partial volume effects are responsible for the observed difference of SUV between superficial invasion and deep invasion. In this study, we found that tumour SUV did not have a significant correlation with the histological grade of uterine corpus cancer, although the number of patients was small. This result differs from that of previously published data for other tumours, such as lung cancer, breast cancer, malignant lymphoma and brain tumour, where FDG uptake had a positive correlation with the histological grade [31–34]. The possible associations of FDG uptake and Glut-1 expression have been investigated in patients with non-small cell lung cancer and breast cancer [31,32]. In the immunohistochemical study of uterine endometrial cancer [35], overexpression of GLUT-1 was consistently observed in tumour tissue but there was no positive correlation between the extent of GLUT-1 expression and tumour differentiation. Further studies may be necessary to clarify what factor may play an important role in the determination of tumour FDG uptake in uterine corpus cancer. Although FDG PET is sensitive for detecting corpus cancer and useful for estimating the myometrial infiltration, MRI is needed to visualize the normal structures of the pelvis and to ensure accurate localization of the hot spots found in PET images. In addition, our results showed that FDG PET could not detect microscopic nodal metastasis in one patient, which was truly positive on the magnetic resonance image. This finding is consistent with the study by Horowitz et al. [16], who demonstrated that FDG PET has a high specificity but is insensitive in predicting microscopic nodal metastases. Thus, lymphadenectomy is still needed when a negative PET finding is encountered. There were limitations in this study. First, the number of patients studied was relatively small. Second, tumour characteristics were various in histological types and

grade. Thus, prospective studies with additional patients in homogenous groups will be necessary to verify our results in more detail. In conclusion, our preliminary study demonstrates that FDG PET may be feasible for predicting the myometrial infiltration of uterine corpus cancer. The cut-off value of SUV ( = 12.0) appears to be a useful index for the differentiation of superficial invasion and deep invasion, especially when indistinct anatomy of the junction zone makes it difficult to assess myometrial invasion at MRI in post-menopausal patients.

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Original article 99m

Tc-labelled red blood cell single-photon emission computed tomography for the diagnosis and follow-up of juvenile nasopharyngeal angiofibroma* Celil Uslua, Mustafa Yildirimb, Hatice Usluc, Yavuz Sutbeyazd, Erhan Varoglub, Bedri Sevenb, Umran Yildirime and Mecit Kantarcif Aim To confirm the usefulness of blood pool scintigraphy with 99mTc-labelled red blood cells (99mTc-RBCs) in the diagnosis and follow-up of juvenile nasopharyngeal angiofibroma. Methods A prospective study design was used. 99m Tc-RBCs were prepared by a modified in-vivo method. After the rapid intravenous injection of 370–740 MBq of 99m Tc-RBCs, dynamic imaging of 1-min duration was performed. After dynamic imaging, static acquisitions at 5 min (second phase: blood pool phase) and 2 h (third phase: static image) were obtained. In addition, single-photon emission computed tomography (SPECT) imaging was performed at 2 h. SPECT images were obtained using a rotating gamma camera (GE-Starcam 4000 XR/T). Results All patients showed no activity in the first phase and mild activity in the second phase (blood pool phase). All patients with juvenile nasopharyngeal angiofibroma showed a prominent increased activity in the third phase and in SPECT images.

Introduction

Conclusions This study shows that blood pool scintigraphy with 99mTc-RBC SPECT is very accurate, easy to perform and a suitable alternative to pre-operative and post-operative imaging techniques, including computed tomography scan, magnetic resonance imaging (MRI) and MRI angiography, for the detection of juvenile nasopharyngeal angiofibroma. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:489–494 Nuclear Medicine Communications 2006, 27:489–494 Keywords: juvenile nasopharyngeal angiofibroma, scintigraphy, 99m Tc-labelled red blood cell a Department of Otorhinolaryngology, Haydarpasa Numune Hospital for Research and Education, Istanbul, bDepartment of Nuclear Medicine, Medical Faculty, Ataturk University, Erzurum, cDepartment of Nuclear Medicine, Dr. Siyami Ersek Hospital for Research and Education, Istanbul, dDepartments of Otorhinolaryngology, ePathology and fRadiology, Medical Faculty, Ataturk University, Erzurum, Turkey.

Correspondence to Celil Uslu MD, Tosunpasa sok, No: 13/19, 34672/Uskudar, Istanbul, Turkey. Tel: + 90 532 4363976; fax: + 90 216 3460582; e-mail: [email protected], [email protected] Received 18 January 2006 Accepted 16 February 2006

Juvenile nasopharyngeal angiofibroma (JNA) is a histologically benign, non-encapsulated, vascular tumour originating in the nasopharynx [1]. Although histologically benign, this tumour is highly aggressive and is associated with significant morbidity and occasional mortality [2]. In most cases, patients present with minor symptoms of a painless, unilateral nasal obstruction and epistaxis [3].

In the diagnosis of JNA, endoscopic nasopharyngoscopy, computed tomography (CT) and magnetic resonance imaging (MRI) are the primary diagnostic tests. In addition to these techniques, because of its high vascularity, angiography is used for vascular mapping and pre-operative embolization of the tumour [5,6]. Blood pool scintigraphy with 99mTc-labelled red blood cells (99mTc-RBCs) is often performed in vascular lesions [7–9].

JNA must be differentiated from nasopharyngeal masses, such as choanal polyp, chordoma, nasopharyngeal carcinoma, rhabdomyosarcoma and adenoid hypertrophy. In such patients, it may be necessary to make the initial diagnosis using non-invasive methods, because this vascular tumour may lead to severe bleeding on biopsy [4].

The aim of this study was to determine the diagnostic accuracy of 99mTc-RBC single-photon emission computed tomography (SPECT) in the investigation of suspected JNA. We evaluated 99mTc-RBC SPECT in comparison with CT/MRI for the detection of JNA.

* Presented at the Annual Congress of the European Association of Nuclear Medicine, Vienna, Austria, August 31–September 4, 2002.

Materials and methods Ten patients (nine males, one female) with JNA were examined in this study. Their ages ranged from 12 to 14 years (Table 1). All patients underwent CT and/or MRI. These imaging methods were used to determine the

c 2006 Lippincott Williams & Wilkins 0143-3636

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490 Nuclear Medicine Communications 2006, Vol 27 No 6

Table 1

Clinical presentation and radiological and scintigraphic findings of the patients Scintigraphic findings

Patient 1, 12 years, male 2, 12 years, male 3, 13 years, male 4, 14 years, male 5, 12 years, male 6, 12 years, male 7, 12 years, male 8, 12 years, male 9, 13 years, male 10, 14 years, female

Diagnosis

Findings

Perfusion

Blood pool

SPECT

Method providing diagnosis

Final diagnosis

JNA JNA JNA JNA JNA JNA JNA JNA JNA JNA

R, E, NO E, NO, S R, NO, S, TD E, NO R, NO, TD E, NO, TD E, NO, S R, NO, S E, NO, S R, E, NO

— — — — — — — — — —

— — — — — — — — — +

+ + + + + + + + + +

CT/MRI CT/MRI CT/MRI CT/MRI CT/MRI CT/MRI CT/MRI CT/MRI CT/MRI CT/MRI

JNA JNA JNA JNA JNA JNA JNA JNA JNA JNA

CT, computed tomography; E, epistaxis; JNA, juvenile nasopharyngeal angiofibroma; MRI, magnetic resonance imaging; NO, nasal obstruction; R, rhinorrhoea; S, snoring; TD, tubal dysfunction.

Fig. 1

99m Tc-labelled red blood cell single-photon emission computed tomography (99mTc-RBC SPECT) findings. Blood pool scintigraphy with 99mTcRBCs shows no increased activity in the dynamic image (a) or in the blood pool phase (b) in the nasopharynx area, but the 99mTc RBC SPECT image shows abnormal activity in the same area.

tumour site and its extension, with special attention being given to skull base involvement, intracranial spread and the relationship to important vascular and neurological structures. For all patients, after completion of CT and/or MRI, 99m Tc-RBC SPECT images were performed. The patients were evaluated scintigraphically with 99mTc-RBCs using a modified in-vivo method [10]. Thirty minutes after the injection of pyrophosphate solution containing SnCl2, 4 ml of venous blood was sampled. The blood was mixed with 99mTc-pertechnetate, and allowed to stand at room

temperature for 20 min; immediately after the rapid intravenous injection of 370–740 MBq of 99mTc-RBCs, dynamic imaging of 1-min duration was performed. After dynamic imaging, a second-phase (blood pool phase) static acquisition at 5 min was performed. A rotating gamma camera (GE-Starcam 4000 XR/T, St Albans, Hertfordshire, UK) was used, with a 64  64 matrix, 3601 rotation and 64 projections of 30 s for each acquisition. The images were pre-filtered using a Butterworth filter. Transaxial, coronal and sagittal slices were reconstructed and displayed on a 64  64 matrix. Two

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Scintigraphy in angiofibroma Uslu et al. 491

Fig. 2

99m

Tc-labelled red blood cell single-photon emission computed tomography (99mTc-RBC SPECT) shows abnormal activity in the nasopharynx area.

Fig. 3

Computed tomography (CT), magnetic resonance imaging (MRI) and MRI angiography findings. Axial CT scan (a) shows a mass in the nasopharyngeal cavity. Axial T-weighted MRI (b) shows a mass in the same area. MRI after gadolinium injection (c) shows a mass in the same area. MRI angiography (d) shows blood supply of maxillary artery with hyperintense uptake of contrast in the mass (probably due to angiofibroma).

experienced nuclear medicine physicians interpreted all SPECT images.

Fig. 4

The scintigraphic appearance of JNA is considered to be typical when the lesion appears ‘hot’ on 99m Tc-RBC SPECT images. All patients’ final diagnoses were based on the findings of MRI/CT and follow-up by surgery.

Results 99m

Tc-RBC SPECT images were performed before definitive imaging (CT/MRI or MRI angiography) in all patients. All patients with JNA had no activity in the first phase, and only one patient had mild activity in the second phase (blood pool phase). All patients with JNA showed a prominent increase in activity on SPECT images (Figs. 1 and 2).

Histopathological findings of juvenile nasopharyngeal angiofibroma (JNA). Cells comprising the microvasculature, fibroblastic (stromal) cells of varying but distinctive morphology, inflammatory cells and dilated, cavernous vascular spaces lined by endothelial cells and separated by fibrostroma with stromal cell nuclei. These findings are compatible with a diagnosis of JNA.

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Fig. 5

Post-operative 99mTc-labelled red blood cell single-photon emission computed tomography (99mTc-RBC SPECT) images of patients with juvenile nasopharyngeal angiofibroma (JNA). Computed tomography (CT) scan with no mass in the nasopharynx (a), and 99mTc-RBC SPECT image with no uptake in the same area (b).

In our patients, only four agreed to operation. On the basis of the clinical and radiological findings (Fig. 3), a pre-operative diagnosis of vascular tumour, most probably JNA, was made, and an incisional transnasal biopsy was taken from the masses. The histopathological findings of JNA (Fig. 4) are as follows: cells comprising the microvasculature, fibroblastic (stromal) cells of varying but distinctive morphology, inflammatory cells and dilated, cavernous vascular spaces lined by endothelial cells and separated by fibrostroma with stromal cell nuclei. These findings were compatible with a diagnosis of JNA.

The post-operative follow-up included clinical examination, CT and 99mTc-RBC SPECT at 1 month for patients with JNA. CT scan performed 1 month after surgery showed no mass in the nasopharynx (Fig. 5a); 99mTc-RBC SPECT images also showed no uptake in the same area (Fig. 5b).

Discussion JNA accounts for less than 0.05% of all benign lesions that originate in the nasopharynx. It is histologically benign, but is characterized by local aggressive growth. Its clinically

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Scintigraphy in angiofibroma Uslu et al. 493

malignant behaviour is a result of its propensity for locally destructive growth, including the nasal cavity, sphenoid sinus and sella, pterygomaxillary fossa, infratemporal space, inferior orbital fissure and intracranial region [2,4]. The diagnosis of angiofibroma is made clinically from the history of a young or adolescent male with nasal obstruction, epistaxis or both, and the presence of a soft tissue mass in the nose or nasopharynx; expansion of the tumour may lead to facial deformity and proptosis if the orbit is invaded [4,11–14]. The symptoms of JNA are numerous and non-specific. As JNA is often more extensive than can be appreciated on physical examination, further diagnostic studies are required [15]. Therefore, surgeons are unwilling to undertake biopsy of a nasopharyngeal mass in an adolescent subject, but prefer to rely on imaging methods to decide whether the mass is likely to be an angiofibroma. In this context, the advent of CT, MRI and selective angiography with embolization has revolutionized the operative management of JNA. CT and MRI are used to determine the tumour site and its extension, with special attention given to skull base involvement, intracranial spread and the relationship to important vascular and neurological structures [16]. MRI is thought to be the best imaging modality for the initial diagnosis and staging of JNA. Angiography is important in determining the pattern of vascular supply, the vascular composition of the tumour and its venous drainage. Blood pool scintigraphy with 99mTc-RBCs is used to image vascular lesions, as it describes the degree of blood supply or vascularity of the mass [4,7,11–14,17–36]. Castor et al. [37] first described the usefulness of radionuclide angiography in angiofibroma with 99mTcpertechnetate. Guneri et al. [38] used 111In-octreotide scintigraphy in angiofibroma. Although 99mTc-RBCs have not been used in JNA previously, Fiore et al. [8] and ElDesouki et al. [18] revealed the use of 99mTc-RBC scintigraphy in head and neck haemangiomas. Licht et al. [39] used 99mTc-RBC scintigraphy for the diagnosis of splenic haemangioma. 99mTc-RBC SPECT has high specificity in detecting haemangioma, and is considered as first choice for differentiating haemangioma from other hepatic lesions [25,40,41]. It has been suggested that, if the mass is smaller than 2 cm, planar imaging is required; if larger than 2 cm, SPECT is necessary [8,39]. In our study, patients with JNA showed increased activity on 99m Tc-RBC scintigraphy. 99mTc-RBC scintigraphy represents a simple and non-invasive technique with low radiation exposure, evaluates the quality and quantity of vascularization, and is very useful in pre-operative detection, localization and follow-up of JNA.

Conclusion 99m

Tc-RBC scintigraphy offers a convenient and safe method to assess the vascularity of a mass and is particularly applicable for the diagnosis and follow-up after operation of angiofibromas of the nasopharynx.

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24 Siegel A, Mazurek R. Early dynamic SPECT acquisition for the imaging of hepatic hemangiomas utilizing Tc-99m labeled red blood cells. Clin Nucl Med 1997; 22:745–748. 25 Ziessman HA, Silverman PM, Patterson J, Harkness B, Fahey FH, Zeman RK, Keyes JW Jr. Improved detection of small cavernous hemangiomas of the liver with high-resolution three-headed SPECT. J Nucl Med 1991; 32:2086–2091. 26 Sayit E, Durak I, Capakaya G, Yilmaz M, Durak H. The role of Tc-99m RBC scintigraphy in the differential diagnosis of orbital cavernous hemangioma. Ann Nucl Med 2001; 15:149–151. 27 Krausz Y, Levy M, Antebi E, Bar-Ziv J, Bocher M, Chisin R. Liver hemangioma. A perioperative Tc-99m RBC SPECT correlation. Clin Nucl Med 1997; 22:35–37. 28 Ji EK, Ryu JS, Kang GH, Moon DH, Auh YH, Lee HK. Pelioid-type hepatocellular carcinoma masquerading as a hepatic hemangioma on technetium-99m red blood cell scintigraphy. Clin Nucl Med 2001; 26:33–35. 29 Tumeh SS, Benson C, Nagel JS, English RJ, Holman BL. Cavernous hemangioma of the liver: detection with single-photon emission computed tomography. Radiology 1987; 164:353–356. 30 Tzen KY, Yen TC, Lin WY, Tsai CC, Lin KJ. Diagnostic value of 99mTc-labeled red blood cell SPET for a solitary solid liver mass in HBV carrier patients with different echogenicities. Hepatogastroenterology 2000; 47:1375–1378. 31 Tamm EP, Rabushka LS, Fishman EK, Hruban RH, Diehl AM, Klein A. Intrahepatic, extramedullary hematopoiesis mimicking hemangioma on technetium-99m red blood cell SPECT examination. Clin Imaging 1995; 19:88–91. 32 Krause T, Hauenstein K, Studier-Fischer B, Schuemichen C, Moser E. Improved evaluation of technetium-99m-red blood cell SPECT in hemangioma of the liver. J Nucl Med 1993; 34:375–380.

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Original article

Quantitative evaluation of salivary gland dysfunction after radioiodine therapy using salivary gland scintigraphy Hasan Razaa, Aakif U. Khanb, Abid Hameeda and Ayub Khanb Aim The most frequent non-thyroidal complication of high-dose 131I therapy for thyroid carcinoma is salivary gland dysfunction, which may be transient or permanent. In this study, we assessed radioiodine-induced permanent salivary gland dysfunction using quantitative salivary gland scintigraphy. Methods Salivary scintigraphy was performed with 99m Tc-pertechnetate on 50 thyroid carcinoma patients who had been given radioiodine for thyroid ablation; 20 normal subjects were imaged as the control population. Dynamic scintigraphy was performed and time–activity curves for four major salivary glands were generated. The glandular functional parameters maximum secretion, time at maximum count and uptake ratio of the parotid and submandibular glands were calculated. Correlation of the administered dose and subjective symptoms with findings of salivary gland scintigraphy was evaluated. Results The maximum secretion and uptake ratio were decreased in 46% and 42% of patients who received radioiodine therapy, respectively. Salivary gland dysfunction correlated well with the administered dose. The parotid glands were more affected than the

submandibular glands. Fifty-two per cent of patients were symptomatic, 69.23% of whom showed salivary gland dysfunction. Conclusion Parenchymal damage to the salivary glands induced by radioactive iodine treatment can be evaluated by salivary gland scintigraphy. The impairment was worse in parotid glands and increased with the total dose. The maximum secretion and uptake ratio were found to be sufficiently sensitive to distinguish the severity c 2006 of the damage. Nucl Med Commun 27:495–499
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:495–499 Keywords: 131I therapy, salivary gland dysfunction, salivary gland scintigraphy, thyroid carcinoma a Atomic Energy Medical Centre, Jinnah Post Graduate Medical Centre, Karachi and bInstitute of Radiotherapy and Nuclear Medicine, Peshawar, Pakistan.

Correspondence to Dr Hasan Raza, 307 Jiwani Homes, 261 Garden East, Karachi, Pakistan. Tel: 92212251739; e-mail: [email protected] Received 10 January 2005 Accepted 17 February 2006

Introduction

Materials and methods

Oral administration of 131I has been a commonly accepted procedure for the treatment of benign and malignant conditions of the thyroid since the 1940s [1]. During this long period of continuous use, favourable therapeutic outcomes have been proven. However, treatment is not without side-effects. The most frequent non-thyroidal complication of high-dose 131I therapy is salivary gland dysfunction, which may be transient or permanent [2,3]. It presents with dry mouth, hypogeusia/dysgeusia and excessive dental caries [4]. The effect is due to radiation damage to the salivary glands, as the salivary glands concentrate iodine by a carrier-mediated system of Na131I, similar to that of the thyroid [4].

This was a cross-sectional study for the evaluation of salivary gland dysfunction. For this, salivary gland scintigraphy was performed using 99mTc-pertechnetate. Prior approval from the Ethical Committee of the Jinnah Post Graduate Medical Centre, Karachi, Pakistan was obtained. Seventy subjects ranging in age from 20 to 70 years (mean age, 41 years) were included in the study after informed and written consent had been obtained. All subjects were divided initially into two major groups: group I consisted of a control population (these subjects had no complaint or evidence of salivary gland or thyroid dysfunction), and group II consisted of patients who had received high-dose radioactive 131I therapy in the past. Group II patients had received treatment for the management of well-differentiated thyroid carcinoma (Table 1), and were further subdivided into three subgroups according to the total radioactive doses received (Table 2).

The purposes of this study were to quantify the damage to the salivary glands produced by 131I treatment for thyroid ablation and to evaluate the damage occurrence rate in relation to dose. We also assessed the correlation between the symptoms related to salivary gland dysfunction and the objective findings observed scintigraphically.

Selection criteria and patient preparation

Patients with differentiated thyroid carcinoma, who had undergone radioiodine therapy at least 6 months prior to

c 2006 Lippincott Williams & Wilkins 0143-3636


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496 Nuclear Medicine Communications 2006, Vol 27 No 6

Characteristic Age (range) (years) Sex (male/ female) Tumour type Papillary Follicular

Table 2

Group I (n = 20)

Group IIa (n = 18)

Group IIb (n = 17)

Group IIc (n = 15)

40 (20–60)

41 (20–65)

39 (23–70)

44 (27–60)

10/10

12/6

6/11

7/8

— —

14 4

10 7

11 4

Subgroups of group II on the basis of cumulative activity

received Subgroup

Cumulative dose received (mCi)

Cumulative dose received (MBq)

Total patients in this group

Group IIa Group IIb Group IIc

50–149 150–299 Z 300

1850–5513 5550–11063 Z 11100

18 17 15

the study, were selected for the study. The control group contained subjects who had no complaints or evidence of salivary gland or thyroid dysfunction. Patients who were taking any drugs that could affect the secretion of saliva (e.g. anticholinergics, antidepressants, antihypertensives, antispasmodics and antihistamines), or who had a tumour, stone or inflammation of the salivary gland, were not included in the study. Patients with autoimmune or severe debilitating disease, and pregnant or lactating women, were also excluded from the study. Patient evaluation

All subjects were evaluated objectively as well as subjectively. For subjective evaluation, all were interviewed about symptoms related to salivary gland dysfunction, such as dry mouth, hypogeusia/dysgeusia and excessive dental caries. The symptoms were not considered to result from radiation effects of 131I if the patients had similar complaints before treatment. For objective evaluation, both groups were imaged in the same manner. All patients were instructed to refrain from eating, drinking or performing any oral hygiene for at least 90 min prior to scintigraphy to ensure a resting secretory state. Imaging protocol

Patients were positioned supine with the neck hyperextended and imaged on a large field of view, singleheaded gamma camera fitted with a low-energy, allpurpose collimator. Each patient received an intravenous injection of 10 mCi (370 MBq) of 99mTc-pertechnetate. Immediately after administration, sequential dynamic images were taken at 1 min/frame on a 128  128 matrix for 25 min. At the 20th minute after the injection, 10 ml of lemon juice (50% concentrated) was administered via a

syringe and butterfly tubing into the patient’s mouth for the stimulation of saliva. Data processing

Composite images of uptake (1–20 min) were obtained. Irregular regions of interest (ROIs) were drawn manually over each of the parotid and submandibular glands. The sublingual glands generally were not visible. A squareshaped uniform background region was drawn inferior and lateral to the left submandibular gland. Time–activity curves of uptake and washout of pertechnetate were generated using counts per minute. As shown in Fig. 1, the following points were designated on the time–activity curve: point a, initial shoulder, representing a vascular perfusion, or, in cases of an unclear shoulder, at 1 min; point b, the maximum count before stimulation; point c, the minimum count after stimulation. On the basis of these ROI counts and subsequent time– activity curve, the following functional indices were derived for each salivary gland; they have been used previously by Adams et al. [5] and Aung et al. [6] in their studies on Sjo¨gren’s syndrome. Tmax ðminÞ ¼time at which cmax was reached on the time  activity curve Maximum secretion ð%Þ ¼ ðcmax  post countsÞ=cmax 100 Uptake ratio ¼ ðcmax =background countsÞ where cmax is the prestimulatory maximum count value and ‘post counts’ is the post-stimulatory minimum count value. When a gland could not be visualized, a value of zero was given for the maximum secretion and uptake ratio, but no value was recorded for Tmax in these cases. Any index that was less than one standard deviation of the mean of the respective index of the control group was considered to be abnormal. Staging was performed according to the number of glands affected in terms of maximum

Fig. 1

(b)

Counts

Table 1 Distribution of control and patient populations in different groups with various characteristics

(a) (c)

Time (min) Time–activity curve of salivary gland scintigraphy.

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Evaluation of Salivary gland dysfunction after radioiodine therapy Raza et al. 497

secretion: Stage I, patients whose glands all showed normal maximum secretion; Stage II, patients in whom one or two glands were affected in terms of maximum secretion; Stage III, patients in whom three or four glands were impaired in terms of maximum secretion. Statistical analysis

The Mann–Whitney U-test was used to assess the dose-dependent changes in salivary gland function. The chi-squared test was used to compare the objective involvement of salivary glands between symptomatic and asymptomatic patients. The chi-squared test was also used to study the relationship between the cumulative dose of radioiodine and the subjective symptoms. Statistical comparisons between right- and left-sided glands within the same group were performed using the Wilcoxon matched pairs signed-rank test. Statistical significance was set at P < 0.05 and vice versa for the Wilcoxon signed-rank test.

(11 100 MBq), severe parenchymal damage in the parotid and submandibular glands was observed in terms of maximum secretion. The uptake ratio was affected similarly, showing a decrease, with a significant deterioration in the parotid glands of group IIb patients and all salivary glands of group IIc patients. When comparing the effects of the various indices, it was found that the maximum secretion was affected more than the uptake ratio and Tmax. Data manipulation in terms of the number of patients indicated that 23 of the 50 patients in group II (46%) showed reduced maximum secretion and 21 of the 50 (42%) showed a reduced uptake ratio. From these values, the difference in the utility of these two indices is not significant, but, when the data were manipulated in terms of individual glands, it was found that eight glands had a normal uptake ratio but reduced maximum secretion.

Results When the data were compared on both sides in the same group, no statistically significant side-to-side difference was observed. Therefore, the data relating to the left and right salivary glands were pooled for further evaluation. However, two patients, one in group IIa and one in group IIb, had the right parotid gland knocked out whilst the left showed some degree of parenchymal function. A comparison of the scintigraphic parameters (i.e. Tmax, maximum secretion and uptake rate) between normal controls and patients is summarized in Table 3 (mean ± SD and P values). The time at which maximum uptake in the glands was achieved (Tmax) was found to be increased in most cases, but a statistically significant difference was not observed when compared with the controls. The maximum secretion of the salivary glands was found to be decreased in all groups with increasing cumulative dose of radioiodine. Group IIa contained patients who received less than 150 mCi (5550 MBq) of radioiodine; their maximum secretion was not affected significantly. However, the parotid glands of group IIb patients showed a marked reduction in maximum secretion (P < 0.05); the submandibular glands were not affected significantly. With a cumulative dose of more than 300 mCi Table 3

When patients were asked about symptoms related to salivary gland dysfunction, 26 of the 50 (52%) were found to be symptomatic. However, only 18 of these 26 patients (69.23%) were objectively proven to have evidence of salivary gland dysfunction in terms of maximum secretion. Only five of the 23 (21.74%) objectively positive patients were found to be asymptomatic (Table 4). A comparison between patients who showed subjective complaints of salivary gland dysfunction and those without, in patients with scintigraphic findings of salivary gland dysfunction, demonstrated a strong association with P = 0.001 (Table 4). However, symptoms were not dosedependent (Table 5). Dry mouth was reported to be the most common discomfort (52.94%), followed by dental caries (47%).

Table 4

Correlation between symptomatology and scintigraphic

findings Symptoms (positive)

Symptoms (negative)

Total

18 8 26

5 19 24

23 27 50

Scintigraphy (positive) Scintigraphy (negative) Total

Degrees of freedom (DF) = 2, w = 11.768, P = 0.001.

Comparison of scintigraphic parameters in controls and patients

Tmax of parotids Tmax of submandibulars Maximum secretion of parotids Maximum secretion of submandibulars Uptake rate of parotids Uptake rate of submandibulars

I (control)

IIa (P )

IIb (P )

IIc (P )

16.70 ± 1.92 11.38 ± 3.26 69.51 ± 8.39 56.32 ± 9.00

16.82 ± 2.02 (0.211) 12.89 ± 2.70 (0.238) 61.32 ± 11.69 (0.287) 53.17 ± 8.42 (0.413)

16.48 ± 2.15 (0.189) 13.95 ± 2.42 (0.255) 56.85 ± 10.01 (0.031) 50.79 ± 11.24 (0.315)

16.04 ± 1.51 (0.361) 14.85 ± 3.46 (0.123) 52.92 ± 11.11 (0.012) 44.70 ± 9.52 (0.027)

50.49 ± 10.25 36.91 ± 8.29

41.79 ± 11.22 (0.072) 27.82 ± 7.72 (0.693)

37.65 ± 12.83 (0.046) 28.41 ± 9.31 (0.224)

31.11 ± 11.23 (0.021) 22.48 ± 8.20 (0.034)

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498 Nuclear Medicine Communications 2006, Vol 27 No 6

Table 5

Correlation between symptoms and cumulative dose

Group Group IIa Group IIb Group IIc Total

Symptoms present Symptoms absent n (%) n (%) 7 11 9 27

(38.89) (64.71) (60) (54)

11 6 6 23

(61.11) (36.36) (37.50) (46.88)

Total n (%) 18 17 15 50

(100) (100) (100) (100)

Degrees of freedom (DF) = 2, w = 2.656, P = 0.265.

Our study demonstrated that, as the dose of radioiodine increased, damage to the salivary glands increased, and this was best manifested by the maximum secretion. The parotid glands were more affected than the submandibular glands. Our study also showed good correlation between objective and subjective findings.

Discussion Radioactive iodine is administered to destroy thyroid tissues, as well as metastatic foci, as they concentrate iodine. However, other tissues also accumulate radioiodine, such as the salivary glands, stomach, intestine, urinary bladder and liver [7]. Because of this localization of 131I, damage may also occur to these extrathyroidal, non-target tissues. The probable cause of salivary gland damage is radiation injury as a result of the concentration of 131I by the glands. The specific mechanism responsible for radiationinduced salivary gland dysfunction is still not understood. A variety of mechanisms, including mitotic and interphase cell death, direct DNA damage or effects of secondary metabolites, damage to progenitor cells or altered gene expression, have been proposed to explain salivary epithelial cell death [8]. It has been well documented that salivary glands concentrate iodide from the blood, the iodide concentration in mixed saliva being 30–40 times the plasma level [9]. Doniach [10] estimated that the salivary glands receive 50 cGy/mCi (135 cGy/ MBq) administered. Goolden et al. [11] measured the salivary and plasma concentrations of 131I, and estimated the radiation dose to the salivary glands to be approximately 700 rad during the first 12 h of therapy from doses of 100–200 mCi (3700–7400 MBq). Absorbed tissue doses, which may reach 700–1500 rad, are thought to induce significant tissue inflammation as a result of an obstructive process, leading to low salivary flow and changes in salivary composition [12]. The effects of irradiation on the salivary glands are irreversible when exposure exceeds 50 Gy [13]. Radiation also induces histological alterations in the salivary glands and changes in the chemical composition of saliva [13]. It is thought that radiation increases aamylase and activated kallikrein in the saliva, and induces an obstructive process leading to reduced salivary flow

[4]. Prostaglandins, which have been implicated in stimulating salivary gland function, have also been found to be decreased in patients who receive radioiodine therapy [14]. Qualitative salivary gland scintigraphy (visual analysis) is useful for the detection of severe parenchymal damage, but quantitative salivary gland scintigraphy can enable the diagnosis of mild damage, which can be missed by visual analysis alone [15]. Sometimes, increased thyroid uptake can influence the salivary gland uptake and thus mislead the diagnosis. For these reasons, we preferred quantitative salivary gland scintigraphy. Furthermore, quantitative analysis can be used for serial follow-up studies. We used three indices: Tmax, maximum secretion and uptake ratio. Tmax is the time at which maximum uptake is acquired by the salivary gland. Maximum secretion is a parameter representing the excretory function of the salivary gland, and the uptake ratio corresponds to the trapping ability of the gland. Statistically, no significant asymmetry was observed in any group, but individual observation of the glands showed that, in some patients, asymmetrical damage to the glands had occurred (two patients showed no visualization of the right parotid gland). Some authors have found more asymmetric patterns: for example, Caglar et al. [2] observed asymmetry in 51% of patients and Malpani et al. [16] noted asymmetry in 48%. We assumed that Tmax would be increased in patients receiving radioiodine therapy, as shown by Albrecht and Creutzig [17], but our study revealed that radioiodine therapy did not have profound effects on Tmax of the salivary glands, thus rendering this index ineffective in our study. However, maximum secretion appeared to be the most effective variable for salivary gland evaluation. It was significantly lower in the parotid glands of group IIb and in all salivary glands of group IIc. The uptake ratio exhibited the same trend. Olmos et al. [18] and Liem et al. [19] concluded that, initially, xerostomia is based predominantly on the failure of the glands to excrete saliva, whereas, later, a decreased trapping ability, together with a loss of secretory function, is also important. They obtained these conclusions from studies performed on 25 patients who received radiotherapy for various head and neck malignancies. Olmos et al. [18] found significant impairment in the excretory function of 50% of the salivary glands in patients who received 25–45 Gy and in almost all glands in patients who received more than 45 Gy after a short (2–7 months) and long (10–50 months) interval after radiotherapy. However, the uptake of the glands was only affected after a longer time interval. Liem et al. [19] observed a

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Evaluation of Salivary gland dysfunction after radioiodine therapy Raza et al. 499

decrease in excretory fraction at 1 month and an influence on uptake at 12 months. Our study is not fully consistent with the abovementioned studies, as post-radioiodine therapy serial follow-up scintigraphy could not be performed in our study. Therefore, comments regarding the disturbance of different parameters as a function of time cannot be made. However, it is assumed that, in due course, as the parenchymal damage worsens, these parameters will deteriorate further. Furthermore, our study was conducted on patients who received radioiodine treatment with considerable dose variation, in contrast with the above-mentioned studies which were carried out on patients who received radiotherapy. As, in radioiodine therapy, the dose received by the salivary glands is mainly dependent on the size of the thyroid gland, it is not possible to predict exactly the amount of radiation received by the salivary glands. This might also contribute to the discordance between our study and the above-mentioned reports. The function of the parotid glands was affected more often than that of the submandibular glands. Liem et al. [19] also made similar findings, as did Albrecht and Creutzig [17]. They found involvement of the parotid glands in 59% of cases and of the submandibular glands in 16%. Variations in parenchymal volume and radiosensitivity may partly explain this. The difference in radiosensitivity may be a result of the higher concentration of serous acinar cells in the parotid glands, which are selectively radiosensitive [20], whereas the mucous tissue of the submandibular glands may contribute to a radioprotective effect. To find a correlation between objective and subjective findings, we used maximum secretion as a predictable variable. We found a good association between objective and subjective findings (Table 4), as 69% of the symptomatic patients were also scintigraphically positive. A similar type of association between the presence of symptoms and objective test results was found by Caglar et al. [2] (P = 0.037) and Solans et al. [4] (P = 0.0001). Although most patients with reduced salivary gland function reported xerostomia, a few had no symptoms. The absence of subjective symptoms in these patients was probably a result of the compensatory function of the other salivary glands. The most common complaint was dry mouth (53%), followed by dental caries (47%). The onset of caries is thought to be secondary to radiationinduced xerostomia, which predisposes to an overgrowth of cariogenic flora. Caglar et al. [2] and Solans et al. [4] also showed dose-dependent symptoms, but this was not found in our study (Table 5). This may be due to variations in the hydration status, pharmacological factors and psychological conditions.

Conclusion Quantitative salivary gland scintigraphy is a useful tool for the evaluation of salivary gland dysfunction induced by radioactive iodine treatment, and the useful indices include the maximum secretion and uptake ratio. It can also be used for the follow-up of patients complaining of dry mouth and dental caries.

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Boice JD Jr. Radiation effects in nuclear medicine. In: Harbert JC, Eckelman WC, Neumann RD, editors. Nuclear medicine diagnosis and therapy. New York: Thieme Medical Publishers; 1996, pp. 279–289. Caglar M, Tuncel M, Alpar R. Scintigraphic evaluation of salivary gland dysfunction in patients with thyroid cancer after radioiodine treatment. Clin Nucl Med 2002; 27:767–771. Bader JB, Schaefer A, Finke C, Kirsch CM. Intermediate and long term side effects of high dose radioiodine therapy for thyroid carcinoma. J Nucl Med 1998; 39:1551–1554. Solans R, Bosch JA, Galofre´ P, Porta F, Rosello J, Selva-O’Callaghan A, Vilardell M. Salivary and lacrimal gland dysfunction (sicca syndrome) after radioiodine therapy. J Nucl Med 2001; 42:738–743. Adams BK, Al Attia HM, Parkar S. Salivary gland scintigraphy in Sjo¨gren’s syndrome: are quantitative indices the answer. Nucl Med Commun 2003; 24:1011–1016. Aung W, Murata Y, Ishida R, Takahashi Y, Okada N, Hitoshi Shibuya H. Study of quantitative oral radioactivity in salivary gland scintigraphy and determination of the clinical stage of Sjo¨gren’s syndrome. J Nucl Med 2001; 42:38–43. Harbert JC. Radioiodine therapy of differentiated thyroid carcinoma. In: Harbert JC, Eckelman WC, Neumann RD, editors. Nuclear medicine diagnosis and therapy. New York: Thieme Medical Publishers; 1996, pp. 975–1010. Fox PC. Acquired salivary gland dysfunction. Drugs and radiation. Ann NY Acad Sci 1998; 842:132–137. Myant NB. Iodine metabolism of salivary glands. Ann NY Acad Sci 1960; 85:208–214. Doniach I. Biologic effects of radiation on the thyroid. In: Werner S, editor. The thyroid. New York: Harper & Row; 1978, pp. 274–283. Goolden AWG, Mallard JR, Farran HEA. Radiation sialoadenitis following radioiodine therapy. Br J Radiol 1957; 30:210. DiRusso G, Kern KA. Comparative analysis of complications from I-131 radioablation for well differentiated thyroid cancer. Surgery 1994; 116:1024. Allweiss P, Braunstein GB, Katz A, Waxman A. Sialoadenitis following I-131 therapy for thyroid carcinoma: concise communication. J Nucl Med 1984; 25:755–758. Rodrigues M, Havlik E, Peskar B, Sinzinger H. Prostaglandins as biochemical markers of radiation injury to the salivary glands after iodine-131 therapy? Eur J Nucl Med 1998; 25:265–269. Bohuslavizki KH, Brenner W, Lassmann S, Tinnemeyer S, Tonshoff G, Wolf H, et al. Quantitative salivary gland scintigraphy in the diagnosis of parenchymal damage after treatment with radioiodine. Nucl Med Commun 1996; 17:681–686. Malpani BL, Samuel AM, Ray S. Quantification of salivary gland function in thyroid cancer patients treated with radioiodine. Int J Radiat Oncol Biol Phys 1996; 35:535–540. Albrecht HH, Creutzig H. Salivary gland scintigraphy after radioiodine therapy. Functional scintigraphy of the salivary gland after high dose radioiodine therapy. Fortschr Roentgenstr 1976; 125: 546–551. Olmos RA, Keus RB, Takes RP, van Tinteren H, Hilgers FJ, Baris G, et al. Scintigraphic assessment of salivary function and excretion response in radiation-induced injury of the major salivary glands. Cancer 1994; 73:2886–2893. Liem IH, Olmos RA, Balm AJ. Evidence for early and persistent impairment of salivary gland excretion after irradiation of head and neck tumors. Eur J Nucl Med 1996; 23:1845–1890. Stephens LC, Schulthesis TE, Price RE, Ang KK, Peters LJ. Radiation apoptosis of serous acinar cells of salivary and lacrimal glands. Cancer 1991; 67:1539.

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Original article

Hand perfusion with 99mTc-HSA in patients expecting to undergo coronary bypass surgery: elaboration of a new complex diagnostic protocol for the safe removal of a radial artery graft Ildiko´ Garai, Zolta´n Csiki, Jo´zsef Varga, La´szlo´ Galuska, Lajos Patonay, Lajos Szabados, A´rpa´d Pe´terffy and Zolta´n Galajda Background The Allen test is used worldwide for radial artery graft removal. The postoperative examination of our patients’ hand function and circulation proved that beside the transient neurological complications chronic hand circulatory disorders may arise. Aim To develop a non-invasive method suitable for an objective evaluation of the hand’s circulation to make it possible to use radial arteries safely for the revascularization of coronary arteries. Methods We examined 35 patients. After selective compression of the radial and ulnar arteries of both hands, we injected 400 MBq 99mTc-HSA intravenously and acquired 240 images, each of 1 s. After 30 s we released the ulnar artery first, and after 120 s the radial artery, too. Then computer analysis was performed.

plateau even after 30 s. Following the release of the radial artery a further increase in the activity could be observed. We assume that the latter patient group is at risk of consequent circulatory disorder of the fingers after the removal of the radial artery, whereas in the former group the artery could be removed safely. Conclusions Hand perfusion with 99mTc-HSA is useful in patients selected for coronary bypass operations, so we recommend the introduction of this method as a routine examination before the removal of the radial artery in patients with an abnormal Allen test. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:501–506 Nuclear Medicine Communications 2006, 27:501–506 Keywords: scintigraphy, coronary bypass, radial harvesting University of Debrecen, Hungary.

Results The patients could be divided into two groups. In the majority of them, releasing only the ulnar artery resulted in a good circulation of the fingers. It meant that the time–activity curve rapidly reached its maximum, and the activity did not change even after releasing the radial artery. In a smaller proportion of the patients the activity of the fingers increased only slowly, and did not reach a

Introduction Since Carpenter first used radial artery grafts for coronary bypass operations, this artery has been widely used for total artery revascularization due to positive early and mid-term results [1,2]. In our university radial arteries have been used for this purpose since 1998. In 2002, 759 coronary bypass operations were performed, out of which in 354 cases of radial artery graft were used for bypass. Despite the large number of interventions, there have been few post-operative complications [3]. However, together with the Allen test there was a justified need for the clinical staff to have a non-invasive screening method that could be used for the safe removal of the radial artery. The aim of our study was to develop a non-invasive procedure suitable for objective evaluation of the hand’s

Correspondence to Dr Ildiko Garai, Nagyerdei KRT 98, Debrecen 4012, Hungary. Tel: + 36 52 422870; fax: + 36 52 422870; e-mail: [email protected] This work was supported by the UDMHSC Mecenatura Grant. Received 16 January 2006 Accepted 24 February 2006

circulation to make it possible to use radial arteries safely for coronary bypass operations.

Patients, materials and methods Patients

We examined 35 patients (nine women and 26 men) waiting for coronary bypass surgery. Their average age was 50.8 years (range, 31–67 years). We excluded patients whose histories included hand injury. Those who reported a sense of disorder in their fingers, discoloration caused by cold, joint pain or movement restriction were also excluded if colour Doppler examination detected a circulation disorder in the radial and/or the ulnar artery. We did not classify the patients according to the dominance of the use of the hands. Before scintigraphy,

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502

Nuclear Medicine Communications 2006, Vol 27 No 6

the same examiner performed the modified Allen test on both hands of all patients. In the Allen test we set the normal recoloration limit to 10 s. Pre-operative examination

The examination required no preparation. Patients placed their hands with the palms down on the collimator of the upward-turned gamma camera. Before injection of the radiopharmaceutical, we blocked the radial and the ulnar artery on the pressure points of the pulse with a special clamp made for this purpose. We controlled the stoppage of the blood flow with a manual Doppler meter distal from the clamp (Fig. 1). We injected about 400 MBq 99m Tc-HSA in a bolus into the right cubital vein. We recorded the exact activity before injecting the activity remaining in the syringe and the time of the measurements. An MB-9200 gamma camera (Gamma Works, Hungary) mounted with a parallel-hole collimator was used. We started the acquisition of 240  1-s dynamic series of images immediately after the injection. After 30 s we released the clamps of the ulnar arteries on both sides, while the radial arteries stayed clamped. We released the radial arteries in the 120th second. The examination was continued for a further 120 s in the same position. Post-operative examination

The patients whose radial artery was removed were sent for a second (control) hand perfusion examination at a mean of 23.8 months (range, 12–28 months) after coronary bypass surgery. The patients, in a similar manner to the pre-operative examination, put their hands on the collimator of the upturned camera. A dose of 400 MBq 99m Tc-HSA was administered through a winged needle inserted into the cubital vein. We registered the amount of activity in the syringe before and after the injection Fig. 1

together with the time of measurement. A series of 240  1-s images was started immediately after the injection. Data analysis Evaluation of the pre-operative examination

We used a dedicated computer program DIAG (Digital Image Analyzer for Gamma Camera), for analysing the hands separately. In the final quantitative data analysis 66 hands were involved (after excluding the hands that moved during the examination, missing data, and the patients who refused to cooperate). In the summed image of the whole series, we outlined the regions of the palm and the fingers (Fig. 2). Finger 1 was excluded from the region of interest (ROI) due to its special blood supply. The shape of the time–activity curve was first visually assessed and then the radial perfusion index (RPI) and ulnar perfusion index (UPI) were calculated as the ratios of the areas under the suitably selected sections of the time–activity curves of both the fingers and the palm, as shown in Fig. 3. Beside these data, we determined the fingers-to-palm ratio (FPR) for each patient, which characterized the microcirculation of the fingers. Post-operative analysis

We introduced a new parameter for the comparison of the conditions before and after the operation. After summing the images in the interval between 150 and 240 s after the injection, we drew the regions of both the palm and fingers II–IV, and expressed the total and maximal activities inside these regions as ‘% of the injected dose/cm2’.

Results The pre-operative examination

Statistical analysis was performed for 66 hands, regardless of the left or right side. With the physical Allen test, the measured recoloration period was under 10 s for 48 hands, while for another 18 it was longer. The perfusion study Fig. 2

Compression of selected radial and ulnar arteries.

Regions of interest drawn around the fingers and palms.

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Perfusion with

Fig. 3

99m

Tc-HSA in prospective coronary bypass patients Garai et al. 503

Fig. 5

0.7

Palms

0.5

60 C

0.4 B

0.3 A

0.2

Cps

Counts/pixel/s

0.6

40

Release

0.1 0

Radial Ulnar 0

30

60

90

120 150 Time (s)

180

210

240

270 20 Fingers

Time–activity curves plotted in accordance with the marked regions (ROIs). The degree of the ulnar and radial perfusion was derived from the ratio of the areas under the curve (the time–activity curves both of the fingers and palm). Ulnar perfusion index = area C/area A. Radial perfusion index = area C/area B, where the interval was 45 s.

0

1

2 Min

3

4

When both ulnar arteries are released the activity of the hands remains low. When the radial artery is released a further increase of the activity occurs.

Fig. 4

Palms

Table 1

Perfusion indexes of the positive and negative Allen’s test

group Perfusion index

Cps

100

75

Palmar ulnar Palmar radial Finger ulnar Finger radial

Fingers

Allen’s test Negative (n = 48)

Positive (n = 18)

53.35 ± 22.206 95.48 ± 10.985 63.94 ± 26.973 97.21 ± 10.697

37.39 ± 28.879 90.67 ± 13.052 36.33 ± 33.020 88.56 ± 12.316

50

the ulnar artery the activity of the fingers and the palm just slowly increased, and after releasing the radial artery it rose further (Fig. 5).

25

0

1

2 Min

3

4

A steady state develops when both ulnar arteries are released.

was unsuccessful in four cases because of incomplete block of the arteries. Based on the shape of the time– activity curves, the patients were divided into two groups. In the majority of patients (55 cases) after releasing the ulnar artery the activity of the fingers and the palm rapidly increased, reaching a plateau in a short time. The activity of the fingers did not change any more after the release of the radial artery (Fig. 4). In the minority of the examinations (11 cases), however, following the release of

The fingers-to-palm ratio (FPR) in the relaxed condition of the microcirculation of the fingers was not different between the patients of the positive and the negative groups. The average FPR in the patients with a negative Allen test was 0.589 ± 0.114, while in the positive group it was 0.597 ± 0.113 (P > 0.05). The perfusion indices calculated for the region of the fingers were as follows: after the release of the ulnar artery (ulnar perfusion index), in patients with negative Allen test: 63.94 ± 26.973, and in those with a positive Allen test: 36.33 ± 33.020. Following the radial release (radial perfusion index) the values were: 97.21 ± 10.697 (negative Allen test) vs. 88.56 ± 12.326 (positive Allen test) (Table 1).

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504 Nuclear Medicine Communications 2006, Vol 27 No 6

Fig. 7

Ulnar perfusion index: Palm Radial perfusion index: Palm Ulnar perfusion index: Fingers Radial perfusion index: Fingers Fingers-to-palm ratio

Allen test

p

n

0

25

50

75

100

20 Mean = 59.14 SD = 11.337 n = 66 15

Frequency

Fig. 6

10

5

125

Box and whisker plot. The coloured rectangles reflect the interquartile ranges which contain 50% of the values. The thin vertical lines represent the lower and higher values, the circles the outline values and the thick black lines inside the rectangle represent the median values.

0 30

We measured FPR values under 0.45 in eight hands, which indicates low finger microcirculation similar to the patients with Raynaud’s syndrome [4] (Fig. 7). However, out of these patients, by using the Allen test we measured prolonged recoloration time in only two hands. In the other cases, in correlation with the results of the Allen test, although we measured low FPR values by the dynamic hand perfusion examinations, by releasing the ulnar artery the activity balance of the fingers recovered before releasing the radial artery (Fig. 8).

50 60 70 80 Fingers-to-palm ratio

90

100

The frequency of fingers-to-palm ratios of patients before the operation.

Fig. 8

80

Palms

60 Cps

The ulnar perfusion rates of the fingers of patients with prolonged recoloration time were significantly lower (P < 0.001), as well as the palmar ulnar perfusion rates (P < 0.05). There was no difference between the fingersto-palm ratios (P > 0.05) (Fig. 6).

40

40

The post-operative examinations (Fig. 9)

All 33 patients underwent coronary bypass surgery. In six patients bilateral, in 12 only left-hand radial artery removal was done based on the hand perfusion scintigraphy. In 15 patients venous grafts were used. All 18 patients with radial artery removal underwent a control hand perfusion study more than 1 year after the heart surgery. Following the operation, perfusions of the fingers remained normal, and we did not find significant differences between the pre-operative and post-operative activity uptake values (expressed in %/cm2) in any region (palm, fingers II–V, finger I, and finger V: P > 0.05) (Figs 10 and 11). One patient of those who had lower FPR values (under 0.45) and prolonged recoloration time, had

Fingers

20

1

2 Min

3

Beneath low digital blood flow (fingers-to-palm ratio, left side: 0.45 and right side: 0.42) with the release of the ulnar artery the steady state had already developed.

undergone left radial artery removal based on hand perfusion scintigraphy. His hand perfusion without radial artery remained normal and did not decrease compared to the opposite side.

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Perfusion with

99m

Tc-HSA in prospective coronary bypass patients Garai et al. 505

Fig. 9

Hand perfusion scintigraphy before (a) and after (b) radial artery harvesting.

Fig. 10

Fig. 11

0.08 0.06

0.001 0.000

Palm

Finger all

I.finger

V.finger

−0.001 −0.002

Change of total activity

Change of maximal activity

0.002

0.04 0.02 0.00 − 0.02

Palm

Finger all

I.finger

V.finger

− 0.04 Confidence intervals of the change of maximal activity at P < 0.05 level. The change of maximal activity could not be observed in the palms and the fingers after radial artery harvesting.

− 0.06 Confidence interval of the change of total activity at P < 0.05 level. The change of total activity could not be observed in the palms and the fingers after radial artery harvesting.

Discussion The arterial blood supply of the hand is provided by the radial and ulnar arteries. There are two larger connections between the two arteries: the superficial and the profound palmar arches. If this anastomotic system is developed, removal of one of the arteries does not cause a significant disorder in the circulation of the hand. In case the connection is not developed or is missing, the removal of one artery (e.g., the radial one) can lead to ischaemia of the hand. Despite the fact that most authors give accounts of mainly neurological and sensory disorders, especially in the early post-operative phase, circulation disorders of the fingers can also be expected [5,6]. Nowadays, the Allen test is the standard screening method to assess if the radial artery can be removed

without damaging the circulation of the hand. It is widely used despite the fact that within the 6-s recoloration limit the sensitivity of the Allen test is only 54.5%, while its specificity is 91.7% and its diagnostic reliability is 78.5% [7]. If we set the recoloration limit longer, the sensitivity can be increased but the specificity becomes worse. So there is a great need for more objective examination methods like quantitative hand perfusion scintigraphy, which are suitable, either individually or in combination with the Allen test, to assess the circulation of the hand

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506 Nuclear Medicine Communications 2006, Vol 27 No 6

[8,9]. Several non-invasive and more invasive techniques have been tried for this purpose [10–12].

objective indicator that is valuable for both planning the removal of the radial artery, and the follow-up monitoring thereafter.

99m

Tc-HSA used in our method remains intravascular after intravenous administration, so in any selected region the measured activity is proportional to its blood content. Based on the shape of the time–activity curves, we divided our patients into two groups. We suppose that in patients whose activity level of fingers II–IV did not increase further after the release of the radial artery, the blood supply through the ulnar artery is dominant, and they have an appropriate collateral network. On the contrary, if following the release of the radial artery the increase in the level of the activity of the fingers is measurable, it indicates that the radial artery contributes significantly to the proper blood supply of the fingers, or the anastomosis is not appropriate between the two systems. However, it does not imply directly that these patient’s radial arteries cannot be removed. We consider that if in such cases the use of the radial artery is inevitable, the follow-up of the patient should be stricter. Although Rafael et al. [13] suggested that the removal of the radial artery reduces the tissue perfusion of the hand, it is remarkable that 1 year after the operation we could not detect any sign of decrease in the blood stream in the regions of the palm, fingers II–IV, or finger I and finger V of our patients considered to have a ‘good ulnar circulation’. Our examinations indicate that there is no direct correlation between ‘cold fingers’ and the ulnar circulation, so the question arises if the removal of the radial artery is really contra-indicated in cases of Raynaud’s syndrome. On the basis of our results we think that the method developed by our team is a sensitive and

References 1

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Hagiwara H, Ito T, Kamiya H, Akita T, Usui A, Ueda Y. Mid-term structural change in the radial artery grafts after coronary artery bypass grafting. Ann Thorac Surg 2004; 77:805–810. Meharwal ZS, Trehan N. Functional status of the hand after radial artery harvesting: results in 3,977 cases. Ann Thorac Surg 2001; 72:1557–1561. Galajda Z, Szentkiralyi I, Peterffy A. Neurologic complications after radial artery harvesting. J Thorac Cardiovasc Surg 2002; 123:194–195. Galuska L, Garai I, Csiki Z, Varga J, Bodolay E, Bajnok L. The clinical usefulness of the fingers-to-palm ratio in different hand microcirculatory abnormalities. Nucl Med Commun 2000; 21:659–663. Budillon AM, Nicolini F, Agostinelli A, Beghi C, Pavesi G, Fragnito C, et al. Complications after radial artery harvesting for coronary artery bypass grafting: our experience. Surgery 2003; 133:283–287. Denton TA, Trento L, Cohen M, Kass RM, Blanche C, Raissi S, et al. Radial artery harvesting for coronary bypass operations: neurologic complications and their potential mechanisms. J Thorac Cardiovasc Surg 2001; 121: 951–956. Jarvis MA, Jarvis CL, Jones PR, Spyt TJ. Reliability of Allen’s test in selection of patients for radial artery harvest. Ann Thorac Surg 2000; 70:1362–1365. Ruengsakulrach P, Brooks M, Hare DL, Gordon I, Buxton BF. Preoperative assessment of hand circulation by means of Doppler ultrasonography and the modified Allen test. J Thorac Cardiovasc Surg 2001; 121:526–531. Rodriguez E, Ormont ML, Lambert EH, Needleman L, Halpern EJ, Diehl JT, et al. The role of preoperative radial artery ultrasound and digital plethysmography prior to coronary artery bypass grafting. Eur J Cardiothorac Surg 2001; 19:135–139. Abu-Omar Y, Mussa S, Anastasiadis K, Steel S, Hands L, Taggart DP. Duplex ultrasonography predicts safety of radial artery harvest in the presence of an abnormal Allen test. Ann Thorac Surg 2004; 77:116–119. Sullivan VV, Higgenbotham C, Shanley CJ, Fowler J, Lampman RM, Whitehouse WM Jr, et al. Can ulnar artery velocity changes be used as a preoperative screening tool for radial artery grafting in coronary artery bypass? Ann Vasc Surg 2003; 17:253–259. Kochi K, Orihashi K, Sueda T. The snuffbox technique: a reliable color Doppler method to assess hand circulation. J Thorac Cardiovasc Surg 2003; 125:821–825. Rafael SJ, Conroy JL, Burniston M, Maughan J, Munsch C. Effect of radial artery harvesting on tissue perfusion and function of the hand. Cardiovasc Surg 2001; 9:378–382.

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Original article

Characterization of radiologically indeterminate lung lesions: 99mTc-depreotide SPECT versus 18F-FDG PET Nuria Ferrana, Yvonne Ricarta, Marta Lopezb, Ignacio Martinez-Ballarinb, Manel Rocaa, Cristina Ga´mezc, David Carrereaa, Sara Guiraoa, Alejandro Fernandez Leon and Jose Martin-Comina Aim To evaluate the diagnostic accuracy of 99m Tc-depreotide vs PET-18FDG scans in patients with suspicion of lung cancer Material and methods Prospective study in 29 patients (age: 38-80 years) diagnosed of inderteminate lung lesions. Diagnosis was established by histology based on samples of surgical resection, fine needle aspiration (FNA) or broncoalveolar lavage (BAL). Within a maximum of 10 days, without pre-established fixed order the following exams were performed: 1) Whole body and chest SPECT-CT with 99mTc-depreo´tide (DEP-SPECT) and 2) PET-CT study with 18F-FDG (PET-FDG). Every exam was evaluated by Nuclear Medicine especialist blinded to patient data. Result Malignancy was confirmed in 20 patients. PET-FDG was positive in all cases. DEP-SPECT was positive in 17 and falselly negative in 3, one carcinoid tumor, one undifferentiated non-small cell adenocarcinoma, and a moderately differentiated adenocarcinoma. In the remaining 9 patients benignancy was confirmed; both studies were normal in 8 and falselly positive in one case of non-specific inflammatory lung process. In 9 out of the 20 cases with malignancy extrapulmonar uptake was seen, with a total number of 19 lesions. In two cases the extrapulmonar uptake were non ganglionar metastasis (bone and adrenal) and in 7 due to mediastinic

Introduction Lung cancer is among the leading causes of death in both men and women [1]. This fact is directly related to the late occurrence of symptoms, which appear when the disease is already advanced and probably non-resectable [2]. Currently used screening procedures include chest X-rays and/or computed tomography (CT), and the probability of malignant/benign disease is estimated based on clinical factors (advanced age, smoking, prior cancer history) and on anatomical and morphological characteristics of the lesions (size, morphology, growth pattern, calcifications) [3,4].

ganglionar involvement. ROC analysis using peak SUV FDG (cut-off point of 3.5) uptake and target/background depreotide uptake (cut-off point of 1.3) provided, sensitivity and specificity values of 95% and 89% of 84% and 88% for PET and SPECT respectively. It does not exist statistically significant differences between both methods (Z-test SPSS). In summary, FDG-PET has a greater sensitivity and diagnostic accuracy for assessing malignancy of indeterminate lung lesions, and for detection of extrapulmonary involvement, DEP-SPECT represents a good diagnostic alternative for centers where PET is c 2006 not available. Nucl Med Commun 27:507–514 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:507–514 Keywords: lung cancer, Pulmonary lesions, 99mTc-depreotide, 18F-FDG-PET a Hospitalet de Llobregat, cS. Medicina Nuclear, PET-IDI and bPneumologia, Hospital Universitari de Bellvitge-Idibell, Spain.

Correspondence to Dr J. Martin-Comin, S. Medicina Nuclear, Hospital U. de BellvitgeFeixa Llarga s/n, 08907 Hospitalet de Llobregat, Spain. Tel: + 0034 932 607620; fax: + 0034 932 607516; e-mail: [email protected] This work was supported by a GE Health care Medical System grant. Received 11 November 2005 Accepted 2 March 2006

The need for early diagnosis leads to an increased detection of radiologically indeterminate lesions. Only a third of lung lesions may be typified based on radiological criteria, providing 50% sensitivity and 89% specificity [2]. In the remaining two thirds, use of invasive techniques (excluding surgical resection) represents a diagnostic alternative. Both lesion accessibility and the associated patient conditions, such as respiratory failure and haemorrhagic diathesis, determine the procedure to be used. Such procedures have a highly variable sensitivity (bronchoscopy 65%, transbronchial biopsy 80%, CT-guided transcutaneous puncture 90%) [5]. It should be remembered that these procedures are not free from complications, as they involve a risk of pneumothorax close to 10% [3].

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508 Nuclear Medicine Communications 2006, Vol 27 No 6

Molecular imaging techniques may play a significant role in optimizing use of such techniques by non-invasively selecting those patients with a greater probability of malignancy. Currently, two methods may mainly be considered. The first of such methods is single photon emission computed tomography using 99mTc-depreotide (DEP SPECT), which is widely available in conventional nuclear medicine centres, and the second is positron emission tomography using [18F]fluorodeoxyglucose (FDG PET). Depreotide is a synthetic peptide, a somatostatin analogue, that has a high affinity for somatostatin receptors (SSTRs) of the subtypes 2, 3 and 5, which are over-expressed by certain tumours, including lung cancer [6]. The high capacity of depreotide to form complexes with 99mTc allows for easy labelling, and also for obtaining high resolution images at a lower cost as compared to scintigraphy with 111In-octreotide [6]. Several studies support the safety and diagnostic accuracy of depreotide [7–9]. The values of sensitivity, specificity and diagnostic accuracy reported in the literature for depreotide range from 93 to 97%, 73 to 88%, and 90%, respectively. Special mention should be made of the multicentre study conducted by Blum et al., which included a large number of cases [9]. These authors studies 114 patients, of whom 88 were diagnosed with a malignant disease. Three false negative results (adenocarcinomas) were obtained with DEP SPECT. Malignancy was adequately excluded in 19 of the 26 benign cases, and there were seven false positive results (six granulomas and one hamartoma). The estimated sensitivity and specificity were 96.6% and 73.1%, respectively, and it was concluded that DEP SPECT was helpful and safe for diagnosing malignancy. On the other hand, over-expression of GLUT 1 receptors and increased hexokinase levels in tumour cells make FDG labelled with 18F an effective tool for diagnosing hypermetabolic malignant lesions [10]. Sensitivity and specificity of FDG PET for assessing a solitary lung lesion range from 82 to 100% and from 63 to 90%, respectively, according to several studies [10–13]. In a meta-analysis by Gould et al. [14], after assessment of 1474 patients with indeterminate lung lesions, mean values of 96% and 74% were obtained for sensitivity and specificity, respectively. In order to evaluate the diagnostic accuracy of both FDG PET and DEP SPECT in assessing malignancy of lung lesions, a study was undertaken at our centre to compare both techniques in patients with suspected lung cancer.

Material and methods This was a prospective study approved by the ethics committee of the centre which was started in February 2004 and completed in May 2005. A total of 29 patients (24 men and five women) with ages ranging from 38 to 80 years (mean age, 52 years) were enrolled into the study. The size of lesions studied ranged from 1 to 6 cm (mean size, 2.6 cm). Patient inclusion criteria were: patients over 18 years and under 80 years of age lung lesion diagnosed in a plain X-ray or/plus TC scan, 6 cm in size or smaller, rounded, ovoid or lobulated in shape, and not associated with visible satellite lesions K subsequent pathological study indicated to establish diagnosis: surgical resection of nodule, fine-needle aspiration (FNA), or bronchoalveolar lavage (BAL). K K

Patient exclusion criteria were: lesions with benign radiological characteristics (e.g., hamartoma, mycetoma) K contraindication for surgical resection, FNA or BAL K concomitant diseases with a life expectancy under 6 months K pregnancy or lactation (women of child-bearing age underwent a pregnancy test before inclusion in the study) K participation in another clinical trial. K

All patients enrolled into the study gave written informed consent. Diagnosis was established based on the result of the pathological study of the specimen obtained by surgery, FNA or BAL. The following studies were performed on each patient within a maximum of 10 days. No pre-established fixed order was followed. Whole-body and chest SPECT scans with CT corecording (120 kV, 2.5 mA) 4 h after administration of 740 MBq of 99mTc-depreotide. VG Millenium Hawkeye equipment from General Electric was used. Images were obtained with low-energy, high-resolution parallel collimators. SPECT acquisition consisted of 60 views of 30 s each, at a 1801 angle, using a 64  64 matrix. K Whole-body PET with CT co-recording (140 kV, 60 mA) 60 min after administration of 370 MBq of FDG. Discovery ST equipment from General Electric was used. K

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Characterization of radiologically indeterminate lung lesions Ferran et al. 509

Table 1

Patients with malignancy

Histology Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Squamous cell carcinoma Adenocarcinoma Adenocarcinoma Adenocarcinoma Adenocarcinoma Adenocarcinoma NSCADC NSCADC NSCADC NSCADC Uterine sarcoma Carcinoid tumour

Size (cm)

Extrapulmonary dissemination

Target/background

Peak SUV (g
ml – 1)

3 2.5 2.5 4 3 4 3 1.5 2.2 1.5 3 2.2 3.8 2 2.7 6 3 2.2 3.1 1.6

No Yes Yes No Yes Yes No Yes** No No No No Yes Yes No No No No Yes** Yes

3.5 3.2 2 2.4 2.7 2.6 2.2 1.6 3.11 1 1.68 1.9 1.5 2.2 1.6 1 1.8 1.9. 1.8 1

15 10.5 17 13.5 18.5 20 7.4 9.6 19 10 5 10.6 7.4 17 7 14 6.7 10.6 6 9.6

DEP SPECT qualitative FDG PET qualitative reading reading Malign Malign Malign Malign Malign Malign Malign Malign Malign Benign* Malign Malign Malign Malign Malign Benign* Malign Malign Malign Benign*

Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign Malign

SUV, standardized uptake value; DEP SPECT, 99mTc-depreotide single photon emission computed tomography; FDG PET, [18F]fluorodeoxyglucose positron emission tomography; NSCADC, non-small cell adenocarcinoma. Values in bold type are the maximum and minimum in each column. * False negative; ** metastasis.

The study results were independently assessed by two nuclear medicine specialists blinded to patient data. Both qualitative and quantitative assessments were made. Qualitative assessment was made based on visual identification of the lesion and on lesion uptake level compared to background. Quantitative assessment was performed in the DEP SPECT study based on the lesion count number. Irregular regions of interest were manually defined on the lesion in several transaxial sections, of which the one showing the greatest activity was selected. The same area in the opposite lung was considered as background activity, and the lesion/background activity ratio was calculated for each patient. In the FDG study, the peak standardized uptake value (SUV) of the lesion, calculated automatically from the radiotracer concentration injected per body weight in the two-dimensional study, was considered. The statistical analysis (SPSS program) of qualitative parameters calculated the sensitivity, specificity and accuracy of both examinations, which were compared to each other. A Z test to compare proportions was used to test for statistically significant differences between both examinations. Quantitative parameters (lesion/background activity–peak SUV) were statistically assessed using receiver operating characteristic (ROC) curves and chi-squared tests (2  2 contingency table).

20 out of 29 patients, malignancy was confirmed by a pathological study of the specimen obtained: 14 after surgery, five after FNA, and one after BAL. The lesions found in the pathological study included nine adenocarcinomas, five squamous cell carcinomas, four undifferentiated, non-small cell adenocarcinomas, one metastasis from a uterine sarcoma, and one carcinoid tumour. Table 1 shows the relevant data for patients with malignancy. Figure 1 shows results from a patient with a metastasis from a uterine sarcoma. All these cases were adequately typified by an FDG PET scan. Three false negative results were obtained in the DEP SPECT scan: a 1.6 cm carcinoid tumour, a 6 cm undifferentiated, non-small cell adenocarcinoma, and a 1.5 cm moderately differentiated adenocarcinoma (Fig. 2). In nine out of 29 patients, the benign nature of the condition was confirmed by a pathological study of the specimen obtained: five after surgery and four after BAL. A histological study showed four non-specific inflammations, two cases with benign cellularity, two inflammatory pseudotumours, and one fibrosis. Table 2 shows the most relevant data for patients with benignancy. Figure 3 illustrates one of the cases of an inflammatory pseudotumour. A coincidental false positive result was found by both techniques. This was a 2 cm, non-specific, inflammatory process in the lung.

Results No patient discontinued the study due to hypersensitivity reactions to radiotracers administered in the trial. In

As regards size, among the 20 tumours diagnosed as malignant, five were greater than 3 cm, three were 3 cm in

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510 Nuclear Medicine Communications 2006, Vol 27 No 6

Fig. 1

(A) Anatomic

24

Physiologic

Fusion

24 Transaxial

36

Transaxial

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Coronal

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Sagittal

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26 Sagittal

A 66-year-old woman with previous endometrial tumour. (Histology: malignancy, uterine sarcoma.) Indeterminate 2 cm lung lesion in the upper right node located by computed tomography. (A) Single photon emission computed tomography using 99mTc depreotide (DEP SPECT): High radiotracer uptake in upper right lobe, suggesting malignancy. (B1) [18F]Fluorodeoxyglucose positron emission tomography (FDG PET): Pleuropulmonary hypermetabolic lesion located in the posterior segment of upper right lobe, suggesting malignancy. (B2) Metastatic hypermetabolic lesion in the right ilium with muscle infiltration.

and 12 were smaller than 3 cm. In the nine cases a benign diagnosis, four tumours were greater 3 cm, two were 3 cm in size, and three were smaller 3 cm.

nine cases mediastinal involvement was identified, while in the final two cases metastatic involvement (bone and adrenal) was detected. A total of 19 lesions were found in the FDG PET study.

As to extrapulmonary involvement, among the 20 cases in which a malignancy was diagnosed, a solitary lung lesion was confirmed in only 11 cases. In seven of the remaining

Eleven of these lesions were located in the chest according to the FDG PET study, and nine of them were also detected by DEP SPECT. Pathological confirmation

size, with than than

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Characterization of radiologically indeterminate lung lesions Ferran et al. 511

(B1)

(B2) S

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S ∗1.59

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was obtained for eight lesions, which were reported to be malignant. These patients were amenable to surgery as a definitive therapeutic procedure. The remaining eight lesions were located in the abdomen, where a DEP SPECT scan was not performed. No confirmation was achieved in the remaining cases, since invasive techniques would have been required. Use of CT co-recording in the DEP SPECT scan allowed for identification of three of the nine extrapulmonary lesions and for an accurate anatomical location of the other six lesions. In the quantitative assessment of the DEP SPECT scan, the mean lesion/background ratio of the total lesions was 1, with a confidence interval (95% CI) 1.5–2 and a range of 2.5. In the FDG PET study, the mean peak SUV was 8.5 g
ml – 1, with a confidence interval (95% CI) 6.1– 10.9 ng
ml – 1 and a range of 19.8 ng
ml – 1. A study of sensitivity and specificity analysing the ROC curves of the lesion/background ratio and peak SUV provided the following results. Considering the peak SUV (green curve) at a cut-off point of 3.5 (green arrow), sensitivity and specificity values of 95% and 89%, respectively, were obtained. Considering the lesion/

−74.22

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Fused sagittals

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P Fused transaxials

background ratio (red curve) at a cut-off point of 1.3 (red arrow), sensitivity and specificity values of 85% and 88%, respectively, were obtained. Figure 4 shows the ROC curves for both indices. The values obtained by the qualitative assessment for sensitivity, specificity and accuracy were 85%, 88% and 85%, respectively, for DEP SPECT, and 100%, 88% and 96% for FDG PET. No statistically significant differences were seen between the methods (P > 0.05) according to the Z test for comparison of means (SPSS).

Discussion In early diagnosis of lung cancer, only one third of pulmonary lesions can be typified based on radiological criteria, while invasive techniques are required in the remaining two thirds. The results obtained after studying 20 malignant lesions and nine benign lesions with both techniques show values for sensitivity, specificity and diagnostic accuracy similar to those reported in the literature. A recently published comparative study by Halley et al. [15] reported specificities of 88.9% for DEP SPECT and 94.4% for FDG PET, with no statistically significant differences. FDG PET identified two cases of adenocarcinoma which

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512 Nuclear Medicine Communications 2006, Vol 27 No 6

Fig. 2

Anatomic

Physiologic

Fusion

R

24

L

24 Transaxial

Transaxial

−274.66

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−274.66

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A 73-year-old woman with ischaemic cardiopathy. Indeterminate 1.5 cm lung lesion in lower right lobe located by CT. (Histology: malign, moderately differentiated adenocarcinoma.) (A) DEP SPECT: Low radiotracer uptake in low right lobe. Malignancy not suspected. (B) FDG PET: Pulmonary hypermetabolic lesion located in the upper part of the low right lobe. Malignancy suspected.

Table 2

Patients with benignancy

Histology Inflammation Inflammation Inflammation Inflammation Benign cells Benign cells Pseudotumour Pseudotumour Fibrosis

Size (cm)

Extrapulmonary dissemination

Target/background

Peak SUV (g
ml – 1)

DEP SPECT qualitative reading

FDG PET qualitative reading

1 1 2 3.2 3 1 3.2 1.8 3.8

No No No No No No No No No

1 1 2 1 1 1 1.8 1.2 1

1.2 1.2 17 1.2 2 1.4 1 1.2 1

Benign Benign Malign* Benign Benign Benign Benign Benign Benign

Benign Benign Malign* Benign Benign Benign Benign Benign Benign

Abbreviations are as given in the footnote to Table 1. Values in bold type are the maximum and minimum in each column. * False positive.

were false negatives for DEP SPECT (sizes, 1.8 cm and 1 cm), and DEP SPECT identified a carcinoid tumour that was a false negative for FDG PET. Two

coincidental false positives were also obtained with both techniques (inflammatory pseudotumour, sclerotic haemangioma).

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Characterization of radiologically indeterminate lung lesions Ferran et al. 513

Fig. 3

Anatomic

Physiologic

Fusion R

L

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25 25 Transaxial

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Transaxial

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37 Coronal

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A 69-year-old man without clinical features of interest. Indeterminate 3.2 cm lung lesion in the upper right node located by CT. (Histology: benign, inflammatory pseudotumour.) (A) DEP SPECT: Physiological radiotracer distribution. (B) FDG PET: Pulmonary lesion with anterior low FDG uptake in the posterior segment of upper right lobe. Malignancy not suspected.

The high proportion of cases (19/29) confirmed by a histological study, considered the most precise method, should be noted. Three false negative results were obtained with DEP SPECT. The lack of uptake by both undifferentiated non-small cell carcinomas and moderately differentiated adenocarcinomas is likely to be due to the probable null/poor expression of membrane SSTRs [10]. The carcinoid tumour was a small lesion (1.6 cm), though this would not be the limiting factor for its detection, as it was greater than 1 cm. Its adequate typing by FDG PET is also strange, because according to the literature such tumours usually show poor glycolytic metabolism [16]. As regards size, it should be noted that one of the tumours showing no uptake was 6 cm in size, and that between the two 1.5 cm carcinomas, the squamous cell carcinoma was detected whereas the small-cell carcinoma was not.

The false positive found by both techniques resulted from an inflammatory process, which is the main reason for false positive results reported in the literature and agrees with other studies [15]. SSTRs have been identified in the epithelioid cells of granulomatous conditions, either from an infectious or non-infectious origin [10], and show a high glycolytic metabolism [17]. While FDG PET had a superior sensitivity and diagnostic accuracy as compared to DEP SPECT in the sample studied (29 patients), no statistically significant differences (P > 0.05) were found between them according to the Z test for comparison of means. No significant size differences were noted between malignant and benign lesions. Mean sizes of malignant and benign lesions were 3.5 cm and 2.5 cm, respectively.

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514 Nuclear Medicine Communications 2006, Vol 27 No 6

It may therefore be concluded that while FDG PET has a greater sensitivity and diagnostic accuracy for assessing malignancy of indeterminate lung lesions, and for detection of extrapulmonary involvement, DEP SPECT represents a good diagnostic alternative for centres where PET is not available.

Fig. 4

1.0

Sensitivity

0.8

Use of hybrid equipment improves anatomical location of extrapulmonary lesions, enhancing staging of these tumours, which is of vital importance for assessing the therapeutic approach.

0.5

Acknowledgements We acknowledge the assistance of Dra. Masuet for statistical analysis.

0.3

References 0.0 0.0

1

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1-Specificity Sensitivity and specificity statistics (ROC curves) on the measured ratio for lesion/background (red line) and SUVmax (green line) (n = 29). Cutoff point for lesion/background is 1.3 (red arrow). Cut-off point for peak SUV is 3.5 (green arrow).

2 3 4

5 6

As regards quantitative assessment, when cut-off points of 3.5 for peak SUV in PET and 1.3 for lesion/background ratio in the DEP SPECT were established, the sensitivity and specificity results obtained did not differ from those seen in the qualitative study. However, these data should be viewed with caution, as they are subject to systematic errors (operator-dependent ROI area and lesion/background, PET facility-dependent calculation of peak SUV), and cannot be strictly extrapolated to other populations.

7

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11

Implementation of CT hybrid equipment results in an increased precision in anatomical location of scintigraphically detected molecular events. Correlation of a CT scan allowed for identification of thoracic extrapulmonary involvement in 33% of cases (3/9) and for accurate mediastinal/hilar location in 66% of them (6/9). These findings show the benefit of hybrid equipment. SPECT CT and PET CT co-recording enhances diagnostic accuracy.

12 13 14

15

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It should also be noted that the cost of the radiotracer for both examinations is high, but the availability of PET is still very limited in many countries.

17

Schrevens L, Lorent N, Dooms C, Vansteenkiste J. The role of PET scan in diagnosis, staging and management of non-small cell lung cancer. The Oncologist 2004; 9:633–642. Tan BB, Flaherty KR, Kazerooni EA, Lannettoni MD, American College of Chest Physicians. The solitary pulmonary nodule. Chest 2003; 123:89S–96S. Ost D, Fein AM, Feinsilver SH. Clinical practice. The solitary pulmonary nodule. N Engl J Med 2003; 348:2535–2542. Swenson SJ, Silverstein MD, Ilstrup DM, Schleck CD, Edell ES. The probability of malignancy in the solitary pulmonary nodules:application to small radiologically indeterminate nodules. Arch Intern Med 1997; 57: 849–855. Goldsmith SJ, Kostnbsakoglu AA, Somrov S, Palestro CJ. Radionuclide imaging of thoracic malignancies. Thoracic Surg Clin 2004; 14:95–112. O’Burme K, Carney D. Somatostatin and the lung. Lung Cancer 1993; 10:151–172. Blum J, Handmarker H, Rinne N. The utility of somatostatin-type receptor binding peptide radiopharmaceutical (P829) in the evaluation of solitary pulmonary nodules. Chest 1999; 115:224–232. Vallabhajosula S, Moyer B, Lister-James J. Preclinical evaluation of technetium-99m-labeled somatostatin receptor-binding peptides. J Nucl Med 1996; 37:1016–1022. Blum J, Handmaker H, Lister-James J, Rinne N. A multicenter trial with a somatostatin analog 99mTc depreotide in the evaluation of solitary pulmonary nodules. Chest 2000; 117:1232–1238. Lowe VJ, Fletcher JW, Gobar L, Lawson M, Kirchner P, Valk P, et al. Prospective investigation of positron emission tomography in lung nodules. J Clin Oncol 1998; 16:1075–1084. Kubota K, Matsuzava T, Fijiwara T, Ito M, Hutazawa J, Ishiwata K, et al. Differential diagnosis of lung tumor with positron emission tomography: a prospective study. J Nucl Med 1990; 31:1927–1932. Gupta NC, Maloof J, Gunel E. Probability of malignancy in solitary pulmonary nodules using fluorine 18-FDG and PET. J Nucl Med 1996; 37:943–948. Bury T, Rigo P. Contribution of positron emision tomography for the management of lung cancer. Rev Pneumol Clin 2000; 56:1506–1511. Gould MK, Maclean CC, Kuschner WG, Rydizak CE, Owens DK. Accuracy of positron emission tomography for diagnosis of pulmonary nodules and mass lesions: a meta-analysis. JAMA 2001; 285:914–924. Halley A, Hugentobler A, Icard P, Porret E, Sobrio F, Lerochais JP, et al. Efficiency of 18F-FDG an 99mTc-depreotide SPECT in the diagnosis of malignancy of solitary pulmonary nodules. Eur J Nucl Med March 2005; Issue on line. ISSN 1619-7089. Roberts PF, Follette DM, von Haag D, Park JA, Valk PE, Pounds TR, et al. Factors associated with false positive staging of lung cancer by positron emission tomography. Ann Thorac Surg 2000; 70:1154–1159. Matthies A, Hickeson M, Cuchiara A, Alvavi A. Dual time point 18F-FDG for the evaluation of pulmonary nodules. J Nucl Med 2002; 43:273–278.

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Original article

Correction of an image size difference between positron emission tomography (PET) and computed tomography (CT) improves image fusion of dedicated PET and CT Wouter V. Vogela, Jorn A. van Dalena, Dominic A.X. Schinaglb, Johannes H.A.M. Kaandersb, HenkJan Huismanc, Frans H.M. Corstensa and Wim J.G. Oyena Aim Clinical work in software positron emission tomography/computed tomography (PET/CT) image fusion has raised suspicion that the image sizes of PET and CT differ slightly from each other, thus rendering the images suboptimal for image fusion. The aim of this study was to evaluate the extent of the relative image size difference between PET and CT and the impact of the correction of this difference on the accuracy of image fusion. Methods The difference in real image size between PET and CT was evaluated using a phantom study. Subsequently, 13 patients with cancer in the head/neck area underwent both CT and [18F]fluorodeoxyglucose PET in a custom-made mask for external beam radiotherapy, with multimodality markers for positional reference. The image size of PET relative to CT was determined by evaluating the distances between the markers in multiple directions in both scans. Rigid-body image fusion was performed using the markers as landmarks, with and without correction of the calculated image size difference. Results Phantom studies confirmed a difference in real image size between PET and CT, caused by an absolute error in PET image size calibration. The clinical scans demonstrated an average relative difference in image size of 2.0% in the transverse plane and 0.8% along the longitudinal axis, the PET images being

Introduction Image fusion of positron emission tomography (PET) and computed tomography (CT) can improve the diagnostic value and diagnostic accuracy in oncological imaging of the head and neck area [1–3]. Image fusion may also be applied to incorporate functional information in external beam radiation treatment [4,5]. When performing image fusion, a high accuracy in anatomical registration of the images is required, because incorrect registration may induce diagnostic errors, such as erroneous localization or characterization of the lesions [6]. In particular, when using image fusion for the definition of target volumes in intensity-modulated radiation therapy (IMRT), the required accuracy is high as the error in dose delivery is in the range of only 2–3 mm [7]. Errors in image registration may influence the outcome of therapy and

significantly smaller. Image fusion using original images demonstrated an average registration error of 2.7 mm. This error was decreased to 1.4 mm after size correction of the PET images, a significant improvement of 48% (P < 0.001). Conclusions A significant deviation in PET image size may occur, either as a real image size deviation or as a relative difference from CT. Although possibly not clinically relevant in normal diagnostic procedures, correction of such a difference benefits image fusion accuracy. Therefore, it is advisable to calibrate the PET image size relative to CT before performing high-accuracy rigid-body image c 2006 Lippincott fusion. Nucl Med Commun 27:515–519 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:515–519 Keywords: image fusion, positron emission tomography (PET), positron emission tomography /computed tomography (PET/CT) Departments of aNuclear Medicine, bRadiotherapy and cRadiology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands. Correspondence to Wouter V. Vogel MD, Radboud University Nijmegen Medical Centre, Department of Nuclear Medicine (565), Postbox 9101, 6500 HB Nijmegen, The Netherlands. Tel: + 31-24-3614048; fax: + 31-24-3618942; e-mail: [email protected] Received 4 August 2005 Accepted 7 March 2006

the level of complications of external beam radiation therapy. For software image fusion of dedicated PET and CT, an accuracy of better than 2 mm has been demonstrated using phantoms [8]. The accuracy that can be achieved in patients will probably be lower as a result of complicating factors, such as small positioning errors, motion artefacts, the time interval between scans and limited comparability between scans due to visualization of different structures and processes on PET and CT. Furthermore, differences may exist in image size. In this article, real image size is defined as the discrepancy between the measured size of an object on an image and the true size of that object. Furthermore, relative differences in image size may occur between scanning modalities. The image

c 2006 Lippincott Williams & Wilkins 0143-3636

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516 Nuclear Medicine Communications 2006, Vol 27 No 6

Fig. 1

phantom was scanned in three directions (i.e. x, y and z) in both PET and CT. The distances between the centres of all marker pairs were measured in three directions in both imaging modalities. Differences in the distances between corresponding marker pairs on PET and CT were evaluated throughout the field of view of the scanners to determine the linearity of deviations. The inter-operator variation of the manual localization of the centre of the markers was evaluated by the analysis of 12 markers in a separate session by two operators. Clinical experiment

Software image fusion of positron emission tomography (PET) and computed tomography (CT). A slice through the head is shown at the level of two multimodality fiducial markers positioned in front of the ears. (a) Original images: the markers on PET are closer to each other than on CT, and hence the PET image is somewhat smaller than the CT image. (b) After correction for image size differences: The markers are centred correctly.

size is often not considered in rigid-body image fusion as, in general, the scanner image sizes are fixed and validated for both PET and CT.

Thirteen patients with newly diagnosed malignancy in the head and neck area were included. All patients had squamous cell carcinoma of the oral cavity or larynx, and were candidates for external beam therapy. None of the patients had a history of diabetes mellitus, and fasting glucose levels were within the normal range. In addition to standard planning CT, an [18F]FDG PET scan was performed to provide biological parameters for optimized target volume definition. Both CT and [18F]FDG PET were acquired in a Fig. 2

During software PET/CT image fusion with multimodality markers for IMRT planning of the head and neck area, we observed a small systematic scaling difference between CT and PET images. Patients were slightly smaller on PET images than on CT images. An example is shown in Fig. 1(a). It was suspected that the image size of the CT scanner and/or PET scanner was inaccurate. Attempts were made to correct these errors with patientspecific manual or automatic scaling procedures, but the results were variable and depended on other factors, such as small positioning differences and deformations. It was hypothesized that correction with an objectively determined systematic scaling factor would provide a better and more elegant solution. The purposes of this study were to determine the extent of image size differences between PET and CT, to evaluate the impact of this problem on image fusion accuracy and, if needed, to determine a systematic scaling correction factor.

Materials and methods Phantom experiment

A linear phantom, 50 cm in length, with 11 multimodality markers positioned at 5 cm intervals, was used to estimate the real image sizes of PET and CT, and to detect the linearity of deviations. Each marker consisted of two glass capillaries positioned under a 901 angle, filled with either iodine-containing X-ray contrast solution or diluted [18F]fluorodeoxyglucose ([18F]FDG) solution. The visualization of a marker is demonstrated in Fig. 2. The

Example of the multimodality markers used for landmark registration and image fusion accuracy evaluation. (a) Markers as placed on the mask. (b) Markers as seen on computed tomography (CT) filled with iodine contrast. (c) Markers as seen on positron emission tomography (PET) filled with [18F]fluorodeoxyglucose solution. The centre of the markers could be determined with an inter-operator variability well below 1 mm on both CT and PET images.

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Correction of PET image size improves PET/CT fusion Vogel et al. 517

custom-moulded mask to provide identical positioning. Four multimodality markers were attached to each mask at corner positions outside the target field (Fig. 2). Image acquisition

CT scans were acquired using a multislice spiral CT scanner (Marconi AcQsim, Marconi Corporation, Cleveland, Ohio, USA). The scanning parameters comprised a scan range from the skull base to the lung top, intravenous contrast in the arterial phase, 100 mA, 130 kV. The pixel size of the images was 0.938 mm2 in the transaxial plane. The voxel size in the axial direction was defined by a slice thickness of 3 mm. PET scans were acquired using a full-ring dedicated PET scanner (Siemens ECAT Exact 47, Siemens/CTI, Knoxville, Tennessee, USA). An activity of 250 MBq of [18F]FDG was injected intravenously. The scans were acquired 1 h post-injection, using three-dimensional emission for 6 min per bed position, and employing attenuation correction based on two-dimensional germanium-68 transmission images for 2 min per bed position. All PET scans were reconstructed using an iterative twodimensional ordered subset expectation maximization (OSEM) algorithm [9] using four iterations, 16 subsets and a three-dimensional Gaussian filter of 5 mm. A zoom factor of 1.5 was applied to generate voxels with a size of 3.432 mm in all directions. Evaluation of image size difference

The difference in real image size between PET and CT was evaluated by measuring the distances between the centres of multiple markers in the transverse and axial planes for both imaging modalities. For each patient, four markers were placed in a rectangular configuration, providing 26 marker pairs in the transverse plane and 26 pairs in the axial direction. Three markers in three separate patients were excluded from evaluation because of poor visibility. Thus, 24 evaluable marker pairs in the transverse plane and 23 pairs in the axial direction were used for the analysis. The relative difference in pixel size between PET and CT was calculated using the formula: (DistancePET – DistanceCT)/DistanceCT  100%. Impact on image fusion

Image fusion was performed twice, with the original PET images and with PET images that had been corrected for the image size difference relative to CT by adaptation of the pixel size in the DICOM header of the source files. Rigid-body landmark-based registration of PET images to CT images was performed with in-house-developed software, based on the visualization toolkit VTK [10]. First, the centre of all markers was identified manually. Subsequently, rigid-body registration, that is based on three translation and three rotation parameters, of PET

to CT images was performed automatically by minimizing the sum of the square distances between corresponding marker centres [11]. Scaling of the images in the image registration software was not necessary, because image size corrections had already been performed in the DICOM source files. The accuracy of the image registration procedure was evaluated mathematically by determining the remaining positional difference between corresponding markers on PET and CT after image registration. Statistical analysis of the difference in image registration accuracy between the two image sets was performed using a two-sided paired t-test. The level of significance was set at 0.05.

Results The inter-operator variability in the determination of the centre of the markers was well below 1 mm in both types of scans. For CT images, the average difference between the two operators was 0.54 mm [standard deviation (SD), 0.28 mm]. For PET images, the average difference was 0.42 mm (SD, 0.19 mm). Image size difference

In the phantom experiment, in the horizontal direction (xaxis), the measured distance between the outermost markers on PET was 490.4 mm. On CT images, the measured distance was 499.2 mm, a relative difference from PET of 1.8%. In the vertical direction (y-axis), the respective distances were 499.0 and 490.1 mm, a relative difference of 1.8%. In the longitudinal direction, the distances were 500.5 and 497.2 mm, a relative difference of 0.7%. For both PETand CT, the markers were distributed evenly along the phantom in all directions. The difference in distance between marker pairs on PET and CT was detected throughout the field of view, indicating a linear image size difference. In the series of clinical scans, in the transverse plane, a significant average relative difference in distance between markers on PET and CT of 2.0% was observed (range, 0.2–3.7%; SD, 0.76%), the patient being smaller on the [18F]FDG PET images than on the CT images in all cases. In the axial direction, an average difference of 0.8% (range, – 0.6 to 2.5%; SD, 0.65%) was found, the patient being smaller on the [18F]FDG PET images than on the CT images in the majority of cases. To compensate for the detected differences in the clinical series, the pixel size of the PET images was increased by 2.0% to 3.501 mm in the transverse plane, and increased by 0.8% to 3.459 mm along the longitudinal axis, by adaptation of pixel size values in the DICOM file header. Repeated evaluation using the corrected PET images showed an image size difference of 0.2% (range, – 1.4 to 1.8%; SD, 0.69%) in the transverse plane, and a

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518 Nuclear Medicine Communications 2006, Vol 27 No 6

difference of 0.0% (range, – 1.5 to 1.7%; SD, 0.68%) in the axial direction. Thus, a significant relative difference in image size between PET and CT could no longer be demonstrated. Accuracy of image fusion

Image fusion using the original uncorrected [18F]FDG PET images demonstrated an average mathematical registration error of 2.7 mm (range, 0.8–5.5 mm) at the position of the markers. When image fusion was performed using PET images corrected for relative image size differences, the error was 1.4 mm (range, 0.3–3.8 mm). This represented a significant decrease in the error in image fusion at the location of the markers of 48% (P < 0.001). Figure 1(b) shows a fused image after pixel size correction.

Discussion The suspected relative difference in real image size between PET and CT was confirmed using both phantom studies and clinical scans. In the clinical scans, a series of separate measurements over multiple patients demonstrated the deviation with statistical significance. The manual identification of the centre of the markers was excluded as a possible source of error in the analysis procedure, because the interoperator variability was negligible. The results of the phantom measurements and the clinical scans were concordant (the size differences in the phantom measurements were well within the standard deviations of the clinical series). The deviation was shown to be linear, and thus the results from the phantom study and the clinical data were theoretically exchangeable. The image size difference derived from the clinical series was considered to be the most accurate, because statistical analysis of a series of measurements is less prone to bias in the manual evaluation of markers than a single measurement in the phantom experiment. Therefore, the image size difference derived from the clinical series was applied as the correction factor. This approach was validated by repeated measurements after correction, which no longer demonstrated significant image size differences. The observed difference in image size between PET and CT was larger in the transverse plane than in the axial direction (2.0% versus 0.8%). The exact cause of this discrepancy remains unclear. There is no apparent reason why image size calibration of PET images would be more difficult in one direction than in another. The relative difference in real image size between PET and CT was caused by an absolute error in calibration of the PET image size. With a pixel size and effective resolution (full-width at half-maximum) in the range of 5 mm in PET imaging, accurate image size calibration

may be difficult. For example, to detect a deviation of 1%, the difference to be found in a marker distance over a length of 20 cm is only 2 mm, which is well below the resolution. Therefore, the procedure to determine the image size of a PET scanner will always be less accurate relative to a CT scanner. Furthermore, in IMRT planning, CT must be the gold standard by default because planning of the radiation fields depends on the electron density information derived from CT. For these reasons, we have adapted the PET images to match CT, regardless of the possibility of a hypothetical small remaining error in CT real image size. Correction of the image size differences alone will not result in perfect image fusion. Other causes of inaccuracies remain, such as slight deformation of the mask between scans (as a result of small patient positioning differences) and patient motion during scanning (for example, swallowing). These factors, in combination with the sampling errors of the marker locations, contribute to the detected remaining error in rigid-body landmarkbased image registration. It can be argued that image size correction should be applied to all PET scanning, but the relevance of an error of this small magnitude in normal diagnostic imaging is probably negligible. Image size corrections need to be advocated only when a high accuracy is required, such as in image fusion for IMRT planning in the head and neck area. Several approaches to rigid-body image registration are available. Examples include manual procedures, automatic methods based on mutual information or the iterative closest point algorithm, and landmark-based registration. We used the latter method for our accuracy evaluation, as landmark registration is a robust and accurate technique when reliable landmarks are available [11]. It seems obvious that other rigid-body registration techniques will benefit similarly from the correction of image size differences. Theoretically, when non-rigid transformations are used, there is no need for additional image size corrections. Multiple approaches may be available to correct the standard image size of PET images. Some PET scanners may allow easy adaptation of the image size on the machine itself, most likely as a parameter in the reconstruction algorithm. Other options include adaptation of the DICOM file that is transported to the fusion software, or adaptation in the fusion software itself. Not all available approaches may support separate adaptation for the transverse and axial directions, as was needed in our specific situation. Otherwise, there are no rational arguments to prefer one approach over another.

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Correction of PET image size improves PET/CT fusion Vogel et al. 519

As an alternative solution to systematic correction of differences in image size, patient-specific scaling may be advocated as it is relatively easy to perform. Image fusion software generally supports scaling, either manual or automatic. Scaling is even considered as a routine procedure in image fusion with magnetic resonance imaging (MRI), which suffers from spatial distortions and size deviations because of magnetic field inhomogeneities. However, when spatial inhomogeneities are absent, we consider this approach to be suboptimal, as the applied size corrections will be influenced by incidental variations, such as positioning differences. The application of ad hoc scaling factors seems to be a less rational procedure, especially when the exact correction values can be derived by relatively simple measurements as discussed in this paper. It seems unlikely that the correction parameters presented in this article can be transferred to other PET systems in general. Variations may occur between systems, especially when using scanners from different manufacturers. Therefore, the extent of image size differences should be assessed locally. Hybrid PET/CT systems may also suffer from relative image size differences between PET and CT, as all currently available hybrid systems are dedicated PET and CT scanners placed in-line. Therefore, the adjustment of pixel size may result in a similar benefit in image fusion accuracy when using hybrid PET/CT scanning.

Conclusion We have demonstrated that a small deviation in PET real image size may occur, as well as a significant difference in

PET image size relative to CT. Although a small deviation in PET image size is not clinically relevant in normal diagnostic procedures, correction of such a difference proves to be beneficial with regard to the accuracy of rigid-body software image fusion. Therefore, it is advisable to re-evaluate PET scanner image size relative to CT images before using high-accuracy rigid-body image fusion with CT.

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Schoder H, Yeung HW, Gonen M, Kraus D, Larson SM. Head and neck cancer: clinical usefulness and accuracy of PET/CT image fusion. Radiology 2004; 231:65–72. 2 Schwartz DL, Ford E, Rajendran J, Yueh B, Coltrera MD, Virgin J, et al. FDGPET/CT imaging for preradiotherapy staging of head-and-neck squamous cell carcinoma. Int J Radiat Oncol Biol Phys 2005; 61:129–136. 3 Goerres GW, von Schulthess GK, Steinert HC. Why most PET of lung and head-and-neck cancer will be PET/CT. J Nucl Med 2004; 45 (Suppl 1): 66S–71S. 4 Scarfone C, Lavely WC, Cmelak AJ, Delbeke D, Martin WH, Billheimer D, et al. Prospective feasibility trial of radiotherapy target definition for head and neck cancer using 3-dimensional PET and CT imaging. J Nucl Med 2004; 45:543–552. 5 Solberg TD, Agazaryan N, Goss BW, Dahlbom M, Lee SP. A feasibility study of 18F-fluorodeoxyglucose positron emission tomography targeting and simultaneous integrated boost for intensity-modulated radiosurgery and radiotherapy. J Neurosurg 2004; 101 (Suppl 3):381–389. 6 Vogel WV, Oyen WJ, Barentsz JO, Kaanders JH, Corstens FH. PET/CT: panacea, redundancy, or something in between? J Nucl Med 2004; 45 (Suppl 1):15S–24S. 7 Low DA, Mutic S, Dempsey JF, Gerber RL, Bosch WR, Perez CA, et al. Quantitative dosimetric verification of an IMRT planning and delivery system. Radiother Oncol 1998; 49:305–316. 8 Lavely WC, Scarfone C, Cevikalp H, Li R, Byrne DW, Cmelak AJ, et al. Phantom validation of coregistration of PET and CT for image-guided radiotherapy. Med Phys 2004; 31:1083–1092. 9 Hudson HM, Larkin RS. Accelerated image reconstruction using ordered subsets of projection data. IEEE Trans Med Imag 1994; 13:601–609. 10 Schroeder W. The visualization toolkit: an object-oriented approach to 3D graphics. 3rd ed. New York: Kitware Inc.; 2003. 11 Maintz JB, Viergever MA. A survey of medical image registration. Med Image Anal 1998; 2:1–36.

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Original article

Anatomical accuracy of hybrid SPECT/spiral CT in the lower spine Anton No¨mayra, Wolfgang Ro¨mera, Daniel Strobela, Werner Bautzb and Torsten Kuwerta Aim The anatomical accuracy of hardware-based registration of skeletal single photon emission computed tomography (SPECT) and X-ray computerized tomography (CT) has as yet not been studied. The aim of this study was to evaluate this variable in the lower spine for a newly introduced hybrid SPECT/spiral-CT camera. Methods In 22 patients referred for degenerative joint disease or tumours, whole-body bone scintigraphy including hybrid SPECT/spiral CT of the lower spine was performed. Subsequent analyses were performed on these pairs of images as well as on data sets obtained after using a rigid automated fusion procedure in addition. Two observers independently measured the distances between the visually determined centres of gravity of the CT and SPECT representation of the fourth and fifth lumbar vertebral body in the X-, Y- and Z-directions (X-, Y- and Z-distances). Results The distances determined by the two observers for the two vertebral bodies correlated significantly and were averaged for further analysis. For hybrid SPECT/spiral CT without consecutive automated registration, the mean X-, Y- and Z-distances were 1.6 ± 1.9 mm, 1.7 ± 1.3 mm and 0.9 ± 0.5 mm, respectively. Additional automated registration lowered these values to 1.2 ± 0.9 mm, 1.1 ± 0.7 mm and 0.8 ± 0.4 mm, respectively. The difference

Introduction Bone scintigraphy has been one of the most frequent nuclear medical examinations for almost 30 years. Its applications include the search for osseous metastases and the assessment of joint disease. Although very sensitive for detecting bone abnormalities, bone scintigraphy suffers from a rather low specificity [1]. In particular, the distinction between metastases to the spine and spondylarthrosis is difficult, even when single photon emission computed tomography (SPECT) is added to the diagnostic work-up. Five years ago, a hybrid system combining a dual-headed SPECT camera with a low-dose computerized tomographical scanner (CT) was introduced [2,3]. Several studies have proven its clinical value [2,4–16]; in particular, Horger et al. [17,18] recently demonstrated that a

for the Y-distance proved statistically significant (P < 0.05). Additional automated registration significantly reduced the number of subjects in whom at least one of the distances determined was greater than the SPECT pixel size of 4.6 mm from 14% (n = 3) to 0% (P < 0.05). Conclusion Hardware-based fusion between skeletal SPECT and CT offers a nearly perfect data match in the lower spine. The additional use of a tool for automated rigid registration has the potential to reduce the error of alignment even further and may be useful in patients with reduced compliance leading to movements between the c 2006 two examinations. Nucl Med Commun 27:521–528 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:521–528 Keywords: SPECT, CT, SPECT/CT, image fusion, registration, bone scintigraphy a

Clinic of Nuclear Medicine and bInstitute of Diagnostic Radiology, University of Erlangen/Nu¨rnberg, Germany. Correspondence to Dr Anton No¨mayr, Clinic of Nuclear Medicine, University of Erlangen/Nu¨rnberg, Krankenhausstr. 12, D-91054 Erlangen, Germany. Tel: + 0049 9131 853 3411; fax: + 0049 9131 853 9262; e-mail: [email protected] Received 23 November 2005 Accepted 9 March 2006

combination of the two modalities improves diagnostic specificity with regard to differentiating benign from malignant lesions to the bone. Nevertheless, a shortcoming of these systems is the limited resolution of their CT component, potentially obviating the detection of small osseous lesions. Consequently, in 2005, a spiral-CT scanner offering high quality diagnostic performance and a SPECT camera were integrated within the same gantry. Evidence reporting the clinical performance of SPECT/spiral CT is still scarce although preliminary clinical data are very promising [19,20]. SPECT/CT systems allow the performance of a CT directly following a SPECT examination, thus limiting misalignment between the two data sets caused by

c 2006 Lippincott Williams & Wilkins 0143-3636

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differences in patient positioning. However, observations from hybrid systems combining positron emission tomography (PET) and CT have shown that minor anatomical inaccuracies of the data match may still occur. The majority of these are caused by respiratory movements and involve the thorax [21]. Mismatches have been reported in the order of 3–4 mm [7,22]. Obviously and, in particular, in critically ill patients, minor body movements over periods of examination lasting at least 20 min cannot be reliably prevented. The aim of this study was to evaluate the anatomical accuracy of hardware-based fusion in the lower spine for a newly introduced hybrid SPECT/spiral-CT camera. Furthermore, we sought to investigate whether additional automated rigid registration could still improve the quality of the match.

Patients, materials and methods Patients

The data base consisted of 22 patients examined consecutively by SPECT/spiral CT involving the lower lumbar spine between March and July 2005. The group included 14 women and eight men, mean age was 66 years. The diagnoses were breast cancer, lung cancer, and spondylosis/spondylarthrosis in 11, two, and nine cases, respectively. SPECT/spiral-CT data acquisition and reconstruction

Three hours after intravenous injection of 7– 10 MBq
kg – 1 body weight 99mTc-diphosphonate planar whole-body scintigraphies were performed using a dualheaded gamma camera (e-cam; Siemens Medical Solutions, Erlangen, Germany). SPECT/spiral CT was performed 3–4 h after tracer using a dual-headed gamma camera in conjunction with a two-slice spiral CT installed within the same gantry (Symbia T2, Siemens Medical Solutions, Erlangen, Germany). The specifications of the gamma-camera heads are those of the dual-head gamma camera that has now been available commercially for several years (e.cam; Siemens Medical Solutions, Erlangen, Germany). The CT component of this system is a dual-detector helical CT scanner (SOMATOM Emotion duo; Siemens Medical Solutions, Erlangen, Germany) with a minimal gantry rotation time of 800 ms.

resolution was 9.9% full width at half maximum (FWHM). The camera heads were equipped with highresolution low-energy parallel-hole collimators. Raw SPECT data were reconstructed into transaxial slices using the e.soft reconstruction software. As published previously, the hounsfield unit (H) values derived from the CT were used to correct the SPECT images for attenuation [20]. Reconstruction was performed iteratively by using the ordered subsets expectation maximization (OSEM 3-D) technique with eight iterations and 16 subsets. Images were smoothed with a threedimensional (3-D) spatial Gaussian filter (FWHM 10 mm). The scan parameters for the CT were: 130 kV; 20 mAs by using real-time tube current modulation (CARE Dose4D, Siemens Medical Solutions, Erlangen, Germany); rotation time 0.8 s; collimation 2  2.5 mm. Image reconstruction resulted in images with a slice thickness of 3 mm using a 1.5 mm reconstruction increment. A bone reconstruction algorithm was used to minimize artefacts from metallic prostheses (window centre, 800 H; window width, 3000 H). Both the SPECT and the CT scans were performed consecutively with the patient lying stable in a supine position with elevated arms. The SPECT was performed with a field of view from the ischiadic tuber to Th12/L1. The field of view of the CT was limited to the pelvis including L4/5 in all cases. For quality control of the SPECT/CT, the physical alignment between SPECT and CT was tested by phantom measurements, allowing the calibration between the fields of view of the two scanners. For this purpose, the Siemens NCO/MHR/NEMA phantom was employed. This phantom uses point sources filled with 99m Tc and iodinated X-ray contrast medium. Using dedicated software, the misalignment of these point sources was measured for three degrees of rotation (X, Y, Z) and three degrees of translation (X, Y, Z). The average physical misalignment ranged between 1.2 (Z) and 3.2 (Y) mm. This hardware-based error was then corrected by the system software, leading to exact registration in control phantom measurements. Data analysis

The details of data acquisition and reconstruction were the following. For SPECT, counts from the 20% energy windows at 140 keV were acquired into a 128 128 matrix leading to a pixel size of 4.6  4.6 mm. A total of 64 frames, each of a duration of 30 s, were acquired over 3601. The field of view of the emission tomographic detectors was 53.3  38.7 cm, the crystal sizes where 59.1  44.5 cm with a thickness of 9.5 mm corresponding to 3/8 inch. The intrinsic spatial resolution was 3.8 mm in the centre of the field of view, the intrinsic energy

CT and SPECT data were transferred in DICOM format to a viewing station (Syngo, Siemens Medical Solutions). The slice thickness of the CT images was 3 mm, the increment 1.5 mm. The CT scans were displayed at bone window settings (level, 500 H; width, 2000 H). The colour scale of the SPECT scans could be manipulated by the reader. Subsequently, the evaluation was performed by two physicians experienced both in radiology and nuclear medicine (A.N. and D.S.), using a commercially available 3-D volume fusion tool (Syngo advanced

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Accuracy of SPECT/spiral CT in the lower spine No¨mayr et al. 523

Fusion VC20H; Siemens Medical Solutions, Erlangen, Germany). This tool allows sub-voxel 3-D rigid-body transformations with six degrees of freedom, three translations and three rotations. Reading was done in 3-D mode with the ability to change the ratio of CT and SPECT in the fused images on %-scales. The observers were able to view the scan data simultaneously in three main planes, i.e., axially, coronally and sagittally. The original SPECT/spiral-CT data sets were subjected to additional automated software-based rigid fusion. For this purpose, the automated fusion tool included in the Syngo advanced Fusion VC20H software package (Siemens, Germany) was used. It employs an algorithm based on normalized mutual information. As we wanted to test the fusion quality of the hybrid SPECT/CT as it is used in clinical praxis, the registration is done between attenuation-corrected (AC) SPECT data and CT. The following measurements were carried out for the original as well as for the additional post-hoc registered data sets. The two observers measured the distances between the centres of the representation of the vertebral bodies L4 and L5 on SPECT and CT in the X-, Y- and Z-directions with reference to a coordinate system centred in the CT representation of the vertebral body. The centre of gravity of the vertebral body was visually estimated by using a manually created ellipsoid, circle or rectangle to fit the contours of the CT and SPECT representation of the vertebral body as illustrated in Fig. 1. The distances between the two centres of gravity were measured in the X-, Y- and Z-directions on axial and sagittal planes (Fig. 1, green lines) using the Syngo advanced 3-D fusion tool.

The values for L4 and L5 were averaged. These mean values are in the following referred to as X-, Y- and Z-distances, respectively. To evaluate the intraobserver reproducibility of this method, the distances between the SPECT and CT lesion were determined in five patients, 20 times for one vertebral body, yielding a mean coefficient of variation (mean/standard deviation  100) of 5.8% in the X-direction, 6.1% in the Y-direction and 5.1% in the Z-direction. This indicates a high intraobserver reproducibility of the measurement of the X-, Y- and Z-distances. The non-parametric Mann–Whitney test disclosed no significant differences between the measurement results of the two observers (P > 0.05; data not shown) so that the interobserver reproducibility of this method seemed reasonably high. To eliminate subjective variations in the data sets even further, the arithmetic means between the distances determined by both observers were used for further analysis. The significance of differences in distances was tested using the non-parametric Wilcoxon test. Differences in distributions were assessed using the chi-squared test. The level of significance was set at P < 0.05. All data are given as the mean value ± standard deviation unless otherwise specified. Misalignment between SPECT and CT may lead to errors in the quantification and visualization of DPD uptake;

Fig. 1

Hardware-based registration between SPECT and CT in a representative patient with significant misalignment in X- and Y-direction (axial and sagittal plane). In this case, the representation of the vertebral body in CT and SPECT is approximated by an ellipsoid on the axial plane and by a rectangle on the sagittal plane. The green-circled crosses highlight the visually determined centres of gravity (COG) of the representation of the fifth lumbar vertebral body in CT and SPECT. The green lines mark the distances between the COG of CT and SPECT in the X-, Y- and Z-directions. In this case, there is no misregistration in the Z-direction. For further explanations, please see text.

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Fig. 2

,,

Fusion

ROI

AC

Non-AC (,,Spectrum ) ,,

AC (,,Edges )

ROI

Non-AC

Hardware-based registration between SPECT and CT in a representative patient (Fig. 1) with significant misalignment in the X- and Y-directions. First row of images. Left: attenuation-corrected (AC) SPECT, displayed with ‘edges’ scale; right: non-attenuation-corrected (non-AC) SPECT, displayed with ‘spectrum’ scale; middle: exact fusion with no significant difference between AC and non-AC with regard to the definition of the boundary between soft tissue and bone (arrows). Second row of images. Hardware-based registration between SPECT (left: AC; right: non-AC) and CT of the same patient with significant misalignment in the X- and Y-directions. Measurement of the uptake values in AC SPECT and non-AC SPECT was performed by placing regions of interest (ROIs) on the misaligned area and in the centre of the vertebral body. For further explanations, please see text.

this potentially affects the definition of the boundary between bone and soft tissue and thus also the determination of the centre of gravity on attenuationcorrected scans. Therefore, in patients exhibiting a misalignment of greater than 4 mm (n = 3), we analysed attenuation-corrected and uncorrected SPECT tomograms to evaluate whether the misalignment really affected the definition of the bone/soft tissue boundary. For this purpose, an edge-finding algorithm was used to facilitate visual evaluation. Furthermore, employing regions of interest (ROIs) placed on the misaligned margin and on the centre of the vertebral body, we measured 99mTc-DPD uptake in the attenuation-corrected and uncorrected SPECT images in cases exhibiting a CT/SPECT misalignment exceeding 4 mm (Fig. 2). Relative uptake values were expressed as the ratio between the misaligned margin and centre of the vertebral body. With a further circular ROI, 99mTc-DPD uptake was also determined in the soft tissue just anterior to the vertebral body (background, BG; not shown). Relative uptake in the vertebral body was then calculated as the quotient between the centre of the vertebral body and background.

Results Figures 1 and 3 give representative patient examples. For the data sets directly gained from hardware-based fusion only, the X-, Y- and Z-distances were 1.6 ± 1.9 mm, 1.7 ± 1.3 mm and 0.9 ± 0.5 mm, respectively (Fig. 4). The corresponding values for data sets corrected by additional automated fusion were 1.2 ± 0.9 mm, 1.1 ± 0.7 mm and 0.8 ± 0.4 mm, respectively (Fig. 4). Only the difference in the Y-direction was statistically significant (P < 0.05). For the original data sets, 9% of the X-distances, 4% of the Y-distances and 0% of the Z-distances were greater than the pixel width of 4.6 mm (Fig. 5). Additional automated registration significantly reduced the number of subjects in whom at least one of the distances determined was greater than the SPECT pixel size of 4.6 mm from 14% (n = 3) to 0% (P < 0.05) (Fig. 5). In the three patients with a CT/SPECT misalignment exceeding 4 mm, the mean difference in relative 99mTcDPD uptake of the misaligned region between the attenuation-corrected and uncorrected SPECT images

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Accuracy of SPECT/spiral CT in the lower spine No¨mayr et al. 525

Fig. 3

A screen shot of hardware-based registration between SPECT and CT in a representative patient. The cross is centred on the visually determined centre of gravity of the fifth lumbar vertebral body. No misalignment is detectable.

disclose differences with regard to the definition of the bone/soft tissue boundary (Fig. 2).

25 20

Discussion

15 10

X

Y

AAF

SPECT/CT

AAF

AAF

0

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Fig. 4

Z

Misregistration in the X-, Y- and Z-directions. Box plots give the mean values for the distances measured, error bars the corresponding standard deviations. AAF indicates the results obtained by additional automated fusion. Only the difference in the Y-direction proved significant (P < 0.05).

was 2.7 ± 0.9%. The mean ratio between 99mTc-DPD uptake in the vertebral body and in background was 7211 ± 679% (AC) and 3631 ± 410% (non-AC). In these three patients, visual inspection of the fusion between the attenuation-corrected and uncorrected scans did not

In the lower lumbar spine, the anatomical accuracy of hardware-based registration between SPECT and spiral CT was well below the pixel width of 4.6 mm in the vast majority of cases studied. To the best of our knowledge, data on the registration quality between bone SPECT and CT using a hardwarebased approach are scarce. Groves and co-workers [23] described a co-registration technique between 16-detector multislice CT and SPECT employing a device to fix the forearm into which fiducial markers were integrated. They reported degrees of misregistration ranging between 2 and 5 mm in three patients; obviously, their technique cannot be applied to the lumbar spine. To allow for registration between 111In-octreotide SPECT and CT, Fo¨rster et al. [24] used a vacuum cushion and internal landmarks for registration; they reported average anatomical inaccuracies between 6.47 and 7.78 mm. Their data do not lend themselves to comparison with our results because the majority of the metastases studied by

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526 Nuclear Medicine Communications 2006, Vol 27 No 6

between the scans; in particular in these patients, additional automated linear registration proved useful by reducing the inaccuracy of the match below pixel size. These data encourage the use of post-hoc correction of the match on a regular basis. Clearly, all pairs of images obtained from SPECT/CT should be visually screened for misregistration before clinical interpretation.

Fig. 5

120

> 1 pixel

80 60 40

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Percentage of patients in whom the X-, Y- and Z-distances were smaller (light columns) or greater (dark columns) than one pixel width of 4.6 mm. AAF indicates results obtained by additional automated fusion. AAF significantly reduced the number of patients with a misregistration greater than 4.6 mm from 14 to 0% (P < 0.05).

Fo¨rster et al. were located in the upper abdomen and thus subject to movements caused by respiration. In PET/CT, the overall inaccuracy has been reported to be 3-4 mm [22]. However, also in this case, the major contribution to these values is from respiratory movements. These do not affect the positions of the lower lumbar vertebral bodies so our data cannot be directly compared to these values. For pelvic lesions, no numeric data of registration accuracy for PET/CT have been published as yet. In general, the anatomical accuracy of image registration seems to be better than that of retrospective image fusion [25,26]. However, so far, the available evidence stems only from studies registering PET and CT; systematic investigations into this question are missing for SPECT/ CT. Theoretically, software-based image fusion for the skeleton should be easier than that between PET with 18 F-fluorodeoxyglucose and CT, for example, because the identification of landmarks visible on both bone SPECT and CT seems more straightforward. In our patients, the anatomical accuracy of hardwarebased registration between SPECT and spiral CT could still be improved by applying linear automated image fusion: with this post-hoc correction, the distances between the centres of gravity of the SPECT and CT representation of the two lower vertebral bodies did not exceed pixel size in any of the patients studied. Clearly, minor movements between CT and SPECT may also occur when a hybrid camera is used, especially in critically ill patients. In our study, three patients showed significant SPECT/CT misregistration due to movements

Hybrid cameras offer the potential to shorten the diagnostic process since two hitherto separate examinations can be performed directly after another. Agreement exists in the field that the correlation between structural and molecular imaging improves diagnosis and subsequent management in roughly one third of patients studied [12]. However, it is as yet unclear how accurate two data sets from different modalities should be matched to guarantee this increase in diagnostic quality. Side-by-side reading of two separately acquired sets of images is easy to perform and has been integrated into the clinical work of probably every nuclear medical institution. Studying this approach more closely, Reinartz and co-workers [27] concluded that hybrid PET/CT cameras would offer no additional benefit in most of the patients studied. Their view is not shared by all authors, however [22,28,29]. The clinical value of software-based retrospective rigid registration has been evaluated repeatedly also for SPECT and CT [2,9,11], and most researchers did not describe the errors in registration inherent in this method as a major drawback. When CT enables detection of clear-cut osteolyses in a patient referred for staging, an exact data match between CT and skeletal SPECT may not be needed. However, it seems conceivable that the match may be useful to exactly correlate enhanced uptake of polyphosphonates with a benign osseous lesion such as spondylarthrosis. If focal uptake did not match exactly to the CT-proven benign lesion, suspicion of malignancy might remain, motivating further examinations such as magnetic resonance imaging. Clearly, more evidence is needed to define the clinical value of the nearly perfect data match offered by a hybrid SPECT/spiral CT system for bone imaging. One possible drawback of our study is the possible imprecision of the manual localization of the centre of the vertebral bodies L4 and 5, since no automated procedure was used for this purpose. However, the X-, Y- and Zdistances measured by the two observers correlated significantly, indicating a high reproducibility of this interactive method. To reduce errors even further, we also averaged the data for the two vertebral bodies so that each data point entered into the statistical analysis represents four independently performed measurements. Another potential drawback of our data are difficulties in defining the boundary between bone and soft tissue on attenuation-corrected scans and hence also the centre of gravity in patients with SPECT/CT misalignment

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Accuracy of SPECT/spiral CT in the lower spine No¨mayr et al. 527

because this may lead to a falsification of uptake values in attenuation-corrected scans. We have studied this question systematically in another set of data; the results of this study will be presented on the DGN Congress (German Society of Nuclear Medicine) in April in Berlin, Germany [30]. We found that deliberate misalignments between SPECT and CT by 1 cm lead to errors in the regional quantification of DPD uptake in the order of 30% [30]. However, the misalignment detected in the group of patients recruited for this study was much smaller, usually well below pixel size so that these findings cannot be extrapolated to the data reported in this manuscript. Three patients exhibited more severe misregistration between SPECT and CT, ranging between 5 and 7 mm. Using an edge-finding algorithm, we could demonstrate that the vertebral boundaries defined on the non-attenuated scans were identical to those for the attenuation-corrected tomograms (Fig. 2). In those three cases, the relative 99mTc-DPD uptake measured in the misaligned region on the attenuation-corrected images differed, on average, by only 2.7 ± 0.9% from that determined in the non-corrected data. This is negligible compared to the mean ratio between 99mTc-DPD uptake in the vertebral body and background which was 7211 ± 679% (AC) and 3631 ± 410% (non-AC). We therefore believe that the minor errors in the visualization of radioactive distribution on the corrected SPECT scans caused by potential misalignment between SPECT and CT did not significantly affect the determination of the centres of gravity in our patients. In this context, it should also be mentioned that our data cannot be extrapolated to the thorax where respiratory movements may lead to greater errors of registration than reported here. However, lower back pain, with a prevalence of 33% [31], is very frequent [32]. Therefore, a study of the quality of image fusion afforded by a SPECT/spiral-CT system in the lower lumbar spine yields data of interest for the clinical application of this technology to a relatively high number of patients with that complaint.

Conclusions Hardware-based fusion between skeletal SPECT and CT offers a nearly perfect data match in the lower spine. The additional use of a tool for automated rigid registration has the potential to reduce the error of alignment even further and may be useful in patients with reduced compliance leading to movements between the two examinations.

Acknowledgements The authors gratefully acknowledge the technical support by Ms Anja Reimann and Mr Willy Amann in performing the SPECT/spiral-CT studies. The Symbia T2 system was kindly provided by Siemens Medical Solutions.

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24 Fo¨rster GJ, Laumann C, Nickel O, Kann P, Rieker O, Bartenstein P. SPET/CT image co-registration in the abdomen with a simple and cost-effective tool. Eur J Nucl Med Mol Imaging 2003; 30:32–39. 25 No¨mayr A, Romer W, Hothorn T, Pfahlberg A, Hornegger J, Bautz W, Kuwert T. Anatomical accuracy of lesion localization. Retrospective interactive rigid image registration between 18F-FDG-PET and X-ray CT. Nuklearmedizin 2005; 44:149–155. 26 Kim JH, Czernin J, Allen-Auerbach MS, Halpern BS, Fueger BJ, Hecht JR, et al. Comparison between 18F-FDG PET, in-line PET/CT, and software fusion for restaging of recurrent colorectal cancer. J Nucl Med 2005; 46:587–595. 27 Reinartz P, Wieres FJ, Schneider W, Schur A, Buell U. Side-by-side reading of PET and CT scans in oncology: which patients might profit from integrated PET/CT? Eur J Nucl Med Mol Imaging 2004; 31:1456–1461.

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Jager PL, Slart RH, Corstens F, Oyen WJ, Hoekstra O, Teule J. PET-CT: a matter of opinion? Eur J Nucl Med Mol Imaging 2003; 30:470–471; author reply 471. 29 Dresel S, Schwenzer K, Brinkbaumer K, Schmid R, Szeimies U, Popperl G, Hahn K. [F-18]FDG imaging of head and neck tumors: comparison of hybrid PET, dedicated PET and CT. Nuklearmedizin 2001; 40:172–178. 30 Ro¨mer W, Schulz V, No¨mayr A, Hornegger J, Herzog H, Bautz W, Kuwert T. Effect of CT-based attenuation correction on uptake ratios in skeletal SPECT. Nuklearmedizin 2006; in press. 31 Skovron ML, Szpalski M, Nordin M, Melot C, Cukier D. Sociocultural factors and back pain. A population-based study in Belgian adults. Spine 1994; 19:129–137. 32 Carragee EJ. Clinical practice. Persistent low back pain. N Engl J Med 2005; 352:1891–1898.

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Original article

Myocardial perfusion imaging in the elderly: a review Olivier De Wintera, Nico Van de Veireb, Filip Gemmela, Ingeborg Goethalsa and Johan De Suttera Coronary artery disease is a major cause of morbidity and mortality in the elderly population. As a result of ageing of the population and better medical, interventional and surgical treatment of patients with coronary artery disease, more and more elderly patients are referred to the cardiology department for diagnostic work-up. Stress testing, in combination with myocardial perfusion imaging, is routinely used in elderly patients, a population in which the diagnosis of significant coronary artery disease is often challenging because of atypical symptomatology. Since the introduction of technetium-99m ligands for myocardial perfusion imaging, it is possible to perform electrocardiogram-gated perfusion imaging. This not only improves the specificity of the test for coronary artery disease detection, but also enables the simultaneous assessment of left ventricular functional parameters. This

Introduction Coronary artery disease (CAD) is a major cause of morbidity and mortality in the elderly population [1–4]. As a result of ageing of the population and better medical, interventional and surgical treatment of patients with CAD, more and more elderly patients are referred to the cardiology department for diagnostic work-up. In the elderly patient population, the diagnosis of CAD is challenging because ischaemic symptoms are often atypical. In addition, the proportion of female patients is much higher in this population and, by the age of 75 years, the rates of coronary morbidity and mortality are similar in men and women [5]. Because of the high prevalence of CAD and the atypical symptomatology in the elderly, accurate non-invasive techniques are needed in this population to identify patients with significant CAD. Stress testing, in combination with myocardial perfusion imaging (MPI), is routinely used in elderly patients to assess CAD. Since the mid-1990s, there has been an increased use of technetium-99m (99mTc)-labelled perfusion agents, such as sestamibi and tetrofosmin, with more favourable imaging characteristics compared with thallium-201 (201Tl). This has made it possible to perform electrocardiogram (ECG)-gated single-photon emission computed tomography (SPECT) during the acquisition of myocardial perfusion [6], which not only improves the specificity for the detection of CAD [7], but also enables the assessment of left ventricular functional parameters,

article briefly overviews the possible stress modalities, diagnostic accuracy and prognostic value of myocardial perfusion imaging in elderly patients. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:529–534 Nuclear Medicine Communications 2006, 27:529–534 Keywords: cardiac function, coronary artery disease, diagnosis, elderly, myocardial perfusion, prognosis a

Nuclear Medicine Division and bCardiology Department, Ghent University Hospital, Ghent, Belgium. Correspondence to Olivier De Winter MD, Nuclear Medicine Division, Ghent University Hospital, De Pintelaan 185, 9000 Ghent, Belgium. Tel: ( + 32) 9 240 30 28; fax: ( + 32) 9 240 38 07; e-mail: [email protected] Received 20 November 2005 Accepted 16 February 2006

including the left ventricular ejection fraction (LVEF) and left ventricular volumes [8,9]. LVEF is one of the most powerful prognostic parameters [10–12], but cardiac volumes also provide important prognostic information in middle-aged CAD patients [13,14]. Therefore, perfusional parameters [15,16] and functional data seem to be important for prognostic assessment in elderly patients [17,18]. This article aims to overview briefly the use of MPI and gated MPI in the elderly patient population for diagnostic and prognostic purposes.

Coronary artery disease in the elderly In contrast with the middle-aged CAD population, women significantly outnumber men in the age group of 65 years and above. Death rates due to CAD have increased in women because of the more sedentary lifestyle in the elderly and the higher prevalence of obesity, diabetes mellitus type 2 and hypercholesterolaemia [19,20]. Data from the Framingham study revealed that women present with their first anginal symptoms 10 years later and sustain their first myocardial infarction 20 years later than men [21] but, by the age of 75 years, the rates of coronary morbidity and mortality are similar in both sexes [5]. This means that, in contrast with the middle-aged CAD population (40–65 years), the proportion of women in the elderly population with suspected CAD is much higher. In addition, the diagnosis of CAD is more difficult in elderly patients because of atypical symptomatology and a higher frequency of comorbidity. For example, exertional angina pectoris is commonly the

c 2006 Lippincott Williams & Wilkins 0143-3636

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first manifestation of CAD in middle-aged persons, but many elderly patients will not experience any exertional angina because of limited physical exercise in daily life. Secondly, other disorders can mask or mimic the ischaemic cascade: shoulder pain is frequently diagnosed as degenerative disease and epigastric pain as peptic ulcer disease. It has been shown that 20–50% of patients over 65 years of age demonstrate silent myocardial ischaemia on stress testing [22]. The detection of this silent ischaemia is important because coronary events are twice as common in patients with silent myocardial ischaemia than in those without ischaemia [23].

Stress testing in the elderly Physical ECG stress testing in the elderly patient

Current American College of Cardiology practice guidelines advise a simple treadmill test as first-choice investigation in elderly patients. However, the frequent presence of resting abnormalities in the ECG of elderly patients and the high proportion of women lower the value of ECG exercise stress testing in this population [24]. Secondly, the frequent presence of neurological, respiratory or orthopaedic disorders makes it more challenging to perform adequate exertional stress testing and to achieve maximal heart rates. Moreover, left bundle branch block and ventricular-paced rhythm, frequently seen in older subjects, are also established indications for pharmacological stress testing [25]. Therefore, ECG stress testing is less useful in elderly patients than in the middle-aged population. Stress testing for MPI in the elderly

The high prevalence of asymptomatic multivessel disease in the elderly population and the fact that a high proportion of patients over 75 years are not able to perform adequate stress testing for MPI, render the application of other stress modalities in this patient population necessary. Iskandrian et al. [26] reported that SPECT perfusion imaging after submaximal exercise is significantly less sensitive than that after maximal exercise in the detection of CAD and in the correct identification of patients with multivessel disease. In these patients, it is possible to perform perfusion imaging after pharmacological stress using agents such as dipyridamole, adenosine or dobutamine. Dipyridamole and adenosine cause vasodilatation in the normal coronary arteries more than in stenotic vessels. This leads to a relative flow difference of the diseased myocardial area as opposed to normal myocardial areas, which is visualized by MPI. Although the achievement of at least 85% of the predicted maximal heart rate is quite difficult to obtain in the elderly due to chronotropic incompetence or the use of b-blockers, the majority of elderly patients are still able to perform low-level exercise. Therefore, it is

possible to combine physical and pharmacological stress: physical exercise stress is started until exhaustion and followed by the infusion of pharmacological vasodilator stress agents (dipyridamole or adenosine). The combination of submaximal stress testing and pharmacological vasodilators not only improves the diagnostic ability of the stress test [27], but also enhances image quality and decreases the frequency of side-effects (compared with the use of pharmacological stressors alone) [28–30]. Although the use of vasodilators is relatively safe, it should be noted that the use of adenosine or dipyridamole is contraindicated in patients with active bronchospasm, acute coronary syndrome, hypotension, grade II– III atrioventricular block, sick sinus syndrome and sinus bradycardia, situations frequently seen in the elderly [31]. It should be noted that low-level exercise should not be added to pharmacological stress in patients with left bundle branch block, as they are susceptible to the same false-positive findings in the interventricular septum as would be seen with exercise. As an alternative to vasodilator stress, it is possible to use dobutamine, a synthetic catecholamine, as stress agent [32]. The infusion of dobutamine increases the heart rate and myocardial contractility, resulting in an increased cardiac output. This causes an increase in myocardial oxygen demand and blood flow of the normal vessels [33]. The chronotropic effect of dobutamine is, however, suboptimal, and therefore the target heart rate may not be reached [34]. In this case, atropine addition can increase the heart rate and the sensitivity for detection of ischaemic heart disease without increasing the sideeffects [35,36]. Contraindications for dobutamine infusion include severe hypertension and hypotension, a recent dissection of the aorta or coronary arteries, uncontrolled atrial fibrillation or atrial flutter and recurrent ventricular tachycardia. Dobutamine cannot be used as a stress agent for MPI in patients with left bundle branch block, because this may result in falsepositive findings in the interventricular septal wall.

Diagnostic accuracy of stress MPI in the elderly Radionuclide perfusion imaging, combined with exercise or pharmacological stress testing, demonstrates a high diagnostic accuracy in the detection of CAD in the middle-aged patient [37,38]. MPI has also been shown to have good accuracy in the elderly. Lam et al. [39] reported similar sensitivities and specificities for 201Tl-dipyridamole imaging in patients aged 70 years or above versus younger patients. More recently, Gentile et al. [40] investigated the accuracy of bicycle stress–rest MPI using 201Tl in 132 patients aged over 65 years who were hospitalized because of cardiac events (angina, dyspnoea, cardiac rhythm disturbances and atypical chest pain), using subsequent coronary angiography as a gold standard. In this study, a lesion on coronary angiography was

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Myocardial perfusion imaging in the elderly De Winter et al. 531

considered to be significant if 60% or more of the lumen diameter was obstructed. The diagnostic accuracy of MPI in this study was 86.3%, with a sensitivity of 93.5% and a specificity of 54.1%. The use of 99mTc-based myocardial imaging agents with more favourable imaging characteristics, resulting in less attenuation and scatter, has improved the specificity and diagnostic value of SPECT imaging in women [41]. Wang et al. [42] studied 75 consecutive patients aged over 80 years who underwent coronary angiography within 6 months of MPI using 99m Tc-labelled sestamibi. The overall sensitivity for detection of a stenosis of more than 75% was 95%, with a specificity of 75%, and the results were similar for pharmacological and exercise stress MPI. The introduction of 99mTc-labelled agents has also made it possible to perform ECG-gated SPECT during the acquisition of myocardial perfusion [6]. This technique facilitates the differentiation between attenuation artefacts and myocardial infarction in the case of fixed myocardial perfusion defects, and therefore results in a higher specificity of the technique. There are, however, no specific data on the possible added value of gated data for diagnostic purposes focused on an elderly patient population. Therefore, further studies are needed to clarify this issue in the elderly. Secondly, there are no data on the diagnostic accuracy of different stress modalities. Therefore, a head-to-head comparison of the diagnostic accuracy of these stress modalities should be the subject of further research.

Risk stratification by MPI in the elderly Prognostic value of myocardial perfusion assessment

Multiple studies have investigated the prognostic value of MPI in patients with known or suspected CAD for the prediction of cardiac events and cardiac mortality [43–48]. These studies have demonstrated the prognostic or incremental prognostic value of MPI, over and above clinical variables, in middle-aged patient populations with known or suspected CAD. Exercise MPI has significant added value for risk stratification in CAD, but most studied patients were middle-aged or younger [49,50]. Iskandrian et al. [15] studied 499 patients with CAD aged 60 years or above using exercise 201Tl planar imaging, and found a prognostic value for MPI in the prediction of future cardiac death or non-fatal myocardial infarction. Steingart et al. [51] investigated 578 patients aged 65 years or above with interpretable ECGs who were able to perform exercise testing with MPI (99mTc ligands and 201Tl). During a 4.4 ± 1.3-year follow-up, there were 39 deaths and 17 non-fatal myocardial infarctions. In this study population, the assessment of stress-induced ischaemia provided only limited prognostic information, over and above clinical parameters, in the prediction of all-cause death and myocardial infarction.

More recently, Schinkel et al. [52] investigated 272 patients aged over 65 years with limited exercise capacity using dobutamine tetrofosmin SPECT, and found that the summed stress score and abnormal myocardial perfusion (fixed or reversible) provided incremental information, over and above clinical data, in the prediction of all-cause mortality, cardiac death and the combined end-point of non-fatal myocardial infarction or cardiac death. As a result of ageing of the general population, a patient population above 60 or 65 years cannot really be considered as an elderly population. In a study by Shaw et al. [16], investigating 348 patients aged over 70 years who underwent dipyridamole planar 201Tl perfusion imaging, an abnormal perfusion scan was the best predictor of cardiac events (cardiac death or non-fatal myocardial infarction). Similar findings were reported by the same group in 120 patients older than 70 years undergoing exercise planar 201Tl perfusion imaging [53]. The first study investigating the prognostic value of MPI in a large population (328 patients) aged 75 years or above with suspected CAD was performed by Lima et al. [54]. In this population, there were 24 cardiac deaths during a follow-up time of 34 ± 15 months. These authors found that an abnormal myocardial perfusion scan (either fixed or reversible) was an independent predictor of cardiac death. More recently, Valeti et al. [55] reported the prognostic value of 201Tl perfusion imaging in 247 patients aged 75 years or above. They found that a higher summed stress score provided incremental information over and above clinical parameters. A higher summed difference score, ventricular enlargement (graded subjectively as present or absent) and increased uptake of 201Tl in the lungs were univariate predictors of cardiac death or myocardial infarction, but these parameters did not demonstrate any incremental value once the summed stress score was entered into the multivariate analysis. One of the most striking aspects of using MPI as a prognostic tool is the extremely low risk in patients with normal scintigraphic images [46,56,57]. Many clinical studies have demonstrated that patients with normal myocardial perfusion images have a very low cardiac event rate (less than 1% cardiac deaths per year). Even when exercise ECG (ST depression) or angiographic (multivessel disease) markers of poor outcome are present, the prognosis in patients with normal MPI is benign [58,59], and these findings seem to be similar in the elderly population [40,51]. In the study by Steingart et al. [51] in 578 patients aged 65 years or above with interpretable ECGs, who underwent exercise testing with MPI, normal scan findings were associated with a good prognosis (97% 3-year event-free survival rate).

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Cardiac death Yes SPECT 294 > 75 De Winter et al. [63]

247 > 75 Valeti et al. [55]

328 > 75 Lima et al. [54]

120 > 70 Hilton et al. [53]

348 > 70 Shaw et al. [16]

272 Schinkel et al. [52]

> 65

99m

Tc-tetrofosmin Dipyridamole and (2 day) bicycle

Cardiac death No SPECT

Cardiac death or MI No SPECT 99m

Tc-sestamibi Dipyridamole and (2 day) treadmill 201 Tl Treadmill

Cardiac death or MI No Planar Treadmill Tl

201

Cardiac death or MI Planar Dipyridamole

No

Cardiac death No SPECT Dobutamine

All-cause death or MI No SPECT Treadmill

Tc ligands and 201 Tl Tc-tetrofosmin (2 day) 201 Tl 99m

578 > 65 Steingart et al. [51]

EDV, end-diastolic volume; ESV, end-systolic volume; LV, left ventricular; LVEF, left ventricular ejection fraction; MI, myocardial infarction; MPI, myocardial perfusion imaging; NA, not available; RR, relative risk; SDS, summed difference score; SPECT, single-photon emission computed tomography; SRS, summed rest score; SSS, summed stress score. Hard event = cardiac death or non-fatal myocardial infarction. The studies by Schinkel et al. [52] and De Winter et al. [63] also included an analysis of total mortality. The study by Steingart et al. [51] did not include an analysis of hard cardiac events (cardiac death or myocardial infarction), only an analysis of all-cause mortality or myocardial infarction.

3 <1

44

SSS, SDS, LV SSS enlargement SSS, SRS, SRS and LV ESV abnormal MPI, LVEF, LV EDV and ESV 7 <1

50

Abnormal MPI <1

Abnormal MPI 11 73

Abnormal MPI <1

Abnormal MPI 9 53

Abnormal MPI 15 43 <1

Abnormal MPI

11 <1

43

LV enlargement, Ischaemia on ischaemia on MPI MPI Abnormal MPI Abnormal MPI NA NA

NA

Abnormal MPI Abnormal MPI 14 54 499 Iskandrian et al. [15]

> 60

99m

201

Tl

Treadmill

Planar

No

Cardiac death or MI

Mean of 25 months Mean of 51 months Mean of 38 months Mean of 23 months Mean of 36 months Mean of 34 months Median of 76 months Median of 26 months

<1

Significant univariate MPI predictors End-points Planar versus Gated SPECT data Stress modality Age Number of Tracer/protocol (years) patients Reference

Table 1

Overview of prognostic radionuclide myocardial perfusion studies in the elderly

Follow-up

Annual hard Percentage RR abnormal event rate of of normal versus normal normal MPI (%) MPI studies study

Significant multivariate MPI predictors

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Prognostic value of left ventricular dilatation at stress

The transient ischaemic dilatation (TID) ratio (TID ratio = ungated volume post-stress/ungated volume at rest imaging) is a marker for the transient enlargement of the left ventricle after stress. It is most commonly known for its diagnostic power as a marker of ischaemia [60]. However, the prognostic value of a high TID ratio has also been extensively investigated in middle-aged patient populations with CAD [60–62]. In the study by Steingart et al. [51], which has been discussed above, ventricular dilatation post-stress was strongly predictive of future death or non-fatal myocardial infarction. These findings were confirmed by Valeti et al. [55] and by our group [63]. Prognostic value of left ventricular functional parameters

Multiple studies have demonstrated the prognostic value of left ventricular functional parameters using gated radionuclide angiography [12,64,65], X-ray angiography [13], echocardiography [66–69] and even cardiac magnetic resonance [70] in the middle-aged population. Using gated SPECT, Sharir et al. [71] found that poststress LVEF and post-stress left ventricular end-systolic volume, assessed during gated SPECT, had an incremental value over and above clinical parameters in predicting cardiac death. Functional data assessed during echocardiography have shown prognostic value in elderly patients [72,73]. However, there is only one report on the prognostic value of combined perfusion and function imaging assessed by myocardial gated SPECT in the elderly. Over the period 1998–2002, we evaluated 294 consecutive patients aged 75 years or above (mean age, 78 ± 3 years) with suspected or known CAD using stress–rest myocardial gated SPECT, and followed these during a median follow-up of 25.9 months [63]. In these patients, left ventricular functional data (LVEF and left ventricular volumes) provided independent and incremental prognostic information over and above clinical and SPECT perfusion data for cardiac and all-cause mortality. More importantly, we found that the value of left ventricular functional measurements assessed by gated SPECT was clearly superior to that of myocardial perfusion parameters in the prediction of future cardiac death or total mortality. Further large studies are, however, needed to confirm these findings in other centres. A brief overview of these prognostic studies is provided in Table 1. Stress modalities

Although it is difficult to achieve at least 85% of the maximal predicted heart rate in elderly patients, the majority of the above-mentioned prognostic studies used physical stress testing in this population. Therefore, two concerns must be noted. First, it is possible

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Myocardial perfusion imaging in the elderly De Winter et al. 533

that patients were injected at submaximal stress, which could result in false-negative MPI studies. Secondly, some groups focused on a subgroup of elderly patients who probably have a different outcome than that of the general elderly patient population, as, for example, the group of elderly patients with an interpretable resting ECG. In our study, we added an intravenous dipyridamole infusion in patients not able to achieve 85% of their maximal predicted heart rate [63]. It is known that this improves the diagnostic abilities of the stress test [27]. Further studies are needed, however, to clarify whether this also leads to better prognostic abilities.

Conclusions The increasing number of elderly patients requiring diagnostic and prognostic assessment for CAD has necessitated accurate, non-invasive techniques applicable to this age group. Exercise testing, either alone or with perfusion imaging, remains a useful tool in elderly patients capable of performing vigorous treadmill or bicycle exercise. Fortunately, for the large elderly subset incapable of such exercise, pharmacological stress testing with dipyramidole, adenosine or dobutamine offers an excellent alternative. Furthermore, recent data support a prognostic approach to the management of middle-aged patients with CAD based on the results of gated myocardial perfusion SPECT imaging. Similarly, in the elderly, the assessment of left ventricular function adds further to the diagnostic and prognostic relevance of this imaging modality. This provides significant incremental information in the prediction of future cardiac death or all-cause mortality, especially in a patient population with multiple comorbidities.

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Original article

Regional cerebral blood flow in the assessment of major depression and Alzheimer’s disease in the early elderly Kazushi Hanadaa, Makoto Hosonob, Takashi Kudoc, Yoshie Hitomia, Yukinobu Yagyub, Eiji Kirimea, Yoshihiro Komeyab, Noa Tsujiia, Kazuhiko Hitomia and Yasumasa Nishimurad Background Alzheimer’s disease and major depression are representative diseases that present forgetfulness and a depressive mood. It is often difficult to make a differential diagnosis between the two in the initial phase. Aim To evaluate the differential diagnosis method using regional cerebral blood flow patterns with a three-dimensional stereotactic surface projection technique. Methods Twenty early-elderly patients with mild and moderate forgetfulness were studied. Among them, 10 were diagnosed as having major depression (the MD group) and the other 10 as having Alzheimer’s disease (the AD group). All patients underwent cerebral perfusion single photon emission computed tomography (SPECT) with [123I]iodoamphetamine. A z-score was calculated for each pixel of the cerebral surface. Twenty-one circular regions of interest (ROIs) were placed on the z-score map. The significance of the statistical difference in ROI values between the two groups was determined by using the two-sided Mann–Whitney U-test. Results The z-scores for the lateral parietal, lateral temporal, bilateral precuneus and bilateral posterior cingulate gyrus were significantly reduced in the AD group compared with those in the MD group. The z-scores for the

Introduction Alzheimer’s disease is the most common form of dementia, and accounts for approximately 60% of old age dementias [1]. It is estimated that it afflicts 5–10% of the population over 65 years of age, and is responsible for the most striking rise in the incidence of dementia in the very old. Scientists are searching for possible causes of Alzheimer’s disease, which may lead to the treatments that delay or even halt the dementing process. On the other hand, geriatric major depression has important medical, social and financial consequences. The overall prevalence of major depression among persons of 65 years of age or older is estimated to be 1.4% in women and 0.4% in men. Major depression is identified in 80% of suicide victims aged 75 years and more, while its frequency ranges from 3.1 to 29.4% in younger victims [2]. Major depression is the psychiatric disorder most likely to

lateral frontal, left thalamus and bilateral medial frontal regions were significantly lower in the MD group than in the AD group. Conclusion Our study demonstrated a difference in regional cerebral blood flow patterns between the early elderly with Alzheimer’s disease and those with major depression. All patients were classified into the appropriate categories using discriminant analysis and z-scores of frontal and parietal regions. Brain perfusion SPECT was a useful tool for the differential diagnosis between Alzheimer’s disease and major depression. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:535–541 Nuclear Medicine Communications 2006, 27:535–541 Keywords: Alzheimer disease, major depression, 3-D SSP, rCBF, linear discriminant analysis Departments of aNeuropsychiatry, bRadiology and, dRadiation Oncology, Kinki University School of Medicine, Osaka, Japan and cPET Unit, Research Institute, Shiga Medical Center, Japan. Correspondence to Dr Kazushi Hanada, Department of Neuropsychiatry, Kinki University School of Medicine, 377-2 Ohnohigashi, Osaka-sayama, Osaka 589-8511, Japan. Tel: + 0081 72 366 0221 (ext. 3285); fax: + 0081 72 366 0277; e-mail: [email protected] Received 23 November 2005 Revised 17 March 2006

increase suicide risks in the elderly. The treatment should be started and continued from the initial phase in both younger and older patients with major depression in order to obtain favourable outcomes [3]. These two diseases, Alzheimer’s disease and major depression, are completely different, but differential diagnosis is often difficult in the initial phase of the disease. This is because about 30% of Alzheimer’s disease patients also have some form of depression, and some elderly major depression patients develop a dementia syndrome that has been termed ‘pseudodementia’ or ‘depression with reversible dementia’ [4]. Studies of brain radionuclide imaging in both diseases have demonstrated important findings. Typically, brain perfusion single photon emission computed tomography (SPECT) in Alzheimer’s disease patients shows bilateral

c 2006 Lippincott Williams & Wilkins 0143-3636

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hypoperfusion in the parietal and posterior temporal lobes. As the disease progresses from mild to severe, the frontal cortex is affected and cognitive decline occurs at the same time [5]. This fact supports the finding that functional imaging deficit spreads from posterior to anterior of the temporal and frontal lobes with progression of the disease. On the other hand, brain SPECT in major depression patients shows various results including hypoperfusion of the prefrontal and temporal lobes [6], anterior cingulate gyrus [7], caudate and thalamus [8], and hyperperfusion of the parieto-occipital [9]. However, these results may depend on imaging modality, radiopharmaceuticals and quantitative methods. Statistical image analysis has merits: various imaging conditions can be unified, and objective and standardized data can be obtained. Minoshima et al. have developed a three-dimensional stereotactic surface projection (3-D SSP) technique [10,11] to statistically map the regions with reduced cerebral blood flow (CBF). 3-D SSP is a fully automated, user independent, data extraction method that has been applied extensively to images of SPECT and positron emission tomography. The aim of this study was to evaluate the method for differential diagnosis between Alzheimer’s disease and major depression in the initial phase of the disease, using regional CBF (rCBF) patterns with the 3-D SSP technique.

Materials and methods Patients

Between January 2003 and March 2004, 34 early-elderly patients (not more than 65 years old) complaining of mild or moderate forgetfulness (Mini-Mental State Examination (MMSE) [12] score of 15–25) underwent cerebral perfusion SPECT at our institution. All patients underwent magnetic resonance imaging (MRI), and the presence or absence of disease was confirmed by radiologists. With exclusion criteria of recent infarction (eight patients), cerebral haemorrhage (three patients), brain tumour (two patients) and brain injury (one patient), 20 patients (nine males and 11 females, age range 51–65 years) were enrolled in this study. The diagnosis of diseases was based on the criteria given in the Diagnostic and Statistical Manual of Mental Disorders, fourth edition (DSM-IV) [13] by psychiatrists using the Structured Clinical Interview for DSM-IV Axis I Disorders [14]. Among the 20 patients, 10 were diagnosed as having major depression (the MD group) and the other 10 as having Alzheimer’s disease (the AD group) (Table 1). No other type of dementia was

Table 1

Patient profile

Patient number Major depression group 1 2 3 4 5 6 7 8 9 10 Alzheimer’s disease group 1 2 3 4 5 6 7 8 9 10

Age (years)

Sex

HDS score

MMSE score

51 52 53 53 56 57 60 62 63 64

F M M M F F F M M F

22 25 21 20 23 25 18 22 24 23

22 22 24 20 19 18 25 20 19 22

53 55 58 59 61 61 62 64 65 65

F F F M F M M F F M

24 17 15 21 23 18 20 18 24 19

18 22 20 19 18 20 17 22 17 20

HDS, Hamilton Depression Scale; MMSE, Mini Mental State Examination.

identified in these patients. Diagnoses were confirmed by follow-up for at least 6 months. Patients also underwent the 17-item Hamilton Depression Rating Scale (HDS) to assess depression status [15]. Patients were not taking psychotic or anti-dementia medications. Five were on stable doses of prescribed medicines (two were on anti-hypertensives, two on gastric medications, and one on anti-hyperuricaemia drugs). All patients were right-handed. We explained the aim of this study to all patients and obtained their informed consent. Imaging procedure

All patients received [123I]iodoamphetamine (123I-IMP) at a dose of 167 MBq. They were then directed to lie in the supine position for 20 min with the eyes closed in a dim, quiet room. A SPECT image acquisition was performed using a double-head HITACHI/ADAC gamma camera ‘FORTE’ equipped with a low-energy, generalpurpose collimator (VXGP). A total of 64 projections (32 per head) were obtained at 25 s per projection into a 128  128 image matrix. A Butterworth filter was used for SPECT image reconstruction at 0.125 cycle/pixel. Attenuation correction was performed using Chang’s method. Data analysis

rCBF was projected onto surface pixels by using a 3-D SSP technique. 3-D SSP involves three steps of re-orientation, projection and transformation. Stereotaxic re-orientation of each brain into a standard shape was performed by non-linear warping with the anterior

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rCBF in assessment of major depression and Alzheimer’s disease Hanada et al. 537

Table 2

Fig. 1

Name

SPoG SPrG IPoG IPrG LPL LOL LPaT LPoT

Lateral

Definition of each region of interest

SFGBA9 SFGBA10 IFG LAT

Pre

CGy

PCG Tha

MFBA8 MFBA9 ACG MFBA10 MTL

Medial

Twenty-one ROIs (a total of 42 regions including both sides of the ROIs) were placed on a z-score map, and analysed by 3-D SSP. (The name of each ROI is given in Table 2.)

commissure–posterior commissure line as the anatomical line of reference. Subsequently, a method of data extraction was applied in which the cortical activity was projected onto the brain surface. Finally, as each brain had been stereotactically transformed into a standard surface image format, it was possible to compare the resultant cortical projections with a normal database by using a zscore subtraction on a pixel-by-pixel basis. Pixel values of an individual’s image set were normalized to the global brain value before the analysis. In this study, we also included data from 20 normal subjects who were matched for age and sex (mean age, 66.5 years; MMSE score, 28.83) on a surface pixel-by-pixel basis. All subjects underwent MRI and were confirmed to have no focal lesions. A z-score was calculated for each pixel of the cerebral surface as follows: z-score = (normal mean – individual value)/normal standard deviation. Twenty-one circular regions of interest (ROIs) consisting of 97 pixels were placed on a z-score map of 3-D SSP (Fig. 1, Table 2) as follows: inferior frontal gyrus including Brodmann areas (BA) 46/45; superior frontal gyrus including BA 10 (SFGBA10); superior frontal gyrus including BA 8/9; superior precentral gyrus including BA 4/6; inferior precentral gyrus including BA 4/6; superior post-central gyrus including BA 1/2/3/40; inferior postcentral gyrus including BA 1/2/3/40; lateral parietal lobe including BA 40/39; lateral occipital lobe including BA 18/19; lateral parieto-temporal including BA 21/22; lateral posterior temporal including BA 21/22; lateral anterior temporal including BA 21/28; precuneus including BA 7; posterior cingulate gyrus including BA 29/30; cingulate gyrus including BA 23/24; anterior cingulate gyrus including BA 24; thalamus; medial temporal lobe including BA 28/36; medial frontal including BA 8/32; medial frontal including BA 9/32; and medial frontal including BA 10/32 (MFBA10). Results obtained from z-scores of the ROIs were expressed as means of each pixel.

Lateral side IFG SFGBA10 SFGBA9 SPrG IprG SpoG IpoG LPL LOL LpaT LpoT LAT Medial side Pre PCG CGy ACG Tha MTL MFBA8 MFBA9 MFBA10

Location

Corresponding Brodmann area

Inferior frontal gyrus Superior frontal gyrus Superior frontal gyrus Superior precentral gyrus Inferior precentral gyrus Superior postcentral gyrus Inferior postcentral gyrus Lateral parietal lobe Lateral occipital lobe Lateral parieto temporal Lateral posterior temporal Lateral anterior temporal

45, 46 10 8, 9 4, 6 4, 6 1, 2, 3, 40 1, 2, 3, 40 39, 40 18, 19 21, 22 21, 22 21, 28

Precuneus Posterior cingulate gyrus Cingulate gyrus Anterior cingulate gyrus Thalamus Medial temporal lobe Medial frontal Medial frontal Medial frontal

7 29, 30 23, 24 24 – 28, 36 8, 32 9, 32 10, 32

BA, Brodmann area.

Statistical analysis

The significance of statistical differences was determined by the two-sided Mann–Whitney U-test in age, the HDS scores, MMSE scores and ROI values between the MD and the AD groups. Results were expressed as mean ± standard deviation, and differences with P values of less than 0.05 were considered to be statistically significant. Linear discriminant function analysis was performed to determine ROI values for the areas that were the best predictors of diseases.

Results The MD group consisted of five men and five women, with a mean age of 57.1 ± 4.9 years, HDS score of 22.3 ± 2.2, and MMSE score of 21.1 ± 2.3. The AD group included four men and six women, with a mean age of 60.3 ± 4.1 years, HDS score of 19.9 ± 3.1, and MMSE score of 19.3 ± 1.8. There were no significant differences in age (P = 0.131, U = 70), HDS score (P = 0.082, U = 27), or MMSE score (P = 0.096, U = 28) between the two groups. Characteristics of regional cerebral blood flow patterns

rCBF in the lateral temporal, parietal and precuneus decreased more in the AD group than in the MD group, while in the lateral and medial frontal it decreased more in the MD group than in the AD group (Figs 2 and 3). The mean of each ROI z-score in the two groups was calculated (Table 3). In the AD group, decreased blood flow was observed compared with normal subjects in the region from the lateral parietal lobe to temporal lobe, anterior cingulate gyrus, pons and medial temporal lobe

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Fig. 2

Rt. LAT

Lt. LAT

SUP

INF

Z

5 4 3

ANT

POST

Rt. MED

Lt. MED

2 1 0

Regions of decreased perfusion of the Alzheimer’s disease (AD) group in comparison with the major depression (MD) group. Values of each pixel of the AD group and MD group were statistically analysed with Student’s t-test to calculate z-scores of mean pixel values, which were then projected and displayed on the brain surface map.

Fig. 3

Rt. LAT

Lt. LAT

SUP

INF

Z

5 4 3

ANT

POST

Rt. MED

Lt. MED

2 1 0

Regions of decreased perfusion of the major depression group in comparison with the Alzheimer’s disease group.

(Fig. 4). On the other hand, in the MD group, decreased blood flow was noted in the medial frontal lobe, lateral inferior frontal lobe, cingulate gyrus, pons the medial temporal lobe (Fig. 5).

Discussion For both Alzheimer’s disease and major depression, there are various diagnostic criteria such as DSM-IV and the International Classification of Disease, Diagnosis Criteria for Research, 10th edition (ICD-10) [16]. These criteria are used in clinical settings to perform the differential diagnosis between Alzheimer’s disease and major depression in the early elderly. Most of diagnostic criteria adopt only clinical symptoms, and not diagnostic imaging or laboratory test values. Therefore, differential diagnosis using these criteria is difficult except for expert psychiatrists.

It has been reported that pseudodementia observed in major depression patients and forgetfulness in Alzheimer’s disease patients show different clinical manifestations [17]. However, the difference is often not clear in the initial phase of the disease, although it becomes remarkable as the disease progresses. Recently, anti-dementia drugs such as cholinesterase inhibitors have been introduced to treat Alzheimer’s disease. It is reported that the drug will be more efficacious when administered from the earlier phase of the disease [18]. The effective use of anti-dementia drugs drives down the cost of care in nursing homes, and is cost-effective for all medical services for the early elderly [19]. It is also known that, even in elderly patients with major depression, therapeutic intervention from the early phase leads to a decrease in suicide attempts [3]. In the present study, the images obtained by brain perfusion SPECT were analysed using 3-D SSP, and ROIs were placed on the z-score map obtained, in order to clarify the features of Alzheimer’s disease and major depression. Comparison of z-scores between the Alzheimer’s disease and major depression groups

z-scores were compared statistically between the AD and MD groups. The z-scores for the lateral parietal (bilateral superior post-central gyrus, lateral parietal lobe and right lateral occipital lobe), lateral temporal (left lateral parieto-temporal and lateral anterior temporal), bilateral precuneus and bilateral posterior cingulate gyrus regions were significantly reduced in the AD group compared with those in the MD group. The z-scores for the lateral frontal (bilateral inferior frontal gyrus and SFGBA10), left thalamus and bilateral MFBA10 regions were significantly lower in the MD group than in the AD group. Compared to normal subjects, our AD group showed reduced blood flow in the lateral parietal lobe, lateral temporal lobe, anterior cingulate gyrus, pons and medial temporal lobe, which was consistent with the results of previous reports [20,21]. On the other hand, the MD group was characterized by decreased perfusion in the medial frontal lobe, lateral inferior frontal lobe, anterior cingulate gyrus, pons and medial temporal lobe [21]. The regions where decreased blood flow was observed in both groups were therefore the anterior cingulate gyrus, medial temporal lobe and pons. Reduced perfusion in the prefrontal area consisting of the medial frontal lobe and lateral inferior frontal lobe was specific to the MD group. It is thought that the regions of the prefrontal area, anterior cingulate gyrus, medial temporal lobe and hippocampus are involved in a network associated with the pathologies of depressive states [22]. These regions of this network corresponded to the regions where reduced perfusion was observed in our study.

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rCBF in assessment of major depression and Alzheimer’s disease Hanada et al. 539

Table 3

z-score of each region of interest in the Alzheimer’s disease and major depression groups

Region

z-score Alzheimer’s disease group

Lateral side IFG SFGBA10 SFGBA9 SPrG IPrG SPoG IPoG LPL LOL LPaT LPoT LAT Medial side Pre PCG CGy ACG Tha MTL MFBA8 MFBA9 MFBA10

Comparison (Mann–Whitney U-test) Major depression group

P

U

1.259 0.913 0.879 0.945 1.388 1.476 0.264 0.209 0.250 0.445 0.308 0.094 0.345 0.388 0.682 0.359 0.080 0.118 0.208 0.380 1.253 0.759 0.848 0.773

0.008 0.016 0.034 0.003 0.762 0.597 0.385 0.496 0.082 0.762 0.019 < 0.001 1.000 0.406 0.001 < 0.001 0.028 0.041 0.045 0.001 0.049 0.070 0.226 0.034

15ww 18w 22w 11ww 46 57 38.5 41 27 46 81* 96** 50 61 93** 97** 79* 77 76.5 92** 76 74 66 78*

0.263 0.377 0.691 0.551 0.690 0.731 1.263 1.180 0.939 1.053 0.723 0.492 1.605 1.289 0.880 0.576 0.726 0.348

0.002 0.007 0.028 0.010 0.112 0.112 0.762 0.940 0.131 0.028 0.326 1.000 0.257 0.650 0.290 0.131 0.028 0.010

91** 86** 79* 84* 71 71 46 51 30 21w 37 50 35 44 36 30 21w 16w

Mean

SD

Mean

SD

right left right left right left right left right left right left right left right left right left right left right left right left

0.402 0.132 0.330 0.283 0.482 0.759 0.148 0.096 0.123 0.372 0.910 0.848 0.508 0.809 3.065 1.856 0.483 0.904 1.181 1.573 1.763 1.532 1.882 1.975

0.773 0.153 0.360 0.203 0.422 0.452 0.167 0.149 0.189 0.481 0.777 0.614 0.651 1.060 1.994 1.129 0.625 0.883 1.405 1.301 1.509 1.646 1.631 1.430

1.499 0.817 0.983 1.123 0.967 1.093 0.242 0.161 0.252 0.332 0.288 0.058 0.372 0.395 0.658 0.173 0.032 0.103 0.176 0.192 0.978 0.460 0.831 0.723

right left right left right left right left right left right left right left right left right left

1.230 1.181 1.416 1.569 1.137 1.233 2.207 1.997 1.299 1.588 2.650 2.820 0.814 0.531 0.843 0.753 0.504 0.503

0.771 0.963 0.796 0.570 0.508 0.453 1.209 1.247 0.742 0.788 1.091 1.407 0.584 0.409 0.569 0.487 0.522 0.428

0.192 0.225 0.657 0.718 0.776 0.841 2.444 2.187 2.051 2.587 3.076 2.702 1.544 1.091 1.476 1.141 1.032 0.914

Refer to Table 2 for name of each region of interest. Alzheimer’s disease group significantly more than major depression group, *P < 0.05, **P < 0.01. Major depression group significantly more than Alzheimer’s disease group, wP < 0.05, wwP < 0.01.

Although impaired rCBF in the anterior cingulate gyrus has been reported as a feature of elderly patients with major depression [7], it is neither specific to major depression nor allows differentiation between Alzheimer’s disease and major depression. It is commonly observed as a part of abnormal perfusion in the network in the various depressive states as shown in this study and previous studies [23]. Rather, the prefrontal perfusion abnormality clearly depicted on the z-score maps should be noted in discriminating major depression and Alzheimer’s disease. In this study the MD group showed a significant decrease in rCBF in the prefrontal area, while the AD group did not. Precise estimation of the distribution of z-scores in the aforementioned ‘depression’ network might enable differential diagnosis of diseases causing depression.

In stereotactic methods such as 3-D SSP and statistical parametric mapping, artifacts resulting from inaccuracy in stereotaxic transformation may occur in atrophic regions of the brain. However, findings obtained by 3-D SSP have been reported to be less affected by such artifacts than statistical parametric mapping [24]. The effect of artifacts on the findings obtained by 3-D SSP in this study was thought to be negligible as our patients, who were of younger age and had good MMSE scores, did not have remarkable brain atrophy. We must mention that in 3-D SSP, the choice of reference region for pixel normalization may affect the accuracy of disease characterization, and is still controversial. In diseases with widespread metabolic or perfusion reduction, the pons and thalamus may be preferable [11,20].

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540 Nuclear Medicine Communications 2006, Vol 27 No 6

Fig. 4

Fig. 6

MD

8

AD

7

Right

Lateral

Medial

Parietal (LPL)

6 5

LPL = 1.24 × SFGBA10 + 1.04

4 3 2

Left

1 0

Less than 1.0

1.0 − 1.5

1.5 − 2.0

2.0 or more

z-scores of each ROI in the Alzheimer’s disease group. The circles deeper in colour indicate the higher z-scores: white, less than 1.0; light grey, 1.0–1.5; dark grey, 1.5–2; and black, 2.0 or more. Decreased blood flow was observed in the regions from the lateral parietal lobe to temporal lobe, anterior cingulate gyrus, pons and medial temporal lobe.

0

0.5

1

1.5 2 2.5 3 3.5 Frontal (SFGBA10) ROI values in the lateral parietal lobe (LPL) and superior frontal gyrus, including Brodmann area 10 (SFGBA10), observed in major depression (K, n = 10) and Alzheimer’s disease (D, n = 10) groups. A scatter plot was presented with SFGBA10 value on the horizontal axis and LPL value on the vertical axis. A straight line, LPL = 1.24  SFGBA10 + 1.04, obtained from a linear discriminant analysis represents the differentiation criteria between Alzheimer’s disease and major depression. Correlation ratio, 0.71; error ratio, 7.04%.

Fig. 5

Right

Lateral

Medial

Left

Less than 1.0

1.0 − 1.5

1.5 − 2.0

2.0 or more

z-scores of each ROI in the major depression group. Decreased blood flow was observed in the medial frontal lobe, lateral inferior frontal lobe, cingulate gyrus, pons and medial temporal lobe.

distribution using linear discriminant function analysis. ROIs, where perfusion was supposed to be observed and a decrease in perfusion specific to Alzheimer’s disease and major depression, were evaluated. Among all the combinations of ROIs in Alzheimer’s disease and those in major depression, the combination of a parietal ROI of the lateral parietal lobe and a prefrontal ROI of SFGBA10 was the most efficient in discriminating the two diseases. The higher z-score at the right and left sides of ROIs were regarded as the ROI values of each individual, and a scatter plot was presented with an SFGBA10 value on the horizontal axis and lateral parietal lobe value on the vertical axis (Fig. 6). The plot area could be divided into two parts, those of major depression and Alzheimer’s disease, by drawing a discriminant line obtained in the linear discriminant function analysis. All the patients were correctly classified into the appropriate group of major depression and Alzheimer’s disease, and the result of the function analysis was highly accurate (correlation ratio, 0.71; error rate, 7.04%).

We used a global brain value as the reference because this reportedly provided high accuracy in discriminating between patients with early Alzheimer’s disease and control subjects [25], and good differentiation between Alzheimer’s disease and major depression showed that the choice of global value was appropriate in our patient groups.

Conclusion

Differentiation between Alzheimer’s disease and major depression using z-scores

References

We constructed an automated method for differential diagnosis between Alzheimer’s disease and major depression on the basis of the above-mentioned z-score

There was a difference in rCBF patterns between the early elderly with Alzheimer’s disease and those with major depression, and the difference became clear when 3-D SSP was used. Brain perfusion SPECT with 3-D SSP might be a useful tool for the differential diagnosis between Alzheimer’s disease and major depression.

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rCBF in assessment of major depression and Alzheimer’s disease Hanada et al. 541

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Correspondence

Correspondence Nuclear Medicine Communications 2006, 27:543–544

Increasing the radiochemical purity of 99mTc-sestamibi commercial preparations results in improved sensitivity of dual-phase planar parathyroid scintigraphy James R. Ballingera and Margaret S. Cooperb a Nuclear Medicine, Guy’s and St Thomas’ Hospital, London, UK and b Nuclear Medicine, St Bartholomew’s Hospital, London, UK. Correspondence to James R. Ballinger, Nuclear Medicine, Guy’s and St Thomas’ Hospital, St Thomas Street, London SE1 9RT, UK. Tel: þ 44(0)207 188 5521; fax: þ 44(0)207 188 4094; e-mail: [email protected] Received 17 January 2006 Accepted 16 February 2006

The recent paper, ‘Increasing the radiochemical purity of 99m Tc-sestamibi commercial preparations results in improved sensitivity of dual-phase planar parathyroid scintigraphy’, by Karam et al. [1], presents an interesting hypothesis that the radiochemical purity of 99mTcsestamibi affects the sensitivity of dual-phase parathyroid scintigraphy. The authors show a direct relationship between the percentage of impurities in the 99mTcsestamibi preparation and the parathyroid washout ratio. The data presented are clearly suggestive, but we feel that there are a number of technical anomalies in the paper which significantly weaken its conclusions. Firstly, the authors state that they achieved a higher radiochemical purity by altering the technetium to sestamibi ratio; however, it is not clear how they achieved this. It is stated that the total quantity of technetium (99mTc plus 99Tc) was reduced from 1 mg/mg sestamibi to 0.34 mg/mg sestamibi, yet the only manipulation mentioned was the addition of stannous chloride dihydrate to a total of 63 mg. This is puzzling as the kit already contains 75 mg of tin [2]. As this is an important aspect of the paper, the authors should describe in detail how this was performed. Secondly, inspection of Figs. 1 and 4 indicates that both the washout ratios and radiochemical purities in Group 1 show a bimodal distribution, with one subgroup superimposing with Group 2 and the other subgroup at higher washout ratios and higher percentage impurities. This suggests that there may be two ‘populations’ of sestamibi preparations. Thirdly, the authors make much of the fact that the United States Pharmacopeia limit is 90% radiochemical

purity, and suggest that off-site commercial radiopharmacies are supplying 99mTc-sestamibi barely above this limit. However, we would suggest that it is their own results which are out of line. In Group 1, the average radiochemical purity was 91.9%. This is at odds with our experience and that of most published papers. Records from our two laboratories show radiochemical purities of 96.771.0% (n ¼ 95) and 96.871.1% (n ¼ 90). In a retrospective review, Decristoforo et al. [3] reported radiochemical purities of 96.3477.66% (n ¼ 249), where the wide range was due to the use of fractionated kits. These results agree with those found in Group 2, but without the requirement for an altered formulation. One possible reason for the low radiochemical purity obtained by the authors could be the excessive amount of 99m Tc added to the kit. The current package insert states a limit of 5.55 GBq of 99mTc [2], but the authors added 18.5 GBq, more than three times higher, although no comment was made on this point. Deviation from a manufacturer’s instructions is not uncommon, but, in such a situation, the responsibility for the quality of the product lies solely with the radiopharmacy and not the manufacturer. The authors suggest that the non-sestamibi tracer is primarily 99mTc-pertechnetate. However, it has been known since the first clinical trials of 99mTc-sestamibi (then RP-30) that the typical radiochemical purity is about 96% with approximately 2% pertechnetate and approximately 2% colloid (NEN DuPont, personal communication, 1988). This is indeed what we obtained using a quality control method which measured the two types of impurities separately [4]: 1.670.7% pertechnetate and 1.870.7% colloid (n ¼ 95). However, it could well be that, at the lower radiochemical purity values which the authors observed, the main impurity may be pertechnetate. Finally, in five of the 42 studies in Group 1, the radiochemical purity was less than 90%, yet the radiopharmaceutical was still administered to the patient. What this paper has shown is that, if one deviates from the manufacturer’s instructions, one risks preparing a substandard product which can produce unreliable clinical results. Thus, the title of the paper is misleading

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544 Nuclear Medicine Communications 2006, Vol 27 No 6

and has raised unnecessary concern about the reliability of parathyroid imaging.

2 3

References 1

Karam M, Dansereau RN, Dolce CJ, Feustel PJ, Robinson LW. Increasing the radiochemical purity of 99mTc-sestamibi commercial preparations results in

4

improved sensitivity of dual-phase planar parathyroid scintigraphy. Nucl Med Commun 2005; 26:1093–1098. Bristol-Myers Squibb. Cardiolites package insert. N. Billerica, MA: Bristol-Myers Squibb Medical Imaging; May 2003. Decristoforo C, Siller R, Chen F, Riccabona G. Radiochemical purity of routinely prepared 99Tcm radiopharmaceuticals: a retrospective study. Nucl Med Commun 2000; 21:349–354. Proulx A, Ballinger JR, Gulenchyn KY. Routine determination of radiochemical purity of 99mTc-MIBI. Appl Radiat Isot 1989; 40:95–97.

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NEWS AND VIEWS JUNE 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up-to-date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 206 6930, E-mail: [email protected]

Nuclear Medicine Communications, 2006, 27, 545–546 Are we ready for the myocardial challenge?

A recent journal article received widespread news coverage, and we believe that it could represent both a significant challenge, and opportunity, for providers of nuclear medicine services. The story is a report on the paper by Nissen et al. in the Journal of the American Medical Association (Volume 295, Number 10, online version only). The article reports on the ASTEROID trial which was a 2-year prospective study of the use of a high-dose statin. Previous studies have shown the value of statins in lowering cardiac hard events by stabilising existing atheromatous disease, but this study presents evidence that the plaques actually reduce in size under the protocol employed. Admittedly, the trial did not demonstrate a ‘cure’ but rather, a fairly modest reduction in crosssectional area of the plaques. Nonetheless, if replicated, this could point in the direction of a switch from interventional to medical treatment of coronary artery disease. If patterns of therapy change in this way, the balance will also tip towards physiological and early diagnostic assessment. Nuclear cardiology should have an advantage here because the decision to treat medically does not require the

identification of specific vessels to treat (i.e. myocardial perfusion studies could become the one-stop diagnostic shop, instead of the gatekeeper to angiography). Similarly, ischaemia that is identified by myocardial perfusion but missed by coronary artery imaging (such as small vessel disease) would have a greater diagnostic value if improved medical therapy becomes available. The nuclear cardiology community needs to rise to this challenge. We need to develop myocardial scanning to be the trusted imaging of choice and so turn the tide against multislice computed tomography, magnetic resonance imaging and low-dose dobutamine echo. The physiological nature of nuclear medicine is a two-edged sword. Despite all the advantages, it lacks the seeing-is-believing feel of highquality anatomical imaging. We must compensate for this by building on our robust reputation for accuracy. Although there are many papers demonstrating good levels of accuracy, there is still space for improvement. Despite developments in instrumentation and tracers subsequent to the days of single-headed thallium single-photon emission computed tomography, it is hard to point to a parallel improvement in reported accuracy,

and a few worrying loose ends remain, such as whether attenuation correction really helps and whether 1801 or 3601 of data acquisition is optimal. Once such questions have been answered and the technique optimised across all centres, we will hopefully have a narrower, more precisely known, range of myocardial counts in normal subjects (i.e. we will be able to create a new normal database with a narrower range) and so be more confident in diagnosing early disease. This aim could be achieved in two steps. Firstly, in anthropomorphic phantoms with a regular left ventricle, we need to optimise the technique and demonstrate the (very narrow) range of count density obtained in images. Phantom data are not convincing on their own; therefore, high-quality studies in humans using the optimised technique will also be required to demonstrate an improved clinical accuracy. It would also help to make myocardial tests more convenient. In the absence of an ischaemia tracer, the next best thing would be a rest–stress protocol completed in say 30 min without moving the patient from the scanning couch. Impossible? With multi-headed cameras and adenosine rather than exercise

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546 Nuclear Medicine Communications 2006, Vol 27 No 6

stressing, this might be an achievable objective. Finally, there must be agreement about the protocol and widespread acceptance through more prescriptive guidelines. To take full advantage of the opportunities ahead, we need to ensure that our services have both the perception and reality of being accurate, reliable, robust and timely. Meeting Announcements BNMS Autumn Meeting

Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk EANM Annual Meeting

Dates: 30 September to 4 October 2006 Venue: Athens, Greece Website: www.eanm.org

9th World Congress of Nuclear Medicine and Biology

Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/congress2006/ index02.htm International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM) Dates: 13–15 November 2006 Venue: Vienna, Austria Website: www-pub.iaea.org/MTCD/ Meetings/ Announcements.asp?ConfID = 146 Education and Training

EANM Learning Courses Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria

Contact: EANM executive Secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected] EANM distance learning in nuclear cardiology This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. http://www.eanm.org/eduOnline/ edu_start.php?navId = 332

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Editorial

Molecular imaging of Takayasu’s arteritis and other large-vessel vasculitis with 18F-FDG PET Myles Webba and Adil Al-Nahhasb Nuclear Medicine Communications 2006, 27:547–549 Keywords: FDG-PET, Takayasu’s arteritis, Large vessel vasculities.

Correspondence to Dr Adil Al-Nahhas, Department of Nuclear Medicine, Hammersmith Hospital, Du Cane Road, London W12 0HS, UK. Tel: + 0044 208 383 4923; fax: + 0044 208 383 1700; e-mail: [email protected]

a Departments of Nuclear Medicine, The Prince Charles Hospital, Brisbane, Australia and bHammersmith Hospital, London, UK.

Introduction The role of positron emission tomography (PET) in oncology, cardiology and neurology is firmly established as an important imaging tool in diagnosis and management. New and emerging roles are also being established for PET in a variety of other medical and surgical specialties. It is in the field of inflammation and infection, however, that PET is becoming noticed for its diagnostic capabilities. One of its emerging applications is the assessment of patients presenting with non-specific symptoms (e.g., fever of unknown origin, arthralgia, weight loss) who may have an underlying malignancy or an inflammatory condition such as vasculitis [1]. 18

assess vessel wall inflammation and provide a measure of disease activity. Angiography is the ‘gold standard’ imaging technique but is invasive and unreliable in the diagnosis of large-vessel vasculitis in the early (potentially reversible) inflammatory phase [2]. Magnetic resonance imaging (MRI) can detect vessel wall oedema but this was found to be an inconsistent guide to subsequent anatomical changes [3]. It is also known that MRI tends to overestimate the degree of vascular lumen stenosis by up to 15% [4]. In a study by our group, we did not find the presence or absence of gadolinium enhancement of the vessel wall a reliable guide to disease activity [2].

F-FDG PET in vasculitis

Vasculitis is a clinico-pathologic process characterized by inflammation of and damage to blood vessels. The vessel lumen is usually compromised, and this is associated with ischaemia of the tissues supplied by the vessel involved. A broad and heterogeneous group of syndromes may result from this process, since any type, size and location of blood vessel may be involved. Broadly speaking, vasculitis can be divided into small (Wegener’s granulomatosis), medium (polyarteritis nodosa, Kawasaki’s arteritis) and large vessel vasculitis (Takayasu’s and giant-cell arteritis). Although the causal role of immune complexes has not been clearly established, their deposition in vessel walls followed by neutrophil infiltration is the most widely accepted pathogenic mechanism of vasculitic syndromes. As the process becomes subacute or chronic, mononuclear/inflammatory cells infiltrate the vessel wall. The clinical diagnosis of early vasculitis is unreliable due to the non-specific nature of the symptoms and absence of specific laboratory markers. A definitive diagnosis of large-vessel vasculitis can be obtained by arterial biopsy but selecting the appropriate vessel to biopsy can be difficult due to the systemic nature of the disease and, as such, false negative results are common. Reliance must therefore be placed on an imaging technique that could

In contrast, the metabolic activity in vasculitis can be demonstrated with 2-[18F]fluoro-2-deoxy-D-glucose (18FFDG) due to its accumulation within inflammatory cells such as macrophages, activated lymphocytes and granulation tissue. This phenomenon is related to increased FDG uptake in relation to augmented oxidative metabolism and has been clearly demonstrated with microautoradiography [5]. In routine 18F-FDG PET scans, the aorta and its immediate branches demonstrate normal background activity due to the low numbers of metabolically active cells dispersed between the many elastic layers of the arterial wall. In large-vessel vasculitis, however, the pattern of uptake becomes distinctly abnormal, extending in a linear pattern along the walls of the involved vessels, as shown in case reports of patients affected with systemic lupus erythematosus [6], Takayasu’s arteritis [7] and giant-cell arteritis (GCA) [8]. These findings were confirmed in publications concerning larger number of patients. Blockmans and colleagues compared the findings of 18F-FDG PET in 25 subjects with GCA or polymyalgia rheumatica with those of 44 age-matched control subjects. They concluded that 18 F-FDG PET had a sensitivity of 56%, specificity of 98%, positive predictive value of 93% and a negative

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predictive value of 80% in the diagnosis of vasculitis associated with polymyalgia rheumatica and GCA [9]. This study also demonstrated that the 18F-FDG PET scan was invaluable in early diagnosis and capable of determining the extent of disease and response to therapy.

inflammation during immunosuppressive therapy [15]. A recent study has demonstrated the value of 18F-FDG in detecting cerebral Takayasu’s arteritis in four out of five patients with Takayasu’s arteritis when the head was included in the imaging field [16].

In a study by Meller et al., 18F-FDG PET was found to better identify the extent of vasculitis when compared with MRI in 15 patients (14 with GCA and one with Takayasu’s arteritis) presenting with fever of unknown origin or elevated ESR/CRP [10]. It was also more sensitive in monitoring disease activity and response to immunosuppressive therapy than MRI, and correlated well with clinical and laboratory findings in all patients. An interesting outcome of this study was the demonstration of a higher frequency of aortic involvement in GCA compared to the published reports of 3–15%, which required a different approach to treatment to prevent life-threatening complications. The authors concluded that a traditional angiographic approach to diagnosing vasculitis in this setting would have missed the diagnosis in all cases. Others have confirmed the role of 18F-FDG PET in the diagnosis of vasculitis presenting with non-specific symptoms [11]. In fact, the high sensitivity of 18F-FDG PET in detecting vasculitis has been used as an argument to characterize polymyalgia rheumatica as a form of vasculitis [12].

Limitations of PET in vasculitis

Takayasu’s arteritis has been the subject of more studies with 18F-FDG PET than other large-vessel vasculitis. Many studies have established the pattern of 18F-FDG PETuptake in active Takayasu’s arteritis, showing mild to severely increased accumulation in larger arterial walls in a continuous linear distribution that may outline the artery as a tubular structure. The activity may also be patchy but always conforms to known arterial territories. Smaller arterial branches may not be visualized due to the resolution limits of the 18F-FDG PET scan. The degree of activity can be quantitatively measured with the standardized uptake value. However, we found that this value was not helpful in assessing disease activity [13] and this opinion is shared by others [14]. It is a very useful marker, however, in the follow-up of the same patient. Using the above criteria in a study of 18 patients with known Takayasu’s arteritis, we compared 18F-FDG PET with cross-sectional imaging, angiography and clinical scoring. We found 18F-FDG PET to have a sensitivity of 92% and specificity of 100% in identifying disease activity and confirmed its value in the management of Takayasu’s arteritis by reducing the need to perform arterial biopsies or repeat angiography [13]. Compared with MRA, MRI and angiography, 18F-FDG PET was shown to detect more lesions particularly in the early phase disease [2,13,15]. Additionally, it is non-invasive, can assess the whole body and can detect resolution of the

The most important limitation of 18F-FDG PET is the small number of PET scanners throughout the world. This unfortunate trend is being reversed with the increasing awareness of the important role of PET in the diagnosis and prognosis of a variety of diseases. General factors affecting the performance of the scanner and patients’ preparation must be considered carefully. False negative results are common due to limited spatial resolution. This problem may be resolved by delaying imaging to more than the standard 1 h post-injection to improve the target-to-background ratio. This is supported by in-vitro studies showing persistent accumulation of FDG by macrophages over 3 h [17]. Another approach to reduce false negative results is to correlate visual grading of vessel wall uptake with CRP and ESR and limiting 18F-FDG PET scans to patients with active inflammation and, if possible, after withdrawal of corticosteroid treatment [18]. However, inflammatory markers may be unreliable [13] while stopping corticosteroids may prove difficult in some patients. False positive uptake of 18F-FDG may occur within arterial walls affected by atherosclerosis and ageing but the pattern of activity is usually focal and non-linear [19]. Intense uptake in the cervical or thoracic arteries is more likely to represent vasculitis as they are less prone to atherosclerosis [20]. Sometimes correlation with other imaging such as CT/MRI or performing PET/CT can provide supportive information [14]. A decrease in the intensity of 18F-FDG uptake following corticosteroid treatment would support a diagnosis of vasculitis [20]. Uptake in mediastinal tumours and lymph nodes is another source of confusion, which may resolve by observing the size, pattern and location of these abnormalities. The increasing use of PET/CT scanners will significantly help in this respect [14]. In summary, positron emission tomography with 18F-FDG is a sensitive, specific and non-invasive diagnostic tool in large-vessel vasculitis and this applies particularly to Takayasu’s arteritis and GCA. It is far superior to clinical assessment and compares favourably with MRI, MRA and angiography in the assessment of early stages of disease. During the course of the disease, it can accurately assess disease activity and monitor the effectiveness of treatment. The facility of whole-body imaging makes it easy to

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Editorial Webb and Al-Nahhas 549

detect unsuspected vascular involvement. Its value is limited in the presence of widespread atherosclerotic disease and other conditions that may produce false negative or false positive results, but these can be minimized with appropriate measures.

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O’Doherty MJ, Barrington SF, Campbell M, Lowe J, Bradbeer CS. PET scanning and the human immunodeficiency virus-positive patient. J Nucl Med 1997; 38:1575–1583. Andrews J, Al-Nahhas A, Pennell DJ, Hossain MS, Davies KA, Haskard DO, Mason JC, et al. Non-invasive imaging in the diagnosis and management of Takayasu’s arteritis. Ann Rheum Dis 2004; 63:995–1000. Tso E, Flamm SD, White RD, Schvartzman PR, Mascha E, Hoffman GS. Takayasu arteritis: Utility and limitation of magnetic resonance imaging in diagnosis and treatment. Arthritis Rheum 2002; 46:1634–1642. Marcos HB, Choyke PL. Magnetic resonance angiography of the kidney. Semin Nephrol 2000; 20:450–455. Ishimori T, Saga T, Mamede M, Kobayashi H, Higashi T, Nakamoto Y, et al. Increased (18)F-FDG uptake in a model of inflammation: concanavalin A-mediated lymphocyte activation. J Nucl Med 2002; 43:658–663. Hiraiwa M, Nonaka C, Abe T, Iio M. Positron emission tomography in systemic lupus erythematosus: relation of cerebral vasculitis to PET findings. Am J Neuroradiol 1983; 4:541–543. Hara M, Goodman PC, Leder RA. FDG-PET finding in early-phase Takayasu arteritis. J Comput Assist Tomogr 1999; 23:16–18. Turlakow A, Yeung HW, Pui J, Macapinlac H, Lieboviz E, Rusch V, et al. Flurodeoxyglucose positron emission tomography in the diagnosis of giant cell arteritis. Arch Intern Med 2001; 161:1003–1007. Blockmans D, Stroobants S, Maes A, Mortelmans L. Positron emission tomography in giant cell arteritis and polymyalgia rheumatica: evidence for inflammation of the aortic arch. Am J Med 2000; 108:246–249.

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Meller J, Strutz F, Siefker U, Scheel A, Sahlmann CO, Lehmann K, et al. Early diagnosis and follow-up of aortitis with [18F]FDG PET and MRI. Eur J Nucl Med Mol Imaging 2003; 30:730–736. Malik IS, Harare O, AL-Nahhas A, Beatt K, Mason J. Takayasu’s arteritis: management of left main stem stenosis. Heart 2003; 89:e9. Blockmans D, Maes A, Stroobants S, Nuyts J, Bormans G, Knockaert D, et al. New arguments for a vasculitic nature of polymyalgia rheumatica using positron emission tomography. Rheumatology (Oxford) 1999; 38:444–447. Webb M, Chambers A, Al-Nahhas A, Mason JC, Maudlin L, Rahman L, Frank J. The role of 18F-FDG PET in characterising disease activity in Takayasu arteritis. Eur J Nucl Med Mol Imaging 2004; 31:627–634. Kobayashi Y, Ishii K, Oda K, Nariai T, Tanaka Y, Ishiwata K, Numano F, et al. Aortic wall inflammation due to Takayasu arteritis imaged with 18F-FDG PET co-registered with enhanced CT. J Nucl Med 2005; 46:917–922. Meller J, Grabbe E, Becker W, Vosshenrich R. Value of F-18 FDG hybrid camera PET and MRI in early Takayasu aortitis. Eur Radiol 2003; 13: 400–405. Weiner SM, Vaith P, Walker UA, Brink I. Detection of alterations in brain glucose metabolism by positron emission tomography in Takayasu’s arteritis. Eur J Nucl Med Mol Imaging 2004; 31:300–302. Deichen JT, Prante O, Gack M, Schmiedehausen K, Kuwert T. Uptake of [18F]fluorodeoxyglucose in human monocyte–macrophages in vitro. Eur J Nucl Med Mol Imaging 2003; 30:267–273. Walter MA, Melzer RA, Schindler C, Muller-Brand J, Tyndall A, Nitzsche EU. The value of [18F]FDG-PET in the diagnosis of large-vessel vasculitis and the assessment of activity and extent of disease. Eur J Nucl Med Mol Imaging 2005; 32:674–681. Yun M, Yeh D, Araujo L, Jang S, Newberg A, Alavi A. F-18 FDG uptake in the large arteries. A new observation. Clin Nucl Med 2001; 26: 314–319. Belhocine T, Blockmans D, Hustinx R, Vandevivere J, Mortelmans L. Imaging large vessel vasculitis with 18FDG PET: illusion or reality? A critical review of the literature data. Eur J Nucl Med Mol Imaging 2003; 30: 1305–1313.

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Original article

Improved quantification in 123I cardiac SPECT imaging with deconvolution of septal penetration Ji Chena, Ernest V. Garciaa, James R. Galta, Russell D. Folksa and Ignasi Carriob Objectives 123I is becoming an important radionuclide for cardiac imaging. Multiple, low-abundance, high-energy photons associated with 123I imaging can cause septal penetration in the collimators and degrade quantification of the 123I cardiac uptake. This study presents a method for the deconvolution of septal penetration (DSP) for improving quantification in 123I cardiac single photon emission computed tomography (SPECT).

using filtered back-projection were statistically significantly higher than those for planar imaging (0.2118 ± 0.0297 vs. 0.0819 ± 0.0070, P = 0.0001). 2-D DSP and 3-D DSP yielded similar HCRs that were close to the true HCR. The slopes of the regression lines for 2-D DSP and 3-D DSP were 0.9203 ± 0.0523 and 0.9101 ± 0.0304, respectively. The DSP HCRs were significantly more accurate than those calculated without DSP (P < 0.0001).

Methods Distance-dependent point spread functions were measured for low-energy high-resolution collimators on a dual-head SPECT system. The measured point spread functions were used in two-dimensional (2-D) and three-dimensional (3-D) models of the collimator response, respectively. 2-D DSP and 3-D DSP were then developed and implemented using iterative reconstruction. A cardiac torso phantom with an internal calibration source was designed with various heart-to-calibration ratios (HCRs) simulating different levels of a patient’s uptake. SPECT acquisitions of the phantom were performed using optimized acquisition and processing parameters for 123 I cardiac SPECT. HCRs were calculated using planar projection and tomographic reconstructions. The paired t-test and regression analysis were used to compare the HCRs given by different calculation methods.

Conclusion DSP significantly improves quantification in 123I cardiac SPECT imaging. 2-D DSP with its less computational burden shows promise for implementation in clinical practice so as to allow the use of the widely available low-energy, high-resolution collimators for quantitative 123I cardiac SPECT imaging. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:551–558

Results SPECT produced more accurate HCRs than planar imaging. The slopes of the regression lines for SPECT

Introduction 123

With increasing global use of m-[ I]iodobenzylguanidine (123I-MIBG) and 15-(p-[123I]iodophenyl)-3-(R,S)methylpentadecanoic acid (123I-BMIPP), 123I is becoming an important radionuclide for cardiac imaging. In 123I cardiac imaging it is important to quantify the cardiac uptake of the tracer. The heart-to-mediastinum ratio, the ratio of the count density in the left ventricle to that in the upper mediastinum, has been widely used as a quantitative index of cardiac 123I-MIBG uptake, and consequently, cardiac sympathetic function [1,2]. The heart-to-mediastinum ratio can be calculated from either planar images or single photon emission computed tomography (SPECT) images [1,2]. It has also been demonstrated that the degree of reduction of regional cardiac 123I-MIBG uptake positively correlated to markers of severity of heart failure and partially correlated to leftventricular regional wall motion and resting myocardial

Nuclear Medicine Communications 2006, 27:551–558 Keywords:

123

I imaging, cardiac SPECT, quantification, septal penetration

a Emory University School of Medicine, Atlanta, USA and bHospital Sant Pau, Barcelona, Spain.

Correspondence to Dr Ji Chen, Department of Radiology, Emory University School of Medicine, 1364 Clifton Road, Atlanta, GA, 30322, USA. Tel: + 001 (404) 712 4024; fax: + 001 (404) 712 7961; e-mail: [email protected] Received 12 December 2005 Accepted 8 March 2006

perfusion abnormalities [3,4]. 123I-BMIPP has been used for assessment of cardiac metabolism. Clinical studies have shown that decreased 123I-BMIPP uptake is related to wall motion abnormality and severe stenosis [5–7]. Thus, acquisition and processing techniques with accurate quantification of cardiac 123I uptake are very important for 123I cardiac imaging. Investigations have shown that the multiple low-abundance high-energy photons in addition to the 159-keV photons from the main 123I photopeak can penetrate the septa of the collimators and degrade the image quantification [8–13]. Imaging these high-energy photons with a scintillation camera on a low-energy collimator results in a 123I spectrum with an extensive high-energy plateau (Fig. 1). This phenomenon raises serious concern that the widely used low-energy high-resolution (LEHR) collimators may not be suitable for 123I cardiac imaging, since

c 2006 Lippincott Williams & Wilkins 0143-3636

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552 Nuclear Medicine Communications 2006, Vol 27 No 7

high-energy photons tend to penetrate the septa of LEHR collimators and degrade the image quality and quantitative accuracy [8–13]. It has been shown that collimator choice substantially influences the estimation of heart-to-mediastinum ratio in 123I cardiac imaging and suggests the use of medium-energy all-purpose collimators for higher quantitative accuracy [14–16]. However, most of the commercial SPECT systems, especially the dedicated small field-of-view cardiac SPECT systems, are currently not equipped with such collimators.

This article presents a deconvolution of septal penetration (DSP) technique, which models collimator response and is implemented using ordered-subsets expectation maximization (OSEM). This technique is applied to a series of phantom experiments to demonstrate its capability to reduce the impact of septal penetration in LEHR collimators and to improve quantification of 123I cardiac uptake.

Materials and methods Measurement of point-spread functions

A GE Millennium VG/Hawkeye SPECT/CT system (Milwaukee, Wisconsin) was used in this study. A point source with approximately 37 MBq (1 mCi) of 123I was imaged with the LEHR collimators of the system at 0 cm (closest), then at 2, 4, 8, 16, 24 and 32 cm away from the camera surface. Each image was acquired for 300 000 counts.

Fig. 1

(a) 32000 26666

cts

21333

A three-dimensional (3-D) PSF was extracted from the acquired data. Linear interpolation was applied to calculate the PSFs for different source-to-camera distances. Then, a two-dimensional (2-D) PSF was approximated from the 3-D PSF to represent the collimator response at a source-to-camera distance of 8 cm, similar to the distance between the heart and the camera.

16000 10666 5333 Kev

0 50

162

274

386

Window Peak Offset Width Lower Upper Set kev kev kev −> 1 159 0 % 15 % 148 172 1

498 Count rate cps

(b) 32000 26666

cts

21333 16000 10666 5333 0

Kev 50

162

274

386

Window Peak Offset Width Lower Upper Set kev kev kev −> 1 159 0 % 15 % 148 172 1

498 Count rate cps

^{123I spectra measured by a single-head camera intrinsically (a) and extrinsically with a low-energy collimator (b). The window placed on the spectra was the 15% photopeak window centred at 159 keV. The relative height of the plateau in the extrinsic spectrum was dramatically higher than that in the intrinsic spectrum due to septal penetration in the collimator.

Deconvolution of septal penetration

Modelling of collimator response has been incorporated into iterative reconstruction for compensation for collimator resolution in cardiac SPECT imaging [17–22] and shown to improve the defect contrast by correcting the blurring effect resulting from the collimator response [23,24]. A similar algorithm was developed in this study for compensation for septal penetration since the measured PSF associated with 123I imaging included both collimator resolution and septal penetration. Figure 2 illustrates iterative reconstruction with DSP. This process started from an initial guess image. A projector, which included 2-D or 3-D convolution of the guess image with the 2-D or 3-D PSF respectively, was used to produce a projection image. Photon attenuation could also be included in the projector to perform attenuation correction. The computed projection image was then compared to the measured projection image, and the back-projection of the error was used to update the guess image. This iterative process was stopped after a preset total number of iterations. Since the measured 123I PSF included septal penetration, this effect would be deconvolved after a number of iterations, and its impact reduced in the reconstructed image. Phantom experiments

A data spectrum anthropomorphic torso phantom with heart, myocardial defect, liver, lung and spine components was used in this study. A calibration bottle (140 ml)

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I cardiac SPECT Chen et al. 553

Fig. 2

Convolution with PSF Initial guess image

New guess image

Project

No

Stop?

Computed projection image Measured projection image

Yes

Output image

Update guess image

Error in projection

Compare

Back project

Error in reconstruction

Iterative reconstruction with deconvolution of septal penetration (DSP). Since in the projector the guess image was convolved with a point-spread function (PSF), which included septal penetration, this effect would be deconvolved after a number of iterations and its impact reduced in the reconstructed image. 2-D and 3-D convolutions were used for 2-D DSP and 3-D DSP, respectively.

–1 Table 1 Compartmental concentrations (in kBq
ml ) of the phantom experiments

Compartment

Heart Defect Liver Calibration Heart-to-calibration ratio*

Experiment A

B

C

D

74.9 N/A 74.7 11.3 6.6

113.2 55.5 19.6 11.2 10.1

222.0 111.0 15.6 15.6 14.2

222.0 111.0 9.5 9.5 23.4

* The heart-to-calibration ratio was the heart concentration divided by the calibration concentration.

with the same concentration as in the background was placed inside the phantom for measurement of the true heart-to-calibration ratio (HCR). The use of the internal calibration bottle allowed more accurate measurement of the true HCR since it was difficult to uniformly distribute the radioactivity in the background of large volume. Table 1 lists the concentrations in the heart, defect, liver and calibration bottle of the phantom experiments. The phantom was imaged with the GE Millennium VG/ Hawkeye SPECT/CT system using a set of optimized acquisition and processing parameters for 123I cardiac SPECT imaging [25]: 1. 15% photopeak window centred at 159 keV, 2. 28% scatter window centred at 128 keV, 3. Energy-window subtraction [26] performed using a scatter compensation scaling factor of 0.3, 4. 1801 acquisition, non-circular orbit, 5. 60 projections, 25 s per projection,

6. X-ray computed tomography transmission scans obtained for attenuation correction (140 kVp, 2.5 mA, 1 cm per slice, four slices per minute, 25 slices in total), 7. 2-D Butterworth pre-filtering with a critical frequency of 0.5 Nyquist and an order of 5 for filtered backprojection (FBP), 8. 3-D Butterworth post-filtering with a critical frequency of 0.5 Nyquist and an order of 5 for OSEM, 9. Three iterations and six subsets of 10 angles each for OSEM.

HCRs were calculated from the photopeak-window projections and the scatter-compensated projections respectively. Regions of interest (ROIs) were placed on the cardiac insert and the calibration bottle to calculate their maximum counts. HCR was defined as the maximum counts in the heart divided by the maximum counts in the calibration bottle. Maximum counts were used instead of average counts in order to reduce the error introduced by sub-optimal ROI definitions. HCRs were also calculated from tomographic reconstructions using a similar ROI analysis. The methods used for tomographic reconstructions were: (1) FBP, (2) FBP with scatter compensation, (3) OSEM, (4) OSEM with 2-D DSP, (5) OSEM with 3-D DSP, (6) OSEM with attenuation correction, (7) OSEM with attenuation correction and 2-D DSP, and (8) OSEM with attenuation correction and 3-D DSP. All of the OSEM reconstructions were performed only on the scatter compensated datasets. Note that the internal calibration bottle was invisible in planar projections and emission reconstructions since it

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had approximately the same concentration of the tracer as background. Nevertheless, the internal calibration bottle was clearly visible in the transmission reconstruction, and its location guided the placement of ROI for the internal calibration bottle in the planar projections and emission reconstructions.

Results The 123I PSFs from the LEHR collimator measured at various distances are shown in Fig. 3. As the camera moved away from the point source, the peak counts decreased and the widths of the PSFs increased. The plateaus resulting from septal penetration existed in all of the PSFs, and their relative heights did not change remarkably with the camera–source distance. Table 2 compares the HCRs calculated using the photopeak-window planar projections to those calculated using the scatter-compensated planar projections. The Fig. 3

10000 Closest d = 8 cm PSF (counts)

1000

d = 16 cm d = 32 cm

100

10

1 0

50

100

150 r (mm)

200

250

300

Closest d = 8 cm Normalized PSF

Figure 4 shows the regression analysis of the HCRs calculated by the planar projection and FBP reconstructions in all of experiments. SPECT yielded statistically significantly higher slopes of the regression lines than planar projection, indicating that SPECT produced better quantification. Figure 5 compares the HCRs calculated by the OSEM reconstructions without and with DSP. The DSP HCRs, 2-D or 3-D, without or with attenuation correction, were statistically significantly closer to the true HCRs (regression slopes were close to 1) than those measured without DSP. 2-D DSP and 3D DSP yielded similar HCRs, shown in Tables 4 and 5, confirming the observation of the PSFs that septal penetration was insignificantly related to the source-tocamera distance. Since 2-D DSP can produce HCR as accurate as 3-D DSP, it should be considered more suitable for clinical implementation since it takes far less computation time. Some phantom images obtained in experiment D (abnormal myocardium, low liver uptake) are shown in Fig. 6. Compared to the images obtained without DSP (Fig. 6(a)), DSP cleared the background activity and improved the defect contrast (Fig. 6(b)). Figure 7 shows the phantom images given by experiment B (normal myocardium, high liver uptake). DSP reduced the myocardium–liver overlap and made the inferior wall more interpretable (Fig. 7(b)).

1

0.1

paired t-test showed that scatter compensation improved HCR when there were not much extra-cardiac activity (experiments C and D). When there was high extracardiac activity (experiments A and B), septal penetration from those activities raised the background counts and decreased the HCR. Scatter compensation could not address this effect so that there were no statistically significant differences between the HCRs given by the photopeak-window planar projections and the HCRs given by the scatter-compensated planar projections. A similar trend can be observed in Table 3, which compares the HCRs given by the FBP reconstructions of the photopeak-window data and the scatter-compensated data. More extra-cardiac activity (experiment A) resulted in less improvement obtained with scatter compensation. Nevertheless, scatter compensation did improve the HCRs in all of the experiments as shown in Table 3.

d = 16 cm d = 32 cm

0.01

0.001

Discussion 0.0001

0

50

100

150 r (mm)

200

250

300

Non-normalized and normalized 123I point spread functions (PSFs) of the low-energy high-resolution collimator at various distances. In the figure, d (in cm) is the distance between the camera surface and the point source, and r (in mm) is the distance in the camera surface from the point source position.

This article presents a DSP technique that was based on 2-D or 3-D modelling of collimator response. This process was built in iterative reconstruction with a projector including 2-D or 3-D convolution of the guess image with measured distance-dependent PSF. Since the PSF included septal penetration, the results of iterative reconstruction would have this effect deconvolved. A typical problem associated with 3-D modelling collimator

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Deconvolution of septal penetration for

Table 2

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I cardiac SPECT Chen et al. 555

Heart-to-calibration ratios measured with planar projection Experiment A

Maximum Minimum Mean STD*** Pw

B

C

D

PP*

SC**

PP

SC

PP

SC

PP

SC

2.17 1.53 1.82 0.22

2.13 1.62 1.83 0.21

2.51 1.79 2.15 0.27

2.52 1.84 2.17 0.25

2.90 2.31 2.64 0.19

3.64 2.69 3.18 0.31

3.30 2.97 3.20 0.12

3.86 3.16 3.59 0.26

0.997

0.564

0.029

0.038

*

PP: heart-to-calibration ratios (HCRs) in this column were calculated using the photopeak-window projections. SC: HCRs in this column were calculated using the scatter-compensated projections. The scatter compensation was done by scaling the scatter-window projections with 0.3 and then subtracting it from the photopeak-window projections. *** STD: numbers in this row were the standard deviations of the measured HCRs. w P: the P values in this row were given by paired t-test, comparing between the PP and SC HCRs. The P values less than 0.05 were considered statistically significant. **

Table 3

Heart-to-calibration ratios measured with filtered back-projection Experiment A

Maximum Minimum Mean STD*** Pw

B

C

D

PP*

SC**

PP

SC

PP

SC

PP

SC

3.90 3.54 3.70 0.16

4.49 3.83 4.13 0.29

4.84 3.85 4.52 0.39

5.54 4.31 5.07 0.45

7.57 6.36 7.19 0.46

8.48 6.89 7.92 0.57

7.36 6.88 7.16 0.21

9.62 8.85 9.22 0.27

0.036

< 0.001

< 0.001

< 0.001

*

PP: heart-to-calibration ratios (HCRs) in this column were calculated using the filtered back-projection (FBP) reconstruction of the photopeak-window projections. HCRs in this column were calculated using the FBP reconstruction of the scatter compensated projections. The scatter compensation was done by scaling the scatterwindow projections with 0.3 and then subtracting it from the photopeak-window projections. *** Numbers in this row were the standard deviations of the measured HCRs. w P: the P values in this row were given by the paired t-test, comparing between the PP and SC HCRs. The P values less than 0.05 were considered statistically significant. **

Fig. 4

8

10

Planar FBP

8 Measured HCR

Measured HCR

6

4

2

0

Planar FBP

6 4 2

5

10

15

20

25

True HCR

0

5

10

15

20

25

True HCR

Comparison of the heart-to-calibration ratios (HCRs) between planar projection and filtered back-projection (FBP). The left panel is with the photopeak-window data. The slopes of the regression lines are 0.0819 ± 0.0070 for planar projection and 0.2118 ± 0.0297 for FBP (P = 0.0001). The right panel is with the scatter-compensated data. The slopes of the regression lines are 0.1079 ± 0.0115 for planar projection and 0.3121 ± 0.0259 for FBP (P < 0.0001).

response in iterative reconstruction is the immense increase in computational burden. Several computational enhancements have been utilized in the developed DSP technique. First, the DSP technique can be implemented in 2-D vastly reducing computational burden. Since

septal penetration changed slightly with the source-tocamera distance (shown in Fig. 3), 2-D DSP has shown to yield similar quantification as 3-D DSP. Second, OSEM instead of maximum likelihood expectation maximization (MLEM) was used to accelerate the reconstruction.

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Fig. 5

25

25

FBPSC OSEM OSEM+2DDSP

15 10 5 0

OSEM+AC OSEM+2DDSP+AC

20 Measured HCR

Measured HCR

20

OSEM

15 10 5

5

10

15 True HCR

20

0

25

5

10

15 True HCR

20

25

Comparison of the heart-to-calibration ratios (HCRs) without and with deconvolution of septal penetration (DSP). The slopes of the regression lines in the left panel are 0.3121 ± 0.0259 for FBP with scatter compensation (SC), 0.4275 ± 0.0289 for ordered-subsets expectation maximization (OSEM), and 0.9203 ± 0.0523 for OSEM with 2-D DSP (OSEM vs. FBP SC: P = 0.0048, OSEM + 2-D DSP vs. OSEM: P < 0.0001). The slopes of the regression lines in the right panel are 0.4275 ± 0.0289 for OSEM, 0.5015 ± 0.0253 for OSEM with attenuation correction (AC), and 0.9101 ± 0.0304 for OSEM with 2-D DSP and attenuation correction (OSEM vs. OSEM + AC: P = 0.0604, OSEM + 2-D DSP + AC vs. OSEM + AC: P < 0.0001). Note that all of the OSEM reconstructions were with the same scatter compensation as that in FBP scatter correction.

Table 4

Heart-to-calibration ratios measured with deconvolution of septal penetration Experiment A 2-D

Maximum Minimum Mean STD*** Pw

*

B 3-D

8.21 6.41 7.03 0.78

**

7.60 5.97 6.54 0.71

C

D

2-D

3-D

2-D

3-D

2-D

3-D

11.35 8.56 9.89 1.03

11.21 8.31 9.54 1.23

17.23 12.84 15.21 1.68

16.84 12.04 14.56 1.75

25.49 19.62 22.22 2.27

25.65 19.12 22.15 2.51

0.236

0.609

0.529

0.961

*

2-D: heart-to-calibration ratios (HCRs) in this column were calculated using the two-dimensional deconvolution of septal penetration (DSP). ** 3-D: heart-to-calibration ratios (HCRs) in this column were calculated using the three-dimensional DSP. *** STD: numbers in this row were the standard deviations of the measured HCRs. w P: the P values in this row were given by paired t-test, comparing between the 2-D DSP and 3-D DSP HCRs. The P values less than 0.05 were considered statistically significant.

Table 5

Heart-to-calibration ratios measured with deconvolution of septal penetration and attenuation correction Experiment A 2-D

Maximum Minimum Mean STD*** Pw

*

B 3-D

7.16 5.72 6.18 0.51

**

7.57 5.96 6.66 0.54 0.143

C

D

2-D

3-D

2-D

3-D

2-D

3-D

10.95 9.49 10.17 0.54

10.50 8.54 9.57 0.77

14.75 11.71 13.35 1.06

15.67 11.99 13.21 1.40

23.45 19.86 21.74 1.33

23.50 19.95 22.29 1.30

0.144

0.848

0.483

*

2-D: heart-to-calibration ratios (HCRs) in this column were calculated using the two-dimensional deconvolution of septal penetration (DSP) and attenuation correction (AC). ** 3-D: heart-to-calibration ratios (HCRs) in this column were calculated using the three-dimensional DSP and AC. *** STD: numbers in this row were the standard deviations of the measured HCRs. w P: the P values in this row were given by paired t-test, comparing between the 2-D DSP + AC and 3-D DSP + AC HCRs. The P values less than 0.05 were considered statistically significant.

Third, the DSP technique only modelled collimator response in the projection step and saved a great number of computations. The valid condition of using unmatched projector and back-projector in iterative reconstruction

has been reported [27]. It has been pointed out that it is less critical in practice if the algorithm stops within the early convergence trend of the algorithm [27]. Care must be taken in choosing reconstruction parameters for this

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Deconvolution of septal penetration for

123

I cardiac SPECT Chen et al. 557

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Example phantom images of experiment D (abnormal myocardium, low liver uptake). Compared to the images obtained without DSP (a), DSP cleared the background activity and improved the defect contrast (b). (Abbreviations as in the legend to Fig. 5).

algorithm. A large number of iterations would lead this algorithm to divergence and magnify noise in the image. This algorithm was tested using a PC with Pentium-4 2.4 GHz CPU and 512 MB RAM. With OSEM (three iterations, six subsets) the reconstruction time for a 64  64  64 image was about 2 min for 2-D DSP. Four phantom experiments were presented in this article with different levels of HCR and extra-cardiac activity. Since the tail of the PSF is quite long because of septal penetration, extra-cardiac activity such as that in the liver would increase the background counts so as to reduce the HCR. This effect was clearly shown by the phantom experiments. The HCRs given by planar projections and tomographic reconstructions without DSP were significantly lower than the true HCRs. Scatter compensation did not help quantification in this situation, since it mostly dealt with the scatter inside the patient body, not the interactions inside the collimator. A similar finding has been reported in a recent publication using the tripleenergy window scatter compensation [16]. The devel-

Example phantom images of experiment B (normal myocardium, high liver uptake). Compared to the images obtained without DSP (a), DSP cleared the background activity, reduced the myocardium-liver overlap, and made the inferior wall more interpretable (b). (Abbreviations as in the legend to Fig. 5).

oped DSP technique was based on the camera-specific PSFs that were measured in air. The degradation to the point images resulted from distance-dependent collimator resolution (increase in the width of the PSFs with the source-to-camera distance) and septal penetration (long plateau, little dependence on the source-to-camera distance). Both 2-D DSP and 3-D DSP, with or without attenuation correction, properly compensated for septal penetration so as to significantly improve quantification and yield the HCRs close to the true HCRs. This finding shows a promise of using the widely available LEHR collimators with the DSP technique for quantification of 123 I cardiac uptake.

Conclusion A DSP technique based on 2-D or 3-D modelling of collimator response to 123I has been developed. It can essentially compensate for septal penetration due to the high-energy photons in 123I cardiac SPECT imaging. This technique shows a promise to allow the use of the widely

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Nuclear Medicine Communications 2006, Vol 27 No 7

available LEHR collimators for quantification of 123I cardiac uptake and broaden the use of the 123I tracers for cardiac imaging.

References 1 2 3

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13

Patel AD, Iskandrian AE. mIBG imaging. J Nucl Cardiol 2002; 9:75–94. Hattori N, Schwaiger M. Metaiodobenzylguanidine scintigraphy of the heart: what have we learnt clinically? Eur J Nucl Med 2000; 27:1–6. Imamura Y, Ando H, Mitsuoka W, Egashira S, Masaki H, Ashihara T, Fukuyama T, et al. Iodine-123 metaiodobenzylguanidine images reflect intense myocardial adrenergic nervous activity in congestive heart failure independent of underlying cause. J Am Coll Cardiol 1995; 26:1594–1599. Parthenakis FI, Prassopoulos VK, Koukouraki SI, et al. Segmental pattern of myocardial sympathetic denervation in idiopathic dilated cardiomyopathy: relationship to regional wall motion and myocardial perfusion abnormalities. J Nucl Cardiol 2002; 9:15–22. Tamaki N, Kawamoto M, Yonekura Y, et al. Regional metabolic abnormality in relation to perfusion and wall motion in patients with myocardial infarction: assessment with emission tomography using iodinated branched fatty acid analog. J Nucl Med 1992; 33:659–667. Franken PR, De Geeter F, Dendale P, et al. Abnormal free fatty acid uptake in subacute myocardial infarction after coronary thrombolysis: correlation with wall motion and inotropic reserve. J Nucl Med 1994; 35:1758–1765. Tateno M, Tamaki N, Yukihiro M, Kudoh T, Hattori N, Tadamura E, et al. Assessment of fatty acid uptake in ischemic heart disease without myocardial infarction. J Nucl Med 1996; 37:1981–1985. Dobbeleir AA, Hambye AS, Franken PR. Influence of high-energy photons on the spectrum of iodine-123 with low- and medium-energy collimators: consequence for imaging with 123I-labelled compounds in clinical practice. Eur J Nucl Med 1999; 26:655–658. Macey DJ, DeNardo GL, DeNardo SJ, Hines HH. Comparison of low- and medium-energy collimators for SPECT imaging with iodine-123-labeled antibodies. J Nucl Med 1986; 27:1467–1474. Geeter F, Franken PR, Defrise M, Andries H, Saelens E, Bossuyt A, et al. Optimal collimator choice for sequential iodine-123 and technetium-99m imaging. Eur J Nucl Med 1996; 23:768–774. Fleming JS, Alaamer AS. Influence of collimator characteristics on quantification in SPECT. J Nucl Med 1996; 37:1832–1836. Dobbeleir AA, Hambye ASE, Franken PR. Influence of methodology on the presence and extent of mismatching between 99mTc-MIBI and 123I-BMIPP in myocardial viability studies. J Nucl Med 1999; 40:707–714. Bolmsjo MS, Persson BR, Strand SE. Imaging 123I with a scintillation camera. A study of detection performance and quality factor concepts. Phys Med Biol 1977; 22:266–277.

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Inoue Y, Suzuki A, Shirouzu I, Machida T, Yoshizawa Y, Akita F, et al. Effect of collimator choice on quantitative assessment of cardiac iodine 123I-mIBG uptake. J Nucl Cardiol 2003; 10:623–632. Inoue Y, Shirouzu I, Machida T, Yoshizawa Y, Akita F, Minami M, Ohtomo K, et al. Collimator choice in cardiac SPECT with I-123-labeled tracers. J Nucl Cardiol 2004; 11:433–439. Verberne HJ, Feenstra C, de Jong WM, Somsen GA, van Eck-Smit BLF, Sokole EB. Influence of collimator choice and simulated clinical conditions on 123I-MIBG heart/mediastinum ratios: a phantom study. Eur J Nucl Med Mol Imaging 2005; 32:1100–1107. Tsui BMW, Hu GB, Gilland DR. Implementation of simultaneous attenuation and detector response correction in SPECT. IEEE Trans Nucl Sci 1988; 35:778–783. Floyd CE, Jaszczak RJ, Manglos SH, Coeman RE. Compensation for collimator divergence in SPECT using inverse Monte Carlo reconstruction. IEEE Trans Nucl Sci 1988; 32:769–785. Formiconi AR, Pupi A, Passeri A. Compensation of spatial response in SPECT with conjugate gradient reconstruction technique. Phys Med Biol 1989; 34:69–84. Penny BC, King MA, Knesaurek K. A projector, backprojector pair which accounts for the two-dimensional depth and distance dependent blurring in SPECT. IEEE Trans Nucl Sci 1990; 37: 681–686. Zeng GL, Gullberg GT, Tsui BMW, Terry JA, et al. Three-dimensional iterative reconstruction algorithms with attenuation and geometric point response correction. IEEE Trans Nucl Sci 1991; 38:693–702. Beekman FJ, Eijkman EGJ, Viergever MA, Born GF, Slijpen ETP. Object shape dependent PSF model for SPECT imaging. IEEE Trans Nucl Sci 1993; 40:31–39. Kohli V, King MA, Pan TS, Glick SJ. Compensation for distancedependent resolution in cardiac-perfusion SPECT: Impact on uniformity of wall counts and wall thickness. IEEE Trans Nucl Sci 1998; 45: 1104–1110. Hutton BF, Lau YH. Application of distance-dependent resolution compensation and post-reconstruction filtering for myocardial SPECT. Phys Med Biol 1998; 43:1679–1693. Chen J, Garcia EV, Galt JR, Folks RD, Carrio I. Optimized acquisition and processing protocols for I-123 cardiac SPECT imaging. J Nucl Cardiol 2006; 13:251–260. Jaszczak RJ, Greer KL, Floyd Jr CE, Harris CC, Coleman RE. Improved SPECT quantitation using compensation for scattered photons. J Nucl Med 1984; 25:893–899. Zeng GL, Gullberg GT. Unmatched projector/backprojector pairs in an iterative reconstruction algorithm. IEEE Trans Med Imag 2000; 19: 548–555.

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Original article

Optimized radioiodine therapy for Graves’ disease: Two MIRD-based models for the computation of patient-specific therapeutic 131I activity Thomas Carliera, Pierre-Yves Salaunb, Marie-Be´atrice Cavarecb, Fre´de´ric Valettea, Alexandre Turzob, Manuel Bardie`sc, Yves Bizaisb and Olivier Couturiera Aim 131I therapy is increasingly used for Graves’ hyperthyroidism. Debate remains about the best method for calculating the activity to administer, as well as about the potential benefit of such computed activity. Several arguments plead, nevertheless, in favour of a personalized computation, such as inter-individual variations of thyroid volume and biokinetics. Methods A MIRD-based dosimetric approach, with an additional extension that takes into account the variation of thyroid mass during the treatment, has been developed. This approach includes the benefits of a personalized determination of biokinetics. Results were compared with those of six methods widely used in routine practice. Forty-one patients were enrolled (34 women, seven men; mean age ± SD: 48.11 ± 6.4 years). 131I uptakes were measured at 4, 24 and 96 h (36.2 ± 14.6%, 42.8 ± 9.7% and 27.6 ± 6.8%, respectively), following administration of the tracer. The kinetics of iodine in the thyroid were evaluated using a two-compartment model (effective half-life of 5.1 ± 1.6 days). Computations of activities to deliver the doses prescribed by the physician were done with the eight formalisms. Results There was no statistical difference between results of the two MIRD-based formalisms (227 ± 148 MBq and

Introduction Surgery and 131I are the two classical therapeutic options for relapsed Graves’ disease, and in many industrial countries, radioiodine has largely replaced surgery because of its safety, convenience and effectiveness. However, there is still controversy about the choice of the best therapeutic 131I regimen, between an empirical, fixed activity and an activity calculated with a dosimetric method. Factors may influence the outcome of radioiodine treatment, such as the importance of thyroid goitre volume, severity of hyperthyroidism, combined antithyroid drug regimen, and individual target tissue sensitivity. Thus, it is not surprising that several studies have reported conflicting results about the ability to predict treatment failure or success with dosimetric methods [1–4]. Likewise, despite numerous methods, it

213 ± 124 MBq), which were also not significantly different from those obtained with the majority of the other methods (from 128 ± 95 MBq to 275 ± 223 MBq). However, a large intra-individual difference up to a factor of 2 between two given methods was found. Conclusion The formalism developed appears to be a good compromise between all the common formalisms already used in many institutions. Furthermore, it allows the exposures of target volumes and non-target volumes to be planned individually and practical individual radiation protection recommendations to be implemented. Nucl c 2006 Lippincott Williams Med Commun 27:559–566
& Wilkins. Nuclear Medicine Communications 2006, 27:559–566 Keywords:

131

I, Graves’ disease, therapy, MIRD-based models

a Nuclear Medicine Department, Nantes University Hospital, bNuclear Medicine Department, Brest University Hospital and cINSERM U601, Nantes, France.

Correspondence to Dr Olivier Couturier, University Hospital of Nantes, Hotel Dieu-Department of Nuclear Medicine, Place Alexis Ricordeau, 44093 Nantes, France. Tel: + 0033 (0)2 40 08 41 36; fax: + 0033 (0)2 40 08 42 18; e-mail: [email protected] Received 24 January 2006 Accepted 30 March 2006

remains impossible to accurately determine the optimal 131 I activity to cure hyperthyroidism while avoiding a permanent secondary hypothyroidism. However, hypothyroidism is not regarded as a serious complication, because of the effective use of thyroid hormone replacement, and even corresponds to the natural longterm evolution of Graves’ disease, irrespective of the amount of radioactivity delivered [5,6]. Nevertheless, the administration of a fixed activity may lead to too much or too little radioactivity being delivered; that is, unnecessary exposure for the patient and for the environment or, conversely, insufficient 131I to effect a cure. Furthermore, because younger patients are now offered radioiodine therapy earlier in the course of their disease, concerns about the long-term effects of

c 2006 Lippincott Williams & Wilkins 0143-3636


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560 Nuclear Medicine Communications 2006, Vol 27 No 7

radioiodine and thyroid hormone replacement therapy suggest the need of caution with large ablative doses of 131 I. Therefore, the optimal approach should be the administration of a computed therapeutic activity that takes into account individual parameters, such as gland size, iodine uptake and iodine turnover. A few studies emphasize that the absorbed dose to the thyroid is related to treatment outcome [7,8] and that only personalized dosimetry is able to take into account some particular cases where non-optimized dosimetry can be inefficient and/or hazardous [9]. However, the choice of the most appropriate dosimetric methodology to calculate therapeutic 131I activity, according to a desired dose to the thyroid, is still highly controversial [7]. Thus, it could be useful to develop a standardized method where the latest advances in internal dosimetry and individual biokinetics are taken into account. An important point must be the reproducibility and the applicability in routine work in a wide range of nuclear medicine departments. In the present study, a formalism based on the MIRD methodology was developed as personalized dosimetry. One of the requirements was the facility to implement it in clinical routine. Results obtained with this new MIRD-based approach were also compared with those derived from other methods widely used in clinical practice.

Patients, materials and methods Patients

This study involved two institutions: the nuclear medicine departments of Brest and Nantes University hospitals. A total of 41 consecutive patients (48.1 ± 16.4 years) suffering from Graves’ disease were included (34 women and seven men). Initial (n = 34) or relapse (n = 7) Graves’ thyrotoxicosis were diagnosed on the basis of clinical findings and laboratory tests showing high value of free thyroxine and low levels of thyroid stimulating hormone for more than 3 months. Relapses occurred after at least 18 months of antithyroid drug treatment. All patients were instructed to stop the antithyroid medication 7 days before undergoing 131I uptake measurements. Uptake measurements methodology and experimental set-up 131

The patients received a capsule of I (2 MBq) as an oral test activity. The 131I capsule was measured at t = 0 and further thyroid uptake measurements were performed after 4, 24 and 96 h. Measurements were performed with a 200  200 NaI(Tl) detector connected to a multichannel analyser. The energy window was set to 312–435 keV. Measurements were corrected from room and patients’ background. They were performed at a constant distance (25 cm)

between the detector and the thyroid (or the capsule for the first measurement). Three independent acquisitions (by displacing and replacing the probe) were done for each counting. The thyroid uptake was set to be the mean count of the three measurements. Additionally, all measurements were performed by the same technologist of each institution in order to minimize the daily probe mispositioning. The detector sensitivity was checked with a 133Ba source each day prior to patient investigations and a correction factor was applied if a counting deviation > 5% was noted; that is, if the ratio between the first and the following 133Ba measurements was > 1.05 or < 0.95, the uptake value was corrected by multiplying the measurement with this ratio. Measurement of thyroid mass

The thyroid mass was evaluated by using two-dimensional ultrasonography methodology. The ellipsoid approximation based on height (h), width (w) and depth (d) of each thyroid lobe was employed. Assuming a density of 1 kgcm – 3, the thyroid mass is given by the relation p m ¼ hwd : 6 Basic theory of the methods used for computing the activity to be delivered

Eight activity computations were chosen: three are based on the formula given by Marinelli et al. [10] (so-called Marinelli 1, Marinelli 2 and Marinelli 3), two were described by Leslie et al. [1] (so-called Leslie 1 and Leslie 2), one was based on the original work of Bockisch et al. [11] and one was developed in this study with an additional extension which takes into account the mass variation during the treatment (approach 1 and approach 2). The two last methods used modelling of the thyroid biokinetics and the MIRD formalism. It should be emphasized that the Leslie 1 and 2 methods do not incorporate information regarding the dose delivered to the thyroid. These methods are only based on a fixed activity administration weighted by the thyroid mass and the 24 h thyroid uptake. Marinelli computation The Marinelli 1 method

The activity to be administered is given by the equation mD ð1Þ A¼ 73:8EFðt ¼ 0ÞTeff in which A is the activity to be administered (in mCi), m is the thyroid mass (in g), D is the absorbed dose (in cGy) delivered to the thyroid, F(t = 0) is the thyroid uptake (as a percentage) extrapolated to time t = 0, Teff the effective half-life (in days), E is the mean electron energy (E = 0.195 MeV) and 1/73.8 is the factor which puts the equation into units of microcuries. The value of F(t = 0) is deduced by linear regression over all thyroid uptake

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MIRD-based models for computing

points corrected from radioactive decay. Teff is computed from an exponential fit including the experimental thyroid uptake (non-corrected from radioactive decay) from 24 to 96 h. Only the electron energy loss is taken into account in Equation 1. Marinelli assumed that the photon contribution was no more than 10% of the electron contribution. This assertion is added in our calculation (D = Delectron + Dphoton = Delectron + 0.1Delectron). The Marinelli 2 method

The only difference between the Marinelli 2 and the Marinelli 1 methods is the determination of the effective half-life. A¼

23:4mD Fðt ¼ 0ÞTeff

ð2Þ

where 23.4 is the factor that puts the equation into units of MBq, Teff is equal to 5 days if F(t = 4)F(t = 24) and 4 days if F(t = 4) r F(t = 24), where F(t = 4) and F(t = 24) are the thyroid uptake measured at 4 h and 24 h, respectively, after oral administration. The Marinelli 3 method

In this method, the activity is given by the equation A¼

23:4mD Fðt ¼ 0ÞTeff

ð3Þ

where 23.4 is a factor that puts the equation into units of MBq, F(t = 24) is the thyroid uptake at t = 24 h (corrected from the radioactive decay) and Teff is equal to a fixed value (5 days). The Leslie method

Two of the original propositions by Leslie et al. are retained [1]. This calculation is based on the administration of a fixed activity weighted by the thyroid uptake at 24 h (F(t = 24)) and the thyroid mass. The first formula is defined as ‘low-adjusted’ (Leslie 1): A¼

2:96m : Fðt ¼ 24Þ

The second formula is defined as ‘high-adjusted’ (Leslie 2): A¼

4:44m Fðt ¼ 24Þ

in which 2.96 and 4.44 are two constants expressed in MBq  g – 1 and F(t = 24) is the thyroid uptake at 24 h corrected from the radioactive decay. The Bockisch method

This formalism is based on the thyroid uptake measurement at 96 h [11]. The activity to be administered is given by the relation

131

I activity in Graves’ disease Carlier et al. 561

A¼

2:46mD Fðt ¼ 96Þ

where 2.46 is a constant expressed in MBq  g – 1  Gy – 1 and defined over a population of 175 patients [11], and F(t = 96) is the thyroid uptake at 96 h without a correction for radioactive decay. Two-compartment modelling associated with the MIRD formalism

The average dose delivered to a target due to activity embedded in a source region D
target source is given by the well-known relation ð4Þ D
target source ¼ A~source Starget source where A~source is the cumulated activity in the source, and Starget’source is the S factor (i.e., the dose per unit of cumulated activity). We assumed that the absorbed dose in the thyroid was only the consequence of the radioiodine uptake (there was no contribution from other possible non-target organs). Besides, we supposed a uniform distribution of 131 I in the thyroid. In this case, the source and the target are identical; therefore the S factor is noted as S. Two S factors extracted from the work of Cristy and Eckerman [12] are considered in order to basically distinguish the computation between an adult male (Smale = 1.57  10 – 6 Gy  MBq – 1  s – 1) and an adult female (Sfemale = 2.59  10 – 6 Gy  MBq – 1  s – 1). Two thyroid geometries are considered: one for male (mph = 20.7 g) and the second for female (mph = 12.4 g). More realistic female anthropomorphic phantoms (as those developed by Stabin et al. [13]) could have been considered. However, the multiplication between the corresponding S factors (found in MIRDOSE3 ¼ 1:90106 Gy  MBq1  s1 ) software [14]: SMIRDOSE3 female and the new mass (mph = 17 g for the female phantom as described by Stabin) leads to almost similar results (0.6% difference). The S factor should be modified to take into account the thyroid patient size. The correction factor for the nonpenetrating component is the ratio of the phantom mass to the estimated actual mass. Thus, the new S factor, Spatient, is related to the S factor determined by Cristy and Eckerman [12]: mph ð5Þ Spatient ¼ S m where m, the thyroid mass, is evaluated by twodimensional ultrasonography. Both penetrating and non-penetrating components are included in the S values but the penetrating component is only about 3% of the total [15] thus justifying the use of the mass-corrected S value.

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Nuclear Medicine Communications 2006, Vol 27 No 7

Fig. 1

t¼


3

þ1 Z

C2 ðtÞdt:

ð8Þ

0

Thyroid (C2)

Blood (C1)

By incorporating Equation 6,


2

t¼
1

l2 : l1 l3

Thus, the activity to administer A0 is given by the equation l1 l3 D
A0 ¼ ð10Þ l2 Spatient

Excretion Simplified two-compartment modelling.

, A0 ¼ The biokinetics were evaluated by a standard twocompartment model (Fig. 1), extracted from the threecompartment model [16]. The compartment which describes the quantity of radioiodine taken from the body and eliminated by faeces was neglected as suggested by Di Martino et al. [17] and Matheoud et al. [18]. The resolution of such a two-compartment system leads to the following differential system: 8 dC > < 1 ¼ l1 C1  l2 C1 þ l 3 C2 dt > : dC2 ¼ l3 C2 þ l2 C1 dt where C1 and C2 are the concentration activities of radioiodine in blood and thyroid and l1, l2, and l3, are the kinetics parameters of the model. Assuming C1(0) = A0 and C2(0) = 0, the system was solved by the Laplace transform: l2 A0 ½exp ða1 tÞ  exp ða2 tÞ C2 ðtÞ ¼ a1  a2 where a1 ¼ and a2 ¼

ðl1 þ l2 þ l3 Þ þ

ð6Þ

qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi ðl1 þ l2 þ l3 Þ2  4l1 l3

2

ð11Þ

where D
is the prescribed dose to the thyroid. The thyroid mass is assumed to be constant during all the treatment. This relation is the statement for approach 1. The second formulation (approach 2)

Many authors have reported a reduction in thyroid mass during treatment [19,20] and a few studies have incorporated this effect into the formalism to compute the activity [21–23]. According to these studies the variation in thyroid mass is dependent, in a first estimate, on the activity according to a first-order equation: dm AðtÞ ¼ k dt m

ð12Þ

where k is a constant. By incorporating Equation 6 into Equation 12, the thyroid mass is given by the relation sffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi ffi   2kA0 l2 exp ða2 tÞ exp ða1 tÞ 1 1 2 mðtÞ ¼  þ  þ þ m0 a2 a1 a1 a2 a2  a1 ð13Þ where m0 is the initial mass of the thyroid (i.e., just before radioiodine therapy).

dD AðtÞSmph ¼ : dt mðtÞ This is equivalent to þ1 Z A0 l2 ðea2 t  ea1 t ÞSmph
rffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi ffi dt: D¼  at  2kA0 l2 2 e2 e a1 t 1 1  þ  þ þ m 0 0 a1 a2 a2 a1 a2 a1

:

The first formulation (approach 1)

The relation between A~ and the initial activity A0 is given by the equation A~ ¼ A0 t


l1 l3 Dm l2 Smph

Equation 4 becomes

2 qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi ðl1 þ l2 þ l3 Þ  ðl1 þ l2 þ l3 Þ2  4l1 l3

ð9Þ

ð7Þ

where t is the residence time. t is expressed by the relation

The integration gives Smph D
¼  k

"sffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi # 2kA l 0 2  m0 : m20  a1 a2

ð14Þ

ð15Þ

ð16Þ

Finally, the appropriate activity to administer is given by the relation

Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.

MIRD-based models for computing

"  2 # a1 a2 2 kD
A0 ¼ m  m0  : Smph 2kl2 0

I activity in Graves’ disease Carlier et al. 563

Fig. 2

ð17Þ

60

The constant k is detailed by Traino et al. [23] and is given by

50 Thyroid uptake (%)

k¼g

131

m20 C

where g = 2.1  10 – 6 (established over a cohort of 40 patients [23]), and C is the maximum thyroid uptake. Experimental data (i.e., the three determinations of the measured thyroid uptake) were fitted with Equation 6 by the least-squares method and were not corrected for radioactive decay because the amount of radioiodine really present in the thyroid was relevant for the dose calculation. Statistical analysis

Multivariate analysis was performed to determine whether the different methods gave similar therapeutic activity. In order to achieve this, a correlated ANOVA test followed by a Tukey honestly significant difference (HSD) test were used to assess the significance of differences. When needed, a univariate analysis was achieved with the t-test. All statistically significant levels were set at 5% (P < 0.05).

Results Thyroid parameters

The mean thyroid mass was 19.4 ± 13.9 g (21.0 ± 14.6 g for women and 11.6 ± 4.2 g for men). The mean uptakes were 36.2 ± 14.6%, 42.8 ± 9.7% and 27.6 ± 6.8%, respectively at 4, 24 and 96 h. Figure 2 shows the fit of the experimental points by means of the two-compartment model. The mean effective half-life was 5.1 ± 1.6 days as calculated by the two-compartment model. The mean effective half-life extracted from the Marinelli 1 method (based on an exponential fit from 24 to 96 h) was 4.8 ± 1.5 days. In this last case, it was impossible to determine the effective half-life for four patients because the computed value exceeded 8 days. Computation of activity to be administered

Activities were computed with the eight previously described methods for the 41 patients with a prescribed dose of 106 ± 22 Gy depending on the aim of treatment (euthyroidism or hypothyroidism). The mean activities calculated for each method are given in Table 1. According to the correlated ANOVA test, at least two methods gave statistically different results (P < 0.001). The results of the Tukey HSD test are summarized in Table 1. The Leslie 1, Leslie 2 and Bockisch methods

40 30 20 10 0 0

20

40 60 80 100 Time (h) Mean thyroid uptake mean at 4, 24 and 96 h with the corresponding two-compartment modelling. Each experimental point represents the mean thyroid uptake (with the standard deviation bar) over the 41 patients. (&), Experimental data; (—), two-compartment model.

Table 1 Mean activity according to the physician prescribed dose for each formalism Method Marinelli 1 Marinelli 2 Marinelli 3 Leslie 1 Leslie 2 Bockisch Approach 1 Approach 2

Mean activity (MBq) 262 ± 176a 275 ± 223b 214 ± 165c 128 ± 95d 193 ± 143e 193 ± 137f 227 ± 148g 213 ± 124h

a

P < 0.01 between Marinelli 3, Leslie 1, Leslie 2, Bockisch and approach 2. P < 0.01 between Marinelli 3, Leslie 1, Leslie 2, Bockisch, approach 1 and approach 2. c P < 0.01 between Marinelli 1, Marinelli 2 and Leslie 1. d P < 0.01 between Marinelli 1, Marinelli 2, Marinelli 3, Leslie 2, Bockisch, approach 1 and approach 2. e P < 0.01 between Marinelli 1, Marinelli 2 and Leslie 1. f P < 0.01 between Marinelli 1, Marinelli 2 and Leslie 1. g P < 0.01 between Marinelli 2 and Leslie 1. h P < 0.01 between Marinelli 1, Marinelli 2 and Leslie 1. b

gave the lowest computed activities and Marinelli 1 and 2 the highest. Results obtained with Leslie 1 were the lowest and were statistically different from all other methods. Marinelli 1 and 2 differed from, respectively five and six other methods. The Marinelli 3, Leslie 2, Bockisch and approach 2 methods were different from three methods. Approach 1 differed only from two methods and its results were in the same range as the majority of the other methodologies.

Discussion When comparing the effectiveness of activities calculated by numerous methods it is necessary to include a very large number of patients, who must be assigned to several

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564 Nuclear Medicine Communications 2006, Vol 27 No 7

groups according to each calculation method, and matched on numerous parameters including at least age, sex, clinical conditions, thyroid volume and medications. Furthermore, the choice of the amount of radioiodine activity to administer in Graves’ disease patients can differ greatly among institutions, depending on the aim of the treatment; that is, to cure hyperthyroidism after one or more treatments or to render the patients either euthyroid or hypothyroid. Therefore, the next point to be considered should be the definition of treatment effectiveness criteria. Eradication of hyperthyroidism could be considered as the only parameter of therapeutic success, but other parameters could be implemented, for instance, related to the risk-to-benefit ratio of radiation with particular concerns about the irradiation of normal tissues. Unfortunately, there is no good indicator of thyroid tissue response to radioiodine therapy, and this latter can vary among individuals, depending on several conflicting factors, including individual thyroid tissue sensitivity, which still remains unknown. As a result, clinical outcome after radioiodine therapy is often disappointing. High rates of hypothyroidism are reported in most series [1,5,6,24,25], whereas over the last decades, much attention has focused on achieving euthyroidism by adjusting the activity of 131I. Thus, although obtaining a sustained euthyroidism appears to be a futile objective, we believe, in line with others, that the optimal approach should be to eradicate hyperthyroidism at the lowest effective radioiodine activity; that is, with an activity calculated by using personalized dosimetry. Moreover, two principal arguments justify a personalized dosimetric approach. Firstly, according to the French regulations laws based on the European Council Directive 97/43 [26], exposures of targets volume shall be individually planned and exposures of non-target volumes shall be as low as reasonably achievable. Secondly, calculating individual biokinetics gives the possibility of providing practical radiation protection guidelines for each patient when he or she leaves the institution. This can be calculated with an appropriate formalism as developed by Cormack and Shearer [27]. Two personalized dosimetric methods for prescribing a therapeutic 131I activity in Graves’ disease patients are presented. These two methods (approaches 1 and 2) are based on MIRD methodology. The main assumption was the uniform distribution of 131I in the thyroid gland, which can be considered only for Graves’ disease, because of a homogeneous thyroid distribution of radioiodine. The amount of therapeutic activity to administer to a patient is based on a computation with a suitable, weighted, S factor depending on both thyroid mass and patient sex [13], and on three-weekly uptake measurements of 131I (after administration of a low, tracer test activity of 2 MBq before treatment), which allowed an accurate determination of radioiodine biokinetics. The kinetics of 131I in the

thyroid and in blood have been evaluated with simplified two-compartment modelling, as suggested by Di Martino et al. [17]. Although this modelling was reliable and robust, mathematically speaking, some studies showed variations up to 15% between the tracer test kinetics and the therapeutic kinetics, assumed to be due to an enhanced iodine turnover under therapeutic conditions [11]. However, the same authors reported a strong correlation between the tracer test kinetics and therapeutic kinetics, except for patients whose medication changed close to the test study or therapy. Finally, although both formalisms take into account the thyroid mass at the time of therapy, the variations of the thyroid volume during the treatment are integrated only into approach 2. In 52 Graves’ disease patients, Murakami et al. observed a reduction of thyroid volume from 1–6 months after radioiodine therapy related to the outcome of thyroid function [19]. A mathematical model of thyroid mass reduction after 131I therapy was proposed by Traino et al., based on patient thyroid masses evaluated by ultrasonography and applied to a general formula for calculation of the thyroid absorbed dose [22,23]. Thus, reduction of the thyroid mass and absorbed dose to the thyroid can be both predicted and depends on the parameter k established on a cohort of patients. In contrast with the findings of Traino et al., we did not find any significant difference between the results of approach 1 and approach 2; that is, between 131I activity calculated considering a reduction of thyroid mass and that obtained without considering a change in thyroid mass. Such a discrepancy could be explained by the fact that the determination of thyroid mass is one of the intrinsic limitations shared by all methods detailed in this work. This measurement is commonly performed with two-dimensional ultrasonography which can bring a 30% error compared to the real volume of the thyroid [28]. Thus, errors in the volume determination by ultrasonography could lower the difference between approaches 1 and 2, and could at least partially explain the difference between our findings and those obtained by Traino et al. However, our methods reflect clinical practice, in which two-dimensional ultrasonography is most often performed without any quality control or calibration of the ultrasound machine. Other methods may be more reliable or accurate, such as threedimensional ultrasonography or single photon emission computed tomography, but are still not implemented in routine [28,29]. Our results showed that the two personalized dosimetric methods (approaches 1 and 2) were not statistically different from Marinelli 3 (using only a measurement at 24 h), Leslie 2 (using only a measurement at 24 h) and Bockisch (using only a measurement at 96 h). It could be argued that personalized dosimetry with one uptake

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MIRD-based models for computing

Ratio deviation for the 41 patients (minimum and maximum values) between activity calculated by the so-called Marinelli 1, Marinelli 3 and Bockisch and activity computed by approach 1

Table 2

Ratio Marinelli 1/approach 1 Marinelli 3/approach 1 Leslie 2/approach 1 Bockisch/approach 1

Minimum–maximum 10.8–1.9 0.5–1.4 0.3–1.5 0.6–1.1

Conclusion Our approaches are more constraining for the patient because three time points (with one at 96 h) need to be measured. However, these new MIRD-based formalisms appear to be a good compromise between the common formalisms already used in many institutions, with the main advantage that they take into account the individual biokinetics and the reduction of thyroid mass after therapy (only for the approach 2). Furthermore, these personalized dosimetries allow the exposures of target volumes and non-target volumes to be planned individually, and practical radiation protection guidelines to be adapted for each patient when he or she leaves the institution. Nevertheless, further studies are required to establish the effectiveness of these new methods, in particular their ability to eradicate hyperthyroidism after only one personalized therapeutic 131I activity (i.e., without later relapses).

I activity in Graves’ disease Carlier et al. 565

References 1

2

3

4

measurement point (24 h or 96 h) was equivalent to a methodology using three uptake measurement points, which is more constraining for the patient. However, this absence of difference can be partly explained by comparing effective half-lives and maximum uptakes. Indeed, the average effective half-life derived from experimental uptake measurements over the 41 patients was 5.1 ± 1.6 days using two-compartment modelling. This value was close to the fixed value of 5 days used in the Marinelli 3 methodology. On the other hand, the mean of the maximum uptake (as determined by the twocompartment model) was similar to the mean of the maximum uptake at 24 h (successively used in the Marinelli 3, Leslie 1 and Leslie 2 methods). The mathematical fit of experimental data gave a value of 0.45 ± 0.12 which was similar to that given at 24 h, i.e. 0.43 ± 0.09 (a Student’s t-test did not show any significant difference). However, the absence of a statistical difference between our formalism and the other methods must be moderated by the presence of large intra-individual differences. For a given patient, it was possible to obtain a ratio of up to 2 (respectively, 0.5) between two methods statistically equivalent (Table 2). These findings were in agreement with previously published data with an average ratio between the two methods which could reach 2.5 [30].

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Leslie WD, Ward L, Salamon EA, Ludwig S, Rowe RC, Cowden EA. A randomized comparison of radioiodine doses in Graves’ hyperthyroidism. J Clinical Endoc Metabol 2003; 88:978–983. Catargi B, Leprat F, Guyot M, Valli N, Ducassou D, Tabarin A, et al. Optimized radioiodine therapy of Graves’ disease: analysis of the delivered dose and other possible factors affecting outcome. Eur J Endocrinol 1999; 141: 117–121. Allahabadia A, Daykin J, Sheppard MC, Gough SCL, Franklyn JA. Radioiodine treatment of hyperthyroidism. Prognostic factors for outcome. J Clin Endocrinol Metabol 2001; 86:3611–3617. Kaplan MM, Meier DA, Dworkin HJ. Treatment of hyperthyroidism with radioactive iodine. Endocrinol Metab Clin North Am 1998; 27: 205–223. Franklyn JA, Daykin J, Drolc Z, Farmer M, Sheppard MC. Long-term follow-up of treatment of thyrotoxicosis by three different methods. Clin Endocrinol 1991; 34:71–76. Hennemann G, Krenning EP, Sankaranarayanan K. Place of radioactive iodine in treatment of thyrotoxicosis. Lancet 1986; 14:1369–1372. Leech NJ, Dayan CM. Controversies in the management of Graves’ disease. Clin Endocrinol 1998; 49:273–280. Reinhardt MJ, Brink I, Joe AY, Mallek D, Ezziddin S, Palmedo H. Radioiodine therapy in Graves’ disease based on tissue-absorbed dose calculations: effect of pre-treatment thyroid volume on clinical outcome. Eur J Nucl Med 2002; 29:1118–1124. Zanzonico PB. Internal radionuclide radiation dosimetry: a review of basic concepts and recent developments. J Nucl Med 2000; 41: 297–308. Marinelli LD, Quimby EH, Hyne GJ. Dosage determination radioactive isotopes. Am J Roentgenol 1948; 59:260–281. Bockisch A, Jamitzky T, Derwanz R, Biersack HJ. Optimized dose planning of radioiodine therapy of benign thyroidal diseases. J Nucl Med 1993; 34:1632–1638. Cristy M, Eckerman K. Specific Absorbed Fractions of Energy at Various Ages from Internal Photons Sources. ORNL/TM-8381 V1-V7. Oak Ridge, Tennessee: Oak Ridge National Laboratory; 1987. Stabin MG, Watson E, Cristy M, Ryman J, Eckerman K, Davis J, et al. Mathematical Models of the Adult Female at Various Stages of Pregnancy. ORNL/TM-12907. Oak Ridge, Tennessee: Oak Ridge National Laboratory; 1995. Stabin MG. MIRDOSE: personal computer software for internal dose assessment in nuclear medicine. J Nucl Med 1996; 37:538–546. Snyder WS, Ford MR, Warner GG. Estimates of Specific Absorbed Fractions for Photon Sources Uniformly Distributed in Various Organs of a Heterogeneous Phantom. MIRD Pamphlet No 5 (revised). New York: Society of Nuclear Medicine; 1978. International Commission on Radiological Protection. Individual Monitoring for External Exposure of Workers. ICRP Publication 78. Oxford, UK: Pergamon Press; 1997. Di Martino F, Traino AC, Brill AB, Stabin MG, Lazzeri M. A theoretical model for prescription of the patient-specific therapeutic activity for radioiodine therapy of Graves’ disease. Phys Med Biol 2002; 47:1493–1499. Matheoud R, Canzi C, Reschini E, Zito F, Voltini F, Gerundini P. Tissuespecific dosimetry for radioiodine therapy of the autonomous thyroid nodule. Med Phys 2003; 30:791–798. Murakami Y, Takamatsu J, Sakane S, Kuma K, Ohsawa N. Changes in thyroid volume in response to radioactive iodine for Graves’ hyperthyroidism correlated with activity of thyroid-stimulating antibody and treatment outcome. J Clin Endocrinol Metab 1996; 81:3257–3260. Nygaard B, Hegedu¨s L, Gervil M, Hjalgrim M, Hansen BM, Soe-Jensen P, Hansen JM, et al. Influence of compensated radioiodine therapy on thyroid volume and incidence of hypothyroidism in Graves’ disease. J Intern Med 1995; 238:491–497. Peters H, Fischer C, Bogner U, Reiners C, Schleusener H. Reduction in thyroid volume after radioiodine therapy of Graves’ hyperthyroidism: results from a prospective, randomized, multicentre study. Eur J Clin Invest 1996; 26:59–63. Traino AC, Di Martino F, Lazzeri M, Stabin MG. Influence of thyroid volume reduction on calculated dose in radioiodine therapy of Graves’ hyperthyroidism. Phys Med Biol 2000; 45:121–129. Traino AC, Di Martino F, Lazzeri M, Stabin MG. Study of the correlation between administered activity and radiation committed dose to the thyroid in the 131I therapy of Graves’ disease. Radiat Prot Dosim 2001; 95: 117–124. Beierwaltes WH. The treatment of hyperthyroidism with iodine-131. Semin Nucl Med 1978; 8:95–103.

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25 Doi SA, Loutfi I, Al-Shoumer KA. A mathematical model of optimized radioiodine-131 therapy of Graves’ hyperthyroidism. BMC Nucl Med 2001; 1:1–10. 26 Commission of the European Communities. Health Protection of Individuals Against the Dangers of Ionising Radiation in Relation to Medical Exposure. Council Directive 97/43, Brussels, 1997. Available from: http://europa.eu.int/ comm/energy/nuclear/radioprotection/doc/legislation/9743_en.pdf 27 Cormack J, Shearer J. Calculation of radiation exposures from patients to whom radioactive materials have been administered. Phys Med Biol 1998; 46:501–516.

Schlo¨gl S, Werner E, Lassmann M, Terekova J, Muffert S, Seybold S, Reiners Chr, et al. The use of three-dimensional ultrasound for thyroid volumetry. Thyroid 2001; 11:569–574. 29 Van Isselt JW, de Klerk JMH, Van Rijk PP, Van Gils AP, Polman LJ, Kamphuis C, et al. Comparison of methods for thyroid volume estimation in patients with Graves’ disease. Eur J Nucl Med 2003; 30: 525–531. 30 Jo¨nsson H, Mattsson S. Excess radiation absorbed dose from non-optimised radioiodine treatment of hyperthyroidism. Radiat Prot Dosim 2004; 108:107–114. 28

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Original article

Efficacy of radioiodine therapy in the treatment of elevated serum thyroglobulin in patients with differentiated thyroid carcinoma and negative whole-body iodine scan Mohsen Saghari, Ali Gholamrezanezhad, Sahar Mirpour, Mohammad Eftekhari, Abbas Takavar, Armaghan Fard-Esfahani, Babak Fallahi and Davood Beiki Introduction In the management of patients with differentiated thyroid carcinoma, serum thyroglobulin levels are often well correlated with whole-body radioiodine scanning (WBS) results. However, occasionally, a mismatched result – increased thyroglobulin with negative WBS – is observed. Radioiodine therapy has been suggested as a therapeutic choice with controversial results. Method We studied 32 differentiated thyroid carcinoma patients with elevated thyroglobulin level and negative WBS who had been treated with high-dose radioiodine. With a mean follow-up of 25.6 months (all follow-ups > 11 months), thyroglobulin and thyroid-stimulating hormone levels, WBS, clinical, radiographic and pathological findings following treatment were recorded. Results The mean pre-therapy off-treatment thyroglobulin was 152 ± 119.0 ng
ml – 1. Although there was a mild trend towards an increase in thyroglobulin in the first post-treatment year, the difference was not significant. At the end of the follow-ups, 22 patients (68.7%) were categorized as non-responders to radioiodine therapy (any change or elevation of thyroglobulin or radiological and pathological evidences of progression), four patients (12.5%) as partial responders (transient reduction but not a normalization of thyroglobulin) and six patients (18.7%) as responders (normalization of thyroglobulin with no

Introduction The diagnostic follow-up of patients with differentiated thyroid carcinoma (DTC) is routinely performed by means of serum thyroglobulin (Tg) measurement and diagnostic whole-body radio-iodine scanning (WBS) [1– 5]. Serum Tg represents a sensitive tool for detection of possible residual/recurrent disease and for early identification of DTC metastases, particularly after the initial radioiodine (RAI) thyroid ablation following total thyroidectomy [1,2,5]. Diagnostic whole-body radioiodine scans are also performed periodically to detect possible residual/recurrent disease or metastases in the follow-up course of DTC patients [1,2,5]. With this widely accepted management protocol, serum Tg levels are often well correlated with WBS results

evidence of remnant disease). In nine of 10 partial and complete responders, reduction or normalization of thyroglobulin had occurred in the first post-treatment year. Conclusion We recommend that in differentiated thyroid carcinoma patients with elevated thyroglobulin and negative WBS, at least one course of radioiodine therapy should be undertaken and if reduction or normalization of serum thyroglobulin is not achieved, repeated courses of radioiodine therapy are not logical and other therapeutic methods should be applied. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:567–572 Nuclear Medicine Communications 2006, 27:567–572 Keywords: differentiated thyroid carcinoma, thyroglobulin, whole-body scanning Research Institute of Nuclear Medicine, Tehran University of Medical Sciences, Iran. Correspondence to Dr Ali Gholamrezanezhad, Research Institute of Nuclear Medicine, Tehran University of Medical Sciences, Shariati Hospital, Northern Kargar St., 14114 Tehran, Iran. Tel: + 0098 21 880 26901; fax: + 0098 21 880 26905; e-mail: [email protected] This research has been supported by Tehran University of Medical Sciences and Health Services. grant 132.10672 Received 6 January 2006 Accepted 30 March 2006

[3,4,6]. Generally, undetectable Tg levels accompany negative WBSs, suggesting complete remission, whereas detectable or elevated Tg concentrations are associated with the presence of one or more foci of abnormal 131I uptake, as a marker of residual/recurrent disease or local or distant metastases [3,4,6]. However, occasionally, a mismatched result – detectable serum Tg levels associated with a negative WBS – is observed. This pattern has been widely studied [7–20] and a few explanations have been suggested [21]. However, the proper and logical therapeutic approach in these settings has not yet been defined exactly. Chemotherapy is generally ineffective in these relatively slow-growing tumours. Surgical intervention and radiotherapy can not be attempted because the exact site of residual/remnant disease or metastatic involvement is not identifiable. Therefore,

c 2006 Lippincott Williams & Wilkins 0143-3636

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568 Nuclear Medicine Communications 2006, Vol 27 No 7

several researchers have suggested that in these patients empirical treatment with high doses of RAI should be considered as a therapeutic choice [19], but its therapeutic effect is controversial. The aim of the present study was to evaluate the efficacy of therapeutic doses of 131I in the treatment of DTC patients with elevated thyroglobulin level but negative diagnostic WBS.

Patients and methods We performed a large retrospective review on medical records of patients who were assessed, followed and treated for DTC (papillary, follicular or Hu ¨rtle cell thyroid carcinomas) in our nuclear medicine ward between 1995 and 2005. The study only included those patients who had history of total thyroidectomy and RAI ablation therapy for DTC and subsequently during their course of follow-up developed high levels of Tg with negative WBS. Hence, these patients had another session of hospitalization in order to treat this elevated serum thyroglobulin and negative WBS state. Hospitalization for RAI ablation therapy after thyroidectomy was called the ‘first hospitalization’, and the session of hospitalization for treatment of elevated Tg and negative WBS was called the ‘target hospitalization’. Moreover, of this limited group of patients only those were selected who had at least 11 months complete follow-up (with measurement of Tg and WBS every 6 months) after target hospitalization. Those who had discontinued their follow-up after discharge from target hospitalization were excluded. No patient was lost to the study because of death from thyroid cancer. Finally, 32 patients (12 men and 20 women) with a mean age of 42.3 ± 17.4 years (range, 10–71 years) entered the study. Twenty-five patients had papillary, six follicular, and one Hu ¨rtle cell cancer. Patients’ records were reviewed for initial surgical and pathological findings; the time and sequence of subsequent RAI treatments; related pathological findings; serum Tg, anti-Tg and thyroid-stimulating hormone (TSH) levels and WBS results during the follow-up period; subsequent clinical findings and surgery for possible recurrences or metastases; and also, in a few cases, the radiographic results obtained throughout the follow-up period. Treatment with thyroxine had been stopped for 4–6 weeks before measurement of Tg and diagnostic whole-body scanning, and these two diagnostic tests were performed 2 weeks after discontinuing a 2-week course of liothyronine. All patients had been advised to have a low-iodine diet and to avoid pharmacological iodine during the period of preparation for the diagnostic scan. Pre-treatment scans were performed 48 h after oral administration of 185 MBq 131I. The pretherapy and post-therapy WBSs were reviewed by nuclear medicine specialists blinded to other clinical data. The

presence of areas with definite or possible abnormal RAI uptake was identified and all patients with either definite or possible abnormal RAI uptake in pre-therapy WBSs were excluded from the study. All WBSs were acquired using a planar gamma camera (Scintronix, UK) equipped with a high-energy collimator, energy setting of 364 keV and a 15% window. The serum levels of TSH (normal value, 0.3–4 mIU
l – 1) and anti-Tg antibody (normal value, less than 100) were measured by IRMA (RADIM, Italy) and Tg was measured by a radioimmunoassay method (CIS Bio International, France). A serum Tg level of more than 1 ng
ml – 1 was considered positive [9,11]. Tests for Tg antibodies were negative in all patients except a 62-yearold lady with an anti-Tg of 212. As noted by Fatourechi et al. [17], this is not surprising as positive anti-Tg antibodies, because of interference with the assay, result in falsely low or undetectable values, and by definition these patients might have been excluded from the study. Statistical analysis

SPSS for Windows software package (Release 11.5.0, SPSS Inc., Chicago, Illinois) was used for statistical analysis. Tg levels before and after 131I therapy were compared by using the Wilcoxon test for all patients. When more than one post-therapy result for Tg was available, the lowest value was used. Both pre-therapy and post-therapy Tg levels were obtained when the patients were off thyroxine suppression. A P value of 0.05 or less was considered significant. All P values were twotailed.

Results The mean time interval between the first hospitalization and the target hospitalization was 23.5 ± 17.1 months. In the 6-month follow-up visits after the first hospitalization, nine patients had a history of positive WBS remnant/ recurrence or metastatic disease before transformation of the disease to the WBS negative form, and they had one or more sessions of hospitalization for its treatment. In seven of these patients, there was only one pre-target hospitalization course of RAI therapy between the first hospitalization and the target hospitalization. Of these seven patients, two had a history of pulmonary metastases and were treated with 7400 MBq radioiodine and the remaining five patients had only thyroid bed remnants and were treated with 3700–5500 MBq radioiodine (mean dose of 5180 MBq). In one patient this hospitalization for treatment of WBS positive disease was repeated three times before target hospitalization (because there was remnant thyroid tissue in the thyroid bed the patient was treated twice with 3700 MBq 131I and once with 4625 MBq) and in another patient it was repeated five times with 6475 MBq 131I (each time because of disseminated pulmonary metastasis, with or without

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Efficacy of radioiodine therapy in thyroid carcinoma Saghari et al. 569

thyroid bed remnant and supraclavicular pre-treatment scans).

131

I uptake in

level comparing to the pre-target hospitalization Tg level, the difference did not reach statistical significance (P>0.5).

At the time of the diagnostic WBS and measurement of the serum Tg level, the mean serum TSH was 46.8 mIU
l – 1, and all patients had a serum TSH levels above 30 mIU
l – 1. The mean pre-target hospitalization serum Tg with the patients off-treatment was 152 ± 119.0 ng
ml – 1 (range, 11–500 ng
ml – 1). All pretherapy scans showed no evidence of functioning thyroid tissue throughout the body. The mean therapeutic 131I dose in target hospitalization was 5661 MBq (range, 3700–7400 MBq).

The second post-target hospitalization follow-up was performed approximately 12 months afterwards (11–13 months) and sufficient data for 22 patients were available. All of these patients were withdrawn from thyroxine therapy for Tg measurement: 16 of these patients (72.7%) had a significant increase, three (13.6%) had a reduction of serum Tg (compared with the baseline pre-target hospitalization level) and two (9%) had normal serum Tg tests. Magnetic resonance imaging of one patient showed cervical lymph node involvement, while X-ray examination and a bone scan showed and evidence of metastatic involvement of both humeri. The pathological analysis showed metastatic papillary carcinoma. Of those five patients who had shown a reduction of Tg level in the first post-target hospitalization follow-up and therefore no further radioiodine treatment, two patients did not show any significant change of Tg level in the second visit and two patients had a negative serum Tg tests on the second follow-up (no RAI treatment was recommended). Data for the remaining patient were not available. In general, 17/22 patients (77.2%) showed no therapeutic benefit, 3/22 (13.6%) showed partial benefit (some degree of reduction but not normalization of the serum Tg level), and 2/22 (9%) of patients showed complete improvement.

Post-treatment diagnostic whole-body scans

Post-therapy scans were available for 26 patients and were done 6–10 days after radioiodine administration. No abnormal radioiodine uptake was noted in 12 patients (46%). Faint uptake was noted in the thyroid bed in four patients, in cervical lymph nodes in two, in lung fields in two, in cervical lymph nodes and lung fields in four, in lung fields and skull in one, and in the mediastinum in one patient. The first-year follow-up after target hospitalization

After approximately 6 months (5–7 months), 25 patients were visited again for the first post-target hospitalization follow-up. All of these patients were withdrawn from thyroxine therapy for diagnostic WBS and Tg measurement: 14 of them had an increase in serum Tg (all of them underwent another course of RAI therapy), and five had a reduction (but not normalization) of serum Tg. In three patients the serum Tg in the first post-target hospitalization follow-up (6 months) was completely negative (no further RAI treatment was recommended). In the remaining three patients in the 6–7 months posttarget hospitalization, clinical evidence of regional recurrence was present and, subsequently, patients underwent surgery again. In two cases the pathology report indicated cervical lymph node involvement with papillary carcinoma of thyroid (these findings were confirmed by elevation of the serum Tg level compared to the baseline pre-target hospitalization level) and in the third case it was consistent with a recurrence of the disease which involved soft tissue of the neck and cervical and supraclavicular lymph nodes. Despite these findings in the latter case, the serum Tg level was decreased compared to the pre-target hospitalization level. In general, 17/25 patients (68%) showed no therapeutic benefit, 5/25 (20%) partial benefit (some degree of reduction, but not normalization of the serum Tg), and 3/25 (12%) of patients showed complete improvement. The mean 6-month post-treatment serum Tg level was 161.7 ± 129.2 ng
ml – 1 (range, 1.0–560.0 ng
ml – 1); Although there was a mild trend toward the increase in serum Tg

The mean 6-month post-treatment serum Tg level was 159.3 ± 132.5 ng
ml – 1 (range, 1.0–620.0 ng
ml – 1); Although there was a mild trend towards an increase in serum Tg level, the difference did not reach statistical significance (P>0.5). Follow-up diagnostic whole-body scan

In the first-year follow-up of patients (5–13 months), all patients were withdrawn from thyroxine therapy for at least one diagnostic WBS. All the follow-up scans were negative, except in the only patient with Hu ¨rtle cell carcinoma whose WBS showed faint uptake in the region of thyroid bed and lungs and two patients with faint uptake in the neck (both had pathological evidence of cervical lymph node involvement after resection). All these three patients were recommended for another course of RAI therapy, the follow-up outcome of which was not available. In general, all 32 patients had at least one post-target hospitalization follow-up visit during the first year. According to the serological, scintigraphic and pathological evidence mentioned above, there were 27 patients (84.3%) categorized as having recurrent/remnant disease. Of these, 23 were non-responders (any change or elevation of the serum Tg level or radiological and pathological evidence of progression, 71.8%) and three

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570 Nuclear Medicine Communications 2006, Vol 27 No 7

patients were partial responders (reduction of serum Tg, but not a normal Tg level, 12.5%) to the first radioiodine treatment) and re-hospitalization for RAI therapy was recommended. Of these 27 patients, three refused further hospitalization. In five patients (15.6%) there was no serological or scintigraphic evidence of the disease and these patients were categorized as responders (normalization of serum Tg with no other evidence of remnant disease) and no subsequent therapy was given. Subsequent follow-ups after the first post-treatment year

The mean follow-up of our patients after target hospitalization was 25.6 ± 7.6 months (range, 11–66 months) and a total of 21 patients have been visited for the second to fifth year follow-ups (14 patients who had been categorized as non-responders at the end of the first year, three partial responders and four responders). During 2–4 years of their follow-up, none of responders showed recurrence of the disease. Two of three partial responders (with 1.5 and 2.5 years of follow-up) showed serological evidence of remnant/recurrent disease as an increase in serum Tg level, while only one of them had negative serum Tg tests and normalization of the serum Tg level during the 2.5 years of follow-up. Twelve of 14 non-responders showed scintigraphic, pathological and serological evidence of disease progression and the presence of remnant/recurrent disease (10 patients with increased serum Tg level, three with regional recurrence in cervical lymph nodes, and one with a pathological fracture in the ribs secondary to a metastatic papillary carcinoma), while in two patients the disease remained stable with no significant changes in serum Tg levels. Of the two groups of non-responders and partial responders (16 patients), 15 underwent re-hospitalization (one to three times) with no evidence of reduction in serum Tg level, but two patients refused to be hospitalized again and had no further follow-up. As mentioned above only one partial responder showed regression of the disease and normalization of serum Tg level. At the end of the follow-ups, and based on the serological, scintigraphic and pathological evidence mentioned above, 22 patients (68.7%) were categorized as non-responders, four as partial responders (12.5%) and six patients (18.7%) as responders to RAI therapy.

Discussion In current practice, it is not unusual to encounter cases of negative WBS and elevated TSH-stimulated or nonstimulated serum Tg levels [6]. In these cases the causes of false negative scans, such as inadequate TSH elevation and iodine contamination, must be excluded initially [21]. If these explanations are not clinically relevant, a true state of Tg positive, WBS negative DTC is present, which may result from a disturbance of the iodine-

concentrating mechanism and dissociation between Tg synthesis and the iodine-trapping mechanism [19,21]. Another possible explanation is that micro-metastases which have dispersed or are too small to be visualized by diagnostic doses of 131I result in elevation of serum Tg level without visualization in WBSs [19,21]. In these cases micro-metastases may be seen with higher therapeutic doses of radioiodine. Accordingly, several researchers have shown that in such cases administration of high 131I activity (3700 MBq or more) increases the sensitivity of a post-therapy diagnostic 131I whole-body scan performed a few days later and allows the detection of neoplastic foci not seen with diagnostic doses of 131I. This fact was confirmed in our study as 53.8% (14/26) of patients had foci of radiotracer uptake in the posttreatment scan, which were not detected in the pretreatment images. Also, it is possible that these foci of abnormal radiotracer uptake become visualized in diagnostic scans after a few months (as was observed in three patients in our study), which could be explained by bulky enlargement of the tumoral mass. In fact, it could be concluded that, in the first scan, the volume of the tumoral mass was so small that it could not be detected by the limited resolution of the imaging system [19,21]. After a few months of tumour growth, the tumoral mass reaches the minimum volume required for detection by the gamma camera. Similar findings were noted in a review by Ma et al. [6] who noted that 62% positive posttherapy scans indicated that a therapeutic dose of 131I could reveal approximately one half of previously undiagnosed lesions in some patients. Also, the similar rate in the study by de Keizer et al. [15] was reported to be 59%. On the other hand, despite of the fact that RAI therapy has so far not been considered harmful [10] (although this assumption can be questioned, as the obligatory rise in serum TSH levels necessary for RAI therapy may have disadvantageous effects on tumour behaviour [10]) TSH receptor expression is retained even in advanced stages of thyroid carcinoma, which may exert proliferative effects on tumour cells. With this assumption it is not logical to impose the risk of tumour growth enhancement without any confirmed beneficial effect of this presumptive treatment. This fact is well documented in our study, as only less than on third of patients have some degree of response to radioiodine treatment. Previous results are also somewhat controversial and it has not yet been determined that this approach is effective or not. Fatourechi and Hay [19] emphasized that, based on presently available information, a generalized recommendation for RAI therapy of Tg-positive, diagnostic scannegative patients should await further studies. Meanwhile, in some high-risk patients, in the absence of alternative therapies, empirical RAI therapy is justified [19]. On the other hand, the need for treatment of these

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Efficacy of radioiodine therapy in thyroid carcinoma Saghari et al. 571

patients has been questioned, as there are some reports indicating that Tg levels may remain stable, decline, or even disappear over time without treatment [22]. In the study by Koh et al. [14], 60 DTC patients with elevated serum Tg levels but negative WBSs were divided into two groups. Twenty-eight patients were treated with RAI and 32 were untreated. In a mean follow-up of 23.8 ± 19.6 months, the percentage decrease in Tg level of the treated group was significantly higher than that of the untreated group and in four cases serum Tg levels became negative. It was concluded that therapeutic doses of RAI have a therapeutic effect, at least for palliation in short-term observation, considering the serum Tg level as an index of tumour burden [14]. In a recent review by Ma et al. [6], the results of 10 serial observations and three non-randomized controlled trials were assessed. The authors mentioned that, on the basis of data from recent studies, 131I therapy should be individualized according to clinical characteristics. In their meta-analysis, a decrease in Tg levels was achieved in 63% of DTC patients with elevated Tg and negative WBSs, suggesting that 131I therapy does have a therapeutic effect when the Tg level is considered an index of tumour burden. Therefore, 131I therapy may be justified in patients with elevated Tg levels and negative WBSs and who are at high risk of any recurrence [6]. However, in our opinion the major drawback of their conclusion is that Tg level could not always be considered as a reliable indicator of treatment efficacy. This fact is clear from the results obtained from one of our patients, in whom the serum Tg level was decreased in comparison to the pretreatment level, although clinical and pathological evidence of locoregional recurrence was apparent. The same drawback is present in the conclusion of de Keizer et al. [15], who stated that blind therapeutic doses result in a decrease in Tg levels in the majority of patients with suspected recurrence or metastases. However, as mentioned previously the results are controversial [23] and similar benefits were not observed in other studies. For example, Schaap et al. [10] did not observe a beneficial effect and they did not advise the continuation of radioiodine therapy in patients with negative post-therapeutic whole-body scintigraphy, unless a positive response is observed in individual cases. Accordingly, in our study more than two thirds of patients showed no benefit of treatment, which was consistent with the results obtained by Kamel et al. [24]. These latter authors showed normalization of Tg levels in only 6/ 38 patients while off thyroxine therapy and concluded that the effect of 131I therapy on long-term survival is not obvious [24]. In general, our findings are in agreement with those of Rosario et al. [25], who had previously concluded that, if

the post-therapy scan is negative or reveals discrete uptake in the thyroid bed, other methods (e.g., fluorodeoxyglucose positron emission tomography) can be performed, and the physician should not insist on radioiodine therapy [25]. If WBSs detect lymph node metastases, surgery is indicated [25,26], while in cases of diffuse lung metastases radioiodine is indicated until the occurrence of a negative WBS or normalization of stimulated Tg levels. These authors have found that patients with a positive post-therapy scan may show a significant reduction in Tg, with even complete remission in some cases after radioiodine, but they emphasized that the impact of this treatment on mortality remains controversial [25]. In our opinion one of the most important findings of our study which is not mentioned in previous reports is that in nine of 10 partial and complete responders, the reduction or normalization of serum Tg occurred in the first post-treatment year. Therefore, in conclusion we recommend that in DTC patients with elevated Tg and negative WBS, at least one course of RAI therapy should be given and if a satisfactory response (reduction or normalization of serum Tg) is not achieved, additional courses of RAI therapy are not logical and other therapeutic methods should be applied. As more than half of these patients will have positive post-treatment scans, this approach can have the advantage of localizing the remnant disease for other possible therapeutic modalities, such as surgical resection of involved cervical lymph nodes.

Acknowledgement Thanks are extended to the nurses at our hospital (especially Ms Javaher Abdollahzadeh) for data collection.

References 1

2 3

4

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6

7

8

Pacini F, Pinchera A, Giani C, Grasso L, Baschieri L. Serum thyroglobulin concentration and 131-I whole body scans in the diagnosis of metastases from differentiated thyroid carcinoma (after thyroidectomy). Clin Endocrinol (Oxford) 1980; 13:107–110. Pacini F, Pinchera A, Giani C, et al. Serum thyroglobulin in thyroid carcinoma and other thyroid disorders. J Endocrinol Invest 1980; 3:283–292. Charles MA, Dodson Jr LE, Waldeck N, et al. Serum thyroglobulin levels predict total body iodine scan findings in patients with treated well differentiated thyroid carcinoma. Am J Med 1980; 69:401–407. Ashcraft MW, Van Herle AJ. The comparative value of serum thyroglobulin measurements and iodine-131 total body scans in the follow-up study of patients with treated differentiated thyroid cancer. Am J Med 1981; 71:806–814. Schneider AB, Line BR, Goldman JM, Robbins J. Sequential serum thyroglobulin determination 131-I scan and 131-I uptakes after triiodothyronine withdrawal in patients with thyroid cancer. J Clin Endocrinol Metab 1981; 53:1199–1206. Ma C, Xie J, Kuang A. Is empiric 131I therapy justified for patients with positive thyroglobulin and negative 131I whole-body scanning results? J Nucl Med 2005; 46:1164–1170. Ong SC, Ng DC, Sundram FX. Initial experience in use of fluorine-18fluorodeoxyglucose positron emission tomography/computed tomography in thyroid carcinoma patients with elevated serum thyroglobulin but negative iodine-131 whole body scans. Singapore Med J 2005; 46:297–301. Pacini F, Agate L, Elisei R, Capezzone M, Ceccarelli C, Lippi F, et al. Outcome of differentiated thyroid cancer with detectable serum Tg and

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negative diagnostic (131)I whole body scan: comparison of patients treated with high (131)I activities versus untreated patients. J Clin Endocrinol Metab 2001; 86:4092–4097. Galligan JP, Winship J, van Doorn T, Mortimer RH. A comparison of serum thyroglobulin measurements and whole body 131I scanning in the management of treated differentiated thyroid carcinoma. J Med Aust NZ 1982; 12:248–254. Schaap J, Eustatia-Rutten CF, Stokkel M, Links TP, Diamant M, van der Velde EA, et al. Does radioiodine therapy have disadvantageous effects in non-iodine accumulating differentiated thyroid carcinoma? Clin Endocrinol (Oxford) 2002; 57:117–124. Pacini F, Lippi F, Formica N, Elisei R, Anelli S, Ceccarelli C, et al. Therapeutic doses of iodine-131 reveal undiagnosed metastases in thyroid cancer patients with detectable serum thyroglobulin levels. J Nucl Med 1987; 28:1888–1891. Mazzaferri EL. Treating high thyroglobulin with radioiodine: a magic bullet or a shot in the dark? J Clin Endocrinol Metab 1995; 80:1485–1487. Schlumberger M, Mancusi F, Baudin E, Pacini F. 131-I therapy for elevated thyroglobulin levels. Thyroid 1997; 7:273–276. Koh JM, Kim ES, Ryu JS, Hong SJ, Kim WB, Shong YK. Effects of therapeutic doses of 131I in thyroid papillary carcinoma patients with elevated thyroglobulin level and negative 131I whole-body scan: comparative study. Clin Endocrinol (Oxford) 2003; 58:421–427. de Keizer B, Koppeschaar HPF, Zelissn PMJ, Lips CJM, van Rijk RP, van Dijk A, et al. Efficacy of high therapeutic doses of iodine-131 in patients with differentiated thyroid cancer and detectable serum thyroglobulin. Eur J Nucl Med 2001; 281:198–202. McDougall IR. 131-I treatment of 131-I negative whole body scan, and positive thyroglobulin in differentiated thyroid carcinoma: what is being treated? Thyroid 1997; 7:669–672. Fatourechi V, Hay ID, Javedan H, Wiseman GA, Mullan B, Gorman CA. Lack of impact of radioiodine therapy in Tg-positive, diagnostic whole-body scan-

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Original article

Quantitative measurement of oxygen metabolic rate in the rat brain using microPET imaging of briefly inhaled 15O-labelled oxygen gas Seong-Hwan Yee, Kihak Lee, Paul A. Jerabek and Peter T. Fox Objective The quantitative measurement of cerebral metabolic rate of oxygen (CMRO2) for rats using positron emission tomography (PET) has been technically difficult. The present study was performed to provide a technique to measure CMRO2 for rats using a dedicated animal PET technique. Methods CMRO2 in the rat brain was quantitatively measured under a-chloralose anaesthesia (30 mg
kg – 1
h – 1, intravenous infusion) using a PET imaging technique. In our experiment, the 15O-labelled gas tracer (O15O) was administered by a bolus insufflation into the lung through a surgically placed cannula in the trachea. The tracer distribution was then dynamically imaged using the microPET. Unlike other conventional PET methods in which a series of arterial blood samples need to be withdrawn for the measurement of an arterial input function, no arterial blood sampling was employed. Instead, the heart was scanned in dynamic mode at the same time of imaging the brain, and the region of interest drawn over the heart was analysed to obtain an arterial input function.

Conclusions Our results suggest that the microPET-based CMRO2 measurement in the rat brain combined with a non-invasive measurement of arterial input function is promising, especially for many applications involving small animals in which repeated measurements of absolute CMRO2 need to be performed. c 2006 Lippincott Williams Nucl Med Commun 27:573–581  & Wilkins. Nuclear Medicine Communications 2006, 27:573–581 Keywords: cerebral metabolic rate of oxygen, CMRO2, microPET, non-invasive, PET, rat

Research Imaging Center, University of Texas Health Science Center, San Antonio, USA. Correspondence to Dr Seong-Hwan Yee, Research Imaging Center, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, TX 78229, USA. Tel: + 001 (210) 567 8100; fax: + 001 (210) 567 8152; e-mail: [email protected] This work was supported by Grant R21 NS050486-01A1 from the NIH/NINDS.

Results The CMRO2 value (lmol
100 g – 1
min – 1) from 10 rats was 208 ± 15 (mean ± SD).

Introduction Accurate measurement of cerebral metabolic rate of oxygen (CMRO2), for both humans and laboratory animals, is of great interest for studying brain functions because the neuronal activity and associated demand for energy, especially provided by oxidative metabolism, are important factors in modulating brain haemodynamics that are detected by widely used functional imaging tools, such as functional magnetic resonance imaging (MRI) and positron emission tomography (PET). The measurement of CMRO2 also has important applications in the study of neurological disorders, not only in the clinical setting but also in the laboratory setting using animal models. There have been several different imaging techniques, especially for PET and MRI, developed to measure CMRO2. For PET, the 15O-labelled gas tracer O15O is commonly used as an imaging agent for a PET scan, and the dynamic tracer distribution in the subject is modelled to obtain an estimation of the absolute CMRO2 value. The PET technique could require two additional PET scans,

Received 20 January 2006 Accepted 30 March 2006

one for the determination of cerebral blood flow (CBF) using H2 15O and the other for the determination of cerebral blood volume (CBV) using C15O. This three-step technique [1] has been widely used as the PET-based absolute CMRO2 measurement technique, especially for humans and large animals. More recently, the one-step method [2] has been developed as a simplification of the three-step method to avoid the requirement for the additional PET scans by utilizing the time-weighted integral method [3,4]. The advantages of the one-step method are two-fold. First, it is suitable for multiple repeated measurements within a limited time, such as for brain activation studies [5–10], because it does not require additional PET scans using different PET imaging tracers. Second, the arterial input function (AIF) measurement procedure can be much simplified, because it does not require separate measurements of AIF for oxygen and for water as required for the three-step method. For MRI, 17O2 can be used as an imaging agent [11–13]. For this technique, 17O2 is delivered to the subject and

c 2006 Lippincott Williams & Wilkins 0143-3636 

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574 Nuclear Medicine Communications 2006, Vol 27 No 7

the metabolic process is modelled using the principle that 17O is detected by MRI only in the form of water (H2 17O) once the 17O2 is consumed inside the body and the H2 17O is produced as a by-product. However, the 17 O-based MRI technique is still of limited use due to the high cost of the 17O tracer, and also due to the sensitivity problem of measurement in low field strength. For the purpose of measuring only relative changes of CMRO2, the blood oxygenation level dependent (BOLD) functional MRI technique based on the hypercapnia calibration and an extensive BOLD biophysical modelling can also be used [14,15]. For the small animals such as rats, which are most widely used in laboratory settings, the 15O PET technique for absolute CMRO2 quantification has not been widely used. One of the reasons is the difficulty of determining an AIF from small animals that have only limited volume of available blood. In addition, the fast dynamics of an AIF during the bolus arrival might be corrupted easily with a conventional method, since the heart rate of rats or mice is much faster than that of humans, requiring much faster blood sampling. In this present study, CMRO2 in the rat brain was measured quantitatively using highresolution dedicated animal PET equipment (microPET R4; CTI, Knoxville, Tennessee, USA) by applying the one-step method [2] and a non-invasive AIF measurement technique developed previously [16].

Materials and methods Theory Determination of cerebral metabolic rate of oxygen

The one-step technique for CMRO2 measurement can be described by the equation [2] Zt h i O2 Ca ðuÞ exp kO2 2 ðu  t Þ du þ V0 Ca ðt Þ ð1Þ M ðt Þ ¼ K 1 0

where M(t) is the tracer concentration (per unit mass of the brain tissue) in the brain at time t and Ca(t) is the tracer concentration (per unit volume of the arterial blood) in the artery supplying fresh blood to the brain. The arterial tracer concentration, Ca(t), is the concentration of the tracer, regardless of the chemical form (either O15O or H2 15O). The constant K1O2 is the clearance of the oxygen and has the same unit as CBF (flow volume per unit mass of the brain per unit time); it is equivalent to the CBF (inflow) multiplied by the oxygen extraction fraction (OEF). It is worthwhile noting that the recirculating water component was assumed to be negligible in this one-step method [2]. The constant 2 is the washout rate of the labelled water as a kO 2 metabolite of O15O and has the unit of reciprocal time; it is equivalent to the CBF (washout flow) divided by the brain–blood partition coefficient l (the ratio of the brain tissue concentration per unit mass to the venous blood concentration per unit volume) of the water tracer. The

constant V0 is related to the CBV, and is used to correct the effect from the non-extracted oxygen and circulating 2 water. The three unknown constants (K1O2 , kO 2 and V0) can be determined by the time-weighted integral method and the measurement of CMRO2, in the unit of millilitres of oxygen per unit mass of tissue per minute, can be determined by the product of the constant K1O2 and the total arterial oxygen content CaO2 (in millilitres of oxygen per unit volume of arterial blood): CMRO2 ¼ K1O2 CaO2 . In the PET-based in-vivo CMRO2 measurement technique, M(t) can be determined using PET imaging of the brain. However, since M(t) in Equation 1 is expressed in the unit of concentration per unit mass, the reconstructed signal intensity of the PET image, which is proportional to the concentration per unit voxel volume, must be converted into the signal intensity proportional to the concentration per unit mass, by multiplying a reciprocal of the brain density. In addition, if the AIF (Ca(t)) is determined by measuring the activities of arterial blood samples from the subject using a separate detector, a calibration constant between the signal measured by PET and the activity measured by a separate detector must also be known. However, since the AIF can be measured using the same PET detector in our method, any calibration between two differently measured signals will be unnecessary. Hence, the equation for CMRO2 measurement using PET can be rewritten as follows, in order directly to utilize the PET signal in the modelling: Zt h i O2 2 Sb ðt Þ ¼ K1;r Sa ðuÞ exp kO ð u  t Þ du þ V0;r Sa ðt Þ ð2Þ 2 0

where Sb(t) is the signal intensity for the brain measured by PET imaging at time t, in the unit of signal intensity per reconstructed voxel volume, Sa(t) is the AIF obtained by analysing and correcting the PET signals in the cardiac ROI, expressed in the same unit as Sb(t), both constants O2 K1;r and V0,r are, respectively, equivalent to constants K1O2 and V0 multiplied by the mean brain density r, and the others are the same as in Equation 1. Therefore, if signal intensities proportional to the tracer concentration are measured in the brain and in the heart as functions of time O2 2 using PET, the three unknown constants (K1;r , kO 2 and V0;r ) can be determined using the time-weighted integral method applied for Equation 2, and the measurement of CMRO2 can be obtained by using the equation CMRO2 ¼

O2 K1;r CaO2

r

:

ð3Þ

Determination of the arterial input function

The AIF was determined by the non-invasive imagederived method as described in our previous study [16]. Briefly, it is based on the assumption that the time– activity curve (TAC) obtained in an ROI drawn over the heart dynamically imaged at the same time of imaging the

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CMRO2 measurement in rats using microPET Yee et al. 575

brain would be a good estimate of the actual AIF, if a correction method to eliminate any non-arterial component in the TAC is applied. The equation to obtain an AIF is Sa ðt Þ ¼ Sh ðt Þ þ Sh ðt Þ
Dðt Þ

ð4Þ

where Sa(t) is the AIF to be determined, Sh(t) is the TAC of the cardiac ROI and D(t) is a weighting function of the form
 ln 2 Dðt Þ ¼ D0 exp 
t ð5Þ Th where D0 and Th are the constants that specify the function. Animal preparation and PET imaging

For the CMRO2 measurement using the microPET, 10 rats (Fischer 344, male) were used. Body weights ranged from 240 to 290 g (261.8 ± 16.4 g). For each animal, surgery was performed under gas anaesthesia established by the isoflurane (1–2%) mixture with oxygen. The surgical procedure consisted of three parts. First, the left femoral vein was catheterized using a polyethylene catheter (PE-10) for infusion of anaesthetic during the imaging procedure. Second, the left external carotid artery was catheterized using a polyethylene catheter (PE-10) for withdrawing arterial blood to measure arterial oxygen content at the end of the experiment. Third, a tracheotomy was performed to provide a passageway for the administration of 15O-labelled oxygen gas directly into the lung through a surgically placed cannula (16 GA); one end of the cannula was left open to the room air to ensure autonomous breathing. After the surgical procedure, intravenous administration of a-chloralose (SigmaAldrich, St. Louis, Missouri) through the catheterized femoral vein was initiated for inducing and maintaining anaesthesia for the imaging procedure: a bolus injection of a loading dose (40 mg
kg – 1) followed by continuous infusion (30 mg
kg – 1
h – 1), and the rat was positioned in the scanner to cover both the brain and the heart in one field of view (FOV). Subsequently, the PET scan was initiated in three-dimensional (3-D) acquisition mode with the use of the O15O as a tracer (74–111 MBq, 5 ml), which was produced by the on-site cyclotron (Scanditronix Model MC17F). The gas tracer, contained in a 10ml syringe, was delivered to the microPET room and insufflated into the surgically placed cannula in the trachea. For the PET scan, the coincidence timing window was set at 6 ns, and lower and upper level energy discriminators were set at 350 keV and 750 keV, respectively. All procedures were approved by the Institutional Animal Care and Use Committee of the University of Texas Health Science Center at San Antonio. Data analysis for CMRO2 calculation

The 3-D PET data acquired for 3 min in list mode was reconstructed into 1-s frames, as described in our

previous study [16]. Each frame has a 3-D data set with a matrix size of 128  128  63 and a voxel size of 0.85 mm  0.85 mm  1.21 mm. The in-plane resolution of the microPET scanner, obtained using 22Na, was reported as approximately 2.5 mm [17]; the resolution (full width at half maximum) obtained using a 15O water line phantom in our laboratory was approximately 3 mm. Sb(t) (the TAC of the brain) and Sa(t) (the corrected cardiac TAC) of Equation 2 were obtained using the ASIPro 4.1 (CTI, Knoxville, Tennessee) and an in-house program developed using MATLAB 6.5 (The MathWorks, Natick, Massachusetts). The ROIs for the brain and the heart were drawn using the image obtained by summing all the 1-s frames. The TAC data from the brain ROI was set to Sb(t) without any correction, but the TAC data from the heart was corrected using the method described in Equations 4 and 5. The volume ratio of the cardiac ROI to the pure arterial space inside the ROI was assumed to be 3:1, resulting in a D0 value of 2. A value of t = 0 was set to the time of initiation of the scan, which is approximately 5 s earlier than the start of tracer administration. The corrected TAC was used as an AIF, equal to Sa(t), in our method. The time-weighted integral method [2] to determine the three unknown constants in O2 2 Equation 2 (K1;r , kO 2 and V0;r ) was then applied using an in-house program developed on MATLAB 6.5. We pffiffi used 1, t and t for the three weighting functions in the time-weighted integral method [2]. Once the constant K1O2 had been determined, the CMRO2 was calculated using Equation 3; the average brain density used was 1.05 [1] and the oxygen content of the arterial blood used in Equation 3 was measured by an oximeter (AVOXimeter 1000E; A-Vox Systems, San Antonio, Texas) using the blood sample withdrawn from the carotid artery at the end of experiment. For the calculation of the voxel-level CMRO2, a TAC of each voxel (Sb(t) of each voxel) was obtained throughout the ROI in the brain. Then, for each voxel (0.85 mm  0.85 mm  1.21 mm) in the ROI drawn O2 over the brain, the constant K1;r was computed and a regional CMRO2 value was calculated using Equation 3.

Results The axial and sagittal sections of the rat brain, imaged by the 3-min PET scan using the intra-tracheal bolus administration of the O15O gas tracer, are shown in Fig. 1(a). The coronal and sagittal sections for the heart, used for drawing a cardiac ROI, are shown in Fig. 1(b). As seen in the images, both the brain and the heart were imaged clearly in the same scanner FOV. The data analysis for the ROI drawn in the heart was further performed to obtain an actual AIF. The TACs of the brain and the heart from the intratracheal bolus administration of O15O are shown in Fig. 2(a). The cardiac TAC shown in Fig. 2a was obtained by the previously described correction method. In

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576 Nuclear Medicine Communications 2006, Vol 27 No 7

Fig. 1

Fig. 2

(a)

(b)

(a)

0.08 L

R

L

Activity (a.u.)

R

0.06

0.04 Heart TAC Brain TAC

0.02 Cross-sectional images, obtained for a rat by summing all the reconstructed frames from the 3-min microPET scan using the O15O tracer administered by a bolus insufflation into the lung. The tracer was administered into the lung through a cannula inserted by a tracheotomy. (a) Axial and sagittal sections, respectively from the left, for the brain. The view of the axial image is from the tail to the head of a rat positioned supine. (b) Coronal and sagittal sections, respectively, for the heart. The cross lines show the corresponding position of the sectional images. The right and left sides of the rat are denoted by R and L in the images. All images are displayed using the same colour scale.

0 0 (b)

3

30

60

90 Time (s)

120

150

180

× 10− 3

Fig. 2(b), data points from the measured brain TAC are shown together with the fitted data points to the model function (Equation 2) using the time-weighted integral method described (R2 = 0.874). It shows a clear similarity to the conventional AIF obtained by withdrawing arterial blood samples for human PET studies. It is noteworthy, however, that the data fitting was poor near the peak of the AIF. The lack of fit might be due to the effect of vascular transit of the excessive oxygen; if the oxygen supply is saturated instantaneously, which might be occurring in this case, the excess oxygen will flow without being taken up during this transit time. This vascular transit effect is common when an intra-carotid bolus injection of a certain tracer (e.g., H2 15O) is employed [18]. In addition, Fig. 2a shows the convergence of both the heart and the brain TACs to the same level as a function of time, which is expected from the fact that the ratio of the two asymptotic values, if the unit of the brain TAC is converted to the signal intensity per unit mass, should be closely related to the water partition coefficient (lB0.96 ml
g – 1). This is because the tracer distribution would reach the equilibrium state while the inhaled labelled oxygen gas is gradually converted into water. This could be verified by plotting the relative signals (brain TAC to cardiac TAC), as shown in Fig. 3, where the brain and the heart TACs were plotted (Fig. 3(a and b)), together with relative signals (Fig. 3(c and d)). It should be noted that the cardiac TAC in Fig. 3, denoted by Sh(t), is not the corrected AIF. In Fig. 3(a and b), the raw TAC data (grey dotted line) were partially (using only data points after 20 s) smoothed (solid line) with a cut-off frequency of 0.2 per sample, in order to reduce the noise level in getting the relative signals. The reason why

Activity (a.u.)

2.5 2 1.5 Measured Fitted (R2 = 0.874)

1 0.5 0

0

30

60

90 Time (s)

120

150

180

The time–activity curves (TACs) of the brain and the heart, obtained for a rat by the 3-min microPET imaging of the O15O tracer administered by a bolus insufflation into the lung. The cardiac TAC was corrected and shown here. The bolus injection of the tracer was made into the lung through the cannula inserted by a tracheostomy. The whole 3-min data was rebinned and reconstructed into multiple 1-s frames. (a) TACs for both the brain and the heart in the same scale. (b) The brain TAC, only, together with a fitted curve using the model function (Equation 2).

smoothing was applied partially only to data points after 20 s is, firstly, because the noise level during the initial period was very low, as seen in the plots; and secondly, because an undesirable significant reduction of the cardiac TAC peak was expected by the smoothing. As seen in Fig. 3(c), the relative signal is approaching 1, verifying the convergence of both the heart and the brain TACs to the same level as a function of time. The measured arterial oxygen contents (CaO2 in Equation 3) for the 10 rats ranged from 15.2 to 18.6 ml
dl – 1, with an average value of 16.87 ± 1.07 ml
dl – 1 (mean ± SD). The values for CMRO2, in units of mmol
100 g – 1
min – 1, obtained for the whole brain from each of the 10 rats

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CMRO2 measurement in rats using microPET Yee et al. 577

Fig. 3

(a)

3

× 10− 3

(b) 0.04

2.5 0.03

1.5

Brain TAC: Sb(t )

PET signal

PET signal

2

1

0.02 Cardiac ROI: Sh(t) 0.01

0.5 0 0

20

40

60

80 100 120 140 160 180 Time (s)

0

(d)

1

0.6

0.6

Sb(t )/Sh(t )

40

60

80 100 120 140 160 180 Time (s)

1-Sb(t)/Sh(t)

0.4

0.2

0.2

0

Ratio

0.8

0.4

20

1

0.8

Ratio

(c)

0

0

20

40

60

80 100 120 140 160 180 Time (s)

0

0

20

40

60

80 100 120 140 160 180 Time (s)

The brain and the heart TACs (a and b, respectively) from one subject, and the relative signals (c and d). The cardiac TAC in b, denoted by Sh(t), is not the corrected arterial input function. In a and b, the raw TACs (grey dotted line) were partially (using only data points after 20 s) smoothed (solid line) with a cut-off frequency of 0.2 per sample, in order to reduce the noise level in obtaining the relative signals. In c and d, grey dotted plots represent the relative signals obtained using the smoothed plots in a and b, while the solid line plots represent exponential model functions. Note that the relative signal shown in c rose to 80% at approximately 100 s.

ranged from 179 to 233, and the average value was 208 ± 15 (mean ± SD). The values of the CMRO2 in units of ml
100 g – 1
min – 1 ranged from 4.31 to 5.60, with an average of 5.00 ± 0.36 (mean ± SD). In addition, values of CMRO2 for each voxel in the 3-D ROI drawn in the brain have been calculated using Equations 2 and 3. For all voxels, the same AIF was used as obtained for the global CMRO2 calculation using the data analysis in the cardiac ROI. The voxel-level CMRO2 map in one of the 10 rats is shown in Fig. 4. The maps are for axial, coronal and sagittal planes, respectively, from the left, for the locations at the cross mark. The values ranged from 47.8 to 342 mmol
100 g – 1
min – 1, with an average value of 184 ± 65.7 mmol
100 g – 1
min – 1 (mean ± SD). As seen in Fig. 4, it is difficult to see the differentiation between grey and white matter, which might be due to the limited resolution.

Discussion The absolute value of CMRO2 in the rat brain in the resting state has been measured using microPET, which is high-resolution, dedicated animal PET for rodents. For each voxel of the 3-D ROI drawn in the brain, a voxellevel regional CMRO2 value was also obtained. In our method, the 15O-labelled oxygen gas tracer was administered by the bolus insufflation of the tracer directly into the lung through a surgically placed cannula. In addition, the arterial blood sampling procedure was eliminated utilizing a data analysis technique in the ROI drawn over the heart which was imaged at the same time of imaging the brain. Our data shown here clearly demonstrate that our developed method for CMRO2 measurement using the microPET is feasible for small animals such as rats. There are several advantages of our technique. Since it does not require any blood sampling procedure for

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578 Nuclear Medicine Communications 2006, Vol 27 No 7

Fig. 4

342 (µM/100g/min)

184 (mean)

47.8

R

L

R

L

D

V

Cross-sectional CMRO2 maps obtained for one of the 10 rats, from the 3-min microPET imaging of the O15O tracer administered by a bolus insufflation into the lung. The maps were overlaid onto a 3-D image obtained by summing all the framed images. They are axial, coronal and sagittal views, respectively from the left, at the locations indicated by the cross marks. The colour bar shows the minimum CMRO2 value of 47.8 mmol
100 g – 1
min – 1, the maximum CMRO2 value of 342 mmol
100 g – 1
min – 1, and the average CMRO2 value of 184 mmol
100 g – 1
min – 1, in the whole 3-D brain. Total number of voxels in the 3-D brain ROI was 4227 for this rat. The right and left, the dorsal and ventral sides are marked by R, L, D and V, respectively.

determining an AIF, the whole procedure for CMRO2 measurement is much simplified in terms of the labour intensity required in the laboratory during the experiment. More importantly, our technique to measure CMRO2 can be applied for small animals in a repeated manner despite the limitation of the available blood volume. Additionally, since the use of a separate detector can be avoided, the possible errors present in the calibration process between the measured PET signal and the activity measured by a separate detector is eliminated, allowing the CMRO2 measurement technique to be less susceptible to calibration errors, which might be an issue when different imaging modes (2-D or 3-D) or different tracers (18F or 15O) are used between the calibration and actual imaging processes. Although our method to obtain an AIF for rats was successfully applied in this study, there is a question that needs to be addressed further. It is whether the use of an exponential function for D(t) as in Equation 5 is acceptable or not, even in the case of the O15O tracer, which might behave differently inside the body from the H15 2 O tracer used in the previous study [16]. To address this, it is helpful to estimate CS/CROI (in Equation 10 in the Appendix) using a reasonable method. If the arterial space A or the surrounding space S, as in the Appendix, could be clearly delineated by PET imaging, CS/CROI can be easily obtained from the two separately drawn ROIs; however, it is impossible to differentiate the arterial space (or the surrounding space) from the ROI space due to the ‘still’ limited spatial resolution and cardiac motion. Alternatively, CS could have been calculated using an approximate model for the oxygen uptake; however, the correct estimation of CS would require prior knowledge of a ‘true’ AIF, which is a conundrum (because CS will be

used for estimating a ‘true’ AIF but CS will need a ‘true’ AIF before being estimated). However, we can reasonably assume that, by noting that CS is the concentration in the surrounding medium, the functional form of CS/CROI would be very similar to the Cbrain/CROI, where Cbrain is the concentration in the brain tissue. The functional form of Cbrain/CROI can be estimated by Sb/Sh, since the PET signal is proportional to tracer concentration. Therefore, the plot in Fig. 3(c) was obtained by dividing Sb (the brain TAC) shown in Fig. 3(a) by Sh (the uncorrected cardiac TAC) shown in Fig. 3(b). The plot in Fig. 3(d) was obtained by 1 – Sb/Sh, which is directly related with the function D(t) in Equation 5. Exponential forms are clearly seen in Fig. 3(c and d). This result demonstrates that the assumption of an exponential function for the weighting function D(t) in Equation 5 is acceptable. The initial amplitude D0 of function D could be determined by VS/VA (or (VROI/VA) – 1, using VROI = VS + VA), if the exact sizes of the ROI and the arterial space A are known. As in our previous study [16], the ratio of VROI to VA was assumed to be approximately 3:1, making D0 equal to 2. The parameter Th was introduced to represent the ‘halflife’ of the weighting function D(t). We assumed that the time point when Sb reached approximately more than 80% of Sh was the time when the weighting function D(t) had almost decayed. In other words, approximately at that time, the error of using the uncorrected cardiac TAC as a true AIF was assumed to be negligible. Therefore, we assumed that approximately 10 half-lives have passed until that time point. As seen in Fig. 3(c and d), approximately 100 s was assumed to be that time (10  Th), resulting in Th = 10 s. Since there are uncertainties in determining D0 and Th, it is worthwhile analysing the error associated with the estimation of D0 and Th. For this purpose, a single set of brain and cardiac TACs were selected from one subject. By individually varying D0 and Th around specific values (D0 = 2 and Th = 10), values of the CMRO2 for the rat were obtained for each possible set of D0 and Th. The percent error at each possible set of D0 and Th from the value when D0 = 2 and Th = 10 are shown in the grey scale (darker for greater error) in Fig. 5. Contour lines were drawn at multiples of 10% from the central 0%. This figure shows that, if the error comes only from determining Th, approximately 20–37% error in determining Th would result in less than 10% error in the calculated CMRO2, and if the error comes only from determining D0, approximately 18–22% error in determining D0 would result in less than 10% error in the calculated CMRO2. In addition, the errors within 10% in both D0 and Th would result in 10% error in the calculated CMRO2. It is also seen that the errors could remain relatively constant toward the direction of either (higher Th, lower D0) or (lower Th, higher D0), as long as both parameters are changing in opposite directions (increase and decrease) at the same time.

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CMRO2 measurement in rats using microPET Yee et al. 579

Fig. 5

Fig. 6

0.035

1.2 20%

1.4

0.03

10%

1.6

Average signal (a.u.)

Initial amplitude (D0) of D(t ) (unitless)

1

0%

1.8 2 2.2 −10%

2.4

−20%

2.6

0.025 0.02 0.015 0.01

2.8

− 30% 0.005

3 5

6

7

8

9 10 11 12 Half-life (Th) of D(t) (s)

13

14

15

The CMRO2 estimation errors associated with determining the parameters D0 and Th in Equation 5. The grey scale here represents the degree of error (darker for greater error), calculated by the percent difference from the reference value obtained when D0 = 2 and Th = 10 s using a set of brain and cardiac TACs from one subject. The possible values of D0 and Th are shown in both axes (note the reverse order of D0). Contour lines were drawn at multiples of 10% from the central 0%. The arrows show the possible ranges of two parameters within 10% error of the estimated CMRO2. The square at the centre shows the range of CMRO2 error when two parameters are in 10% error ranges.

In our experiment, the whole FOV was fully employed, since the brain and the heart need to be covered by a single PET scan. However, different axial planes in the same FOV may have different sensitivities and noise levels due to the 3-D mode of reconstruction (especially between those at the ends of FOV and those at the centre). The plot in Fig. 6 shows a profile across all the axial planes to illustrate this effect: the profile was obtained from an image of a 15O water line phantom (length = 15 cm, diameter = 0.3 mm) reconstructed in 3D mode. As seen in Fig. 6, the noise levels toward the ends of the FOV are clearly increasing. Nevertheless, if we exclude the edge plane in each side, the noise levels are relatively uniform. Considering also the fact that the higher activity object (either the brain or the heart) positioned toward the ends can reduce the statistical noise levels, the image quality is not much degraded by positioning the heart and the brain toward the ends within the same FOV. Since our approach to obtain an image-derived AIF seems to be different from that used by other investigators (i.e., the method based on recovery and spill-over constants, [19,20]), it is helpful to compare the two approaches. For that purpose, an equation that contains recovery and spill-over constants (which we will call r and m, respectively) can be written as follows, by substituting the notations used in other studies for those used in our

0

10

20 30 40 Axial plane number

50

60

A profile across all the axial planes of microPET, obtained from an image of a 15O water line phantom (185 MBq, length = 15 cm, diameter = 0.3 mm) reconstructed in 3-D mode.

method: CROI ¼ rCA þ mCS : If we rearrange the equation, then 
 1r m CS 1
CA ¼ CROI þ CROI r 1  r CROI

ð6Þ

ð7Þ

If m + r = 1, as is usually assumed [19], then m/(1 – r) = 1, and therefore, 

 1r CS
1 ð8Þ CA ¼ CROI þ CROI r CROI Equation 8 is equivalent to Equation 9 (and also to Equation 4) in our method. Therefore, the basic principle of the two methods is the same. However, our method further assumes the functional form of (1 – CS/CROI), which might be an inevitable approach since it is not practical to have separate ROIs for myocardium and left ventricle spaces (as in other studies, [19]) for rats with such a small cardiac dimension. Likewise, it is not feasible, for small animals, to draw and test several ROIs on different regions (e.g., left ventricle, left atrium, ascending aorta or descending aorta) for searching the best possible candidate in determining an AIF [21]. Only few imaging studies, in which absolute CMRO2 values were measured in the rat brain, have been reported: one from the 17O MRI study [13] and the other from a 15 O PET study [22] are two of them. When compared to these studies, our results are in good agreement. In the study where the 17O MRI method was applied, the value of 2.19 ± 0.14 mmol
g – 1
min – 1 was reported for the brains of rats anaesthetized with a-chloralose [13]. The reported value is in excellent agreement with our result (208 ± 15 mmol
100 g – 1
min – 1). The other reported

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580 Nuclear Medicine Communications 2006, Vol 27 No 7

CMRO2 values obtained by using a small animal PET and 15 O tracers is 4.3 ± 1.3 (ml
100 g – 1
min – 1 for the brains of rats anaesthetized with pentobarbital [22]). The reported CMRO2 value is 14% lower than ours (5 ± 0.36 ml
100 g – 1
min – 1), but the range of measurements is still in good agreement: the effect of different anaesthetics used for the imaging procedure might have to be considered as well. Additionally, it is interesting to note that our result is a little higher than a stoichiometrically expected value from a reported cerebral glucose metabolic rate (CMRglc) of 26 mmol
100 g – 1
min – 1 for rats anaesthetized with a-chloralose [23]; the expectation was 1:6 for CMRglc to CMRO2. To our knowledge, our work is the first reported CMRO2 measurement study in the rat brain that used (1) highresolution animal PET, (2) the non-invasive method for an AIF determination and (3) the one-step method which requires only one PET scan (therefore allowing repeatable measurements at a much faster rate). In the previous study [22] where high-resolution animal PET was also used, the conventional invasive method for determination of AIF, and the three-step method which requires additional PET scans for CBF and CBV measurements, were still used (although only one additional PET scan for CBF measurement was performed in the study by importing CBV values from another reference). Based on our present development for the CMRO2 measurement technique utilizing a non-invasive technique for the determination of AIF, more complicated studies, in which the CMRO2 needs to be measured repeatedly, can be planned with fewer complications. However, in future studies, the limitations of our technique, which include the invasiveness of tracheostomy (although not significant) for administration of the gas tracer, the size limitation of the animal (even rats if the size is bigger) imposed by the scanner FOV due to the need to have a simultaneous heart–brain scan, the possibility of a poor signal-to-noise ratio of the cardiac TAC due to the smaller ROI size, and the possibility of having a poorly estimated weighting function D(t), should not be overlooked. Further improvements might be needed to overcome these limitations. For further validation of our technique which utilizes a data analysis method in the cardiac ROI, a method that does not require any AIF for CMRO2 calculation can be employed [24]. It would involve labelling blood with O15O for injecting the labelled blood into the internal carotid artery. This validation, however, was not performed in our present study. It is our intention to perform this validation in a rigorous way as our next step. A rigorous validation would have to employ physiological challenges to shift the variables (e.g., CBF or CMRO2) in several different states either by using hypercapnia which would change CBF substantially without changing CMRO2 [25] or by using core temperature challenge which would change both CMRO2 and CBF [26,27].

Conclusion A study for quantitative measurements of CMRO2 in rat brain was successfully performed using the short-lived isotope 15O, as O15O, and dedicated high-resolution animal PET. In our study, a new non-invasive method for obtaining an AIF was used for the CMRO2 measurement in rat brain. Based on our results, we demonstrated that the microPET technique utilizing 15O-labelled oxygen (O15O) gas as an inhalant imaging tracer for the quantitative measurement of CMRO2 could be done without requiring the invasive, labour-intensive and technically demanding arterial sampling procedure.

References 1

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16

17

18

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Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.

CMRO2 measurement in rats using microPET Yee et al. 581

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Iida H, Miura S, Shoji Y, Ogawa T, Kado H, Narita Y, et al. Noninvasive quantitation of cerebral blood flow using oxygen-15-water and a dual PET system. J Nucl Med 1998; 39:1789–1798. Chen K, Bandy D, Reiman E, Huang S, Lawson M, Feng D, et al. Noninvasive quantification of the cerebral metabolic rate for glucose using positron emission tomography, 18F-fluoro-2-deoxyglucose, the Patlak method, and an image-derived input function. J Cereb Blood Flow Metab 1998; 18:716–723. van der Weerdt AP, Klein LJ, Boellaard R, Visser CA, Visser FC, Lammertsma AA. Image-derived input functions for determination of MRGlu in cardiac (18)F-FDG PET scans. J Nucl Med 2001; 42:1622–1629. Magata Y, Temma T, Iida H, Ogawa M, Mukai T, Iida Y, et al. Development of injectable O-15 oxygen and estimation of rat OEF. J Cereb Blood Flow Metab 2003; 23:671–676.

Appendix Detailed descriptions of our method can be put forward using the left-hand diagram in Fig. 7, where an ‘ideal’ arterial space is designated by ‘A’, and the remaining, nonarterial, surrounding space in the ROI is designated by ‘S’. It should be noted that only the ROI space is what we can draw (the space A is an ‘ideal’ space that is assumed to exist inside the drawn ROI). If the concentrations in each space are CA, CS and CROI, respectively, for arterial (A), non-arterial surrounding (S) and ROI spaces, CA can be written as Equation 9, where VA and VS are volumes of spaces A and S:




 VS CS
1 : VA CROI

ð9Þ

This equation suggests that the arterial concentration can be obtained by knowing the concentration in the ROI and a weighting function as enclosed by the outer brackets in the second term. If this weighting function is designated by D, the following equation can be written for D:
 VS CS D¼
1 : VA CROI

24

25

26 27

Sibson NR, Dhankhar A, Mason GF, Rothman DL, Behar KL, Shulman RG. Stoichiometric coupling of brain glucose metabolism and glutamatergic neuronal activity. Proc Natl Acad Sci U S A 1998; 95:316–321. Raichle ME, Grubb RL Jr., Eichling JO, Ter-Pogossian MM. Measurement of brain oxygen utilization with radioactive oxygen-15: experimental verification. J Appl Physiol 1976; 40:638–640. Horvath I, Sandor NT, Ruttner Z, McLaughlin AC. Role of nitric oxide in regulating cerebrocortical oxygen consumption and blood flow during hypercapnia. J Cereb Blood Flow Metab 1994; 14:503–509. Klementavicius R, Nemoto EM, Yonas H. The Q10 ratio for basal cerebral metabolic rate for oxygen in rats. J Neurosurg 1996; 85:482–487. Mateescu GD, Cabrera ME. In vivo 17O magnetic resonance spectroscopy. Determination of temperature effects on metabolic rates (Q10 factor). Adv Exp Med Biol 1997; 411:585–590.

Fig. 7

ROI drawing and concentrations

CA ¼ CROI þ CROI

23



ð10Þ

Therefore, the determination of the weighting function D will enable an AIF to be determined by using Equation 4. Obtaining a corrected AIF will be a matter of determining the correct time-dependent weighting function D. In the right-hand diagram of Fig. 7, the volumes designated by 1, 2 and 3 are possible locations (due to cardiac motion) of the ‘ideal’ arterial space at three arbitrary time points during the whole scan time. Since a cardiac ROI is drawn using the summed image, this is easily the case. However, even this motion is allowed in our method as long as the ‘ideal’ arterial space remains inside the drawn ROI and the concentrations of the non-arterial space, S, in cases 1, 2 and 3 are effectively the same. Therefore, our method is less susceptible to the cardiac motion or ROI position problem.

ROI

Arterial (A) The rest (S)

A

1

2

3

S ROI = A+S

Schematic diagrams for the cardiac ROI. The cardiac ROI space is composed of the arterial space (‘A’) and the surrounding space (‘S’). 1, 2 and 3 in the right diagram show the possible locations of the arterial space inside the cardiac ROI.

Boundary conditions and constraints

It would be helpful to speculate upon the behaviour of the weighting function D at extreme conditions (t = 0 and t = 8). Since the concentration in the surrounding space will be initially zero, CS will be zero at t = 0 and, therefore, D(t = 0) = VS/VA. When t = 8, the tracer (now all the tracer is in the form of H2O) will be in equilibrium states in both spaces, and, practically, there is no space distinction (i.e., CS = CROI) since the tracer will be uniformly distributed throughout the whole ROI space (in theory, there would be a concentration difference between two spaces expected by the water partition coefficient between two spaces. However, the water partition coefficient is close to 1). This means that D(t) will asymptotically approach to zero as time t increases (i.e., D(t = N) = 0). In addition, it is not difficult to foresee that CS will always be smaller than CROI, since CS will always be smaller than CA. Based on these three conditions (D(t = 0) = VS/VA, D(t = N) = 0, CS = CROI), we can easily predict the behaviour of the weighting function as a function of time: it must continuously decrease from the initial value of VS/VA to the asymptotic value of zero. It is because of this speculation that the time-dependent weighting function in the manuscript was assumed to be an exponentially decaying function that can be specified by two parameters (initial amplitude D0 and half life Th) as in Equation 5.

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Original article

Sensitivity and positive predictive value of CT, MRI and 123 I-MIBG scintigraphy in localizing pheochromocytomas: A prospective study Franco Lumachia, Alberto Tregnaghib, Pietro Zucchettac, Maria Cristina Marzolac, Diego Cecchinc, Gaia Grassettoc and Franco Buic Aim To establish a standardized non-invasive imaging protocol for patients with pheochromocytoma undergoing surgery. Methods A series of 32 consecutive patients (16 men, 16 women; median age 43 years, range 15–71 years) with biochemically confirmed pheochromocytoma underwent computed tomography (CT) scanning, magnetic resonance imaging (MRI) and meta-[123I]iodobenzylguanidine (MIBG) whole-body scintigraphy prior to adrenalectomy or excision of extra-adrenal tumour (paraganglioma).

results with both CT and MRI, and no false positive MIBG scintigraphy, which had the highest (100%) positive predictive value. The combination of MRI + MIBG scintigraphy reached 100% sensitivity and positive predictive value. Conclusion Our data suggest that this imaging protocol should be used in all patients with biochemically confirmed pheochromocytoma. Nucl Med Commun 27:583–587

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:583–587

Results At final pathology no malignant pheochromocytomas were found. The tumour was right-sided in 16 (50%) patients, left-sided in 13 (41%), extra-adrenal (sympathetic ganglia, upper abdomen) in two (6%) and bilateral in one (3%) patient. Overall, the median greatest diameter (size) of the tumour was 35 mm (range, 15–90 mm). The sensitivity of CT, MRI and MIBG scintigraphy was 90%, 93% and 91%, and the specificity was 93%, 93% and 100%, respectively. The three patients with false negative scintigraphy had an intra-adrenal tumour, ranging from 20 to 50 mm in size. The presence of necrosis within the mass might justify the lack of significant uptake of radiopharmaceutical in two patients, and the small size (15 mm) of the mass in the other. There were two false positive

Keywords: pheochromocytoma, CT, MRI, MIBG scintigraphy, arterial hypertension a

Endocrine Surgery Unit, Department of Surgical & Gastoenterological Sciences, Radiology and cNuclear Medicine Service, Department of Diagnostic Medical Sciences, School of Medicine, University of Padua, Italy.

b

Correspondence to Dr F. Lumachi, University of Padua, School of Medicine, Endocrine Surgery Unit, Department of Surgical & Gastroenterological Sciences, Via Giustiniani 2, 35128 Padua, Italy. Tel: + 0039 049 821 1812; fax: + 0039 049 656 145; e-mail: [email protected] This paper was presented at the 85th Annual Meeting of the Endocrine Society, Philadelphia, PA, USA, 19-22 June 2003. Received 26th January 2006 Accepted 30th March 2006

Introduction

Patients and methods

Pheochromocytomas are rare neuroendocrine tumours, and one of the few causes of arterial hypertension that can be treated surgically [1]. It is estimated that 0.1% of hypertensive patients have pheocromocytomas [2]. The majority of cases are sporadic, with only 10–15% having a history of associated endocrine disorders [1,3]. In 10–15% of cases the tumours are bilateral or localized in extraadrenal sites (paragangliomas) [4]. Although the sensitivity of biochemical tests, including a 24-h urinary catecholamine and metanephrine assay, is nearly 100%, imaging techniques are necessary for localizing the tumour and, subsequently, to exclude extra-adrenal or bilateral lesions before surgery [5]. The aim of this study was to establish a standardized imaging protocol for patients with pheochromocytoma undergoing surgery.

Study population

A series of 32 consecutive patients (median age 43 years, range 15–71 years) with functioning pheochromocytomas were prospectively enrolled in the study. Written informed consent was obtained from each patient according to the full ethics approval by the institutional review approval. Table 1 reports the main clinical and biochemical data of the study population. Arterial blood pressure was recorded using an automatic device. Three recordings were made at intervals of 2–3 min the day before the operation. Eight (25%) patients had both persistent and paroxysmal hypertension. Measurement of urinary catecholamines and metanephrines was obtained using high-pressure liquid chromatography.

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584 Nuclear Medicine Communications 2006, Vol 27 No 7

Table 1 Demographics, and main clinical and biochemical data of the study population Parameter Age (years) Men/women Persistent hypertension Paroxysmal hypertension Systolic blood pressure (mmHg) Diastolic blood pressure (mmHg) 24-h urinary epinephrine (nmol
d – 1) 24-h urinary norepinephrine (nmol
d – 1) 24-h urinary metanephrines (mmol
d – 1) Number of patients

Overall 44 ± 16 16/16 27 (84%) 13 (41%) 146 ± 17 92 ± 12 122 ± 98 732 ± 568 5.2 ± 1.6 32

Normal values: 24-h urinary epinephrine = 5–110 nmol
d – 1; norepinephrine = 40–600 nmol
d – 1; metanephrines = 1.0–3.4 mmol
d – 1.

All the patients underwent computed tomography (CT) scanning, magnetic resonance imaging (MRI), and meta[123I]iodobenzylguanidine (123I-MIBG) whole-body scintigraphy prior to adrenalectomy or excision of extraadrenal pheochromocytoma (paraganglioma). Pregnancy was excluded in each premenopausal woman (n = 6) both before CT and 123I-MIBG scintigraphy. Percutaneous venous catheterization with blood sampling and norepinephrine and epinephrine measurement was not performed. Patients were carefully followed for at least 2 years after surgery, as suggested by Goldfien [2]. No relapse of the disease was observed during follow-up. Computed tomography

Helical CT examinations were carried out with a singledetector Siemens Somatom Emotion scanner. The volume was acquired with 3 mm collimation, pitch 1.5 and 3 mm reconstruction intervals. Thicker collimation values may reduce the ability to identify small lesions and still reduce the sensitivity of the CT examination. Moreover, thick slices (> 5 mm) make density measurement of small adrenal masses more difficult and not so accurate. The direct examination, before injection of contrast media, has been followed by a later one, 70 ms after injection of 100–120 ml of iodinated contrast media at a flow rate of 3 ml
s – 1. Administration of contrast media demonstrates the vascular pattern of the lesion and shows more clearly the presence of avascular or necrotic areas, and may help in differentiating between the adrenal gland and surrounding vessels. Standard parameters used were 130 kV, 150 mAs and a gantry rotation time of 0.8 s. Even if a CT scan demonstrates high accuracy in detecting adrenal pheochromocytoma, it lacks specificity. Lesions generally appear round or oval, and hyperdense after contrast media with wide non-homogeneous areas due to necrosis or haemorrhage. This pattern is common to other adrenal malignant lesions. Scattered parenchymal calcification may be observed. Magnetic resonance imaging

The MRI study was performed with 1-T Siemens Harmony equipment with a gradient system of

20 mT
m – 1 and a slew rate of 25 mT
m – 1
ms – 1. The protocol included turboflash localizer sequences of 12 s acquired without breath synchronization. Then a coronaloriented, breath-hold true fast imaging with steady-state precession (trueFISP) sequence of 12 slices (4 mm thickness) is localized on the adrenal region. Diagnostic T1 (echo time (TE) 13 ms, repetition time (TR) 384 ms) and T2 (TE 108 ms, TR 4000 ms) turbo spin echo sequences were oriented on axial planes from the diaphragm to the pelvis. A phase array body coil was used with 6–7 mm slices thickness with a gap of 0.6– 0.7 mm. Breath hold or respiratory triggered axial T2 sequences were localized on trueFISP and turboflash images, respectively. The pheochromocytomas are usually hypo-intense or iso-intense on T1-weighted and hyperintense on T2-weighted sequences. Small tumours are homogeneous, while large lesions frequently present areas of dishomogeneity within the mass due to haemorrhage or necrosis. The hypervascularity of the mass is confirmed after Ga-DTPA intravenous injection (0.1 mmol
kg – 1 at a flow rate of 2.5 ml
s – 1). However, this non-homogeneous pattern generates problem of differential diagnosis with primary or metatstatic malignant lesions. 123

I-MIBG Scintigraphy

Medications that could interfere with MIBG uptake (i.e., calcium-channel blockers, sympathomimetics, reserpine) were suspended for 5–6 days before administration of radiopharmaceuticals. Patients also received either potassium iodine or sodium perchlorate orally (1 day before tracer injection, for 7 days) with the aim of blocking the thyroid uptake of unbound iodine. Anterior and posterior whole-body planar scans containing 400–1000 kcounts per image (256  1024 matrix) were obtained 4 and 24 h after intravenous administration of 300–370 MBq of 123IMIBG. Spot images (256  256 matrix) of the suspicious areas (350–400 kcounts) were also recorded. A singleheaded, large field-of-view gamma camera (Sopha Medical DSX) equipped with a low-energy high-resolution parallel-hole collimator was used. Images of kidney using 99m Tc-DTPA were usually obtained for a better localization of the mass, at the end of the last acquisition. The intensity of tumour uptake was evaluated with regard to hepatic uptake at 24 h. Scintigrams were considered as positive when (1) adrenal uptake more intense than liver or (2) extra-adrenal focal uptake were observed. Statistical analysis

The reported data are expressed as mean ± standard deviation (SD) and comparisons between different groups were performed using two-tailed Student’s t-test or the chi-squared test, as appropriate. Pearson’s correlation coefficient (R) was used to evaluate the linear relationship between pairs of variables. A value of P < 0.05 was considered to be statistically significant. Sensitivity was defined as true positive (TP)/TP + false

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CT, MRI and

123

I-MIBG scintigraphy in localizing pheochromocytomas Lumachi et al. 585

negative (FN), and the positive predictive values (PPV) as TP/TP + false positives (FP).

Table 3

Results

CT MRI MIBG CT + MIBG MRI + MIBG

Table 2 reports the main differences between men and women. Women were significantly (P < 0.05) older than men, and their tumour was smaller. At final pathology no malignant pheochromocytomas were found. The tumour was right-sided in 16 (50%) patients, left-sided in 13 (40.6%), extra-adrenal (sympathetic ganglia, upper abdomen) in two (6.3%) and bilateral in one (3.1%) patient. Overall, the median greatest diameter (size) of the tumour measured by the pathologist was 35 mm (range, 15–90 mm). As expected, a significant (R = 0.78, P < 0.001) relationship between systolic and diastolic blood pressure was found. Moreover, there was a weak relationship between age and both systolic blood pressure (R = 0.30, P = 0.037) and urinary catecholamine levels (R = 0.34, P = 0.014), with no differences between men and women (P = NS).

Results of imaging techniques

Technique

True positive

False negative

False positive

27 28 29 30 32

3 2 3 2 0

2 2 0 0 0

Sensitivity PPV (%) (%) 90.0 93.3 90.6 93.7 100

93.1 93.3 100 100 100

PPV, positive predictive value; CT, computed tomography; MRI, magnetic resonance imaging; MIBG, meta-[123I]iodobenzylguanidine scintigraphy. Fig. 1

The results of imaging techniques are shown in Table 3. MRI had the highest sensitivity (93.7%) compared with both CT and MIBG scintigraphy, but the difference was not significant (P = NS, chi-squared test). The MRI did not show a typical signal trend in chemical shift images in two patients (Fig. 1). The three patients with false negative scintigraphy had an intra-adrenal tumour, ranging from 20 to 50 mm in size. Figs 2 and 3 show the scintigraphic pattern in patients with bilateral and extra-adrenal pheochromocytomas. The presence of necrosis within the mass might justify the lack of significant uptake of radiopharmaceutical in two patients, and the small size of the mass in the other. There were two false positive results with both CT and MRI and no false positive with MIBG scintigraphy, which had the highest PPV (100%). The combination of MRI + MIBG scintigraphy reached 100% sensitivity and PPV. Differences between the subgroups of men and women (Student’s t-test and the *chi-squared test)

Table 2

Parameter Age (years) Persistent hypertension Paroxysmal hypertension Systolic blood pressure (mmHg) Diastolic blood pressure (mmHg) 24-h urinary epinephrine (mmol
day – 1) 24-h urinary norepinephrine (nmol
day – 1) 24-h urinary metanephrines (mmol
day – 1) Side of the tumour (right/left) Extra-adrenal/bilateral Greatest diameter of the tumour (mm) Number of patients

Men

Women

P value

38 ± 14 16 (50%) 7 (44%) 149 ± 20 94 ± 13 130 ± 71 781 ± 424 5.5 ± 1.5 9/5 1/1 53 ± 27

50 ± 15 11 (69%) 6 (38%) 144 ± 16 91 ± 10 97 ± 43 568 ± 259 4.9 ± 1.7 7/8 1/0 38 ± 9

0.03 NS* NS* NS NS NS NS NS NS* – 0.04

16

16

–

(a) T2-weighted magnetic resonance image coronal plane reveals a left adrenal mass in a 45-year-old man with a biochemically confirmed pheochromocytoma. T1-weighted transverse contrast-enhanced magnetic resonance image shows central necrosis (b), while the in-phase (c) and out-of-phase (d) chemical shift images result in a doubtful signal trend.

Discussion Several studies have found that measurements of plasma or urinary catacholamines and metanephrines is the most sensitive laboratory test in the diagnosis of pheochromocytoma, reaching 100% sensitivity [5–7]. The clonidine suppression test is necessary in a few cases [8,9]. When the diagnosis has been established, the localization of the tumour is required with the aim of both facilitating its surgical removal and excluding unsuspected multicentric tumours [2]. CT, MRI and MIBG scintigraphy are the imaging procedures usually available. The sensitivity of CT and MRI ranges from 88% to 100%, and from 96% and 100%, respectively, but unfortunately both techniques lack specificity [5,10–12]. MRI is superior to CT for detecting familial and extra-adrenal pheochromocytomas, and should be the initial imaging technique in pregnant women and children [12,13]. MIBG scintigraphy has 95–100% specificity but a sensitivity lower than both CT and MRI, ranging from 80% to 100% mainly depending on the size of the tumour

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586 Nuclear Medicine Communications 2006, Vol 27 No 7

Fig. 2

gamma camera, a shorter half-life, and lack of beta emission [15,16]. Positron emission tomography (PET) with 2-[18F]fluoro2-deoxy-D-glucose (18F-FDG) represents an important method of imaging in cancer patients, especially in those with lymphomas and lung cancer, but FDG PET is not recommended as a first-line localization of pheochromocytomas [6,9,17–20]. Usually, the neuroendocrine tumours take up decarboxylate and store amino acids and their biogenic amines, and the use of 18F-labelled dihydroxyphenylalanine (18F-DOPA) PET is based on this capability [6,21,22]. Unfortunately, this radiopharmaceutical agent is more expensive than 123I-MIBG, and not yet registered in Europe for localizing pheochromocytomas. However, 18F-DOPA PET seems to be a promising imaging procedure which offers a sensitivity similar to MRI at 100% specificity [9,12,22,23].

Meta-[123I]iodobenzylguanidine (123I-MIBG) scan in a 32-year-old man with a bilateral pheochromocytoma. Planar anterior and posterior view 6 and 12 h after injection of the radiopharmaceutical.

Fig. 3

Few studies analyse the diagnostic accuracy of two or more procedures together in patients with pheochromocytoma [5,10,11,24]. CT has good sensitivity in patients with adrenal tumours, but its accuracy decreases for extra-adrenal pheochromocytomas [3,9]. MRI has similar sensitivity, and both imaging procedures have poor specificity with respect to MIBG scintigraphy [9,12,25]. In our experience the sensitivity of CT, MRI and MIBG scintigraphy was 90%, 93% and 91%, and the specificity was 93%, 93% and 100%, respectively. However, the combination of MRI and MIBG scintigraphy was 100% sensitive at a specificity of 100%. In conclusion, our data suggest that this imaging protocol should be used in all patients with biochemically confirmed pheochromocytoma.

Acknowledgement The authors are particularly grateful to Dr S. Dall’Acqua for help in preparing the manuscript and for reviewing the English.

References 1

2

3

4

Planar MIBG scintigraphic images showing an extra-adrenal pheochromocytoma in a 43-year-old woman.

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6

[10–12,14]. Compared with 131I-MIBG, better results are usually obtained using 123I-MIBG, which offers the advantages of having a more suitable energy for the

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Bentrem DJ, Pappas SG, Ahuja Y, Murayama KM, Angelos P. Contemporary surgical management of pheochromocytoma. Am J Surg 2002; 184: 621–625. Goldfien A. Adrenal medulla. In: Greenspan FS, Gardner DG (editors): Basic & Clinical Endocrinology. New York: Lange Medical Books/McGrawHill; 2001, pp. 399–421. Goldstein RE, O’Neill Jr JA, Holcomb 3rd GW, Morgan 3rd WM, Neblett 3rd WW, Oates JA, et al. Clinical experience over 48 years with pheochromocytoma. Ann Surg 1999; 229:755–766. Havlik RJ, Cahow, Kinder BK. Advances in the diagnosis and treatment of pheochromocytoma. Arch Surg 1988; 123:626–630. Wittels RM, Kaplan EL, Roizen MF. Sensitivity of diagnostic and localization tests for pheochromocytoma in clinical practice. Arch Intern Med 2000; 160:2521–2524. Brink I, Hoegerle S, Klisch J, Bley TA. Imaging of pheochromocytoma and paraganglioma. Familial Cancer 2005; 4:61–68. Sheps SG, Jiang NS, Klee GG. Diagnostic evaluation of pheochromocytoma. Endocrinol Metab Clin North Am 1998; 17:397–414. Lenz T, Ross A, Schumm-Draeger P, Schulte KL, Geiger H. Clonidine suppression test revisited. Blood Press 1998; 7:153–159.

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Pacak K, Linehan WM, Eisenhofer G, Walther MM, Goldstein DS. Recent advances in genetics, diagnosis, localization, and treatment of pheochromocytoma. Ann Intern Med 2001; 134:315–329. Berglund AS, Hulthen UL, Manhem P, Thorsson O, Wollmer P, To¨rnquist C. Metaiodobenzylguanidine (MIBG) scintigraphy and computed tomography (CT) in clinical practice. Primary and secondary evaluation for localization of pheochromocytomas. J Intern Med 2001; 249:247–251. Williams DT, Dann S, Wheeler MH. Phaeochromocytoma – views on current management. Eur J Surg Oncol 2003; 29:483–490. Pacak K, Eisenhofer G, Carrasuillo JA, Chen CC, Whatley M, Goldstein DS. Diagnostic localization of pheochromocytoma. The coming of age of positron emission tomography. Ann NY Acad Sci 2002; 970:170–176. Manger WM, Gifford Jr RW. Pheochromocytoma: current diagnosis and management. Cleveland Clin J Med 1993; 60:365–378. Maurea S, Klain M, Caraco` C, Ziviello M, Salvatore M. Diagnostic accuracy of radionuclide imaging using 131I norcholesterol or metaiodobenzylguanidine in patients with hypersecreting or non-hypersecreting adrenal tumours. Nucl Med Commun 2002; 23:951–969. Maurea S, Lastoria S, Cuocolo A, Celentano L, Salvatore M. The diagnosis of non-functioning pheochromocytoma. The role of I-123 MIBG imaging. Clin Nucl Med 1995; 20:22–24. Solanski KK, Bomanji J, Moyes J, Mather SJ, Trainer PJ, Britton KE. A pharmacological guide to medicines which interfere with the distribution of radiolabelled meta-iodobenzylguanidine (MIBG). Nucl Med Commun 1992; 13:513–521. Sisson JC, Shulkin BL. Nuclear medicine imaging of pheochromocytoma and neuroblastoma. Q J Nucl Med 1999; 43:217–223.

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Review

A meta-analysis of scintimammography: an evidence-based approach to its clinical utility Rahain Hussaina and John R. Buscombeb Background Scintimammography using 99mTc-labelled isonitriles, sestamibi and tetrofosmin, has become a mature technique in the adjunctive setting for the diagnosis of primary breast cancer. To establish an evidence base for its use, clinically, a meta-analysis was performed on both single-site and multi-centre trials performed since January 1997.

Conclusion There is evidence that this is a robust imaging technique delivering high sensitivities and specificities in patients studied in both single-centre and multi-centre trials and, as such, can be relied on as an adjunctive method for the investigation of primary breast cancer. c 2006 Lippincott Williams Nucl Med Commun 27:589–594 & Wilkins.

Methods Using an on-line literature search all such trials containing 100 or more studies were identified. To prevent double counting of patients only the last published report from any centre was used.

Nuclear Medicine Communications 2006, 27:589–594

Results A total of 2424 patients were identified in the single-site trial group, the smallest study having 105 patients and the largest 353 patients. The overall sensitivity was 85% and the specificity was 84%. In the multi-centre trial studies, published data from 3049 patients were included. The overall sensitivity in this group was also 85% and the specificity was 83%.

Introduction Scintimammography is the method by which breast pathology is identified by using a radiopharmaceutical. The agent used can be tumour specific such as 99mTcsestamibi (99mTc-MIBI) or a non-specific tracer such as 99m Tc-methylene diphosphonate (99mTc-MDP). However, as this technique is used to find breast cancer this review will only look at results of studies which are used to identify primary breast neoplasms (which may include MALT breast lymphomas). Although 201Tl has been used with some good results [1] the high radiation dose means that it is not an ideal agent to use in these patients, some of whom may not have cancer. Likewise, 99mTc-MDP has been shown to find the extent of tumour in patients in whom a breast cancer is present; it is not really tumour specific [2,3]. In almost all studies the imaging has been performed within 15 min injection of the tracer. However, bone scintigrams should not be performed on patients with no diagnosis of breast cancer and with small primaries of 50 mm or less. Therefore this technique was only used in those patients in whom the probability of extensive disease was high. Therefore we will only review those cases in which the imaging has been performed using the 99mTc-labelled isonitriles 99mTc-MIBI and 99mTc-tetrofosmin. Both these agents are licensed for identification of breast cancer in

Keywords: scintimammography,

99m

Tc-MIBI meta-analysis

a Institute of Nuclear Medicine and Ultrasound, BSM Medical University Campus, BAEC, Dhaka, Bangladesh and bNuclear Medicine, Royal Free Hospital, London, UK.

Correspondence to Dr John Buscombe, Nuclear Medicine, Royal Free Hospital, Pond Street, London NW3 2QG UK. Tel: + 0044 207 830 2470; fax: + 0044 207 830 2469; e-mail: [email protected] Received 24 February 2006 Accepted 26 March 2006

most European countries. The technique for imaging was devised and popularized by Khalkhali et al. [4,5]. In this technique the tracer is injected in the arm away from the site of the expected tumour, or the feet [6]. Within 15 min the patient is imaged by taking a prone lateral image. This is really the innovation of the approach devised by Khalkhali et al. [4,5]. The breast tissue, especially in more mature women, tends to overly the heart on the left and the liver on the right, both organs with high uptake of 99mTc-MIBI or 99mTc-tetrofosmin. Supine imaging will mean any cancer in the lower segments of the breasts may not be differentiated from underlying activity. With the patient lying on her front the breast is allowed to fall through a cut-out in a mattress. If the breast is not crimped or crushed a good quality image can be obtained with 500 000–600 000 counts in a 10-min image. To ensure no interference or cross-talk of activity, the contralateral breast can be squeezed flat against the mattress. By separating the two breasts with a lead divider, both can be imaged on a double-headed gamma camera without moving the patient. When the main image has been taken a nipple marker view can be performed and then the contralateral breast is imaged along with its nipple marker. Then, the mattress is removed and a supine image is obtained: the patient raises both arms over her head so that the axilla and axillary tail of the breasts can be seen. The whole

c 2006 Lippincott Williams & Wilkins 0143-3636

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590 Nuclear Medicine Communications 2006, Vol 27 No 7

procedure takes about 45 min. The images can then be displayed on a single sheet of X-ray film. By reviewing the literature published up to December 1999, Liberman et al. [7] produced an extensive review of the accuracy of this technique in breast cancer. These authors found 106 papers on scintimammography, and 83 of these were reviewed by using two methods. Firstly, the results of sensitivity, specificity, positive and negative predictive values, and overall accuracy were considered. The authors also scored the quality of the papers in terms of whether or not they were prospective or retrospective, the number of readers, the ‘gold standard’ used (e.g., mastectomy was better than fine-needle aspiration), although they did not include core biopsy, which has been standard practice in our centre for 10 years and is known to be more accurate than fine-needle aspirate [8,9]. The type of patients studied and the number per study were also included. Secondly, Liberman et al. [7] then graded the studies as excellent, average and poor. However, all the studies were then included in the metaanalysis. The best sensitivity and specificity was seen in those studies described as excellent or poor with the lowest in those described as average. There were only two multi-centre trials in the results and two retrospective trials; one was the European multi-centre trial and the other was a single-site study. One paper looked at recurrent cancer and another used single photon emission computed tomography (SPECT) alone. Agents other than 99mTc-MIBI and 99mTc-tetrofosmin were used in many studies; for example, 201Tl in five, 99mTc-diethylenetriaminepentaacetic acid (99mTc-DTPA) in three, 99m Tc-MDP in one, 111In-octreotide in one, 99mTc-citrate in one and antibodies in one. Only 17 studies had more than 100 patients and a sample size of 30–40 was common. With these caveats Liberman et al. [7] reported that the overall sensitivity of radionuclide breast imaging was 85.2% and specificity was 86.6%, with better figures for sensitivity occurring in those tumours stated as palpable. However, palpability had little influence on specificity. Since this last study North American and Spanish multicentre trials have been published [10,11]. Furthermore, the method of imaging based on that devised by Khalkhali et al. [4,5] has been universally accepted and forms part of published guidelines. Therefore it was felt that another review of scintimammography results would be timely, now that this additional information is available.

Methods A search of the medical literature using Pubmed was initiated using the keywords ‘breast’, ‘cancer’, ‘scintimammography’, ‘99mTc-MIBI’ (and its derivative names)

and ‘tetrofosmin’. To ensure data were acquired for centres only after the technique became mature, studies were only included if they had been published after 1997. Studies including fewer than 100 patients were excluded from the assessment. Other studies were excluded if the results did not assess the final pathology by cytology or there was no pathological assessment of tissues. To prevent a possible re-count of patients who appeared in multiple publications only the study with the greatest number of patients was chosen. Patients were only included if the aim of the study was to look for primary breast cancer. In all cases, although SPECT may have been used as part of the acquisition, the study was only included if planar imaging had been undertaken and the resulting data had been used. Only data from images performed using 99mTc-MIBI and 99mTc-tetrofosmin were included in the assessment. Results from prospective and retrospective single-site trials were included but these were assessed separately from the results arising from prospective multi-centre trials. Outside the USA few trials differentiated between palpable and non-palpable lesions and therefore results were not divided between these two groups except in the multi-centre trials when this information was available. Results were tabulated in an Excel database (Office 2000, Microsoft, Redmond, Washington, USA). A total weighted mean was calculated for sensitivity and specificity, and the positive and negative predictive values were then determined. As mammography and ultrasound were often used as the entry criteria to the study, data on these are included but they do not represent independent variables. An attempt was made to identify meta-analyses of mammography, ultrasound and magnetic resonance imaging (MRI) in the diagnosis of suspected cancer (as against screening) but no useful information could be obtained. In addition, the results of two studies performed to assess MRI in breast cancer were included partly for comparison. More studies were not used as the number of studies looking at the accuracy of MRI in a multi-centre trial and with more than 100 patients is very limited.

Results The overall sensitivity of 2424 patients undergoing scintimammographies imaged in single-centre trials was 85% and the specificity was 84% (Table 1). The range of sensitivities was 69–90% and the range of specificities was 71–94%. The majority of these trials were retrospective and many had only a single reader for the data, which may not have been read blind from the results of imaging by mammography and ultrasound.

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Meta-analysis of scintimammography Hussain and Buscombe

591

The data for the multi-centre trials show that by 2005, 3049 patients had been studied in prospective multicentre trials (Table 2). The sample sizes were larger than in single-centre trials – the smallest having 246 patients and the largest 1243. The overall sensitivity and specificity were 85% and 83%, respectively. These studies employed up to three readers, who were not provided with any clinical information. The range of sensitivities was 71–93% and range of specificities was 69–90%. There was no clear division between the sensitivities and specificities of individual studies (Fig. 1).

for palpable lesions and 0.82–0.89 for non-palpable lesions.

Although not all studies recorded palpable and nonpalpable lesions separately, when separate results had been recorded, the sensitivity of non-palpable tumours was generally lower than for palpable tumours. Also, the North American study noted no real difference between the accuracy of the technique in women with fatty or dense breasts. This study also noted a slightly better sensitivity in younger compared to older women. Also, it was the only study to look at variability in results reported by the three readers, with a recorded kappa of 0.93–0.99

Discussion

Table 1

Results of multi-centre trials*

Reference (first author and number)

Year

n

Sensitivity (%)

Specificity (%)

PPV (%)

NPV (%)

Scopinaro [12] Palmedo [13] Palpable Non-palpable Prats [10] Palpable Non-palpable Khalkhali [11] Palpable Non-palpable Sampalis [14]

1997 1998

499 246

97

61

1998

388

90 69 75 50 70 61 81 86 85 93 87

75 82 80 58

83 78 80 98

83

71

85

2000

673

2003

1243

85 71 83 30 90 94 75 80 76 47 93

3049

85

Overall

Scintimammography using 99mTc-MIBI or 99mTc-tetrofosmin appears to be a method that retains a high level of accuracy. Lesions under 10 mm may not be seen easily and this reduced sensitivity, as does some tumour types such as lobular cancers [29]. There may also be technical aspects such as the distance of the tumour from the gamma camera in medial tumours which can result in non-appearance of the cancer on the image. SPECT may help but the results are somewhat mixed with any advantage in sensitivity and specificity only being a few percent points [16,17,30]. Specificity depends on the patient population studies and may be related to the age of the patients because younger patients tend to have a high degree of highly active fibroadenomas, which can take up 99mTc-MIBI or 99mTc-tetrosfosmin [31]. However, by considering the number of studies compared and the patients in both groups, the possibility of bias selection in patients should be reduced.

10

*

All trials listed were prospective. n, number of patients with data studied; PPV, positive predictive value; NPV, negative predictive value.

Table 2

In the two MRI studies (Table 3) the study by Bluemke et al. [27] was the closest in design to the multi-centre trials developed for scintimammography: the sensitivity was 88% and specificity 68%. The MARBIS study was a little smaller and was also a screening study looking at patients with a high risk of breast cancer but there was a sensitivity of 71% and a specificity of 81%.

The multi-centre trials yielded a similar sensitivity and specificity to single-centre trials and blind reading seemed to be similar to results read unblinded. This is different from normal experience in similar trials performed with other imaging techniques [32,33]. In the North American multi-centre trial the inter-observer agreement was also measured and this was very high, with a kappa of 0.82–0.99 – again a remarkably high figure.

Results of single-centre trials

Reference (first author and number)

Year

R or P

n

Sensitivity (%)

Specificity (%)

PPV (%)

NPV (%)

Lind [15] Schillaci [16] Becherer [17] Helbech [18] Mekhmandarov [19] Arslan [20] Howarth [21] Danielsson [22] Buscombe [23] Lumachi [24] Myslivecek [25] Tiling [26]

1997 1997 1997 1997 1998 1998 1999 1999 2000 2001 2005 2005

R R R R R R R P R R P P

137 198 174 150 140 105 137 121 353 239 303 252

90 90 77 69 84 81 84 84 89 88 82 84

80 94 87 84 86 87 84 74 71 94 91 85

71 76 71 69 90 86

93 86 90 84 77 82

89 79 99

65 84 55

2424

85

84

79

80

Overall

R, retrospective; P, prospective; n, number of patients with data studied; PPV, positive predictive value; NPV, negative predictive value.

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Nuclear Medicine Communications 2006, Vol 27 No 7

Fig. 2

Fig. 1

Single-centre trial Triple assessment of examination, imaging and biopsy

Multi-centre trial 1

Sensitivity

0.8 Equivocal/ disagreement

Cancer

0.6

Benign

Scintimammography

Positive

0.4 0

0.2

0.4 0.6 1 - Specificity

0.8

Negative

1 Surgery

Follow-up

Plot of sensitivity and 1 – specificity of results in single-centre (*) and multi-centre trials (^). Suggested use of scintimammography in the diagnosis of primary breast cancer.

Table 3

Results of prospective trials using magnetic resonance

imaging Reference (first author and number)

Year

P/R

n

Sensitivity (%)

Specificity (%)

Bluemke [27] MARBIS [28]

2004 2005

P P

821 649

88 77

68 81

R, retrospective; P, prospective; n, number of patients with data studied; PPV, positive predictive value; NPV, negative predictive value.

The results of both the single-centre and multi-centre trials are almost identical to those from a previous metaanalysis performed by Liberman et al. [7] but in this previous meta-analysis only one of the studies was a prospective multi-centre trial and therefore the data provided from six multi-centre trials including over 3000 patients should provide a more accurate and robust assessment of the technique. There is some emerging information, however, as to when scintimammography will be at its most accurate if used in patients with tumours of more than 10 mm and when it should not be used in patients with lobular cancers. Therefore as the technique clearly is efficacious and can be used with confidence, when should it be used? There is some evidence from studies from the UK and Sweden that the role of scintimammography should be used as an adjunctive method to mammography and ultrasound. Combining functional and anatomical imaging will result in a sensitivity of over 90% whilst retaining a high specificity [22,23]. The adjunctive role of scintimammography, when

mammography and ultrasound are equivocal, is confirmed by the Royal College of Radiologists in their booklet advising the best role for radiology [34] and in European practice and procedure guidelines [35,36]. This can be described visually in a clinical algorithm (Fig. 2). Why then has scintimammography not become the method of choice for adjunctive imaging whilst MRI is widely used? In part this appears to be due to a lack of a prospective head-to-head trial of the two techniques allowing a true comparison. In the multi-centre trials in which results have been published the sensitivity of MRI is very similar to that of scintimammography with a lower specificity [27,28]. The failure of scintimammography to be used instead of MRI may be due to two factors. Firstly, there may be a wish to reduce the number of studies performed without ionizing radiation. Secondly, where nuclear medicine departments are not integrated into radiology there may be a reluctance to refer patients out of the department especially if this will result in a delay in patient management, which can result in financial penalties in the UK. There is little evidence for the extended role of scintimammography in identifying lymph node disease where there is a significant reduction in sensitivity and also specificity [37,38]. Therefore the use of scintimammography for axillary staging can not be recommended and sentinel node studies should be used in smaller primary breast cancers.

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Meta-analysis of scintimammography Hussain and Buscombe

There has been much interesting work using 99mTcMIBI, particularly, in the assessment and prediction of response to neoadjuvant chemotherapy [39,40]. However, although the results are interesting, they fail to provide consistency as to what change of uptake predicts and confirms response to chemotherapy. Another area where scintimammography may be of use is to look at recurrent cancer because post-surgical or postradiotherapy changes mean that anatomical methods of imaging can be of limited use. The studies performed show interesting results [41,42] but the number of patients studied remains small and therefore scintimammography cannot be recommended for this indication.

Conclusion

13

14

15

16

17

18

19

This meta-analysis shows that scintimammography with 99m Tc-MIBI or 99mTc-tetrofosmin can be recommended for use as an adjunctive method for imaging the breast when mammography and ultrasound prove inconclusive or equivocal.

21

Acknowledgements

22

Dr Rahain Hussain was supported by the International Atomic Energy Agency and the British Council.

20

23

Conflict of interest statements Dr Buscombe has received honoraria from Bristol Myers Squibb (formerly DuPont) makers of MIBI and research monies and honoraria and acted as an advisor to GE Healthcare (formerly Amersham Healthcare) makers of tetrofosmin.

24

References

25

1

Cimitan M, Volpe R, Candiani E, Gusso G, Ruffo R, Borsatti E, et al. The use of thallium-201 in the preoperative detection of breast cancer: an adjunct to mammography and ultrasonography. Eur J Nucl Med 1995; 22:1110–1117. 2 Lee JK, Jao CH, Sun SS. Technetium-99m methylene diposphonate scintimammography for evaluation of palpable breast masses. Oncol Rep 1999; 6:659–663. 3 Piccolo S, Lastoria S, Thomas R, Muto P, Apicella A, Petrosino T, et al. Scintimammography with 99mTc-MDP: experience of the National Cancer Institute of Naples. Eur J Radiol 1998; 27:S275–S281. 4 Khalkhali I, Mena I, Jouanne E, Diggles L, Benegas R, Block J, et al. Prone scintimammography in patients with suspicion of carcinoma of the breast. J Am Coll Surg 1994; 178:491–497. 5 Khalkhali I, Diggles LE, Taillefer R, Vandestreek PR, Peller PJ, Abdel-Nabi HH. Procedure guideline for breast scintigraphy. Society of Nuclear Medicine. J Nucl Med 1999; 40:1233–1235. 6 Buscombe JR. Scintimammography Imaging 2001; 13:163–170. 7 Liberman M, Samplais F, Mulder D, Sampalis JS. Breast cancer diagnosis by scintimammography: a meta-analysis and review of the literature. Breast Cancer Res Therapeutics 2003; 80:115–126. 8 Ballo MS, Sneige N. Can core needle biopsy replace fine-needle aspiration cytology in the diagnosis of palpable breast carcinoma. A comparative study of 124 women. Cancer 1996; 78:773–777. 9 Pijnappel RM, van den Donk M, Holland R, Mali WP, Peterse JL, Hendriks JH, Peeters PH. Diagnostic accuracy for different strategies of image-guided breast intervention in cases of nonpalpable breast lesions. Br J Cancer 2004; 90:595–600. 10 Prats E, Asia F, Abos DM, Villavieja L, Garcia-Lopez F, Asenjo MJ, et al. Mammography and 99mTc-MIBI scintimammography in suspected breast cancer. J Nucl Med 1999; 40:296–301. 11 Khalkhali I, Villaneuva-Meyer J, Edell SL, Connolly JL, Schnitt SJ, Baum JK, et al. Diagnostic accuracy of 99mTc-sestamibi breast imaging: Multi-center trial results. J Nucl Med 2000; 41:1973–1979. 12 Scopinaro F, Schillaci O, Ussof W, Tiberio NS, De Vincentis G, Mezi S, et al. A three-center study on the diagnostic accuracy of 99mTc-MIBI scintimammography. Anticancer Res 1997; 17:1631–1634.

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Taillefer R, Robidoux A, Turpin S, Lambert R, Cantin J, Leveille J. Metastatic axillary lymph node technetium-99m-MIBI imaging in primary breast cancer. J Nucl Med 1998; 39:459–464. Mankoff DA, Dunnwald LK, Gralow JR, Ellis GK, Drucker MJ, Livingston RB. Monitoring the response of patients with locally advanced breast carcinoma to neoadjuvant chemotherapy using [technetium 99m]-sestamibi scintimammography. Cancer 1999; 85:2410–2423.

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Buscombe JR. Monitoring therapy in breast cancer. Nucl Med Commun 2002; 23:619–624. Katayama N, Inoue Y, Ichikawa T, Harada M, Itoh T, Shimoyamada K, et al. Local recurrence of breast cancer demonstrated on 99mTc-MIBI scintigraphy. Radiat Med 1999; 17:389–391. Cwikla JB, Kolasinska A, Buscombe JR, Hilson AJ. Tc-99m MIBI in suspected recurrent breast cancer. Cancer Biother Radiopharm 2000; 15:367–372.

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Correspondence

Correspondence Nuclear Medicine Communications 2006, 27:595–596

90

Y-Ibritumomab tiuxetan: new drug, interesting concept, and encouraging in practice Andreas Otte Center of Clinical Trials, University Hospital Freiburg, Germany. Correspondence to Prof. Dr Andreas Otte, Center of Clinical Trials, University Hospital Freiburg, Elsa¨sser Str. 2, D-79110 Freiburg, Germany. e-mail: [email protected]

From the perspective of a nuclear medicine physician, I have noticed with great interest the launch of 90Ylabelled ibritumomab tiuxetan (Zevalin; Biogenldec, San Diego, USA, and Schering AG, Berlin, Germany) for the treatment of patients whose CD20 þ follicular/low-grade or transformed non-Hodgkin’s lymphoma (NHL) has failed to respond to chemotherapy and rituximab. So far, more than 6000 patients have been treated in over 30 countries. In addition, clinical studies are being conducted using Y-labelled ibritumomab tiuxetan as front-line therapy and in relapsed disease for indolent and aggressive NHL, relapsed diffuse large B-cell lymphoma, mantle cell lymphoma, paediatrics, multiple myeloma, primary CNS lymphoma, and for Hodgkin’s disease. 90

In 2002, Witzig et al. [1] demonstrated that radioimmunotherapy with 90Y-ibritumomab tiuxetan produces statistically and clinically significant higher overall response rates (ORRs) and complete response rates (CRRs) as compared to standard immunotherapy using rituximab [1]: ORRs were 80% for the 90Y-ibritumomab tiuxetan group versus 56% for the rituximab group (P ¼ 0.002), and CRRs were 30% in the 90Y-ibritumomab tiuxetan group versus 16% in the rituximab group (P ¼ 0.04). Reversible myelosuppression was the primary toxicity noted with 90 Y-ibritumomab tiuxetan in this study. The non-comparative trials by Witzig et al. [2] and Wiseman et al. [3] were also encouraging: the ORR for the 54 patients with follicular NHL in the study by Witzig et al. [2] was 74% (15% complete responses and 59% partial responses), and the time to progression estimated by the Kaplan–Meier method was 6.8 months for all patients and 8.7 months for responders. Adverse events were primarily haematological and reversible. In the study by Wiseman et al. [3] with a

reduced-dose 90Y-ibritumomab tiuxetan treatment in 30 patients with relapsed or refractory NHL and mild thrombocytopenia, ORR was 83% (37% complete response, 6.7% complete response unconfirmed, and 40% partial response), and the time to progression estimated by the Kaplan–Meier method was 9.4 months in all patients and 12.6 months in responders. Also in this study, toxicity was primarily haematological, transient and reversible. Furthermore, in updated evaluations (as published by Gordon et al. [4]), with a median follow-up of 44 months, the data are even more mature than in the aforementioned studies from 2002, since all ongoing patients of both groups exceeded the median Kaplan– Meier estimated time to progression (TTP), duration of response (DR), and time to next therapy. Although this comparative study was not powered to detect differences in time-to-event variables, the updated results demonstrate clear trends toward longer median TTP (15 vs. 10.2 months; P ¼ 0.07), DR (16.7 vs. 11.2 months; P ¼ 0.44) and time to next therapy (21.1 vs. 13.8 months; P ¼ 0.27) in follicular NHL patients treated with Zevalin compared with the rituximab control arm. In patients achieving a complete response/complete response unconfirmed (CR/ CRu), the median TTP was 24.7 months for patients treated with Zevalin compared with 13.2 months for rituximab-treated patients (P ¼ 0.41), and ongoing responses of 45 years have been observed. Non-haematological toxicity after Zevalin and rituximab is comparable, and in the vast majority mild or moderate. Only the haematological toxicity is more expressed in Zevalin patients when compared to rituximab. It is not unusual that more effective regimens (as in the comparative trial with Zevalin) are mirrored by a more expressed side-effect profile. However, the haematological toxicity is predictable and easy to manage (e.g., with platelet transfusions). The handling and administration of 90Y-labelled ibritumomab tiuxetan are subject to qualified personnel in an authorized healthcare establishment; normally, this is done by a nuclear medicine physician and for this discipline applications like 90Y-labelled ibritumomab tiuxetan are no more complex than any other routine application. The study and market

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596 Nuclear Medicine Communications 2006, Vol 27 No 7

experience with Zevalin shows that ‘complex logistics’ quickly turn into routine after the first patient has been treated. In summary, the existing data on 90Y-ibritumomab tiuxetan encourage the use of this new treatment option in NHL, as it is safe and well tolerated and has significant clinical activity in this patient population.

2

3

4

References 1

Witzig TE, Gordon LI, Cabanillas F, Czuzman MS, Emmanouilides C, Joyce R, et al. Randomized controlled trial of yttrium-90-labeled ibritumomab

tiuxetan radioimmunotherapy versus rituximab immunotherapy qfor patients with relapsed or refractory low-grade, follicular, or ransformed B-cell nonHodgkin’s lymphoma. J Clin Oncol 2002; 20:2453–2463. Witzig TE, Flinn IW, Gordon LI, Emmanouilides C, Czuzman MS, Saleh MN, et al. Treatment with ibritumomab tiuxetan radioimmunotherapy in patients with rituximab-refractory follicular non-Hodgkin’s lymphoma. J Clin Oncol 2002; 20:3262–3269. Wiseman GA, Gordon LI, Multani PS, Witzig TE, Spies S, Bartlett NL, et al. Ibritumomab tiuxetan radioimmunotherapy for patients with relapsed or refractory non-Hodgkin lymphoma and mild thrombocytopenia: a phase II multicenter trial. Blood 2002; 99:4336–4342. Gordon LI, Witzig T, Molina A, Czuczman M, Emmanouilides C, Joyce R, et al. Yttrium 90-labeled ibritumomab tiuxetan radioimmunotherapy produces high response rates and durable remissions in patients with previously treated B-cell lymphoma. Clinical Lymphoma 2004; 5:98–101.

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NEWS AND VIEWS JULY 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up-to-date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 206 6930, E-mail: [email protected]

Nuclear Medicine Communications, 2006, 27, 597–599 BNMS spring meeting 2006

Once again the decaying– rejuvenating industrial splendour of Manchester was the venue for this year’s principal meeting of the BNMS. As in recent years, the presentations were dominated by PET papers, although cardiology and endocrinology were also well represented. An interesting twist to the PET offerings was that tracers other than 18F came to prominence. Indeed the Annual Lecture by Mathew Thakur gave us an exciting insight into his work with 68Galabelled molecules in imaging oncogenes and peptides overexpressed in cancer cells. In the cardiac sessions, the Newcastle team gave us food for thought about the parameters we should report from gated SPECT studies, and the Royal London team showed very convincing evidence that a large number of patients unable to reach target heart rate went on to have positive perfusion studies, indeed a much higher proportion than amongst those that did reach target. With the NPfIT deployment of PACS and RIS now in full swing it was inevitable that these subjects would assume greater importance, and Danny McCool gave a thorough, if rather daunting, invited review on the challenges ahead. Finally, the exhibition provided evidence that

gamma cameras now come in all shapes and sizes from those designed for cardiac with ever decreasing footprints, to those combining modalities. All the major manufacturers now offer SPECT–CT but there are major differences between the trade-offs that they make between size and CT quality. The first commercial PET scanner incorporating time-of-flight into the algorithm was also unveiled, showing impressive improvements in the image quality of clinical images. The format of the meeting was slightly altered, concluding the meeting with lunch on the final day. This earlier finish (possibly combined with the withholding of CPD certificates until lunchtime) resulted in a much increased audience for the excellent highlights lecture given by Adil Al-Nahhas. There were also an increased number of prizes awarded, mostly sponsored by industry, and it is hoped that this will act as an incentive to encourage submission of papers and posters in future meetings. Details of those who won prizes may be found below. This year was the last year that the meeting was organised by Professor Malcolm Frier as chair of the scientific committee. He has done a wonderful job and the BNMS is

most grateful for all his hard work. Professor David Williams has now stepped into the role and he would be very interested to hear from anyone who has suggestions about the content or format of the meeting in the future. Prize-winners at the BNMS spring meeting 2006

Oral presentations 1st Sponsored by Philips Medical Systems Radio-immunotherapy in advanced pancreatic cancer. Is it feasible? (Paper 23) A. Sultanaa, R. Jayanb, S. Chauhana, S. Shorea, J.C. Evansc, C. Garveyb, P. Ghaneha, S. Vinjamurib, J.P. Neoptolemosa a Division of Surgery and Oncology, and Departments of bNuclear Medicine and cRadiology, Royal Liverpool University Hospital 2nd 68Ga DOTATE PET–CT in primary and recurrent neuroendocrine tumours (Paper 26) I. Kayani, A.M. Groves, S.M. Gacinovic, P.J. Ell, I. Bomanji The Institute of Nuclear Medicine, UCLH 3rd Usefulness of pre-operative MIBI dual phase scanning in patients with primary hyperparathyroidism (Paper 6) N. Hamilton, G. Ellul, G. Al-Bahrani Manchester Royal Infirmary

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598 Nuclear Medicine Communications 2006, Vol 27 No 7

BNMS young investigator Simple scatter correction methods for gamma camera images: do they work? (Paper 18) A. Wright, S. Chandler Darlington Memorial Hospital Siemens multimodality oral presentation The direct use of combined 18F-FDG PET–CT images for radiotherapy treatment: Planning for patients with non-small cell lung cancer (Paper 63) K. Carsona, V. Cosgroveb, A. Zatarib, R. Eakinc, J. Clarked, D. Stewardc, J. McAleesec, L. Flemingc, A. b a Hounsell , P. Jarritt a NI Regional Medical Physics Agency, Nuclear Medicine, Royal Victoria Hospital, Belfast, bNI Regional Medical Physics Agency, Radiotherapy, NI Cancer Centre, c Department of Oncology, NI Cancer Centre, and dRadiology Department, Royal Victoria Hospital, Belfast Siemens multimodality poster Does respiratory effort have any significant influence on the spatial registration of PET/CT on lung lesions? (P12) H.K. Cheow, A. Winship, S. Rankin, D. Landau, M.J. O’Doherty Technologist (oral or poster)

1st Sponsored by Tyco Healthcare 68 Ga-DOTATOC PET–CT: A technologist’s perspective (Paper 29) G. Heath, P. Blanchard, J. Dickson, I. Kayani The Institute of Nuclear Medicine, UCLH 2nd A study to assess the feasibility of introducing radiographer reporting in a nuclear medicine department (Paper 37) K. Custis Princess Royal University Hospital, Bromley, Kent 3rd How should we measure the diagnostic accuracy in reporting? (Paper 32) K.G. Holmesa, M.E. Welshb a St Martins College Lancaster, b Morecambe Bay Hospitals Trust

Posters 1st Sponsored by GE Healthcare Bone mineral density in children (P39) D.E. Simpson, S. Stevens, V.S. Dontu, V. Low, L.J. Archbold, H. Bashir, M.J. O’Doherty, N. Marin, A.J. Coakley East Kent Hospitals NHS Trust, Kent and Canterbury Hospital 2nd Cardiac volume determination using tomographic equilibrium radionuclide angiography with threshold edge detection (P9) I.P. Clements, B.P. Mulla, J.F. Breen, C.G. McGregor Mayo Clinic, USA 3rd Audit of paediatric DMSA scanning: Are we still scanning too many too prematurely? A local experience N.R. Jefferson, E.J. Owens, D. Dunlop Royal University Hospital, Bath Student prizes 1st Computer simulation of gamma camera images of the kidney (Paper 19) A.H. Dawson, J.S. Fleming, S.M.A. Hoffmann, L. Papspyrou, S. Peel Southampton General Hospital 2nd Comparison of radioiodine with radioiodine plus lithium in the treatment of hyperthyroidism (Poster 1) K. Ahmed, G.M.K. Nijher, A. Banerjee, J. Frank, K. Meeran Charing Cross Hospital 3rd Modelling and correcting for respiratory motion in PET (Paper 17) N.S. Vyas, P. Schleyer, D.L.G. Hill, P.K. Marsden Guy’s, King’s, St Thomas’ and UCL, London School prizes 1st Matthew Kasoar – Wilson’s Grammar School, Wallington Runners up Michal Kwasigroch – Altrincham Grammar School for Boys Natalia Tereshchenko – Tormead School, Guildford Ariadne Whitby – Wimbledon High School for Girls

Patient information leaflets 1. ‘Paediatric scans’, Barking, Havering and Redbridge NHS Trust 2. ‘Children’s scans’, Royal Devon and Exeter Healthcare NHS Trust 3. ‘Your visit – Nuclear Medicine Department’, North Glasgow University Hospitals Division, Glasgow Royal Infirmary 4. ‘Radio-iodine therapy for thyrotoxicosis’, Central Manchester Children’s University Hospitals NHS Trust Meeting Announcements

BNMS Autumn Meeting Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk EANM Annual Meeting Dates: 30 September to 4 October 2006 Venue: Athens, Greece Website: www.eanm.org 9th World Congress of Nuclear Medicine and Biology Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/congress 2006/index02.htm International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM) Dates: 13–15 November 2006 Venue: Vienna, Austria Website: www-pub.iaea.org/MTCD/ Meetings/Announcements.asp?ConfID = 146 Design of Radionuclide Facilities with Reference to Radiation Protection (Revisited) Date: 14 November 2006 Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT Website: www.bir.org.uk Education and Training

EANM learning courses Dates: Weekend courses throughout 2006

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Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected]

EANM distance learning in Nuclear Cardiology This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical ele-

ments needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications.

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Abstracts

Abstracts for the Autumn Meeting of the British Nuclear Medicine Society Glasgow University, UK, 8–9 September 2005

OPENING LECTURE Nuclear medicine in drug delivery development Clive G. Wilson Strathclyde University, Glasgow, UK. Although many chemicals bind to drug receptors and trigger or block transmitter action, all such ligands are not drugs or medicines. This is because a useful drug must exert an action generally in a reversible or least controllable extent (efficacy) with a minimum of unwanted side-effects. If introduced directly into the body a lack of efficacy may be explained by poor exposure and may result from hepatic or renal clearance; a side-effect by the rapid upward swing of plasma concentrations providing diffusional drive. For other non-systemic routes (i.e. as by mouth) there are additional parameters to consider; in particular, release of the drug, as appropriate control of dissolution helps to provide more predictable concentrations. This, however, is only one of the possible improvements that could be made, as directed exposure increases efficacy whilst reducing risk. Drug targeting allows the use of more potent drugs with an increased margin of safety but sampling of body fluid provides the vaguest of indications of the processes which are competing to absorb, distribute and remove the drug. What is termed as a ‘medicine’ is generally a formulation of a drug. This formulation can be tested in vitro to ensure quality control using fluids which mimic the contents of the gastrointestinal lumen and apparatus which mimics peripheral and deep lung disposition. Whilst such systems provide comfort for the manufacturing chemist, the performance outside the body may be a poor mimic of what happens within. The growth of non-invasive imaging techniques, particularly planar scintigraphy, single photon emission computed tomography (SPECT) and positron emission tomography (PET) have added to the library of techniques that nuclear medicine has provided to the pharmaceutical scientist. In particular, patient variability becomes a tool that explains the overall pattern of drug disposition because the relationship between the distribution or anatomical location of the dose and the effect or plasma concentration–time profile can be understood. It is perhaps no exaggeration that the gamma camera has been the most important analytical instrument in the development of sophisticated medicines, increasing the utilization of compounds and helping us to understand the relationship between physiology and physicochemical processes. SENTINEL NODE SESSION A1 SLN localization: a surgeon’s perspective A. McKay Glasgow, UK.

A2 A practical guide to SLN imaging and localization W. Waddington London, UK. A3 Early experience of lymphatic mapping in vulvar malignant melanoma in comparison with previous studies I. Heinle, Q. Siraj and J. Hurren Departments of Nuclear Medicine and General Surgery, Portsmouth Hospitals NHS Trust, UK. Background The poor prognosis of the relatively rare vulvar malignant melanomas is mainly related to the tumour thickness and the status of lymph nodes. There are many controversies about the therapeutic management of these tumours and there is an ongoing debate on the value of radical vulvectomy with complete lymphadenectomy, despite the considerable morbidity, versus the less extensive surgical procedure and elective groin dissection with an associated high risk for locoregional recurrence. Aim To report our early experience of lymphatic mapping for identifying the sentinel lymph nodes in cases of vulvar malignant melanoma and to review the literature. Methods All patients had no palpable lymphadenopathy. Interstitial vulvar injection of 99mTc-nanocolloid was followed by dynamic and static imaging, with subsequent skin marking of the position of sentinel nodes identified. Intraoperative gamma probe and patent blue dye were used to identify the nodes draining the tumour site. We have also reviewed the medical literature on lymphatic mapping in vulvar malignant melanomas. Results and conclusion Sentinel lymph nodes were successfully localized in all patients. The technique appears to be effective in tumour staging and surgical management. A4 Sentinel node imaging: a drain on resources? A. Burns, R. Bury and M. Waller Leeds Teaching Hospital Trust, UK. Background We have faced increasing demands for sentinel node (SLN) imaging from our very busy breast cancer service. Implementation of the full BNMS protocol for SLN imaging made coping with the workload difficult, so we devised an abbreviated version of the protocol which allowed us to provide all the information required by the surgeons without overloading our resources. Methods As visualization of lymphatics and nodes is rapid following intradermal injection, we stop the dynamic acquisition as soon as a lymph node is visible, sometimes less than a minute after injection. We only perform one set of static images. If no node is visible at the end of the dynamic run, the patient is sent

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A2 Nuclear Medicine Communications 2006, Vol 27 No 7

away for an hour, or until a camera slot is available, and it is very unusual to fail to visualize a node or nodes at this stage. We acquire transmission and emission images simultaneously, usually achieving adequate results within 2–3 min. The breast surgeons we work with do not find skin marking useful, so we have discontinued this practice. Results To date, there have been no failures to demonstrate nodes on imaging, or to find them in theatre, and we have been able to accommodate the increased workload with no increase in resources. This claim will be supported by audit data. CARDIOLOGY A5 Image assessment of hibernating myocardium S. Woldman Ayr, UK. A6 Development of a myocardial perfusion imaging service in a district general hospital using the VG Hawkeye gamma camera H. Blundell, S. Hutchinson, A. Lee and L. Griffiths Nevill Hall Hospital, Gwent Healthcare NHS Trust, Abergavenny, UK. Objectives To show how a myocardial perfusion imaging service has been developed at Nevill Hall Hospital, Abergavenny, using the VG Hawkeye gamma camera, incorporating ECG gating and attenuation correction. Methods One hundred and fifty patients (79 male, 71 female) have undergone stress/rest myocardial perfusion imaging using 99m Tc-MIBI and a 2-day protocol. The first 80 patients were imaged using a non-gated acquisition protocol and sequential CT Hawkeye attenuation correction (AC). A gated protocol was then introduced at stress and rest in addition to AC. Results Using AC alone, overall image quality increased for obese patients. Counts were increased in male patients in regions of suspected inferior wall attenuation and in some female patients in regions of suspected anterior wall breast attenuation. However, mal-registration of the emission and transmission data caused anterior artefacts to be generated on either the stress or rest image in 29% of patients. Gating the emission images allowed wall motion and thickening to be observed, so differentiating between true perfusion defects and artefacts. Conclusions The combination of gated acquisition and attenuation correction, whilst increasing imaging time from approximately 12 to 20 min, provides valuable functional information and greatly improves clinical confidence compared with no correction or attenuation correction alone. A7 Feasibility of gated SPECT myocardial perfusion scintigraphy using 201Tl G.A. Wright, A.C. Tweddel, G. Avery and G. Davies Departments of Nuclear Medicine and Cardiology, Hull & East Yorkshire Hospitals, UK. Background The choice of perfusion agent for myocardial perfusion scintigraphy remains controversial. One generally accepted advantage of 99mTc perfusion agents is the ability to perform gated SPECT (G-SPECT). This is considered impractical with 201Tl. Aim To investigate the feasibility of G-SPECT using 201Tl.

Methods Twenty-five patients underwent stress–redistribution G-SPECT imaging with 201Tl. Patients with body weight < 90 kg (mean, 67 ± 10 kg) were selected. Mean administered activity was 80 ± 5 MBq. Images were acquired on a GE Infinia gamma camera with LEGP collimators. Imaging time was 15 min. Perfusion and wall motion (WM) quality were scored using 5-point scales, higher scores indicating better quality. Net left ventricular (LV) counts were assessed using the summed LAO40 projection of the SPECT data. These were compared to LV counts from 20 tetrofosmin studies in patients of similar weight (mean, 67 ± 9 kg), acquired for 10 min with LEHR collimators. Results Mean quality scores at stress were: perfusion 4.5 ± 0.7, WM 4.6 ± 0.6; at redistribution: perfusion 3.8 ± 0.7, WM 3.5 ± 0.5. No studies were considered unacceptable for assessment of WM. Mean LV counts at stress were slightly higher for 201 Tl studies (3640 ± 1343 vs. 3244 ± 1429, P = 0.4) despite administered activities of 5–6 times higher for tetrofosmin (P = 0.002). Conclusion Good quality G-SPECT images can be readily obtained with UK administered activities of 201Tl in patients under 90 kg. A8 A short pilot study into the feasibility of mycocardial viability assessment by gamma camera PET F.I. McKiddiea, H.G. Gemmella, E.J. Davidson and M.J. Metcalfeb Departments of aNuclear Medicine and bCardiology, Aberdeen Royal Infirmary, UK. Background 201Tl myocardial perfusion imaging involves high radiation dose to the patients and produces low count images. This study was undertaken to assess whether an FDG/NH3 gamma camera PET protocol could replace thallium imaging for assessment of myocardial viability. This would resolve these problems, remove the need for stressing and halve the time required for imaging from 4 to 2 h. Methods Ten patients who had undergone thallium imaging volunteered for FDG/NH3 imaging. All patients produced adequate quality NH3 images. The quality of the FDG images was more variable. Six patients produced good quality images and three produced adequate images for quantitative analysis. One patient produced FDG images which were so poor as to be unusable. This most likely indicates variability of the blood glucose levels at the time of injection. Results Quantitative analysis of the thallium and FDG/NH3 images used a 70% threshold between normal and abnormal perfusion and metabolism. In nine patients the analyses of the thallium and FDG/NH3 images showed good agreement. In the patient where there was a disagreement the FDG/NH3 analysis demonstrated a larger area of viability than with thallium. Conclusion This preliminary work has shown that the proposed protocol is practicable and produces data of sufficient quality for analysis. However, more attention must be paid to the monitoring of glucose levels prior to FDG imaging. A9 Myocardial perfusion imaging (MPI) in preoperative evaluation for non-cardiac surgery in a district hospital S. Hana, M.H. Tuna, C. Hosieb, A.C. Pella and M.C. Vishua

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BNMS meeting: Abstracts A3

Departments of aCardiology/General Medicine and bNuclear Medicine, Monklands District Hospital, Airdrie, UK. Introduction Effective preoperative evaluation is vital to minimize perioperative morbidity and mortality. We present our experience of MPI in preoperative fitness assessment for elective non-cardiac surgery. Patients and methods Thirty patients (16 male, 14 female; age, 48–75, mean, 68 years) who had preoperative SPECT MPI during 2003–2004 were retrospectively reviewed. All patients had intermediate or major cardiovascular risk and were planned for intermediate or high-risk operations. Exercise tolerance was poor in the majority. Results Twelve patients had normal MPI. Ten of the 12 (83%) were recommended for surgery (7 had uncomplicated operations and 3 were on the waiting list). Two were treated conservatively. Eight patients had low-risk MPI. Six of these 8 (75%) were offered operations. Two of the 8 proceeded to coronary angiogram, which confirmed MPI results. Three had uneventful operations and 4 were on the waiting list. One patient chose conservative treatment. Ten patients had intermediate–highrisk MPI. Coronary angiogram was recommended in 6 and revascularization in three. Three of the 10 (30%) have been offered operations (2 underwent uncomplicated operations and 1 was on the waiting list). Two of the 10 were having a further cardiology review. Five of the 10 were treated conservatively. Conclusion MPI provides reliable non-invasive preoperative cardiovascular risk stratification for non-cardiac surgery. Normal or low-risk MPI indicates minimal perioperative complication whereas an intermediate–high-risk scan merits further evaluation. A10 Is there a difference between myocardial meta[123I]iodobenzylguanidine uptake in patients with chronic heart failure of ischaemic versus non-ischaemic aetiology? D.W. Motherwell, A.M. Fletcher, A.D. Small, G. McCurrach, W. Martin and S.M. Cobbe Royal Infirmary, Glasgow, UK. Background meta-[123I]iodobenzylguanidine (123I-MIBG) uptake by the heart is a marker of uptake-1 function and noradrenaline re-uptake in the sympathetic nerve terminal. Patients with chronic heart failure (CHF) have reduced global MIBG uptake as measured by heart to mediastinum ratio. Aim To assess MIBG uptake in patients with ischaemic and nonischaemic CHF and similar degrees of left ventricular dysfunction. Methods MIBG uptake (185 MBq) and radionuclide ventriculography (RNVG) were performed in 20 patients with clinical evidence of CHF, and objective evidence of left ventricular dysfunction. Aetiology of CHF was determined from the coronary arteriogram, myocardial perfusion scintigraphy and clinical history. Analysis of MIBG uptake allowed calculation of the heart to mediastinal ratio (H:M) from early images at 10 min. Left ventricular ejection fraction was obtained from the RNVG study. Results Thirteen patients were found to have an ischaemic aetiology (group 1), and 7 patients a non-ischaemic aetiology (group 2). There was no significant difference between the myocardial uptake of MIBG in group 1 (range 1.86–2.86) and

group 2 (range 2.61–3.21). LVEF ranged from 14 to 39 in group 1 and 10 to 32 in group 2. There was no significant difference in ejection fractions between the two groups. Conclusion Patients with CHF of ischaemic and non-ischaemic aetiology and comparable LVEF have a similar MIBG uptake. A11 Stroke index, ejection fraction and cardiac index from gated SPECT MIBI scans in symptomatic patients P.J. Wheeler, J.P. Coffey and J.C. Hill Department of Nuclear Medicine, Royal Preston Hospital, Lancashire Teaching Hospitals NHS Trust, Fulwood, Preston, UK. Background Stroke index (SI), defined as ratio of stroke volume (SV) to body surface area, is a recognized parameter in evaluating left ventricular function. Aim To record SI from gated SPECT studies and to correlate with cardiac index, CI (cardiac output (CO)/ body surface area) and compare with ejection fraction (EF). Method Seventy symptomatic patients (mean age 57 years; 35 male) were studied prospectively in an outpatient setting. Following a post-stress injection of 450 MBq 99mTc-MIBI, a gated SPECT study was performed and SV, EF and average resting heart rate were recorded. SI and CO (heart rate  SV) were calculated together with CI. Results Mean SI was 28.5 ml
m – 2, (SD 7.46; max SI 44 ml
m – 2). Eighteen (25%) patients had abnormal SI (defined as less than 25 ml
m – 2) whereas only 7 (10%) had abnormal ejection fraction below 45%. Forty-four (62.8%) patients had abnormal CI defined as less than 2.5 l
min – 1
m – 2. Correlation of SI with CI was 0.57, while correlation of ejection fraction with CI was 0.27. Conclusion Stroke index is readily determined from gated SPECT perfusion data, correlates with cardiac output/index and when reduced, may be a more sensitive indication of ventricular dysfunction than ejection fraction. TECHNOLOGISTS TRAINING SESSION: THINK ABOUT WHITE CELL IMAGING A12 White cell labelling: How do you do yours? A. McCartney Department of Nuclear Medicine, Glasgow Royal Infirmary, UK. A13 White cell imaging: a nursing perspective including a paediatric service G. McEwan Department of Nuclear Medicine, Glasgow Royal Infirmary, UK. A14 White cell imaging: Problems RADIOBIOLOGY AND THERAPY/PHYSICS A15 Cell signalling, genetics and the in-vivo responses to ionizing radiation E. Wright Dundee, UK. A16 Randomized trial of high-dose vs. low-dose radioiodine, with or without rhTSH, for remnant ablation in differentiated thyroid cancer A. Nicol, on behalf of NCRI Thyroid Cancer Group Victoria Infirmary, Glasgow, UK.

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A4 Nuclear Medicine Communications 2006, Vol 27 No 7

Thyroid cancer is the most frequently occurring malignant endocrine tumour. Radioiodine is used to ablate residual thyroid tissue following thyroidectomy for differentiated thyroid cancer. However, there is controversy over the activity of radioiodine that is used. A multicentre trial is presented that will compare low activity (1.1 GBq) with high activity (3.7 GBq) 131I for remnant ablation. Four hundred and sixty-eight patients will be included in the trial. Patients will be randomized to receive low or high activity. Further randomization will be performed where patients discontinue thyroid hormone replacement or receive recombinant human thyroid stimulating hormone (rhTSH). The aims of the trial are to show that (1) a low dose of radioiodine has a similar remnant ablation success rate as a high dose; and (2) patients given rhTSH have a similar ablation rate to those who discontinue thyroid hormone replacement. The success of ablation will be determined 8 months after treatment using a 131I diagnostic scan and measurement of thyroglobulin. Recruitment is expected to start in January 2006 and last for 3 years. A17 Developing services for thyroid cancer patients who require radioactive iodine ablation (131I): the impact of user involvement I. Driver, M. Vincent, K. Farnell and J. Sinclair Newcastle General Hospital, UK. Thyroid cancer is the most common endocrine malignancy in the UK with 0.7 per 100 000 men and 1.9 per 100 000 women diagnosed with the disease every year. The most common treatment is surgery (thyroidectomy) with a high proportion of patients requiring 131I to reduce the risk of local recurrence and improve survival [1]. The disease is associated with significant physical and psychological morbidity and this is exacerbated by short-term radiation protection issues associated with the administration of 131 I. A specific area of concern for patients revolves around the requirement to be nursed in a shielded room for several days with minimal contact with staff and visitors [2]. The central theme underpinning the reform of health care services in the past decade has been the emphasis on consulting patients, involving them directly in the shaping and redesigning of local services [3,4]. Our aim was to provide an insight into how user involvement could be used to facilitate service modifications. We audited access to patient information and support and undertook a service review in collaboration with all key stakeholders. Changes were then made in the following areas: Improved treatment environment and visiting Assessment of risk for patients who are nursed by ‘neglect’ K Improved education and training for all staff groups K Improved access to patient information and support services, including the inception of the first UK support group for thyroid cancer patients, Butterfly NorthEast. K K

References 1. Northern Cancer Network. Northern Cancer Network Guidelines: Thyroid Cancer 2000. Available on http://www.cancernorth. nhs.uk/

2. Stajduhar KI, et al. Thyroid cancer: patients’ experiences of receiving iodine-131 therapy. Oncol Nurs Forum 2000; 27:1213–1218. 3. Department of Health. The NHS Plan: A Plan for Investment, a Plan for Reform. 2000. Available on http://www.dh.gov.uk/Publications AndStatistics/Publications/PublicationsPolicyAndGuidance/ PublicationsPolicyAndGuidanceArticle/fs/en?CONTENT_ID = 4002960&chk = 07GL5R 4. Department of Health. The NHS Cancer Plan: A Plan for Investment, a Plan for Reform. 2000. Available on http://www. dh.gov.uk/PublicationsAndStatistics/Publications/Publications PolicyAndGuidance/PublicationsPolicyAndGuidanceArticle/fs/ en?CONTENT_ID = 4009609&chk = n4LXTU A18 The value of a 123I thyroid uptake study in the management of subclinical hyperthyroidism and other thyroid disorders F. Sundrama, P. Deb and A. Notghia Departments of aNuclear Medicine and bEndocrinology, City Hospital, Birmingham, UK. Aim To determine the value of 123I thyroid uptake studies in the management of subclinical hyperthyroidism (SCH) and other thyroid disorders, particularly whether it influenced therapeutic decisions. Methods A retrospective review of uptake studies over a 3-year period. Initial diagnoses and outcomes were noted. Uptake values at 24 h were classified as high uptake (HU), normal uptake (NU) or low uptake (LU). Results Fifty-three studies were performed. There were 19 HU, 22 NU and 12 LU. Eleven patients had SCH. Of 4 HU, 2 had radioiodine and 2 no therapy. Of 5 NU, 2 had antithyroid therapy and 3 no therapy. Two had LU; both had no therapy. Forty-two other diagnoses included 13 multinodular goitres (2 HU had radioiodine, 8 NU, 3LU), 9 Graves’ disease (all HU), 6 toxic adenomas (3 HU; 1 had surgery, 2 antithyroid therapy and 3 NU; 2 had radioiodine, 1 no therapy), 6 thyroiditis and 8 various other thyroid disorders. Overall, 11/53 (21%) of patients received therapy aided by uptake values. Conclusion In SCH, appropriate therapy can be considered for patients with high or normal thyroid uptake values. In multinodular goitres, uptakes identify hot nodules suitable for therapy. A19 Issues associated with 90Y-Zevalin therapy T. Murray, G.J. Gillen, A.T. Elliott and N. O’Rourke Western Infirmary, Glasgow, UK. Aim To report on our experience of radiochemical purity and radiation protection issues associated with the preparation and administration of 90Y-Zevalin. Methods Ten patients have been treated to date. Radiochemical purity was carried out on each patient dose sample using ITLC-SG and 0.9% saline. The strips were imaged and RCP calculated using a Packard Instant Imager. The material was administered using a syringe driver for the first 7 patients. For the remaining patients a manual push system was used. Finger doses were measured using a TLD on the tip of each finger.

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BNMS meeting: Abstracts A5

Results Mean RCP at 2 h post-preparation was 98.9% (range 95.7–99.7%), but we are aware of failures at other centres. During preparation the average dose to the finger tip was 9.4 mSv for the first 4 procedures (max 27 mSv). However, this was reduced to 1.7 mSv for the remainder (max. 8.8 mSv). The average dose to the operator when using a syringe driver for administration was 6 mSv (max. 47 mSv). For the manual push system the average dose was 1.8 mSv (max. 6.5 mSv). Conclusions Centres providing Zevalin treatment should be aware that failure to meet the specified radiolabelling efficiency can occur and contingency plans are required. Using some simple measures finger doses for the preparation and administration of 90 Y-Zevalin can be kept at acceptable levels. A20 Evaluation of detectors for ambulatory monitoring of renal reflux S.-J. Hyde, M.G. Coulthard and M.J. Keir Departments of Paediatric Nephrology and Medical Physics, Royal Victoria Infirmary, Newcastle upon Tyne, UK. Purpose (1) To evaluate the performance of CsI detectors suitable for ambulatory monitoring of radioactivity in patients, and (2) to evaluate their potential for continuous monitoring of the renal pelvis to detect vesico-ureteric reflux. Methods (1) Measurement of a range of physical parameters of the detectors including energy resolution, dead time, sensitivity. (2) Determining detector response to a moving point source in a scattering medium with varying degrees of collimation. (3) Measuring CT scans on children of different sizes to assess the depth and separation of the renal collecting systems. (4) Combining these data to predict the relative sensitivities and cross-talk from the ipsi- and contra-lateral kidney over a range of detector positions. Results (1) Energy resolution (30% for 99mTc and dead time (30% at an observed 4 kcps) are poor compared to a NaI system. (2) Renal pelvis depths ranged from 3 to 8.6 cm posteriorly, 9 to 31 cm anteriorly. Lateral depths ranged from 5 to 11 cm (3) Predicted cross-talk (4–20%) and sensitivities (over a factor of 3) varied with detector position and patient size, and these results define the optimum positions for the detectors. Conclusions Despite some limitations these detectors show promise for the ambulatory monitoring of radioactivity on the renal pelvis. A21 An analysis if current SEHCAT procedure: is patient background important? E. Pearce and M. Newell Royal London Hospital, UK. Aim Accurate subtraction of room background is essential when performing SeHCAT studies for bile salt malabsorption (BSM). However, the interposition of the patient will affect background counts. This paper analyses any effect this will have on final outcome and compares errors in results between whole-body counter (WBC) and gamma camera (GC) techniques. Methods One hundred and forty-two patient SeHCAT studies using a WBC were analysed retrospectively. Based on published

guidelines [1], 40% were normal, 11% had mild BSM and the remainder moderate to severe BSM. Adjustment of room background by the patient was routinely taken into account and room background was measured. Results If only room background counts had been used, the result would only have changed in 1 patient (from mild BSM to normal), but the percentage altered by more than 1% in 4 other cases. If only anterior or posterior views were used, results changed significantly. For the case of only 1 detector, including patient background altered the final value by more than 1% in 11 cases, and the final result in 5. Conclusion For the WBC case, the inclusion of patient background is statistically significant, (t-test, P = 0.0195) and extremely statistically significant for a single detector where 5 results were changed (P < 0.0001). The errors in results from a WBC and GC were comparable. Reference 1. Thomas PD, et al. Gut 2003; 1:52. TECHNOLOGISTS SESSION A22 Clinical technologist: This is your life C. Segasby A23 Extremity dose optimization in the radionuclide dispensary D.E. Anstee, M. Whitby and G. Gillen Glasgow Western Infirmary and Gartnavel Infirmary, UK. Purpose Extremity doses in a large centralized radionuclide dispensary, which prepares over 40 000 patient injections per year, are approaching and occasionally exceeding local dose constraint levels. As a result the effectiveness of simple and cost effective changes in staff technique/procedures to reduce extremity dose to personnel have been evaluated. Methods Finger doses have been measured using an electronic extremity dose monitor (AEGIS). AEGIS allows the dose received from individual actions to be quantified and patterns of radiation exposure determined. Finger doses were measured during various manipulations involving the dispensing of kit vials and the sub-dispensing of individual patient vials using different methods of shielding, small changes in existing technique and protocol. Results The change of syringe size from a 1 ml to 3 ml during the making up of kit vials could potentially result in an overall dose saving of approximately 15–20%. This associated with slight changes in dispensing technique and the use of various lead, protective syringe inserts have the potential to reduce technician hand dose. Conclusions Only small changes in dispensing protocol can potentially make significant dose savings over a complete dispensing session. Small changes in technique and the use of various devices to improve local shielding, which can easily be replicated by other dispensary/nuclear medicine departments, are also discussed in relation to dose optimization. A24 The sentinel node: A challenge for the technologist S.L. Johns, V.B. Batty and J. Smallwood

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A6 Nuclear Medicine Communications 2006, Vol 27 No 7

Department of Nuclear Medicine, Southampton University Hospitals NHS Trust, UK. Background The emerging need for nuclear medicine to play a role in the detection of the sentinel node has provided an opportunity for technologists at Southampton to develop their skills. To meet demand from surgeons and overcome medical staffing problems we have developed a technologist-led service. Technologists are responsible for the entire investigation including report. Method To ensure quality is maintained continual audit is performed between the number of nodes detected by the technologist and the surgeon. As a comparison and a standard, data is compared to studies performed by radiologists prior to the new technologist service. Results Radiologist agreement, 50% (9/18 studies); technologist agreement, 64% (47/74 studies). Variance is found due to: close proximity of multiple nodes surgeon’s decision depending on count rate K inaccessible nodes K pooling in lymphatic vessels. K K

Benefits This service provides greater procedure availability. Technologists have extended their responsibilities into medical roles e.g. injecting, marking and reporting. The challenge has demanded that technologists have researched and reviewed protocols. Audit allows review to investigate why differences occur between surgeon and technologist. The patient has improved continuity of care because one person performs the entire procedure. Lymphoscintigraphy can be the first opportunity the patient has post-diagnosis to actually ask questions, which an informed technologist can answer. This new development has enabled the workload to increase whilst advancing the technologist’s role. A25 The design and implementation of a virtual learning environment for nuclear medicine students M. Griffiths and S. Messer University of The West of England, Bristol, UK. Purpose The nuclear medicine programme at the University of the West of England encompasses a geographical range of students. In order to promote practitioner autonomy, develop IT skills and provide asynchronous learning material, a web-based platform was utilized. Method A virtual learning environment (VLE) was utilized, which enabled students to learn by their preferred style [1] and supported their overall experience of the programme in the periods between synchronous communication within the classroom. Students accessed the VLE via a password system and were able to utilize an on-line discussion board and upload/ download course material. The VLE also provided core supporting material for the students’ assessments, in particular the use of video streaming technology for module experiments. Results Due to the geographical nature of the nuclear medicine students, the use of the VLE was well received by students, who embraced the various aspects of the asynchronous learning environment. The nature of the learning environment

enabled the students to access the on-line material/discussion board at any time. Conclusion The development of the nuclear medicine practitioner is fundamental to the future provision of the clinical service. The development and utilization of a web-based learning resource promotes autonomy and encourages ownership of the programme. Reference 1 Ally M. Foundations of Educational Theory for Online Learning. Athabasca, Alberta, Canada: Athabasca University, 2004. A26 Experiences in validating MUGA scan processing on new platforms F.I. McKiddie, E.J. Davidson, R.T. Staff and H.G. Gemmell Nuclear Medicine Department, Aberdeen Royal Infirmary, Aberdeen, UK. Background Following installation of a new gamma camera we determined to find the optimal method of processing MUGA scan results on the available platforms. These were a Siemens eSoft and a Nuclear Diagnostics Hermes. The low threshold of normality on the existing Link Medical MAPS system has caused some problems with site acceptance into multi-centre clinical trials. Methods and results Thirty-nine patient studies were processed on all 3 systems. Bland–Altman analysis demonstrated absolute systematic errors of around 12% between the Hermes and Siemens systems and the MAPS technique. Regression analysis demonstrated the 40% normal threshold on the MAPS translated to 58% with the new techniques, which seemed rather high. Therefore, an analysis of the resulting sensitivities was carried out. The MAPS technique produced an abnormal rate of 12.8%. The 58% threshold with the new techniques produced rates of 25.6% for the Hermes and 28.2% for the Siemens. The thresholds were varied to produce abnormal rates closer to those obtained with the MAPS software. Thresholds of 55% with the Siemens software and 53% with the Hermes were found to produce the closest results. The IPEM audit data was then used as a test of inter and intra-observer variability. This found the Hermes system to be the most robust. Conclusion This study demonstrates that straightforward parametric analysis may not be optimal for tasks of this sort. Discussion Forum: Technologists Training GENERAL SESSION A27 Radiation protection and the design of radionuclide facilities P. Marsden London, UK. A28 Lung ventilation scintigraphy used to assess aerosol nebulizer deposition and repeatability in CF C.M. Waudbya, G.A. Wrightb, J.D. Seymoura and A.H. Moricec a Department of Comparative and Applied Social Science, The University of Hull; and the Departments of bNuclear Medicine and cAcademic Medicine, Castle Hill Hospital, East Yorkshire, UK. Introduction Cystic fibrosis (CF) patients use nebulizers regularly for delivering drugs to the lungs. The quantity and

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BNMS meeting: Abstracts A7

pattern of deposition can vary greatly. Lung ventilation scintigraphy can be used to assess deposition from nebulizers. The aim of this study was to compare two commonly used nebulizers (Pari LC Plus and Profile Prodose) and investigate the repeatability of the technique. Methods Ten CF patients were recruited for the study. Patients used one of the nebulizers for 2 visits and the remaining nebulizer once. Each nebulizer delivered 5 mg salbutamol in 3 ml saline followed by approximately 20 MBq of 99m Tc-DTPA in 3 ml saline. Anterior and posterior images were taken immediately following nebulization. ROIs were drawn manually by two operators and used to quantify lung deposition as a percentage of original nebulizer activity. Bland–Altman analysis was used to assess agreement between repeated measurements and processing. Results The Pari delivered less activity to the lungs than the Prodose ( – 1.11% ± 0.76, P = 0.002). Inter- and intra-operator agreement was 0.025% ± 0.07 (P = NS) and 0.13% ± 0.1 (P = s). The difference in deposition between visits for the same nebulizer was 0.26% ± 0.33 (P < 0.05). Conclusions Lung ventilation scintigraphy allows comparison of nebulizer performance. However, variations in deposition can occur with the same nebulizer in the same patients. The magnitude of such differences must be borne in mind when comparing between nebulizers. A29 Technegas or Smartvent for lung ventilation? G. Gardner and G. Ainslie Department of Nuclear Medicine, Glasgow Royal Infirmary, UK. This study compared the relative effectiveness of the new Smartvent aerosol lung ventilation system with the QUADOS Technegas system. We have looked at issues regarding ease of use, patient compliance, staff dose and image artefacts, as well as the image quality. With a randomized study of 100 patients from each system, a team of 5 observers scored each patient on a scale of 1 (poor) to 5 (good). Findings show that both techniques produced a mean score of 3.8 over the entire patient group, with a similar number of technically poor results, due to patient compliance or COAD, being demonstrated. The Smartvent system was considered simpler to operate, although patient administration times were longer. No significant difference in staff dose during preparation or administration was noted and there was no evidence of airborne contamination with the Smartvent system while the Technegas system, being pressurized, could contaminate staff and cameras. In conclusion, the Smartvent system is an acceptable substitute for lung ventilation studies. A30 Divided function: Are two heads better than one? J.L. Dennis, A.A. Bolster and H.W. Gray Glasgow Royal Infirmary, Glasgow, UK. Background Using both posterior and anterior image data in renography allows a geometric mean divided function to be calculated, providing some correction for renal depth differences. This method has been used in static renal imaging for many years, but the traditional method of using posterior image data alone is still used routinely for dynamic renogram investigations. This study was performed to assess the feasibility

of obtaining both anterior and posterior image data to calculate the geometric mean divided function and to assess any differences between calculation methods. Methods Both posterior and anterior dynamic image data were acquired using a double-headed gamma camera for 110 renography investigations on adults. Geometric mean images were created from this data. Regions of interest were drawn on the posterior image and the divided function calculated by integrating the renogram curve from 1 to 2 min for both the posterior image data only and the geometric mean images. These results were then compared. Results It was possible to calculate the divided function using the geometric mean. The mean difference between the calculation methods was 0.91% ± 3.56%. Clinically significant differences ( Z 5%) existed between the methods in 11.8% of cases. The intra- and inter-observer variability were assessed and found to be high for both calculation methods. Conclusion The results of this study indicate that the geometric mean method can successfully be used to calculate renal divided function in dynamic renography. A31 Indirect radionuclide cystography in children: Physiological parameters associated with renal reflux H. Bashira, I. Gordona, L. Biassonia, P.J. Andersona and D. Ridoutb a Department of Nuclear Medicine, Great Ormond Street Hospital London, and bCentre for Paediatric Epidemiology and Biostatistics, Institute of Child Health, University College London, UK. Aim To assess the physiological parameters associated with renal reflux in children undergoing indirect radionuclide cystograms (IRCs). Methods One thousand and thirty children (524 boys and 506 girls, aged between 2.6 and 17.6 years) underwent 1113 MAG3 renograms followed by IRC studies. Reports were reviewed for differential renal function (DRF), hesitancy on micturition, extent of micturition (complete, incomplete or failure-to-void) and presence or absence of renal reflux. Results Unilateral reflux was seen in 12% of voids on the left and 8% on the right. Bilateral reflux was seen in 6% of voids. Seventeen percent of the boys and 9% of the girls showed reflux. In single-void studies (n = 549) 20% refluxed while in multiple-void studies (n = 564) 47% refluxed. In multiple-void studies 13% displayed reflux on all acquisitions while 20% refluxed intermittently. Hesitancy was longer (mean 36 s vs. 29 s (P < 0.05)) and voided volumes smaller (mean, 209 ml vs. 237 ml (P < 0.05)) when reflux was present compared to voids with no reflux. With unilateral reflux the DRF was reduced on the affected side (35%). Discussion Our patient population is unusual in the high proportion of boys who underwent MAG3 and IRC. Reflux is associated with smaller voided volume and longer hesitancy. Unilateral reflux is associated with reduced function of the affected kidney. The more often the child voids, the greater the chance of discovering reflux. A32 ‘Counter Intelligence’: A Class IIa Medical Device D.E. Simpson, D.T. Attwell, A.G. Kettle and C.P. Wells Medical Physics Department, Kent and Canterbury Hospital, Canterbury, Kent, UK.

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A8 Nuclear Medicine Communications 2006, Vol 27 No 7

Over the past 10 years our centre has developed a software package using Microsoft Visual Basic called Counter Intelligence. The current version (version 23) calculates glomerular filtration rate (GFR) from patient plasma samples following an injection of 51Cr-EDTA. This figure is reported along with GFR normalized to a body surface area of 1.73 m2 and an age-matched normal range. The software calculates the GFR and normal range in accordance with the 2004 BNMS guidelines. Version 23 also calculates GFR according to the previous methodology of the department at Kent and Canterbury Hospital. This differed from the BNMS guidelines in that a Chantler one-pool correction was applied and the Du Bois & Du Bois body surface area formula was used. The department is registered under ISO 9001:2000, ISO13485:2003 and the Medical Devices Directive. The software has been CE marked as a Class IIa Medical Device. The technical file produced was recently examined by our external auditors and found to be satisfactory. The experience of designing software to comply with MDD essential requirements will be discussed, together with the process of testing, validation and maintenance under our quality management system. POSTER SESSION P1 Does a little ECG knowledge improve gated cardiac acquisition? P. Sorrell and C. Randall Royal London Hospital, UK. Background Investigation of myocardium by single photon emission computed tomography (SPECT) myocardial perfusion imaging (MPI) is set to increase. National Institute for Clinical Excellence technology appraisal 73 (section 3) describes ECG gating as a ‘technical improvement’ to SPECT MPI. The number of examinations gated will rise alongside the increased use of MPI. Gating of MPI adds quantitative information. However, reliability of this information is limited by the quality of the data obtained [1]. It is common to obtain gated data from patients with rhythm abnormalities. One study showed over 50% of gating errors arising from inconsistent or transient fluctuations in rhythm [2]. Audit, combined with education, has been used to improve the gated data acquired. Results There is improved operator confidence at solving gating problems, an apparent reduction in gating errors, and a software auto-centring approach showed no advantage over operator ECG selection. Conclusions The following recommendations were made for those acquiring gated MPI: Better location and preparation of sensor sites Reassurance/relaxation of the patient K Read the supplied information K Record and seek advice on unusual traces K Do not allow a patient to sleep, as the heart rate falls K Changing the lead may eliminate artefactual problems K Re-audit and hold teaching sessions annually, or on introduction of new staff. K K

References 1 Nichols K, et al. Influence of arrhythmias on gated SPECT myocardial perfusion and function quantification. J Nucl Med 1999; 40. 2 Nichols K, et al. Clinical impact of arrhythmias on gated SPECT cardiac myocardial perfusion and function assessment. J Nucl Cardiol 2001. P2 Evaluation of cardiac index in obese and normal weight patients using post-stress gated SPECT MIBI perfusion imaging P.J. Wheeler, J.P. Coffey and J.C. Hill Department of Nuclear Medicine, Royal Preston Hospital, Lancashire Teaching Hospitals, Fulwood, Preston, UK. Background Cardiac output is elevated in obese subjects owing to increase in overall mass and relative increase in fat-free mass. Objective To compare cardiac index CI, (cardiac output, CO, corrected for body surface area), stroke volume and ejection fraction of obese and normal weight symptomatic patients. Methods Thirty-two obese patients (body mass index, BMI > 30, mean 35) with 32 age-matched patients with BMI < 30, (mean 26), referred for evaluation of atypical chest pain, dyspnoea or inconclusive exercise ECG, were studied following injection of 400 MBq MIBI. Gated SPECT imaging was performed 40 min after treadmill stress, in the supine position, 8-interval cardiac cycle with normal time–volume ventricular curves. CO was calculated as the product of stroke volume and resting heart rate and CI was calculated. Results Mean CI for both groups was 2.28 l
m – 2
min – 1 for obese and 2.24 l
m – 2
min – 1 for normal weight patients. T tests showed no significant difference in mean CI or in mean stroke volume (SV). Ejection fraction was significantly increased for the obese group (mean, 65.2%), P < 0.03. Conclusion Haemodynamic function, represented by CI, is equivalent for obese and normal weight patients. Increased ejection fraction is present in obese patients, possibly related to subclinical changes in myocardial function. P3 Do we receive clinically important information on cardiac request forms? A. Nafati, J.R. Buscombe and A.J.W. Hilson Nuclear Medicine, Royal Free Hospital, London, UK. Aim To evaluate whether clinically important information was provided by referrers on the request forms for myocardial perfusion imaging (MPI). Method A retrospective audit was performed of 100 MPI request forms. We looked for information on a history of asthma, hypertension, diabetes, hypercholesterolaemia, a family history of cardiac disease, history of smoking and a previous history of CABG. This was compared to the information gained by direct questioning of the patients when they attended the department for their MPI stress test. Results There were 14 patients with asthma; 10 (71%) were notified on the request form. Only 63% with a previous history of CABG had it recorded. Only 50% of the 26 diabetics were noted on the request form; high cholesterol was recorded in 25%; only 3/40 (7%) of smokers were notified.

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BNMS meeting: Abstracts A9

Conclusion There is severe lack of information presented from referrers to practitioners, necessitating more effort to ensure clinically important information is provided on the request form. P4 The effect of SPECT reconstruction corrections on the quantitative accuracy of myocardial perfusion studies K.L. Dixon, E.J. Vandervoort, S. Blinder, A. Fung and A. Celler Vancouver Hospital & Health Sciences Centre, Canada. Aim To investigate the performance of SPECT corrections for photon attenuation (AC), scatter (SC) and collimator blurring (DRC), on the absolute and the relative quantification of myocardial perfusion studies. The absolute measurement used is the myocardial wall thickness, and the relative measurement is myocardial infarct size. Method A series of phantom studies were performed and additional information was gathered from normal patients. Each set of data was reconstructed with (1) filtered back-projection (FBP), (2) ordered subset expectation maximization (OSEM), (3) OSEM plus AC, (4) OSEM plus DRC, (5) OSEM plus AC and DRC, and (6) OSEM plus AC, DRC and SC. Results Both patient and phantom data showed SPECT corrections to have a significant effect on myocardial wall thickness. Phantom data showed the measurement of the anterior and lateral wall infarct sizes to be consistent for all reconstruction techniques, and the measurement of the inferior and septal walls to be more accurate when the reconstruction included AC. Conclusion This analysis suggests that in order to achieve accurate, absolute quantification, AC, DRC and SC should be applied. However, for the intermediate step of relative quantification, the application of AC and DRC is sufficient. P5 Quantification from SPECT MPS: An ejection fiction? K.C. Cockburn, G.A. Wright, A.C. Tweddel, G.R. Avery and G. Davies Departments of Nuclear Medicine and Cardiology, Hull and East Yorkshire Hospitals NHS Trust, UK. Background and aim Measurement of LVEF from perfusion studies requires a reproducible method of defining the left ventricular (LV) surfaces. Inconsistencies will influence the calculated LVEF. Opinion varies on the reliability of commercial software packages (e.g. QGS, ECTb). The aim of this study was to investigate the effect of various parameters on such measurements. Methods A 99mTc-filled static phantom (with and without defects) was imaged gated to an ECG simulator on a GE Infinia Camera. LVEFs and volumes were calculated using ECTb and QGS. The effect of collimator and simulated breast attenuation was investigated. Statistical significance was assessed using the Wilcoxon test. Results True LV volume (65 ml) was underestimated by both programs (QGS, 56.8 ± 5.25 ml; ECTb, 51.4 ± 6.25 ml). Volumes were smaller with the LEGP vs. LEHR collimator (QGS, 3.96 ± 2.82 ml, P < 0.002; ECTb, 2.4 ± 4.54 ml, P < 0.05). Variations in calculated volume throughout the ‘cardiac cycle’ resulted in apparent LVEFs of up to 16% in QGS (mean 7.4 ± 2.8%) and 38% in ECTb (mean 16.2 ± 6.8%),

compared to the expected 0%. The addition of ‘breasts’ resulted in lower volumes (4.46 ± 3.6 ml, P < 0.002) and higher LVEFs with QGS (1.79 ± 3.73%, P < 0.05), but not ECTb (P = NS). Conclusion Quantitative parameters from these programs should be interpreted with caution. Both programs produced non-zero LVEFs in the absence of changing LV volume. The presence of breast attenuation increased the apparent LVEF. P6 Clinical audit of myocardial perfusion scans: How NICE are we to our patients? S.A. Mostafa, F. Sundram and A. Notghi Department of Physics and Nuclear Medicine, City Hospital, Birmingham, UK. Aim NICE guidelines (Technology Appraisal Guidance 73, November 2003) recommend a waiting time of 6 weeks and 1 week, respectively, for routine and urgent myocardial perfusion studies. We reviewed our working practices to see if we were compliant with the guidelines. In addition, we examined the time interval between injection and going home in our patients. Method A retrospective review of 109 patients who had myocardial perfusion scans over a 3-month period in 2004 was undertaken. The date the referral forms were received and the scan date were recorded. Of the patients referred, details from 81 were recorded on the scan day: this included the time of injection, the time of the scan and the time the patient left the department. Results The mean waiting time for a scan was 125 days (approximately 18 weeks, range 6–205 days). Fifty percent of patients waited between 126 and 143 days, as shown in Table 1 below. Table 1

Waiting times during perfusion studies

Interval Referral to scan (days) Injection to departure (min) Scan to departure (min)

Mean ± SD

Range

125 ± 38 112 ± 29

6–205 62–181

46 ± 16

20–100

Conclusions We are currently not adhering to NICE guidelines for myocardial perfusion studies. We anticipate the waiting list time to further increase due to increased number of referrals. We are looking into a feasible solution to reduce waiting list times to comply with the guidelines. P7 What is the influence of food and water on the quality of myocardial perfusion images (MPI)? S.A. Mostafa, F. Sundram and A. Notghi Department of Physics and Nuclear Medicine, City Hospital, Birmingham, UK. Aim Extra-cardiac GI tract activity can interfere with inferior myocardial wall uptake and make the interpretation of images difficult. This study examined the effect of food and water on the image quality. Method A retrospective review of 116 MPI studies was undertaken. Patients injected with 99mTc-tetrofosmin were imaged 45 min later. Before scanning, they were asked if they had consumed food (snack, including sandwich, pastry, crisps or chocolate) or water or nothing and this was noted. Three independent observers examined the short-axis slices for gut

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A10 Nuclear Medicine Communications 2006, Vol 27 No 7

uptake. Images were given a qualitative consensus score of 1 (no gut uptake) to 5 (major uptake, scan repeated). Results The results are shown in Table 1 below. Only 2/18 (11%) of patients who had food had gut uptake, 12/24 (50%) of patients who had no food/drink and 39/74 (53%) of patients who had water had some gut uptake. A chi-squared test between the food and water groups, and between the food and no food/water groups showed highly significant P values of < 0.001 and < 0.01, respectively. Table 1

Effect of food and water on scores for image quality

Score 1 2 3 4 5

Food (n = 18)

Water (n = 74)

No food/water (n = 24)

16 0 1 0 1

35 12 18 5 4

12 4 6 2 0

Conclusion In MPI, the ingestion of food appears to be associated with reduced gut uptake compared with water or no food/water. We recommend that all patients consume a snack before their scan. P8 Screening for bone metastases in breast cancer patients: Correlation with local surgical staging K. Gray, E. Kalkman and C. Wilson Western Infirmary, Glasgow, UK. Aim To evaluate the diagnostic use of isotope bone scans (BS) to screen for asymptomatic metastases in breast cancer patients and correlate with loco-regional surgical staging. Methods Retrospective review of surgical database and corresponding BS reports. Surgical staging was scored using the Nottingham Prognostic Index (NPI), using tumour size, histology and number of lymph nodes involved. Results Seven hundred and forty-six patients were identified in the surgical database of 2002 and 2003. Seven hundred and seven patients had been surgically staged, 111 of whom, because of large tumour size, nodal spread or inflammatory type histology, were referred for BS. Thirty-nine patients were inoperable. Eighty-two full sets of data were obtained (Table 1, below). Table 1

Relationship between NPI scores and bone scan results

NPI 3—5 (n = 28) 5—6 (n = 27) Z 6 (n = 27) All (n = 82)

Negative BS (%)

Equivocal BS (%)

Positive BS (%)

78.6 66.7 74.2 73.2

7.1 11.1 14.8 11.0

14.3 22.2 11.1 15.8

Conclusion Bone scanning is useful to stage breast cancer patients selected by the NPI, even if there are no symptoms suggestive of bone metastases. Within the study group, the range of NPIs was too small to allow further correlation between the local surgical stage and BS result.

Aim To ascertain whether ALP has an additional benefit to using Gleason score and PSA level in predicting bone metastases in prostate cancer patients. Methods One hundred and five patients (mean age, 70 years) diagnosed with prostate cancer over a period of 12 months were referred for bone scans. Only 77 included in this retrospective study, as they had Gleason scores, PSA and ALP values available in addition to the bone scan results. High Gleason score and PSA > 11 mg
l – 1 are already known as strong indicators for bone metastases. In this study ALP values > 130 U
l – 1 were evaluated as a possible third indicator of positive bone scans. Results Thirty of 77 patients had positive bone scans. All had PSA values > 11. However, only 13/30 had ALP > 130 (average 683) and Gleason scores of 8–10. The remaining 17 had normal ALP values (average 92) and a tendency towards lower Gleason scores. Forty-seven of 77 patients had negative bone scans. However, 37 of them had PSA levels > 11. All except 2 had a normal ALP. Conclusion Total serum ALP offers no additional benefit in predicting positive bone scan results in the presence of high PSA and Gleason score values. No significant correlation was identified between PSA, ALP values and Gleason scores in patients with negative bone scans. P10 Metastatic prostate carcinoma: Should clinicians ‘please stop asking’ for a bone scan in patients with a low PSA? S.A. Mostafa and F. Sundram Department of Physics and Nuclear Medicine, City Hospital, Birmingham, UK. Aim Suspected bone metastases from prostate carcinoma warrant an isotope bone scan. We investigated the patterns of bone scans in patients with prostate cancer and associated them with levels of prostate specific antigen (PSA). Method A retrospective study of 46 bone scans of patients with biopsy-proven prostate carcinoma over a 1-month period in 2004. Whole-body scans were performed using 99mTc-MDP. Scans were classified as follows: normal, presence of metastases and abnormal (due to other pathology). Patients had PSA levels measured around the time of the scan. Results The majority of patients with prostate carcinoma in our study group had normal bone scans (mean PSA 31). The mean PSA for patients with bone metastases was higher (87). Patients with other concomitant bony pathology (degenerative, osteoporosis and Paget’s disease) had mean PSA of 18. An unpaired ttest for PSA values between normal and metastatic groups showed a t value of 2.56 and P value of < 0.029 (significantly different). Groups showed a t value of 2.56 and a P value of < 0.029 (significantly different). Results are shown in Table 1, below.

Table 1

Relationship between scan classification and mean PSA

result

P9 Total serum alkaline phosphatase (ALP) and bone metastases in patients with prostate cancer T. Toma and V. Shah Department of Nuclear Medicine, Southend Hospital NHS Trust, UK.

Scan classification Normal (n = 20, 43%) Metastases (n = 14, 30%) Abnormal (n = 12, 27%)

Mean PSA ± SD

Range

31 ± 44 87 ± 79 18 ± 11

4–168 17–210 0.4–35

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BNMS meeting: Abstracts A11

Conclusions There is a strong link between elevated PSA values and the presence of bone metastases. However, a low PSA does not exclude metastases. Therefore, bone scans should be done in all patients. P11 Interobserver variability in the interpretation of V/Q lung scintigrams S.C. Chua, F. Cooke, D. Green, D. Rose, D. Walk and A. O’Connor Nottingham City Hospital, UK. Aim Despite using the PIOPED criteria to standardize reporting, the basis of V/Q scan interpretation remains identification and assessment of size of a perfusion defect, a somewhat subjective task. It is important that there should be a high rate of inter-observer correlation in their reporting. This study evaluates the degree of agreement by 5 observers. Material and methods Sixty lung scintigrams(V/Qs) were interpreted by 3 nuclear medicine radiologists, 1 SpR and 1 nuclear medicine radiographer with accompanying chest radiographs but without clinical data. V/Qs were read according to the PIOPED criteria (normal, low, intermediate and high probability). Correlation was assigned a numerical score: a score of 0 indicates the same PIOPED category as other observer; a score of 1 indicates one level of difference (e.g. normal to low or intermediate to high); a score of 2 or 3 indicates two or three levels of difference (e.g. low to high, which is regarded as two levels). Kappa statistics were used to evaluate the degree of agreement between individual observer interpretations beyond that expected by chance alone. Results 38.3% of the studies showed 0 level of difference between observers; 41.7% with one level of difference and 20% two levels of difference. None had three levels of difference. The mean kappa value (k) was 0.57 (0.46–0.69) as compared to a literature standard: average of k = 0.54 (0.27–0.70) [1–4]. Conclusions This study showed results comparable to previously published data.

1 2 3 4

References Christiansen F, et al. Acta Radiol 1996; 37:754–758. Christiansen F, et al. Nucl Med Commun 1997; 18:112–117. Infante JR, et al. Nucl Med Commun 2002; 21:93–98. Hagen PJ, et al. J Nucl Med 2003; 44:739–744.

P12 The importance of chest radiograph abnormalities in the choice of lung scintigraphy versus CT pulmonary angiography V. Prakash, M.S. Thyagarajan and S. Matthews Sheffield Teaching Hospitals NHS Trust, UK. Aim To assess the awareness of radiology specialist registrars of the significance of abnormal chest radiographs within a local pulmonary embolism imaging protocol. Method At our tertiary hospital we have local guidelines for the evaluation of chest radiographs prior to further imaging with perfusion scans and/or computed tomography for the investigation of pulmonary embolism. We submitted a questionnaire to all radiology registrars at our institution. This asks what would be the next best imaging step when the chest radiograph reveals an abnormality. The completed questionnaires were compared with the recommended guidelines.

Results Seventy percent of the guidelines were not being followed. Thirty percent of the guidelines which were being followed did so without appropriate justification. Conclusion A severe lack of awareness amongst the radiology registrars concerning the recommended guidelines for the investigation of pulmonary embolism has resulted in inappropriate use of imaging modalities; specifically, perfusion scanning in a tertiary care teaching hospital. P13 An audit of clinical effectiveness of ventilation– perfusion lung scans in the investigation of pulmonary embolism J. McDonagh, E. Kalkman and G. Gillen Dept of Nuclear Medicine, Western Infirmary, Glasgow, UK. Introduction Ventilation–perfusion (V/Q) scanning is a sensitive method for detecting pulmonary embolism (PE). However, many other cardio-pulmonary diseases can also cause perfusion defects, resulting in an increasing percentage of indeterminate scans. This percentage can be reduced by appropriate patient selection, using CT pulmonary angiography (CTPA) for patients with abnormal chest radiographs (CXR); also by scanning patients as quickly as possible after the onset of symptoms. This audit aimed to provide practical information into the clinical practices of VQ scanning within the Nuclear Medicine Department, Western Infirmary Glasgow. Methods A retrospective review of V/Q scan reports from the first quarter of 2003–2005 was performed. Clinical information and CXR were used to select patients for V/Q scanning or CTPA. V/Q scans were performed as soon as possible after receiving the request. Scans were reported by a consultant radiologists experienced in nuclear medicine using the revised PIOPED criteria [1]. Results Three hundred and twenty V/Q scans were performed within the audit period, the distribution of probabilities reported are displayed in Table 1. Ninety-four percent of the patients were scanned within 2 days of request.

Table 1

Distribution of probabilities reported

V/Q result Normal Low probability Intermediate High probability

2003 (%) 28.5 30.8 25.4 15.4

(n = 37) (n = 40) (n = 33) (n = 20)

2004 (%) 27.2 43.9 16.7 10.5

(n = 25) (n = 41) (n = 18) (n = 7)

2005 (%) 29.3 39.4 16.2 15.1

(n = 29) (n = 39) (n = 16) (n = 15)

Conclusion Departmental protocols to exclude unsuitable patients for V/Q scanning and to expedite scans following the onset of symptoms resulted in a high diagnostic effectiveness in all 3 audit periods. PE was excluded or confirmed in 79% of the population being screened. Reference 1 Gottschalk A, et al. J Nucl Med 1993; 34:1119–1126. P14 Technegas: Where does it go? M. McDade, S. Allan, D. Brown, E. Donald, J. Morrison and R. Williamson Nuclear Medicine Dept., Southern General Hospital, Glasgow, UK.

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A12 Nuclear Medicine Communications 2006, Vol 27 No 7

Background Lung ventilation scans at our centre are performed using Technegas. This consists of a fine suspension of 99m Tc-labelled carbon particles in an argon gas flow. During patient administration there is the possibility of both internal and external contamination to the operator. Aim To investigate the levels of contamination using a wholebody monitor. Method Operators wore gloves and paper gowns during patient administration procedures. The gowns and gloves were subsequently counted in the whole-body monitor in order to calculate possible contamination. The staff member was also counted to determine internal contamination. The monitor was calibrated using known 99mTc standards. Patients were rated subjectively by the operator for probability of contamination on a scale of 1–3 (1 = high, 3 = low). Results Staff contamination has been monitored on 22 occasions. Average internal activity was 11 kBq (range, 1–53 kBq) and average external activity was 13 kBq (range, 0.4–104 kBq). This suggests that both internal and external contamination may occur during ventilation procedures using Technegas. P15 A comparison of tumour to background ratios at 2 and 4 h in 99mtc-depreotide Imaging of lung cancer G. Ainslie, A. Bolster and J.B. Neilly Department of Nuclear Medicine, Glasgow Royal Infirmary, UK. Preliminary data is presented from a larger on-going multicentre Glasgow-wide trial into the use of 99mTc-depreotide in patients with known lung cancer. In the first instance this trial involves 50 patients recruited from around Glasgow. These patients have known lung cancer and will have a 99mTcdepreotide SPECT image acquired at 2 and 4 h post-injection. To date, 20 out of 50 acquisitions have been performed, of which 10 were performed at GRI. The data presented provides a quantitative comparison of the tumour/background ratio between 2-h and 4-h 99mTc-depreotide SPECT images of the 10 patients scanned at GRI. Images were reconstructed using the traditional filtered backprojection technique and then OSEM iterative technique and a comparison made between both techniques with regards to qualitative and quantitative analysis. Initial results show significant differences in tumour/background ratios calculated using the FBP and OSEM reconstruction techniques. Phantom data has also been undertaken which will support the reconstruction technique choice for this multicentre study. Tumour/background ratios from 10 patients imaged to date showed either no change between the 2-h and 4-h images or an increase at 4 h. Previous studies suggest no change to be indicative of benign disease. This hypothesis will be tested in this small patient group. P16 Octreotide therapy preventing radiolabelled octreotide therapy: 2 case reports M. Kana, R.H. Reid, W. Gillis, W. Kocha and R. Davidsonb a London Middlesex Community Care Access Centre; and bLondon Health Sciences Centre, UK. Carcinoid crisis in patients with metastatic carcinoid tumour can be a serious life-threatening situation. Synthetic somatostatin

analogues, both short- and long-acting forms, have significantly improved the clinical course in carcinoid syndrome. These patients can also be treated with the radiopharmaceuticals 111In-octreotide and 131I-MIBG. It is recommended that patients stop taking sandostatin before diagnostic scanning with 111 In-octreotide, although many patients are unable to do this because of clinical instability. In a recent retrospective audit of 85 patients at our centre, who were on sandostatin therapy, 83 showed no significant reduction in uptake of 111In-octreotide, but uptake was completely blocked in the other 2. These 2 patients had previously received radioisotope therapy, and still had severe residual carcinoid symptoms requiring increasing sandostatin infusion. One other patient with similar disease severity and sandostatin requirements did not have the block. Attempts to wean the 2 patients off sandostatin to allow therapy were not successful in 1 of them, as it precipitated a severe carcinoid crisis just before therapy. In the second patient, control of the symptoms was possible with i.v. sandostatin in increasing doses, and radio-frequency ablation of metastasis in the liver was attempted in order to reduce the tumour burden and thereby control the carcinoid crisis. These two patients form the basis of this clinical review, which describes the unusual situation where one life-saving therapy prevents the administration of another. In both cases other therapy options were precluded because of the underlying medical condition. P17 123I-MIBG in the investigation of neuroendocrine tumours: is imaging at 4 and 24 h after injection beneficial? J. Prossera, E. Kalkmanb and S. Vinjamuric Departments of aNuclear Medicine and bRadiology, Western Infirmary, Glasgow, and cNuclear Medicine, Royal Liverpool University Hospital, UK. Background In the investigation of tumours derived from the neural crest using 123I-MIBG, guidelines published by the EANM recommend imaging at 24 h after injection. In our institution, we perform imaging at 4 h and 24 h after injection. Aim To assess whether whole-body imaging at two time points (4 and 24 h) after injection is beneficial for image interpretation. Method Thirty-five patients underwent 123I-MIBG imaging at 4 and 24 h after injection. Images were shown individually to 2 observers, blinded to patient identity and image time. Overall scan appearances were recorded on a 5-point scale (definitely abnormal to definitely physiological uptake). Images were then shown side by side and any change in confidence was noted. Results Interpretation of images is shown in the Table 1 below for one observer. The second observer considered 29/35 studies to be definitely abnormal/physiological after viewing 4- and 24-h

Table 1

Interpretation of images

Image presented 4-h scan 24-h scan 4- and 24-h scans

Definitely abnormal Probably abnormal or physiological or physiological 9 11 13

22 19 19

Uncertain 4 5 3

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BNMS meeting: Abstracts A13

images simultaneously compared to 11/35 at 4 h. The main area of uncertainty was activity in the renal area. Conclusion Simultaneous viewing of images acquired at 4 and 24 h resulted in some improvement in confidence of interpretation.

Result The results indicated that the in-vivo incorporation of [methyl-14C]choline in rat mammary tumour MCF-7 cells strongly correlated (P < 0.001) with cell proliferation. Conclusion [Methyl-11C]choline PET may be used as a measure of cell proliferation.

P18 Value of co-registered imaging in octreotide scintigraphy S.C. Geary, S.C. Chua, R.M. Smith and R. Ganatra University Hospital Nottingham, UK.

P20 Comparison of the finger dose received when injecting patients with and without using syringe shields M.J.M. Pearson, L. Welsh and I. Driver Newcastle General Hospital, UK.

Abdominal carcinoid and other neuro-endocrine neoplasms can be difficult to diagnose with 111In-octreotide planar scintigraphy due to overlapping structures below the diaphragm and heterogeneous liver uptake. Furthermore, liver metastases from carcinoid tumours are poorly localized to anatomical segments even with single photon emission tomography (SPET). Conventional cross-sectional imaging with computed tomography (CT) and magnetic resonance imaging (MRI) may give a false negative result due to the small size and frequent intra-luminal location of the primary tumour. Liver metastases under 1 cm are poorly characterized by CT. Identification of the primary tumour and exclusion or correct localization of liver metastases is crucial in determining whether the patient has a chance of surgical cure. At our institution routine use of SPET with non-contrast CT at the same sitting (Hawkeye, GE) in these patients has aided in the correct diagnosis and staging. We present 5 cases of carcinoid tumours where diagnostic accuracy was improved by coregistration.

Purpose To compare the dose received with and without syringe shields when administering radiopharmaceuticals to the patient, taking into account the increased time involved in using syringe shields. Method The AEGIS ED2 Personal Extremity Dosimeter was used to monitor the dose received in 1-s windows when injecting 19 bone scan patients. Each injection of 99mTc-HDP had an activity of 600 ± 60 MBq. The same technologist carried out all the injections. The probe was worn under the technologist’s glove, taped to the middle finger of the right hand. Results The mean dose without the syringe shields was 35.0 ± 7.6 mSv, compared with 10.2 ± 2.5 mSv when using the syringe shields. This shows that although the procedure may take longer, use of a syringe shield results in a significant reduction in finger dose. A t-test for unequal variances gave a P value of 9  107. The majority of the dose obtained when using the syringe shield occurs when transferring the syringe from the lead pot to the shield. The mean dose for the injection itself while using the syringe shield was 2.3 ± 0.7 mSv. Conclusion The use of syringe shields results in a significant reduction in finger dose.

P19 Incorporation of [methyl-14C]choline into rat mammary tumour cells: correlation with proliferation F. Al-Saeedi Department of Nuclear Medicine, Faculty of Medicine, Kuwait University (Health Sciences), Kuwait. Purpose Recently, [methyl-11C]choline PET was introduced to image many tumours, which are characterized by uncontrolled increase in cell proliferation. The aim of this study was to investigate whether cell proliferation could be detected by/or related to choline incorporation or not. In this study, the relationship between in-vivo [methyl-14C]choline incorporation and cell proliferation was investigated in rat mammary tumour (MCF-7) cells using the common cell proliferation assay, [methyl-3H]thymidine incorporation. Method Pieces of breast cancer (MCF-7) tissue (1 mm3) were implanted into the backs of two groups of female Wistar rats. The tumour in one group was allowed to grow to 0.5 cm3 (rapidly proliferating cells), whilst the other group to 1 cm3 (slowly proliferating cells) in diameter. When the tumour from each group reached the required volume, the rats from each group were injected simultaneously under light anaesthesia with 3.7 MBq (100 mCi) [methyl-14C]choline and [methyl-3H]thymidine into a tail vein for 10 min. This is to simulate the incorporation of [methyl-11C]choline in tumour cells during a PET scan. After 10 min, rats were dissected and the tumours were harvested, weighed and dissolved in Solvable (DuPont, Boston, MA) and counted for 3H/14C activity in a liquid scintillation counter.

P21 A functional approach to designing a new nuclear medicine department D.A. Ibbett Derby Hospitals NHS Foundation Trust, UK. Two departments with very different physical layouts are being combined into a single department in a new PFI hospital at Derby. A functional design method has been used to develop the new department’s design. Staff have been consulted at all stages. Initially, the following set of design goals were defined: simple and minimal patient/staff flow; high quality environment for patients and staff; high safety standards; good communication facilities; easy manual handling of patient around the department; high standards of patient confidentiality, dignity and care; compliance with legislation (IRR99, RSA93 etc.). The design process resulted in the development of the following suites of rooms: reception & waiting, radiopharmacy, dispensing & administration, measurement, imaging, staff and administration (clerical). The final design contains many new features that should enable ionizing radiation and other operational risks to be reduced. The most significant design constraints have been the shape and size of the space allocated and a lack of knowledge, by our PFI partners, of relevant legislation and operational practice. Establishing clear design concepts at the start of the design process has given us a strong negotiating position with our PFI partner and has enabled us to achieve many of our design goals.

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P22 Influence of gamma camera characteristics on the quantitative analysis of 99mTc-depreotide images in lung cancer staging N. Dulai and A. Nicol The Victoria Infirmary, Glasgow, UK. Background A pilot multicentre trial is under way to assess Tc-depreotide (NeoSPECT) in the staging of lung cancer. Imaging of solitary pulmonary nodules (SPNs) and lymph nodes (LNs) is being performed using different gamma cameras. Dualtime imaging is being investigated to differentiate between malignant and benign uptake. Aims To perform a phantom study to (1) determine the influence of gamma camera characteristics on the quantitative analysis, and (2) investigate whether changes in uptake can be reliably detected during dual time-point imaging (2 h + 4 h). Method A chest phantom containing lung, SPN and LN (0.25– 9 ml) inserts was used. The effect of changing the gamma camera characteristics (spatial resolution and sensitivity altered by changing the orbit of rotation and acquisition time respectively) was examined using a series of SPECT scans. Scans were performed for two different levels of SPN/LN activity concentration. ROIs were drawn around the SPN/LN and the counts per pixel per second (cpp
s – 1) noted. The quantitative analysis was repeated when the SPN/LN activity was changed by ± 10% and ± 20%. Results Significant differences in the cpp
s – 1 were obtained for the orbits of rotation investigated (P < 0.001). No significant difference in cpp
s – 1 was detected for the acquisition times examined. No significant difference in cpp
s – 1 was detected when SPN/LN activity uptake was changed by ± 10% and ± 20%. Conclusion Quantitative measures (cpp
s – 1) were found to be influenced by spatial resolution but independent of the acquired counts investigated. As changes in activity of ± 10% or ± 20% could not be detected, it is proposed that dual-time imaging is unlikely to be beneficial. 99m

P23 A normal range for gallbladder ejection fraction using a fatty meal stimulus P.D. Hay, A.A. Bolster, J.B. Neilly and H.W. Gray Department of Nuclear Medicine, Glasgow Royal Infirmary, UK. Aim To estimate a ‘normal’ range for a 60-min gallbladder ejection fraction following ingestion of a standard stimulus. Methods In the absence of data from asymptomatic volunteers the normal range was calculated using symptomatic patients who underwent imaging but were subsequently diagnosed with non-biliary disease in the following year. The case records of patients, scanned between May 2002 and December 2003, were reviewed. Each patient had received a standard clinical work-up for chronic upper abdominal pain before attending for a gallbladder scan. The scanning protocol was as follows: injection with 150 MBq of 99mTc-labelled brom-IDA, acquisition of a dynamic series of 1-min frames over 1 h, ingestion of 40 ml of a liquid containing 20 g fat, and acquisition of a 1-min static image 1 h later. A decay corrected gallbladder ejection fraction was then calculated. Results Patients who were subsequently demonstrated to have histopathologically confirmed cholecystitis were excluded, as

were patients diagnosed with chronic diseases known to affect hepatobiliary function. The case records of 81 patients were requested for review, of which 51 were excluded from the calculation for a variety of reasons. The normal range was estimated to be 49–100%, median 76%, 1st quartile 65%, 3rd quartile 88%. Conclusion The results suggest that a 60-min gallbladder ejection fraction of greater than 50% would be expected in ‘normal’ subjects. P24 Tumour localization using markers and an interoperative PET probe P.A. Beaumonta, R. Mortonb, D. Murraya, M. Kissinb, E. Bellamyc, G. Plantd and J.R.W. Halle a Guildford Diagnostic Imaging, bRoyal Surrey County Hospital, Guildford, cSt. Peter’s Hospital, dNorth Hampshire Hospital, and e Frimley Park Hospital NHS Trust, UK. Sentinel node localization using a gamma probe is becoming an established technique, particularly in breast cancer. In this study a specialized intra-operative PET probe was used to localize a thoracic abnormality using 18F-FDG. The patient with previous bilateral breast cancer demonstrated a thoracic abnormality on a PET/CT scan. Even with the CT registration it was difficult to determine if the area of increased FDG uptake involved the rib or internal mammary node due to the patient’s bilateral breast prosthesis. A further scan was performed which included a limited (1 bed position) view to position (metal) markers. The patient then proceeded to surgery where an intra-operative 18F PET-Probe (IntraMedical Imaging, LLC, USA) was used to localize the uptake. The probe successfully aided the surgeon in locating the tracer in a tumour deposit behind the rib. Prior to surgical intervention a full risk assessment was performed and appropriate precautions taken to limit dose to all staff involved. Whole-body dose to the surgeon and anaesthetist was 32 mSv and 20 mSv, respectively, and the surgeon’s finger dose was 0.4 mSv. The 18F intra-operative probe proved extremely helpful in localizing the malignant deposit identified on PET/CT and warrants further evaluation. P25 The impact of reduced diagnostic reference level on results for the investigation of lymphoedema A.M. Harris, C.M. Taylor and W.B. Tindale Department of Medical Physics and Clinical Technology, Sheffield Teaching Hospitals NHS Trust, UK. The ARSAC notes for guidance state that the DRL for the use of 99mTc-labelled colloids for the investigation of lymphoedema is 40 MBq, which was interpreted locally as applying per limb investigated. Recent information from the ARSAC suggests that the whole-body DRL should be 40 MBq per patient investigation. The impact of reducing the injected activity from 40 MBq to 20 MBq per limb was considered prior to changing the local DRL. Static images of 500 s each are normally acquired at four different time points post-injection and the percentage nodal uptake is calculated at each. Three consecutive patients were scanned using a 2-frame dynamic image (250 s per frame) to simulate the image quality and count statistics produced by

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BNMS meeting: Abstracts A15

20 MBq injected activity with a 500-s imaging time, and allowing the summing of the data to represent the 40 MBq data. The differences between the percentage nodal uptake calculated from the simulated 20 and 40 MBq data were assessed. This showed a maximum absolute difference in uptake of 0.4% for a lymph node uptake of 6.1%. Additional

analysis demonstrated that the differences were of the order expected due to statistical variability and the inherent intra-operator variability of the processing. None of the differences in the calculated nodal uptake between the results of the 20 and the 40 MBq data were considered to be significant.

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Editorial

PACS man: Questioning nuclear medicine and PET integration Edward J. Somer Nuclear Medicine Communications 2006, 27:601–602

Received 7 April 2006 Accepted 7 April 2006

Correspondence to Mr Edward J. Somer, MSc, The PET Imaging Centre, St Thomas’ Hospital, Lambeth Palace Road, London. SEI 7EH. Tel: þ 0044 207 188 4988; fax: þ 0044 207 620 0790; e-mail: [email protected]

At its conception the principal requirements of a picture archiving and communication system (PACS) were a robust server that could archive and distribute computed tomography (CT), magnetic resonance and CR/DR images within a departmental or enterprise-wide environment and workstations with the necessary tools to view these images. The problems faced by PACS were the same as those faced by all IT systems – the problems of archiving and communication.

a useful starting point although these have been overtaken by the development of PET-CT and SPECT-CT and fused image display is a particular problem. The PET and CT components of a PET-CT are acquired at different times, with a different field-of-view, different matrix size, different pixel size and even different slice thickness yet the relevant parts of the DICOM standard that handle the registration, interpolation and fusion of multimodality data are still under discussion.

Redundant array of inexpensive disks (RAID) and network-attached storage (NAS) technology, coupled with increased network speeds and the development of internet protocols, have largely resolved these issues and, in all but the most specialized cases such as mammography, the problems of ‘radiological PACS’ are more fiscal than technological.

A solution to the problem of displaying fused images has been proposed [3] and implemented by some manufacturers but this ‘ready-for-display’ data cannot be rethresholded, fusion opacity is fixed and, as ‘secondary captures’, lack the image geometry information required for further processing. Such images are non-diagnostic and have little more value than a single representative screen print.

As the original digital imaging modality, nuclear medicine had already developed its own alternatives to standards such as DICOM, and has been reluctant to give these up in favour of the limited advantages offered by PACS integration. Similarly, PACS systems have always been built on the assumption that images are two-dimensional, static and monochrome. To handle positron emission tomography (PET) and nuclear medicine data correctly would involve a complete system redesign and with no demand to drive what is a relatively small market compared to conventional radiology, the large PACS vendors have largely ignored the requirements of nuclear medicine and PET. The principal hurdles to PACS and nuclear medicine/ PET integration have been listed as [1]: K loss of information stored in manufacturer’s specific private fields K poor support for the PET or nuclear medicine portions of the DICOM standard (IODs) K the assumption that the PET and nuclear medicine IODs are the same thing K poor image display design and functionality In terms of defining the minimum display functionality the Procedural Guidelines for Telenuclear Medicine [2] provide

More worryingly, there are still issues with manipulation of display thresholds, colour scales, display of files containing more than one view or a dynamic series, postage-stamp size images, semi-quantitative display and support for multi-planar reslicing (MPR) within generic PACS workstations. Without entering the debate on the propriety of releasing complete data sets for display on a PACS workstation where it may be re-interpreted, there are few instances where trust-wide PACS access may be justified. One example is the multidisciplinary meeting where full integration allows the interactive display, even projection of images under discussion. However, to be useful the PACS workstation would need to replicate the basic functionality of the modality workstation and, while phrases like ‘multimodality PACS’ have a certain verisimilitude, wide-scale implementation remains unrealistic. While there are many suitable visualization applications available, the companies that supply and maintain large PACS systems are reluctant to allow the installation of third-party applications, citing security, integrity and

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maintenance response conflicts. This lack of support for nuclear medicine and PET in trust-wide PACS has led to the adoption of a departmental mini-PACS with dedicated ‘modality workstations’. These systems, often supplied by smaller, independent, specialist vendors are more easily tailored to individual needs and can preserve legacy data and applications while offering the advantages of DICOM connectivity. A PACS cannot work effectively in isolation and it makes little sense to invest in PACS while still passing paper requests and reports to and from the office. Unfortunately, the integration of PACS and radiology/hospital information systems (HIS/RIS) with imaging modalities introduces yet another set of standards. Rather than setting new standards the IHE (‘Integrating the Healthcare Enterprise’) is an international organization that promotes the co-ordinated use of established standards such as DICOM and HL7 through the adoption of ‘Integration Profiles’. The implementation of these profiles can then be validated at events called ‘connectathons’ where vendors get together to check their understanding of the profiles against at least three other systems. Today, nobody would buy imaging equipment without a DICOM conformance statement even though this offers no guarantee that the expected functionality will be included or enabled. We should also be asking for IHE Integration statements and ‘connectathon’ results, remaining aware that the first are product specific but not externally verified while the second are externally verified but not necessarily product specific. In terms of implementation, the nuclear medicine profile and, more recently, the image fusion profile are helpful steps but the radiology technical framework is not modality specific and the same workflow and profiles apply to all modalities. Any differences that nuclear medicine or PET may have to CT and magnetic resonance are minuscule compared to the similarities, so there is no reason why nuclear medicine and PET usage should not follow the existing profiles just like other modalities. Nuclear medicine and PET need to look towards embracing the entire technical framework although there is no reason why this cannot be implemented in stages, starting with the ‘scheduled workflow’.

The IHE is good for users but informal discussion at the recent BNMS meeting gave the impression that manufacturers are a little sceptical of the whole IHE process, although most of them had a presence on the UK IHE technical committee. Why would a company that sells both PACS and imaging equipment openly share sensitive information so customers can equip their site with components from alternative suppliers without compatibility concerns? Perhaps because even sites where a single vendor has a monopoly often have trouble integrating their systems, perhaps because the size of the nuclear medicine/PET market is such that nobody can afford to turn down a camera sale just because the customer wants alternative image reporting software. Politically driven projects such as ‘NHS PACS’, ‘Choose and Book’ and the increasing deployment of mobile imaging units will increase the pressure upon nuclear medicine and PET departments to integrate with largescale HIS/RIS and PACS. However, until these systems offer the most basic level of functionality it is only sensible that departments adopt the mini-PACS model. Where mini-PACS is already in place, IT project managers must recognize its necessity and be flexible in the application of the ‘Connecting for Health’ connection template. Nuclear medicine departments must maintain their functionality and if trust/NHS PACS integration is limited to the sending of screen captures reconciled with a HIS/RIS workflow, that remains a giant leap forward. Where image data is sent out to referring clinicians via CD, NHSnet/N3 or even the Internet, it must be accompanied by a fully functional viewing application to facilitate meaningful discussion and set a standard to help illustrate the shortcomings of generic PACS for nuclear medicine and PET.

Acknowledgement With thanks to Andrew Kettle of East Kent Hospitals NHS Trust for his helpful comments.

References 1 2 3

Wallis JW. Nuclear PACS: problems, solutions. Decisions in Imaging Economics 2004; 6:21–25. Parker JA, Wallis JW, Jadvar H, Christian P, Todd-Pokropek A. Procedure Guidelines for Telenuclear Medicine 1.0. J Nucl Med 2002; 43:1410–1413. Vogel WV, Oyen WJG, van Dalen JA, Huisman H, Karssemeijer N. Sliced alternating DICOM series: convenient visualisation of image fusion on PACS. Eur J Nucl Med Mol Imaging 2005; 32:247–248.

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Original Article

Applied radioactivity in radiation synovectomy with [188Re]rhenium sulfide suspension Peiyong Lia, Junfeng Yub, Gang Chena, Xufeng Jianga, Zhonghua Tanga, Suyun Chena, Lei Jianga, Lin Tangb and Duanzhi Yinb Background Early experience demonstrated absorbed dose in radiation synovectomy is about 100 Gy. For reaching this dose, the applied radioactivity should be calculated. Method Twenty-nine synovitic models of rabbit were treated by intra-articular injection of [188Re]rhenium sulfide and histological changes of synovium and cartilage were examined. The applied radioactivity was calculated by method of absorbed dose factor. In clinical, eleven haemophilic patients with haemarthrosis were performed radiation synovectomy with treated [188Re]rhenium sulfide. The synovial thickness was evaluated by MR and its value was used to calculate the applied radioactivity. After radiation synovectomy, all patients were followed up by synovial thickness, regional inflammation, and clinical course including bleeding frequency. Results In rabbit models, the synovitic membrane can be eliminated by calculated radioactivity as planed without damaging the joint cartilage. In patients study, all patients exhibited significant reductions in synovial thickness and inflammation after radiation synovectomy with the planed radioactivities of [188Re]rhenium sulfide. Post-procedure bleeding frequency reduction in excellent and good reached to 63.6% by 18 months. In the cases of joint

Introduction Haemophilia A is a congenital blood defect responsible for a partial or total absence of clotting factor VIII. The abnormal bleeding originates from the subsynovial venous plexus underlying the capsule, where a lack of thromboplastic activity has been demonstrated. Synovial hypertrophy and eventual destruction of articular cartilage can ensue if recurrent haemarthrosis is not arrested at an early stage. Radiation synovectomy by injection of radioactive materials was first introduced in the United States in 1963 for rheumatoid arthritis [1]. Ahlberg and Pettersson began using radioactive gold in haemophilic patients in Sweden in 1971 [2]. Since then it has been developed as an alternative to surgical synovectomy by intra-articular injection of a beta-emitting radionuclide in colloidal or particulate form. Phagocytic lining cells along the synovial surface quickly absorb some of the injected radioactivity from the capsule. Provided the amount of radioactivity injected is sufficiently large to deliver a therapeutic dose to the synovium, it will be destroyed.

bleeding, the need for antihaemophilic factor treatment decreased immensely. Most of the recurrent episodes of bleeding were mild, subsiding with local means. Conclusion The applied radioactivity in radiation synovectomy could be calculated according to thickness of inflamed synovium. Further study including comparison therapeutic results from calculated individual activities with results from fixed activities and long-term follow-up is c 2006 warranted. Nucl Med Commun 27:603–609
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:603–609 Keywords: [188Re] rhenium sulfide, radiation synovectomy, radioactivity, absorbed dose a Department of Nuclear Medicine, Rui Jin Hospital, Shanghai, and bShanghai Institute of Applied Physics, Chinese Academy of Sciences, P.R. China.

Correspondence to Dr Peiyong Li, Department of Nuclear Medicine, Rui Jin Hospital, Shanghai 200025, P.R. China. Tel: + 0086 21-53064539; fax: + 0086 21 643 33548; e-mail: [email protected] This work was supported partially by Key Project of Knowledge Innovation Program of Chinese Academy of Sciences (Contract No. KJCXI-SW-08), and by the National Natural Science Foundation of China (No. 20501022). Received 22 January 2006 Accepted 31 March 2006

The several potential advantages of radiation synovectomy over surgical synovectomy include (1) a minimal requirement of anti-haemophilic factor, (2) performance on an outpatient basis, (3) concurrent treatment of several joints, and (4) low risk of haemorrhage in patients with inhibitors. 188

Re is suitable for treatment of the knee owing to its deep tissue penetration (maximum 11 mm, average 3.8 mm) and relatively short physical half-life (16.9 h). It is available from an in-house generator system or its gamma ray also can be used to imaging for monitoring its leakage. In the past, radiation synovectomy with [188Re] rhenium sulfide colloid and 188Re tin colloid have been performed [3–5]. Early experience demonstrated a target dose of approximately 100 Gy to be necessary for the therapeutic effect of radiation synovectomy. The activity to be applied to obtain a dose of 100 Gy was often derived from the distribution volume of the radiation sources [6,7]. However, it is very difficult to calculate the volume

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of an inflamed synovial tissue as described by Johnson et al. [8]. The hypothesis of this study is whether the applied radioactivity could be calculated personally in radiation synovectomy according to the thickness of a patient’s synovitic membrane. In radiation synovectomy, the synovitic membrane should be eliminated without damaging the joint cartilage. In previous work, beta-particle dosimetry in radiation synovectomy has been estimated by absorbed dose factors described by Johnson et al. [8]. However, this method was used only in phantom experiments and has not been used in an animal experimental study with histological examination or in clinical practice of radiation synovectomy. In this study, the experimental radioactivity of a [188Re]rhenium sulfide suspension was calculated. The effects of [188Re]rhenium sulfide were evaluated by histology. In clinical work, the thickness of the synovium in haemophic arthroses was evaluated by magnetic resonance and its value was used to calculate the applied radioactivity of [188Re]rhenium sulfide. The therapeutic effect of radiation synovectomy was then evaluated in clinical follow-up.

Materials and methods [188Re]rhenium sulfide suspension

Carrier-free 188Re was produced by a 188W/188Re generator (Oak Ridge, Tennessee, USA). As our previous study reported, the [188Re]rhenium sulfide suspension was prepared by the reaction of sodium thiosulfate with potassium perrhenate in acid solution [9]. The particle size used in the study was 1–10 mm as determined by microscopy. In-vitro stability

The method for stability testing was described by Wang et al. [10]. Briefly, aliquots of [188Re]rhenium sulfide suspension were added to tubes containing 3 ml of synovial fluid, 1% bovine serum albumin and phosphate buffered saline (0.1 M, pH 7.4), respectively. The synovial fluid was drawn from the inflamed joints of patients with rheumatoid arthritis. The tubes were stoppered and mixed continuously on a rotator. At intervals of 4 h, 1 day and 3 days, the tubes were removed and centrifuged at 500  g for 5 min. Aliquots of the supernatants were counted by a gamma counter. All counts were corrected for radioactive decay, and expressed as a percentage of the total radioactivity measured at the beginning of the experiment. Calculation of applied radioactivity

The applied radioactivity was calculated by the equation suggested by Johnson et al. [8]. For the long retention time of [188Re]rhenium sulfide in joints relative to its half-life, the radiation dose throughout treated joints can

be estimated as follows: Di ¼

A0 Fi lp Ssyn

where Di is the total dose (cGy); in this study it was set to four values: 33 Gy, 66 Gy, 99 Gy and 165 Gy. The 99 Gy was in line with the early experience of absorbed dose in radiation synovectomy; A0 is the applied radioactivity initially deposited in the source region (MBq), which is to be calculated; and lp is the physical decay constant for the injected radionuclide. For 188Re, lp is 0.14 10 – 4s – 1. Fi is the mean absorbed factor (cGycm2MBq – 1s – 1) and is related to the thickness of synovium. This value was set at 0.05 as the thickness of rabbit synovium is less than 1 mm; Syn is the total synovial surface area of the joint and was set at 8.0 cm2 in this experiment. According to the above equation and setting, the corresponding applied radioactivity was 7.4, 14.8, 22.2 and 37 MBq. Animal studies

By using the method described by Steinberg et al. [11], an active synovitis model of rheumatoid arthritis was induced in 29 mature New Zealand white rabbits by injecting ovalbumin antigen into the knee joint. The animals were obtained from the Shanghai Institute of Experimental Animals and were fed with a standard diet and water. The experiments were conducted according to the Principles of Care and Use of Laboratory Animals, which are set by the Shanghai Second Medical University. Each rabbit weighed approximately 3 kg and was injected via an anterior subpatellar approach into the knee joint with 0.2 ml sterile [188Re]rhenium sulfide suspension containing different doses of radioactivity. Twenty-three rabbits were divided into five groups, including a negative control group, which received 0.2 ml of cold rhenium sulfide in three rabbits; four treatment groups with five rabbits each received a radioactive [188Re]rhenium sulfide suspension at 7.4, 14.8, 22.2 and 37 MBq, respectively. Another six rabbits were used for an in-vivo biodistribution study with 22.2 MBq. The rabbits were cultured without immobilization after injection for 24 h. Whole-body imaging analysis of the distribution of [188Re]rhenium sulfide in each rabbit and retention of radioactivity in the knee joint was performed at 1, 24 and 72 h after injection, using an ADAC Vetex gamma camera adjusted to 155 keV gamma rays (ADAC Laboratories, Milpitas, USA) with a low-energy collimator: the distance between the table and the detector maintained constant. Data were stored by a computer for subsequent analysis. The percentage of radioactivity retained in the injected knee was calculated by the method of Sledge et al. [12]. Briefly, the radioactivity in the injected knees was counted and the amount was calculated as a percentage of the [188Re]rhenium sulfide suspension injected.

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Radiation synovectomy with [188Re]rhenium sulfide suspension Li et al. 605

For in-vivo analysis of the biodistribution of the [188Re]rhenium sulfide suspension, three rabbits were killed at each of 4 and 24 h. Radioactivity in the thyroid, liver, spleen, kidney and iliac lymph node at the injected lateral side was counted. The leakage of radioactivity in tissue was calculated by the percentages of activity counts of tissues in total injected radioactivity. Three months after injection, 23 rabbits were killed. Knee joints of the dead rabbits were dissected and sectioned for histological examination. Changes in synovium and cartilage were examined using haematoxylin and eosin stain or toluidine blue stain, respectively. Primary clinical studies

A total of 11 haemophilic patients (male, aged 19–35 years) with symptoms of haemarthrosis and pain in the knee were enrolled in this study. The coagulation FVIII in all patients ranged from 1 to 4% by coagulation assay. Patients had a bleeding frequency of at least two episodes per month in a target joint and conservative treatment failed, which included a combination of clotting factor concentrate and physical therapy. There are no major contraindications for this procedure. Written informed consent was obtained from each patient before entry into the study. Approval from the Ethics Subcommittee of Rui Jin Hospital for Research Involving Human Subjects was received. All patients had been followed clinically for 6–18 months. Three parameters were reviewed to evaluate the results, as measured by a reduction in frequency of haemarthroses, requirements for anti-haemophilic factor, and number of days of treatment. The frequency of haemarthroses was evaluated by comparing average joint bleeds per month before and after radiation synovectomy. Post-procedure haemarthroses were recorded after 3, 12 and 18 months. A total absence of joint bleeding was rated excellent. A reduction in bleeding between 75 and 99% was rated good, between 50% and 74% as fair and below 50% as poor. Radiation synovectomy

The management protocol is (1) to increase patients’ anti-haemophilic factor to 15–20%; and (2) to inject [188Re]rhenium sulfide suspension, containing different amounts of radioactivity, via the intra-articular route according to the synovium thickness, as shown by magnetic resonance. The synovium thickness was classified into three groups: < 4 mm between 4 and 5 mm and thicker than 5 mm. After intra-articular injection of the [188Re]rhenium sulfide suspension, the injection needle and the puncture channel have to be flushed so that radiation necrosis along the puncture channel can be avoided. After removing the needle, the puncture site was squeezed off with a gauze, fixed with an elastic bandage and the joint, under manual pressure on the puncture site, was carefully moved twice. Then a splint or cast is formed along the physiological position of the joint and the joint immobilized. Four days after the injection, the plaster of Paris is removed.

Imaging study

Magnetic resonance and gamma scintigraphy with 99m Tc-IgG were used to evaluate synovial thickness, articular cartilage and inflammation of the joint. The magnetic resonance examinations were performed on a 1.5-T magnetic resonance unit (Signa 1.5, GE company) with a circular polarized knee coil. The following sequences were applied: a sagital T1-weighted spin-echo sequence, repetition time (TR)/echo time (TE) 400/14 ms and saggital T2-weighted spin-echo sequence (TR/TE 2000/ 89 ms). Other relevant parameters were: NEX, 4; field of view, 200; matrix, 256  190; slice thickness, 2 mm and slice space 1 mm. Synovial measurements were performed three times in each patient in three regions (retropatellar, lateral and medial). The mean of synovial thickness was calculated. The synovial structure (smooth, cushion-like, fine or rough villous, oedematous, coarse) was also evaluated. Gamma scintigraphy for detecting inflammation was performed in each patient by using the routine procedure. Seven hundred and forty MBq of 99mTc-IgG was injected intravenously and the same camera as in the rabbit study was used. The accumulation of radioactivity in affected joints was compared before and after radiation synovectomy over 2 months. Estimation of activity for therapy

Earlier experience showed that a target dose of about 100 Gy was necessary for the therapeutic effect of radiation synovectomy. The radioactivity to be applied to obtain a dose of 100 Gy was calculated by the above equation used in experimental study. Here, the Fi value was set to 0.5  10 – 2, 0.4  10 – 2 and 0.3  10 – 2, corresponding to < 4 mm, 4–5 mm and more than 5 mm distances, respectively, defined in relation to the synovium and radioactive source interface. Ssyn is the total synovial surface area of the joint; for the knee it is 250 cm2. The patients were assigned to three groups receiving 550, 687 or 917 MBq of [188Re]rhenium sulfide, corresponding to synovial thicknesses of < 4 mm, 4–5 mm and more than 5 mm, respectively, as measured by magnetic resonance. Statistical analysis

Student’s t-test was used for the intra-group comparison between the baseline and post-treatment measurements and the Kruskal–Wallis test for the inter-group comparison among three treatment groups. Data were analysed using the Statistical Analysis System package (SAS version 6.12) and P < 0.05 was regarded as statistically significant.

Results Animal study

In-vitro stability tests revealed that after incubating [188Re]rhenium sulfide with different solutions, the

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Table 1

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The in-vivo biodistribution of [188Re]rhenium sulfide after intra-articular injection (tissue ID ¾ 10 – 2) in rabbits

Time after injection

4 h (n = 3) 24 h (n = 3)

Tissue Liver

Kidney

Spleen

Lung

Iliac lymph node

Thyroid

0.15 ± 0.017 0.62 ± 0.11

0.06 ± 0.34 1.22 ± 0.28

0.23 ± 0.011 0.78 ± 0.17

0.11 ± 0.009 0.25 ± 0.06

0.62 ± 0.14 1.02 ± 0.11

Undetectable Undetectable

n is the number of rabbits.

Fig. 1

Histological finding of cartilage at three months after [188Re]rhenium sulfide intra-articular injection. (Toluidine blue,  100, small area in angle,  200). (a) With non-radioactive rhenium sulfide, the chondrocytes are arranged in a regular manner, forming the tide mark. (b) With 37 MBq of [188Re]rhenium sulfide, the osteoblasts have denatured and are arranged irregularly. The tide mark had disappeared.

labelling efficiency of the sulfide was still greater than 98% over a 3-day period. Imaging the distribution of [188Re]rhenium sulfide in vivo at 1 and 24 h after injection, showed that the radioactivity was highly concentrated in the injected joint. No sign of radioactivity could be seen in other parts of the rabbit body, including the kidney and lymph nodes. The mean percentages of retained radioactivity in the injected knee, determined by gamma camera imaging, were 98.5 ± 1.2%, 95 ± 2.4% and 92 ± 2.6%, at 1, 24 and 72 h after injection, respectively. The biodistribution of [188Re]rhenium sulfide in vivo showed that 1.22% of injected radioactivity was in the kidney and 1.02% was in the iliac lymph node. The amount of radioactivity in other tissues was negligible (Table 1). The synovial membrane from synovitic knee joints of rabbits showed hyperaemia, synovial lining cell hyperplasia and a remarkable inflammatory cell infiltration (Fig. 1(a)). With 7.4 MBq of [188Re]rhenium sulfide intra-articular injection, there was no effect on the synovial membrane, and the synovial lining cell was still hyperplasic. With 14.8–22.2 MBq of [188Re]rhenium sulfide, the synovial membrane became thin and the infiltration of inflammatory cells decreased. The cartilage under the synovium had no sign of injury (Fig. 1(a)). However, when the dose of radioactivity was increased to 37 MBq of [188Re]rhenium sulfide, the cartilage under the synovium

was injured, the arrangement of chondrocytes became irregular and denatured cells occurred (Fig. 1(b)). Patient study Imaging study

Synovial thickness and inflammation All patients except three exhibited significant reductions in synovial thickness after radiation synovectomy. Magnetic resonance imaging (MRI) showed that the pathological changes in the synovitic joint are joint space enlargement, thick synovium, rough villous and oedema. In follow-up, MRI indicated that the synovectomy with [188Re]rhenium sulfide thinned the synovium, reduced the synovia, and removed the proliferated villi (Fig. 2). Gamma scintigraphy for detecting inflammation showed that radioactivity in the affected joint was decreased. This revealed the inflammation was reduced after the radiation synovectomy with [188Re]rhenium sulfide (Fig. 3). Articular cartilage There was no evidence of any immediate damage to the articular cartilage. After an observation period of 18 months, the homogeneity of the articular cartilage was unaltered in all but three patients in whom the distribution of radioactivity was different. MRI of these three patients revealed previously undetectable superficial erosions.

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Radiation synovectomy with [188Re]rhenium sulfide suspension Li et al. 607

Fig. 2

A decrease in synovial thickness and villi (arrows) on sagittal T2-weighted magnetic resonance (MR) images before (a) and after (b) radiation synovectomy.

Fig. 3

Gamma scintigraphy with 99mTc-IgG showed the radioactivity in affected right knee was decreased obviously after the radiation synovectomy with 687 MBq of [188Re]rhenium sulfide intravenous injection. (a) Before radiation synovectomy; (b) 6 weeks after radiation synovectomy with 687 MBq of [188Re]rhenium sulfide intravenous injection.

Adverse effects

Moderately increased swelling and local tenderness were observed in three patients within a few days after treatment, with a maximum duration of 5 days. The adverse effects have no relation to applied dose of radioactivity. No special treatment was undertaken except the physical treatment to resolve the symptoms. Most of the patients did not complain of any discomfort after the administration, such as pain in the superficial tissues around the injection point.

Clinical course

Before radiation synovectomy, patients were bleeding, on average, 2.9 times per month (range, 2–5; SD = 3.1). The average frequency at 3 and 12 months after the radiation synovectomy were 0.4 (range, 0–2; SD = 1.1) and 0.7 (range, 0–3; SD = 0.9) bleeds per month, which represents, respectively, a reduction of 90.2% (P < 0.001) and 73.6% (P < 0.001). By 18 months, patients reported an average of 1.2 (range, 0–3; SD = 1.9) bleeds per month, representing a reduction of 63.2% (P < 0.001). Excellent results (total absence of joint bleeding) were obtained in 63.6% of cases (7/11) at 6 months of follow-up. This number decreased over time, corresponding to 45.4% (5/11) by 18 months. Good results (reduction in joint bleeding between 75.0 and 99.0%) were found in 27.2% (3/11) of cases at 6 months and 18.2% (2/11) cases at 18 months. Poor and fair results (reduction in joint bleeding lower than 75%) were found in 9.1% and 36.4% (4/11) of cases at 6 and 18 months, respectively. In the cases of joint bleeding, the need for antihaemophilic factor treatment decreased immensely; less than 75% of the factor was required compared to the amount before radiation synovectomy (Table 2). Most of the recurrent episodes of bleeding were mild, subsiding with local means, including ice, compression bandages, rest and days of treatment. Recurrent haemarthroses that required coverage needed only 1–4 days of treatment, whereas previously they required as many as 3 weeks, with a mean of 1 week.

Discussion Usually, the vicious cycle of haemarthrosis–synovitis– haemarthrosis becomes established in haemophilic arthroses. Conservative treatment is usually inadequate. Synovectomy is the procedure of choice to break this

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608 Nuclear Medicine Communications 2006

2006, Vol 27 No 8

188 Table 2 The results of radiation synovectomy with [ Re]rhenium sulfide in three groups of synovium thickness

Synovial Number of thickness joints (mm) <4 4–5 >5

3 5 3

Radioactivity (MBq)

555 687 917

Reduction of bleeding frequency per month (%) Month 3 86 93 91

Month 12 73 84 63

Month 18 64 69 56

cycle. Radiation synovectomy is a regularly practised nuclear medicine therapy in haemophilic patients suffering from pain and haemarthroses: it is safe and of great benefit to them. The frequency of haemarthroses evaluated in this study showed that excellent and good results have reached 63.6% by 18 months after treatment. Beta-particle dosimetry in radiation synovectomy is difficult for a variety of reasons. These include the physical difficulties associated with the beta dosimetry of low-energy electrons in heterogeneous media, and biological and anatomical difficulties associated with the non-uniform distribution of radioactivity within the media. The calculation methods of absorbed dose include the medical internal radiation dosimetry (MIRD) scheme [13], radiation source distribution volume and absorbed dose factors which were used in this study. In practice, it is almost impossible to calculate the volume of an inflamed synovial tissue. A very wide range of values has been quoted for the knee synovial area. In human knee it is often regarded as 250–500 cm2. Howson et al. estimated that the synovial area of rabbit knee was 1/20– 1/30 of the human knee [14]. We had tried to measure the rabbit knee synovial area by dyeing the synovium and found its value varied largely from 5.0 to 11 cm2. So we adopted the reported value. Another limitation of this approach is the fact that synovial thickness is not consistent through the knee joint in patients. However, we chose synovial thickness as the key factor because it reflects the severity of inflamed synovium. In a previous study, a fixed amount of radioactivity was used in radiation synovectomy. Radioactivity of 74– 111 MBq 32P was applied to the knee joints in each patient. The result of synovectomy by this approach was about 70–80%, similar to that reported by Silva et al. reported that their 32P chromic phosphate study in 170 radiosynovectomies for chronic haemophilic synovitis [15]. Excellent and good results (haemarthrosis reduction from 75 to 100%) were obtained in 79.2% of cases at 6 months to 8 years. Recently, a different approach to the problem has been the use of an equation to calculate the applied radioactivity in radiation synovectomy. Pirich et al. reported that a median activity of 8.4 GBq 165Dy ferric hydroxide was applied in knee joints of 13 patients.

During clinical follow-up to 13 months, nine of 13 patients (69%) continued to respond to radiation synovectomy [16]. Our study showed an excellent and good result was 90% follow-up to 6 months but a drop to 63.6% at 18 months. Although it is too early to obtain the conclusion that the synovectomy result by calculated applied radioactivity is better than the fixed radioactivity, the more radiation synovectomy study by approach of calculated activity including more patient number and longer follow-up should be performed. The quotation approach in nuclear therapy needed to be considered by nuclear medicine physicians. Besides the 188Re, many radionuclides have been used for radiation synovectomy, including 166Ho, 90Y, 165Dy, 32P, 198 Au and 186Re. It has been confirmed that different radionuclides should be used in different joint according the physical characters of radionuclides like its mean energy and maximum soft tissue penetration. Other formulations of 188Re have 188Re–tin colloid [17]. 188Re– sulfur colloid [4] and 188Re–microspheres [10]. The main difference among these 188Re formulations is their particle size. The particle size of the 188Re sulfide suspension prepared in this study is about 1–10 mm, suitable for the radiation synocectomy. It was reported that although both 188Re–tin colloid and 188Re–sulfur colloid might be useful for radionuclide therapy; the 188 Re–tin colloid had more advantages compared to the 188 Re–sulfur colloid, due to its higher labelling efficiency, control of the particle size, and lower residual activity in the injection syringes [17]. The rational for choosing absorbed dose factors to calculate the applied radioactivity in MRI has been used to assess the activity of arthritis and its response to therapy. Synovial thickness determines the probability of therapeutic success of synovectomy [16]. By this method, the thickness of the synovium is the one of the key factors in determining the radioactivity. Whether the reduction of the synovial thickness and inflammatory activity is achieved at the price of any damage to the articular cartilage is concerned. Our synovectomy experiment with [188Re]rhenium sulfide in rabbits indicated that 14.8–22.2 MBq radioactivity intra-articular injection, the absorbed dose of synovium which calculated by absorbed dose factors is 56–84 Gy. It only has an effect on the synovium and does not affect the articular cartilage. But when the absorbed dose is raised to 150 Gy (37 MBq of [188Re]rhenium sulfide intra-articular injection), the cartilage under synovium was damaged (Fig. 2). In patients study, none of the 11 treated joints had MRI evidence of immediate cartilage damage at the early follow-up study and five patients had isolated superficial lesions of articular cartilage on the later follow-up MRI, although some patients had basal articular cartilage lesions before treatment. It is quite possible that the course of the underlying disease, rather than the

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Radiation synovectomy with [188Re]rhenium sulfide suspension Li et al. 609

treatment, caused these changes. Radiation-induced damage leads to a diffuse reduction in the volume of articular cartilage, whereas only isolated erosions were found in the retropatellar region. Moreover, radiationinduced damage should be detectable a few days after treatment, particularly when considering the fact that we used a short-lived radionuclide (half-life 16.9 h). These clinical results are also in agreement with those of other trials [6,18,19] and reveal reductions in joint pain and effusion as well as an increased range of motion in most patients.

Conclusion We concluded from this study that applied radioactivity in radiation synovectomy could be calculated according to thickness of inflamed synovium. Further study, including a comparison of the therapeutic results from calculated individual activities, with results from fixed activities and long-term follow-up is warranted.

Acknowledgement The authors thank Dr Hongli Wang for reviewing the manuscript.

References 1

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4

Ansell BM, Crook AJM. Evaluation of intra-articular colloidal gold 198Au in the treatment of persistent knee effusions. Ann Rheum Dis 1963; 22:435–439. Ahlberg A, Pettersson H. Synoviorrthesis with radioactive gold in hemophiliacs. Clinical and radiological follow-up. Acta Orthop Scand 1979; 50:513–517. Venkatesan PP, Shortkroff S, Zalutsky MR, Sledge CB. Rhenium heptasulfide: a potential carrier system for radiation synovectomy. Nucl Med Biol, Part B 1990; 17:357–362. Wang SJ, Lin WY, Hsieh BT, Shen LH, Tsai ZT, Ting G, et al. Rhenium-188 sulphur colloid as a radiation synovectomy agent. Eur J Nucl Med 1995; 22:505–507.

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Lee EB, Shin KC, Lee YJ, Cheon GJ, Jeong JM, et al. 188Re–tin colloid as a new therapeutic agent for rheumatoid arthritis. Nucl Med Commun 2003; 24:689–696. Sledge CB, Zuckerman JD, Shortkroff S, Zalutsky MR, Venkatesan P, Snyder MA, et al. Synovectomy of the rheumatoid knee using intra-articular injection of dysprosium-165-ferric hydroxide macroaggregates. J Bone Joint Surg 1987; 69A:970–975. Havlik E, Pirich C, Preitfellner J, et al. Radiation exposure from patients treated with 165Dy-ferric hydroxide. Nucl Med Commun 2001; 22:79–82. Johnson LS, Yanch JC, Shortkroff S, Barnes CL, Spitzer AI, Sledge CB. Beta-particle dosimetry in radiation synovectomy. Eur J Nucl Med 1995; 22:977–988. Yu JF, Yin DZ, Min XF, Guo ZL, Zhang J, Wang YX, et al. Preparation of [188Re]rhenium sulfide suspension and its biodistribution following intra-tumor injection in mice. J Label Compound Radiopharm 1999; 42:233–243. Wang SJ, Lin WY, Chen MN, Chen J-T, Ho W-L, Hsieh B-T, et al. Histologic study of effects of radiation synovectomy with rhenium-188 microsphere. Nucl Med Biol 2001; 28:727–732. Steinberg ME, McCrae CR, Bershelli RA, Cram B, et al. Intra-articular 5-fluorouracil in antigen induced arthritis. J Bone Joint Surg Am 1971; 53:514–522. Sledge CB, Noble J, Hnatowich DJ, Kramer R, Shortkroff S, et al. Experimental radiation synovectomy by 165Dy ferric hydroxide macroaggregate. Arthritis Rheum 1977; 20:1334–1342. Neves M, Waerenborgh F, Patricio L. Palladium-109 and holmium-166 potential radionuclides for synoviotherapy – Radiation absorbed dose calculations. Appl Radiat Isot 1987; 38:745–749. Howson MP, Shepard NL, Mitchell NS. Colloidal chromic phosphate 32 P synovectomy in antigen-induced arthritis in the rabbit. Clin Orthop 1988; 229:283–293. Silva M, Luck Jr JV, Siegel ME. 32P chromic phosphate radiosynovectomy for chronic haemophilic synovitis. Haemophilia 2001; 7(suppl 2):40–49. Pirich C, Schwameis E, Bernecker P, Radauer M, Friedl M, Lang S, et al. Influence of radiation synovectomy on articular cartilage, synovial thickness and enhancement as evidenced by MRI in patients with chronic synovitis. J Nucl Med 1999; 40:1277–1284. Jeong JM, Lee YJ, Kim YJ, Chang YS, Lee DS, Chung JK, et al. Preparation of rhenium-188–tin colloid as a radiation synovectomy agent and comparison with rhenium-188-sulfur colloid. Appl Radiat Isot 2000; 52:851–855. Clunie G, Ell PJ. A survey of radiation synovectomy in Europe. Eur J Nucl Med 1995; 22:970–976. Edmonds J, Smart R, Laurent R, Butler P, Brooks P, Hoschl R, et al. A comparative study of the safety and efficacy of dysprosium-165 hydroxide macro-aggregate and yttrium-90 silicate colloid in radiation synovectomy: A multicentre double blind clinical trial. Br J Rheumatol 1994; 33:947–953.

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Original article

Assessment of 123I-FIAU imaging of herpes simplex viral gene expression in the treatment of glioma Mary F. Dempseya, David Wypera, Jonathan Owensb, Sally Pimlottb, Vakis Papanastassioua, James Pattersona, Donald M. Hadleya, Alice Nicola, Roy Ramplingc and S.M. Brownd Background Herpes simplex virus 1716 (HSV1716), a selectively replication competent mutant of HSV1, is under investigation as an oncolytic viral therapy in human malignant glioma. As with similar therapies, a technique for measurement of viral replication and distribution over time following virus administration is required. Imaging expression of the HSV-thymidine kinase (HSV-tk) gene offers an opportunity for non-invasive assessment of viral distribution in living subjects. This is the first study to explore the use of HSV-tk as a reporter gene and radiolabelled thymidine analogue 5-[123I]iodo-1-(2-deoxy2-fluoro-b-D-arabinofuranosyl) uracil (123I-FIAU) as a marker substrate to non-invasively monitor HSV1716 replication in humans during treatment of high-grade glioma. Methods 123I-FIAU brain SPECT imaging was undertaken in eight patients receiving intra-tumoural injection of HSV1716, before and after administration of the virus. Baseline images were acquired 3 days prior to virus administration and between 1 and 5 days following virus administration. Region of interest analysis was used to investigate whether there was an increase in 123I-FIAU

Introduction Genetic modification to sensitize brain tumour cells for therapy shows great potential and is the basis of many ongoing clinical trials. Whilst many studies have demonstrated encouraging results in animal models such findings have yet to be achieved in the clinic. This is thought to be due to insufficient levels of gene expression and/or insufficient numbers of cells being targeted to gain a clinical response. A potential method of achieving a greater level of gene expression and infecting a greater number of cells within a tumour is to use replication-selective agents. This approach uses microbial agents such as viruses that replicate in human tissue. Replication leads to high level gene expression from both the infecting agent and it’s progeny. Replication selective agents can also be utilized as ‘oncolytic’ agents. Herpes simplex virus 1716 (HSV1716), a modified version of herpes simplex virus (HSV) type 1, replicates in actively dividing cells but not in terminally differentiated cells [1]. The virus lyses human glioblastoma

concentration following virus administration due to HSV-tk expression. Result Increased 123I-FIAU accumulation due to HSV-tk expression was not detected in this study. The possible explanations for this finding are explored and design options for future studies are discussed. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:611–617 Nuclear Medicine Communications 2006, 27:611–617 Keywords: glioma, herpes simplex virus, FIAU, molecular imaging a

Institute of Neurological Sciences, bWest of Scotland Radionuclide Dispensary, Beatson Oncology Centre, Glasgow, UK and dS. Moira Brown Crusade Laboratories Glasgow, UK c

Correspondence to Mary F. Dempsey, Clinical Physics, Institute of Neurological Sciences, Southern General Hospital, 1345 Govan Road, Glasgow G51 4TF, UK. This work was funded by a Scottish Executive CSO grant. Tel: + 0044 141 201 2120; fax: + 0044 141 201 2060; e-mail: [email protected] Received 16 December 2005 Accepted 4 April 2006

cells in vitro [2] and in model systems and has therefore the potential to eliminate brain tumours whilst failing to replicate in, and therefore damage, normal brain cells. One of the most important factors for making oncolytic viral therapy widely applicable, however, is the development of a method for non-invasive assessment of virus replication and distribution in tumour. Great progress has already been made in the field of nuclear reporter gene expression imaging with the most frequently studied reporter gene for nuclear imaging happening to be the herpes simplex virus type 1 thymidine kinase gene (HSV1-tk). Although almost a decade has elapsed since nuclear imaging of HSV1-tk gene expression was successfully demonstrated in vitro and in vivo [3–5], translation of this paradigm for use in humans has been limited [6]. Of the potential marker substrates for the enzyme HSV1-tk, the radiolabelled nucleoside pyrimidine derivative 5-iodo-1(2-deoxy-2-fluoro-b-D-arabinofuranosyl) uracil (FIAU) has

c 2006 Lippincott Williams & Wilkins 0143-3636

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612 Nuclear Medicine Communications 2006, Vol 27 No 8

been reported on most often and has shown superior sensitivity when compared to other probes [3,7,8]. Radiolabelled FIAU (using 123I, 124I or 131I) has been used successfully to image HSV1-tk expression in stably transfected cells both in vitro and in animal models [7–11]. This work has been extended into the clinical realm by Jacobs et al. [12] who demonstrated the in-vivo kinetics of 124IFIAU and its uptake in human glioblastoma. The same group then showed that following intratumoural infusion of liposome–gene complex (LIPO-HSV1-tk), one of the five patients demonstrated 124I-FIAU accumulation that was significantly above the pre-vector baseline and therefore consistent with successful imaging of HSV1-tk expression in gene therapy in man [13]. Evidence of successful imaging of gene expression in oncolytic viral therapy, as opposed to gene therapy (stable transfection), has been less forthcoming. To date, only two animal and in-vitro studies have demonstrated FIAU imaging of HSV1-tk gene expression in infected rather transfected cells [13,14] with the levels of 124I-FIAU accumulation being comparatively lower after in-vivo infection [13]. Thus far, radiolabelled FIAU imaging of HSV1-tk expression during viral therapy in man has not been investigated. Our study was undertaken with the aim of using HSV-tk as a reporter gene and 123I-FIAU as a marker substrate to provide evidence of HSV1716 replication for oncolytic brain tumour therapy in humans. Figure 1 illustrates trapping of radiolabelled FIAU in an HSV1716 interfaced cell. Fig. 1

Diagram for accumulation of 5-[123I]iodo-1-(2-deoxy-2-fluoro-b-Darabinofuranosyl) uracil (123I-FIAU) in a cell infected with HSV1716. Following HSV1716 infection of a cell, viral replication begins and the HSV1716-tk gene is expressed. This reporter gene is transcribed to HSV-tk mRNA and then translated to a protein, HSV1716-TK. Following transport of 123I-FIAU probe into the infected cell, the probe is phosphorylated and trapped.

Patients and methods Patients

Patients eligible for this study were those recruited to the HSV1716 trial for treatment of malignant glioma (either newly diagnosed or recurrent glioblastoma) being undertaken at this centre [15]. Each patient received an intratumoural dose of 106 pfu of HSV1716 in 1 ml by stereotactic injection in a number of small aliquots along one trajectory. The tumour was localized using a ‘Lecksell’ stereotactic frame and image guided surgery. An initial biopsy was taken to confirm positioning of the needle within active tumour. This mode of delivery was as reported previously [15,16]. An immediate post-virus administration computed tomography scan was performed to identify the injection site and exclude any complications. 123

I-FIAU imaging was undertaken in eight of these patients prior to and following intra-tumoural injection of HSV1716. The study protocol was approved by the local ethics committee and the Administration of Radioactive Substances Advisory Committee (UK), and all experiments undertaken for this study comply with current UK laws.

Radiosynthesis

Synthesis of the necessary alkyltin precursor (5-trimethylstannyl-1-(2-deoxy-2-fluoro-b-D-arabinofuranosyl) uracil (FTAU)) and the cold iodinated standard (FIAU) was contracted to ABX Advanced Biochemical Compounds, Radeberg, Germany. The radiochemical synthesis of 123I-FIAU from the FTAU precursor by electrophilic iododestannylation was achieved using peracetic acid as the oxidant, via a slight modification of the method described by Vaidyanathan et al. [17]. Utilizing small-scale reactions designed to mimic a full-scale clinical preparation, a radiochemical yield of 92.50% ± 0.96 (n = 4) was achieved. On work-up to a full scale clinical synthesis this translated to an isolated radiochemical yield of 60.25% ± 3.64% (n = 4). These yields are in line with most other radioiodinated ligands in production. The reaction was purified by preparative high-performance liquid chromatography and the radioactive eluate corresponding to 123I-FIAU collected. The eluate was then evaporated to dryness and reconstituted in 0.9% saline prior to sterilization by terminal filtration. The final product had a radiochemical purity of 99.16% ± 0.43% (n = 4) with a specific activity of 973.1 ± 466.2 GBq
mmol – 1 (26.3 ± 12.6 Ci mmol – 1). Investigations into the stability of the prepared 123IFIAU were carried out and only minimal degradation was observed over 48 h (to date these studies have only been carried out at ambient temperature). Imaging

Single photon emission computed tomography (SPECT) imaging of the brain was performed using a dedicated

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Imaging of herpes simplex gene expression Dempsey et al. 613

Neurofocus 900 SPECT scanner, with a spatial resolution of 6 mm full width at half maximum. 123

I-FIAU injection and imaging was undertaken prior to (baseline) and following HSV1716 therapy. For each patient (n = 8), baseline 123I-FIAU images were acquired 3 days prior to viral administration. Following administration of virus, 123I-FIAU images were acquired at 1 day (n = 3), 2 day (n = 2), 3 day (n = 2) and 5 days (n = 1). 123 I-FIAU imaging involved brain SPECT acquisition at 1.5 h, 4 h and 18–24 h post-injection of 123I-FIAU (60-min image acquisition with timing of images matched pre- and post-HSV1716 on an individual patient basis). 123 I-FIAU (150 MBq) was administered intravenously for the first five patients and this was increased to 300 MBq for the last three patients. Pre- and post-viral administration 123I-FIAU images were compared qualitatively and quantitatively using region of interest (ROI) analysis. ROI analysis was used to measure 123 I-FIAU concentrations in tumour and background brain at the various time points to allow comparison of 123 I-FIAU uptake pre-viral administration to that postviral administration. Background brain concentration was measured by placing ROIs on the normal contralateral hemisphere. This background concentration was then used to set a threshold to objectively delineate tumour. Whole-body images were also acquired for three patients using a Philips Axis dual-headed gamma camera at 15 min, 5 h and 25 h post-injection of 123I-FIAU. Blood analysis

Where clinically possible a blood sample was obtained at the same time as imaging. Blood samples were centrifuged using an ALC International PK110 Centrifuge at 3000 rpm for 10 min to separate red blood cells from plasma. Plasma activity measurements were obtained using a Packard Cobra gamma counter and a 123I standard. Due to low levels of activity in plasma, accurate determination of tracer metabolism was difficult. Therefore, the %ID plasma values presented are total plasma activity (non-metabolite corrected).

Results Whole-body accumulation of

123

I-FIAU

Whole-body images (0.25, 5 and 25 h p.i. 123I-FIAU) obtained in three subjects had similar kinetics and demonstrated no evidence of specific trapping in any organ of 123I-FIAU by mammalian-tk (Fig. 2). Analysis of blood samples obtained post-injection of I-FIAU showed rapid clearance from the circulation with less than 0.005%ID
ml – 1 in the blood by 1.5 h postinjection.

123

123

I-FIAU in pre-virus brain tumours

Baseline brain SPECT imaging (i.e., pre-viral therapy) showed increased 123I-FIAU uptake in areas corresponding to disrupted blood–brain–barrier/tumour (in line with areas of Gd-diethylenetriaminepentaacetic acid (Gd-DTPA) contrast enhancement on magnetic resonance imaging) compared with background brain activity (top row, Fig. 3). Whilst tracer delivery to the tumour bed was demonstrated, measurements revealed accumulation to be low (in the order of 0.001% injected dose per millilitre). The 123I-FIAU tumour to background brain concentration ratio was of the order 3:1. Figure 4 shows the average baseline brain tumour concentration following injection of 123I-FIAU (corrected for radioactive decay). 123I-FIAU demonstrated slow clearance from the tumour bed. Radioactive decay (the half-life of 123I is 13.2 h) and the low initial delivery of isotope into brain tumour resulted in poor counting statistics in the delayed images. 123

I-FIAU uptake in HSV1716 infected brain tumours

To detect HSV-tk expression during viral replication, 123 I-FIAU imaging was undertaken before and after administration of 106 pfu HSV1716 for each patient (Fig. 3). As averaging the results may prevent detection of small differences, the pre- versus post-HSV1716 FIAU concentrations (corrected for radioactive decay) are shown (Fig. 5) for the eight patients (a–h) at 1.5 and 18–24 h post-injection of FIAU. Patients a–c were imaged at 1 day post-infection, d and e at 2 days, f and g at 3 days and patient h at 5 days. Evidence of increased uptake of FIAU due to viral replication was not detected on the 1.5-h images. The 18-h to 24-h images were used to investigate whether there was increased FIAU retention due to viral replication but non-parametric Mann– Whitney analysis showed no statistically significant difference. 123

I-FIAU concentration in plasma (parent compound plus metabolites) at 1.5 h ranged from 0.0002 to 0.0041% ID
ml – 1 (mean measurement = 0.0014 ± 0.0010%ID
ml – 1) and in tumour ranged from 0.0006 to 0.0015%ID/ml (mean measurement = 0.0009 ± 0.0003%ID
ml – 1).

Discussion In this first clinical study of non-invasive imaging of HSV-tk expression using the SPECT tracer 123I-FIAU in oncolytic virus therapy of brain tumours, we demonstrate that this technique is feasible in the clinical setting. Whilst not yet providing evidence of viral replication in humans, this study should inform researchers on the limitations of the current approach and therefore accelerate improvements of the method to achieve the goal of monitoring non-invasively oncolytic viral therapy in humans.

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614 Nuclear Medicine Communications 2006, Vol 27 No 8

Fig. 2

Representative example of anterior whole body gamma camera image of (Abbreviations as in the legend to Fig. 1).

Tumour uptake of

123

I-FIAU

One of the main findings of this study was that the initial delivery of 123I-FIAU into the brain tumour was low and this hampered accurate measurement of 123I-FIAU accumulation in the late images (18–24 h after injection). One previous study investigated radiolabelled FIAU accumulation in a single glioblastoma patient [12] and

123

I-FIAU distribution at (a) 0.25 h (b) 5 h and (c) 25 h post-injection.

imaging with PET at 10 min after injection of 124I-FIAU, the measurements revealed a similar level of 124I-FIAU derived activity of 0.001%ID/g. The effect of low delivery is highlighted in this SPECT study because of the shorter half-life of 123I compared to 124I (13.2 h vs. 4.2 days), and at 24 h our count rate was less than optimal for imaging. As well as 123I having a shorter half-life than 124I, the

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Imaging of herpes simplex gene expression Dempsey et al. 615

Fig. 3

^{123I-FIAU brain single photon emission computed tomography images with corresponding T1-weighted magnetic resonance imaging showing contrast enhancing tumour. Top row images were acquired prior to HSV1716 therapy (baseline) and show 123I-FIAU accumulation within the tumour bed at 1.5 h with substantial FIAU uptake at later time points (4 and 24 h) after injection showing washout to be slow. Bottom row images were acquired at similar time points after HSV1716 therapy for comparison with baseline images but demonstrated no discernible difference in 123I-FIAU uptake or retention.

%ID/ml

Fig. 4

0.0020 0.0018 0.0016 0.0014 0.0012 0.001 0.0008 0.0006 0.0004 0.0002 0

Average FIAU uptake pre-HSV1716

initial delivery to the tumour bed is therefore dependent on the degree of disruption of the barrier in individual gliomas. Strategies of circumventing the barrier either in the design of the reporter probe [19,20], in the use of a carrier vehicle or by osmotic opening all merit attention for future trials [21,22]. Expression of HSV-tk

Pre-1.5h

Pre-4h Time post-injection

Pre-18 − 24h

Time course of 123I-FIAU in human brain tumour (prior to HSV1716 therapy) after systemic administration of 123I-FIAU. Values are corrected for radioactive decay and averaged over eight patients.

reduced sensitivity of SPECT compared to PET may also be an issue [4]. Whilst the SPECT imaging system used for this study has a lower sensitivity than PET, it has a significantly greater sensitivity than a gamma camera and even at these low levels of activity we have the ability to accurately detect a 10% difference between pre- and post-virus 123I-FIAU uptake on the early images. A greater difference in retention (> 25%) would be required to detect a change on the late images. FIAU and other nucleoside analogues do not appear to penetrate the intact blood–brain barrier [12,18] and

It is possible that increased levels of HSV-tk expression are required to enable detection of viral replication in human glioma with radiolabelled FIAU. Previous work has demonstrated that levels of HSV-tk expression in virally infected cells is significantly less than the levels of expression achieved with stably transfected cells in animal models [13]. The incorporation of alternative promoters into HSV1716 may be required to increase HSV-tk expression and FIAU accumulation [14]. In addition to this, a reporter probe with increased HSV-tk sensitivity would aid the detection of viral replication. Recent research into reporter probe chemistry shows radiolabelled FEAU to have increased potential in this area [20,23]. Extent of viral replication

Delivery of HSV1716 in this study relied on administration of 106 pfu in 1 ml. A previous study involved administration of HSV1716 4–9 days prior to tumour resection in 12 glioblastoma patients. Following resection, the tumour specimens were examined for evidence of HSV1716 replication and it was shown that HSV1716

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616 Nuclear Medicine Communications 2006, Vol 27 No 8

Fig. 5

Brain tumour FIAU uptake at 1.5h

(a) 0.0020 0.0018 0.0016 0.0014 0.0012 0.0010 0.0008 0.0006 0.0004 0.0002 0 %ID/ml

pre HSV1716 post HSV1716

sion to the levels required for trapping of sufficient 123IFIAU for imaging. With only one patient imaged at this time point and with such low level FIAU accumulation, more convincing data is required before such a positive result should be claimed. Cell lysis

a (b) 0.0020 0.0018 0.0016 0.0014 0.0012 0.0010 0.0008 0.0006 0.0004 0.0002 0

b

c

d

e

f

g

h

Brain tumour FIAU uptake at 18− 24h

%ID/ml

pre HSV1716 post HSV1716

a

b

c

d

e

f

g

h

Accumulation of 123I-FIAU activity in brain tumour at (a) 1.5 h and (b) 18–24 h after injection for eight patients imaged before and after HSV1716 therapy. Post-HSV1716 1.5-h images were not acquired for patients c and g due to ill health. Patients a–c were imaged at 1 day post-infection, d and e at 2 days, f and g at 3 days and patient h at 5 days.

selectively replicated in tumour tissue [15]. Nevertheless, in this study, there was possibly insufficient viral replication to enable accumulation of 123I-FIAU to the level required for detection by SPECT imaging. FIAU accumulation is known to be dose-dependent [14] with evidence of a possible threshold effect and it may be argued that this initial viral load is too low, but as the intrinsic replication capacity of HSV1716 means that the input dose is likely to increase 200-fold for each replication cycle, the input dose should not be critical. In addition to the input dose, the method of viral administration used in this study may not have been ideal as efficacy depends on infectious virus coming in contact with as many cells in active division as possible. In future studies 107 pfu of HSV1716 will be administered by convection enhanced delivery.

The effect of oncolysis in viral therapy may play a critical role with respect to imaging replication. Replicationconditional HSV1716 is cytotoxic and should cause lysis in replicating host cells. This phenomenon may have two major consequences affecting imaging studies. Firstly, the effect on the 123I-FIAU molecule itself, following trapping and then lysis of the host cell, is not known and this may confound the delayed images. Secondly, such lysis of host cells will result in a reduction of the total number of actively dividing cells, imposing a limit on the extent of HSV1716 infection, replication and ultimately HSV-tk expression. Jacobs et al. found the timing window of HSV-tk expression to be 8 h to 8 days post-viral administration in animal studies with oncolytic herpes simplex virus type 1 [13]. The use of 123I-FIAU presented here is novel and imaging of this type has not been attempted previously with viral therapy in patients with malignant glioma. This work did not provide direct evidence of viral replication in the eight patients studied but the data produced informs other groups of the potential limitations of this technique for future clinical use.

Acknowledgement Crusade Laboratories Ltd provided the virus.

References 1

2

3

4

5

Number of replication cycles

We have imaged patients at several time points post-viral administration (1 day (n = 3), 2 day (n = 2), 3 day (n = 2) or 5 days (n = 1)). Whilst no significant timing effect was apparent, the FIAU uptake at 20 h of the patient imaged at 5 days post-infection (patient h) could perhaps be interpreted as positive for HSV-tk. Such a result would concord with the theory that 5 days allows for several viral replication cycles resulting in increased HSV-tk expres-

6

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8

Brown SM, Harland J, MacLean AR, Podlech J, Clements JB. Cell type and cell state determine differential in vitro growth of non-neurovirulent ICP34.5-negative herpes simplex virus types 1 and 2. J Gen Virol 1994; 75:2367–2377. McKie EA, MacLean AR, Lewis AD, Cruickshank G, Rampling R, Barnett SC, et al. Selective in vitro replication of herpes simplex virus type 1 (HSV-1) ICP34.5 null mutants in primary human CNS tumours – evaluation of a potentially effective clinical therapy. Br J Cancer 1996; 74:745–752. Tjuvajev JG, Stockhammer G, Desai R, Uehara H, Watanabe K, Gansbacher B, et al. Imaging the expression of transfected genes in vivo. Cancer Res 1995; 55:6126–6132. Tjuvajev JG, Finn R, Watanabe K, Joshi R, Oku T, Kennedy J, et al. Noninvasive imaging of herpes virus thymidine kinase gene transfer and expression: a potential method for monitoring clinical gene therapy. Cancer Res 1996; 56:4087–4095. Tjuvajev JG, Avril N, Oku T, Sasajima T, Miyagawa T, Joshi R, et al. Imaging herpes virus thymidine kinase gene transfer and expression by positron emission tomography. Cancer Res 1998; 58:4333–4341. Penuelas I, Mazzolini G, Boan JF, Sangro B, Marti-Clement J, Ruiz M, et al. Positron emission tomography imaging of adenoviral-mediated transgene expression in liver cancer patients. Gastroenterology 2005; 128:1787–1795. Tjuvajev JG, Doubrovin M, Akhurst T, Cai S, Balatoni J, Alauddin MM, et al. Comparison of radiolabeled nucleoside probes (FIAU, FHBG, and FHPG) for PET imaging of HSV1-tk gene expression. J Nucl Med 2002; 43:1072–1083. Brust P, Haubner R, Friedrich A, Scheunemann M, Anton M, Koufaki ON, et al. Comparison of [18F]FHPG and [124/125I]FIAU for imaging herpes simplex virus type 1 thymidine kinase gene expression. Eur J Nucl Med 2001; 28:721–729.

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Deng WP, Yang WK, Lai WF, Liu RS, Hwang JJ, Yang DM, et al. Non-invasive in vivo imaging with radiolabelled FIAU for monitoring cancer gene therapy using herpes simplex virus type 1 thymidine kinase and ganciclovir. Eur J Nucl Med Mol Imaging 2004; 31:99–109. Haubner R, Avril N, Hantzopoulos PA, Gansbacher B, Schwaiger M. In vivo imaging of herpes simplex virus type 1 thymidine kinase gene expression: early kinetics of radiolabelled FIAU. Eur J Nucl Med 2000; 27:283–291. Blasberg RG, Tjuvajev JG. Herpes simplex virus thymidine kinase as a marker/reporter gene for PET imaging of gene therapy. Q J Nucl Med 1999; 43:163–169. Jacobs A, Braunlich I, Graf R, Lercher M, Sakaki T, Voges J, et al. Quantitative kinetics of [124I]FIAU in cat and man. J Nucl Med 2001; 42:467–475. Jacobs A, Tjuvajev JG, Dubrovin M, Akhurst T, Balatoni J, Beattie B, et al. Positron emission tomography-based imaging of transgene expression mediated by replication-conditional, oncolytic herpes simplex virus type 1 mutant vectors in vivo. Cancer Res 2001; 61:2983–2995. Bennett JJ, Tjuvajev J, Johnson P, Doubrovin M, Akhurst T, Malholtra S, et al. Positron emission tomography imaging for herpes virus infection: Implications for oncolytic viral treatments of cancer. Nat Med 2001; 7:859–863. Papanastassiou V, Rampling R, Fraser M, Petty R, Hadley D, Nicoll J, et al. The potential for efficacy of the modified (ICP 34.5( – )) herpes simplex virus HSV1716 following intratumoural injection into human malignant glioma: a proof of principle study. Gene Ther 2002; 9:398–406. Rampling R, Cruickshank G, Papanastassiou V, Nicoll J, Hadley D, Brennan D, et al. Toxicity evaluation of replication-competent herpes simplex

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Original article

Synthesis of trans-1,2-cyclohexyldinitrilo tetramethylene phosphonic acid and its radiolabelling with 99mTc for the detection of skeletal metastases Puja Panwara,b, Krishna Chuttania, Pushpa Mishraa, Rajnish Sharmaa,c, Anupam Mondala and Anil Kumar Mishraa Background and aim The development of bone-seeking radiopharmaceuticals for the detection of malignant bone lesions could further improve the diagnostic accuracy of routine bone scanning. This study aimed to provide a convenient synthesis of trans-1,2-cyclohexylenedinitrilo tetramethylene phosphonic acid (CDTMP) and an improved preparation of its 99mTc complex. Methods CDTMP was prepared from trans-1,2-cyclohexyldinitrilotetraacetic acid by reaction with phosphorus trichloride and it was labelled with 99mTc. Toxicity and biodistribution studies were carried out in BALB/c mice, while blood clearance and bone scintigraphy studies were carried out in rabbits. 99mTc-CDTMP was evaluated for the detection of malignant bone lesions in 11 patients. Bone scintigraphy (a methylene diphosphonate scan) was performed to detect metastases at diagnosis and follow-up.

of 99mTc-CDTMP in bone was 7.69 ± 0.65%ID/g at 1 h. The mean ratio of bone lesion to soft tissue was 6.8 ± 0.69 and of bone lesion to normal bone was 5.67 ± 0.82. Visual image analysis of 99mTc-CDTMP was clinically comparable to the interpretation of imaging studies with 99mTc-MDP. Conclusion These preliminary data support increased bone uptake by the tetraphosphonate complex of 99mTc. This suggests that CDTMP complexed with therapeutic radionuclides should be evaluated for therapy of skeletal c 2006 metastases. Nucl Med Commun 27:619–626 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:619–626 Keywords: Bone imaging, CDTMP,

99m

TC

a Institute of Nuclear Medicine and Allied Sciences, Brig. SK Mazumdar Road, Delhi 110054. India, bDivisions of Cyclotron and Radiopharmaceutical Sciences and cDepartment of Nuclear Medicine.

Results The radiolabelling efficiency was found to be > 97% and the stability in serum indicated that 99mTc remained bound to the chelate, CDTMP, for up to 24 h. Blood clearance showed a quick wash-out from the circulation and the biological half-lives (t12) were 55 min (F) and 8 h 48 min (S). The LD50 was 110 mg
kg – 1 as determined by toxicity studies. The drug was excreted mainly through renal route and the accumulation

Correspondence to Dr Anil K. Mishra, Institute of Nuclear Medicine and Allied Sciences, Division of Cyclotron and Radiopharmaceutical Sciences, Brig. SK Mazumdar Road, Timarpur, Delhi 110054, India. Tel: + 0091 23914374; fax: + 0091 23919509; e-mail: [email protected]

Introduction

99m

Polyphosphates are known to localize in the metastatic lesions in the bone. A variety of 99mTc-labelled phosphate compounds have been evaluated for possible use in scintigraphy and therapy [1,2]. The tracers used routinely for clinical bone scanning are methylene diphosphonate (MDP), dicarboxypropane diphosphonate (DPD), hydroxythylidine diphosphonate (HEDP) and ethylenediaminetetramethylene phosphonate (EDTMP) [3–9]. Phosphonates are characterized by the presence of one or more —CH2—PO3—H groups. Phosphonate compounds containing more than one phosphonate group are effective sequestrants and possess other useful properties such as high water solubility and chemical stability. Larger ligands with greater number of phosphonate groups per molecule should produce

Received 17 August 2005 Revised 19 December 2005 Accepted 14 March 2006

Tc chelates with fewer ligands per complex than the diphosphonates. Multidentate phosphonate ligands exhibit good metal ion control properties with highly selective bone uptake in vivo. Most phosphonates are synthesized from phosphorous acid by reaction with formaldehyde and either ammonia or amines [10]. It is thus an object of the present work to provide a modified synthetic route of preparation of more stable agent for radiotherapy and diagnosis based on phosphonic acid derivatives. The compound trans-1, 2-cyclohexyldinitrilo tetramethylenephosphonic acid (CDTMP) was synthesized by a reaction with phosphorous acid in a one-step process [11]. CDTMP is a strong (multidentate) complexing agent for metal ions and is sufficiently hydrophilic to be soluble in water.

c 2006 Lippincott Williams & Wilkins 0143-3636

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620 Nuclear Medicine Communications 2006, Vol 27 No 8

Extra-osseous accumulation of bone imaging agents has been reported in the literature. Factors such as bone blood flow, enzyme activity (especially alkaline phosphatase), and active bone mineralization of hydroxyapatite crystals play a part in the accumulation of radionuclides [12–16]. Normally, many bone lesions have higher concentrations of amorphous calcium phosphate, which has a greater affinity for bone-imaging agents. This is due to the greater surface area of the matrix that facilitates adsorption of the bone agent [15]. Specifically, the OH radical on the phosphonate group is responsible for this matrix affinity. Another suggested mechanism is disruption of the cell membrane with deposition of the bone agent on calcium already fixed to mitochondria [16]. A major problem encountered in oncology is that patients may have intractable pain secondary to bone metastases. This is most often seen in cancer of the prostate, breast and lung. When the pain has become unresponsive to all available analgesics, a multimodal approach involving radiation, hormonal, and even surgical therapy becomes necessary. The presence of multiple, scattered, bone lesions is ideal for radionuclide therapy because the lesions can be targeted selectively through a single systematic administration of the radiopharmaceutical. The available list of radionuclides has been growing, and now includes 89Sr, 32P, 131I, 90Y, 186/188Re and 153Sm [17–20]. The utility of radioisotopes in the treatment of advanced metastatic cancer is that it results in a significantly larger proportion of patients who are free of new pain sites and a lower incidence of adverse effects, particularly gastrointestinal. We report here the work carried out in the synthesis of CDTMP and the radiochemical studies with 99mTc and biodistribution studies of the complex.

Material and methods Chemicals

All the chemicals – trans-1,2-cyclohexyldinitrilotetraacetic acid, monohydrate salt (CDTA), phosphorous acid, phosphorous trichloride, stannous chloride, ethanol and methanol (HPLC grade) – were purchased from SigmaAldrich Co. 99mTc was provided by the Regional Center for Radiopharmaceuticals (Northern Region), Board of Radiation and Isotope Technology (BRIT), Department of Atomic Energy, India. Instrumentation

The 1H NMR and 13C NMR spectra were recorded by a Bruker 250 MHz system for 1H and 62 MHz for 13C NMR. Mass spectra were obtained in-house from an ion trap SL 1100 system (Agilent, Waldbronn, Germany) using the ESI positive and negative mode.

Animal models

New Zealand rabbits, each weighing 2–3 kg, and BALB/c mice, each weighing 25–30 g, were used for toxicity, blood clearance, imaging and biodistribution studies. Animal protocols were approved by institutional animal ethics committee. Clinical studies

Eleven patients (age, 71 ± 14 years; nine men with prostate cancer and two women with breast cancer) who were suspected of having skeletal metastasis, as determined by a 99mTc-MDP bone scan took part in the study. Each patient received a dose of 555 MBq of 99m Tc-CDTMP. Synthesis of trans-1,2-cyclohexyldinitrilo tetramethylene phosphonic acid

Phosphorous acid (0.44 g, 40.0 mmol) was added to a solution of trans-CDTA (0.5 g, 9.49 mmol) dissolved in 10 ml toluene. The reaction mixture was refluxed while phosphorus trichloride in a volume of 342 ml (0.55 g, 0.44 mmol) was added dropwise to the refluxing mixture. After 3 h, the toluene was removed following addition of deionized water. The filtrate was concentrated under vacuum. The concentrated product was precipitated on addition of methanol/ethanol. Analytical data 1

H NMR (250 MHz) D2O: d 3.59–2.88 (8H, complex multiplet, —CH2—PO3H2—), 2.1 (2H, m), 1.74 (2H, m), 1.71–1.21 (4H, q). 13C NMR (62 MHz): d 50.7, 48.0, 45.7, 24.1, 22.8 ESI-MS + : m/e calcd. 490, found [M + H + ] 490.61. Radiochemical synthesis of

99m

Tc-CDTMP

Stannous chloride (1.0 mmol) was added to 10.0 mmol of CDTMP. The pH of the reaction mixture was adjusted to 6.0–6.5 with 0.5 M NaHCO3. The mixture was passed through a 0.22 mm millipore filter into a sterile vial. The 99m Tc eluate containing 370–800 MBq activity was added and the complex was incubated for 30 min at room temperature to obtain optimum labelling yield. The labelling efficiency was estimated chromatographically using ITLC-SG (instant thin layer chromatography–silica gel) paper as the stationary phase and 100% acetone as the mobile phase. The percentage of colloids was determined using pyridine:acetic acid:water (3:5:1.5) as the mobile phase and an ITLC-SG strip as the stationary phase. The in vitro stability of the complex was ascertained chromatographically by estimating the labelling efficiency at different time intervals up to 24 h. The complexation was determined by titration of the chelating groups with 111In. A stock solution of carrier-free 111 InCl3 (20 ml, in sodium acetate buffer, pH 5.5 adjusted with 1 M sodium acetate) was added to a stock solution (20 ml of 50 mM, 0.1 < mol). Then 16 ml of 0.1 M sodium acetate buffer, pH 7, was added. The pH was checked to

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99m

Tc-CDTMP for the detection of skeletal metastases Panwar et al. 621

ensure that is was 7. After 3 h, the mixture was analysed by using a TLC system (MeOH/10% NH4OAc, 1:1).

in each organ was measured. The data were expressed as the percent administered dose per gram of the organ.

Animal studies Toxicity studies

Clinical studies

Toxicity studies were carried out in male BALB/c mice. In the experiment, a single dose of CDTMP was administered intravenously to groups of five mice. Each mouse weighed 28 ± 3 g. The dosage levels were 30–185 mg
kg – 1 body weight. The animals are weighed weekly and observed for clinical signs of toxicity and the number of deaths that occurred in 4 weeks was recorded after treatment. Blood clearance

For studies of blood clearance the labelled complex (10 MBq) was administered intravenously through the dorsal ear veins of normal rabbits weighing about 2.5–3.2 kg. The remaining activity in blood, in terms of percentage administered dose, was determined by using a gamma camera at various time intervals between 5 min and 24 h post-injection. Bone scintigraphy

Imaging of rabbit was carried out at different time intervals after administering 10 MBq of the labelled compound intravenously to normal rabbits. A comparison study was also conducted with 99mTc-MDP. Biodistribution studies of mice

99m

Whole-body images were obtained using a dual-head gamma camera. For all patients, a whole-body 99mTcMDP scan was done first. After 1 week, 99mTc-CDTMP imaging was performed after i.v. administration of 555 MBq (15 mCi)) radiotracer. A whole-body scan was carried out 1 h after injection of the radiotracer as a standard protocol and five patients were also imaged at 5, 10, 30, 45, 60 and 180 min. Semiquantitative analysis was generated from regions of interest (ROIs) placed over areas counting average counts per pixel with maximum radiotracer uptake on the skeletal metastases and compared to symmetric counterparts in normal bone, and with the ROIs in soft tissues. The ROI of soft tissue was placed on the medial aspect of the thigh region. The data obtained by ROI analysis were expressed as ratios of bone lesions to normal bone (BL/NB) and bone lesion to soft tissue (BL/ST) Statistical analysis

Data are reported as mean ± standard deviation (SD). A mean of 10 lesions per patient were obtained. The overall mean ± SD for each of the 11 patients was calculated.

Results

Tc-CDTMP in BALB/c

Synthesis

Biodistribution was studied in male BALB/c mice weighing 25 ± 2 g. 99mTc-CDTMP (3.7 MBq) was injected intravenously through the tail vein of each mouse. The animals were killed at 1, 4 and 24 h post-injection and dissected. The various organs were removed, made free from adhering tissue and weighed. The radioactivity

The starting point for the synthesis of the chelating agent CDTMP was the trans isomer, which provides a semi-rigid structure, (1S*,2S*,4R*)-1,2-dinitrilotetramethlyene phosphonic acid with six coordination centres for complexation. The synthesis of the tetraphosphonate chelator CDTMP is shown in Fig. 1. The final product was obtained in 92%

Fig. 1

O COOH

P

OH

OH N

N COOH

OH

i) H3PO3 ii) PCl3

P

> 80°C

O

N

OH

N O P

COOH HOOC

P O

trans-1,2-cyclohexyldinitrilotetraacetic acid

OH

OH OH

OH

CDTMP

Synthesis of trans-1,2-cyclohexyldinitrilo tetramethylene phosphonic acid (CDTMP).

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622 Nuclear Medicine Communications 2006, Vol 27 No 8

yield and the [M + H] + was found to be 490.61 in positive mode. In 1H NMR the —CH2—P chemical shift appeared at 3.59–2.88 ppm. 13C NMR suggests cyclohexyl carbons (3C) and —CH2—P—carbons (2C). Quality control

The radiochemical purity of 99mTc-CDTMP was estimated chromatographically using ITLC-SG paper as the stationary phase and 100% acetone as the mobile phase. The complex remained at the point of spotting. Under identical condition 99mTcO4– moved towards the solvent front. Thus the yield of free and complex ligand could be estimated. The labelling yield was found to be more than 97%. A metal binding assay confirmed the radiochemical purity of CDTMP to be > 97%. Phosphor imaging showed only a single spot with an RF = 0.3 (10% ammonium acetate/methanol) which suggests the formation of only one species. Stability studies

Prior to biodistribution studies of the radiopharmaceutical different factors were considered, particularly chelate stability in serum, this depends on various parameters such as medium pH, the metabolism of the metal and the presence of binding proteins. The stability study for 99mTc-CDTMP showed no trans-chelation phenomena for the metal toward the physiological components of the blood because there was no release of radioactivity to complex in a more or less stable manner with albumin and transfering. This study showed that 99m Tc-CDTMP is highly stable in serum medium. The complex thus prepared was highly stable in vitro as only 3% detachment of the radioisotope was observed at 24 h (Fig. 2).

reaction was observed. Doses > 110 mg
kg – 1 were found to be lethal. A dose of 30-65 mg
kg – 1 was well tolerated and no apparent adverse reaction was seen. At a dose of 80–108 mg
kg – 1 an immediate adverse reaction was observed but after tremors and an increased heart rate for 2–3 min the reaction ceased. No adverse signs were observed in these groups after 30 days. At a dose of 110 mg
kg – 1 50% of the mice died within 30 days. The lethal dose was found to be 126 mg
kg – 1. Myelotoxicity studies were also performed and no significant decrease in blood counts of all the animals that received the radioactivity up to 2 weeks was observed. Histopathological examinations showed no evidence of cell damage of vital organs, which showed that the drug is quite safe. Blood clearance

Blood clearance studies in rabbits showed that the complex was washed out very quickly from the circulation and the blood activity curves show that the clearance of 99m Tc-CDTMP is comparable to that of 99mTc-MDP. There was less activity present in the blood at 3 h postinjection. This accounts for high bone/blood and bone/ muscle ratios exhibited by 99mTc-CDTMP. These findings in rabbits are an indication of the excellent in-vivo stability of the 99mTc-CDTMP. The biological half-lives were found to be 55 min (F) and 8 h 48 min (S) (Fig. 3). Imaging and biodistribution studies

Fig. 2

Fig. 3

Animal studies Toxicity studies

% Intact chelate

100 90 80 70 60 50 40 30 20 10 0 0

5

10

15

20

99m

30 25

t1/2 (F) = 55 min and t1/2 (S) = 8 h 48 min

20 15 10 5 0 0

240

480

720

960

1200

1440

Time (min)

Time (h) In-vitro human serum stability studies of physiological conditions.

25

% Radioactivity remaining in blood

The acute i.v. LD50 was determined to be 110 mg
kg – 1. Doses < 110 mg
kg – 1 were well tolerated and no adverse

Imaging of animals was carried out at different time intervals after the labelled compound had been administered intravenously. Within 15 min the labelled phosphonate ligand accumulated in the shoulder region. After 1 h the ligand was localized in the whole skeleton of rabbit. A comparison study conducted for CDTMP and MDP in normal rabbits of the same age and weight demonstrated comparable quality at 3 h with 99mTc-MDP and at 1 h with 99mTc-CDTMP post-injection. The scintigrams of 99mTc-CDTMP are of comparable quality to a

Tc-CDTMP under

Blood clearance in normal rabbits following intravenous injection of 99m Tc-CDTMP, administered through the ear vein.

Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.

99m

Tc-MDP scintigram in a 3.1-kg rabbit and the images also show that non-osseous tissue uptake of 99m Tc-CDTMP is minimal and when assessed with its rapid blood clearance accounts for the low soft-tissue activity (Fig. 4). In the early images of the 99mTcCDTMP scan, major accumulation was visualized in the kidneys. This is accounted for by the presence of an aminophosphonate group in the compound, which has higher affinity for the kidney cells, and the time taken for it to clear from the kidneys. Biodistribution studies in mice showed approximately 7.69 ± 0.65% D/g skeletal uptake at 1 h post-injection in CDTMP (Table 1). Localization of 99mTc-CDTMP in non-osseous tissues was low. The bone-to-muscle uptake ratios of 99mTcCDTMP were not significantly different from those of 99mTc-MDP (Fig. 5) but the bone uptake to blood ratio and bone/muscle ratios of 99mTc-CDTMP were 40.4

Tc-CDTMP for the detection of skeletal metastases Panwar et al. 623

Fig. 5

9 8 7 % Injected dose/g

99m

6 5 4

99mTc-MDP

3

99mTc-CDTMP Soft tissue

2 1 0 0

5

10 15 Time (h)

Comparison of %ID/g of 99mTc-CDTMP and in bone and soft tissue in BALB/c mice.

99m

20

Tc-MDP accumulated

Fig. 4 99mTc-CDTMP

99mTc-MDP

(a)

(b)

and 54.9, respectively, which were higher than those observed with 99mTc-MDP. Clinical studies

Whole-body gamma-scintigraphic images of (a) and (b) 99mTc-MDP in rabbit (1 h).

99m

Tc-CDTMP (1 h);

99m Table 1 Biodistribution of Tc-trans-1,2-cyclohexyldinitrilotetramethylene phosphonic acid (99mTc-CDTMP) in BALB/c mice following intravenous injection

Organ

Blood Heart Lungs Liver Spleen Kidneys Stomach Intestines Brain Muscles Bone

Percentage of injected dose per gram of tissue 1h

4h

24 h

0.19 ± 0.01 0.09 ± 0.01 0.17 ± 0.02 0.20 ± 0.03 0.16 ± 0.01 1.04 ± 0.05 0.22 ± 0.01 0.11 ± 0.96 0.12 ± 0.01 0.14 ± 0.01 7.69 ± 0.65

0.15 ± 0.02 0.07 ± 0.01 0.16 ± 0.01 0.11 ± 0.01 0.12 ± 0.01 1.52 ± 0.10 0.15 ± 0.01 0.05 ± 0.01 0.07 ± 0.005 0.08 ± 0.01 6.93 ± 0.36

0.02 ± 0.003 0.04 ± 0.005 0.08 ± 0.005 0.09 ± 0.006 0.06 ± 0.040 0.28 ± 0.040 0.07 ± 0.010 0.06 ± 0.005 0.07 ± 0.003 0.06 ± 0.007 6.22 ± 0.180

Data from groups of five mice are expressed as mean %ID/g ± SD.

Whole-body scanning revealed numerous malignant lesions displaying increased uptake of 99mTc-CDTMP. A group of five patients aged 62 ± 12 years; three with prostate cancer and two women with breast cancer, were imaged at 5, 10, 30, 45, 60 and 180 min. Following i.v injection the radiotracer cleared from the blood pool within 5 min. Subsequently at 5, 10 and 45 min and later, images showed faint accumulation in the liver. The kidneys were visualized at 45 min, while the bladder became visible on the 1h image. From 10 min onwards, the amount of tracer increased in the skeleton and significant activity accumulated in the lesions. The tracer was still present in the skeletal system even at 180 min. Lesions were used to enumerate the bone lesion to soft tissue (BL/ST) and bone lesion to normal bone (BL/NB) ratios (Fig. 6). Table 2 shows the results of ROI analysis. A mean ± SD of 6.8 ± 0.69 for BL/ST and 5.67 ± 0.82 for BL/NB was obtained. The ratios have an overall range of 5.6–7.8 for BL/ST and 5.36–7.21 for BL/NB. The kidneys and bladder were also visualized on the bone scan because 99mTc-CDTMP is efficiently excreted primarily via the renal route.

Discussion Accurate interpretation of a bone scan for metastatic disease requires an image quality sufficient to demonstrate subtle abnormalities in bone uptake (increased or decreased). The sensitivity of bone scanning is outstanding with 99mTc-CDTMP, the compound prepared in

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624 Nuclear Medicine Communications 2006, Vol 27 No 8

Fig. 6

Whole-body imaging using 99mTc-CDTMP. The patient has multiple bone metastases as a result of prostate cancer. Multiple lesions on the ribs, sternum, left clavicle, right clavicle, vertebral column and both femurs can be seen.

Table 2 Semiquantative region of interest analysis showing bone lesion to soft tissue and bone lesion to normal bone ratios in 11 patients Patient 1 2 3 4 5 6 7 8 9 10 11 Mean ± SD Range

Bone lesion/soft tissue ratio

Bone lesion/normal bone ratio

7.1 ± 2.43 7.7 ± 1.86 5.6 ± 3.48 6.7 ± 1.71 7.4 ± 1.85 6.2 ± 1.28 5.9 ± 2.76 7.3 ± 0.94 6.5 ± 1.20 7.8 ± 1.10 6.6 ± 2.05 6.8 ± 0.69 5.6–7.8

6.40 ± 2.04 7.21 ± 0.68 5.09 ± 2.24 6.03 ± 1.58 6.70 ± 1.88 5.60 ± 1.93 5.36 ± 3.12 6.60 ± 2.2 5.90 ± 1.67 7.09 ± 1.83 6.02 ± 0.20 5.67 ± 0.82 5.36–7.21

Results are expressed as mean ± SD.

this study. The tetraphosphonates used in this study have unusual metal ion control properties with very high invitro stability.

Previously reported methods for the synthesis of tetraphosphonates are not only time consuming but also provide very poor yield [10]. We have modified the procedure to provide a product with high purity and greater yield (92%). The one-step process eliminates the formation of unwanted intermediate products thereby obviating the need for further purification [11]. The old methodology for synthesizing the tetraphosphonate chelator, CDTMP, involves a Mannichtype reaction using ethylamines, phosphorous acid and formaldehyde [10]. In the reported methodology phosphorus trichloride is used which reacts violently with water to give phosphorous acid. After synthesis, it was fully characterized on the basis of spectral studies and labelled with 99mTc to evaluate it for bone scintigraphy and biodistribution. The polydentate nature of the CDTMP favours the formation of complexes with only one ligand per metal atom, therefore minimizing the formation of multiple species in solution, which is shown by metal binding assays.

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99m

Tc-CDTMP for the detection of skeletal metastases Panwar et al. 625

The phosphonates are not metabolized in vivo. This is due to the stability of their —CH2—P bond to heat and most chemical reagents, as well as to their resistance to hydrolysis by the enzymes found in the body. Phosphonates are excreted unchanged via the renal route. Blood clearance studies showed the rapid clearance of the drug. We have examined the in vivo biodistribution of 99mTcCDTMP in BALB/c mice. A high bone lesion to normal bone uptake ratio would be an effective means of enhancing the contrast between calcifying lesions and normal bone. The high lesion to bone ratios recorded in Table 2 reflects the specificity of CDTMP to selectively localize the lesions in a manner analogous to that seen with 99mTc-MDP. High skeletal uptake was observed, with the peak uptake at 1 h.

complexes that exhibit high and selective in vivo bone uptake in rats and rabbits despite the presence of large organic backbones. This type of ligand forms stable complexes with metal ions with fewer structural forms because four methylene phosphonic acid groups form highly negatively charged chelates with multidenticity, which is essential to wrap up transition-metal ions with higher stability. This factor, coupled with the polydentate nature of the ligand, favours the formation of a complex with only one ligand per metal atom, therefore minimizing the formation of multiple species in solution, which is not desirable for biological activity. The rapid accumulation of the diagnostic agent in bone metastases and a high lesion-to-normal bone ratio showed that this drug has therapeutic potential when coupled to beta-emitting radionuclides. Further investigation of this property would be useful.

In all the patients studied, no symptoms of immediate side effects were seen after administration of 99mTcCDTMP and the infusion was not associated with other subjective or objective side effects, such as nausea or vomiting. This might be explained by the fact that phosphonates are characterized by a rapid and strong binding to the hydroxyapatite crystals [12–16] and its rapid incorporation into calcified tissue leads to their short presence in the circulation [12]. The amount of radiotracer (0.09 ± 0.015 mg
kg – 1) administered to the patients was 1000 times less than the LD50 (110 mg
kg – 1) compared to that of the mice. Bone metastases are demonstrated most commonly as ‘hot spots’ distributed unevenly throughout the skeleton. Pelvic region metastases were observed at 10 min, lesions in skull, shoulders, and ribs were visualized after 10 min showing its rapid accumulation in bone in the study conducted with five patients. Blood pool images showed concentration of radiotracer in the liver and heart. Significant wash-out was observed in delayed images. A standard study protocol of 1 h in all patients demonstrated uptake of 99m Tc-CDTMP by all the lesions as seen in the 3 h 99mTcMDP scintigraphic image.

References 1

2 3

4 5

6

7

8

9 99m

Since we were evaluating Tc-CDTMP for clinical studies we took the images at different time intervals to validate the uptake of the new bone-seeking radiotracer. In the case of 99mTc-MDP, at 1 h the tracer was mostly accumulated in soft tissue and there was no appreciable uptake in the lesions. We compared the images of 99mTcCDTMP with 99mTc-MDP which is routinely used for bone imaging to demonstrate that it was possible to visualize all the lesions at 1 h compared to the wellestablished diagnostic agent MDP. Most importantly, CDTMP has multiple denticity compared to MDP which will allow beta-emitting radionuclides to be labelled for therapy.

10

11

12

13

14

15

This investigation shows that multidentate tetramethylene phosphonate ligands are capable of forming stable

Hamdy NA, Papapoulos SE. The palliative management of skeletal metastases in prostate cancer: use of bone-seeking radionuclides and bisphosphonates. Semin Nucl Med 2001; 31:62–68. Body JJ. Effectiveness and cost of bisphosphonate therapy in tumor bone disease. Cancer 2003; 97(suppl):859–865. Pauwels EK, Blom J, Camps JA, Hermans J, Rijke AM. A comparison between the diagnostic efficacy of 99mTc-MDP, 99mTc-DPD and 99mTc-HDP for the detection of bone metastases. Eur J Nucl Med 1983; 8: 118–122. Vorne M, Vahatalo S, Lantto T. A clinical comparison of 99mTc-DPD and two 99m Tc-MDP agents. Eur J Nucl Med 1983; 8:395–397. Palmedo H, Manka-Waluch A, Albers P, Schmidt-Wolf IG, Reinhardt M, Ezziddin S, et al. Repeated bone targeted therapy for hormone refractory prostate carcinoma: randomized phase II trials with the new high-energy radiopharmaceutical rhenium-188 hydroxythylidinediphosphonate. J Clin Oncol 2003; 21:2869–2875. Franzius C, Schuck A, Bielack SS. High-dose samarium-153 ethylenediaminetetramethylene phosphonate: low toxicity of skeletal irradiation in patients with osteosarcoma and bone metastases. J Clin Oncol 2002; 20:1953–1954. Van Rensburg AJ, Alberts AS, Louw WK. Quantifying the radiation dosage to individual skeletal lesions treated with samarium-153 EDTMP. J Nucl Med 1998; 39:2110–2115. Bayouth JE, Macey DJ, Kasi LP, Fossella FV. Dosimetry and toxicity of samarium-153-EDTMP administered for bone pain due to skeletal metastases. J Nucl Med 1994; 35:63–69. Ahonen A, Joensuu H, Hiltunen J, Hannelin M, Heikkila J, Jakobsson M, et al. Samarium-153-EDTMP in bone metastases. J Nucl Biol Med 1994; 38(4, suppl 1):123–127. Moedritzer K, Irani R. The direct synthesis of a-aminomethylene phosphonic acids: Mannich type reaction with orthophosphorous acid. J Org Chem 1966; 31:1603. Bailya T, Burgada R, Lecouvey M, Neuman A, Prange´ T. Racemic trans-1,2diaminocyclohexane as a template in the synthesis of ligands for transition metal and actinide in vivo detoxification. Phosphorous, Sulfur, Silicon 1995; 101:131–140. Bisaz S, Jung A, Fleisch H. Uptake by bone of pyrophosphate, diphosphonates and their technetium derivatives. Clin Sci Mol Med 1978; 54:265–272. Jung A, Bisaz S, Fleisch H. The binding of pyrophosphate and two diphosphonates by hydroxyapatite crystals. Calcif Tissue Res 1973; 11:269–280. Fitton A, McTavish D. Pamidronate. A review of its pharmacological properties and therapeutic efficacy in resorptive bone disease. Drugs 1991; 41:289–318. Francis MD, Ferguson DL, Tofe AJ, Bevan JA, Michaels SE. Comparative evaluation of three diphosphonates in vitro adsorption (C-14 labeled) and in vivo osteogenic uptake (Tc-99m complexed). J Nucl Med 1980; 21: 1185–1189.

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626 Nuclear Medicine Communications 2006, Vol 27 No 8

16 Richards AG. Metastatic calcification detected through scanning with Tc-99m polyphosphate. J Nucl Med 1974; 15:1057–1060. 17 Val J, Lewington BM. Bone-seeking radionuclides for therapy. J Nucl Med 2005; 46(suppl):38S–47S. 18 Nair N. Relative efficacy of P-23 and Sr-89 in palliation in skeletal metastases. J Nucl Med 1999; 40:256–261.

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Breen SL, Powe JE, Porter AT. Dose estimation in strontium-89 radiotherapy of metastatic prostatic carcinoma. J Nucl Med 1992; 33: 1316–1323. Laing AH, Ackery DM, Bayly RJ, Buchanan RB, Lewington VJ, McEwan AJ, et al. Strontium-89 therapy for pain palliation in prostatic skeletal malignancy. Br J Radiol 1991; 64:816–822.

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Original article

Recombinant human thyroid-stimulating hormone is effective for radioiodine ablation of post-surgical thyroid remnants Daniele Barbaroa, Giuseppe Bonib, Giuseppe Meuccia, Umberto Simia, Paola Lapia, Paola Orsinia, Cristina Pasquinia, Anna Turcoa and Giuliano Marianib Objective To investigate whether recombinant human thyroid-stimulating factor (rhTSH) is effective for the radiometabolic ablation of post-surgery thyroid remnants, using low doses of 131I. Patients and methods The study included two groups of patients enrolled consecutively: group 1 consisted of 52 patients with papillary cancer or minimally invasive follicular cancer (stage I and II), and group 2 consisted of 41 patients with the same stage of disease. All patients underwent a total thyroidectomy. Group 1 received 1.11 GBq (30 mCi) 131I for post-surgical remnants ablation with the aid of rhTSH, while group 2, in the hypothyroid state, received the same amount of radioiodine. To minimize iodine interference, all patients remained on a low iodine diet for 2 weeks and L-thyroxine (L-T4) was stopped for 4 days in the group of patients treated with the aid of rhTSH. To investigate 131I uptake in this group, a tracer dose was administered 3 h after the second injection of rhTSH and the uptake was evaluated at 24 h just before administration of the therapeutic dose. 131I was also measured in the patients treated in the hypothyroid state just before the therapeutic dose was given. Results After 1 year both groups were studied by using whole-body scintigraphy (WBS) and measuring thyroglobulin after rhTSH. In group 1, WBS was negative in 76.9% (40 patients), while thyroglobulin-stimulated

Introduction After a total thyroidectomy, the whole strategy in the treatment and follow-up of differentiated thyroid cancer is based on the capability of normal and tumoral thyroid cells to take up iodine and synthesize thyroglobulin. These extremely important points are dependent upon thyroid-stimulating hormone (TSH) and, hence, maximum stimulation of TSH must be achieved. Powerful stimulation of TSH can be achieved by interrupting L-thyroxine (L-T4) for at least 30 days and causing the consequent hypothyroid state and, at present, by using recombinant human TSH (rhTSH). rhTSH is now used in the follow-up of differentiated thyroid cancer [1–4]. Additionally, rhTSH appears to be useful for the treatment of advanced diseases in patients

levels were < 1.0 ng
ml – 1 in 86.5% (45 patients). In Group 2, WBS was negative in 75.6% (31 patients), while thyroglobulin-stimulated levels were < 1 ng
ml – 1 in 78.0% (32 patients). 131I uptake was 2.29 ± 0.45 in the group treated with the aid of rhTSH, and 3.30 ± 0.7 in the group treated in the hypothyroid state (P = 0.2). No patients treated with the aid of rhTSH and with the short stoppage of L-T4 experienced symptoms of hypothyroidism, and free thyroxine (FT4) and thyroid-stimulating hormone levels remained normal. Conclusions Our data confirm that, when the interference of iodine is minimized, rhTSH is highly effective for the treatment of post-surgical thyroid remnants using a low c 2006 dose of 131I. Nucl Med Commun 27:627–632 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:627–632 Keywords: rhTSH, differentiated thyroid cancer, radiometabolic treatment a Spedali Riuniti di Livorno, Italy and bU.O. Nuclear Medicine, University of Pisa, Italy.

Correspondence to Dr Daniele Barbaro, Endocrinology Section, Spedali Riuniti di Livorno, viale Alfieri 36, 57100 Livorno, Italy. Tel: + 0039 (0)586 223017; fax: + 0039 (0)586 223367; e-mail: [email protected] or [email protected] Received 1 April 2006 Accepted 3 May 2006

who can not tolerate the withdrawal of L-T4 [5–8]. The use of rhTSH, together with high doses of 131I, for the ablation of post-surgical thyroid remnants appears to be effective [9,10], although the literature is not definitive about the outcome of ablation when rhTSH is used with low doses (1.1 GBq (30 mCi)) of 131I [11]. Our previous data in a small group of patients suggested a possible role of iodine coming from exogenous L-T4. In this study, treatment with rhTSH and 1.11GBq of 131I, combined with briefly stopping L-T4 and maintaining a low iodine diet for 2 weeks, has shown an outcome of ablation in the same range as that obtained by treating patients in the hypothyroid state [12]. Using the same protocol we have treated a further, larger group of patients. In this study we have also evaluated

c 2006 Lippincott Williams & Wilkins 0143-3636

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628 Nuclear Medicine Communications 2006, Vol 27 No 8

Fig. 2 131

TSH, FT3, FT4,50 µg I ,, 24 h uptake 131I Tg and AbTg Tg serum serum measurement measurement ,,

the uptake of 131I before treatment. Moreover, we have evaluated the outcome of ablation and compared it with the group of patients treated in hypothyroidism.

Patients and methods The comparison was made between patients treated with the aid of rhTSH (group 1) and patients treated in the hypothyroid state (group 2). Group 1 included 52 patients (aged 19–71 years; 35 women and 17 men) who were enrolled consecutively, with their written consent, after rhTSH was made available in Italy. This group also included the small group of patients enrolled in the first year and previously reported [12]. All patients had papillary cancer (except for the more aggressive variants) or minimally invasive follicular cancer, and could be considered patients at low risk of recurrence (stages I and II). All patients underwent a total thyroidectomy and lymphectomy of the central compartment of the neck. Patients in whom we found positive laterocervical lymph nodes at fine-needle aspiration before surgery also underwent modified ipsilateral laterocervical lymphectomy. After surgery, all these patients began treatment with a TSH-suppressive dose of L-T4. After at least 30 days, TSH and free thyroid hormones were checked and the dose of L-T4 eventually modified. Subsequently, with the approval of the local ethics committee, patients received 131I treatment (38–62 days later) using rhTSH as reported in Fig. 1 and in the following paragraph. To evaluate 131I uptake, 30 consecutively enrolled patients were investigated by using a tracer dose of 131I given before the therapeutic dose, as reported in Fig. 2. Group 2 was represented by all patients (41 patients aged 24–69 years; 26 women and 15 men) with the same features of diseases (histology and stage) and treated consecutively in the 3 years before Fig. 1

TSH, FT3, FT4, Tg and AbTg serum measurement Tg serum measurement Total thyroidectomy

Day 0 LT4 treatment

Total thyroidectomy

LT4 treatment

Hr TSH

30

Day 0

0

LT4 treatment Check of LT4 treatment

1

2

Stop LT4

3

4 5

6

131

I 1.11 GBq

7 WBS

Low-iodine diet

Protocol for the administration of rhTSH and for studying the uptake of 131 I.

Table 1

Histology and clinical stage of both groups of patients

studied Group and age (years) Group 1 < 45 > 45 Group 2 < 45 > 45

pT1–2, Nx

pT1–2, N1

CVPC

FVPC

Follicular cancer

15 27

10

17 19

7 7

1 1

11 23

7

10 19

4 5

2 1

CVCP, classical variant of papillary cancer; FVPC, follicular variant of papillary cancer.

rhTSH was made available in Italy. Also, this group included the small group of patients consecutively enrolled the year before, as previously reported [12]. These patients underwent thyroxine withdrawal and 131I treatment was carried out in the hypothyroid state, the patients having adhered to a low iodine diet for the preceding 2 weeks. The time between thyroidectomy and 131I treatment was 42–91 days, depending on the availability of 131I for treatment.

Hr TSH LT4 treatment

30 Check of LT4 treatment

0

1

Stop LT4

2 131

3

4

5

6

I 1.11 GBq

7 WBS

Table 1 summarizes the pTNM stage and histology of the cancers in both groups of patients. TSH, free thyroxine (FT4), free triiodothyronine (FT3), thyroglobulin, antithyroglobulin antibody and anti-thyroperoxidase (antiTPO) were measured by Immulite 2000 (Diagnostic Products, Los Angeles, California). Functional sensitivity of Tg was 0.5 ng
ml – 1.

Low-iodine diet Protocol for the administration of recombinant human thyroidstimulating hormone (rhTSH).

Protocol for the administration of rhTSH

The protocol for the administration of rhTSH, both for therapeutic and diagnostic purposes, was the same as

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rhTSH is effective in radioiodine ablation of thyroid remnants Barbaro et al. 629

represented in Fig. 1 and reported below. After adhering to a low iodine diet for 2 weeks patients underwent stimulation with rhTSH (Thyrogen, Genzyme). rhTSH (0.9 mg) was administered intramuscularly for two consecutive days. L-T4 was stopped the day before the first administration of rhTSH and given again the day after administration of 131I. Serum samples for the determination of TSH, FT4, FT3, thyroglobulin and anti-thyroglobulin antibody were taken the day before the first administration of rhTSH and the day after the second administration of rhTSH. A serum sample for thyroglobulin was also taken 2 and 3 days after the last administration of rhTSH. Administration of 131I for therapeutic and diagnostic purposes and whole-body scintigraphy

Between 38 and 62 days after surgery, 1.11 GBq of 131I was administered, for therapeutic purposes, the day following the last injection of rhTSH, and post-therapy WBS was carried out after 4–6 days. In 30 consecutively enrolled patients a tracer dose of 185 MBq was given 3 h after the last injection of rhTSH, and 131I uptake was evaluated at 24 h just before the administration of the therapeutic dose (Fig. 2). Diagnostic 131I WBS was performed (after 12 months) according to the same protocol for stimulation by rhTSH. Images were obtained 48 h after oral administration of 185 MBq 131I with a double-headed gamma camera (Millenium MG; GE Medical Systems, Massachusetts, USA) using a 3/8-inch thick crystal and a high-energy, general-purpose collimator. Anterior and posterior views of WBS were acquired after scanning for a minimum of 30 min. Anterior neck/chest spot views with and without markers (99mTc) on the suprasternal notch and chin were acquired after scanning for a minimum of 15 min or after obtaining 150 000 counts.

Statistical analysis

Results are expressed as mean ± SE for laboratory data and as percentage for the groups of subjects. Student’s ttest was used to compare laboratory data. The chisquared test was used to detect differences in the proportion of cases.

Results Laboratory data in patients who underwent 131I treatment were as follows. In the first group TSH was 0.04–0.35 mU
l – 1 before rhTSH and 74–210 mU
l – 1 (126 ± 10) after the second injection of rhTSH, and in the second group (hypothyroid patients) 36–82 mU
l – 1 (50 ± 3). At that time thyroglobulin was 1.1–22.3 ng
ml – 1 (8.8 ± 1.71) in patients treated with rhTSH stimulation ( < 0.5–11.0 ng
ml – 1 before rhTSH) and < 0.5– 21.1 ng
ml – 1 (7.48 ± 1.67) in hypothyroid patients. Free T4 was normal in patients treated with rhTSH before (1.48–1.91 ng
dl – 1, 19.04–24.58) and after (1.24–1.58, 14.11–20.33) the 4-day cessation of L-T4. After the therapeutic dose, WBS showed residual thyroid tissue in the thyroid bed at the post-treatment scan both in patients treated in hypothyroidism and in patients treated by rhTSH. Also, three patients treated with rhTSH showed minor abnormal areas of uptake in the neck, and three hypothyroid patients showed a similar minor abnormal uptake in the neck.

In all patients, the serum levels of TSH, FT4, FT3, thyroglobulin and anti-thyroglobulin antibodies were assessed periodically. All patients had undetectable levels of thyroglobulin during TSH-suppressive treatment. Patients who were positive for anti-thyroglobulin antibodies were excluded from the study.

At the 1-year follow-up, 40 patients (76.9%) treated with I after rhTSH showed negative WBS; in six cases there was a minor uptake in the thyroid bed, and in six other cases WBS was clearly positive for a residual uptake in the thyroid bed. In patients treated with 131I in the hypothyroid state WBS was negative in 31 patients (75.6%); in three cases (7.3%) there was a minor uptake in the thyroid bed and in the other seven cases (17.0%) WBS was clearly positive for a residual uptake in the thyroid bed. In the group of patients treated by rhTSH, thyroglobulin was < 1.0 ng
ml – 1 in 45 patients (86.5%), between 1 and 2 ng
ml – 1 in three patients (5.7%) and > 2 ng
ml – 1 in four patients (7.6%). In the group of hypothyroid patients, thyroglobulin was < 0.5 in 32 patients (78.0%), between 1 and 2.0 ng
ml – 1 in one patient (2.4%) and > 2.0 ng
ml – 1 in eight patients (19.5%). In Figs 3 and 4 data of WBS and serum thyroglobulin in both groups of patients are reported. The statistical comparison between the two groups showed no difference. Data for TSH and free thyroid hormones at this time were similar to those before treatment with 131I and are not reported.

After 1 year the outcome of thyroid ablation was assessed in both groups by ultrasonography, a conventional 131I scan and serum thyroglobulin measurements using rhTSH, as stated above.

Comparison of 131I uptake showed a value of 2.29 ± 0.45 in the group that received rhTSH, and a value of 3.30 ± 0.7 in the group treated in hypothyroid state (P = 0.2).

Uptake into the thyroid bed was diagnosed on wholebody or spot-view images that showed only visible uptake between the suprasternal notch and thyroid cartilage and after exclusion of pathological lymph nodes by ultrasonography. Follow-up

131

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630

Nuclear Medicine Communications 2006, Vol 27 No 8

Fig. 3

100% Patient treated in hypothyroidism No residue 31 41

Patient treated with rhTSH

40 52 Residue Minimal uptake 3 41

7 41

6 52

6 52

Whole-body scanning with a tracer dose of 131I at the first follow-up after 1 year (number of patients is indicated inside the columns).

Fig. 4

100%

Tg < 1.0

Patient treated in hypothyroidism

However, a correct comparison between treatment after L-T4 withdrawal and treatment with the aid of rhTSH can be made only by creating the same presuppositions for the outcome of ablation.

Patient treated with rhTSH 32 41

rhTSH for the preparation of radioiodine treatment in advanced disease [4–8], fewer reports have investigated the use of rhTSH with radioiodine for the post-surgical ablation of thyroid remnants. The use of high standard doses (not less than 3.7 GBq) after preparation by rhTSH has been shown to ablate post-surgical remnants [9,10,16], and the use of rhTSH is now reported in indications regarding prescription information. In contrast, the situation is less clear for the use of rhTSH with 1.11GBq of radioiodine. In one study in which standard low doses (1.11 GBq) were used, it was shown that the preparation by rhTSH is not as effective as withdrawal of L-T4 [11]. In this study the therapeutic dose of 131I was delayed by 24 h to allow the measurement of a 24-h uptake with a tracer dose of 1.85 MBq, and this could explain, at least partially, the worse outcome of 131I ablation. However, in the same study 131I uptake was lower after administration of rhTSH than after withdrawal of L-T4, hence suggesting a true lower effectiveness of rhTSH, at least for the stimulation of this first step.

45 52

Tg > 2 Tg 1 - 2 1 41

3 52

8 41

4 52

Serum thyroglobulin values at the first follow-up after 1 year (number of patients is indicated inside the columns).

After administration of rhTSH, no patient had any significant side effects. No symptoms, even mild, of hypothyroidism were reported by patients during the 4 days of L-T4 withdrawal.

The importance of keeping a low iodine intake is well described in many reports [17–19], and for this reason the possible interference of iodine from the metabolism of L-T4, which is at least 60 mg
day – 1, could represent an important factor. Our previous report on a small group of patients showed that the preparation by rhTSH with a cessation in L-T4 administration for 4 days before giving 131 I showed the same rate of post-surgical ablation [12]. In this study, where the iodine level in the diet was under careful control, the short interruption of T4 was capable of decreasing ioduria (mean, 47.2 vs. 76.4) with respect to a control group in which a brief cessation of L-T4 was not carried out.

There is general agreement that the use of 131I for the ablation of post-surgical thyroid remnants after total thyroidectomy appears to reduce recurrences and death in patients with differentiated thyroid cancer [13–15].

Our present results show that post-surgical thyroid remnant ablation with 1.11 GBq after preparation with rhTSH has the same percentage of success as found in patients prepared in hypothyroidism. When we considered both thyroglobulin values and WBS criteria for complete ablation, the outcome was the same.

The use of empirical fixed doses is most widespread due to their simplicity; standard doses usually employed range from 1.11 to 3.7 GBq or more when there is evidence of local spreading or of metastatic disease. However, most patients can be treated with lower doses and 1.11 GBq appears to be effective for post-surgical thyroid ablation, at least in ‘low risk cancer’. It is interesting to note that, although several reports have shown the effectiveness of

As has been reported in many previous studies, stimulated thyroglobulin levels represent the best means of detecting normal or pathological thyroid tissue [2–4]. Thyroglobulin levels of < 1.0 ng
ml – 1 suggest the absence of disease while levels > 2.0 ng
ml – 1 indicate the possible presence of metastatic disease. In group 1 our results have shown a clear trend to a lower percentage of cases with stimulated thyroglobulin levels

Discussion

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rhTSH is effective in radioiodine ablation of thyroid remnants Barbaro et al. 631

> 2.0 ng
ml – 1 and although this result did not reach statistical difference we could suppose that treatment with the aid of rhTSH could decrease the number of patients who have metastatic spread of the disease.

could be that the acute stimulation by rhTSH fails to stimulate suitably the last steps in hormone production (i.e., thyroid hormone secretion), making for a longer period of persistence of radioiodine.

The therapeutic action of 131I is based on complex mechanisms; it depends on the amount of 131I uptake, on the amount of 131I which is organified and stored, and on the amount of hormone secretion. So a high 131I can not necessarily determine whether there will be a severely damaging effect if, for example, 131I is not stored inside cells and follicles for a sufficiently long time. As is known, a very high turnover of iodine in Graves’ disease may be a cause of failure of radiometabolic treatment. However, 131 I uptake represents the first step in the action of 131I and can be considered as a good indicator of the possible action of radioiodine.

Conclusion

The study of 131I uptake was not easy in patients prepared with rhTSH because, in order to investigate the correct 131I uptake, we should administer the tracer dose 24 h after the last injection of rhTSH, and so delay the therapeutic dose. On the other hand, administration of the tracer dose soon after the last injection of rhTSH could show a lower uptake because, after rhTSH stimulation, synthesis of Na/I Symporter (NIS) begins after 3–6 h, reaching a maximum at 24 h. Furthermore, an increased thyroid iodine uptake is detected 12 h after rhTSH with a maximum after 72 h [20,21]. Regarding these considerations our protocol of administration of the tracer dose of 131I could permit correct treatment, but there is a possibility that it could underestimate the value of 131I uptake. Notwithstanding all these considerations, our data have shown only a slight difference between values of 131I after rhTSH and the hypothyroid state, without statistical difference. This confirms that at least the first step of the complex process that leads to the action of radioiodine is suitably stimulated by rhTSH when the levels of dietary iodine are kept low and when the iodine coming from T4 is minimized. The clearance of iodine in euthyroidism is faster than in the hypothyroid state and this produces less radiation to the body [22,23]; therefore the possibility of decreasing the exposure to radiation without significantly decreasing 131I uptake and the therapeutic action of 131I represents unique advantages of radioiodine treatment with rhTSH preparation. As already stated, our data appear to show a better result for the outcome of radioiodine ablation using rhTSH, at least when we consider the values of thyroglobulin. Further study concerning the persistence of 131I inside the cells and follicles should be done, but a possible explanation for the better action of 131I after rhTSH

Our study shows that, when a correct comparison between radioiodine treatment after rhTSH stimulation and during the hypothyroid state is made, we obtain the same percentage of ablation. The condition of increased clearance that we have in euthyroidism in respect to the hypothyroid state probably makes the iodine intake of primary importance in the outcome of thyroid ablation. The possibility of effective treatment for thyroid cancer, without symptoms of hypothyroidism and with less bone marrow exposure to radiation, represents unique advantages of the use of rhTSH. Further methods of reducing the iodine pool (e.g., use of diuretics) and perhaps a slight modification of the protocol for the administration of rhTSH in order to increase 131I uptake and its retention inside thyroid tissue could, in many cases, permit a further reduction in the doses of 131I employed.

References 1

Robbins RJ, Tuttle RM, Sharaf RN, Larson SM, Robbins HK, Ghossein RA, et al. Preparations by recombinant human thyrotropin hormone withdrawal are comparable for the detection of residual differentiated thyroid carcinoma. J Clin Endocrinol Metab 2001; 86:619–625. 2 Mazzaferri EL, Kloos RT. Is diagnostic iodine-131 scanning with recombinant human TSH useful in the follow-up of differentiated thyroid cancer after thyroid ablation? J Clin Endocrinol Metab 2002; 87:1490–1498. 3 Mazzaferri EL, Robbins RJ, Spencer CA, Braverman LE, Pacini F, Wartofsky L, et al. A consensus report of the role of serum thyroglobulin as a monitoring method for low-risk patients with papillary thyroid carcinoma. J Clin Endocrinol Metab 2003; 88:1433–1441. 4 Schlumberger M, Pacini F, Wiersinga WM, Toft A, Smit JV, Sanchez Franco F, et al. Follow up and management of differentiated thyroid carcinoma: a European perspective in clinical practice. Eur J Endocrinol 2004; 151: 539–548. 5 Pellegriti G, Scollo C, Giuffrida D, Vigneri R, Squatrito S, Pezzino V. Usefulness of recombinant human thyrotrophin in the radiometabolic treatment of selected patients with thyroid cancer. Thyroid 2001; 11: 1025–1030. 6 Chiu AC, Delpassand ES, Sherman SI. Prognosis and treatment of brain metastases in thyroid carcinoma. J Clin Endocrinol Metab 1997; 82: 3637–3642. 7 Adler ML, Macapinlac HA, Robbins RJ. Radioiodine treatment of thyroid cancer with the aid of recombinant human thyrotropin. Endocr Pract 1998; 4:282–286. 8 Robbins RJ, Voelker E, Wang W, Macapinlac HA, Larson SM. Compassionate use of recombinant human thyrotropin to facilitate radioiodine therapy: case report and review of literature. Endocr Pract 2000; 6:460–464. 9 Robbins RJ, Tuttle RM, Sonenberg M, Shaha A, Sharaf R, Robbins H, et al. Radioiodine ablation of thyroid remnants after preparation with recombinant human thyrotropin. Thyroid 2001; 11:865–869. 10 Berg G, Lindstedt G, Suurkula M, Jansson S. Radioiodine ablation and therapy in differentiated thyroid cancer under stimulation with recombinant human thyroid-stimulating hormone. J Endocrinol Invest 2002; 25:44–52. 11 Pacini F, Molinaro E, Castagna MG, Lippi F, Ceccarelli C, Agate L, et al. Ablation of thyroid residues with 30 mCi (131)I: a comparison in thyroid cancer patients prepared with recombinant human TSH or thyroid hormone withdrawal. J Clin Endocrinol Metab 2002; 87:4059–4062. 12 Barbaro D, Boni G, Meucci G, Simi U, Lapi P, Orsini P, et al. Radioiodine treatment with 30 mCi after recombinant human TSH stimulation in thyroid cancer: effectiveness for post-surgical remnants ablation and possible role of iodine content in L-T4 in the outcome of ablation. J Clinical Endocrinol Metab 2003; 88:4110–4114.

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13 De Groot LJ, Kaplan EL, McMormick M, Strauss F. Natural history, treatment and course of papillary thyroid carcinoma. J Clin Endocrinol Metab 1990; 71:414–424. 14 Mazzaferri EL, Jhiang SM. Long-term impact of initial surgical and medical therapy on papillary and follicular thyroid cancer. Am J Med 1994; 97:418–428. 15 Sclumberger M. Papillary and follicular thyroid carcinoma. N Engl J Med 1998; 338:297–306. 16 Ralli M, Cohan P, Lee K. Successful use of recombinant human thyrotropin in the therapy of pediatric well-differentiated thyroid cancer. J Endocrinol Invest 2005; 28:270–273. 17 Park JT, Hennessey JV. Two-week low iodine diet is necessary for adequate outpatient preparation for radioiodine rhTSH scanning in patients taking levothyroxine. Thyroid 2004; 14:57–63. 18 Loffler M, Weckesser M, Franzius C, Kies P, Schober O. Iodine excretion during stimulation with rhTSH in differentiated thyroid carcinoma. Nuclearmedizin 2003; 42:240–243.

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Maruca J, Santner S, Miller K, Santen RJ. Prolonged iodine clearance with a depletion regimen for thyroid carcinoma: concise communication. J Nucl Med 1984; 25:1089–1093. Kogai T, Endo T, Saito T, Miyazaki A, Kawaguchi A, Onaya T. Regulation by thyroid-stimulating hormone of sodium/iodidesymporter gene expression and protein levels in FRTL-5 cells. Endocrinology 1997; 138:2227–2232. Duick DS, Baskin HJ. Utility of recombinant human thyrotropin for augmentation of radioiodine uptake and treatment of non-toxic and toxic multinodular goiters. Endocrine Practice 2003; 9:204–209. De Keiser B, Hoekstra A, Konijnenberg MW, De Vos F, Lambert B, Van Rijk PP, et al. Bone marrow dosimetry and safety of high 131I activities given after recombinant human thyroid-stimulating hormone to treat metastatic differentiated thyroid cancer. J Nucl Med 2004; 45:1549–1554. Menzel C, Kranert WT, Dobert N, Diehl M, Fietz T, Hamscho N, et al. RhTSH stimulation before radioiodine therapy in thyroid cancer reduces the effective half-life of 131I. J Nucl Med 2003; 44:1069–1071.

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Review paper

Nuclear medicine imaging of bone infections Napoleone Prandinia, Elena Lazzerib, Brunella Rossic, Paola Erbab, Maria Gemma Parisellad and Alberto Signored The inflammation and infection of bone include a wide range of processes that can result in a reduction of function or in the complete inability of patients. Apart from the inflammation, infection is sustained by pyogenic microorganisms and results mostly in massive destruction of bones and joints. The treatment of osteomyelitis requires long and expensive medical therapies and, sometimes, surgical resection for debridement of necrotic bone or to consolidate or substitute the compromised bones and joints. Radiographs and bone cultures are the mainstays for the diagnosis but often are useless in the diagnosis of activity or relapse of infection in the lengthy management of these patients. Imaging with radiopharmaceuticals, computed tomography and magnetic resonance are also used to study secondary and chronic infections and their diffusion to soft or deep tissues. The diagnosis is quite easy in acute osteomyelitis of long bones when the structure of bone is still intact. But most cases of osteomyelitis are subacute or chronic at the onset or become chronic during their evolution because of the frequent resistance to antibiotics. In chronic osteomyelitis the structure of bones is altered by fractures, surgical interventions and as a result of bone reabsorption produced by the infection. Metallic implants and prostheses produce artefacts both in computed tomography and magnetic resonance images, and radionuclide studies should be essential in these cases. Vertebral osteomyelitis is a specific entity that can be correctly diagnosed by computed tomography or magnetic resonance imaging at the onset of symptoms but only with radionuclide imaging is it possible

Peripheric post-traumatic and prosthetic joint infection Background

Osteomyelitis is an infection of bone caused by a pyogenic organism, primarily the Staphylococcus aureus. Histologically, osteomyelitis is categorized as acute, subacute or chronic, with the presentation of each type based on the time of disease onset (i.e., occurrence of infection or injury). Acute osteomyelitis develops within 2 weeks after disease onset, subacute osteomyelitis within one to several months and chronic osteomyelitis after a few months. Other classification systems of osteomyelitis, beyond the general categories of acute, subacute and chronic, are preferred clinically. The Waldvogel classification system divides osteomyelitis into the categories of haematogenous, contiguous and chronic

to assess the activity of the disease after surgical stabilization or medical therapy. The lack of comparative studies showing the accuracy of each radiopharmaceutical for the study of bone infection does not allow the best nuclear medicine techniques to be chosen in an evidence-based manner. To this end we performed a meta-analysis of peer reviewed articles published between 1984 and 2004 describing the use of nuclear medicine imaging for the study of the most frequent causes of bone infections, including prosthetic joint, peripheric post-traumatic bone infections, vertebral and sternal infections. Guidelines for the choice of the optimal radiopharmaceuticals to be used in each clinical condition and for different aims is provided. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:633–644
Nuclear Medicine Communications 2006, 27:633–644 Keywords: osteomyelitis, spondylodiskitis, prosthesis-related infections, radionuclide imaging a Struttura Complessa di Medicina Nucleare, Azienda Ospedaliero-Universitaria, Ferrara, Italy, bIstituto di Medicina Nucleare, Universita` degli Studi di Pisa, Italy, c Struttura Complessa di Medicina Nucleare, Ospedale Umberto I, Azienda Ospedaliera, Ancona, Italy and dIstituto di Medicina Nucleare, 2a Facolta` di Medicina, Universita` ‘‘La Sapienza’’, Rome, Italy.

Correspondence to Dr Napoleone Prandini, Nuclear Medicine Department, Corso Giovecca 203, 44100 Ferrara, Italy. Tel: + 39 0532 236387; fax: + 39 0532 237553; e-mail: [email protected] Received 27 June 2005 Accepted 5 May 2006

(Table 1) [1–3]. In the same way in the more recent Cierny–Mader system the terms acute and chronic are not used still (Table 2) [4]. The stages in this system are dynamic and may be altered by changes in the medical condition of the patient (host), successful antibiotic therapy and other treatments. On the other hand, the classification systems for osteomyelitis describe the infection and determine the need for surgery, but the categories do not apply to special circumstances (i.e., infections involving prosthetic joints, implanted materials) or special types of infection (e.g., sternal or vertebral osteomyelitis). Acute haematogenous osteomyelitis occurs predominantly in children, usually involving the metaphysis of long bones. Patients have signs of systemic illness,

c 2006 Lippincott Williams & Wilkins 0143-3636


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Table 1 Classification system for osteomyelitis according to Waldvogel et al. [1]

of the bone to open fractures or the implants of joint prostheses are the most common causes of osteomyelitis.

K Haematogenous osteomyelitis K Osteomyelitis secondary to contiguous focus of infection

3 No generalized vascular disease 3 Generalized vascular disease K Chronic osteomyelitis (necrotic bone)

Table 2

Staging system for osteomyelitis, adapted from Cierny

et al. [4] Anatomical type K Stage 1 K Stage 2 K Stage 3 K Stage 4 Physiological class K A host K B host Bs Bl Bls K C host

Medullary osteomyelitis Superficial osteomyelitis Localized osteomyelitis Diffuse osteomyelitis

Healthy Systemic compromise Local compromise Local and systemic compromise Treatment worse than the disease

Factors affecting immune surveillance, metabolism and local vascularity Systemic factors (Bs): malnutrition, renal or hepatic failure, diabetes mellitus, chronic hypoxia, immune disease, extremes of age, immunosuppression or immune deficiency Local factors (Bl): chronic lymphoedema, venous stasis, major vessel compromise, arteritis, extensive scarring, radiation fibrosis, small-vessel disease, neuropathy, tobacco abuse

including fever, irritability and lethargy with tenderness over the involved bone and decreased range of motion in adjacent joints. The subacute and chronic forms of osteomyelitis usually occur in adults, generally secondary to an open wound, most often an open injury or a surgical intervention to bone and surrounding soft tissue. Localized bone pain, erythema and drainage around the affected area are frequently present with deformity, instability and local signs of impaired vascularity. The infection usually spreads from bone to soft tissues and vice versa. So the incidence of deep musculoskeletal infections from open fractures is high (23%) [5]. In bone infection, usually the pus is produced within the medulla, and may result in a swollen or in a typical abscess of marrow space. As the swollen tissue presses against the rigid outer wall of the bone, the blood vessels in the bone marrow may become compressed, reducing or cutting off the blood supply to the bone. Parts of the bone may die and remain excluded by the haematic supply of medical treatment. In these areas infection can persist for months or years causing complication with necrosis and fractures of the infected bone. Surgical treatment of osteomyelitis involves debridement of necrotic bone and tissue, and, the implant of metallic hardware to obtain bone stability. Moreover, the exposure

Diagnosis

The diagnosis of osteomyelitis is based on the clinical findings, with physical examination and laboratory tests: elevations in the erythrocyte sedimentation rate and C-reactive protein are the most frequent signs of infection. Leukocytosis may be noted and blood cultures could be positive more frequently in acute osteomyelitis. In osteomyelitis plain film radiography shows a typical evidence of bone destruction with deep soft-tissue swelling but radiographic signs may not appear until approximately 2 weeks after the onset of infection [6]. This could be a problem in paediatric age groups where osteomyelitis of long bones can lead to damage of growing cartilage with a cessation of bone lengthening [7]. The radiological diagnosis of haematogenous osteomyelitis is quite simple because of the intact structure of surrounding normal bone [8]. In secondary and chronic osteomyelitis bony changes include osteolysis, periosteal reaction, sequestra of necrotic bone and sub-periosteal new bone formation and these signs are less specific for the activity of the infection [9]. Ultrasonography may be helpful in the diagnosis of muskuloskeletal infections detecting fluid collections (e.g., an abscess) and periostitis or guiding the biopsy but it is limited in deep tissues and in bone marrow infections [10,11]. The culture of joint aspiration and fluid collection is necessary to assess the aetiology of an infection and to set a specific treatment. Moreover, in approximately 30% of all osteomyelitis the results of culture are equivocal or negative because of antibiotic treatment or of the difficulty in obtaining sufficient or adequate material [12–14]. The computed tomography (CT) signs of osteomyelitis include osteolysis in cortical bone, small foci of gas and minute foreign bodies or increased vascularity after administration of contrast media. CT cannot detect early functional processes in bone infections because of the delay in detecting bone structure alterations but it is less limited by artefacts than is ultrasonography [15]. Magnetic resonance imaging (MRI) has a high sensitivity for osteomyelitis and infection of soft tissues and also provides greater spatial resolution in delineating the anatomical extension of osteomyelitis. Moreover, both CT and MRI lack specificity in chronic infection and in prosthetic joint replacements when metallic devices are implanted because of the artefacts due to metallic implant and the difficulty of detecting active infection

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Nuclear medicine imaging of bone infections Prandini et al. 635

in altered bone after surgical treatments or repairing bone processes [16,17]. MRI is particularly useful in haematogenous osteomyelitis and in diskitis involving the axial skeleton and pelvis. The bone structure alterations in chronic infection can persist intermittently for years. Even with intravenous administration of contrast media it is difficult to distinguish latent infection from simple bone remodelling by both CT and MRI. The role of nuclear medicine

Three-phase bone scintigraphy with 99mTc-disphosphonates (MDP, HDP, DPD, HEDP) is the most widely diffuse nuclear imaging procedure in the study of bone infections because of its low cost and high availability [18–20]. In acute haematogenous osteomyelitis and in children bone scintigraphy becomes positive 24–48 h after the onset of symptoms of infection. The bone scintigraphy signs of osteomyelitis are an increased vascularity both in dynamic and in blood pool images and an increased metabolic uptake of 99mTc-disphosphonates in late images (2–3 h after i.v. injection). Moreover, the specificity of bone scintigraphy is not high and diagnosis of osteomyelitis in many clinical situations is difficult. Bone scintigraphy cannot correctly distinguish osteomyelitis from a neurotrophic lesion, post-surgical changes, a healing fracture or a chronic infection from a simple mobilization of joint prosthesis. Radiolabelled autologous leukocytes (WBCs) are widely used to image infection and inflammation because of their excellent performance in most cases of osteomyelitis [21–24]. Scintigraphy with WBCs or granulocytes labelled with both 111In and 99mTc-hexamethylpropylene amine oxime (99mTc-HMPAO) are preferred in the diagnosis of bone infections secondary to traumas and fractures, and in the study of prosthetic joint implants [25–28]. The main problem of leukocyte scintigraphy is the normal margination of leukocytes and granulocytes in bone marrow: in central bone this produces a lack of specificity because of a high percentage of ‘cold defects’ due to lower blood supply areas and to necrotic bone. In contrast, in peripheral bones, especially after orthopaedic surgery, there are often displacements of bone marrow with ‘hot spots’ that must be distinguished from focal infection [29]. Therefore, in chronic osteomyelitis there is a faint migration of leukocytes from vascular spaces which can result in reduced accuracy of WBC scintigraphy [30,31]. The method of colloid subtraction of bone marrow proposed by Palestro et al. [32] in hip prostheses and the quantitative or semi-quantitative analysis of serial images of labelled WBCs, proposed more recently, has resolved most of these interpretation problems. All recent papers give the comparison with bone marrow activity as

the fundamental method of interpretation of WBC scintigraphy in osteomyelitis [33]. The mechanism of action of labelled monoclonal antibodies (MoAbs) is the binding to surface antigens on granulocytes giving an in-vivo labelling of cells that does not require blood manipulation [34]. It has also been suggested that the antibody may preferentially bind to granulocytes that have migrated from vascular spaces and become activated. A part of the injected tracer may bind to inflamed tissues for increased capillary permeability, due to increased diffusion of proteins into the interstitial space. Recently, Skehan published a study demonstrating that 99mTc-Fab0 directed against NCA-90 surface antigen does not localize in inflammation as a result of binding to circulating granulocytes but is cleared into inflammation non-specifically via increased vascular permeability [35]. Human polyclonal immune globulin (HIG) accumulates in infectious and inflammatory foci by non-specific extravasations, facilitated by locally enhanced vascular permeability. Usually, HIG is labelled by both 99mTc and 111 In and the longer half-life of 111In-HIG compared to that of 99mTc permits images to be obtained after 24– 48 h, thus improving the target:non-target uptake in chronic inflammation where the permeability is only moderately increased [36,37]. 67

Ga was the first tracer to be introduced in nuclear medicine for the detection of inflammation, although the mechanism of uptake into inflamed and neoplastic tissues is still not completely understood [38]. Diffusion from blood by transportation as gallium-transferrin and/or by increased endothelial permeability are the most likely hypotheses. Uptake into lymphoma and other malignancies (e.g., lung cancer) or in chronic granulomatous processes (e.g., sarcoidosis or tuberculosis) and bowel activity are the most frequent problems in the interpretation of gallium studies. Positron emission tomography (PET) using 2-[18F]fluoro2-deoxy-D-glucose (18F-FDG) is a promising technique for the diagnosis of bone infections and inflammation based on the intensive consumption of glucose by mononuclear cells and activated granulocytes [39]. The method may have limitations in distinguishing uncomplicated bone healing from osteomyelitis. Bone healing involves an inflammatory phase that represents a highly activated state of cell metabolism and glucose consumption, mimicking infection on PET images [40]. Meta-analysis

Eighty-nine studies published between 1984 and 2004 have been analysed and considered for a metanalysis of published data concerning the radionuclide imaging of bone infections [13,14,16–20,22–103]. Only the papers

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regarding clinical figures in humans were included: special attention was given to papers dealing with radionuclide techniques that are widely diffused and papers reporting comparative results among the different radionuclide imaging techniques or among nuclear medicine and other procedures to diagnose bone infections (plain film radiography, MRI, CT, ultrasonography, guided aspiration biopsy). Case reports and reviews without clinical results were considered for the discussion but not included in the meta-analysis. Also not included were experimental techniques that are not commercially available in most of countries (e.g., use of peptides, labelled antibiotics, cytokines, liposomes, avidin/biotin). In the first phase, papers regarding the most common causes of bone infections (peripheral open fractures and prosthetic joint infections) were considered. The papers regarding infections of the vertebral column or sternum (which require a particular diagnostic approach) were excluded in the first meta-analysis and collected in the second and third phases. In the same way, all papers regarding infections of the diabetic foot, which are characterized by a singular pathogenesis requiring a specific diagnostic and therapeutic approach, were excluded from this meta-analysis. Most of the papers included in the meta-analysis concern three-phase bone scintigraphy (29 papers) followed by scintigraphy with mixed WBCs or granulocytes labelled with 111In (26 papers) and, more recently, with 99m Tc-HMPAO (22 papers). In the papers published in the past decades the combined technique of bone marrow subtraction (obtained by simultaneous sulfur colloid injection) is preferred for leukocytes labelled by indium or technetium. Other papers included radionuclide imaging of granulocytes labelled by MAbs (eleven), or the use of gallium (seven), HIG (seven), fluid aspiration biopsy (six), MRI (five) and 18F-FDG PET (six) used alone or with different combinations of techniques.

Table 3 reports the results of the meta-analysis collecting the single tracers considered independently by the techniques used (for example, the combination of bone scintigraphy and gallium or WBCs and colloids) and independently by the time after i.v. injection (for example, labelled WBC imagings collected after 2, 4, 8 or 24 h were put together). The first column of Table 3 gives the number of lesions reported in the patients. Where there was more than one lesion for a patient (e.g., in the study of joint prosthesis infections) these were also included. Approximately 8180 lesions were studied with the different methods. The greatest number of lesions were studied with leucocytes (WBCs and granulocytes) labelled with 111In (2147 lesions) and 99mTc-HMPAO (1453 lesions) [62–67]. Both techniques allow a diagnostic accuracy of around 90%. Despite of the highest number of patients studied, labelled WBC scintigraphy is not the most accurate technique. Amazingly, the use of WBCs labelled with 99mTc had a greater sensitivity (89%) and specificity (90.1%) than WBCs labelled with 111In (82.8% and 83.8%, respectively, for sensitivity and specificity) event though most cases of osteomyelitis are typically chronic infections requiring a longer time than acute infections to obtain a significant accumulation of cells. This can be explained by considering the more recent publications dealing with 99mTc labelling (after 1990) in comparison with 111In. Much of the recent work has benefitted from the experience of Palestro, and a combination of 99mTccolloid bone marrow imaging and 111In-oxine leukocyte imaging at 24 h has been proposed [70,71]. For WBCs labelled with 99mTc several authors proposed prolonging the scans until 24 h, which would give a result that was as effective as the comparison with colloid images. The accuracy of labelled WBCs is high in peripheral bone and in acute or sub-acute infections [72]. In the axial skeleton or in chronic infections ‘cold areas’, with a lack of specificity of WBC scintigraphy, are often reported [73].

Table 3 Results of the meta-analysis of data published between 1984 and 2004 regarding the infections of peripheric bone and of prosthetic joint implants Technique 67

Ga In-WBCs Tc-WBCs 99m Tc-BS 99m MoAb BW 250/183 99m MoAb MN3 99m Tc-HIG and 111In-HIG 18 F-FDG PET 99m Tc-nanocolloid MRI Aspiration Total 111

99m

Lesions 569 2147 1453 1527 258 129 537 413 154 95 786 8180

Sensitivity (%) Specificity (%) 70.1 82.8 89.0 85.4 81.7 92 95.2 94.1 89 88.2 50.7

81.8 83.8 89.1 75.2 80.1 86.0 78.7 87.3 80.5 84.7 93.9

Lesions

Accuracy

Lesions

PPV

NPV

396 1327 960 866 365 106 288 273 97 54 663 5207

78.2 84.6 89.1 75.5 83.2 86 86.0 91.9 80.7 88.7 88.4

343 845 768 403 159 NA 323 294 19 19 614 3907

50.0 60.9 75.1 62.9 79.9 NA 72.7 86.9 33 69.0 91.3

89.5 92.4 91.6 95.8 88.5 NA 96.1 94.2 90 100.0 55.5

WBC = labelled white blood cells and granulocytes; BS = bone scan (multiphase); MoAb = monoclonal antibodies against granulocytes antigens; HIG = polyclonal human immune globulin; MRI = Magnetic resonance imaging; CT = computed tomography; Lesions = number of lesions for which the data of sensitivity, specificity accuracy, PPV and NPV were available. PPV = positive predictive value; NPV = negative predictive value; NA = Not available data. All parameters have been weighted for the number of lesions available in each study (sometimes more than one for each patient). The numbers in bold refer to the best performing technique.

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Nuclear medicine imaging of bone infections Prandini et al. 637

Immunoscintigraphy with MAbs against granulocytes has the advantage, according to most authors, of greater simplicity in comparison to in-vitro labelling of WBCs [74,75]. Currently, there are three different antibodies available for clinical use: the whole antibody 99mTc-IgG (Granuloscint (BW 250/183)), 99mTc-Fab0 (Leukoscan (MN3)) and CD15, IgM (LeuTech, Palatin Technologies, now NeutroSpec, marketed by Tyco in the USA). Our meta-analysis included only the papers that refer to the two older of these three MAbs. The accuracy of 99mTcFab0 is higher than the accuracy of whole antibody BW250/183 (86% vs. 83.2%) [1,76–83]. Moreover all murine antibodies injected intravenously can be perceived as foreign proteins that provoke an allergic response from the patient’s immune system, leading to the production of human anti-mouse antibodies. The possibility of producing such antibodies is a serious concern because the chance of a severe allergic reaction may be increased when scintigraphy is repeated, as is frequent in orthopaedic infections. The small Fab0 fragment significantly reduces the chance of a severe immune reaction. Nevertheless, these agents give a result that is somewhat less accurate than that obtained by using in vitro labelled leukocytes [84,85]. HIGs labelled by either 99mTc or 111In do not induce antibody reactions and are as accurate as the MAbs in the diagnosis of bone infections [86–89]. The sensitivity of these tracers is the highest in our meta-analysis (95.2%) but the specificity and the predictive positive index are rather low (78.7% and 72.7%, respectively). This suggests the first indication of immunoglobulin should be to confirm the activity of an inflammation, especially of chronic inflammation of joints, while they are useless in the differential diagnosis between infection and antiseptic inflammation. Nanocolloids labelled with 99mTechnetium are initially employed in the study of bone marrow: the accumulation of colloids in inflammation is related to the increased vascular permeability in inflamed tissues [90]. The papers included in our analysis show an accuracy of 80.7% in imaging of bone inflammation [91–93]. Although the tracer is economic and supplies the result in less than 1 h after the i.v. injection, its use has been almost abandoned because of its low specificity (80.5%). 67

Ga citrate is the oldest tracer used to image infection: it has a half-life of 3.26 days and this allows imaging to be obtained until 3–4 days after i.v. injection [6,50]. Despite this useful time span the accuracy of this method is low (78.2%) in comparison with other radionuclide methods (Table 3). The specificity for infection is 81.2% and the positivity of gallium is reported in most lymphoproliferative processes, in solid tumours and in chronic granulomatous flogosis (excluding tuberculosis and sarcoidosis). Most authors recommend the addition of bone scinti-

graphy to increase the accuracy of the results by combining metabolic with infective imaging [94–96]. Despite accumulating in malignant tissues or in areas of bone remodelling, gallium still maintains a primary role in imaging chronic osteomyelitis (secondary osteomyelitis, spinal infections, tubercular infections) [97]. In our meta-analysis, 18F-FDG PET was the most accurate method (91.9%) for the study of bone infections. The sensitivity is 94% and the positive predictive value is 94.2%. The specificity is lower than 99mTc-WBCs (89.1%) but higher than all other imaging methods. So, 18 F-FDG PET appears to be a valuable tool for the assessment of inflammation during follow-up of secondary osteomyelitis and for the study of infections treated by antibiotics [98]. Clinical indications

Primary and haematogenous osteomyelitis must be studied by three-phase bone scintigraphy due to the high sensitivity of this method in differentiating increased metabolic uptake of inflamed bone from that in intact bone. Haematogenous osteomyelitis is typically multifocal and a whole-body study is essential. Red bone marrow occurs widely in the bones of all children and most adolescents and the accuracy of all radionuclide tracers (labelled WBCs, granulocytes, MAbs, HIG, microcolloids and gallium) is reduced by the physiological distribution of the tracer in bone marrow. In secondary bone infections, bone scintigraphy has limited usefulness because of its low specificity in non-consolidated fractures or in chronic secondary osteomyelitis, and more specific tracers such as labelled WBCs or monoclonal antibodies are preferred [99–101]. In patients surgically treated with metallic devices or with joint prostheses bone scintigraphy alone is not sufficient. Radiographs and bone cultures are the first approach to the diagnosis of a suspected infection in a painful device to assess the position and the connection between bone and metallic devices [95]. In the same way the usefulness of MRI and CT are questionable because of the metallic artefacts [96]. By means of colloid subtraction or quantitative analysis, WBC scintigraphy is the most accurate investigation to establish if the infection is present or if it persists or it is reduced. Moreover, steroids and antibiotics can reduce the uptake of leucocytes because of inhibition of cytokines and could lead to false negative results because of a reduction of cellular migration in the inflamed areas. In our experience, in chronic infections, and if antibiotics have been given for more than 4 weeks, treatment should be suspended for 2 weeks before WBC scintigraphy in order to increase the sensitivity of the method. In acute infections, and if antibiotics have been given for less than 1 week, it is not necessary to discontinue the treatment.

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In chronic bone infections the migration of granulocytes from vascular space is reduced and the accuracy of WBC scintigraphy is lower than in acute osteomyelitis. Gallium, associated with bone scintigraphy, is an excellent method for studying the persistence or activity of chronic infection in bone and in the diagnosis of osteomyelitis activity. 18

F-FDG PET is a promising technique for diagnosing the activity of a doubtful foci of osteomyelitis but is less specific in distinguishing infection from inflammation: its application should be in the diagnosis of chronic osteomyelitis where the accuracy is higher than gallium. Moreover, this imaging technique is not widely available and is more expensive than all others [102–104].

Infections of the vertebral column Introduction

Spondylo-diskitis is an infection of two or more contiguous vertebrae and the intervertebral disks, occasionally with soft-tissue extension. The posterior extension of infection can be delayed; in an epidural, subdural abscess, in meningitis, the anterior and/or lateral extension in paravertebral, retropharyngeal, mediastinal and retroperitoneal abscesses. In clinical practice spondylodiskitis is divided into primary or secondary. The secondary form is principally of iatrogenic aetiology (post-surgical operation). The course of the disease depends on early diagnosis and early introduction of effective antibiotic therapy [105]. The diagnostic imaging modality for detecting spondylodiskitis is often radiological. MRI can easily detect primary spondylo-diskitis [106]. Instead, in the presence of secondary spondylo-diskitis, MRI may present some limitations due to aspecific signal characteristics [107], and the lack of fat suppression sequences [108] particularly in the immediate post-operative period, when onset of the disease is common. The determination of a microbiological diagnosis by using a CT-guided biopsy is not routinely performed and presents a specificity of 100% and variable sensitivity from 58 to 91% [109–113]. The conventional radiopharmaceutical for infectious disease (labelled leukocytes) can fail in the detection of vertebral osteomyelitis so 18F-FDG PET or combinations of 99mTc-diphosphonate bone scintigraphy and 67Ga citrate scintigraphy are used to complement MRI [97,114–116]. Meta-analysis

Thirty papers published between 1984 and 2004 have been evaluated [96,105–134]. Some original articles and reviews report higher sensitivity and specificity of

scintigraphic imaging compared to radiological imaging. Nevertheless, some observations evaluate data from unselected disease populations [135–137]. One paper incorrectly describes as false negative the presence of a cold area in relation to an infectious vertebral body detected by 99mTc-WBC scintigraphy [138]. A ‘case report’ suggests that only nuclear medicine imaging is useful in diagnosing spondylo-diskitis [139]. 18F-FDG seems to be a good radiopharmaceutical showing high sensitivity (100%) and specificity of 88% [140,141]. From a literature analysis we note that the sensitivity of infection tracers used in the study of spondylo-diskitis varies from 63 to 100%, the specificity varies from 36 to 100%, and the accuracy from 62 to 90% (Table 4). In the table we report only the results of single infection tracer; two papers presenting data obtained by the combination of two different radiopharmaceuticals (99mTc-MDP and 67 Ga) show higher values of both sensitivity and specificity of the tracers [97,114]. Is important to mention that there are some papers on experimental tracers not included in the table but which have already been published in human trials (111In-biotin [142,143] and 99mTc-ciprofloxacin [144,145]). These tracers that seem to present higher values of specificity compared to conventional radiopharmaceuticals were not considered. Clinical indications

Nuclear medicine procedures are necessary in the diagnosis of secondary spondylo-diskitis, especially in post-surgical forms where sensitivity and specificity of radiological imaging maintain an important decrement. A suspicious primary spondylo-diskitis represents an additional indication in the presence of doubtful radiological imaging (MRI and/or CT). Conclusions

From the literature meta-analysis we conclude that in suspected spondylo-diskitis the use of labelled leukocytes is not recommended because of their usual inability to reach the site of vertebral infection and because of their lack of utility in the follow-up pathology. The first-choice scintigraphic procedure is 18F-FDG PET. The presence of 18F-FDG uptake in the vertebral body will be diagnostic for the presence of infection. Nevertheless, clinical interpretation of any 18F-FDG accumulation is essential because the radiopharmaceutical lacks specificity. The combination of bone and 67Ga scintigraphy represents a valid alternative for revealing the presence of an infectious vertebral process. If the uptake of 67Ga is higher than the uptake of 99mTc-disphosphonate the scintigraphy will be positive for infection whereas if the 67Ga uptake is less than 99mTc-disphosphonate the

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Nuclear medicine imaging of bone infections Prandini et al. 639

Table 4

Infections of the vertebral column: results from meta-analysis of data published between 1984 and 2004

Technique 67

Ga Tc-MoAbs In-WBCs 99m Tc-WBCs 99m Tc-HDP 18 F-FDG PET X-rays CT MRI Total 99m 111

Patients

Sensitivity (%)

Specificity (%)

Patients

Accuracy

Patients

PPV

NPV

223 22 163 26 324 147 115 53 300 1373

86.3 91.8 83.8 63.4 81.4 99.9 64.2 82.4 81.8

35.8 84.4 54.9 100 40.7 87.9 37.8 12.8 57

64 22 145 26 197 104 69 17 160 804

88.5 88.5 65.5 80.1 69.3 90 78.2 82.4 78.8

4 NA 123 NA 121 57 28 17 28 378

66.6 NA 63.5 NA 67.7 NA 95.4 80 95.6

100 NA 86.7 NA 60.6 100 66.6 66.7 80

Abbreviations as in the footnote to Table 3. Table 5

Sternal wound infection: results from a meta-analysis of data published between 1982 and 2004

Technique 99m

Tc-WBCs 111 In-WBCs 99m Tc-MoAbs 67 Ga 99m Tc bone scan + 111In-WBCs CT Aspiration Total

Patients 74 369 23 724 32 24 24 1270

Sensitivity (%) Specificity (%) 100 83.9 87.0 74.3 84.0 66.7 100.0

88.4 67.3 95.0 94.0 100 71.4 92.3

Patients

Accuracy

Patients

PPV

NPV

50 369 23 0 32 24 24 522

91.8 75.3 93.0 NA 42.9 54.0 69.2

74 24 0 60 32 24 24 238

82.2 100 NA 93.7 100 50.0 85.7

60.8 94.7 NA 91.0 63.6 83.3 100

Abbreviations as in the footnote to Table 3.

scintigraphy will be positive for bone fracture or other bone pathology.

Sternal wound infections Introduction

Cardiothoracic surgery for aortocoronary by-pass is the most common surgical technique in western Europe. Sternal wound infection after surgical approach is a potential life-threatening complication (1–3%) in openheart operations that requires prompt diagnosis for optimal treatment. Sternal wound infection may be superficial or deep [146]. The extent of superficial infection involves skin and subcutaneous tissues. Deep infection may involve the sternum and deeper tissues such as peri-sternal and retrosternal spaces. The differential diagnosis is crucial for a correct therapeutic approach since superficial infection has a good prognosis with prompt and aggressive antibiotic therapy. On the other hand, in cases of deep infection, the prognosis may be unfavourable with increased morbidity and the percentage of death for mediastinitis and osteomyelitis may be 50–70% [147]. The clinical suspicion of infection is based on clinical examination by the presence of erythema, oedema, purulent drainage, local pain and general signs such as fever, tachycardia and leukocytosis. In some patients, correct diagnosis between superficial and deep infection is distinguished by positive cultures of deep sternal wound aspirates. However, in some patients deep infection may be occult with low fever and normal sternal wound. In these cases culture from aspirates may be doubtful or negative [146].

For this purpose it is necessary for the use of specific diagnostic techniques able to (1) visualize and localize the infective process and (2) evaluate the extent of infection on deep tissues. The results obtained from these tests are therefore used for therapy decision making: antibiotic therapy or surgical revision of wound. The role of nuclear medicine

The diagnostic modalities available to study sternal wound infection and their applications are listed in Table 5. The ‘gold standard’ for distinguishing superficial from deep infection is microbiological culture of deep sternal wound aspirate. But diagnosis may be difficult in the early post-operative stage due to general inflammatory reaction after operative trauma and extra-corporeal circulation, which includes fever, leukocytosis and elevated C-reactive protein. Patients with signs of sternal wound infection or sternal dehiscence are rapidly brought to operation, but those with unclear clinical presentation present a diagnostic dilemma. The first method of ‘imaging’ that is utilized has been plain film radiography of the thorax but it cannot aid diagnosis of sternal wound soft-tissue infection [148]. Radiography visualizes bone disruption, focal osteopenia and periosteal reaction, usually associated with enlargement of retrosternal tissue. However, its value is limited in differentiating between post-operative complications, haemorrhage, oedema and mediastinitis. The X-ray is normal in the early course of infection and when osteomyelitis is evident the risk of mediastinitis is greatly increased [149]. Therefore, in some patients the

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640 Nuclear Medicine Communications 2006, Vol 27 No 8

presence of fluid and gas under the wound for a few days after a surgical procedure could be normal [150]. CT is able to demonstrate adjacent soft-tissue changes and gas, which are non-specific signs of infection, and anatomical details such as subtle erosions, reactive periosteal new bone formation and sharply marginated sclerosis [151]. However, this method is characterized by low sensitivity because signs of infection are clearly evident in the late stage. In addition, CT has a low specificity because the anterior mediastinum is generally abnormal after sternotomy and deeper sternal wound infections are difficult to diagnose [152]. By contrast, ultrasonography is useful in detecting superficial infection, in particular the extent to soft tissues. Several different radiopharmaceuticals have been used to detect sternal wound infections: 67Ga, 99mTc-methylene diphosphonate (99mTc-MDP), 111In-WBCs, 99mTc-HMPAOWBCs and 99mTc-MoAb anti-granulocyte antibodies. However, these techniques are often not included in published diagnostic flow charts and diagnostic guidelines from clinical societies, even though their utility has been shown in experimental studies. Meta-analysis

We evaluated 11 original papers and two reviews (published from 1982 to 2004) on imaging of sternal wound infections with 67Ga, bone scintigraphy with 99m Tc-MDP, WBCs labelled with 111In oxine or 99mTcHMPAO and 99mTc-MoAb anti-granulocytes (results shown in Table 5) [147–159]. From different papers the emerging problem was to identify a method able to detect the extent of infection as superficial or deep. The 67Ga scan showed a sensitivity from 70 to 93% (weighted mean, 74%) and specificity from 93 to 100% (weighted mean, 94%). These values depend on the pretest likelihood of patients studied as described by Salit et al. [157]. In particular, if the clinical pre-test likelihood of sternal osteomyelitis is 30%, then the gallium scan will have a 90% positive predictive value and a 93% negative predictive value. The usefulness of a gallium scan is doubtful in patients with a high suspicion of underlying sternal bone osteomyelitis or poorly decisive for diagnosis in borderline cases [158]. The accuracy of a gallium scan is increased with SPECT images [159]. However, this method is not able to distinguish from osteomyelitis and cellulites, as demonstrated from meta-analysis results. In the last decade WBCs labelled with indium or technetium have been used as a radiopharmaceutical. Studies performed with 111In-WBCs showed a sensitivity of 83.9%, a specificity of 67.3% and an accuracy of 75.3% in 369 patients. More recently, WBCs were labelled with

technetium with better results in detecting foci of infection outlined from values of sensitivity (100%), specificity (88.4%) and accuracy (91.4%) calculated in 50 patients. As in the meta-analysis reported in Table 3, 99m Tc-WBCs appear more effective than 111In-WBCs in terms of sensitivity, specificity and accuracy. This difference between 111In and 99mTc-WBCs may be explained by the lower activity used in indium labelling in comparison with technetium labelling that cannot consent SPECT imaging for example. In addition, from the literature analysis we observed a high variability in the sensitivity and specificity for WBC scans amongst various papers. On the one hand this can be explained by the different acquisition times chosen in different studies, and on the other hand by the presence or not of antibiotic therapy at the time of the scan [146]. Both aspects required standardization. Overall, the labelled leukocyte scan is the most useful technique to differentiate superficial from deep wound infection being able to detect the involvement of bone and retrosternal space. We found very few papers that dealt with other radiopharmaceuticals and this did not allow us to reliably calculate sensitivity and specificity. In particular, monoclonal anti-granulocytes antibody labelled with technetium has been described in only one paper and demonstrated high sensitivity, specificity and accuracy (87%, 95% and 93%, respectively). However, because of high uptake in bone marrow (55% at 4 h and 40% at 20 h), this method can be used only in selected patients, especially with granulocytopenia. As far as a bone scan with 99mTc-MDP is concerned, it is well known that this radiopharmaceutical has a very high sensitivity and low specificity. For imaging sternal wound infection, as demonstrated by Bessette et al. [150], the bone scan is useful only in addition to other methods. Finally, it must be mentioned that in many papers it was impossible to obtain data regarding overall sensitivity and specificity of the technique used because of the absence of a reference ‘gold standard’ for proving infection. Clinical indications for performing white blood cell scintigraphy

Evaluation of infected complications may occur after sternotomy in patients with symptoms and signs of infections such as leukocytosis, an increase of ESR and CRP and doubtful cultures of aspirates. In the follow-up, for the management of patients after specific therapies, SPET images are not necessary although could be helpful in some doubtful case. Early (3 h) and late (24 h) acquisition of planar images

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Nuclear medicine imaging of bone infections Prandini et al. 641

(with acquisition time corrected for Tc decay) are essential.

2

3

Conclusions

Results of this meta-analysis showed that 99mTcHMPAO-WBC scintigraphy is the most reliable imaging modality for differential diagnosis between superficial and deep sternal wound infection. A 99mTc-WBC scan overcomes the limitations of cultures from sternal wounds, a method that is unable to detect infection when the process involves bone tissue. Results obtained from scintigraphy allow adequate management of patients compared with other radiopharmaceuticals, radiological imaging and microbiological studies. Technetium-labelled WBCs is preferable for the shorter physical half-life, lower cost, higher image resolution and better dosimetry compared with indium-labelled WBCs. However, 111In/99mTc-WBCs has a lower sensitivity than microbiological culture of aspirates for the diagnosis of superficial sternal wound infection. As an alternative to WBCs, it is possible to use 99mTc-labelled anti-granulocyte MoAbs, particularly in patients with granulocytopenia, although its diagnostic accuracy needs to be confirmed by other studies.

Acknowledgements This work is part of a large multicentric study conducted by the Italian Study Group on Inflammation–Infection Imaging by the Italian Society of Nuclear Medicine (AIMN) co-ordinated by Dr Alberto Signore. Members of the group are: Marco Agnolucci, Alessio Annovazzi, Giorgio Ascoli, Carla Augeri, Bruno Bagni, Marilena Bello`, Sergio Bissoli, Nicola Boccuni, Sergio Boemi, Paolo Braggio, Luca Burroni, Dario Cantalupi, Gabriela Capriotti, Giuseppe Cascini, Marco Chianelli, Arturo Chiti, Micaela D’Alberto, Diego De Palma, Giovanni D’Errico, Narcisa De Vincentis, Salvatore di Rosa, Paola Erba, Antonio Ferrarese, Guido Ferretti. Chiara Gallini, Elena Lazzeri, Lorenzo Maffioli, Giulia Manfredini, Rita Mannino, Luigi Mansi, Giuliano Mariani, Mario Marinelli, Pietro Marinelli, Luigi Martino, Federica Matteucci, Marino Mele, Angelo Mita, Monica Mori, Maria Gemma Parisella, Valentina Picardi, Carlo Poti, Michele Povolata, Napoleone Prandini, Pierfrancesco Rambaldi, Brunella Rossi, Domenico Rubello, Vittoria Rufini, Orazio Schillaci, Alberto Signore, Alberto Spina, Luca Tagliabue, Maria Cristina Tappa, Daniela Turrin, Venanzio Valenza, Anna Viglietti and Alberto Vignati. Our gratitude and acknowledgements extend to all the members for their collaboration and helpful discussions in the preparation and progress of this study.

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Original article

Osteomyelitis: clinical update for practical guidelines Ercole Conciaa, Napoleone Prandinib, Leo Massaric, Franco Ghisellinid, Vincenzo Consolie,*, Francesco Menichettif and Elena Lazzerig Bone infections represent a diagnostic or therapeutic challenge for the infectivologist, orthopaedic surgeon, radiologist and nuclear medicine physician. Staphylococcus aureus is the major bacterium responsible for bone infections although Mycobacterium tuberculosis is emerging as an infectious agent in Italy because of immigration from Africa and Asia. Osteomyelitis requires long and expensive antibiotic treatment, including rifampicin administered parenterally for several weeks and the use of antimicrobial-impregnated cement in prosthesis substitution. Sometimes it is necessary to carry out surgical debridement of a necrotic bone or the consolidation of compromised bones and joint prosthesis implants. Radiographs and bone cultures are mainstays for the diagnosis of bone infections but are often useless in the lengthy management of these patients. Diagnosis of skeletal infections still includes conventional radiography but magnetic resonance imaging is essential in haematogenous and spinal infections. Bone scans are still useful in acute osteomyelitis whereas scintigraphy using labelled white blood cells is preferred in infections of peripheral bone segments or joint prosthesis. In the axial skeleton a combination of an agent for detecting inflammation (67Ga citrate) and a metabolic agent (99mTc-methylene diphosphonate) enables an infection and an area of increased metabolic activity to be distinguished.

[18F]Fluorodeoxyglucose positron emission tomography, where available, has a significant impact in the study of infections using radionuclides: high-resolution tomographic images represent an effective alternative to gallium in the assessment of inflammation of spine lesions but a comparison with morphological examinations (computed tomography or magnetic resonance imaging) is c 2006 essential. Nucl Med Commun 27:645–660 Lippincott Williams & Wilkins.

Introduction

results. The identification of the causative microorganisms is essential for specific antibiotic treatment but evidence from swabs of ulcers or fistulae is often misleading. Infections of joint prostheses, where bio film microorganisms are involved, should be treated with a combination of antibiotics, which must include rifampicin.

Bone is normally highly resistant to infection, which can occur after large inoculums, trauma or in the presence of metal hardware or in the case of immunocompromised hosts. Infection associated with prosthetic joints is typically caused by microorganisms growing in bio film into organized communities. Bio film protects bacteria from antimicrobial agents and host immune responses. Staphylococcus aureus is the major cause of bone infections: microorganisms adhere to bone and to devices surgically implanted by expressing bone matrix receptors and a phenotypic resistance to antimicrobial treatment [1]. Mycobacterium tuberculosis is an emerging agent of vertebral column infections in Italy as a consequence of immigration from Africa and Asia. Early and specific treatment of osteomyelitis, before extensive bone destruction or necrosis, produces the best *

In memory of Professor Vincenzo Consoli–March 2006.

Nuclear Medicine Communications 2006, 27:645–660 Keywords: osteomyelitis, spondylodiskitis, prosthesis-related infections, radionuclide imaging, antibiotics, antitubercular, arthroplasty, replacement a Istituto di Malattie Infettive e Tropicali, Universita` di Verona, Italy, bStruttura Complessa di Medicina Nucleare, Azienda Ospedaliero-Universitaria, Ferrara, Italy, cClinica Ortopedica e Traumatologica, Universita` di Ferrara, Italy, dDivisione Ortopedica Ospedale Maggiore, Novara, Italy, eClinica Ortopedica 3, Azienda Ospedaliero-Universitaria, Pisa, Italy, fMalattie Infettive and gIstituto di Medicina Nucleare, Universita` degli Studi di Pisa, Italy.

Correspondence to Dr Napoleone Prandini, Struttura Complessa di Medicina Nucleare, Azienda Ospedaliero Universitaria, Corso Giovecca 203, 44100 Ferrara, Italy. Tel: + 39 0532 236387; fax: + 39 0532 237553; e-mail: [email protected] Received 22 June 2005 Accepted 5 May 2006

Surgery is usually unnecessary in acute haematogenous osteomyelitis and in diskitis but a combined antimicrobial and surgical approach should be common after injury with an open fracture or in infections associated with joint prostheses. Surgical treatments include debridement with retention of the prosthesis, or two-stage exchange with re-implantation of the new prosthesis delayed for a variable period of time. The use of antimicrobialimpregnated cement is suggested to correct length and allows partial joint mobility. The diagnosis of skeletal infections includes a variety of imaging methods, but conventional radiography is still

c 2006 Lippincott Williams & Wilkins 0143-3636

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necessary at presentation of acute osteomyelitis. It is of less importance during follow-up or in secondary and chronic infections. Ultrasonography and computed tomography (CT) are very useful for guiding needle biopsies in closed infections of either soft tissues or bones. CT and magnetic resonance imaging (MRI) have excellent resolution and can reveal oedema and any periosteal reaction or soft-tissue involvement: MRI is the preferred diagnostic imaging method for spinal osteomyelitis, but is not suitable for all patients and has certain limitations in the presence of metallic implants [2]. Nuclear medicine imaging procedures to evaluate osteomyelitis include three-phase bone scans, the use of leukocytes (white blood cells, WBCs) labelled with either 99m Tc-hexamethylpropylene amine oxime (99mTcHMPAO) or 111In-oxime, and the use of 2-[18F]fluoro2-deoxy-D-glucose (18F-FDG) positron emission tomography (PET) and 67Ga citrate [3–5]. The three-phase bone scan is the test of choice in evaluating acute osteomyelitis and doubtful diskitis but the specificity of this method falls in secondary osteomyelitis. The finding of increased metabolic activity in osteomyelitis is indistinguishable from post-traumatic injury or following surgery or cancer. WBCs accumulate at sites of infection and in bone marrow. The combination of the 111In-oxime WBC scan with a 99mTc-sulfur colloid bone marrow scan is considered the ‘gold standard’ method for the study of infections of hip prostheses but can be also helpful in the study of peripheral bone segments. Three-phase 99mTcHMPAO WBC scintigraphy is widely used in Italy as a helpful alternative to the combined method of WBC/ sulfur colloid: a WBC scan is less sensitive for imaging those bones where red marrow is present (i.e., the axial skeleton and spine). In these cases, by combining 67Ga citrate and bone scintigraphy it is possible to distinguish the activity of secondary vertebral osteomyelitis from other causes of increased bone metabolism. 18F-FDG PET, where available, has a significant impact in the radionuclide study of infections: the high-resolution tomographic images represent an effective alternative to gallium in the assessment of inflammation of spinal lesions but a comparison with morphological examinations (CT or MRI) is essential.

The pathophysiology of osteomyelitis Osteomyelitis is an inflammatory suppurative process of the bone marrow in which both the endosteum and periosteum participate actively whereas the trabeculae and Haversian system participate passively with necrosis and osteolysis. Bone is a tissue that is resistant to bacterial colonization and, in effect, to cause a bone infection, other negative events must occur such as traumas, the presence of foreign bodies, prostheses or an

inoculation of aggressive bacteria or other bacteria that generally have characteristics that inherently favour implants; characteristics of adherence, for example [6]. Classification of osteomyelitis

The classification by Waldvogel et al. [7] is rather old, but still topical, and is based on the genesis of osteomyelitis (haematogenous or secondary) and on the modality of onset. Acute haematogenous osteomyelitis symptoms last no more than 10 days. The chronic form is by far the more frequent and includes all the remaining cases (Table 1). The classification by Cierny et al. [8] is more recent and relevant as it proposes the anatomical and histological subdivision of osteomyelitis (medullary, superficial, located or diffused) and introduces the important concept of host immunocompetence, which is highly relevant regarding the onset and diffusion of infections (Table 2). Haematogenous osteomyelitis

Haematogenous osteomyelitis affects children in 85% of all cases and it often originates from unknown primary foci (nasopharynx) via direct inoculation but can also be contiguous. The most affected group varies from 2 to 5 years of age and the elective location is the lower limbs (femur 27%, tibia 22%). The incidence of osteomyelitis is between 1:1000 and 1:20 000 and mortality reached 50% in the pre-antibiotic era. Nowadays, mortality through osteomyelitis is almost zero. The primary focus is not Classification of osteomyelitis according to Waldvogel et al. [7] (modified)

Table 1

K Haematogenous osteomyelitis K Secondary

Infection next to focal point With vascular impairment Without vascular impairment K Acute Suppuration and oedema, vasal congestion and small vessel thrombosis Vascularization is compromised if nearby soft tissue becomes involved If haematic and periosteal flow rates are reduced, large areas of dead or isolated tissue form K Chronic A knot of necrotic bone tissue or scar tissue surrounded by ischaemic soft tissue

Table 2

Classification of osteomyelitis according to Cierny et al.

[8] Anatomical type I II III IV Physiological class A–Host B–Host C–Host

Clinical stage Type + class Example

Medullary osteomyelitis Superficial osteomyelitis Localized osteomyelitis Diffuse osteomyelitis Good immune system and delivery Compromised locally (BL) or systemically (BS) Requires suppressive or no treatment; minimal disability; treatment worse than disease; not a surgical candidate = Clinical stage Stage IV BS osteomyelitis = a diffuse lesion in a systemically compromised host

BL: local host compromised; BS: sistemic host compromised.

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Osteomyelitis: clinical update for practical guidelines Concia et al. 647

found in 70% of cases and even when present, may be far from clear. It can be from a simple boil, an infected ulcer, a urinary tract infection or other soft-tissue focal infection. In adults, the infection locations are often in the vertebrae. S. aureus is the most frequent microorganism but in the 2–3 year age group, streptococci such as Haemophilus influenzae can predominate. In haematogenous osteomyelitis, Gram-negative microorganisms (Escherichia coli, Klebsiella, Salmonella and Proteus) are found, and in some immunocompromised patients (e.g., drug addicts) Pseudomonas aeruginosa and, occasionally, Candida aspergillus [9,10]. Secondary osteomyelitis

Secondary osteomyelitis includes post-traumatic osteomyelitis after compound fractures involving a mass of bacteria, the nature of which depends on the environment in which contamination occurred. Post-operative osteomyelitis partly includes the above for open-fracture reduction but also other operations on bones such as the cranium, vertebral column and sternum, and for prosthesis implants and ozone therapy. Contiguous osteomyelitis

Contiguous osteomyelitis includes, besides the classical diabetic foot, osteomyelitis of the heel following unrecognized traumas (puncture and scratch injuries) and osteomyelitis of the jaw following radiotherapy for neoplasms of the head and the neck. S. aureus is the most frequent microorganism of postsurgical osteomyelitis and it is multi-resistant to antibiotics: in Italy, the global incidence of methicillinresistant staphylococci exceeds the 50% of the isolated bacteria. Nevertheless, the surgical prophylaxis guidelines still foresee the use of cephazolin, which is not effective over two of these infections. It should therefore be necessary for each hospital to assess its own incidence of methicillin-resistant infections, using glycopeptides in surgical prophylaxis only when the incidence of these microorganisms exceeds 50%. Hence, the indiscriminate use of teicoplanin and vancomycin can be avoided. Periprosthetic osteomyelitis

Periprosthetic osteomyelitis is the new osteomyelitis. Nowadays, the elderly insist on more mobility and an ever-increasing number of prostheses are being fitted; at present there are more than 600 000 in the USA (Table 3). Post-operative infections of prosthetic joints have decreased from 5.9% ( ± 1.8) in the 1970s to 1.2 ( ± 0.5) since 2000 [11]. In Italy, between 45 000 and 50 000 prosthetic hip joints and between 9000 and 10 000 knees are fitted each year. Although the incidence of hip prosthesis infections in 1999 was approximately 1.5% per year for new implants, this incidence tripled in reimplantation cases even if the re-implant was not

Table 3

Incidence of device-associated infections in the United

States Device Joint prostheses Fracture fixators Dental implants

Table 4

Usage/year

Infection risk (%)

600 000 2 million 1 million

1–3 5–10 5–10

Classification of prosthesis infections

Onset of infection Precocious ( < 1 month)

Delayed (2–12 months) Late ( > 12 months)

Infecting organism Staphylococcus aureus Staphylococcus coagulase-negative Aerobic Gram-negative bacteria Staphylococcus coagulase-negative Skin bacteria (S. epidermidis) Staphylococcus aureus (methicillin-resistant) Staphylococcus coagulase-negative Skin bacteria (S. epidermidis) Anaerobes Streptococcus species Escherichia coli

required for a previous infection of the prosthesis. We can therefore estimate that 10–15 significant infections will develop in each 100 prosthetic operations over a 10year period, which is the average life span of a prosthesis. The orthopaedic surgeon’s worst enemy is still S. aureus, even if other clinically significant bacteria such as P. aeruginosa and S. epidermidis appear [12,13]. The early forms of periprosthetic osteomyelitis develop in the first month, while the delayed form arises in the first year following surgery and the late form can arise many months or years after the initial event. The symptoms are pain, fever, oedema and fistula. Table 4 gives a classification of prosthesis infections. In precocious prosthetic hip infections, infection tends to set in within a month of surgery and stems from bacterial activity during surgery, or in the immediate postoperative period, around the operation site, then develops in the periprosthetic soft tissues with little involvement of the bone–prosthesis interface. This means that the infection is located within the surgical access area and to identify it we can use a method that thoroughly studies soft tissues. If diagnosed early, these infections have a good prognosis and save the use of another implant. In delayed or late infections that have occurred from 2 to 15 months after an operation, the rejection of, or reaction to, a foreign body (the prosthesis) can offer an excellent point of attack for bacteria that lodge in the body. During the course of 1 day, there can be episodes of bacteraemia in the blood which are easily overcome by healthy subjects. In patients with prosthesis, however, if the bacteraemia is severe, perhaps because the patient has a

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648 Nuclear Medicine Communications 2006, Vol 27 No 8

dental abscess or a chronic urinary tract infection, osteomyelitis develops on the prosthesis. This is also due to the presence of a reactive phenomenon to the use of polyethylene in prostheses. This results in a subsequent reaction with the appearance of periprosthetic osteolysis or of less vascularized centres and the possible occurrence of circulating bacterial colonies. In this infection, the seat is typically the prosthesis/bone interface, with the soft tissue involved secondarily. In this case, the diagnostic approach must be to use a method that allows this to be carefully studied. Prognosis in these cases is rather poor and the nuclear physician and radiologist must therefore be fully aware of the patient’s clinical background and have all the relevant laboratory data [14–16]. Infections of the vertebral column

Infections of the vertebral column are haematogenous infections but primarily strike adults over 50 years of age. Drug addicts are an exception and are particularly exposed to infections of the vertebral column just like diabetics and dialysis patients. The infections present as spondylitis or vertebral osteomyelitis, diskitis and secondary diskitis where the infection shows an effect either by itself or in association with adjacent vertebrae and intervertebral disks. Diskitis is a specific or non-specific bacterial infectious process (very occasionally mycotic) of two or more adjoining vertebrae and of the respective intervertebral disk. The infection sometimes spreads to the surrounding soft tissue. Backwards extension of the infection can result in an epidural or subdural abscess or in meningitis, whereas forward and/or lateral extension can result in para vertebral, retropharyngeal, mediastinal or retroperitoneal abscesses [6,17,18]. Infections of the vertebral column can be divided into spontaneous and iatrogenic forms, these last due to invasive manoeuvres or to surgical interventions. The spontaneous forms, which are supported by an arterial haematogenous dissemination in almost all cases, are unspecific bacterial or mycotic and they represent the 2– 4% of all vertebral osteomyelitis. The source of infection can be presented both from venous inoculation and from different kinds of sources (e.g., genitourinary apparatus, cutaneous or subcutaneous, respiratory, dental) or from vascular devices. In 24–37% of cases, the source of infection remains unknown. The most susceptible vertebrae are the lumbar (45%), followed by the dorsal, above all the inferior, (35%) and the cervical (20%). A greater incidence of the cervical forms has been noticed among the drug addicts. S. aureus is the most frequently isolated agent (55–85% of cases), followed by coagulasenegative staphylococci, enterobacterias (Salmonella spp, E. coli, Klebsiella spp, Serratia spp), P. aeruginosa (frequent among drug addicts) and Candida (among drug addicts and in infections of vascular devices) [19,20].

Spontaneous specific diskitis

Spontaneous specific diskitis is represented by vertebral tuberculosis or Pott’s disease supported by a haematogenous dissemination originating from the lung or from other lymph nodes or from genitourinary infections, often within disseminated tuberculosis. Vertebral infections are increasing, especially among immigrants from Romania, India, Sri Lanka and Africa, as well as in patients over 80 years of age who have compromised immune systems. The tubercular infection begins from the anterior part of the vertebral body and usually involves the subcondral region, spreading then to the cortical region and to the adjacent disk. It can result in vertebral abscesses involving the ileopsoas muscles. The classic symptomatology of spontaneous specific diskitis is represented by ‘back pain’ with accentuation of the painful symptomatology in the passage from the clino to the orthostatic position and from the seated to the erect position. Because of the insidious symptomatology, whose progress can last weeks or months, the moment of the diagnosis it is often delayed (it may be between 3 weeks and 3 months). The fever, usually around 37.51C, is present in 50% of patients. Laboratory data show a modest leukocytosis in 50% of subjects while erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) are usually increased [19,21]. Iatrogenic diskitis

Iatrogenic diskitis follows direct inoculation of microorganisms after spinal anaesthesia, chemonucleolysis and local infiltrations with analgesic purpose. Above all, it follows surgical interventions for slipped disk, spondylolysis and spondylolisthesis. Also, in such cases, staphylococci play the principal aetiological role, with the prevalence of coagulase-negative strains. The painful symptomatology, similar to that in spontaneous diskitis, involves the site of the intervention and can sometimes be associated with subcutaneous infections noticeable during physical examination. The onset of the painful symptomatology can vary from a few days to 2–3 months from the invasive or surgical manoeuvre. The fever is not constant, while an increase of ESR and CRP is typical, with or without variations in granulocytes [22].

Assessment of the disease Symptoms of osteomyelitis

The symptoms of osteomyelitis are variable: in the typical form, fever, pain, motor limitations and local inflammation or septicaemia, are found, as is usual among infectious diseases. A bone infection leads to tissue destruction, which can result in the functional loss of the involved bone and the surrounding soft tissues [23]. Results of laboratory tests (CRP, ESR, neutrophilia) are used in the diagnosis of the infection, but a precise diagnosis and effective therapy cannot be formulated unless the microorganism is isolated and the culture examined [6,7,17,18].

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Osteomyelitis: clinical update for practical guidelines Concia et al. 649

Onset of infection is variable: a thick inoculation of virulent bacteria generates an acute infection but a thick contamination in a guest immunocompetent patient produces a faint infection. In these cases, the appearance is delayed, even after years: there are often fewer virulent bacteria but with adhesive abilities. A third possibility is that there is a low intra-operating contamination, with low virulence of the organism, and it can be easy to maintain under controlled conditions. Possibly, it also exists in a haematogenous manner, it is not frequent and is a consequence of oral, endoscopic or urinary surgical interventions. The infection should be prevented by effective antibiotic prophylaxis before the intervention. Radiological investigation

In the initial phases of the haematogenous osteomyelitis, we find a medullar inflammation characterized by the hyperaemia, the oedema, the leukocytes infiltration and the purulent transformation. In this phase the radiological diagnosis is limited because there are no definite signs of infection in the first 2 weeks after onset. There are some specific signs such as soft-tissue oedema and the disappearance of the fascial levels, which are easily found by nuclear medicine techniques and MRI [24,25]. The abscess of Brodie is a unique focal infection, with chronic evolution, generally located at the proximal metaphysis of the tibia or femur. X-rays show an oval osteolytic area that is better seen by MRI with spin-echo T2-weighted sequences and a heterogeneous signal hyperintensity. The stir sequences of the MRI provoke the suppression of the fat, and they confirm the presence of oedema and the involvement of the soft tissues [26,27]. The infection can spread to the cortical with lifting of the periosteum and interruption of the vascular support. X-ray examination shows osteopenia and a worm-hole aspect of the bone with a meaningful re-absorption up to the osteolysis. The consequence is bone death, which can lead to alterations such as swelling of the surrounding skeletal soft tissues, periosteitis, endosteitis and the disappearance of the medullar canal up to the lamellar osteonecrosis and thickened bone. In the following phase, sequestration with clear demarcation of necrotic bone from healthy bone can be seen; this is due to the phenomenon of osteoclastic demolition. After the sequestrum, a chronic process is followed with the overproduction of bone and therefore a demarcation from the surrounding bone. Conventional X-rays show osteolytic areas of infection delimited by a diffusely and heterogeneously thickened bone. The MRI spin-echo T1-weighted sequences demonstrate sequestration of bone and the activity of the infection. If a patient receives gadolinium, the MRI shows hyperaemia, definite expression of the process activity [28–30]. The CT shows very well the partly corpuscular swelling of the soft tissues around bones and joints.

Radionuclide imaging

Radionuclide imaging is usually employed after the radiological investigations, in the diagnosis of haematogenous osteomyelitis and of soft-tissue infections. The three-phase bone scan with 99mTc-disphosphonates shows an increase of bone perfusion and of the surrounding soft tissues in the dynamic and blood pool images. In delayed images bone uptake appears blurred in the boundaries of healthy bone, and can easily be distinguished from cellulites in which the involvement also includes soft tissues. However, the efficacy of a three-phase bone scan decreases in follow-up or after antibiotic or surgical treatment because the modifications of bone metabolism, and the normalization of scintigraphic images, are very slow. An increased uptake of labelled diphosphonates can persist for months or years after recovery of a bone fracture or after osteomyelitis. In paediatric patients the water content and perfusion supply of bone are increased in comparison with adults. So, septic bone necrosis is not rare and can result in cold areas on methylene diphosphonate bone scintigraphy (Fig. 1). Haematogenous osteomyelitis can involve more bones, which is the reason why a whole-body bone scan is essential in these cases. The diagnosis of bone infections becomes difficult in cases of recently implanted prostheses, in delays of consolidation of exposed fractures and after repeated surgical or therapeutic interventions. The principal problem in osteomyelitis remains the search for infections in a bone with altered structure caused by re-absorption or new apposition processes where there is a loss of specificity of bone scintigraphy, CT and MRI. Radiological methods are also limited by the presence of synthesis materials or by implants or prostheses that provoke important artefacts in both CT and MRI scans. In these contexts the most reliable investigation for verifying the presence of bone infection remains scintigraphy with labelled leucocytes. In this technique, either WBCs or Fig. 1

Bone scan in a right foot osteomyelitis in a 6-year-old child. The cold area corresponding to the navicular bone of the right foot is expressing septic necrosis.

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pure granulocytes labelled with either 111In-oxime or 99m Tc-HMPAO can be used: the choice of cell or radiolabel is not critical. The most important factor influencing the accuracy of this examination is the time required to follow the diapedesis of the granulocytes, prolonging the examination to at least 24 h. In this way it is possible to observe labelled granulocytes migrating from the blood to the soft tissues and concentrating around the prostheses, in the cavities, in fistulas, and, sometimes, in the regional lymph nodes [4,7,16–18]. Moreover, the scintigraphic accuracy of labelled leucocytes is modified by the presence of the red marrow in bones involved in osteomyelitis. In haematogenous osteomyelitis of the axial skeleton or of the proximal appendicular skeleton 30–75% of cold areas are found with WBC scintigraphy. These are areas where there is zero or very low blood flow and are the equivalent of sequestered bone on X-ray examinations. In these cases scintigraphy with leucocytes (or even monoclonal antibodies) labelled in vitro, is useless in assessing the activity of inflammation because cold areas can persist indefinitely, maintaining this situation for years. In young patients the bone marrow is extended in all the skeleton segments up to the distal appendicular ones. Therefore, in paediatric age groups we have, more or less, the same limitations in the use of labelled WBCs that we have in the axial skeleton of adults and labelled WBCs in vitro (or monoclonal antibodies) are often useless. Unspecific radiopharmaceuticals for detecting inflammation, such as 67Ga or 18F-FDG, with an uptake proportional to the vascular permeability or to metabolic activity, should be preferred in secondary bone infections of axial skeleton. In these cases the scintigraphy is ordered only for assessing whether the infection is still active or not after therapeutic approaches (e.g., surgery, antibiotics). Prosthetic joints

Orthopaedic surgeons and clinicians must identify the position and extent of infection when dealing with prosthetic joints. Furthermore, they have to provide accurate information on the prosthesis, taking into account patient history, the clinical possibility of infection, X-ray images, and laboratory data. Most importantly, they must identify the kind of prosthesis with which they are working. Cemented prostheses are custom-made because the cement hardens within 20 min and gives immediate mechanical stability to the prosthesis. However, as the cement hardens it produces an exothermic reaction, which may bring about an endosteal necrosis that affects vascular flow in the bone–cement interface. The polyethylene waste product that builds up over time because of wear and tear of the cotyloid cavity may lead to osteolysis near the cement of the prosthesis showing a

characteristic accumulation of labelled diphosphonate. Non-cemented prostheses, on the other hand, inevitably bring about a reshaping of adjacent tissues that may either erode the bone or cause new bony deposits. This reshaping depends on the material used in the prosthesis, on its design, its primary anchoring, and on whether the prosthesis is coated with osteo-inductive materials. Chrome–cobalt–molybdenum alloys, once commonly used, are very rigid, and produce significant reshaping of the bone, which induces a considerable necrosis in bone tissue due to the accumulation of deposits of both bone and metal. For this reason, titanium alloys are now being used, as they are less rigid and produce fewer deposits and less bone necrosis [31]. Design and primary anchoring are equally important. The so-called distal press-fit involves long prostheses while proximal press-fit and distal filling have completely different osteo-metabolic characteristics. Another key factor is the potential presence of an osteo-inductive coating. The purity and porosity of this coating have an impact on the extent of reshaping that takes place around the prosthesis, and consequently on the degree of diphosphonate uptake at bone scintigraphy. Traditional X-ray reveals specific details of the bone– prosthesis interface but it is of little help if the infection is in soft tissue. Characteristic signs are small and unclear, and conventional X-rays often give negative results. In soft-tissue infection, ultrasonography will provide highly detailed information; for example, in identifying whether an abscess is relative to vascular and nerve bundles. This is essential to plan access, intervention and debridement. If this infection is extensive, it can spread to the abdomen or the pelvis. The limits of the technique are that, in the early phase, it fails to distinguish post-surgical haematoma from septic haematoma, which may affect soft tissues [32]. CT and MRI are excellent techniques but, when dealing with infections near prostheses, lose some of their effectiveness because the quality of the resulting images is severely affected by the presence of the metal hardware. CT can provide information about the movement of the prosthesis, but fails to distinguish between mechanical and infectious loosening. However, CT can play a role if used with WBC scintigraphy, which allows a morphological examination of their location and accumulation. Likewise, CT–PET can reveal the morphology of 18 F-FDG accumulation. By using CT, the surgeon can obtain important information on the extent to which an infection has spread through muscular tissues, while CT fistulography is useful if abdominal tissue is involved. CT allows the biopsy to be guided to the location of infection and an antibiogram, which is essential for curing the infection with specific antibiotics, to be obtained.

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Osteomyelitis: clinical update for practical guidelines Concia et al. 651

With infections in hip prostheses, a three-phase bone scan shows an increase in early perfusion and a delayed metabolic accumulation of diphosphonate all around the prosthesis, marking out its contour. The specificity of bone scintigraphy in hip prostheses ranges from 50 to 70% according to case specific circumstances [33] because the bone scan signal requires months if not years before returning to normality even if the clinical problem has been resolved. In the case of simple instability, on the other hand, there is no early increase in perfusion while the delayed accumulation of diphosphonate is typically concentrated in the load-bearing points at the top of the acetabulum, in the minor and major trochanter and at the top of the prosthesis stem. A three-phase bone scan has a sensitivity of about 85% in hip prostheses infection [34]. In knee prostheses, the role of three-phase bone scintigraphy is less definite and it is more difficult to differentiate a case of movement from a case of infection [35]. Scintigraphy with WBCs or granulocytes labelled with 111 In-oxime [36] or with 99mTc-HMPAO [37] is the most accurate method for studying bone infections. Leukocytes accumulate by diapedesis in musculoskeletal infections, attracted by chemotaxis, thus the type of white-cell accumulation changes over time. Successive scintigraphies will reveal these movements and migration of labelled cells within the tissues leads to a progressive concentration in the fistula or spaces near the prostheses, in joints, and sometimes in local lymph nodes (Fig. 2). White-cell distribution in the body, however, sharply limits clinical use of WBC scintigraphy. For example, the study of infections of the dorsolumbar area and of the lower ribs is difficult because labelled white cells tend to concentrate in the liver and spleen with higher resulting

activity of the overlapping abdominal and thoracic wall. Furthermore, in 30–75% of cases of infection in the axial skeleton and nearby areas, cold areas are observed with WBC scintigraphy linked to the low flow of cells within the inflamed bone tissue [38,39]. This is equivalent to radiological sequestra, which increase with the duration of the infection, and with repeated antibiotic treatment. In the same way, granulomatous chronic infections (i.e., tubercular osteomyelitis, or mycosis) are characterized by a low percentage of granulocytes and are hardly revealed using labelled WBCs or pure granulocytes [40,41]. Antibiotics and immunosuppressants can reduce the accumulation of white cells because these drugs reduce the diapedesis of granulocytes by reducing the concentration of cytokines in tissues. Their inhibitory action is proportional to the duration and efficacy of antibiotic therapy as well as to the bacterial population causing the infection. Pyogenics, such as S. aureus, bring about the highest white-cell accumulation but the intensity of uptake is also more susceptible to the antibiotic effect. Patients suffering from acute infections should not suspend antibiotic treatment, however, as this would worsen their clinical state. In these cases, scintigraphy should be carried out as soon as possible after the onset of symptoms. In patients with chronic infection or undergoing long-term antibiotic treatment, scintigraphy should be delayed until at least 2 weeks after the end of therapy. In this way, it is possible to determine accurately whether therapy has been successful or if the infective foci remain.

In adults with prosthetic joints and post-traumatic infections, surgical intervention may bring about a

Fig. 2

Infection of a left-hip prosthesis. Three-phase 99mTc-HMPAO white blood cell (WBC) scintigraphy shows increased activity of labelled WBCs in soft tissues of the left thigh in the first images (1 h after i.v. injection), which significantly increases in the images after 4 and 20 h. The path to the hip prosthesis is also well demonstrated.

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652 Nuclear Medicine Communications 2006, Vol 27 No 8

peripheral displacement of the bone marrow towards surrounding spaces, which may then be mistakenly interpreted as septic. To avoid this inconvenience, Palestro et al. [42] suggested comparing 111In-leukocyte scintigraphy with a bone marrow scan (with 99mTc-sulfur colloids). The discrepancy between the two scintigraphic images (leukocytes greater than colloids) very accurately identifies the presence of infection. As an alternative, repeating scintigraphic observations at 1, 4 and 20 h after the re-injection of cells makes it possible to distinguish the invariant accumulation with time (that is, bone marrow) from progressively rising accumulation in osteomyelitis. Comparing the three images, it may be possible to follow the path taken by the white-cell diapedesis from the location of the infection to the fistulas or other periprosthesis spaces [43]. The two methods have equivalent sensitivity at around 95% for peripheral bone tissue with a specificity of 97% [44]. The same figures are lower for axial skeleton and in chronic infections [45].

Management of patients with osteomyelitis A multidisciplinary and structured approach to the management of bone infections is important. Antibiotic therapy must rest on the identification of the pathogen. This allows an accurate antibiogram, which will pinpoint the group of antibiotics to be used in the treatment. Among these, we are then able to choose the antibiotic that can best penetrate bone tissue and which has low toxicity. Staphylococcus is enemy number one. Rifampicin, macrolides and teicoplanin have excellent diffusion profiles, thus whenever possible this group of antibiotics should be favoured. On the other hand, we know that teicoplanin has a good degree of coverage for methicillinresistant S. aureus and of vancomycin-resistant Enteroccoccus, although vancomycin is associated with a higher risk of relapse when compared to teicoplanin. As for quinolones, the paired quinolone–rifampicin has an excellent pharmacokinetic profile and therefore can be a useful combination in treating some forms of staphylococci [46,47]. Aetiological therapy

Aetiological therapy cannot do without rifampicin, which has an optimal intercellular concentration, a very good sensitivity profile for methicillin-resistant staphylococci and when used with teicoplanin offers significant clinical advantages that have been demonstrated both in the laboratory and in practice [48]. Among new drugs, linezolid inhibits bacterial protein synthesis and is very effective on methicillin-resistant staphylococci and on vancomycin-resistant Enteroccoccus. Linezolid is also easily absorbed which is a great advantage when associated with 100% bioavailability and has excellent bone penetration. For the time being, however, there are no agreed protocols for the use of this antibiotic in osteomyelitis

[49,50]. Quinupristin–dalfopristin is sometimes used. This antibiotic also blocks protein synthesis, spreads easily amongst macrophages, and has a good tissue distribution. However, its use in osteomyelitis is advisable only in particularly selected cases [51,52]. Empirical therapy

Empirical therapy has to be resorted to when it is impossible to isolate the root of the infection. In this case local epidemiological factors must be taken into account, together with determining whether the infection has been contracted in hospital or elsewhere. Infections contracted outside hospitals are usually from methicillinsensitive staphylococci, and are frequently polymicrobic infections with the presence of Gram-negative bacteria. In these cases, the clinician should start with an amino penicillin associated with a beta-lactamase inhibitor, on the following antimicrobial associations: rifampicin + quinolone, oxacillin or teicoplanin or clyndamicin + ceftriaxon or cefepime or quinolone [53]. Hospitalcontracted infections, on the other hand, have a high probability of being derived from methicillin-resistant staphylococci, which should be dealt with through a glycopeptide, particularly teicoplanin, together with rifampicin, possibly associated with antibiotics active on Gram-negatives [14,54]. The antibiotics should preferably be administered parenterally for several weeks. This raises the problems of cost, patient cooperation, and morbidity. Most patients will have to remain in hospital because it is not always possible to identify an antibiotic that can be administered parenterally at home, given that dosages are seldom intended for individual home use. Oral therapy has been considered but we do not yet have enough data. For the time being, oral therapy is indicated only in children. Furthermore, oral therapy can only be used with patients whose compliance is certain. Treatment duration is not standardized at present. In any event, it must be based on the type of infection. For instance, haematogenous infections must be dealt with by determining whether bone sequestration is occurring and whether debridement is necessary. Depending on the specific case, therapy for 4–6 weeks may be enough or it may prove necessary to increase it to 6 weeks or longer [55,56]. Often osteomyelitis fails to improve because bacteria have the ability to resist to antibiotics. S. epidermidis sticks to the prosthesis and is enclosed in the bio film: a polymeric matrix acts as a protective mantle to impede phagocytosis and the delivery of the antibiotic [57,58]. The reduced growth of bio film bacteria is responsible for their resistance to many antibiotics that are only active in this phase. Rifampicin acts on the bio film and must therefore always be used with other antibiotics when a prosthesis infection is present. Failure to use rifampicin

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within a month or a few weeks of treatment will allow infection to start again. Treatment of prosthesis infections

Antibiograms in prosthesis infections should be performed, mixing the materials that replicate the impact of the prosthesis itself. For example, ofloxacin affects the Gram-positives of a fluid culture but adding some polystyrene beads to the broth better simulates real conditions. In this case, antibiograms change completely and bio film producing bacteria develop. It follows that the duration of the antibiotic treatment is not standardized. It has to be defined according to infection type: in haematogenous infection, it is necessary to check for bone sequestration. If debridement is called for, therapy for 4–6 weeks can be enough or it may prove necessary to prolong therapy for 6 weeks or more [19]. The causes of pain in hip prostheses can be mechanical or ‘biological’. Mechanical causes are load bearing, excessive periprosthesis re-absorption, possible sinking following the original placement, fractures (macro or micro) in the femur near or under the prosthesis. Biological causes are reactions to polyethylene deposits (in cases of antiseptic movements) and infection (in the case of septic movement). In cases of infection, a two-stage intervention must be accompanied by medical therapy of up to 6 weeks, comprising collecting biotic samples and examining cultures in the removal stage and resorting to antibiograms to fine-tune the antibiotic. In the case of prosthesis infection, different surgical interventions are possible: from simple debridement, to one-stage or two-stage intervention, to outright removal. Different interventions will, of course, demand different therapies. In one-stage interventions, antibiotic therapy must be particularly long, at least 4 weeks before the intervention and up to 16 weeks afterwards. Note that, especially in this type of intervention, resorting to empirical therapy will be simpler than the more rigorous aetiological approach. Therapies lasting less than 4 weeks have been shown to carry a high risk of re-infection; thus, they should be continued for at least 6 weeks. While it is difficult to be certain that the bone has healed, combining clinical, radiological and biochemical, information (such as carefully monitored CRP follow-up) can definitely be of assistance. As for CRP data, it is well to keep in mind that recurrent inflammation episodes may over-ride all other considerations. Use of antibiotic-loaded acrylic cement

In 1970, Buchholz and Engelbrecht [59] introduced antibiotic-loaded acrylic cement to the treatment of prosthesis infections, a technique that has since been used frequently. Simply put, its advantages are higher concentrations of antibiotic in the soft tissues and in the

bone than would be possible by alternative delivery methods, low serum concentration and, consequently, lower toxicity. In our opinion, the technique has benefited from the well-known suggestion, in 1988, by Wilde and Ruth [60] of a space block and multiple stage treatment. The space block has a double advantage: mechanical and biological. The first is in preventing joint head fusion, while maintaining the correct length of muscular structures and reducing post-surgery blood pooling. The second is in assisting the disinfection of localized septic points and maintaining high local concentrations of antibiotic. In addition, the two-stage technique allows for a repeat of surgical cleaning and the use of un-anchored prostheses. The most important problem remains the choice of dosage of the antibiotic associated with the cement. Bactericidal tests performed on various stocks of pathogenic agents have shown the absolute ineffectiveness of some antibiotics. Our conclusion is that for S. aureus and S. epidermidis and Pseudomonas, the most effective bactericide was a combination of vancomycin and imipenem–cilastatin, most likely because their actions were mutually reinforced to the greater porosity of the cement, which leads to a greater release of the antibiotic. Furthermore, two-stage procedures offer better control of the infection because they give the opportunity of introducing a prosthesis without cement, while the onestage intervention forces the use of a cemented prosthesis [61,62]. This treatment requires cooperation between epidemiologists, microbiologists and nuclear physicians. Its high cost may discourage hospital administrations and private nursing homes, however [63].

Diagnosis of patients with suspected osteomyelitis Clinical examination of the patient always allows the detection of functional impairment and laboratory tests can provide bone infection data. Conventional X-ray is the first imaging procedure in the diagnostic flow chart of osteomyelitis: if the results are positive, this is sufficient to begin the appropriate therapy. Haematogenous osteomyelitis

In haematogenous osteomyelitis, a three-phase bone scan can provide a result within 24-48 h after the onset of osteomyelitis symptoms and can be used very profitably in radiologically negative cases or for whole-body studies because haematogenous osteomyelitis is often multifocal. In children and adolescents, pain is the main symptom of osteomyelitis, accompanied by other joint diseases such as arthritis, aseptic necrosis or epiphysiolysis of the hip, which can be quickly identified and distinguished from osteomyelitis by using bone scintigraphy. Finding cold areas on the scintigraphic image of a septic necrosis that complicates haematogenous osteomyelitis is more

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common in children due to their higher bone water content [64]. The interpretation of conventional radiological results becomes complicated where there are prosthetic joints and secondary or post-traumatic osteomyelitis, or where there has been bone re-modelling, previous operations and the presence of synthesis materials or metal support devices. Nevertheless, such tests are carried out to assess the bone condition. If there are any doubts about the Xray results, CT and MRI provide a more detailed morphological study and offer the orthopaedic surgeon a guide regarding surgical drainage of abscesses or the debridement of necrotic bone. The presence of synthesis materials, however, hinders or limits the detection of persistent infection by CT or MRI imaging. In these cases, where a surgical approach is difficult, it is necessary to establish whether the secondary osteomyelitis is truly cured or whether it is merely present following consolidation surgery. Scintigraphy with WBCs, in association with the appropriate suspension of antibiotic therapy, is the most accurate way of determining the persistence of an infection [65].

Post-traumatic infections

In our experience, post-traumatic infections are studied with laboratory tests and conventional radiography. If the results of these tests all suggest infection, medical therapy is initiated, guided by a culture test and an antibiogram and possibly by pulsed magnetic therapy especially in the presence of late consolidation or a pseudoarthrosis (Fig. 3). An alternative is to begin antibiotic medication followed by surgery, a clinical follow-up, radiography and scintigraphy with radiolabelled leucocytes to assess recovery. If the conventional radiography produces doubtful or negative results, and there are positive clinical and laboratory indications of inflammation, we carry out scintigraphy with radiolabelled leucocytes, which may give positive, doubtful or negative results. If positive, the next step is to prescribe antibiotic therapy for 2–4 months and pulsed magnetic therapy or surgery. In the event of a doubtful positive result with scintigraphy, antibiotics are prescribed along with magnetic field therapy and a subsequent clinical and radiological follow-up after 4 weeks. This is followed by scintigraphy with labelled leucocytes after 2 weeks of suspension of the antibiotic therapy. If the results of

Fig. 3

Post-traumatic infection

Physical exam ERS, C-reactive protein

X-rays

Doubtful or negative

WBC scintigraphy

Positive

Medical therapy and pulsed magneto TH

Positive

Medical therapy or surgery

Follow-up clinical EX X-rays WBC scintigraphy

Doubtful

Medical therapy and pulsed magneto TH

Negative

Follow-up clinical EX X-rays WBC scintigraphy

Follow-up clinical EX X-rays WBC scintigraphy

Diagnostic flow chart of peripheral post-traumatic and secondary bone infections.

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Osteomyelitis: clinical update for practical guidelines Concia et al. 655

WBC scintigraphy are negative, the antibiotics can be restarted to consolidate the results and the clinical and radiological follow-up can take place after 4 weeks followed by labelled-leucocyte scintigraphy at 2 months. Infections of prosthetic joints

With infections of prosthetic joints, however, a different approach is required for hip and knee. As the knee is closer to the surface and more easily reached it allows a quicker arthrocentesis and thus the possibility to obtain a specific diagnosis and implement a targeted therapy regime. In these cases, in addition, we need to know the extent of the infection and degree of activity but in the event of a painful prosthesis, the first thing to be done is to obtain conventional X-rays and carry out laboratory tests (Fig. 4). If the results of these examinations are negative but the prosthesis is still causing pain, we order a three-phase bone scan. If the scan results are normal in all three phases, we begin aetiological therapy and observation regarding pain and inflammation. If the bone scan results are positive in all three phases (perfusion, blood pool and metabolic phase) and there is bone remodelling, but the laboratory tests are negative for infection, we institute a regimen of medication and physiotherapy along with pulsed magnetic therapy and subsequent follow-up with radiography and a three-phase bone scan.

On the other hand, if the initial laboratory tests (ESR or C-reactive protein) and radiological examinations are positive for an infection, we order WBC scintigraphy. If the result of scintigraphy is negative for infection, we implement physiotherapy with pulsed magnetic therapy, or consider re-implanting the prosthesis, although this is not an easy clinical decision to make. If the scintigraphy results are positive, we have the opportunity to conduct antibiotic therapy followed by radiography then radiological and labelled leucocytes scintigraphic checks (Fig. 5). If the results of WBC scintigraphy are negative, physiotherapy is again the option. If the results remain positive, we prefer explanting the prosthesis and put in the antibiotic-impregnated cement spacer along with systemic administration of antibiotics for 4–6 weeks. Only when the follow-up with labelled WBCs has become negative do we proceed with the re-implantation of the joint prosthesis. There is a third possibility: that of the doubtful positive of the radiographic images and the laboratory tests. In this case, we prefer to carry out WBC scintigraphy, which is a better determinant. If this is negative, we return to medical therapy and physiotherapy. If it is positive we proceed to antibiotic therapy or even removal of the joint prosthesis, depending upon the case.

Fig. 4

Painful prosthesis

Negative

Medical therapy

Three phase Bone scan

Negative ESR C-reactive protein Rx Doubtful

Positive Negative

WBC scan

Positive

Positive

Positive

Negative

Three phase Bone scan

Antibiotics therapy

Medical and physical therapy

Physical therapy and prosthesis reimplantation

Negative

Rx WBC scan

Clinical follow-up Rx and Bone scan

Negative Prosthesis reimplantation

Positive

Surgery and antibiotics

Clinical follow-up and WBC scan

Diagnostic flow chart of the diagnosis of suspect infection in painful joints prostheses.

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Fig. 5

Example of a woman with infection of a left-knee prosthesis. (a) Biphasic 99mTc-MDP scintigraphy on 3 September 2002. The 5 minutes images show an increased perfusion in the tissues surrounding the knee prosthesis. The 3 hours images show that the uptake of MDP prevails in the tibial epiphysis and in patella. (b) Three-phase 99mTc-HMPAO WBC scintigraphy of the same patient on 18 September 2002 (i.e., 2 weeks after the images in panel a). There is significant increase of labelled WBCs both in femur and tibia periprosthetic bone, in surrounding soft tissues and in the articular cavity. (c) Three-phase 99mTc-HMPAO WBC scintigraphy check-up on 5 December 2002, after removal of the infected knee prosthesis, with 6 weeks of antibiotic therapy and the use of antimicrobial-impregnated cement in the articular cavity to allow partial joint mobility and length of leg.

Vertebral osteomyelitis

In diagnosing vertebral osteomyelitis, a series of haematochemical assessments must be carried out, including indices of inflammation, culture tests on biological liquids and instrumental examinations, including biopsies. Early diagnosis is the key to resolving septic spondylodiskitis as it can prevent the onset of permanent neurological deficits and the formation of vertebral deformities. MRI is, without doubt, the most important and most sensitive tool and allows the formulation of a differential diagnosis with degenerative and metastatic processes as it supplies data on the anatomy and the extension of the infectious process in haematogenous spondylodiskitis. In the study of post-operative spondylodiskitis, the sensitivity and specificity of radiological methods suffer a substantial loss due to the presence of scar tissue and/or post-operative reactions. From the diagnostic point of view, biopsies are very important examinations in identifying pathogenic agents and can be affected by the transpedicular or disk routes or by open surgery. This procedure can produce negative results in 30–50% of cases, however, and is therefore not particularly useful in diagnosing spondylodiskitis, which in 90% of cases is postoperative, with bones that have been altered by nonspecific re-modelling and is thus difficult to diagnose using radiological methods. In traditional diagnostic algorithms (Fig. 6), scintigraphy with 99mTc-hydroxymethylene diphosphonate (99mTcHDP) and 67Ga citrate are often mentioned. Both

methods use tracers that accumulate in inflamed areas and are highly sensitive. Nowadays, even if this use has not been fully referred to in diagnostic algorithms, nuclear medicine makes great use of this technique in diagnosing post-operative spondylodiskitis. The radiopharmaceuticals to be used in a case of suspected spondylodiskitis are, amongst those available in all the centres, 99mTc-HDP, 111In- and 99mTc-labelled WBCs, immunoscintigraphy with monoclonal antibodies and 67 Ga citrate. Others still in the experimental stage include 111In-biotin and 99mTc-ciprofloxacin (Fig. 7). In descending order of accuracy of scintigraphic tracers, the data from the study of spondylodiskitis is as follows: 18FFDG (90%) (Fig. 8), 67Ga citrate (88.50%), antigranulocyte antibodies (88.5%) and labelled WBCs (65.5–80%). The data regarding accuracy with experimental markers is 111In-biotin (95.2%) and 99mTcciprofloxacin (81.5%) [66–72]. This demonstrates therefore that the radiopharmaceuticals to be used in cases of suspected spondylodiskitis are 99m Tc-HDP and 67Ga citrate, which are available in all the centres, 18F-FDG is available in selected centres and 111 In-biotin and 99mTc-ciprofloxacin where possible (Fig. 9). Scintigraphy with inflammation tracers must be used as a complementary examination to current radiological methods (CT or MRI) where haematogenous diskitis is suspected. It may be chosen, however, as the primary investigative tool in cases of suspected post-operative diskitis and its follow-up.

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Osteomyelitis: clinical update for practical guidelines Concia et al. 657

Scintigraphy with inflammation markers must always be ordered where there is suspected post-operative diskitis or diskitis of another type, where there are doubtful or Fig. 6

Bone pain

Septic diskitis conventional approach

Physical examination

X-ray + LAB (ESR, C-reactive protein) Positive

Negative or doubtful

Positive

Therapy Positive

Positive

CT e/o MRI Negative or doubtful

99mTc-MDP + 67Ga Negative

Doubtful

Bone biopsy

Negative

Follow-up

Diagnostic flow chart of infective diskitis with conventional nuclear imaging procedures.

negative CT and/or MRI results in the event of positive results from a bone scan. In vertebral osteomyelitis the use of a specific therapy against the agent demonstrated in cultures of blood or in biopsy is preferable. More frequently, the antibiotic treatment is empirical and this may be justified in primitive diskitis without tubercular marks or in iatrogenic infections in which S. aureus is the most common aetiological agent. If there is no clinical improvement or significant reduction of inflammation indices, after at least 4–6 weeks of therapy against Staphylococcus aureus, it is necessary to repeat the antibiogram by a biopsy or by an open surgical incision. The therapeutic choice against Staphylococcus will be decided because of its methicillin resistance: we use oxacillin if the bacteria are methicillinsensitive or teicoplanin if the bacteria are methicillinresistant. The therapeutic treatment generally begins during hospitalization with an association of oxacillin– rifampicin or teicoplanin–rifampicin administered by intravenous injection for 4–6 weeks or in selected cases with oral therapy with linezolid to obtain a regression of clinical signs and normalization of laboratory tests. The treatment continues with oral administration of antibiotics, including amoxicillin–clavulanic acid, minociclin, moxifloxacin in association with rifampicin). The treatment of diskitis by Gram-negative aetiology consists of the utilization of protected ureidopenicillin (piperacillin– tazobactam), carbepenems, third-generation cephalosporins, aztreonam with aminoglycosides for 4 weeks and then

Fig. 7

Lumbar diskitis in a 65-year-old woman. 99mTc-MDP, 99mTc-HMPAO WBC scintigraphy and 99mTc-Infecton were used in the study. In the upper row, the images of the lumbar spine are reported as spot views, while in the lower row, the images are the corresponding coronal SPECT views for each tracer. There is a typical linear hot spot in the space between L5–S1 vertebral bodies in the bone scan images which corresponds to a cold area in the WBC scan. The Infecton images show a focal uptake in the spine without involvement of soft tissues.

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Fig. 8

(a) Bone scan taken at 5 min and 3 h after i.v. injection of 99mTc-MDP, and a 67Ga scan of a diskitis of the whole lumbar column treated by antibiotics and surgical stabilization of multiple vertebral bodies. In comparison with the normal uptake of the dorsal column increased uptake of 99mTc-MDP and decreased uptake of 67Ga by the lumbar spine was found. In our experience, such a result is unusual. (b) Fusion of the lumbar SPECT with 67Ga and 18 F-FDG PET of the same patients 2 weeks later. Uptake of 18F-FDG in two linear areas located at the second (filled arrow) and fourth (unfilled arrow) intervertebral space of lumbar spine. The uptake of gallium is limited to the fourth space.

quinolones. The duration of therapy is based on clinical response, inflammation indices and diagnostic imaging. In our experience, we found full recovery from disease after 3–18 months of therapy. The diskitis of Candida aetiology is treated initially with amphotericin B liposomal or caspofungin i.v. with a switch to oral fluconazole or voriconazole for at least 6 months. Tubercular diskitis needs an association therapy with isoniazid, rifampicin, pyrazinamide and ethambutol at least for 2 months followed by isoniazid and rifampicin for the successive 7 months.

Conclusions Osteomyelitis is an infection with multiple aspects but it is always difficult to treat because of the characteristics of

the microorganisms involved (adherence to bone and prostheses or bio film production). In fact, most bone infections are chronic or become chronic with complications as sequestra or bone destruction, which can require orthopaedic interventions for debridement or consolidation. The management of these patients is complicated and needs the cooperation of clinicians, orthopaedic specialists, radiologists and nuclear medicine physicians. The objective of the diagnosis of bone infections is to identify the agent in order to provide an aetiological antibiotic therapy. When this is not possible, it becomes necessary to assess if there is an infection or a simple inflammation or reaction to bone injury.

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Osteomyelitis: clinical update for practical guidelines Concia et al. 659

5

Fig. 9

Septic diskitis Experimental approach

Bone pain

6 7

Physical examination 8

X-ray + LAB (ESR, C-reactive protein)

Negative or doubtful

CT e/o MRI

9 10

Positive

Positive

Negative or doubtful

Therapy

99mTc-MDP

+

Positive

11

67Ga

12

Positive

Negative Doubtful

Bone biopsy

Positive

18F-FDG

PET

111In-Biotin

Negative

13

Follow-up

99mTc-Infecton

14

Diagnostic flow chart of infective diskitis with experimental nuclear imaging procedures.

15

16

As a start, the diagnostic flow charts of osteomyelitis require physical examination, laboratory tests and then radiological imaging. Nuclear medicine plays an important role in the diagnosis of bone infections. A threephase bone scan is more advanced than other imaging modalities in haematogenous osteomyelitis. In the followup or after surgical interventions on bones, the importance of nuclear medicine procedures increases. In prosthetic-joint infections or in peripheral fractures radionuclide studies with WBCs labelled with either 111 In-oxime or 99mTc-HMPAO are the primary imaging modality for determining if an infection is present and to differentiate it from a simple inflammation or other bone alterations. The tracers available for assessing the activity of a bone infection also include 67Ga and 18F-FDG which are essential in the assessment of persistence or activity of a secondary infection of the axial skeleton.

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Summers M, Misenhimer GR, Antony SJ. Vancomycin-resistant Enterococcus faecium osteomyelitis: successful treatment with quinupristin–dalfopristin. South Med J 2001; 94:353–355. Conrad DA, Williams RR, Couchman TL, Lentnek AL. Efficacy of aztreonam in the treatment of skeletal infections due to Pseudomonas aeruginosa. Rev Infect Dis 1991; 13(suppl 7):S634–S639. Mader JT, Shirtliff ME, Bergquist SC, Calhoun J. Antimicrobial treatment of chronic osteomyelitis [Review]. Clin Orthop Relat Res 1999; 360:47–65. Tice AD, Hoaglund PA, Shoultz DA. Outcomes of osteomyelitis among patients treated with outpatient parenteral antimicrobial therapy. Am J Med 2003; 114:723–728. Swiontkowski MF, Hanel DP, Vedder NB, Schwappach JR. A comparison of short- and long-term intravenous antibiotic therapy in the postoperative management of adult osteomyelitis. J Bone Joint Surg Br 1999; 81: 1046–1050. Ceri H, Olson ME, Stremick C, Read RR, Morck D, Buret A. The Calgary Biofilm Device: new technology for rapid determination of antibiotic susceptibilities of bacterial biofilms. J Clin Microbiol 1999; 37:1771. Anderl JN, Zahller J, Roe F, Stewart PS. Role of nutrient limitation and stationary phase existence in Klebsiella pneumoniae biofilm resistance to ampicillin and ciprofloxacin. Antimicrob Agents Chemother 2003; 47: 1251–1256. Buchholz HW, Elson RA, Hengelbrecht E, Lodenkaemper H, Roettger J, Siegel A. Management of deep infection of total hip replacement. J Bone Joint Surg 1981; 63-B:342–353. Wilde AH, Ruth JT. Two stages reimplantation in infected total knee arthroplasty. Clin Orthop 1988; 236:23–35. Ghisellini F, Ceffa R. Trattamento delle infezioni di protesi articolari. Roma: Mediprint Ed.; 1997. Cerretani D, Giorgi G, Fornara P, Bocchi L, Neri L, Ceffa R, et al. The ‘in vitro’ elution characteristics of vancomycin combined with imipenem– cilastatin in acrylic bone cements: A pharmacokinetic study. J Arthroplast 2002; 17:5. Gomis M, Barberan J, Sanchez B, Khorrami S, Borja J, Garcia-Barbal J. Oral ofloxacin versus parenteral imipenem–cilastatin in the treatment of osteomyelitis. Rev Esp Quimioter 1999; 12:244–249. Mader JT, Ortiz M, Calhoun JH. Update on the diagnosis and management of osteomyelitis [Review]. Clin Podiatr Med Surg 1996; 13:701–724. Boutin RD, Brossmann J, Sartoris DJ, Reilly D, Resnick D. Update on imaging of orthopaedic infections. Orthop Clin North Am 1998; 29:41–66. Atkinson RN, Paterson DC, Morris LL, Savage JP. Bone scintigraphy in discitis and related disorders in children. Aust NZ J Surg 1978; 48: 374–377. Lazzeri E, Manca M, Molea N, Marchetti S, Consoli V, Bodei L, et al. Clinical validation of the avidin/indium-111 biotin approach for imaging infection/ inflammation in orthopaedic patients. Eur J Nucl Med 1999; 26:606–614. De Winter F, Gemmel F, Van Laere K, De Winter O, Poffijn B, Dierckx RA, Van de Wiele C. 99mTc-ciprofloxacin planar and tomographic imaging for the diagnosis of infection in the postoperative spine: experience in 48 patients. Eur J Nucl Med Mol Imaging 2004; 31:233–239. Gratz S, Dorner J, Oestmann JW, Opitz M, Behr T, Meller J, et al. 67Gacitrate and 99Tcm-MDP for estimating the severity of vertebral osteomyelitis. Nucl Med Commun 2000; 21:111–120. Lazzeri E, Pauwels EK, Erba PA, Volterrani D, Manca M, Bodei L, et al. Clinical feasibility of two-step streptavidin/111In-biotin scintigraphy in patients with suspected vertebral osteomyelitis. Eur J Nucl Med Mol Imaging 2004; 31:1505–1511. E-pub 6 July 2004. Love C, Patel M, Lonner BS, Tomas MB, Palestro CJ. Diagnosing spinal osteomyelitis: a comparison of bone and Ga-67 scintigraphy and magnetic resonance imaging. Clin Nucl Med 2000; 25:963–977. Prandini N, Feggi L, Panareo S, Ciprian A, Massari L, Galla A, et al. 99mTcciprofloxacin in the imaging of central bone infections. Nucl Med Commun 2001; 22:1156–1157.

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Original article

Preparation and preliminary biological evaluation of 177 Lu-labelled hydroxyapatite as a promising agent for radiation synovectomy of small joints Sudipta Chakrabortya, Tapas Dasa, Sharmila Banerjeea, Haladhar Dev Sarmab and Meera Venkatesha Aim Lutetium-177 (177Lu) is considered to be a promising radionuclide for use in radiation synovectomy of small-sized joints owing to its favourable decay characteristics [t1/2 = 6.73 days, Eb(max) = 0.49 MeV, Ec = 113 keV (6.4%), 208 keV (11%)] and feasible and cost-effective production route. Hydroxyapatite particles are regarded as one of the most suitable carriers for applications in radiation synovectomy, and labelling with 177 Lu has been envisaged. The present work describes the preparation and preliminary biological evaluation of 177 Lu-labelled hydroxyapatite particles. Methods 177Lu-labelled hydroxyapatite particles were prepared using 177Lu produced by thermal neutron irradiation of a natural (2.6% 176Lu) Lu2O3 target and hydroxyapatite particles (particle size, 2–10 lm) prepared in-house. The biological efficacy of the radiolabelled preparation was tested by recording serial gamma scintigraphic images after injecting the agent in both normal and arthritic knee joints of Wistar rats. Results 177Lu-hydroxyapatite was prepared with high yield and high radiochemical purity (B99%) and the radiolabelled particles showed excellent in-vitro stability at

Introduction Rheumatoid arthritis (RA) is a chronic inflammatory disorder of joints that is characterized by the inflammation and proliferation of synovial tissues [1]. Approximately 1% of the adult population worldwide is affected by this disease, and it is manifested in pain, joint immobility and disability [2–5]. If unchecked, it often results in the destruction of articular cartilage, eventually leading to complete deformation and destruction of the involved diarthrodial or synovial joints [2–5]. Radiation synovectomy (RSV) is a radiotherapeutic modality wherein a b – -emitting radionuclide is administered locally by intra-articular injection in the form of a colloid or radiolabelled particulate. This mode of therapy has been successfully employed as a viable alternative to surgical and chemical synovectomy for more than 50 years for the treatment of RA and other types of inflammatory arthropathies after failure of systemic pharmacotherapy and intra-articular steroid injections [1–3,6–9].

room temperature. Serial scintigraphic images of normal and arthritic Wistar rats showed complete retention of activity within the synovial cavity, with no measurable activity leaching out from the joint until 168 h post-injection. Conclusion Studies with 177Lu-hydroxyapatite indicate its potential for use as an agent for radiation synovectomy of digital joints, as a viable alternative to 169Er-based agents. The results also demonstrate the possibility of preparing a large number of patient doses of 177Lu-hydroxyapatite from indigenously produced 177Lu using a natural target. Nucl c 2006 Lippincott Williams & Med Commun 27:661–668
Wilkins. Nuclear Medicine Communications 2006, 27:661–668 Keywords: digital joints, hydroxyapatite,

177

Lu, radiation synovectomy

a Radiopharmaceuticals Division and bRadiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai, India.

Correspondence to Sharmila Banerjee, Radiopharmaceuticals Division, Bhabha Atomic Research Centre, Mumbai 400 085, India. Tel: 91-22-2559 0616; fax: 91-22-2550 5345; e-mail: [email protected] Received 5 April 2006 Accepted 8 May 2006

It has been well documented that radiolabelled particulates and radionuclide-loaded colloid particles are rapidly phagocytosed by macrophages in the inflamed synovial membrane, which is eventually ablated by radiation from the b – particles [2,6]. However, the primary disadvantage of the RSV procedure is the unacceptable radiation dose delivered to non-target organs, such as the liver, spleen and lymph nodes, as a result of leakage of the radioactive material from the synovial cavity [6,9–11]. Leakage usually results from either the very small size of the particulates or from the poor in-vivo stability of the preparation leading to the dissociation of activity from the particles [6,9,11,12]. Of the different types of agent used for RSV so far, the majority are radiocolloid or radiolabelled macroaggregates in nature. However, the basic problem associated with these agents is leakage of radioactivity from inflamed joints, mainly because of a lack of control of particle size [6,10,11]. Therefore, an ideal agent for RSV would be one in which the

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Nuclear Medicine Communications 2006, Vol 27 No 8

radionuclide is irreversibly attached to pre-formed particles of appropriate size. The ideal size of such particles is reported to be 2–10 mm, so that they are small enough to be phagocytosed, but not so small that they may leak out of the cavity before being phagocytosed [2,3,6,13]. In addition, the particles should be biodegradable, and the biological half-life of such particles should be longer than the physical half-life of the radionuclide tagged with them [6,11]. Hydroxyapatite [Ca10(PO4)6(OH)2] is one of the preferred particulates as it is the natural mineral constituent of bone matrix and can be synthesized easily with the desired particle size [3,11]. Hydroxyapatite particles are converted to Ca2 + and PO34 – ions by natural metabolic processes and are eliminated over a period of 6 weeks, thereby providing excellent biocompatibility [3,11]. A wide variety of radionuclides have been proposed and a number have been tested in animal models and humans for RSV [3,4,6,10,11,14]. It is generally believed that b – emitting particles, after being phagocytosed, are evenly distributed in the synovium; therefore, the energy of the radionuclide must be proportional to the size of the synovial joint being irradiated in order to obtain therapeutic efficacy [2,6,15]. For an ideal radionuclide, the b – particle energy should be sufficient to penetrate and ablate the proliferating synovial tissue, but the radiation dose to the underlying articular cartilage should be minimal [6]. This is evident from the fact that 90Y-labelled [Eb(max) = 2.21 MeV] silicate/citrate colloid is used to carry out RSV of large joints (hip, shoulder and knee), whereas 186 Re-labelled [Eb(max) = 1.07 MeV] colloids are used in the treatment of medium-sized joints (wrist, elbow and ankle). Similarly, for the treatment of digital joints, 169Erlabelled [Eb(max) = 0.35 MeV] citrate colloid has been recommended [2,16–19]. To our knowledge 169Er is the only radioisotope used so far clinically for the RSV of digital joints. However, the production of 169Er is costly owing to the fact that it requires the irradiation of highly enriched 168 Er target so that the desired radionuclide can be produced with sufficient radionuclide purity. The use of an enriched target is also essential to produce 169Er with adequate specific activity to be commercially viable, as the thermal neutron capture cross-section of 168Er is only 1.95 10 – 28 m2 [20]. Moreover, for isotope production, the use of enriched targets with low activation cross-section is not economical, as a significant part of the costly target remains unused. In this context, 177Lu could be considered as a viable alternative to 169Er for developing potential agents for RSV of small-sized joints owing to its suitable nuclear decay characteristics [t1/2 = 6.73 days, Eb(max) = 0.49 MeV, Eg = 113 keV (6.4%), 208 keV (11%)] [20]. The presence of g photons of imageable energy with low abundance provides the additional benefit of carrying out simultaneous scintigraphy and dosimetry studies. Moreover, 177Lu can be produced with adequately high specific activity using a natural lutetium target in a

moderate flux reactor owing to the high thermal neutron capture cross-section of 176Lu (s = 2100  10 – 28 m2) [20], and this would be sufficient to prepare a large number of patient doses for use in RSV [21]. Therefore, 177Lu-based RSV agents will be economically more viable than their 169 Er-labelled analogues. Extensive efforts are being made at our institution to develop suitable agents for RSV of different types of joints, and our experiences with hydroxyapatite particles labelled with 166Ho, 90Y and 186Re have shown encouraging results in animal models [11,22,23]. Of these, 166Hohydroxyapatite has undergone extensive clinical trials, and is an approved product for the RSV of large joints in India. In this paper, we describe the preparation of 177Lulabelled hydroxyapatite particles for RSV of digital joints using 177Lu produced by the irradiation of a natural Lu2O3 target. The biological behaviour of the radiolabelled particles is studied in normal and diseased animal models.

Experimental details Materials and methods

Natural Lu2O3 (spectroscopic grade, more than 99.99% chemically pure, 2.6% 176Lu) powder, used as the target for the production of 177Lu, was procured from American Potash Inc., Delaware, Ohio, USA. Hydroxyapatite particles used in this study were synthesized and characterized in our laboratory following the procedure reported previously [11]. The particle size distribution of hydroxyapatite particles, determined using a laser diffraction particle size analyser, is shown in Fig. 1. It is evident from Fig. 1 that most of the particles used in this study were in the size range of 2–10 mm, the most appropriate particle size range for radiolabelled particulates used in RSV [2,3,6,13]. Diethylenetriamine pentaacetic acid (DTPA) was obtained from Aldrich Chemical Co., Milwaukee, Wisconsin, USA. All other chemicals used were of AR grade and were supplied by reputable chemical manufacturers. LuCl3 solution, used as a carrier for the preparation of 177Lu-labelled hydroxyapatite particles, was prepared by dissolving natural Lu2O3 powder in 0.1 M HCl by gentle heating. The radionuclide purity of the 177Lu produced was determined by high-resolution g-ray spectrometry using an HPGe detector (EGG Ortec/Canberra detector, Oak Ridge, Tennessee, USA) coupled to a 4 K multichannel analyser (MCA). The 152Eu reference source, used for both energy and efficiency calibration of the detector, was obtained from Amersham Inc., Piscataway, New Jersey, USA. All other radioactivity measurements were carried out using a welltype NaI(Tl) scintillation counter after adjusting the baseline at 150 kev and keeping a window of 100 kev, thereby utilizing the 208 keV g photon of 177Lu. Whatman 3 MM chromatography paper (Whatman, Maidstone, Kent, UK) was used for paper chromatography studies.

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177

Lu-hydroxyapatite for radiation synovectomy of small joints Chakraborty et al. 663

Fig. 1

A HA∗5

FIXED

VOLUME

15

50

0 0.1

1

10

U%

F%

100

0 100 200

SIZE
m % on DIA : 10.0
m = 95.5% DIA on % : 90.0% = 8.31
m SP.AREA = 16135 cm2/cm3 MEDIAN = 4.33
m

Particle size distribution of hydroxyapatite particles used for the preparation of 177Lu-hydroxyapatite.

Scintigraphic images were recorded using a single-head digital single photon emission computed tomography (SPECT) gamma camera (MPS GE, Milwaukee, Wisconsin, USA). Production and radiochemical processing of

177

Lu

177

Lu was produced by irradiation of a natural Lu2O3 (2.6% 176 Lu) target in the Dhruva reactor at a thermal neutron flux of approximately 3  1013 neutrons/cm2/s for 7 days. A weighed amount of Lu2O3 powder was placed into a quartz ampoule, which was subsequently flame sealed and irradiated after placing inside an aluminium can. Following irradiation, the target was dissolved in 1 M HCl by gentle warming inside a lead-shielded plant, after allowing a cooling period of approximately 6 h. The resultant solution was evaporated to near-dryness and reconstituted in doubledistilled water. An assay of the total activity produced was carried out by measuring the ionization current obtained when an aliquot of the processed 177LuCl3 solution was placed inside a pre-calibrated well-type ion chamber. The radionuclide purity of the 177Lu produced was ascertained by recording the g-ray spectra using an HPGe detector coupled to a 4 K MCA system. Energy and efficiency calibrations of the detector were carried out using a 152Eu reference source prior to the recording of g-ray spectra. Several spectra were recorded for each batch at regular time intervals. Samples measured initially for the assay of 177Lu were preserved for complete decay of 177Lu (8–10t1/2 of 177 Lu, i.e. for a period of 50–70 days) and re-assayed to determine the activity of long-lived 177mLu (t1/2 = 160.5 days). Appropriately diluted aliquots of the processed 177 LuCl3 solution were counted for 1 h. Preparation of

177

Lu-hydroxyapatite

The preparation of 177Lu-labelled hydroxyapatite particles was carried out by adding 0.1 ml of 177LuCl3 solution

(B10 MBq 177Lu) containing 200 mg of lutetium carrier to a suspension of 5 mg of hydroxyapatite in 0.8 ml of normal saline after the addition of 0.1 ml of 0.5 M NaHCO3 buffer (pHB9). The reaction mixture was vortexed thoroughly after adjusting to pHB7 using 0.1 M NaOH solution, and was mixed continuously at room temperature for 30 min in a rotary shaker. Subsequently, the reaction mixture was centrifuged at 1000 g for 5 min. The supernatant was separated from the precipitate carefully. The radiolabelled hydroxyapatite particles obtained as a precipitate were subjected to further washings (three times), using 1 ml of normal saline on each occasion, to ensure the removal of free 177Lu activity in case of possible leaching from the labelled particles. Finally, the 177Lu-labelled particles were suspended in sterile saline, autoclaved and used for biological studies. Determination of radiolabelling yield and radiochemical purity of 177Lu-hydroxyapatite

For the determination of the radiolabelling yield, the reaction mixture was vortexed thoroughly and centrifuged at 1000 g for 5 min after the completion of the reaction. Subsequently, half the volume of the supernatant solution was carefully pipetted out into a test-tube and the 177Lu activity was measured. In a similar manner, the 177 Lu activity associated with the hydroxyapatite particles together with the remaining half of the supernatant solution was also measured. From these data, the percentage radiolabelling yield and radiochemical purity of 177Lu-hydroxyapatite were calculated using the following equation: Radiolabelling yield ð%Þ ¼ ½ðY  XÞ=ðY þ XÞ100 where X is the background-corrected count rate associated with half the volume of the supernatant solution and Y is the background-corrected count rate of the precipitated hydroxyapatite particles together with the remaining half of the supernatant. The radiochemical purity of the 177Lu-hydroxyapatite prepared was further confirmed by carrying out paper chromatography using suitable eluting solvent. In this technique, 5 ml aliquots of the reaction mixture were applied at 1.5 cm from the lower end of Whatman 3 mm chromatography paper strips (12 cm  2 cm). The strips were developed in a 5 mM aqueous solution of DTPA, dried, cut into segments of 1 cm each and the radioactivity associated with each segment was measured. Similar experiments were carried out with 177LuCl3 under identical conditions (without hydroxyapatite particles) for comparison. Optimization studies

In order to obtain the maximum radiolabelling yield of 177 Lu-hydroxyapatite, several experiments were carried out by varying different reaction parameters, such as the ligand (hydroxyapatite) concentration, pH of the reaction mixture,

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664 Nuclear Medicine Communications 2006, Vol 27 No 8

incubation time and carrier lutetium concentration. In a reaction volume of 1 ml, the amount of hydroxyapatite used for radiolabelling was varied between 1 and 20 mg and the radiolabelling yields were determined in each case. The effect of the variation of pH on radiolabelling yield at room temperature was studied by adjusting the reaction mixture to different pH values in the range 1–9 using either 1 M HCl or 1 M NaOH solutions. The incubation time required to obtain the maximum labelling yield was optimized by carrying out reactions for different time periods (0, 10, 20, 30, 60 and 120 min) at room temperature and determining the yields in each case. The radiolabelling yields of 177Lu-hydroxyapatite were also determined for different concentrations of carrier lutetium (20, 50, 100, 200, 500 and 1000 mg/ml) using the optimized amount of hydroxyapatite for radiolabelling. In-vitro stability studies

The in-vitro stability of 177Lu-labelled hydroxyapatite particles was studied to determine the leaching of 177Lu activity, if any, from the radiolabelled hydroxyapatite particles on storage. In order to ascertain the in-vitro stability, 177Lu-labelled hydroxyapatite particles prepared under optimized conditions were suspended in 1 ml of normal saline and stored at room temperature for up to 14 days (more than two half-lives of 177Lu). At the end of different time intervals (1, 2, 4, 7 and 14 days), the suspension was vortexed thoroughly and centrifuged at 1000 g for 5 min. Following this, half the volume of the supernatant solution was removed and counted to detect the possible leaching out of the radioactivity from the particles. The 177Lu activity associated with the hydroxyapatite particles together with the remaining supernatant solution was also determined to ascertain the radiochemical purity of the labelled preparation. The supernatant was returned to the original suspension after counting. This procedure was repeated at different time intervals. Biological studies

The biological behaviour of the prepared 177Lu-labelled hydroxyapatite particles was studied by injecting the radiolabelled preparation into both normal and arthritic knee joints of Wistar rats and determining the retention and leakage of the activity from the synovium. Serial scintigraphic images of the animals were recorded using a single-head digital SPECT gamma camera to determine the in-vivo localization of the injected radioactivity. Arthritis induction was carried out in 20-week-old male Wistar rats weighing approximately 300 g following anaesthetization using a combination of xylazine hydrochloride and ketamine hydrochloride. The left knee area of the animals was clipped and prepared aseptically for the intra-articular injection of 175 ml of complete Freund’s adjuvant (CFA) directly into the knee joint above the meniscus. The knee joint was approached craniolaterally in between the ligamentum collaterale laterale genus and

musculus gastrocnemius lateralis, below the condulis lateralis osis femoris. Inflammatory symptoms were noticed within 3–4 days of CFA injection. A sterile preparation of 177Lu-hydroxyapatite particles (7.4 MBq), suspended in 100 ml of normal saline, was injected intraarticularly in one of the knee joints of a healthy Wistar rat weighing approximately 300 g. A similar volume of the preparation was also injected in a Wistar rat with an arthritic knee joint. Prior to the acquisition of the images, the animals were anaesthetized using a combination of xylazine hydrochloride and ketamine hydrochloride. Sequential scintigraphic images were acquired using a single-head digital SPECT gamma camera at 30 min and 3, 24, 48, 96 and 168 h post-injection using a low-energy, high-resolution (LEHR) collimator. The gamma camera had previously been calibrated for the 208 keV g photon of 177Lu with a 20% window. All the images were recorded by acquiring 500 kilocounts using a 256  256 matrix size. Estimation of the activity arising from possible in-vivo leaching of the radiolabelled particulates injected in the knee joints was also carried out by drawing blood samples at the same time intervals from the tail vein of the animals and counting for any 177Lu activity present. All the animal experiments were carried out in compliance with the relevant national laws relating to the conduct of animal experimentation.

Results and discussion Production of

177

Lu

Irradiation of a natural Lu2O3 target at a thermal neutron flux of approximately 3  1013 neutrons/cm2/s for 7 days yielded 177Lu with a specific activity of approximately 4 GBq/mg (approximately 108 mCi/mg) at 6 h after the end of bombardment (EOB). This is considerably higher than the theoretically calculated value of the specific activity (2.9 GBq/mg) under the same irradiation conditions. This could perhaps be attributed to the contribution from epithermal neutrons (resonance integral, 1087  10 – 28 m2) [20], which is not accounted for in the theoretical calculations. The specific activity of 177Lu could be enhanced manifold by irradiating a commercially available enriched lutetium target (more than 60% of 176 Lu) instead of natural lutetium. For practical purposes, however, the specific activity of 177Lu obtained using a natural lutetium target is adequate for the preparation of agents for RSV, and hence the use of an enriched target is not essential. The specific activity could be increased to some extent, however, by carrying out longer irradiations (approximately 3–4 weeks) using the natural target. The radionuclide purity of the 177Lu produced was 99.985%, obtained by analysing the g-ray spectrum of the irradiated target after radiochemical processing. The major g peaks observed were at 72, 112, 208, 250 and 321 keV, all of which correspond to the photopeaks of 177Lu [20]. This was further confirmed from the decay followed by monitoring

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177

Lu-hydroxyapatite for radiation synovectomy of small joints Chakraborty et al. 665

Radiolabelling yield and radiochemical purity of 177 Lu-hydroxyapatite

concentration did not show any appreciable increase in the complexation yield, and hence 5 mg/ml was considered to be the optimum ligand concentration for further studies. The effect of the variation of the hydroxyapatite concentration on the radiolabelling yield of 177Luhydroxyapatite is shown in Fig. 3. Variation of the pH of the reaction mixture within the range 3–8 showed that it did not have any significant effect on the complexation yield, which was observed to be 97.2 ± 1.9%. Below pH 3, the radiolabelling yield was found to be poor because, at acidic pH, hydroxyapatite particles are unstable and undergo gradual dissolution. Above pH 8, a slight decrease in the radiolabelling yield was observed. The optimum pH for radiolabelling was

Fig. 2

(a) 100

80 % Radioactivity

the peak area count per second values at these peaks according to the half-life of 177Lu. It is worthwhile to note that there is a possibility of the formation of 176mLu (t1/2 = 3.7 h), via the 175Lu(n,g)176mLu process (s = 16.4  10 – 28 m2), and of 177mLu (t1/2 = 160.5 days), via the 176 Lu(n,g)177mLu process (s = 7.0  10 – 28 m2), together with 177Lu, on thermal neutron bombardment of a natural Lu2O3 target [20,21,24,25]. As 176mLu has a half-life of 3.7 h, 24 h of cooling ensures the complete decay of the 176m Lu activity produced. However, long-lived 177mLu is a likely radionuclide impurity present in processed 177Lu. Nevertheless, the g-ray spectrum did not show any significant peak (at 128, 153, 228, 378, 414 and 418 keV) corresponding to 177mLu [20]. This is expected as the radioactivity from the 177mLu produced will not be significant on 7 days of irradiation owing to its long halflife and comparatively low cross-section of formation. The 177m Lu activity produced was determined by recording the g-ray spectrum of an aliquot of the sample, initially having high radioactive concentration, after complete decay of 177 Lu activity (8–10t1/2 of 177Lu, i.e. for a period of 50–70 days). The average level of radionuclide impurity burden in 177 Lu due to 177mLu, determined by this technique, was found to be 5.5 kBq of 177mLu/37 MBq of 177Lu (150 nCi/ 1 mCi) at EOB, which corresponds to 98.985% radionuclide purity of 177Lu at EOB.

60

40

177

The radiolabelling yield of Lu-hydroxyapatite, determined by measuring the radioactivity associated with half the volume of the supernatant solution and that associated with precipitated hydroxyapatite particles together with the remaining half of the supernatant, as described in the experimental section, was found to be approximately 99% under optimized reaction conditions.

Optimization studies

A complexation yield of 85.2 ± 1.1% was obtained with 1 mg of hydroxyapatite in 1 ml of reaction volume, which increased to 98.6 ± 0.8% when 5 mg of hydroxyapatite was used. A further increase in the hydroxyapatite

0

0

1 2 3 4 5 6 Migration from point of spotting (cm)

7

0

1 2 3 4 5 6 Migration from point of spotting (cm)

7

(b) 100

80 % Radioactivity

The radiochemical purity of 177Lu-hydroxyapatite was further ascertained by employing a paper chromatography technique. In paper chromatography, using a 5 mM aqueous solution of DTPA as the eluting solvent, it was observed that 177Lu-hydroxyapatite remained at the point of spotting (Rf = 0), whereas unreacted 177LuCl3 moved towards the solvent front (Rf = 1) due to the formation of an 177Lu–DTPA complex under identical conditions. Typical paper chromatography patterns of 177Lu-hydroxyapatite and 177LuCl3 (blank) are shown in Figs. 2(a) and 2(b), respectively. As no radioactivity could be detected at Rf of 177Lu–DTPA in the reaction mixture of 177Lu-hydroxyapatite, the presence of free 177LuCl3 in the radiolabelled preparation could be ruled out.

20

60

40

20

0

Paper chromatography patterns of 177Lu-hydroxyapatite (a) and 177 LuCl3 (b) using a 5 mM aqueous solution of diethylenetriamine pentaacetic acid (DTPA) as eluting solvent.

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666 Nuclear Medicine Communications 2006, Vol 27 No 8

therefore considered to be pHB7, which is advantageous as it is similar to the physiological pH.

Fig. 3

100

It was observed that the radiolabelling yield gradually increased with incubation time, and 98.4 ± 1.0% of complexation was achieved after 30 min of incubation at room temperature. Hence, 30 min of incubation at room temperature was considered to be the optimum time for radiolabelling.

% Radiolabelling yield

95

90

85

80 0

4

8 12 16 Concentration of HA (mg/ml)

20

Effect of variation of the hydroxyapatite (HA) concentration on the radiolabelling yield of 177Lu-hydroxyapatite.

The variation of the carrier lutetium concentration was also found to have an effect on the radiolabelling yield of 177 Lu-hydroxyapatite. It was found that a radiolabelling yield of 98.1 ± 1.4% could be obtained up to the addition of 200 mg of carrier lutetium. The yields were found to decrease gradually when the carrier concentrations were increased beyond this. Considering that approximately 3.7 GBq/mg (100 mCi/mg) specific activity of 177Lu can be obtained, 200 mg of lutetium corresponds to approximately 740 MBq (20 mCi) of 177Lu activity, which, in turn, is sufficient for the preparation of a number of

Fig. 4

Scintigraphic images of a normal Wistar rat injected with the 177Lu-hydroxyapatite preparation in one of the knee joints at various time points postinjection.

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177

Lu-hydroxyapatite for radiation synovectomy of small joints Chakraborty et al. 667

Fig. 5

Scintigraphic images of a Wistar rat with an arthritic left knee joint injected with the 177Lu-hydroxyapatite preparation in the diseased joint at various time points post-injection.

patient doses of 177Lu-labelled particulates for RSV of small-sized joints [21]. This activity is achievable in a single batch of 177Lu-hydroxyapatite prepared using hydroxyapatite particles in amounts as low as 5 mg. This implies that even a moderate specific activity of 177Lu, obtained from reactors with relatively low thermal neutron flux and using a natural lutetium target, can be used for the preparation of a large number of patient doses of 177Lu-hydroxyapatite particulate, thereby making it a more economically viable alternative than 169Erbased RSV agents.

In-vitro stability of

177

Lu-hydroxyapatite

In-vitro stability studies showed that the 177Lu-hydroxyapatite preparation was highly stable at room temperature in saline up to a time period of 14 days (more than two half-lives of 177Lu). This was evident from the retention of the radiochemical purity of the radiolabelled preparation to the extent of approximately 99% up to that period.

Biological studies

The scintigraphic images of normal Wistar rats obtained at various time intervals (3, 24, 48 and 168 h) after the intra-articular injection of the 177Lu-hydroxyapatite preparation in one of the knee joints are shown in Fig. 4. Similar scintigraphic images of animals with an arthritic knee joint, in which the agent was injected, obtained at the same time points, are depicted in Fig. 5. It is evident from the figures that all the injected activity remained localized in the synovium even at 6 days post-injection. The whole-body images recorded for the normal and diseased animals also did not show any detectable activity in any other organs, thereby confirming that no leakage of instilled particles occurred. This observation was further corroborated by observing the activity in blood samples taken at different time points. The blood samples did not show any radioactivity above the background activity post-injection at any of the time points studied. Moreover, the similar behaviour of the agent in normal and diseased animals confirms that, despite the induced inflammation, which results in increased vascularity and

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668 Nuclear Medicine Communications 2006, Vol 27 No 8

permeability [11], no leakage of the instilled particles occurred.

Conclusion This work describes the preparation of 177Lu-labelled hydroxyapatite particles in high yield and with excellent radiochemical purity using 177Lu produced by thermal neutron irradiation of a natural lutetium target in a moderate flux reactor and hydroxyapatite particles prepared and characterized in our laboratory. The radiolabelled particulates showed excellent in-vitro stability at room temperature. Biological studies carried out by injecting the radiolabelled preparation in both normal and arthritic knee joints of Wistar rats showed complete retention of the injected radioactivity within the synovial cavity until 168 h post-injection in both cases. These studies reveal that 177Lu-hydroxyapatite particles offer potential as a suitable agent for RSV, and further investigations are warranted to investigate their use as a viable alternative to 169Er-labelled particulates/radiocolloids for RSV of small joints.

5

6 7

8 9

10

11

12

13

14

Acknowledgements

15

The authors gratefully acknowledge Dr V. Venugopal (Director, Radiochemistry and Isotope Group) for his keen interest and encouragement. The authors also acknowledge the sincere help received from the staff members of the Animal House Facility, Bhabha Atomic Research Centre, during animal experimentation. The authors are grateful to Dr S. V. Thakare and Mr K. C. Jagadeesan for their valuable help in carrying out the irradiation of the lutetium targets.

16

17 18

19

20

References 1

2 3

4

Song J, Suh CH, Park YB, Lee SH, Yoo NC, Lee JD, et al. A phase I/IIa study on intra-articular injection of holmium-166-chitosan complex for the treatment of knee synovitis of rheumatoid arthritis. Eur J Nucl Med 2001; 28:489–497. Schneider P, Farahati J, Reiners C. Radiosynovectomy in rheumatology, orthopedics, and hemophilia. J Nucl Med 2005; 46:48S–54S. Chinol M, Vallabhajosula S, Goldsmith SJ, Klein MJ, Deutsch KF, Chinen LK, et al. Chemistry and biological behaviour of samarium-153 and rhenium186-labeled hydroxyapatite particles: potential radiopharmaceuticals for radiation synovectomy. J Nucl Med 1993; 34:1536–1542. Davis MA, Chinol M. Radiopharmaceuticals for radiation synovectomy: evaluation of two yttrium-90 particulate agents. J Nucl Med 1989; 30:1047–1055.

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O’Dell JR. Rheumatoid arthritis: the clinical picture. In: Koopman WJ, editor. Arthritis and allied conditions. A textbook of rheumatology. 14th ed. Philadelphia: Lippincott Williams & Wilkins; 2001. pp. 1153–1186. Deutsch E, Brodack JW, Deutsch KF. Radiation synovectomy revisited. Eur J Nucl Med 1993; 20:1113–1127. Shortkroff S, Sledge CB. Radiation synovectomy. In: Wagner HN Jr, Szabo Z, Buchanan JW, editors. Principles of nuclear medicine. 2nd ed. Philadelphia: W. B. Saunders; 1995: pp. 1021–1028. Siegel ME, Siegal HJ, Luck JV. Radiosynovectomy’s clinical applications and cost effectiveness: a review. Semin Nucl Med 1997; 4:364–371. Zuckerman JD, Sledge CB, Shortkroff S, Venkateshan P. Treatment of rheumatoid arthritis using radiopharmaceuticals. Nucl Med Biol 1987; 14:211–218. Sledge CB, Noble J, Hnatowich DJ, Kramer R, Shortkroff S. Experimental radiation synovectomy by 165-Dy ferric hydroxide macroaggregates. Arthritis Rheum 1977; 20:1334–1342. Unni PR, Chaudhari PR, Venkatesh M, Ramamoorthy N, Pillai MRA. Preparation and bioevaluation of 166Ho labeled hydroxyapatite (HA) particles for radiosynovectomy. Nucl Med Biol 2002; 29:199–209. Pandey U, Bapat KN, Samuel G, Sarma HD, Chaudhari PR, Dhami PS, et al. Evaluation of 90Y phosphate particles as a possible radiation synoviorthesis agent. Nucl Med Commun 2005; 26:459–463. Noble J, Jones AG, Davis MA, Sledge CB, Kramer RI, Livni E. Leakage of radioactive particle systems from a synovial joint studied with a gamma camera: its application to radiation synovectomy. J Bone Joint Surg 1983; 65A:381–389. Clunie G, Lui D, Cullum I, Edwards JCW, Ell PJ. Samarium-153-particulate hydroxyapatite for radiation synovectomy: biodistribution data for chronic knee synovitis. J Nucl Med 1995; 36:51–57. Harbert JC. Radiocolloid therapy in joint diseases. In: Harbert JC, editor. Nuclear medicine therapy. New York: Thieme Medical; 1987: pp. 169–186. Modder G. Radiosynoviorthesis – involvement of nuclear medicine in rheumatology and orthopedics. Meckenheim: Warlich Druck und Verlagsges; 2001. Siegel HJ, Luck VJ Jr, Siegel ME. Advances in radionuclide therapeutics in orthopedics. J Am Acad Orthop Surg 2004; 12:55–64. Kahan A, Modder G, Menkes CJ, Verrier P, Devaux JY, Bonmartin A, et al. 169Erbium-citrate synoviorthesis after failure of local corticosteroid injections to treat rheumatoid arthritis-affected finger joints. Clin Exp Rheumatol 2004; 22:722–726. Kampen WU, Hellweg L, Massoudi-Nickel S, Czech N, Brenner W, Henze E. Clinical efficacy of radiation synovectomy in digital joint osteoarthritis. Eur J Nucl Med Mol Imaging 2005; 32:575–580. R. Firestone. In: Shirley VS, editor. Table of isotopes. 8th ed. New York: John Wiley; 1996. pp. 2112–2114. Pillai MRA, Chakraborty S, Das T, Venkatesh M, Ramamoorthy N. Production logistics of 177Lu for radionuclide therapy. Appl Radiat Isot 2003; 59:109–118. Pandey U, Mukherjee A, Chaudhary PR, Pillai MRA, Venkatesh M. Preparation and studies with 90Y-labelled particles for use in radiation synovectomy. Appl Radiat Isot 2003; 55:471–475. Kothari K, Subramanian S, Sarma HD, Venkatesh M, Pillai MRA. 188Re labeled hydroxyapatite particles for radiation synovectomy. Appl Radiat Isot 2003; 58:463–468. Neves M, Kling A, Lambrecht RM. Radionuclide production for therapeutic radiopharmaceuticals. Appl Radiat Isot 2002; 57:657–664. Nir-El Y. Production of 177Lu by neutron activation of 176Lu. J Radioanal Nucl Chem 2004; 262:563–567.

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Original article

Relationship between cumulative radiation dose and salivary gland uptake associated with radioiodine therapy of thyroid cancer Walter Jentzen, Elke Schneider, Lutz Freudenberg, Ernst G. Eising, Rainer Go¨rges, Stefan P. Mu¨ller, Wofgang Brandau and Andreas Bockisch Aim To estimate the individual absorbed dose to the parotid and submandibular salivary glands in radioiodine therapy and its dependence from the previous cumulative therapy. Methods Fifty-five patients with differentiated thyroid carcinoma after thyroidectomy received 1–21 GBq 131 I using single activities of 1–6 GBq. The patients were stratified according to the cumulative activities into low-activity (1–2 GBq), middle-activity (3–7 GBq), and high-activity groups (9–21 GBq). The time–activity curves over the respective salivary glands were derived from multiple static calibrated images measured for each patient up to 48 h after ingestion of the radioiodine therapy capsule with a gamma camera. Manually drawn regions of interests were used to determine the background activities and the activities arising from the salivary glands. The gland volumes were determined by ultrasonography using appropriate volume models.

lower for the high-activity than for the low-activity groups and correlated with the mean cumulative administered activity of the activity groups. Conclusion The iodine uptake of salivary glands is significantly reduced, whereas the absorbed dose per administered 131I activity was not significantly decreased during the course of therapy. Comparing the well-known dose–effect relationships in external radiation therapy, the absorbed dose per administered 131I activity is too low to induce comparable radiation damage, suggesting an inhomogeneous distribution of 131I in human salivary c 2006 Lippincott glands. Nucl Med Commun 27:669–676 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:669–676 Keywords: salivary glands, radioiodine therapy, cancer

131

I, dosimetry, thyroid

Clinic for Nuclear Medicine, University of Duisburg/Essen, Germany.

Results The median absorbed dose per administered activity of each single parotid and submandibular gland was about 0.15 Gy
GBq – 1 (range, 0.1–0.3 Gy
GBq – 1) and 0.48 Gy
GBq – 1 (range, 0.2–1.2 Gy
GBq – 1), respectively. The maximum uptake of both gland types was significantly

Introduction After high-dose radioiodine therapy of differentiated thyroid carcinoma, dysfunction of the salivary glands may be seen in some patients finally causing swelling, sialadenitis or xerostomia [1–6]. Both after external radiation therapy and radioiodine therapy the functional and histological damage occurs more frequently and is more evident for the parotid than for the submandibular glands [4,7–9]. A detailed investigation of the dose–effect relationships is needed that certainly requires accurate estimates of the individual absorbed dose to the salivary glands and glandular dysfunction following radioiodine therapy. The functional alterations in salivary glands after radioiodine therapy were proven by quantitative salivary gland scintigraphy and were found to correlate with the cumulative administered activity of 131I [7,10,11].

Correspondence to Dr W. Jentzen, Clinic for Nuclear Medicine, University of Duisburg/Essen, Hufelandstrasse 55, D-45122 Essen, Germany. Tel: + 0049 201 723 4173; fax: + 0049 201 723 4143; e-mail: [email protected] Received 31 May 2005 Accepted 24 March 2006

According to the literature, a treatment with an activity of 6 GBq reveals a 30% loss of parenchymal function; a cumulative administered activity of 24 GBq results in a 90% loss of the glandular function [4,5,10]. Until now, estimates of radiation dose delivered to the salivary glands by radioiodine therapy remain uncertain. Reviewing the published data, much confusion about the dose estimates seems evident. For example, Albrecht and Creutzig [4] reported that absorbed doses in the parotid glands for patients after thyroidectomy vary between 0.2 and 15.0 Gy after administration of 7.4 GBq 131I, whereas Doniach [12] reported a dose estimate of 13.5 Gy after ingestion of 1 GBq without specifying the type of the gland. Thus, the published absorbed dose per administered activity exhibits a large variation with a range of 0.03–14 Gy
GBq – 1. This variability may hardly be explained by biological effects but more likely reflects

c 2006 Lippincott Williams & Wilkins 0143-3636

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670 Nuclear Medicine Communications 2006, Vol 27 No 8

technical problems. Thus, accurate and reliable radiation dosimetry is required. The aim of this study was to determine the absorbed dose to the parotid and submandibular glands by measuring the individual iodine kinetics in salivary glands during radioiodine therapy and its dependence from previously administered amount of radioiodine.

Methods and materials Patients

Fifty-five patients (35 females, 20 males) with differentiated thyroid carcinoma (37 papillary, 15 follicular, three Hu ¨rthle cell) after thyroidectomy were investigated. The data of three patients were acquired over two or more cycles of radioiodine therapy, representing a total of 60 time–activity curves. The ages of the female patients ranged from 25 to 77 years (mean 53 years, standard deviation 11 years); the ages of the male patients varied from 31 to 77 years (mean 58 years, standard deviation 15 years). After their first radioiodine therapy, the patients in this study underwent at least one additional radioiodine therapy, thereby effectively eliminating overlap of the salivary glands by the large thyroid remnants. The single administered activity ranged from 1 to 6 GBq as listed in Table 1. (It has to be noted that the cumulative activity given does not include the last single 131I administration.) For dosimetric purposes, the patients were stratified into three groups depending on their previously administered activity [10,11]: low-activity (1–2 GBq, presumably little damage), middle-activity (3–7 GBq, presumably moderate damage), and high-activity groups (9–21 GBq, presumably high damage) (Table 1). Radioiodine therapy was performed under conditions of salivary stimulation. On the first day, patients chewed lemon slices at several different times and were instructed to drink at least Table 1

Detailed information about the activity groups

Activity group

n*

F/M**

Activity (GBq) Cumulative***

Low Middle

High

*

9

5/4

42

25/17

9

8/1

1.0 (9) 3.0 5.0 7.0 9.0 10.5 13.0 14.8 15.0 21.0

(9) (3) (30) (3) (1) (1) (1) (2) (1)

Singlew 4.0 6.0 4.0 6.0

(8) (1) (30) (12)

1.0 (1) 4.0 (1) 6.0 (7)

Number of patients within the corresponding activity group. Ratio of female to male. *** Cumulative administered activity prior to the radioiodine therapy are listed and numbers of patients are in parentheses. The cumulative activity listed does not include the last single 131I administration (last column). w Numbers of patients receiving the corresponding single activity are in parentheses. **

2 litres of water. The patients ate lunch after 0.5–1.5 h, a snack after 2–3 h, and dinner 5–6 h after administration of radioiodine. Scintigraphy

The time-dependent activity in the salivary glands was determined by serial planar gamma camera imaging (Siemens Orbiter ZLC 370) using 131I energy window of 364 keV ± 15% and high-energy collimator. The patient sitting on a chair was positioned anteriorly to cover an area from the brain to the level of the thyroid gland. The time points chosen after pilot studies were 0.5, 1.0, 1.5, 2.0, 3.0, 4.5, 5.5, 6.0, 6.5, 7.5, 10.0, 24, and 48 h after ingestion of the radioiodine therapy capsule. Images of 5 min duration were acquired in a 128  128 matrix. Dead-time correction was not necessary and scatter correction was not performed. Since the measurements were carried out over 2 days, the stability of the equipment was checked with a 131I standard. For evaluation of the images, five regions–of–interests (ROIs) were drawn. As can be seen in Fig. 1, the first rectangular ROI was placed over the cerebrum serving as a background as proposed by Bohuslavizki and co-workers [13] and the other four ROIs were positioned over the parotid and submandibular glands. These manually drawn regions–of–interests (ROI) were found to be optimal both for the background as well as for parotid and the submandibular salivary glands obtained by a comprehensive curve fitting procedure [14]. The size and position of these ROIs were kept constant for each time point. Subsequently, the count rates obtained from the ROIs were used to calculate the activity within the salivary glands. An efficiency factor was necessary to convert the count rates into the absolute activity. For this purpose, a head phantom with lesions comparable in size and location with the parotid and submandibular glands was placed in a similar position as the patient’s head. The instrument set-up for these phantom measurements was identical with the patient measurements. The lesions were filled with known activity to measure the system calibration factor for converting net count rates to activities. This calibration factor takes into account the instrument detection efficiency and the tissue attenuation. The ROIs used for analysis of the phantom images corresponded in size to those used for the analysis of the patient images. The percentage uptake of the administered activity was calculated and used in the time–activity curves. Instead of using a single or multiple exponential decay function, the area under the time–activity curves representing the cumulated activity was determined by applying the trapezoid rule (numerical integration) because of the strong fluctuation caused by food intake. The count rates

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Dose–uptake relationship in salivary glands Jentzen et al. 671

Fig. 1

(Siemens Symphony 1.5 T, 4-mm slice thickness), delineating each gland with a ROI on each tomographic section and summing the ROI areas over the sections covering the gland. The MRI-based and US-based volumes were compared to evaluate the volume models. Calculation of absorbed dose

The mean self-irradiation (gland-to-gland) absorbed dose was calculated using the MIRD formalism assuming a uniform 131I activity distribution. Only the absorbed dose deposited by the non-penetrating b-ray radiation was considered, whereas the contribution of the penetrating g-ray radiation was neglected, which contributes 10% of the total absorbed dose for target regions comparable in volume (about 20 ml) to human salivary glands. Therefore, the self-absorbed dose, D, was calculated as follows: D

D ~ A rV

The symbol A~ represents the cumulated activity, D the equilibrium dose constant for non-penetrating radiation, r the gland mass density assuming the value of water (1 g
ml – 1), and V the volume of the gland (ml). For 131I, the value of the equilibrium dose constant D is 0.11 (Gy g)
(MBq h) – 1 [15]. Statistics

Scintigram of the salivary glands acquired 0.5 h after ingestion of 6 GBq 131I. The five regions of interest (ROIs) are shown. The top one over the brain was used to correct for background. The other four are the ROIs over the parotid (middle) and submandibular glands (bottom).

beyond 48 h were small and its contribution to the cumulated activity (less than 1%) was neglected.

The data were given as the mean and standard deviation as well as the median and, graphically only, the 10th, 25th, 75th and 90th percentile values. Differences among the activity groups were evaluated by the Kruskal–Wallis one-way analysis of variance (ANOVA) and the pairwise differences (between-group analysis) are tested by Dunn’s method. Statistically significant differences were assumed at P-values < 0.05.

Results For a complete athyreotic patient, the reliability of the gamma camera-based salivary gland uptake measurements was checked by comparing the total salivary gland uptake measured by the gamma camera method with the results of NaI(Tl) thyroid uptake probe; the two measurements agreed within 10%. Gland volume

Gland dimensions were determined by ultrasonography (US) using a B-mode device (CS 9150 Picker) and a linear 7.5-MHz transducer, with its axis held at two orthogonal directions to measure the width, length and depth of the gland. The gland volume was calculated assuming an ellipsoidal shape – accurate for the submandibular but not for the parotid gland. The volume of the irregularly shaped parotids was estimated assuming either an ellipsoid or a cylindrical shape. The volume models were evaluated using magnetic resonance imaging (MRI) serving as reference. Specifically, the gland volumes of five patients were determined with MRI

Volume of the salivary glands

The volumetric data of the parotid and submandibular glands are listed in Table 2 using MRI or US. The mean parotid and submandibular gland volumes of five patients evaluated by MRI were 25 and 9 ml, respectively. The parotid volume based on US using the cylindrical model was 21 ml but 61 ml using the ellipsoidal model. The submandibular volume determined by US (ellipsoidal model only) was 7 ml. It has to be noted that US-based volumes for both gland types was, except two of ten, smaller than the MRI-based volumes. The maximum differences calculated between the MRI and US volumes were – 40% for the submandibular gland (ellipsoidal model) and – 37% for the parotid gland using the cylindrical model. Therefore, the cylindrical model for the parotid gland and the ellipsoidal model for the submandibular gland were applied to determine the US-based volumes of each patient. These individual US-based volumes were used in

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672 Nuclear Medicine Communications 2006, Vol 27 No 8

the dosimetric calculation. The mean individual volumes and standard deviations obtained from 55 patients (including left and right sides) were 18 ± 5.8 ml (range 4–35 ml) and 9.9 ± 5.0 ml (range 3–27 ml for the parotid and submandibular salivary glands, respectively. Time–activity curve

Representative time–activity curves for the three groups are presented in Fig. 2. The large variation in the uptake values appears to be related to food intake, decreasing with time to a greater extent with chewing and salivation.

Table 2 Volumetric data of the salivary glands by using different methods and volume models obtained from a subset of five patients* Salivary gland

MRI (ml)

Parotid Submandibular

left right Left

24.4 ± 8.5 24.8 ± 9.2 8.6 ± 2.2

right

8.8 ± 2.8

US (ml) Cylinder**

Ellipsoid

21.2 ± 7.5 20.7 ± 5.9

60.2 ± 9.4 60.8 ± 7.6 8.7 ± 1.1 7.3 ± 2.6

*

The mean values and their corresponding single standard deviations are listed. A volume underestimation of the US-based volumes except two of ten was observed. ** Because of the irregular shape of the parotid glands, both the ellipsoidal and cylindrical formulae were used. The volumes of the submandibular glands were only approximated by the ellipsoidal formula.

As a rule the submandibular glands appeared earlier than the parotid glands after ingestion of the radioiodine capsule, indicating differences in iodine kinetics between the two-paired glands. Figure 3 shows box plots of the maximum uptake obtained from the corresponding time–activity curves of each activity group. For the parotid glands (Fig. 3a), the average (median) maximum uptake and their single standard deviation for the low-activity, middle-activity and high-activity group was 0.28 (0.26) ± 0.07%, 0.23 (0.22) ± 0.06%, and 0.19 (0.18) ± 0.05%, respectively. The maximum uptake showed a significant difference between the activity groups (P-value < 0.0002, Kruskal– Wallis analysis of variance). A significant difference between the low-activity and high-activity group (Pvalue < 0.01, Dunn’s test) as well as low-activity and middle-activity group (P-value < 0.05, Dunn’s test) can be stated. For the submandibular gland (Fig. 3b), the average (median) maximum uptake and their single standard deviations for the low-activity, middle-activity and high-activity groups was 0.26 (0.26) ± 0.04%, 0.26 (0.25) ± 0.07%, and 0.21 (0.20) ± 0.05%, respectively. The maximum uptake also showed a significant difference between the activity groups (P-value < 0.006, Kruskal–Wallis analysis of variance). Significant differences were calculated between the low-activity and high-

Fig. 2

(a) 0.5

(b)

0.5 Left Right

Gl. parotid

0.4

Uptake (%)

(c)

0.4

0.3

0.3

0.2

0.2

0.1

0.1

0.0

0.0 Low

Gl. subm.

0.4

Middle

High

0.4

0.3

0.3

0.2

0.2

0.1

0.1

0.0

0.0 0

2

4

6

8 10

30 40

0

2

4

6

8 10

30 40

0

2

4

6

8 10

30 40

Time (h) Representative uptake curves of 131I in the left (&) and right (*) parotid (top) and submandibular glands (bottom) of the low (panel a), middle (panel b), and high-activity group (panel c).

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Dose–uptake relationship in salivary glands Jentzen et al. 673

Fig. 3

Fig. 4

0.30

0.40 (a) 0.35

3 −7 GBq 9 −21 GBq

0.25

Maximum uptake (%)

0.20

1 GBq

0.15 Gl. parotid

Median maximum uptake (%)

0.30

0.25

0.20

0.15 Gl. subm. 0.10

0.10 (b)

0.05

0.35 0.30

Gl. parotid

3 −7 GBq 1 GBq

9 −21 GBq

0.25

0.00 0

5

10 15 20 25 30 35 Mean cumulative activity (GBq)

40

Median maximum uptake of the single glands versus the mean cumulative administered activity of the low-activity, middle-activity and high-activity group. The extrapolated line (dotted) was obtained from data points between 1 and 10 GBq (solid line) using linear regression.

0.20 0.15 Gl. subm. 0.10 Low

Middle Activity group

High

Box plot of the maximum 131I uptake (average of the left and right glands) for the parotid (top) and submandibular glands (bottom) of the low-activity, middle-activity and high-activity group. The values were taken from the time–activity curves. The errors bars are the 10th and 90th percentiles, the grey box itself is the boundary of the 25th and 75th percentiles, and the solid line indicates the median and the dashed line the mean value.

activity group (P-value < 0.05, Dunn’s test) as well as between middle-activity and high-activity group (P-value < 0.01, Dunn’s test). The plot of the median (or average) maximum uptake versus the mean cumulative administered activity exhibits a linear relationship for both the parotid (r = 0.99) and submandibular gland (r = 0.91) (Fig. 4). By extrapolation using data points between 1 and 10 GBq, the abscissa intercepts indicate the complete elimination of salivary gland function (dotted line). The complete loss of radioiodine uptake roughly occurs at an absorbed dose of about 35 Gy. In the absence of parameters for a realistic sigmoidal dose response, a simple linear relationship was assumed. As a consequence, this underestimates the dose necessary for the complete loss of radioiodine uptake in agreement with clinical experience.

Absorbed dose per administered activity

Figure 5 shows the absorbed dose per administered individual activity for the parotid and submandibular glands separated in low-activity, middle-activity and highactivity group. The absorbed dose per administered activity especially for the submandibular glands showed a large variation. Unexpectedly, there was no statistically significant difference among the activity groups for both the parotid (P-value < 0.244, Kruskal–Wallis analysis of variance) and submandibular glands (P-value < 0.052, Kruskal–Wallis analysis of variance). The mean (median) and standard deviation for each single parotid and submandibular gland were 0.18 (0.15) ± 0.09 Gy
GBq – 1 (range, 0.10–0.30 Gy
GBq – 1) and 0.57 (0.48) ± 0.40 Gy
GBq – 1 (range, 0.20–1.20 Gy
GBq – 1), respectively. Error estimates of the absorbed dose per administered activity

Several parameters had an influence on the absorbed dose calculation. The main errors were related to volumetric determination of the gland as well as the background ROIs. As described above, the maximum error in the volumetric determination was about – 50% (an underestimation) for both gland types. The maximum error in the background activity was estimated to be about 30%. Thus, both errors can yield a combined error of about 60% using Gaussian error propagation. However, this error estimate is an upper limit and, on average, is certainly

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674 Nuclear Medicine Communications 2006, Vol 27 No 8

Fig. 5

(a)

Gl. parotid 0.4

0.2

0.0 (b)

Absorbed dose (Gy/GBq)

Gl. subm. 3 −7 GBq

1.2

9 − 21 GBq

1.0

0.8

or 4 Gy for the parotid gland and 3 or 12 Gy for the submandibular gland. It should be noted that the applied cumulative dose is not the only factor with possible impact on tissue damage because the activities were administered at variable time intervals. Using the dose–response relationship for salivary glands after external radiation therapy, it is also possible to estimate the absorbed dose in radioiodine therapy based on the observed functional effect on the glands. A complete loss of salivary gland function from external radiation therapy occurs at a dose level of 60 Gy [16–18]. The corresponding cumulative administered activity of radioiodine would be about 35 GBq (Fig. 4) that an absorbed dose per administered 131I activity of 1.7 Gy
GBq – 1.

1 GBq

0.6

0.4

0.2

0.0 Low

Middle Activity group

High

Box plot of the absorbed dose per administered activity (average of the left and right glands) for the parotid (top) and submandibular glands (bottom) of the low-activity, middle-activity and high-activity group. Symbols are as in Fig. 3.

lower. In addition, the uncertainty arising from the spikes caused by the food intake in the time–activity curves produced statistical errors in the cumulated activity within the population.

Discussion It is well known that there is a correlation between gland dysfunction and cumulative administered activity and/or absorbed dose [7,10,11]. Specifically, if the patient receives a single activity of 6 GBq, results in a 30% loss of parenchymal function and a cumulative administered activity of 24 GBq results in a 90% loss of the glandular function as determined by salivary gland scintigraphy [4,5,10]. As can be seen from Fig. 5, the median absorbed dose using the MIRD formalism is 0.15 Gy
GBq – 1 and 0.48 Gy
GBq – 1 for the single parotid and submandibular gland, respectively. Thus, the corresponding absorbed doses assuming 6-GBq or 24-GBq activities were about 1

Since the biological effectiveness of external radiation and radioiodine therapy are not equivalent, the estimated absorbed dose per administered activity is even larger. In terms of growth delay of tumour xenografts in mice, radioimmunotherapy with 131I labelled ‘anti-tumour’ antibodies is only about a third as effective as external radiation therapy [19]. Thus, an absorbed dose per administered activity of about 5 Gy
GBq – 1 would actually be required to account for the observed functional effect of radioiodine therapy on salivary glands. This estimated value of 5 Gy
GBq – 1 is significantly larger than the measured value between 0.15 and 0.48 Gy
GBq – 1. Attempting to resolve the discrepancy between the dose– effect relationship for radioiodine therapy and external radiation therapy, a possible explanation may be inhomogeneous distribution of radioiodine, as demonstrated by autoradiography of animal tissues [20–22] within the salivary glands. In guinea pig glands, for example, iodine and technetium were concentrated in the duct cells but not in the acinar cells. In addition, because about 90% of the gland mass is accounted for by acinar cells, the mass of the radioiodine-targeted volume (i.e., the duct cells) is, in the first order, about 90% smaller and the absorbed dose therefore about 90% larger than for the salivary gland as a whole. A small-scale dosimetry analysis is likely required to derive reliable dose–response relationships for salivary gland dysfunction associated with radioiodine therapy. Another possible explanation for the obvious discrepancy could be the large uncertainties associated with the measurements. However, even if the maximum error is about 60% primarily caused by uncertainties in gland volume and background activity, the absorbed doses are still an order of magnitude too small to account for the observed radiation damage in radioiodine therapy. Moreover, the strong fluctuation of the time–activity curves caused by food intake cannot explain the large deviation

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Dose–uptake relationship in salivary glands Jentzen et al. 675

between the observed and expected absorbed dose per administered activity. As a consequence, further investigations are necessary with more reliable quantitative imaging (see below). Another pertinent issue is the correlation between salivary gland dysfunction and the cumulative administered activity. The functional impairment of the glands as indicated by maximum radioiodine uptake correlates well with the cumulative administered activity (Figs 3 and 4) and is consistent with published data [4,23]. However, there is no significant difference in the absorbed dose per administered activity among the three activity groups. Thus, a significantly decreasing maximum uptake with cumulative activity is observed, yet the absorbed dose per administered activity after increasing iodine dose seems to be similar in the activity groups. It is likely that individual changes in absorbed dose are too variable to detect altered gland function among the administered-activity groups. Note, for example, the large ‘spikes’ in the salivary gland time–activity curves (Fig. 2). Nevertheless, the absorbed dose to the parotid and not to the submandibular glands has a tendency, although not significant, to decrease from the low-activity to the highactivity group, presumably reflecting radiogenic impairment of function (Fig. 5). This tendency, including the significant larger change in the maximum uptake from the low-activity to the high-activity group, is more likely to occur for the parotid gland. It has been reported that the radiosensitivity (in external radiation and radioiodine therapy) for the parotid gland is higher than for the submandibular gland [4,7–9]. There are at least two hypotheses regarding the apparent difference in radiosensitivity between the parotid and submandibular salivary glands (Fig. 5) [24–27]. The first is related to anatomical differences between the two glands. Specifically, the parotid gland mainly contains serous cells, whereas the submandibular gland consists of a mixture of serous and mucinous cells. Because the mucinous-secreting glands increase their secretion rate after irradiation, the submandibular gland is deemed to be less radiosensitive than the parotid glands. The second hypothesis is based on the difference in the kinetics of the two types of salivary glands, i.e., the submandibular glands have a higher continuous unstimulated secretion than the parotid gland, possibly reducing sensitivity. The measured absorbed dose cannot explain the apparent difference in radiosensitivity (absorbed dose to the submandibular gland is evidently higher than that to the parotid gland) and thus corroborates the hypothesis that the differences in radioiodine secretion and cell composition between the parotid and submandibular salivary glands have predominant impact on relative radiosensitivities.

Admittedly, the results and interpretations in this study appears to be limited because of the uncertainties regarding gland volume, background activities, food intake pattern as well as the poor quantitative accuracy of the scintigraphic 131I image. Nonetheless, taking into account all possible errors, the absorbed dose per administered activity calculated with the MIRD formalism is at least an order of magnitude too small to account for the observed radiation damage in radioiodine therapy. Further investigations are still required to substantiate the findings in this paper. The use of multimodality imaging such as positron emission and computed tomography, PET/CT, for the determination of the activity concentration and delineation of target volumes is a promising concept for estimating the absorbed dose to the salivary glands using the non-pure positron emitter 124 I.

Conclusion The maximum uptake of the parotid and submandibular gland as a functional marker is significantly lower for the high-activity than for the low-activity group. Thus, an accurate estimate of the individual radiation burden of salivary glands requires measurements and cannot be deduced from the measured absorbed dose found in this study. Comparing the well-known dose–effect relationships in external radiation therapy with radioiodine therapy, the absorbed dose per administered activity calculated with the MIRD formalism is at least an order of magnitude too small to account for the observed radiation damage in radioiodine therapy. This finding probably indicates an inhomogeneous activity distribution of 131I in human salivary glands.

Acknowledgements The assistance of the engineers Wilfried Sonnenschein and Dietmar Wedeleit supporting the measurement on the gamma camera is gratefully acknowledged. We thank Dr J. Kanja for helpful suggestions and discussions.

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Allweiss P, Braunstein GD, Katz A, Waxman A. Sialadenitis following I-131 for thyroid carcinoma: concise communication. J Nucl Med 1984; 25:755–758. Goolden AWG, Mallard JR, Farran HEA. Radiation sialitis following radioiodine therapy. Br J Radiol 1957; 30:210–212. Hilton G. The role of radioiodine in the treatment of carcinoma of the thyroid. Br J Radiol 1956; 29:297–310. Albrecht HH, Creutzig H. Funktionsszintigraphie der Speicheldru¨sen nach hochdosierter Radiojodtherapie. Fortschr Ro¨ntgenstr 1976; 125:546–551. Reiners C, Eilles C, Eichner R, Spiegel W, Bo¨rner W. Speicheldru¨senFunktionsszintigraphie zur Verlaufskontrolle bei der Therapie des Schilddru¨sen-Karzinoms mit Radiojodid. Nuklearmedizin 1980; 3:281–286. Spiegel W, Reiners C, Bo¨rner W. Einschra¨nkung der Speicheldru¨senfunktion nach hochdosierter Radiojodtherapie. Nuklearmediziner 1986; 9:159–166. Caglar M, Tuncel M, Alpar R. Scintigraphic evaluation of salivary gland dysfunction in patients with thyroid cancer after radioiodine treatment. Clin Nucl Med 2002; 27:767–771. Kashima HK, Kirkham WR, Andrews JR. Postirradiation sialadenitis. Am J Roentgenol 1965; 94:271–291.

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Sinn DF, Stocker NG, Epker BN. Effects of fractionated doses of cobalt 60 irradiation on rabbit submandibular glands: light microscopic studies. J Org Surg 1972; 30:277–283. Bohuslavizki KH, Brenner W, Lassmann S, Tinnemeyer S, Kalina S, Clausen M, Henze E. Quantitative salivary gland scintigraphy – a recommended examination prior to and after radioiodine therapy. Nuklearmedizin 1997; 36:103–109. Bohuslavizki KH, Brenner W, Lassmann S, Tinnemeyer S, Tonshoff G, Sippel C, et al. Quantitative salivary gland scintigraphy in the diagnosis of parenchymal damage after treatment with radioiodine. Nucl Med Commun 1996; 17:681–686. Doniach I. Biological effects of radiation on the thyroid. In: Werner S (editor): The Thyroid. New York: Harper & Row; 1978, pp. 274–283. Bohuslavizki KH, Brenner W, Lassmann S, Sippel C, Wolf H, Clausen M, Henze E. Value of quantitative salivary gland scintigraphy in early stage of Sjo¨rgren’s syndrome. Nucl Med Commun 1995; 16: 917–922. Schneider E. Zusammenhang zwischen den kumulativen absorbierten Dosen und den Funktionssto¨rungen der Speicheldru¨sen bei der Radiojodtherapie von differenzierten Schilddru¨senkarzinomen. Dissertation, University of Duisburg/Essen, Germany, 2006. Snyder WS, Ford MR, Warner GG, Watson SB. ‘‘S,’’ Absorbed Dose Per Unit Cumulated Activity for Selected Radionuclides and Organs. MIRD Pamphlet 11. New York, NY: Society of Nuclear Medicine; 1975. Valdez IH, Atkinson JC, Ship JA, Fox PC. Major salivary gland function in patients with radiation-induced xerostomia: flow rates and sialochemistry. Int J Radiat Oncol Biol Phys 1993; 25:41–47. Franzen L, Funegard U, Ericson T, Henriksson R. Parotid gland function during and following radiotherapy of malignancies in the head and neck, a consecutive study of salivary flow and patient discomfort. Eur J Cancer 1992; 28:457–462.

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Original article

Evaluation of early versus delayed lymphoscintigraphic imaging in detecting internal mammary sentinel lymph nodes in breast cancer: A multicenter study to establish an optimal lymphatic mapping protocol Esther M. Heutsa, Fred W.C. van der Enta, Harry A.G. van der Polb, Joop M.H. Debetsc, Roland A.M. Kengend, Joost M.A. Verkeyne, Karel W.E. Hulsewe´a and Anton G.M. Hoofwijka Objective Metastases in the internal mammary lymph nodes have an important prognostic value in breast cancer. Lymphatic mapping and sentinel node biopsy of internal mammary nodes improves staging and permits specific therapeutic strategies, thereby possibly improving final outcome. Therefore, optimal lymphoscintigraphic results are needed. Visualization of internal mammary lymph drainage, however, is influenced by several factors. We evaluated the effect of different time intervals between radioactive tracer injection and lymphoscintigraphy on visualization of internal mammary sentinel lymph nodes.

showed internal mammary hotspots in 21% in group A, compared to 27% in group B. The mean number of internal mammary hotspots per patient was 1.9 in group A and 1.8 in group B. Conclusions We found no significant differences between early and delayed lymphoscintigraphic imaging in visualizing internal mammary sentinel lymph nodes. Nucl c 2006 Lippincott Williams & Med Commun 27:677–681 Wilkins. Nuclear Medicine Communications 2006, 27:677–681

Methods From February 1997 to August 2001 a total of 682 eligible breast cancer patients underwent sentinel lymph node mapping. The technique involved the injection of 370 MBq (10 mCi) 99mTc-nanocolloid peritumorally. In 470 patients (group A) the time interval between injection of the radiocolloid and lymphoscintigraphy was 16 h, compared to 2.5 h in 212 patients (group B). Results Patient characteristics showed no statistically significant difference between both groups for age and location of the tumour. Axillary hotspots were visualized in 97% in group A and 96% in group B. Lymphoscintigraphy

Introduction Since the introduction in 1993 of the sentinel node concept, it quickly evolved into a generally accepted and worldwide applied method for staging in breast cancer [1]. Many validation studies confirmed the accuracy of the sentinel node biopsy in predicting the status of the axillary lymph nodes [2–6]. Axillary lymph node dissection is nowadays abandoned in cases of negative pathological findings in the axillary sentinel node. The technique of lymphatic mapping inevitably confronts us with the visualization of non-axillary lymphatic draining patterns, and the question of their clinical relevance. Previous studies showed that the tumour status of the internal mammary sentinel nodes represents a major independent prognostic factor in breast cancer

Keywords: sentinel node biopsy, breast cancer, lymphoscintigraphy, internal mammary lymph node Departments of aSurgery and, bNuclear Medicine, Maaslandziekenhuis, Sittard, Departments of cSurgery and, dNuclear Medicine, Laurentius Ziekenhuis, Roermond and eDepartment of Surgery, St Jans Gasthuis, Weert, the Netherlands. Correspondence to Dr Esther M. Heuts, Department of Surgery, Maaslandziekenhuis, Walramstraat 23, 6130 MB Sittard, The Netherlands. Tel: + 0031 464 597777; fax: + 0031 464 597975; e-mail: [email protected] Received 12 April 2006 Accepted 16 May 2006

[7]. Although axillary sentinel nodes are routinely evaluated, internal mammary lymph nodes are not routinely biopsied or included in staging. As for axillary sentinel node biopsy, harvesting of the internal mammary nodes, based on lymphoscintigraphy, will have implications for staging and treatment [8]. The results of preoperative lymphoscintigraphy therefore should be optimal. However, the visualization rate of internal mammary nodes on lymphoscintigraphy is influenced by several different factors. In order to establish an optimal internal mammary imaging protocol within the complex logistics of planning preoperative lymphoscintigraphy and sentinel node surgery, we conducted a study to determine whether different time intervals between injections of a radioactive tracer and

c 2006 Lippincott Williams & Wilkins 0143-3636

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678 Nuclear Medicine Communications 2006, Vol 27 No 8

performing lymphoscintigraphy affect the visualization rate of internal mammary hotspots.

Table 1

Methods

Mean age (years)

Based on a prospective database, following approval of the local ethics committee and after obtaining informed consent, a cohort study was performed in 682 clinically node-negative T1–3 breast cancer patients. All patients underwent preoperative lymphatic mapping for sentinel node biopsy from February 1997 to August 2001. Data for group A, consisting of 470 consecutive patients, were collected from a single institution. Group B consisted of 223 patients from two other hospitals in the same region. In the latter, insufficient data for analysis were available in 11 patients, resulting in 212 eligible patients in group B. A 2-day protocol for sentinel node biopsy was administered for both groups. Lymphatic mapping was performed by the same nuclear medicine physicians for both groups, preventing bias in individual technical performance. Our technique of sentinel node biopsy has been described in detail elsewhere [2]. The day before surgery, in both groups 370 MBq (10 mCi) 99mTc-nanocolloid in 4 ml saline was injected peritumorally in three to four depots around the tumour or in the breast parenchyma surrounding the cavity of a previous excisional biopsy. For group A, delayed lymphoscintigraphy was performed the next day, after a mean period of 16 h (range 16–18 h ) following radiotracer injection, and shortly before surgery. For group B, early lymphoscintigraphic images were obtained the same day after a mean period of 2.5 h (range 2–4 h) after the injection of the radiocolloid tracer and surgery was performed the next day. Scintigraphic images in all patients were obtained in three positions: anterior, anterior–oblique and lateral in order to visualize the number of sentinel nodes and their axillary and non-axillary position. Statistical analysis was performed using Student’s t-test for continuous variables and for discrete variables chisquared analysis was used. A P-value < 0.05 was considered statistically significant.

Results Lymphatic mapping and sentinel node biopsy was performed in 470 patients in group A and in 212 patients in group B. Patient characteristics are listed in Table 1. Comparing the two groups, no statistical significance was reached for age and location of the tumour. Tumour diameter was also similar, but histology showed more lobular tumours in group A and more ductal carcinomas in group B. The results of preoperative lymphoscintigraphy with visualization of axillary and internal mammary hotspots

Patient characteristics of group A and B

Characteristic

Tumour characteristics Diameter < 20 mm 20–49 mm > 49 mm Ductal carcinoma Lobular carcinoma Other Left Right Breast quadrant Upper outer Upper inner Lower outer Lower inner Central Unknown

Table 2

Delayed imaging group A (n = 470)

Early imaging group B (n = 212)

59 (range 28–96) SD 12.0

58 (range 30–87) SD 12.9

221 (47%) 214 (46%) 35 (7%) 324(69%) 112 (24%) 34 (7%) 262 (56%) 208 (44%) 216 102 58 55 36 3

115 90 7 177 27 8 106 106

(54%) (42%) (3%) (83%) (13%) (4%) (50%) (50%)

99 41 29 17 22 4

(47%) (19%) (14%) (8%) (10%) (2%)

(46%) (22%) (12%) (12%) (7%) (1%)

Lymphoscintigraphic visualization of hotspots in group A

and B Characteristic Patients with axillary hotspots Total axillary hotspots Mean axillary hotspots (range) Patients with additional internal mammary hotspots Total internal mammary hotspots Mean internal mammary hotspots (range) Only internal mammary hotspots

Delayed imaging group A (n = 470)

Early imaging group B (n = 212)

P-value

454 (97%)

203 (96%)

0.75

629 1.4(1–7) SD 1.0

369 1.7(1–5) SD 1.0

99 (21%)

58(27%)

191

107

1.9 (1–6) SD 1.0

1.8 (1–5) SD 0.9

0

0

0.09

in both groups are listed in Table 2. Detection of axillary hotspots on lymphoscintigraphy was almost equal in both groups (97% in group A and 96% in group B). The lymphoscintigraphic detection of internal mammary hotspots was 21% (99/470) after delayed imaging (group A) and 27% (58/212) after early imaging (group B). Drainage to the internal mammary lymph nodes originated from laterally located tumours in 41% (41/99) and in 59% (58/ 99) for central or medially located tumours in group A. For group B, internal mammary drainage originated from laterally located tumours in 28/58 (48%) patients and from central or medially located tumours in 30/58 (52%). The mean number of internal mammary hotspots per patient was 1.9 in group A and 1.8 in group B. The differences in the rate of internal mammary visualization did not reach statistical significance (P-value > 0.05). The mean age of patients showing internal mammary drainage was 54 years in group A and 56 years in group B. The detection of internal mammary hotspots was influenced by age, as is shown in Table 3. The proportion

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An optimal lymphatic mapping protocol in breast cancer Heuts et al. 679

Table 3 Correlation between age and internal mammary hotspots on lymphoscintigraphy in group A and B Age (years)

Delayed imaging group A (n = 470) n

< 40 41–50 51–60 61–70 > 70

32 114 126 95 103

Internal mammary hotspots 12 40 19 15 13

(38%) (35%) (15%) (16%) (13%)

Early imaging group B (n = 212) n 11 47 64 50 40

Internal mammay hotspots 4 19 18 8 9

(36%) (40%) (28%) (16%) (23%)

of patients with internal mammary hotspots decreases with age. This effect was seen in both groups.

Discussion The lymphatic drainage of the breast has been studied intensively. Halsted identified the internal mammary chain as a route of metastases in breast cancer in 1907. Many subsequent studies have confirmed the importance of the internal mammary basin as a second draining route in breast cancer [9,10]. As a consequence, internal mammary lymph node dissection was part of the standard surgical treatment in the 1950s and 1960s. Studies concerning this radical surgical treatment are the main source of information on the extent of nodal involvement in the internal mammary basin. Survival rates proved to be worse in case of internal mammary metastases [7,11]. During the 1970s internal mammary dissection was abandoned, based on patient outcome studies, which showed unfavourable survival rates, but no improvement on prognosis after radical dissection of the internal mammary chain. Following the introduction of the sentinel node technique, however, the internal mammary basin has regained interest. In patients with documented internal mammary lymphatic drainage, internal mammary sentinel node biopsy adds to adequate staging by identifying internal mammary node-tumour positive patients. The less favourable prognosis in these patients thus can be influenced by tailoring their individual adjuvant treatment programmes [8], in order to benefit final outcome. Optimal visualization of both axillary and non-axillary lymph drainage patterns is essential in reaching accurate lymphatic mapping and staging. Whether a 1-day protocol or a 2-day protocol is used is often a matter of logistic preference, but neither should have a negative effect on the lymphoscintigraphic results. In a 1-day protocol, tracer injection, lymphoscintigraphy and sentinel node biopsy are performed on the same day, thereby avoiding the patient’s need for an extra hospital visit. In a 2-day protocol, tracer injection can be followed by lymphoscintigraphy on the same day or the next day, i.e., the day of surgery. A 2-day protocol has advantages both in scheduling multiple patients for lymphoscintigraphic imaging as

well as in planning the operating room schedule, facilitating the planning of multiple sentinel node procedures at the same day. Moreover, a 2-day protocol has radiation safety advantages, showing a significantly decreased dosage of radioactivity in the resected specimen [12] and a smaller absorbed radiation dose to the hands of the medical staff due to radioactive decay [13,14]. Late lymphoscintigraphic imaging on the day of surgery has the advantage of visualizing the sentinel node without the risk of registering confusing temporary radiocolloid accumulations in lymphatic tracts. It has been suggested that a longer interval between radioisotope injection and scanning results in a greater number of internal mammary sentinel nodes on the lymphoscintigraphy [15,16]. While using a 2-day protocol in both group A and group B, we analysed the effect of early lymphoscintigraphic imaging (group B) as compared to delayed imaging (group A) in visualizing internal mammary sentinel nodes. In this study, we found no statistical significant differences between internal mammary sentinel node visualization in group A (21%) and group B (27%). Hence, we could not demonstrate a correlation between timing of the radiotracer injection and the lymphoscintigraphic detection of internal mammary sentinel nodes. However, there is a wide range of reported visualization of internal mammary hotspots in lymphatic mapping for sentinel node biopsy. Published results differ mainly, because the detection rate is influenced by other several technical, anatomical and biological factors. The most important factor is the site of tracer injection. While aiming for improved accuracy, various injection sites for the radiocolloid are used, such as intracutaneous, subareolar, peritumoral, intratumoral, subtumoral and periareolar injections. Although axillary lymph drainage patterns are independent of the site of tracer injection [5,17], internal mammary lymph drainage is not. Visualization of the internal mammary nodes is rarely seen following intra-cutaneous or subcutaneous, subareolar or periareolar tracer injections, as compared to peritumoral, intratumoral or subtumoral tracer injections. This suggests that the breast parenchyma drains to both the axilla and the internal mammary chain, but the skin overlying the breast only has a lymphatic drainage to the axilla. Shimazu et al. [18] and Paganelli et al. [19] compared superficial and deep parenchymatous injection of a tracer. Internal mammary hotspots were more frequently seen after deep tracer injections. This was associated with more internal mammary metastases found for deep located tumours, compared to superficially located tumours. A second factor that influences the visualization of internal mammary lymphatic drainage is the localization

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680 Nuclear Medicine Communications 2006, Vol 27 No 8

of the tumour. Anatomical considerations suggest that medially located intraparenchymatous injections will result in an increased rate of internal mammary hotspots. However, the results in the literature are inconsistent. Early anatomical studies by Hultborn et al. [9] and Turner-Warwick [10] conclude that both the medial and lateral halves of the breast show similar drainage rates to the internal mammary chain. Our own previously published results showed internal mammary drainage in 65 (25.3%) of 256 patients, in 63% draining medial or central tumours, and in 37% draining lateral tumours [8]. These results are in accordance with other studies [19,20], which confirm that tracer administration in the medial or central quadrants of the breast parenchyma is associated with increased rates of internal mammary hotspots. However, it is possible that these findings have no relationship with tumour location itself, but can be explained by what Krynyckyi et al. [21] called a proximity effect. These authors noted that in small breasts the chances of detecting internal mammary sentinel nodes were much higher, since the lesion and therefore the tracer injection would be much closer to the chest wall and the deeper lymphatic channels that course to the internal mammary sentinel nodes. This hypothesis is concordant with the effects of deep subtumoral tracer injection as mentioned by Shimazu et al. [18]. A third factor influencing parasternal hotspot visualization is the particle size of the different tracers. After injection, a colloid tracer is absorbed into the lymphatics and transported through the lymph channels. Smaller particles flow more easily through the lymphatics and are retained in the macrophages of the lymph node for a relatively short period, as compared to larger colloids [18,22]. While using radioactive gold (198Au), which has a small particle size, Hultborn et al. [9] described parasternal radioactive uptake in all patients. Using 99m Tc-antimony sulfide, Uren et al. [23] reported parasternal drainage in 35% of his patients. In Europe, 99mTcnanocolloid is used with a particle size larger than radioactive gold and 99mTc-antimony sulfide colloid. The 99mTc-sulfur colloid, commonly used in the United States, has the largest particle size of the radiocolloid tracers mentioned. Consequently, these differences in particle size contribute in the varying reports regarding lymphoscintigraphic visualization of internal mammary hotspots. Yet another factor that influences the pattern and speed of lymph drainage is increasing age [24–26]. It appears that patient age is inversely correlated with the ability to identify axillary sentinel nodes. This is related to the induction of fatty degeneration of breast parenchyma in postmenopausal women. The lymphatic capillaries of the

breast are confined to the ducto-lobular complexes, in contrast to fatty tissue which contains scarcely any lymphatics. We evaluated the effect of age on the visualization of internal mammary hotspots. Our results confirmed that, like axillary drainage, internal mammary drainage is affected by increasing age (Table 3). Besides all the above-mentioned issues that influence visualization of parasternal hotspots, it is suggested that the ability to retain radioactivity is different for axillary lymph nodes, as compared to parasternal lymph nodes [18]. Radiocolloid is said to be retained for a shorter time in the internal mammary sentinel node than in the axillary sentinel node. If so, this would suggest that less internal mammary nodes would be seen in group A patients, based on the longer time interval between radioactive tracer injection and lymphoscintigraphy. Indeed, we found fewer internal mammary hotspots in group A (21%) as compared to group B (27%), but the difference between both groups was not statistically significant. Another variable which could affect internal mammary node visualization is the amount of radioactivity used. However, our results can not distinguish whether increased tracer activity is associated with node visualization because all patients received the same dose (i.e., 370 MBq) of 99mTc-nanocolloid. Published data by Estourgie et al. [27], who used an intratumoral injection of 111 MBq (3 mCi) 99mTc-nanocolloid showed internal mammary hotspots in 22% (150/691), which is comparable to our results.

Conclusion Internal mammary metastases are increasingly being recognized as having important prognostic significance in breast cancer. Since internal mammary sentinel node biopsy is feasible, and capable of identifying these high-risk internal mammary node-positive patients, factors influencing parasternal hotspot visualization have regained interest. In search of the optimal logistical and technical protocol for internal mammary lymphatic mapping, we evaluated the effect of different time interval between radioactive tracer injection and lymphoscintigraphy on visualization of internal mammary sentinel nodes. We found no statistically significant differences between early and delayed lymphoscintigraphic imaging protocols in this study.

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Mansel RE, Goyal A, Newcombe RG. Internal mammary node drainage and its role in sentinel lymph node biopsy: the initial ALMANAC experience. Clin Breast Cancer 2004; 5:279–284. Zavagno G, Rubello D, Franchini Z, Meggiolaro F, Ballarin A, Casara D, et al. Axillary sentinel lymph nodes in breast cancer: a single lymphatic pathway drains the entire mammary gland. Eur J Surg Oncol 2005; 31:479–484. Shimazu K, Tamaki Y, Taguchi T, Motomura K, Inaji H, Koyama H, et al. Lymphoscintigraphic visualization of internal mammary nodes with subtumoral injection of radiocolloid in patients with breast cancer. Ann Surg 2003; 237:390–398. Paganelli G, Galimberti V, Trifiro G, Travaini L, De Cicco C, Mazzarol G, et al. Internal mammary node lymphoscintigraphy and biopsy in breast cancer. Q J Nucl Med 2002; 46:138–144. Bevilacqua JL, Gucciardo G, Cody HS, MacDonald KA, Sacchini V, Borgen PI, et al. A selection algorithm for internal mammary sentinel lymph node biopsy in breast cancer. Eur J Surg Oncol 2002; 28:603–614. Krynyckyi BR, Chun H, Kim HH, Eskandar Y, Kim CK, Machac J. Factors affecting visualization rates of internal mammary sentinel nodes during lymphoscintigraphy. J Nucl Med 2003; 44:1387–1393. Borgstein PJ, Pijpers R, Comans EF, Van Diest PJ, Boom RP, Meijer S. Sentinel lymph node biopsy in breast cancer: guidelines and pitfalls of lymphoscintigraphy and gamma probe detection. J Am Coll Surg 1998; 186:275–283. Uren RF, Howman-Giles RB, Thompson JF, Malouf D, Ramsey-Stewart G, Niesche FW, et al. Mammary lymphoscintigraphy in breast cancer. J Nucl Med 1995; 36:1775–1780. McMasters KM, Tuttle TM, Carlson DJ, Brown CM, Noyes RD, Glaser RL, et al. Sentinel lymph node biopsy for breast cancer: a suitable alternative to routine axillary dissection in multi-institutional practice when optimal technique is used. J Clin Oncol 2000; 18:2560–2566. Valdes-Olmos RA, Jansen L, Hoefnagel CA, Nieweg OE, Muller SH, Rutgers EJ, et al. Evaluation of mammary lymphoscintigraphy by a single intratumoral injection for sentinel node identification. J Nucl Med 2000; 41:1500–1506. Gulec SA, Moffat FL, Carroll RG, Krag DN. Gamma probe guided sentinel node biopsy in breast cancer. Q J Nucl Med 1997; 41:251–261. Estourgie SH, Tanis PJ, Nieweg OE, Valdes Olmos RA, Rutgers EJ, Kroon BB. Should the hunt for internal mammary chain sentinel nodes begin? An evaluation of 150 breast cancer patients. Ann Surg Oncol 2003; 10:935–941.

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NEWS AND VIEWS AUGUST 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up-to-date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 206 6930, E-mail: [email protected]

Nuclear Medicine Communications, 2006, 27, 683–684 What do we tell the patients?

UK readers may recall that a weighty questionnaire came round last summer asking for examples of all the information we give to patients about their diagnostic tests and therapies. The work was commissioned by the Health and Safety Executive (HSE) and carried out by the Department of Medical Physics, Royal Surrey County Hospital and the Department of Psychology, University of Surrey. The report resulting from this survey has now been published as HSE Research Report 416 with the title ‘Information to accompany patients undergoing nuclear medicine procedures’ (http://www.hse.gov.uk/ research/rrpdf/rr416.pdf). As befits the size of the survey, the report is weighty and comprehensive. It makes interesting, sometimes even amusing reading. We would like to share a few views about the document, though obviously reading this is no substitute for reading the report. The overall aim of the report is to present a snapshot of the status quo vis-a`-vis written and oral information presented to patients attending for diagnostic and therapeutic procedures. The information offered both at the time of booking and at the time of attendance is considered. The report does not appraise information content on a technical radiation protection level though it

does appraise on a patient communication level, since each of the information sheets was examined using the Ensuring Quality Information for Patients tool, EQIP (Moult, et al. Health Expectations 7:165–175). The authors carried out two further interesting exercises for the report: a telephone audit of patients’ satisfaction and use of information they received, and focus group discussions with professionals from within and without nuclear medicine. The latter groups included hospital nurses, community nurses, cleaners, porters, a radiologist, general practitioners, an embalmer, a pathologist, mortuary staff and a funeral director. The indisputable (and not surprising) conclusion is that there is great variability in the categories of information supplied and advice in each category. The categories included risks to patient and contacts, what happens during the procedure, contact and contamination restrictions, rationale for contact and contamination restrictions, special situation such as breast feeding and pregnancy, who to contact for advice and external information resources (e.g., trusted web sites). It makes a useful checklist of things we should at least consider when revising our information to patients.

A major variable in the advice offered concerned the explanation of risk. There are no prizes for guessing that the advice was somewhat inconsistent between centres, but what we found most astonishing was what some providers of the advice felt might be reassuring, for instance ‘The same as having 100 X-rays on your back’. Most of us are quite poor at comprehending low risks, so some sort of comparison is probably unavoidable, but we agree with several of the patients who stated that comparing one poorly understood medical procedure risk with another was unhelpful. On the other hand, we do believe that explaining, quantitatively, the risks of cancer is likely to cause undue alarm due to our poor comprehension of small risk coefficients and because many of us irrationally weight more heavily the risks of cancer than risks of other diseases or accidents. Our own favourite is road use. Like radiation risk it is genuinely stochastic, linear and widespread but patients have a much better feel for it. Overall, the report is thought provoking and demonstrates a need for a generic set of advice leaflets available to the community. Writing them well is a time-consuming job and it makes no sense for us all to do separate sets, quite apart from the waste of man-hours. Dr Ann Tweddel,

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on behalf of BNMS is currently gathering a multi-disciplinary group to complete this task, and in due course we expect to be able to make available model leaflets which could either be adopted by departments or form the core for locally developed material. Meeting Announcements BNMS Autumn Meeting

Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk EANM Annual Meeting

Dates: 30 September–4 October 2006 Venue: Athens, Greece Website: www.eanm.org

Website: www.wfnmb.org/ congress2006/index02.htm

Venue: EANM PET Facility, Vienna, Austria

International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM)

Contact: EANM executive Secretariat on + 43 1 2128030, fax + 43 1 21280309

Dates: 13–15 November 2006 Venue: Vienna, Austria Website: www-pub.iaea.org/MTCD/ Meetings/Announcements.asp? ConfID = 146

Website: www.eanm.org/education/ esnm/esnm_intro.php

Design of Radionuclide Facilities with Reference to Radiation Protection (Revisited)

Date: 14 November 2006 Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT Website: www.bir.org.uk

9th World Congress of Nuclear Medicine and Biology

Education and Training EANM Learning Courses

Dates: 22–27 October 2006 Venue: Seoul, South Korea

Dates: Weekend courses throughout 2006

Learning

Email: [email protected]

EANM Distance Learning in Nuclear Cardiology

This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications.

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Editorial

Clinical PET in oncology: not only FDG Cristina Nannia, Domenico Rubellob, Adil Al-Nahhasc and Stefano Fantia Nuclear Medicine Communications 2006, 27:685–688 a Nuclear Medicine Service, PET Unit, Policlinico S. Orsola-Malpighi, University of Bologna, Italy, bNuclear Medicine Service, Pet Unit, S. Maria della Misericordia Hospital, Rovigo, Italy and cDepartment of Nuclear Medicine, Hammersmith Hospital, London, UK.

Correspondence to Dr Adil Al-Nahhas, Department of Nuclear Medicine, Hammersmith Hospital, Du Cane Road, London W12 0HS, UK. Tel: + 44 208 383 4923; fax: + 44 208 383 1700; e-mail: [email protected] Received 8 May 2006 Accepted 12 May 2006

Introduction

11

Since positron emission tomography (PET) started to play a major role in clinical oncology, 2-[18F]fluoro-2-deoxy-Dglucose (18F-FDG) has become the most commonly used radiopharmaceutical both for PET and PET/computed tomography (CT) studies. FDG is a glucose analogue that has a very high sensitivity for detecting the majority of malignant tumours and is useful for staging, re-staging, assessing therapy response and regular follow-up.

Well-differentiated Prostate cancer is recognized as a low-glucose-consumer malignant tumour demonstrating low 18F-FDG uptake. In addition, the frequent inflammatory processes of the prostate gland and its close proximity to the urinary bladder make it difficult to use 18 F-FDG for diagnosing primary prostate cancer and, more importantly, to detect local recurrence or assess response to therapy [1,2].

However, a small number of malignant tumours (prostate cancer, neuroendocrine tumours and hepatic tumours amongst others) do not show a significantly increased 18FFDG uptake and are therefore difficult to assess with PET. The reduced sensitivity of 18F-FDG is noted in some histological types of cancer with high differentiation and low growth rate, such as prostate cancer and neuroendocrine tumours, resulting in less glucose consumption and low FDG uptake. Another factor contributing to low sensitivity is the inability of certain types of tumours, such as hepatocellular carcinoma, to trap FDG inside their cells. One explanation of this phenomenon is the clash between cytoplasmic glucose phosphorylase and de-phosphorylase, resulting in a negative balance between intracellular and extracellular FDG concentration and its eventual loss from the cell. Despite its widespread use, 18F-FDG has well-known disadvantages, such as its inability to evaluate malignant lesions located in or close to tissues with high metabolic activity (e.g., the brain) or areas of intense physiological accumulation (e.g., the urinary bladder). Furthermore, 18 F-FDG has reduced specificity when it comes to differentiating malignancy from infected or highly inflamed lesions. To improve the sensitivity and specificity in PET imaging, several new positron-emitting radiopharmaceuticals have been developed and introduced into clinical practice, although they still play a minor role compared to FDG. The most important non-FDG compounds for oncological studies include 11C-choline, 11C-methionine, 18 F-DOPA, 68Ga-DOTA-NOC and 11C-acetate.

C-choline

11

C-choline is a small molecule that, once injected intravenously, is integrated in the cell membrane as phosphatidylcholine and acts as a marker of membrane metabolism. Unlike FDG, 11C-choline has a high affinity for malignant prostate tissue, albeit of a low grade. Its late urinary excretion ensures that the whole pelvis is free from urinary radioactivity at the time of image acquisition. 11

C-choline PET/CT has been shown to be cost-effective in patients with prostate cancer relapse following radical therapy with surgery and or radiotherapy. This is particularly true when a rising serum level of prostatespecific antigen (PSA) is associated with negative conventional imaging procedures such as trans-rectal ultrasound, bone scintigraphy and pelvic magnetic resonance imaging. It has a sensitivity, specificity and accuracy for early nodal involvement detection of 80%, 96% and 93%, respectively, and can detect early secondary bone lesions. It is also useful for detecting local recurrence but with a lower sensitivity [3,4]. However, caution is advised when interpreting 11 C-choline PET. Farsad and colleagues have shown its efficacy for detection of intraprostatic primary cancer but have also demonstrated that, for this indication, it is non-specific and must be used only for selected patients at high risk with multiple negative biopsies [5]. 11

C-methionine

Malignant brain tumours are usually treated with different combinations of surgery, radiotherapy and chemotherapy to achieve an optimal response. After such

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686 Nuclear Medicine Communications 2006, Vol 27 No 9

combination therapy, conventional imaging procedures (contrast CT and magnetic resonance imaging) are useful for detecting disease relapse but differentiation from non-malignant conditions such as post-surgical fibrosis, radionecrosis or oedema can be extremely difficult.

areas of high protein metabolism inside a brain mass to guide a biopsy [8] or to evaluate the early effect of radiotherapy on head and neck tumours. However, most of these applications are in the research domain and are not yet validated for clinical use [9].

The brain has high physiological uptake of 18F-FDG because glucose is the main metabolic substrate for the central nervous system. Consequently, small malignant lesions are very difficult to detect with 18F-FDG PET as they may be masked by the hypermetabolic background [6,7].

18

11

C-methionine is an amino acid radiopharmaceutical for PET and has no physiological tendency to accumulate in the brain. However, malignant lesions present as areas of increased 11C-methionine uptake due to increased protein metabolism and vascular permeability (Fig. 1). Benign conditions such as fibrosis, necrosis or oedema show uptake similar to normal brain tissue or even appear as photon deficient areas. In these conditions, the contrast between low or absent background activity and low-grade increased uptake in malignant lesions makes 11 C-methionine very useful in assessing malignant brain tumours. Its most appropriate indication is when conventional imaging procedures do not discriminate between the aforementioned benign conditions and disease relapse allowing for prompt and timely second line treatment to be introduced. Other applications of 11C-methionine PET include its use to define the radiotherapy field for the central nervous system or head and neck tumours, to localize

Fig. 1

F-DOPA

18

F-deoxyphenylalanine (18F-DOPA) is a radiolabelled amino acid precursor of dopamine. It was originally synthesized for the in-vivo evaluation of receptor uptake in the caudate and putamen in Parkinson’s disease. More recently, it has been employed in oncology for the detection of malignant tumours derived from the neural crest [10]. These tumours (carcinoid, pheochromocytoma, neuroblastoma, medullary thyroid carcinoma, microcytoma, carotid glomus tumours and melanoma) are usually very well differentiated and 18F-FDG PET has proved to be very insensitive in their detection. When there are clinical and/or biochemical indications of recurrence or relapse of these tumours, in association with negative cross-sectional imaging, then 111In-OctreoScan or meta-[123I]iodobenzylguanidine (123I-MIBG) (planar and SPECT views) are performed for localization purposes. These tests have good sensitivity and specificity, but small lesions could be missed due to their poor spatial resolution compared to what can be obtained with PET scans.

One of the features of these tumours is the capacity to accumulate amino acids inside the cytoplasmic space through a metabolic pathway, allowing radiopharmaceuticals such as 18F-DOPA to detect their presence. Various studies have shown good sensitivity and specificity, probably higher than conventional imaging procedures [11–16]. Despite promising preliminary results, 18 F-DOPA has not yet been fully implemented in clinical practice due to complicated production methods and the relative rarity of neuroendocrine tumours. 68

21

−84.00 mm

Brain 11C-methionine scan showing relapse of a CNS malignant cancer (black arrow).

Ga-DOTA-NOC

During the last few years, 68Ga-tetra-azacyclododecane tetra-acetic acid-Nal3-octreotide (68Ga-DOTA-NOC) has been successfully synthesized for PET imaging [17]. 68 Ga-DOTA-NOC has a high affinity for human somatostatin receptor 2 (hSSR2) and hSSR5 and is being used for the detection of neuroendocrine tumours in preliminary studies. Like 111In-OctreoScan, the use of 68 Ga-DOTA-NOC in tumour detection is based on uptake in the highly expressed receptors in the surface membrane of a tumour cell, but it has added advantages, including the superior PET spatial resolution, the shorter uptake time (60 min) and aquisition procedures (no need for 24- or 48-h imaging). 68Ga is eluted from a 68Ge–68Ga generator, which has a half-life of 270 days, making it readily available on a daily basis.

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Editorial Nanni et al. 687

Since 68Ga-DOTA-NOC binds to neuroendocrine tumours through a receptor mechanism, its uptake is correlated with the density of somatostatin receptors expressed on the cell surface, and therefore its sensitivity may be lower than that of 18F-DOPA, which relies on a metabolic mechanism. Overall sensitivity and specificity of 68Ga-DOTA-NOC as compared to 18F-DOPA has still to be assessed, but it may be realistic to predict a complementary role for the two tracers, because they can be used to explore different aspects of the behaviour of neuroendocrine tumours. 68Ga can also be used as a label for other members of the DOTA family such as DOTATATE and DOTA-TOC. 11

C-acetate

11

C-acetate is a PET tracer that accumulates as an intermediate molecule both in the glucose catabolism pathways and in membrane metabolism, partially resembling the behaviour of FDG and choline. In fact, acetate can be transformed into acetyl-CoA, entering the tricarboxylic acid cycle, or used as a precursor of membrane fatty acids. As for 18F-DOPA, the original application of 11 C-acetate was not in oncology but in cardiology as its accumulation in the myocardium is proportional to fatty acid oxidation and is commonly used to evaluate cardiac metabolism. From the oncological point of view, 11C-acetate was at first used as a choline analogue for the evaluation of prostate cancer, showing a similar sensitivity and specificity [18], but more recently Delbeke and colleagues Fig. 2

18F-FDG

11C-Acetate Patient with hepatocellular carcinoma: negative FDG PET scan and positive 11C-acetate PET scan.

used 11C-acetate and 18F-FDG for the evaluation of liver masses. Preliminary results demonstrate that 11C-acetate has good sensitivity for low-grade hepatic tumours but not for high-grade tumours, while FDG has the opposite behaviour, being insensitive for low-grade cancer (differentiated and still producing a cytoplasmic de-phosphorylase) but very sensitive for high-grade tumours (Fig. 2). The two radiopharmaceuticals, therefore, cover the whole spectrum of hepatic cancers [19,20].

[18F]Fluoro-levo-thymidine [18F]Fluoro-levo-thymidine (18F-FLT) was one of the most promising radiotracers synthesized for PET oncological studies. Thymidine is a DNA base and is actively included into DNA when it replicates. 18F-FLT is, therefore, a specific marker of cell proliferation and was thought to be more sensitive and specific than FDG for oncological studies, particularly because it does not accumulate into inflammation and can theoretically be an optimal marker to assess therapy response. However, further experience has shown that 18F-FDG is more sensitive than 18F -FLT for the evaluation of several malignant tumours [21]. 18F-FLT is preferred for smallanimal PET scanning in experimental settings. Small animals such as mice and rats express very low levels of thymidine kinase compared to humans, causing delayed 18 F-FLT metabolism, which is optimal for in-vivo proliferation measurements in pre-clinical studies. Other PET radiopharmaceuticals include hypoxia markers such as [18F]fluoromisonidazole (18F-MISO) and 64 Cu-ATSM, which highlight the presence of hypoxic areas in tumours and are very useful in patients treated with radiotherapy. It is well recognized that hypoxia is one of the factors responsible for resistance to treatment and hypoxic areas should be recognized and overtreated for optimum results [22]. Despite promising results, 18 F-MISO was proved not to be highly accurate and 64 Cu-ATSM has a rather complicated production procedure.

Conclusions Although 18F-FDG is widely used in oncological PET imaging, with the sensitivity to detect the majority of malignant tumours, there are several situations in which it is not appropriate or cost-effective. Other radiopharmaceuticals, targeting cell membranes, hSSR and hypoxia have been developed and tested for use with other types of malignant tumours that are FDG negative. In addition to 18F, the new radiopharmaceuticals are labelled with 11C and 68Ga. However, most of these have not been fully introduced into clinical practice being either specific for tumours with low incidence, or are difficult to prepare or be labelled with short half-life isotopes, requiring an on-site cyclotron. Nevertheless,

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these new PET radiopharmaceuticals are important and destined to grow in number and specialization to allow for wider and more effective use of PET imaging.

References 1 2

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Sanz G, Rioja J, Zudaire JJ, Berian JM, Richter JA. PET and prostate cancer. World J Urol 2004; 22:351–352. Picchio M, Messa C, Landoni C, Gianolli L, Sironi S, Brioschi M, et al. Value of [11C]choline positron emission tomography for re-staging prostate cancer: a comparison with [18F]fluorodeoxyglucose-positron emission tomography. J Urol 2003; 169:1337–1340. Picchio M, Landoni C, Messa C, Gianolli L, Matarrese M, De Cobelli F, et al. Positive [11C]choline and negative [18F]FDG with positron emission tomography in recurrence of prostate cancer. AJR Am J Roentgenol 2002; 179:482–484. de Jong IJ, Pruim J, Elsinga PH, Vaalburg W, Mensink HJ. 11C-choline positron emission tomography for the evaluation after treatment of localized prostate cancer. Eur Urol 2003; 44:32–38. Farsad M, Schiavina R, Castellucci P, Nanni C, Corti B, Martorana G, et al. Detection and localization of prostate cancer: correlation of (11)C-choline PET/CT with histopathologic step-section analysis. J Nucl Med 2005; 46:1642–1649. Kim S, Chung J-K, Im S-H, Jeong JM, Lee DS, Kim DG, et al. 11C-methionine PET as a prognostic marker in patients with glioma: comparison with 18 F-FDG PET. Eur J Nucl Med Mol Imaging 2005; 32:52–59. van Laere K, Ceyssens S, van Calenbergh F, de Groot T, Menten J, Flamen P, et al. Direct comparison of 18F-FDG and 11C-methionine PET in suspected recurrence of glioma: sensitivity, inter-observer variability and prognostic value. Eur J Nucl Med Mol Imaging 2005; 32:39–51. Pirotte B, Goldman S, Massager N, David P, Wikler D, Vandesteene A, et al. Comparison of 18F-FDG and 11C-methionine for PET-guided stereotactic brain biopsy of gliomas. J Nucl Med 2004; 45:1293–1298. Geets X, Daisnea J-F, Gregoire V, Hamoir M, Lonneux M. Role of 11-C-methionine positron emission tomography for the delineation of the tumor volume in pharyngo-laryngeal squamous cell carcinoma: comparison with FDG-PET and CT. Radiother Oncol 2004; 71:267–273.

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Ahlstrom H, Eriksson B, Bergstrom M, Bjurling P, Langstrom B, Oberg K. Pancreatic neuroendocrine tumors: diagnosis with PET. Radiology 1995; 195:333–337. Hoegerle S, Altehoefer C, Ghanem N, Koehler G, Waller CF, Scheruebl H, et al. Whole-body 18F Dopa PET for detection of gastrointestinal carcinoid tumors. Radiology 2001; 220:373–380. Hoegerle S, Altehoefer C, Ghanem N, Manz T, Brink I, et al. Pheochromocytomas: detection with 18F DOPA whole body PET – initial results. Radiology 2002; 222:507–512. Hoegerle S, Ghanem N, Altehoefer C, Schipper J, Brink I, Moser E, Neumann HP. 18F-DOPA positron emission tomography for the detection of glomus tumours. Eur J Nucl Med Mol Imaging 2003; 30:689–694. Brink I, Hoegerle S, Klisch J, Bley TA. Imaging of pheochromocytoma and paraganglioma. Fam Cancer 2005; 4:61–68. Hoegerle S, Altehoefer C, Ghanem N, Brink I, Moser E, Nitzsche E. 18 F-DOPA positron emission tomography for tumour detection in patients with medullary thyroid carcinoma and elevated calcitonin levels. Eur J Nucl Med 2001; 28:64–71. Gourgiotis L, Sarlis NJ, Reynolds JC, Vanwaes C, Merino MJ, Pacak K. Localization of medullary thyroid carcinoma metastasis in a multiple endocrine neoplasia type 2A patient by 6-[18F]-fluorodopamine positron emission tomography. J Clin Endocrinol Metab 2004; 88:637–641. Wild D, Macke HR, Waser B, Reubi JC, Ginj M, Rasch H, et al. 68 Ga-DOTANOC: a first compound for PET imaging with high affinity for somatostatin receptor subtypes 2 and 5. Eur J Nucl Med Mol Imaging 2005; 32:724. Dimitrakopoulou-Strauss A, Strauss LG. PET imaging of prostate cancer with 11C-acetate. J Nucl Med 2003; 44:556–558. Delbeke D, Pinson CW. 11C-acetate: a new tracer for the evaluation of hepatocellular carcinoma. J Nucl Med 2003; 44:222–223. Ho CL, Yu SC, Yeung DW. 11C-acetate PET imaging in hepatocellular carcinoma and other liver masses. J Nucl Med 2003; 44:213–221. Been LB, Suurmeijer AJ, Cobben DC, Jager PL, Hoekstra HJ, Elsinga PH. [18F]FLT-PET in oncology: current status and opportunities. Eur J Nucl Med Mol Imaging 2004; 31:1659–1672. Lewis JS, Welch MJ. PET imaging of hypoxia. Q J Nucl Med 2001; 45: 183–188.

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Original article 99m

Tc-methylene diphosphonate single photon emission tomography of the knees: Intensity of uptake and its correlation with arthroscopic findings Yoel Siegela, Haim Golanb,* and Refael Theinc Objective To examine whether single photon emission computed tomography (SPECT) can determine the severity of knee pathology, based on intensity of uptake and, therefore, possibly substituting this technique for more invasive and expensive diagnostic procedures, such as arthroscopy, in certain patient populations. Methods The study results of patients referred for knee SPECT followed by an arthroscopy at our institution were evaluated retrospectively. The scintigraphic and arthroscopic findings for the menisci and femoral condyles were each graded on a numeric scale. One numeric scale corresponded to the level of uptake in the SPECT and the other to the severity of the pathology visualized at arthroscopy. Statistical correlation between both scales, representing the SPECT and arthroscopy findings, was performed.

Conclusion The degree of uptake in the knee, as determined by SPECT, positively correlates with the severity of pathology seen at arthroscopy. The data can potentially be used to assist in decision-making before proceeding to surgery, especially where there is severe pathology that may be less amenable to arthroscopic c 2006 Lippincott therapy. Nucl Med Commun 27:689–693 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:689–693 Keywords: knee SPECT, knee arthroscopy, knee pain

a Department of Radiology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel, bDepartment of Nuclear Medicine and cUnit of Arthroscopy and Sports Injuries, Rabin Medical Center, Golda Campus, Petach Tikva, Israel.

Results Forty-one patients were included in the study. A positive and statistically significant correlation was found between the intensity of uptake on the SPECT and the severity of the arthroscopic findings in the menisci and medial femoral condyle.

Correspondence to Dr Yoel Siegel, Department of Radiology, Rabin Medical Center, Beilinson Campus, Petach Tikva 49100 Israel. Tel: + 972-3-9376344; fax: + 972-3-9376387; e-mail: [email protected]

Introduction

implications because more severely damaged knees are less amenable to arthroscopic therapy [1]. Typically, four types of cartilage lesions are described (from mild to severe): softening, fibrillation, fragmentation and eburnation (when the bone can be seen through the shredded cartilage). Although arthroscopy is considered the diagnostic gold standard it is an invasive and costly procedure that is best reserved for cases that are mainly therapeutic.

Knee pain poses a diagnostic and therapeutic dilemma because of its diverse aetiologies. The clinician not only has to diagnose the cause of the pain but must also devise a treatment plan tailored to the specific patient and pathology. Arthroscopy is considered the ‘gold standard’ method for diagnosis, and is frequently the treatment of choice for knee pathologies such as meniscal tears and cartilage disease. The pathologies seen at arthroscopy can be defined according to shape, size and anatomical location. For instance, a meniscal tear is usually described as being simple, such as a radial and flap tear, or more complicated, such as a longitudinal tear and its subtype the buckethandle tear. In some cases more than one type of tear may co-exist; this is known as a complex tear. Knee cartilage joint disease (CJD) can also be subdivided according to the severity of the chondral damage seen at arthroscopy. The type and severely of joint pathology has surgical * Dr Golan is now at the Department of Nuclear Medicine, Assaf Harofeh Medical Center, Zerifin, Israel.

Received 18 December 2005 Accepted 6 February 2006

In order to minimize diagnostic invasive procedures there are a number of modalities such as conventional X-ray, magnetic resonance imaging (MRI) and scintigraphy that may aid in evaluating the knee before planning therapeutic strategies. The use of bone scans for diagnosing knee pathology has been reported in a number of studies [2–7]. Single photon emission computed tomography (SPECT) has shown high sensitivity and specificity for detecting meniscal tears [2–6]. Knee-bone scintigraphy can also identify cruciate ligament tears and cartilage insult

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[2,7,8]. The majority of the literature addressing the use of scintigraphy for investigating knee pain has established the advantages of bone scans, both planar and SPECT, in diagnosing the different types of pathology. Less emphasis has addressed the role scintigraphy may have in predicting the severity of pathology found at arthroscopy. The purpose of our study was to examine in what way SPECT can predict the severity of common knee pathologies and provide the clinician with information that can assist in deciding whether or not to proceed with more invasive procedures such as arthroscopy.

Fig. 1

(a)

(b)

Materials and methods Medical records of patients who had a SPECT examination for knee pain and who later underwent arthroscopy at our medical center during 2002–2003 were reviewed retrospectively. The data collection was in accordance with the Helsinki ethics committee guidelines. We included patients whose complaints were of at least 1month duration prior to the SPECT study. Demographic and relevant clinical data was recorded. Bone SPECT studies were performed using a triple-head gamma camera (Irix, Philips) with low-energy, high-resolution, parallel-hole collimators at 120 min after intravenous injection of 740 MBq (20 mCi) 99mTc-methylene diphosphonate (MDP). Data acquisition was performed using 128  128 frame matrix, for 3601 orbit at 31 step size at 22.5 s per step, for a total acquisition time of 15 min. Iterative reconstruction at 1 pixel per slice with a Butterworth post-reconstruction three-dimensional filter (power = 4, cut-off = 0.4 Nyquist frequency) was used for tomographic image reconstruction. A single nuclear medicine physician blinded to the arthroscopy results analysed the SPECT images according to the following criteria. Pathological SPECT uptake in either the femoral condyles or menisci was noted. Any increase in uptake, regardless of size, was considered pathological, including meniscal uptake that was not crescent in shape. Intensity of uptake was graded on a scale of 0–3 (Figs 1–3). Anatomical sites that were scintigraphically normal were designated as grade zero. Anatomical sites that had a high degree of uptake were designated as grade 3. Moderate uptake was considered to be grade 2 and mild uptake was designated as grade 1. An arthroscopy surgeon who was blinded to the SPECT results was retrospectively presented with the patients’ arthroscopy reports and pictures taken during the arthroscopy. The surgeon evaluated the data and defined the meniscal tears in increasing severity as radial, flap, complex and longitudinal (bucket handle). A grade of zero was designated to normal findings, a grade of 1 for a radial tear, a grade of 2 for a flap tear, grade 3 for a

Two serial axial images from a knee SPECT study of a 25-year-old man complaining of knee pain 6 months after a patellar fracture. (a) High intensity uptake is seen in the right patella (short arrow) and trochlea region (long arrow). (b) High intensity uptake is seen in the right patella (short arrow) and low intensity uptake is seen in the right lateral femoral condyle (long arrow).

complex tear, and grade 4 for a longitudinal tear. The surgeon also graded on a scale of 1–4 the cartilage damage of the femoral condyles according to the appearance at arthroscopy. Again, a designation of zero was given to normal findings, grade 1 for cartilage softening, grade 2 for fibrillation and so on. The graded arthroscopy and SPECT results were compared. Data were analysed using SAS commercial software. Pearson’s r value and its P value were calculated. A P value of less than or equal to 0.05 was considered statistically significant.

Results Between 2002 and 2003 42 patients had a SPECT scan of the knees followed by arthroscopy at our institution. One patient’s complaint was less than 1-month duration and was excluded from the study. The ages of the patients ranged from 18 to 73 years (average 44.7 ± 14 years). Sixteen of the patients were female and 25 male. Fifteen of the patients had a history of trauma while the rest either had a history of knee over-use or pain without an identifiable cause. The time that elapsed between the beginning of the complaint and the SPECT study ranged from 1 to 84 months (average 11.8 months, median 6 months). The average time that elapsed between the SPECT and the arthroscopy was 2 ± 2 months.

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99m

Tc SPECT of the knees Siegel et al. 691

Complex tears were the most common type of meniscal tear. CJD of the femoral condyles was noted relatively frequently and arthroscopic grade 3 severity of chondral damage was the most commonly seen. The scintigraphic and arthroscopic findings of the medial meniscus, lateral meniscus, medial femoral condyle (MFC) and lateral femoral condyle (LFC) are listed in Tables 1–4. Sensitivity, specificity, positive and negative predictive values for the SPECT results when compared to arthroscopic findings are presented in Table 5. The correlation between the SPECT intensity and the severity of meniscal and chondral damage was as follows:

Table 2

Lateral meniscus tears and intensity of uptake

Arthroscopy findings

SPECTintensity

Total

Normal

Mild

Moderate

High

Normal Radial Complex

25

5 4

2 1 2

2

32 1 8

Total

25

9

5

2

41

Table 3 Medical femoral condyle cartilage damage and its intensity uptake Arthroscopic findings

SPECT findings

Total

Normal 14

Mild 2

Moderate 1

High

Normal Grade 1 Grade 2 Grade 3

2 1 1

1 3 5

1

4 1

5

4 8 12

18

7

10

6

41

Fig. 2

Total

Table 4

17

Lateral femoral condyle cartilage damage and its intensity

uptake An axial image from a knee SPECT study of a 28-year-old woman with a history of prior trauma and complaining of knee pain. High intensity uptake can be seen in the right medial femoral condyle (arrow).

Arthroscopic findings

SPECT findings

Total

Normal

Mild

Moderate

High

Normal Grade 1 Grade 2 Grade 3

24 1

6

2

2 1

2

1 1

34 2 3 2

Total

26

8

4

3

41

1

Fig. 3

Table 5 Statistical analysis of SPECT findings in the knee compared to arthroscopy Statistical analysis

Anatomical location Medial me- Lateral meniscal tear niscal tear

An axial image from a knee SPECT study of a 70-year-old man complaining of right knee pain for a number of years with no history or initial trauma. High intensity uptake is seen in the right medial meniscus (short arrow) and moderate uptake in the left medial meniscus (long arrow).

Table 1

Sensitivity (%) Specificity (%) Positive predictive value (%) Negative predictive value (%)

Medial femoral condyle chondral damage

Lateral femoral condyle chondral damage

100 75 86

100 78 56

83 82 87

71 71 33

100

100

78

92

Medial meniscus tears and intensity of uptake

Arthroscopy findings

SPECT intensity

Total

Normal

Mild

Moderate

High

Normal Radial Flap Complex Bucket handle

12

1

2

2 1

6 5 1

1 1 4 3 2

16 1 12 8 4

Total

12

4

14

11

41

for the medial meniscus an r value of 0.663 (P < 0.001), for the lateral meniscus an r value of 0.67 (P < 0.001) and for the medial femoral condyle an r value of 0.7 (P < 0.001), indicating that the uptake intensity positively correlated with the severity of pathology seen at arthroscopy. In the LFC the r value was 0.086 (P > 0.05) indicating that no correlation was found between the intensity of uptake and the severity of findings at arthroscopy.

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Discussion Knee pain caused either by trauma or over-use is a common complaint among patients presenting to the primary physician or orthopaedic surgeon. Clinicians have a number of modalities at their disposal to reach a diagnosis. Ideally, the work-up should include the most appropriate studies at the lowest possible cost. MRI has shown a high sensitivity and specificity in detecting knee pathology, such as meniscal tears, although some studies have suggested that SPECT can be used as an alternative to MRI in suitable cases and when MRI is not available [5,9]. In addition, some practitioners have found that MRI is relatively insensitive in detecting articular cartilage damage [10,11]. CJD is also a frequent cause of pain that can be a diagnostic challenge. In fact, there may be occult cartilage damage that is not seen by direct visualization at surgery [11–16]. Therapy for articular cartilage damage is controversial although the degree of severity seems to be of importance, because in cases of grade 4 osteoarthritis the efficacy of arthroscopic treatment is low and less likely to be of benefit [1,17]. Therefore, it is important to identify these severe cases in which a conservative approach may be more appropriate, and an imaging modality that can stage the damage may contribute to therapeutic strategies. The study presented here attempted to address this issue. The correlation between the severity of damage seen at arthroscopy and the uptake seen on SPECT in the menisci and medial femoral condyle was relatively high and reached statistical significance. This would indicate that at these locations a high level of uptake would favour more complex and severe damage as seen at arthroscopy. In addition, there was a relatively high negative predictive value found especially in the menisci and in the MFC. This suggests that a scan interpreted as normal would probably be normal at arthroscopy as well. In contrast to the findings in the MFC the correlation of the SPECT and the arthroscopy in the LFC was low and was not statistically significant. This finding is inconsistent with the other anatomical sites that we examined and probably can be attributed to the few cases that had LFC pathology. Another possibility that can explain this discrepancy is that the SPECT findings in the LFC were mainly of subchondral bone changes caused by mechanical trauma secondary to knee instability and therefore not appreciated on arthroscopy.

There are relatively few attempts described in the literature to quantify knee scintigraphic findings. EvenSapir et al. [8] used an intensity grading system that correlated with the presence of bone injury accompanying an anterior cruciate ligament tear. Ryan et al. [6] found that enlarging the area of activity as the definition of a meniscal tear, thus increasing uptake area, helps in detecting more severe tears yet reduces sensitivity. In

our study the ability to determine severity of pathology did not reduce sensitivity because any increased uptake was considered a possible lesion. The study has a number of limitations. Most patients had relatively severe findings on arthroscopy. This may limit the ability of the study to investigate the role of SPECT in diagnosing mild meniscal tears and minimal articular cartilage damage. The study group was relatively heterogeneous with a wide range of pathology and complaint periods. Also, correlation with other modalities such as MRI was not performed in our study although this was not our purpose since we compared our results to the gold standard, i.e., arthroscopy. Keeping these caveats in mind, the main objective of this study was to attempt to quantify and correlate pathological findings on knee SPECT with actual arthroscopic findings. By grading the scintigraphic findings a correlation with the surgical findings could be established. The semi-quantitative grading system of uptake intensity used in this study correlated positively with the findings at arthroscopy. This correlation further supports the use of SPECT alone or in conjunction with other modalities to estimate the type and severity of meniscal tears prior to deciding on arthroscopic intervention. Possibly, the most important implication of the findings is the correlation of high uptake with the severity of articular cartilage damage. In cases of severe articular cartilage damage, with or without meniscal tears, the benefit of arthroscopy is frequently marginal since arthroscopic surgical options for pain relief are relatively limited. In conclusion, the grade of uptake intensity in knee SPECT appears to correlate with the severity of meniscal and articular cartilage pathology seen at arthroscopy, while a normal SPECT study will usually predict normal findings. This correlation may aid in decision-making prior to choosing a surgical approach especially in cases of severe cartilage damage, which usually indicates a poorer prognosis. Further investigation with a larger and more homogeneous cohort is needed to confirm these findings.

References 1

2

3 4

5

Jackson WR, Dieterichs C. The results of arthroscopic lavage and debridement of osteoarthritic knees based on the severity of degeneration: A 4 to 6 year symptomatic follow-up. Arthroscopy 2003; 19:13–20. Collier BD, Johnson RP, Carrera GF, Isitman AT, Veluvolu P, Knobel J, et al. Chronic knee pain by SPECT: comparison with other modalities. Radiology 1985; 157:795–802. Murray IP, Dixon J, Kohan L. SPECT for acute knee pain. Clin Nucl Med 1990; 15:828–840. Fajman WA, Diehl M, Dunaway E. Tomographic and planar radionuclide imaging in patients with suspected meniscal injury: arthroscopic correlation. J Nucl Med 1985; 26:77. Ryan PJ, Reddy K, Fleetcroft J. A prospective comparison of clinical examination, MRI, bone SPECT, and arthroscopy to detect meniscal tears. Clin Nucl Med 1998; 23:803–806.

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Ryan PJ, Taylor M, Grevitt M, Allen P, Shields J, Clarke SE, et al. Bone single emission tomography in recent meniscal tears: an assessment of the diagnostic criteria. Eur J Nucl Med 1993; 20:703–707. 7 Gary JR, Cook GJ, Ryan PJ, Clarke SE, Fogelman I. SPECT bone scintigraphy of anterior cruciate ligament injury. J Nucl Med 1996; 37: 1353–1356. 8 Even-Sapir E, Arbel R, Lerman H, Flusser G, Livshitz G, Halperin N. Bone injury associated with anterior cruciate ligament and meniscal tears: assessment with bone single photon emission computed tomography. Invest Radiol 2002; 37:521–527. 9 Ryan PJ, Chauduri R, Bingham J, Fogelman I. A comparison of MRI and bone SPECT in the diagnosis of knee pathology. Nucl Med Commun 1996; 17:125–131. 10 Levy AS, Lohnes J, Sculley S, LeCroy M,Garrett W. Chondral delamination of the knee in soccer players. Am J Sports Med 1996; 24:634–639. 11 Thein R, Eichenblat M. Concealed knee cartilage lesions: is arthroscopic probing therapeutic? Am J Sports Med 1999; 27:495–499.

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Speer KP, Spritzer CE, Goldner JL. Magnetic resonance imaging of traumatic knee articular cartilage injuries. Am J Sports Med 1991; 19:396–402. Johnson DL, Urban Jr WP, Caborn DN, Vanarthos WJ, Carlson CS. Articular cartilage changes seen with magnetic resonance imaging- detected bone bruise associated with acute anterior cruciate ligament rupture. Am J Sports Med 1998; 26:409–414. Mink JH, Deutsch AL. Occult cartilage and bone injuries of the knee: detection, classification, and assessment with MR imaging. Radiology 1989; 170(3, pt 1):823–829. Goodfellow J, Hungerford DS, Woods C. Patello-femoral joint mechanics and pathology. 2. Chondromalacia patellae. J Bone Joint Surg B 1976; 58:291–299. Thein R, Eichenblat M. Concealed knee cartilage lesions: is arthroscopic probing therapeutic? Am J Sports Med 1999; 27:495–499. Moseley JB, O’Malley K, Petersen NJ, Menke TJ, Brody BA, Kuykendall DH, et al. A Controlled trial of arthroscopic surgery for osteoarthritis of the knee. New Engl J Med 2002; 347:81–88.

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Original article

The tissue distribution of Evans blue dye in a sheep model of sentinel node biopsy Michael Greena, Gelareh Farshidb, James Kolliasc, Barry E. Chattertond and Chris Tsopelasd Background 99mTc-Evans blue is a ‘single dose’ agent for lymphatic mapping combining radioactivity and blue dye for sentinel node identification. The mechanism and distribution of blue dye retention in the lymph node is not clearly understood. Objective To demonstrate the cellular distribution of 99m Tc-Evans blue in sheep sentinel lymph nodes by measuring the radioactivity of different tissue components and correlating this with pathological examination. Methods 99mTc-Evans blue was used to identify sheep lymph nodes. Part of each node was sent for pathological examination including imprint cytology, and frozen and permanent section examination. Sections were examined without stains, with only red stains and conventional haematoxylin & eosin staining. The remaining nodal tissue was homogenized and components separated by enzymatic digestion and density gradient centrifugation. Fractions representing each tissue component were counted in a gamma counter and the distribution of 99m Tc-Evans blue calculated. Results A dispersed population of blue staining cells was found. Their distribution, number and size indicated that they were histiocytes such as macrophages or antigen

Introduction Sentinel lymph node biopsy is a valuable technique to assess the nodal stage of patients with early breast and other malignancies such as melanoma. This mode of focussed lymph node sampling has distinct advantages over lymph node clearance where all lymph nodes are removed in the surgical management of patients with cancer [1]. Techniques for localizing sentinel lymph nodes are well described in the literature. In general, two procedures are required: scintigraphic images obtained following injection of a radioactive colloidal agent and operative lymphatic tracing with a blue dye and a handheld isotope probe. The radiopharmaceutical is routinely injected several hours before surgery is planned. The blue dye is injected several minutes prior to surgery. These agents are administered at different times because of their different rates of migration through the lymphatic channels. Their rates of migration are different because

presenting cells. Radioactivity was distributed throughout the lymph node. Over 70% remained in the plasma, 19% in the leukocyte layer, and 10% was associated with erythrocytes and undigested tissue. Conclusion The accumulation of radioactivity and blue colour in the lymph nodes indicates the mechanism of retention is a result of the binding interaction between 99m Tc-Evans blue–protein and lymph node histiocytes including macrophages and antigen presenting cells. Nucl c 2006 Lippincott Williams & Med Commun 27:695–700 Wilkins. Nuclear Medicine Communications 2006, 27:695–700 Keywords: radioactive blue dye, Evans blue, lymphoscintigraphy, sentinel node biopsy, lymph node, histiocyte a Royal Hampshire County Hospital, Winchester, UK, bTissue Pathology Section, Institute of Medical and Veterinary Science, Adelaide, Australia, cBreast and Endocrine Unit and dNuclear Medicine Department, Royal Adelaide Hospital, Australia.

Correspondence to Dr Michael Green, Royal Hampshire County Hospital, Romsey Road, Winchester, Hampshire, S022 5D9. UK. Tel: + 44 (0)1962 824889; fax: + 44 (0)1962 824640; e-mail: [email protected] Received 15 December 2005 Accepted 23 May 2006

the radiocolloid is comprised of particles and the blue dye is a soluble substance. A ‘single dose’ agent combining both radioactivity and blue staining ability has been successfully prepared with a vital dye, Evans blue [2]. 99mTc-Evans blue (99mTc-EB) has been validated as a lymphoscintigraphic agent in a rabbit model [3], and its lymphatic uptake mechanism, lymphatic migration properties and physiological modes of excretion are also described [4]. 99mTc-EB is a soluble molecule, like other blue dyes used for the sentinel node biopsy, which transits the lymphatic system faster than radiocolloid particles such as 99mTc-antimony trisulphide. This property may facilitate the sentinel lymph node biopsy technique because lymphoscintigraphy can be performed immediately before surgery such that the lymphatic channels remain visible at surgery. Furthermore, since 99mTc-EB contains radioactive and blue

c 2006 Lippincott Williams & Wilkins 0143-3636

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696 Nuclear Medicine Communications 2006, Vol 27 No 9

signals, it is expected that the sentinel nodes will be both ‘hot’ and blue, contrary to those occasions where signals in the lymph nodes are discordant when different agents are used [5]. When Evans blue dye is injected into soft tissue most binds rapidly to endogenous proteins, primarily albumin. It is then transported in the lymph to the regional lymph nodes, beginning with the sentinel lymph node, where it may bind to collagen and other proteins of the lymph node stroma [6]. Brilliant blue G, which is structurally similar to EB, has been shown to associate with the outer surface cell wall of mouse B cells prior to diffusion via pores into the intracellular space [7]. Radiocolloid particles have been shown to be associated with macrophages or antigen presenting cells (APCs) within lymph nodes [8], but the mechanism of uptake and distribution of EB lymph nodes following locoregional injection has not been extensively studied. This information could potentially explain why certain agents are useful for highlighting the lymphatic system during the sentinel node biopsy. An ideal lymphatic mapping agent should be retained in the sentinel lymph node, such that it is better discriminated from less intense second and third tier lymph nodes, thereby lowering the false rate of negative biopsy. The aim of this study was to define the tissue distribution of 99mTc-EB dye, to enhance an understanding of its pharmacobiology, and as a prelude to further validation as a lymphatic mapping agent.

Methods General

Dulbecco’s modified Eagle’s medium (DMEM), bovine serum albumin (BSA; 7.5% w/v), fetal calf serum (FCS), hyaluronidase, collagenase, and centrifugation gradient solutions (Histopaque-1119; Histopaque-1077) were used in tissue processing (Sigma-Aldrich, NSW, Australia). DMEM stock contained DMEM:BSA (1:9). Evans Blue Sterile Injection (0.5% w/v; Pharmalab, NSW, Australia) was used as the raw material to prepare cold kits. EB cold kits containing dye (10 mg) and stannous chloride (144 mg) in water for injection (2 ml), were reconstituted with [99mTc]pertechnetate (90–100 MBq in 0.5 ml saline) and the product was analysed as described previously [4]. Only 99mTc-EB with greater than 95% radiochemical purity was used. All experiments were repeated in quadruplicate unless stated otherwise. Radiochemical analyses

Radiochemical analyses were performed using ascending instant thin-layer chromatography (ITLC) with silica gel impregnated glass fibre strips (1  16 cm; Gelman Sciences) developed in saline (0.9%). RF values were determined based on the definition: distance of radiotracer migrated divided by the distance travelled by the solvent. The strip sections (1 cm) were counted in a

gamma counter (Cobra II Auto-Gamma, Canberra Packard, Victoria, Australia) over a 99mTc window (90–190 keV), or in a large volume gamma counter (Biosentry, AEI-EKCO, Australia) linked to a multichannel analyser (Model 3100, Canberra Industries Inc., USA) over a 99m Tc window (70–210 keV). Sheep surgical procedure

Experiments were performed with male Merino sheep (4–6 tooth; 40–54 kg) and were compliant with the ‘Australian Code of Practice for the Care and Use of Animals for Scientific Purposes NHMRC’ and according to a protocol approved by the Animal Ethics Committee of the Institute of Medical and Veterinary Science, Adelaide, South Australia. A total of three sheep were used in the study. Each was pre-medicated by intravenous injection of thiopentone sodium (Pentothal, 12 mg
kg – 1) via the jugular vein. Anaesthesia was maintained by intubation of halothane in air (2.5 l
min – 1) plus oxygen (4 l
min – 1) throughout the entire procedure. Each sheep was secured by adhesive tape to an operating table in the supine position, the hind limbs were shaved of wool and then 99mTc-EB (5 ml) was injected subdermally at the hoof. The injection site was massaged for 2 min to facilitate migration. An incision was made over the popliteal and inguinal regions and each lymph node was identified by the blue colouration of the afferent lymphatic vessels and node as well as the radioactive counts identified with a hand-held gamma probe (SRP Mk II, Gammasonics, NSW, Australia). A total of three lymph nodes was then excised after 10–13 min and the sheep were killed by intracardiac puncture of pentobarbitone (Lethobarb; 325 mg
ml – 1, 25 ml). In a separate procedure with one sheep, a popliteal lymph node was harvested without using a radiotracer or blue dye. The node was cooled on ice for < 15 min until the in-vitro cannulation experiments with 99mTc-EB. Lymph node preparation and analysis

The harvested blue and radioactive inguinal lymph nodes (0.7–2.7 g) were processed to remove the adipose tissue, and then macerated with a scalpel. The finely divided tissue was incubated for 30–60 min at 371C in DMEM stock (17 ml) containing collagenase (15 mg) and hyaluronidase (1.7 mg) to release cells from the fibrous parenchyma. The digest was forced through a 100 mm nylon membrane filter (Steriflip unit, Millipore, Massachusetts, USA), the tissue mass was then washed with DMEM stock (2 25 ml) and the combined filtrates centrifuged at 400  g for 15 min (Cellsep 6/720R centrifuge; Sanyo Gallenkamp PLC, Loughborough, UK). After this time, the tissue pellet that incorporated all the major constituents of the lymph node was isolated and re-suspended in DMEM stock (5 ml), then carefully loaded on top of centrifugation gradient solutions of

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Distribution of Evans blue in a sheep model of SNB Green et al. 697

Histopaque-1077 (top layer, 1.7 ml; density = 1.077 g
ml – 1) plus Histopaque-1119 (bottom layer, 1.5 ml; density = 1.119 g
ml – 1). The constituents were separated by centrifugation at 800  g for 20 min at 221C to give plasma, leukocyte and gradient layers. Supernatant layers of each tube were each isolated using an insulin syringe and then counted in the gamma counter. The leukocyte layer was centrifuged (400  g for 15 min) and the resulting pellet was washed with saline (2  1 ml). All fractions were counted in a gamma counter. The distribution of 99mTc-EB in each layer was calculated as a percentage of the total radioactivity for each tube. In another experiment, the harvested popliteal lymph node was cleaned of adipose tissue and 99mTc-EB solution was injected at the hilum. The node (1.84 g) was incubated in DMEM stock (20 ml) at 371C. After 40 min, the node was bisected and half was submitted for pathological examination while the second half was macerated with a scalpel and similarly treated as above (30 min enzyme digest) to ultimately calculate the percent 99mTc-EB in each layer per tube. Control centrifugations 99m

Tc-EB (B2 MBq; 0.1 ml) was added to BSA solution (2 ml) and incubated at 371C for 10 min. A sample (0.1 ml) diluted in saline (0.5 ml) was loaded onto the gradient solutions Histopaque-1077 (1.7 ml) plus Histopaque-1119 (1.5 ml), and then centrifuged at 800  g for 20 min at 221C. The supernatant layers of each tube were each isolated using an insulin syringe and then counted in the gamma counter. The distribution of 99mTc activity in each layer was calculated as a percentage of the total radioactivity for each tube. Radiochemical analysis of the layers was also performed using ITLC–silica gel paper/ saline solvent. In another experiment, 99mTc-EB ( < 1 MBq; 0.04 ml) was centrifuged and analysed in the same way as above.

Histological methods

The fresh lymph node was sliced at 2-mm intervals. By pressing dry glass slides onto the cut surfaces of the lymph node, imprints of the node were prepared. Some of the imprints were examined without further processing. Others were stained rapidly with a modified Haem-Qwik stain (HD Scientific, NSW, Australia), using only the fixative, first staining solution (red stain), but omitting the second stain (coloured blue) to prevent masking of any 99mTc-EB-stained cells. Part of the nodal tissue was frozen in liquid nitrogen, and sections were prepared from this portion of the node. Some of these frozen sections were examined histologically without fixation or staining, and others were stained only with eosin, or only with haematoxylin & eosin (H&E). The remaining portions of the node were fixed in 4% buffered formalin and subsequently embedded in paraffin. Histological

sections from this material were examined after staining with H&E.

Results Surgical exposure

The lymphatic anatomy in the hind limb of one sheep was exposed to show blue (and radioactive) contiguously linked popliteal and inguinal lymph nodes at 13 min after injection. Distribution of

99m

Tc-EB in lymph nodes

After centrifugation of the lymph node tissue samples, three distinct aqueous layers were visible, and a sticky pellet deposited at the bottom of each tube. The top band was identified as the plasma layer, containing free 99m Tc-EB and as protein-bound 99mTc-EB. This was shown in the control centrifugation experiments such that 100% colour and radioactivity for both 99mTc-EB and 99m Tc-EB–proteins were located there. The percent 99m Tc-EB in lymph node tissue is summarized in Table 1. At least half of the total radioactivity was found in the plasma layer containing both 99mTc-EB and 99mTc-EB– protein. A chromatographic analysis of one of these fractions (60 min incubation), using ITLC-SG paper developed in saline, did separate the minor component 99m Tc-EB (RF = 0.64–0.92) from the 91.8% of the coloured complex 99mTc-EB–protein (RF = 0.0–0.42). The leukocyte fraction was comprised of monocytes, neutrophils, lymphocytes and platelets, and it accounted for 12–19% of the radioactivity. When the leukocytes were repeatedly washed with saline, there was 87.1 ± 0.6% (n = 4) 99mTc-EB associated with cells and 13.4 ± 0.7% (n = 4) in the soluble fraction. The bottom layer (the density gradient solution) comprised a minor proportion of the total radioactivity. The pellet was comprised of erythrocytes and fibrous tissue fragments from the lymph node stroma. With an extended incubation to 60 min, far less pellet was evident, and the pellet activity was low enough (6%) to suggest this period was optimal for lymph node tissue digestion. A lower pellet activity resulted in a higher plasma activity at 60 min, Table 1

Percent

99m

Tc-Evans blue dye in lymph node tissue Percent

Centrifugation gradient layer

Popliteal lymph node*

99m

Tc activity (mean ± SE, n = 4) Inguinal lymph node**

30 min incubation*** 30 min incubation*** 60 min incubation*** Plasma Leukocytes Bottom Pellet

51.3 ± 0.3 13.4 ± 0.3 6.1 ± 1.1 29.2 ± 2.1

52.0 ± 0.4 11.9 ± 0.4 6.1 ± 0.5 30.0 ± 3.2

71.5 ± 1.3 19.3 ± 1.3 3.2 ± 0.5 5.9 ± 0.6

*

In-vitro injection via hilium. Harvested from sheep at 10 min post-injection. Tissue digested with collagenase–hyaluronidase at 371C in DMEM.

**

***

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698 Nuclear Medicine Communications 2006, Vol 27 No 9

indicating that most of the pellet activity at 30 min was due to 99mTc-EB–protein trapped inside the stroma. The cannulated popliteal lymph node gave a similar 99mTc-EB distribution as the inguinal node after incubation with enyzmes at 371C for 30 min.

Fig. 2

Histological analyses

Unstained imprints and frozen sections from the popliteal node (in-vitro injected sample) demonstrated blue staining in some cells (Fig. 1). These were dispersed as single cells that constituted no more than 5% of the population of the lymph node. They varied little in their size and shape, and all of these cells were stained blue, without accentuation of either the nuclear or the cytoplasmic regions. In frozen sections, cells were more abundant in the subcapsular area of the node (Fig. 2). The sections stained with the modified Haem-Qwik stain and the eosin stain, showed retention of the blue stained cells. The background lymphocytes highlighted by the stain, were found to be slightly smaller and more abundant than the blue staining cells (Fig. 3a and b). There was little variability in cell size or shape of the blue stained cells, suggesting that they were a single population, probably histiocytes. The H&E stained sections showed nodal tissue with retention of the normal architecture and histology. The blue-stained cells were no longer identifiable in these preparations.

Discussion Blue dyes used in lymphatic mapping are known to transit the lymphatics rapidly, and for them to be useful in assisting surgical identification and excision of the sentinel lymph node, they must significantly concentrate Fig. 1

Frozen section of lymph node not stained with laboratory reagents. A population of blue staining cells (A) is identified, accentuated in the perimeter of the tissue, consistent with a subcapsular location, and also seen more sparsely in the central portion of the node (B). The tissue cracking is an artefact of frozen section preparation.

Frozen section of lymph node not stained with laboratory reagents. Higher magnification. The blue staining cells (A) are accentuated at the perimeter of the node. Compared to the pale (unstained) lymphocytes seen in the background, they are far less numerous and slightly larger. There is little variability in nuclear size or shape among the blue cells, suggesting that they may constitute a single population.

in the lymphatics and then fix within the microanatomy of the sentinel lymph node. In principle, the same should apply to the radiotracer used for scintigraphic detection. However, previous studies have demonstrated as few as 70% of sentinel lymph nodes to be both hot and blue when the staggered, two agent technique was used [5]. If the dye transits the sentinel lymph node too quickly it may be confused with second or third tier lymph nodes. Thus for 99mTc-EB, it is important to determine the distribution and proportion of the blue colour and radioactivity that is retained in a lymph node. Certainly from the surgeon’s point of view, in this simple lymphatic drainage model, there were adequate visual and radioactive signals present in the lymph nodes at operation at 20 min after subdermal injection. Our in-vitro results showed a significant proportion of 99mTc-EB was indeed incorporated into the cells of the lymph node or became bound to the parenchyma. The lymphatic uptake of 99mTc-EB is linked with protein binding ability. Dye injected into the dermis diffuses through the cellular matrix, and non-specifically enters the blood capillaries and lymphatic vessels. Evans blue interacts with lymphatic proteins and the resulting complex is retained within the lymphatic system [9]. The dye–protein complex migrates with flowing lymph to a lymph node in the sequence, where it becomes concentrated. From our results, this radioactive complex was found to be associated with leukocytes to the extent of B20%. The 99mTc-labelled dye was tightly bound to these cells such that washing them repeatedly with saline released a minor amount of the isotope.

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Distribution of Evans blue in a sheep model of SNB Green et al. 699

Fig. 3

these cell types were surface stained, which is a prerequisite condition for phagocytosis. This follows other work where fluorescein-labelled human serum albumin was shown to be associated with APCs in porcine sentinel lymph nodes using the fluorescenceactivated cell sorter technique [8]. The 99mTc-dye– protein interaction with histiocytes results in retention of radioactive and blue signals within the lymph node. Evans blue is known to be metabolized in the liver by reductive cleavage with cytochrome C (P-450) reductase to yield aromatic amines, which are then excreted in the urine [10]. Following the initial surface interaction between 99mTc-EB–protein and histiocyte, the course of the radioactive dye thereafter is not clear. The P-450 enzyme is known to exist in lymph node cells, highly concentrated in the nuclei of lymphoid cells and macrophages [10,11]. If the 99mTc-EB is indeed phagocytosed by macrophages in the lymph node, it would be expected to lose the blue colour, but not radioactivity, due to scission of the azo chemical bond. The time taken to achieve engulfment of this antigen is unknown, but it may not be important from a sentinel node biopsy perspective.

Frozen sections of lymph node stained briefly with eosin examined at low (a) and high (b) power. The blue staining cells are seen against a larger background population of pale eosinophilic cells, consistent with lymphocytes.

The histological examination of cells from the second layer of the spin density gradient was limited by the processing technique, requiring numerous aqueous and alcohol-based reagents, that ultimately extracted proteinbound Evans blue from the tissue samples. Nevertheless, there was sufficient dye present to permit visual analysis, when compared with unstained imprints and frozen sections of nodes. 99mTc-EB was clearly localized to a small number of dispersed cells in the lymph node, the size, number and distribution of these being consistent with histiocytes or antigen presenting cells. An immunohistochemical analysis of these cells could prove useful, although the additional processing steps required may render the cells of interest visually indistinguishable from their neighbours. From the results in this study, it cannot be elucidated whether 99mTc-EB is internalized by APCs, although the histological observations indicated that

A low level of 99mTc-EB identified in the plasma layer of the centrifugation gradient confirmed most of the 99mTcEB was protein-bound, and perhaps the free 99mTc-EB was liberated during enzymatic processing of the lymph node specimens. The plasma activity was considerably higher than leukocytes activity (3.7-fold), indicating the concentration of 99mTc-EB–protein exceeded that of histiocytes in the node. This correlates with the observation that blue colour was visible beyond the first (popliteal) lymph node onto the next tier (inguinal) node in the sheep. Those factors that govern the surface attachment of 99mTc-EB–protein to histiocytes appear to be the binding affinity of antigen to cell, as well as residence time in the lymph node. The latter concept is the subject of current investigation in a sheep model by this laboratory.

Conclusion This study has demonstrated that although much of the 99m Tc-EB remains bound to endogenous proteins in lymph, a significant proportion of it is bound to histiocytes present in the subcapsular area of the sentinel lymph node. The evidence is based on centrifugation density gradient separation to isolate the constituents of the lymph node, and microscopic analysis of those cells. The 99mTc-EB–protein complex is believed to be attached onto the surface of histiocytes and may be internalized, and this is the mechanism of retention that concentrates radioactive and blue signals during the sentinel node biopsy procedure.

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700 Nuclear Medicine Communications 2006, Vol 27 No 9

Acknowledgements We are grateful to the staff of the Animal Care Facility, Institute of Medical and Veterinary Sciences, Adelaide, South Australia, for their assistance in preparing sheep prior to surgery.

References 1

2 3

4 5

Luini A, Gatti G, Ballardini B, Zurrida S, Galimberti V, Veronesi P, et al. Development of axillary surgery in breast cancer. Ann Oncol 2005; 16: 259–262. Tsopelas C. Technetium-99m labeling of a molecule bearing easily reducible groups. Nucl Med Biol 2000; 27:797–802. Sutton R, Tsopelas C, Kollias J, Chatterton BE, Coventry BJ. Sentinel node biopsy and lymphoscintigraphy with a technetium 99m labeled blue dye in a rabbit model. Surgery 2002; 131:44–49. Tsopelas C, Penglis S. Visualisation of lymphatic flow in a rabbit model using 99m Tc-labelled dyes. Hellenic J Nucl Med 2000; 3:96–101. Martin 2nd RC, Edwards MJ, Wong SL, Tuttle TM, Carlson DJ, Brown CM, et al. Practical guidelines for optimal gamma probe detection of sentinel

lymph nodes in breast cancer: results of a multi-institutional study. For the University of Louisville Breast Cancer Study Group. Surgery 2000; 128:139–144. 6 Heinle H, Lindner V. The binding of Evans blue to collagen and elastin in elastic tissue. Arch Int Physiol Biochim 1984; 92:13–17. 7 Neumann E, Toensing K, Kakorin S, Budde O, Frey J. Mechanism of electroporative dye uptake by mouse B cells. Biophys J 1998; 74: 98–108. 8 Faries MB, Bedrosian I, Reynolds C, Nguyen HQ, Alavi A, Czerniecki BJ. Active macromolecule uptake by lymph node antigen-presenting cells: a novel mechanism in determining sentinel lymph node status. Ann Surg Oncol 2000; 7:98–105. 9 Tsopelas C, Sutton R. Why certain dyes are useful for localizing the sentinel lymph node. J Nucl Med 2002; 43:1377–1382. 10 Muskhelishvili L, Thompson PA, Kusewitt DF, Wang C, Kadlubar FF. In situ hybridization and immunohistochemical analysis of cytochrome P450 1B1 expression in human normal tissues. J Histochem Cytochem 2001; 49:229–236. 11 Borodin YI, Safina AF, Maiborodin IV, Grishanova AY. Benzo[a]pyrene induction of cytochrome P450 1A1/1A2 in the lymph nodes of rats. Bull Exp Biol Med 2003; 136:611–614.

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Original article

A technique for analysis of geometric mean renography John S. Fleming Background and objectives Renography is used routinely to assess relative right to left renal function. Quantification is usually carried out using posterior images. Errors in relative renal function may occur if the kidneys are at different depths. Geometric mean images from combined anterior and posterior views are much less affected by kidney depth and offer the opportunity of more accurate and precise quantification. Background subtraction is a key part of the analysis process and validated protocols for geometric mean imaging have not been devised. This study aims to derive a suitable background subtraction protocol for geometric mean imaging. Methods Simultaneous anterior and posterior renography using 99mTc mercaptoacetyltriglycine (MAG3) was performed on 16 adults. Analysis was carried out using both geometric mean and posterior images. The geometric mean background subtraction protocol was modified to give the same results as a posterior method, which had previously been validated by correlation with measurements of glomerular filtration rate. Absolute and relative uptakes were then obtained from both geometric mean and posterior analyses. For each analysis values were obtained both with and without depth correction. Results A revised background subtraction protocol for geometric mean renography was devised which operated

Introduction Radionuclide renography is used extensively in the assessment of renal function [1]. Parameters describing both the ability of the kidneys to extract radiopharmaceuticals from the blood and also their rate of transit through the kidney are obtained. One particular parameter of interest that is regularly used clinically is the relative, right to left, renal function (RRF). There are a number of factors that affect the accuracy with which RRF can be obtained from renography [2,3]. If only RRF is required, then static renal imaging using 99mTcdimercaptosuccinic acid (99mTc-DMSA) is preferable. However, where renal transit is being measured it is often useful to have an assessment of RRF, and therefore methods of improving the accuracy of its assessment from renography are of value. Relative renal function is estimated by measuring the relative uptake of radiopharmaceutical at about 2 min following injection. The limited time for assessment of uptake is due to the requirement that the measurement

successfully on all studies. Both absolute renal uptake and relative function values obtained from geometric mean analysis were not systematically different from those obtained using posterior analysis with depth correction. Values of the relative renal function from posterior analysis after depth correction were closer to the geometric mean values than estimates obtained before correction. Conclusion A technique for analysing geometric mean renography data has been developed which gives results consistent with a previously validated posterior-only c 2006 Lippincott method. Nucl Med Commun 27:701–708
Williams & Wilkins. Nuclear Medicine Communications 2006, 27:701–708 Keywords: renography, geometric mean imaging, quantitative analysis

Departments of Medical Physics and Bioengineering and Nuclear Medicine, Southampton University Hospitals NHS Trust, UK.

Correspondence to Professor John S. Fleming, Department of Nuclear Medicine, Southampton General Hospital, Southampton SO16 6YD, UK. Tel: + 44 (0)2380 796202; fax: + 44 (0)2380 796927; e-mail: [email protected]

Received 12 April 2006 Accepted 31 May 2006

must be taken before the minimum transit time through the kidney [4]. In fact for centres using furosemide injection prior to or at the same time as injection, the transit times are even shorter, which means that this assessment needs to be brought still earlier, probably to around 1.5 min [5]. The short time period following injection during which the estimate of renal uptake needs to be made influences the accuracy with which RRF can be assessed. At this stage in the renogram procedure, there is a relatively high contribution to counts in the renal region of interest (ROI) from background activity and this provides a particularly important error component in the assessment of RRF [2]. There is also a significant component of error due to attenuation of gamma rays in tissue. Most renography studies are carried out using the posterior view. Counts in the region of interest will be attenuated by a varying amount, depending on the thickness of tissue between the kidney and camera. It is often assumed that the depths of the kidneys are similar so that the attenuation

c 2006 Lippincott Williams & Wilkins 0143-3636


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702 Nuclear Medicine Communications 2006, Vol 27 No 9

factor can be taken as the same for both kidneys. If this is true then the relative counts in the kidney will give an accurate estimate of RRF. While this is often accepted as a reasonable approximation in small children [6], studies, primarily with DMSA, have shown that for a reasonable proportion of adult patients this assumption leads to significant errors in RRF [7]. Methods of correcting for this have used either ultrasound [8], computed tomography (CT) images [9] or lateral radionuclide images taken at the end of the dynamic renography study [10]. All these methods have their drawbacks. Ultrasound or CT images are not always available and also would only apply to supine renography studies. Lateral images taken at the end of the renography study suffer from a variety of complicating factors: (1) the images are often of poor quality due to low counts and contrast, as the kidneys may have low activity at the end of the study; (2) the spatial distribution of activity at the end of the study will be different from that in the first 2 min following injection when kidney uptake is being evaluated; (3) it is often difficult to distinguish the two kidneys as their images on a lateral view overlap; and (4) determining the posterior edge of the patient is subject to some uncertainty due to patient positioning and low counts. Nevertheless measurements of kidney depth using the lateral image technique have been used to calculate absolute values of radiopharmaceutical uptake, which correlate well with glomerular filtration rate in the same patients [11]. While this study provides some confidence that the systematic errors of the method are reasonably low, there are clearly still considerable random errors on individual measurements for the reasons described above.

for use with geometric mean renography by calibrating them against a validated posterior-only method.

It is therefore reasonable to consider the use of geometric mean renography as an alternative method for allowing for depth differences when calculating RRF. Geometric mean imaging is commonly used with DMSA imaging [12]. Geometric mean counts are dependent on the total attenuation thickness through the body [13] and the assumption is made that this thickness is the same for both kidneys. It is of note that while this seems intuitively a reasonable assumption, the errors involved have never been rigorously evaluated. However, total renal uptake from geometric mean measurements in renography with 99mTc-diethylenetriaminepentaacetic acid (99mTc-DTPA) has been used to calculate glomerular filtration rate [14]. The values obtained were shown to correlate well with blood sample measurements.

The posterior dynamic study was analysed using the standard departmental analysis technique. Four regions were drawn manually covering (1) right and (2) left kidneys, (3) a background area between the upper poles of the kidneys and (4) the left ventricle. Time–activity curves were then created and the renal and background curves smoothed by a variable amount depending on the maximum count [2]. The smoothing protocol used n passes of a (1-2-1) filter, subject to a minimum of two, where

Background subtraction is known to be a key component of error in assessing renal function from renography [2]. Protocols for background correction of counts in both renal and cardiac regions have been derived for posterior images. Their application has been validated against measurements of glomerular filtration rate from 99m Tc-DTPA renography [11]. The purpose of this study was to devise suitable background subtraction protocols

Methods Acquisition of patient data

A series of 16 adult patients referred to our department for routine renography were selected for study. The age range varied from 21 to 78 years. Subjects were hydrated prior to injection of 80 MBq 99mTc-labelled mercaptoacetyltriglycine (MAG3). They were imaged supine between the two heads of a Philips Genesys gamma camera (Philips Nuclear Medicine, Milpitas, California) positioned to provide both anterior and posterior views of the heart and kidneys. Diuretic was given at the time of injection. Digital acquisition of 100 frames of 12 s duration was carried out and the data transferred to a Link MAPS 10000 computer where the analysis was performed. A geometric mean dynamic study was created from the anterior and posterior views. The software flipped the anterior image about the centre of the matrix and formed the geometric mean image with the posterior on a pixel-by-pixel basis. It was assumed that the camera had been calibrated for the centre of rotation so that the images were perfectly aligned. The alignment of the centre of rotation with the image matrix was checked at regular intervals and the camera was re-calibrated if necessary. At the end of the dynamic acquisition, the gamma camera was rotated through 901 and a 60-s static acquisition obtained which gave lateral views of the left and right kidneys. Derivation of background subtraction factors for geometric mean analysis

1

c =2 ð1Þ n ¼ 12  15 and c is the count per frame in the kidney curve at 2 min. Extra-organ background was subtracted from both renal and cardiac curves as described previously [2,15]. Prior to subtraction, the background curve was corrected for the relative areas of the ROIs and by a further factor to allow for the fact that the extra-renal background tissue did not extend the full thickness of the subject whereas that of the background region did. These factors (0.87 for the right kidney, 0.79 for the left kidney and 0.57 for the heart) had been derived for imaging with 123I-hippuran but are assumed to apply to other radiopharmaceuticals as

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Technique for analysis of geometric mean renography Fleming 703

they principally depend on the relative thickness of the kidney compared to the thickness of the patient. The assumption depends on the distribution in vascular and extravascular space being similar for the different radiopharmaceuticals. The peak time of the cardiac curve was determined automatically and taken as zero time for the analysis. Background subtraction using this regime has been validated for DTPA by comparing the percentage uptake at 2 min with glomerular filtration rate (GFR). The resulting regression curve passed close to the origin indicating the accuracy of the subtraction technique [11]. Kidney depth was derived from the lateral views by manually defining the renal region of interest and two points on the posterior border of the body. The depth (d) was calculated from the difference in the anterior– posterior direction between the centre of gravity of the kidney ROI and the mean position of the two points [16]. The counts in each renogram after extra-renal background subtraction were summed for the two frames covering 1.2–1.6 min following the heart peak. This count was taken as a measure of the uptake of MAG3 in each kidney. The time period for assessing this uptake was earlier than the conventional time of 2 min, due to the use of the F0 protocol. It has been observed that this protocol can lead to very rapid transit through the kidney [5]. In the current study, peak times of the renogram following the heart peak were as low as 1.6 min. The background subtracted count (Cpbs) was corrected for renal depth using the equation Cpk ¼ Cpbs expðmd Þ

ð2Þ

where Cpk is the corrected count and m is the broad-beam attenuation coefficient for 140 keV gamma rays, which was taken as 0.12 cm – 1. This was taken as the estimate of the true count rate in the kidney. Although the value will be subject to random error due to the difficulty of subtracting background and estimating kidney depth, the systematic error should be small by virtue of the good correlation between renal uptake of 99mTc-DTPA assessed in this way and GFR [11]. Geometric mean renogram curves were produced using the same regions of interest as for the posterior image analysis. The counts in the renogram between 1.2 and 1.6 min uncorrected for background were calculated for the renal (Cgmr) and background (Cgmb) ROIs. The kidney counts corrected for extra-renal background (Cgmkbs) were considered as: Cgmkbs ¼ Cgmr  fCgmb

Ak Ab

ð3Þ

where f is the fraction by which the geometric mean background count needs to be multiplied to give the extra-renal contribution in the renal ROI. Ak and Ab were the areas of the kidney and background regions, respectively. This count was corrected for attenuation

using measurement of the total thickness of the body at the level of the kidneys from the lateral image (T) [13]. The anterior and posterior borders were marked manually from a visual inspection of the images. Both right and left lateral images were used and the values averaged.
 mT : ð4Þ Cgmk ¼ Cgmkbs exp 2 It was then possible to estimate the optimum value of the geometric mean background factor f by equating the two estimates of the count rate in the kidney from the posterior and geometric mean views, Cpk and Cgmk, respectively.    Cpk Ak f ¼ Cgmr  : ð5Þ Cgmb expðmT=2Þ Ab The fraction f was estimated from Equation 5 for each subject separately for the right and left kidneys and the mean values calculated: fr and fl, respectively. For renal analysis using either deconvolution or the Patlak–Rutland plot, it is necessary to estimate the input function. This is usually done from the time–activity curve over the cardiac region. It has been shown previously that the most accurate estimation of the input curve is obtained when the cardiac curve is corrected for background [17,18]. Previous studies showed that it was necessary to multiply the area corrected background curve by a factor to obtain the correct variation of vascular activity [17]. It was assumed that the factor for the geometric mean cardiac curve, fc, would be different from that of the posterior. To calculate this factor, it was assumed that the systematic error in the background subtracted posterior cardiac curve was small and that the value of the 15 min to 2 min count ratio from this curve Rp was a true representation of the vascular activity ratio. The background subtracted geometric mean cardiac curve, Cgmcbs(t), was defined as Ac Cgmbcs ðt Þ ¼ Cgmc ðt Þfc Cgmb ðt Þ Ab where fc is the unknown fraction of the background curve to be subtracted from the cardiac curve for geometric mean images. The geometric mean cardiac and background curves were evaluated at 2 min and 15 min. The geometric mean 15 min to 2 min vascular activity ratio Rgm was a function of fraction fc. The correct value of fc was taken as that which gave a value of Rgm equal to Rp. Thus   fc ¼ Cgmc ð15Þ  Cgmc ð2ÞRp   Ab=A c  : ð6Þ Cgmb ð15Þ  Rp Cgmb ð2Þ fc was evaluated for all patients who had the left ventricle fully visualized in the field of view of the gamma camera. This applied to ten of the patients in this study. The mean value of the fraction was calculated.

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Technique for geometric mean analysis

The software currently used for renogram analysis for posterior view renography was modified slightly for the analysis of geometric mean renograms. The background subtraction correction factors of each kidney and the cardiac regions were replaced with values based on the above experiments. In clinical practice it was found that application of the mean background subtraction correction factors for the renal regions occasionally resulted in over-subtraction of the background when renal function was very poor. Over-subtraction was recognized by the presence of negative counts in the renogram following subtraction. Therefore the value of the factor used was the mean multiplied by 0.8. This represented the mean minus two standard deviations of the average coefficient of variation of all factors. The mean and applied factors for left and right kidneys for both posterior and geometric mean images are shown in Table 1. This process introduces a small systematic error in extra-renal background subtraction, but this is almost completely corrected in the subsequent vascular subtraction, which is performed as a part of deconvolution analysis described below. The net errors are therefore very small and use of the technique has the advantage of avoiding the relatively large errors that can occasionally occur in poorly functioning kidneys due to over-subtraction. Background subtraction of the cardiac curve used the mean correction factor and no obvious cases of over-subtraction were observed for this curve. The number of smooths required for the renal time– activity curves was defined by the equation pffiffiffi pffiffiffiffiffiffiffi 2 cgm n ¼ 12  ð7Þ 15 where cgm is the geometric mean count per frame in the renal region at 2 min. This is analogous to Equation 1 with the signalpffiffito ffi noise ratio assumed to be increased by a factor of 2 compared to the posterior-only situation. The background-subtracted renograms were then analysed by deconvolution using the matrix method, with the background subtracted cardiac curve as the input function. The plateau (P) of the retention function was Table 1 Background subtraction correction factors for left and right kidney and cardiac regions for posterior and geometric mean image analysis Image type

Factor value

Right kidney

Left kidney

Left ventricle

Posterior

Mean Standard deviation Applied

0.87 0.11 0.70

0.79 0.06 0.63

0.57 0.08 0.57

Geometric mean

Mean Standard deviation Applied

0.79 0.08 0.63

0.72 0.08 0.57

0.36 0.14 0.36

Both the mean factors and those applied in clinical practice are shown. These factors apply to a background area between the upper half of the kidneys

calculated from the mean value of the curve between 1.2 and 1.6 min. Intra-renal vascular background was subtracted by back-extrapolation of the plateau value to zero time and replacement of the retention function values before 1.2 min with the plateau level P1. The corrected retention function was then reconvolved and renal uptake assessed by integrating the curve between 1.2 and 1.6 min. The relative function was calculated from the ratio of counts in the right kidney to the sum of the counts in both kidneys. In calculating the RRF for the posterior data, the counts were corrected for attenuation using the measured depth of the kidney in Equation 2. In contrast, when determining the relative function from geometric mean analysis, it is not necessary to correct for attenuation. It is assumed that the total thickness of the body at the level of the kidneys is equal on both sides and therefore that the attenuation is the same. In order to compare absolute counts, the geometric mean data were also corrected for attenuation using Equation 4. The total thickness was obtained from the lateral images as described above. The mean transit time and renal outflow efficiency were also calculated for each kidney [4] for both the posterior and geometric mean data. Calculation of specific uptake size index

Background subtraction has been shown to be a major source of error in renography analysis [2]. To assess the relative importance of background in posterior and geometric mean analyses, the specific uptake size index (SUSI) was calculated [19]. This is a measure of the total uptake in an organ relative to the surrounding background. It is defined by SUSI ¼ Cs=b where Cs is the specific uptake in the kidney in counts per second summed between 1.2 and 1.6 min. These are the counts remaining after background subtraction but before attenuation correction. b is the corresponding counts per second per square centimetre in the central area background corrected by the appropriate background subtraction correction factor. SUSI values were calculated for both the posterior and geometric mean images. In a two-dimensional image SUSI is the area of background activity that has the same uptake as the specific uptake in the organ. In order to compare the statistical quality of the data obtained in the posterior-only and geometric mean views, the summed counts in the renal ROI for the posterior only view before attenuation correction were compared with the summed counts from anterior and posterior views. Statistical analysis

Systematic differences between parameters were assessed by the paired t-test. Differences in random uncertainties were compared by the F-test.

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Technique for analysis of geometric mean renography Fleming 705

Background correction fractions were determined for 15 of the 16 subjects in the study. In the other subject it was considered that the lateral depths could not be determined with sufficient confidence for one of the kidneys due to overlying activity from a ‘hot’ contralateral kidney and other organs. The average background subtraction correction fractions are shown in Table 1. All values for the geometric mean analysis were significantly lower (P < 0.05) than the posterior values determined in earlier experiments [15,17]. The geometric mean values were higher for the right kidney than for the left by a similar degree to that found for the posterior images. The standard deviations for geometric mean analysis were similar to those previously found for the posterior. Values were also consistently lower for the left ventricle ROI compared to the renal ROIs.

Fig. 1

(a) Relative renal function right geometric mean (%)

Results

100 Line of identity

80

60

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Relative renal function right posterior (%)

The comparison between absolute count rates at 1.4 min obtained from depth corrected posterior and geometric mean counts is shown in Fig. 2. There was no systematic difference between the values but the random coefficient of variation was 9.6%. The comparison between mean transit time values calculated from posterior and geometric mean analyses is shown in Fig. 3. The geometric mean values were systematically higher by 6% (P < 0.005). The renal outflow efficiency values from the geometric mean analysis were significantly lower by 2% (P < 0.05). The apparent reduction in renal outflow efficiency (ROE) is consistent with the apparent increase in mean transit time (MTT) due to the expected inverse relationship between these parameters [4]. The comparison of the SUSI values for posterior and geometric mean images is shown in Fig. 4. The SUSI values for the posterior image were systematically higher than for the geometric mean. The mean values were 164 cm2 and 104 cm2, respectively. These were signifi-

(b) Relative renal function right posteriorno depth correction (%)

The comparison of right RRF from the geometric mean analysis with that from the posterior-only analysis with depth correction is shown in Fig. 1(a). There was no systematic difference between the values, and the root mean square difference was 2.3 percentage points (coefficient of variation 4.3%). The comparison between posterior analysis with and without depth correction is shown in Fig. 1(b). There was a systematic difference between the values (P < 0.05). This was associated with the average right kidney depth (9.7 cm) being significantly greater than the left (9.0 cm) (P < 0.05). The root mean square difference between posterior RRF with and without depth correction was 4.1 percentage points (coefficient of variation 7.9%). This was significantly higher than for the comparison between geometric mean and depth-corrected posterior values (P < 0.05).

100 Line of identity

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Relative renal function right posterior (%) (a) Variation of relative renal function (RRF) evaluated from the geometric mean analysis with that from posterior-only analysis with correction for depth. (b) Variation of RRF from posterior analysis with and without depth correction.

cantly different (P < 0.001). The reduced SUSI for the geometric mean image was caused by the renal counts on the anterior image being lower than for the posterior giving a reduced value to the geometric mean. The background counts were not significantly different: 0.81 countss – 1cm – 2 and 0.83 countss – 1cm – 2, respectively. The mean total summed counts contributing to the renal geometric mean value was however higher than for the posterior, 197 countss – 1 compared to 138 countss – 1 (P < 0.001) (Fig. 5). The total contribution to the geometric mean background count was approximately twice that of the geometric mean value and therefore twice that of the posterior alone (P < 0.001).

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Fig. 2

Fig. 4

400 Line of identity SUSI Geometric mean (cm2)

Renal uptake at 1.4 min geometric mean (c/s)

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(cm2)

Comparison of attenuation-corrected counts in each kidney between posterior-only and geometric mean analyses.

Comparison of specific uptake size index (SUSI) for each kidney between posterior-only and geometric mean analyses.

Fig. 3

Fig. 5

14000

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Mean transit time posterior (min) Comparison of mean transit time through each kidney between posterior-only and geometric mean analyses.

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Posterior counts at 1.4 min Comparison of total counts in each kidney between posterior-only and geometric mean analyses.

Discussion One of the principal sources of error in assessing both absolute and relative renal function from renography using posterior imaging is due to attenuation of gamma rays. It has been shown that reasonably accurate assessment of renal depth is possible from lateral imaging.

However, there are a number of problems associated with the measurement, which result in a loss of precision in individual measurements. In some cases the measurements cannot be made at all due to overlying activity from the contra-lateral kidney. This was the situation for

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Technique for analysis of geometric mean renography Fleming 707

one of the patients studied. Geometric mean imaging represents a relatively simple adaptation to conventional posterior renography which helps to overcome this issue. The reduction in counts in a geometric mean image due to attenuation is dependent on the total thickness of the body [13]. To obtain absolute renal counts the attenuation factor can either be obtained by measuring the total body thickness [13] or by making a transmission measurement [20]. If only relative function is required then it can be assumed to a reasonable approximation that attenuation of left and right kidney counts is equal. This should, in principle, simplify the accurate assessment of relative function. However, background subtraction techniques are often specifically tailored for posterior-only imaging. This study has developed a background subtraction protocol specifically for geometric mean imaging that is consistent with a previously validated posterior-only technique. The methodology for developing the background subtraction protocols was successful in that when the technique was applied, there was no systematic difference in the renal counts or relative function between posterior and geometric mean images. This was not surprising as the same set of data was used in deriving and applying the technique. Further testing of the technique on different subjects will be required to fully evaluate the new method and this will be the subject of future work. The background subtraction factors for left and right kidneys were systematically lower than the values previously derived for the posterior image alone (Table 1). There is no obvious reason for this. In fact, intuitive reasoning would suggest that the factor should be higher for the geometric mean image. Since the average depth of the extra-renal background is less for the anterior view than the posterior, the background contribution should be higher. When this is combined with the posterior view to give the geometric mean this would be expected to increase rather than decrease the background subtraction fraction. The most probable alternative cause of this effect is that distribution of activity in vascular and extravascular space was different for the MAG3 used in this study compared to hippuran on which the historical background factors were based [21]. Both posterior and geometric mean factors were higher for the right kidney than for the left. This reflects the higher contribution of activity in the liver which partially overlays the right kidney region. The factors for the left ventricle region were lower than those for the kidneys for both posterior and geometric mean images. This can be explained by the presence of air in the background tissue over and underlying the heart. A smaller fraction of the counts in the background region between the kidneys was therefore required to represent the extra-cardiac tissue in the left ventricular ROI. As for the kidney regions the geometric mean factor

for the cardiac region was lower than that of the posterior view alone. If the difference in background subtraction factors for posterior-only and geometric mean analyses is due to the difference in distribution pattern between hippuran and MAG3, then the validity of using the posterior factors derived for hippuran to analyse MAG3 renography may be in question. However, this issue is only likely to be of academic interest. It is unlikely that it will result in any major errors in assessing renal uptake and therefore on relative renal function. Any small systematic error in extra-renal background subtraction due to this factor will be compensated in practice by the subsequent vascular background subtraction step. The relatively low coefficient of variation of 2.3% between estimates of RRF from geometric mean images and posterior images with depth correction suggested that both techniques were operating with reasonable precision. However, it should be noted that the depth correction technique for the posterior-only analysis could not be applied in one of the 16 studies. This is consistent with our clinical experience where the lateral images are difficult to interpret in a small percentage of patients. The interoperator variability of the geometric mean technique should also be better than for the posterior-only method as the determination of renal depth is performed manually in the posterior analysis. A study of inter-operator variability will be the subject of further work. There was somewhat unexpectedly a small but significant difference between RRF assessed by posterior analysis with and without attenuation correction (Fig. 1(b)). The RRF on the right was higher for the corrected results. This was explained by the significantly greater depth for the right kidney in this patient group. The tendency for the right kidney to be deeper has been observed previously [22]. However, the mean kidney depth in that study was less than in the current investigation (7.5 cm compared to 9.4 cm) and the corresponding difference in left and right depths was only 1 mm compared to 7 mm. Although of academic interest, this small systematic difference is not of great practical importance. The important practical finding was that the RRF values from posterior analysis after attenuation correction were closer to the geometric mean values than estimates obtained before correction. This demonstrated the important effect of attenuation on RRF values (up to 9 percentage points in this study) and that both methods of correction were producing consistent results. The assessment of absolute counts in each kidney also gave no systematic difference between the geometric mean technique and the attenuation corrected posterior assessment. The coefficient of variation of 9.6% was significantly higher than for the RRF. There were random errors in both cases due to the assessment of attenuation

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thickness from lateral measurements. In principle, errors for the geometric mean technique should be less than this if transmission measurements were used to determine attenuation correction factors [14]. This study has not been able to assess the accuracy of either method, as the true value of renal uptake is not known. The most promising approach to evaluation of accuracy is to use realistic simulated renograms and work has commenced towards the production of such data [23]. The influence of technique on assessment of transit was also investigated by assessing the MTT and ROE by both methods. There were small systematic differences for both parameters with the geometric mean technique predicting slightly slower transit. However, the differences were small: 6% in the case of MTT and 2% for ROE. These are considered sufficiently small not to have any significant clinical impact. Two sources of error were also compared: background and statistical noise. The ratio of renal uptake to background was assessed by the SUSI and found to be lower for the geometric mean technique (Fig. 4). In a previous simulation study, errors in background subtraction were shown to be an important factor in determining the precision of relative function measurements [2]. Therefore errors due to this factor will be higher for the geometric mean method and this represents probably the only technical disadvantage of the geometric mean method. The total count acquired in the assessment of a geometric mean count is considerably higher than that from the posterior alone (Fig. 5). Both the total renal count and background count are improved giving better precision in the subtracted counts. There are some minor practical considerations of geometric mean renography. It means using a dual-head gamma camera, which will generally increase the cost of the study slightly. It also means that the study will probably have to be performed supine. Erect or semirecumbent renography may be possible with some designs of dual-head camera but will certainly be practically more difficult than the supine position. Supine renography may result in poorer drainage compared to a study performed in the erect position. It should therefore be useful to include in the imaging protocol a delayed static image in the erect position to allow for hold-up resulting from this factor. In conclusion, this paper has described a method of analysis for geometric mean renography data, which gives results consistent with a previously validated posterioronly method.

Acknowledgement I would like to acknowledge the help of Sandra Johns in setting up the acquisition protocol for this investigation.

References 1

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Oei HY. Dynamic and static renal imaging. In: Murray IPC, Ell PJ (editors): Nuclear Medicine in Clinical Diagnosis and Treatment. Edinburgh: Churchill Livingstone; 1994, pp. 213–227. Gullquist RR, Fleming JS. Error analysis by simulation studies in renography deconvolution. Phys Med Biol 1987; 32:383–395. Houston AS, Whalley DR, Skrypnuik JV, Jarritt PH, Fleming JS, Cosgriff PS. UK audit and analysis of quantitative parameters obtained from gamma camera renography. Nucl Med Commun 2001; 22:559–566. Fleming JS, Kemp PM. A comparison of deconvolution and the Patlak–Rutland plot in renography analysis. J Nucl Med 1999; 40:1503–1507. Donoso G, Kuyvenhoven JD, Ham H, Piepsz A. 99mTc-MAG3 diuretic renography in children: a comparison between F0 and F + 20. Nucl Med Commun 2003; 24:1189–1193. Tindall A, Prescod N, Clarke SE. The value of geometric mean divided function measurements in routine 99mTc-DMSA studies [Abstract]. Nucl Med Commun 1998; 19:376. Wujanto MB, Lawson RS, Prescott MC, Test HJ. The importance of using anterior and posterior views in the calculation of differential renal function using Tc-99m DMSA. Br J Radiol 1987; 60:869–872. Gruenewald SM, Collins LT, Fawdry RM. Kidney depth measurement and its influence on quantitation of function from gamma camera renography. Clin Nucl Med 1985; 6:398–401. Maneval DC, Magill HL, Cypess AM, Rodman JH. Measurement of skin-tokidney distance in children: implications for quantitative renography. J Nucl Med 1990; 31:287–291. Kohn HD, Mostbeck A. Value of additional lateral scans in renal scintigraphy. Eur J Nucl Med 1979; 4:21–25. Fleming JS, Keast CM, Waller DG, Ackery DM. Measurement of glomerular filtration rate with Tc-99m DTPA – a comparison of gamma camera methods. Eur J Nucl Med 1987; 13:250–253. Fleming JS, Cosgriff PS, Houston AS, Jarritt PH, Skrypniuk JV, Whalley DR. United Kingdom audit of relative renal function measurement using DMSA scintigraphy. Nucl Med Commun 1998; 19:989–997. Fleming JS. A technique for the absolute measurement of activity using a gamma camera and computer. Phys Med Biol 1979; 24:176–180. Carlsen O. The gamma camera as an absolute measurement device: determination of glomerular filtration rate in 99mTc-DTPA renography using a dual head gamma camera. Nucl Med Commun 2004; 25:1021–1029. Kenny RW, Ackery DM, Fleming JS, Goddard BA, Grant RW. Deconvolution of the scintillation camera renogram. Br J Radiol 1975; 48:481–486. Steinmetz AP, Zwas ST, Macadziob S, Rotemberg G, Shrem Y. Renal depth estimates to improve the accuracy of glomerular filtration rate. J Nucl Med 1998; 39:1822–1825. Fleming JS. Measurement of hippuran plasma clearance using a gamma camera. Phys Med Biol 1977; 22:526–530. Fleming JS. Estimation of organ input function and plasma clearance from the cardiac curve in dynamic scintigraphy. Eur J Nucl Med 1992; 19:248–253. Fleming JS, Bolt L, Stratford JS, Kemp PM. The specific uptake size index for quantifying radiopharmaceutical uptake Phys Med Biol 2004; 49:N227–N234. Macey DJ, Marshall R. Absolute quantitation of radio tracer uptake in the lungs using a gamma camera. J Nucl Med 1982; 23:731–735. Taylor A, Eshima D, Fritzberg AR, Christian PE, Kasina S. Comparison of iodine-131 OIH and technetium-99m MAG3 renal imaging in volunteers. J Nucl Med 1986; 27:795–803. Taylor A, Curtis C, Giacometti A, Hall EC, Barefield KP. Improved formulas for the estimation of renal depth in adults. J Nucl Med 1993; 24:1766–1769. Dawson AH, Fleming JS, Hoffmann SMA, Papaspyrou L, Peel S. Computer simulation of gamma camera images of the kidney [Abstract]. Nucl Med Commun 2006; 27:285.

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Original Article

Minimally invasive radio-guided parathyroidectomy: long-term results with the ‘low 99mTc-sestamibi protocol’ Domenico Rubelloa,*, Giuliano Marianib,*, Adil Al-Nahhasc and Maria R. Pelizzod,* Background This paper reports the results of minimally invasive radio-guided surgery (MIRS) performed in a large group of 280 consecutive patients affected by primary hyperparathyroidism (PHPT) and with a high probability of being affected by a solitary parathyroid adenoma before surgery. Methods The probability of a solitary parathyroid adenoma was established by evaluating the patients with a single-day imaging protocol based on parathyroid double-tracer scintigraphy and high resolution neck ultrasonography. MIRS was performed successfully in 269 (96.1%) patients. MIRS consisted of a very low dose (37 MBq (1 mCi)) of 99mTc-sestamibi, given by intravenous injection, in the operating theatre a few minutes before surgery, thus allowing the radiation exposure dose to the patient and operating theatre personnel to be minimized (< 1.5 lSv
h – 1 to the surgeon). Results No major intraoperative complication was recorded in our series. Transient hypocalcaemia was observed in 8% of patients. MIRS required a mean duration time of 33 min and a mean hospital stay of 1.2 days. Local anaesthesia was successfully performed in 71 patients, 63

Introduction Bilateral neck exploration (BNE) is still considered the ‘gold standard’ approach in some centres for treating patients with primary hyperparathyroidism (PHPT) [1]. However, the surgical approach to PHPT has changed in many surgical centres during the last decade, moving from BNE to unilateral or limited neck exploration, followed by minimally invasive approaches in the form of minimally invasive radio-guided surgery (MIRS). This change has been brought about by (1) the knowledge that PHPT is caused by a solitary parathyroid adenoma in the majority of cases (85% or more), and (2) the technical improvements achieved in surgical practice. The latter includes the introduction of microsurgical instruments such as the endoscope, the intraoperative gamma probe, and the measurement of intraoperative quick parathyroid hormone (QPTH). For these reasons, the current practice of more than half of endocrine surgeons is * On behalf of the Italian Study Group on Radioguided Surgery and Immunoscintigraphy (GISCRIS)

of whom were elderly and with concomitant invalidating diseases contraindicating general anaesthesia. No case of disease relapse was observed during the subsequent follow-up. Conclusion MIRS using the ‘low sestamibi protocol’ is a safe and effective treatment in PHPT patients with a high likelihood of a solitary parathyroid adenoma at preoperative imaging. Nucl Med Commun 27:709–713

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:709–713 Keywords: primary hyperparathyroidism, sestamibi, high-resolution neck ultrasonography, intraoperative probe, minimally invasive radio-guided surgery a

Nuclear Medicine Service, ‘S. Maria della Misericordia’ Hospital, Rovigo, Italy, Regional Center of Nuclear Medicine, University of Pisa Medical School, Italy, c Department of Nuclear Medicine, Hammersmith Hospital, London, UK and d Department of Surgery, University of Padova Medical School, Italy. b

Correspondence to Dr Adil Al-Nahhas, Department of Nuclear Medicine, Hammersmith Hospital, Du Cane Road, London W12 0HS, UK. Tel: + 0044 208 383 4923; fax: + 0044 208 383 1700; e-mail: [email protected] Received 25 April 2006 Accepted 8 June 2006

minimally invasive parathyroidectomy based on the removal of the solitary parathyroid adenoma throughout a small skin incision of 1–2 cm [2]. Unlike in BNE, the minimally invasive approach depends on preoperative imaging and it is essential to (1) establish whether the parathyroid adenoma is solitary, (3) precisely locate the depth of the parathyroid adenoma, and (3) evaluate the presence of possible concomitant nodular thyroid diseases [3,4]. The present study deals with MIRS obtained in a large group of 280 consecutive patients homogeneously investigated preoperatively and operated on by the same surgeon using the ‘low sestamibi protocol’ with an intraoperative gamma probe.

Patients, materials and methods Two hundred and eighty consecutive patients affected by PHPT were included in the study. Clinical presentation included bone pain with osteoporosis (63), bilateral renal stones (15%) or a combination of osteoporosis and renal stones (14%). The remaining 12% of patients were

c 2006 Lippincott Williams & Wilkins 0143-3636

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asymptomatic and hyperparathyroidism was incidentally diagnosed at routine biochemical screening. PHPT was biochemically proven with raised total and ionized serum calcium levels, and raised parathyroid hormone serum levels. There were 179 females and 101 men, mean age 55.8 years (range 18–83 years). Fifty-five patients (20%) had undergone previous thyroid or unsuccessful parathyroid surgery at other centres. We adopted the following inclusion criteria for MIRS: (1) evidence at [99mTc]pertechnetate/99mTc-sestamibi subtraction (sestamibi) scintigraphy of a solitary parathyroid adenoma, (2) a clear sestamibi uptake in the parathyroid adenoma with a parathyroid adenoma to thyroid ratio > 1.2, (3) absence at sestamibi scintigraphy and high resolution – 10 MHz – neck ultrasound of concomitant thyroid nodules, (4) no history of familial hyperparathyroidism or multiple endocrine neoplasia, and (5) no history of neck irradiation. Seventy-eight patients were excluded from MIRS due to concomitant thyroid nodules (49), negative or inconclusive sestamibi scan (21), familial hyperparathyroidism (three), history of multiple endocrine neoplasia (four) or history of neck irradiation (one). No patient who had been offered MIRS refused this treatment. Preoperative imaging procedures included a singlesession sestamibi scintigraphy and neck ultrasound as previously described [5–8]. In patients with concordant sestamibi/ultrasound findings (both positive or negative) no further imaging was performed, while in discordant cases (sestamibi positive but ultrasound negative) a sestamibi tomographic (SPECT) examination (n = 73 patients) was also obtained to investigate a possible ectopic or deep position of the parathyroid adenoma. SPECT was obtained just after the completion of the planar sestamibi scintigraphy, thus using the same sestamibi dose; negating the need for reinjection and avoiding additional radiation exposure to the patient and nuclear medicine personnel. The scintigraphic images were interpreted by two skilled nuclear medicine physicians (D.R. and G.M.), and in cases of discrepancy (2%), final diagnosis was reached by consensus. In patients with a normal thyroid gland confirmed by [99mTc]pertechnetate scintigraphy and ultrasound, a single focus of sestamibi uptake was judged to be consistent with a solitary parathyroid adenoma while in patients in whom at least two foci of sestamibi uptake were shown, a multi-gland disease was diagnosed. In patients with concomitant sestamibi positive thyroid nodules, ultrasound and [99mTc]pertechnetate thyroid scintigraphy were used to distinguish parathyroid adenoma from sestamibi-avid thyroid nodule(s).

Scintigraphic imaging

Planar sestamibi scintigraphy was acquired with a largefield-of-view (LFOV) gamma camera (Orbiter, 7500, Siemens, Hoffman Estates, Illinois, USA) equipped with a parallel-hole, low-energy, high-resolution collimator. Images were stored in a 128  128 matrix and processed using a dedicated computer. Tomographic sestamibi SPECT acquisition was performed by a dual-head gamma camera (Axis, Picker International, Cleveland, Ohio, USA) equipped with a parallel-hole, low-energy, ultra-high-resolution collimator. The following parameters were adopted: elliptical orbit, 120 steps, 30 s per step and 64  64 matrix. Images were reconstructed using a low-pass filter, cut-off 0.2–0.25, order 5.0–6.0, and processed using a dedicated computer. Three-dimensional analysis was also obtained. Neck ultrasound

Neck ultrasound was performed by high-resolution (7.5– 10 MHz) transducer (Technos, Esaote, Italy). Longitudinal and axial neck scans were obtained from the angle of the mandible to the sternal notch. The parathyroid adenoma was identified on grey-scale imaging as a hypo-echoic nodule distinct from the thyroid gland. After operation, the surgeon was asked to judge the degree of probe usefulness during operation according to the following four-point scale: ‘not useful’, ‘slightly useful’, ‘useful’ and ‘very useful’. Steps of the

Steps for minimally invasive radio-guided surgery for solitary parathyroid adenoma using the ‘low sestamibi dose’ protocol

Table 1

Step number Procedure 1

2 3

4

5

6

7 8 9 10

Blood samples are drawn from a peripheral vein both before commencing surgery and 10 min after removal of parathyroid adenoma to measure intraoperative quick parathyroid hormone levels Sestamibi (37 MBq) is injected in the operating room 10 min before the start of surgery Prior to surgical incision, the patient’s neck is scanned with an 11-mm collimated probe to localize the high count rate area corresponding to the cutaneous projection of the parathyroid adenoma A transverse midline neck access (approximately 1 cm above the sternal notch) is preferred because conversion to bilateral neck exploration is easily obtainable if necessary The probe is repeatedly inserted through a 2-cm skin incision guiding the surgeon to the maximum count rate area corresponding to the parathyroid adenoma In some patients with a parathyroid adenoma located deep in the neck, ligature of the middle thyroid vein and inferior thyroid artery is required Radioactivity is measured with the probe on the parathyroid adenoma, thyroid gland and background Radioactivity is measured on the ex-vivo parathyroid adenoma to evaluate successful removal of parathyroid tissue Radioactivity is checked on the empty operating basin to evaluate completeness of parathyroid tissue extirpation Tissue ratios are calculated (parathyroid-to-background ratio and parathyroid-to-thyroid ratio)

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Tc-sestamibi in minimally invasive radio-guided parathyroidectomy Rubello et al. 711

intraoperative technique used in our centre for MIRS are summarized in Table 1. Intraoperative probe

A hand-held commercially available collimated gamma probe (Scintiprobe MR 100, Pol.hi.tech, Italy) was used. The probe had the following technical characteristics: linear collimated probe with an external diameter of 11 mm, mounting a NaI scintillation detector; lateral shielding efficiency of 99% for 99mTc; energy range of 30–385 keV; gamma detecting efficiency of 99% for the 99m Tc peak energy (140 keV); sensitivity threshold of 37 Bq (10 nCi) and spatial resolution of 5 mm at a distance of 2 mm. Intraoperative quick parathyroid hormone

Intraoperative QPTH was measured by imunochemoluminescent assay (Liason, Byk Gulden, Italy). A fall of 50% or more in PTH levels 10 min after removal of parathyroid adenoma in comparison with the baseline pre-excision value was considered indicative of a successful parathyroidectomy. Additional blood samples for QPTH measurement were obtained in patients with multi-gland disease after removal of any hyperfunctioning parathyroid gland. All operations were performed by the same surgeon (M.R.P.). Post-surgical follow-up ranged between 20 and 79 months, mean ± SD = 16.4 ± 8.2 months.

cases (71.3%) and ‘very useful’ in 55 cases (20.5%). The probe was very useful in 18 ectopic parathyroid adenomas located in the mediastinum, two ectopic parathyroid adenomas located at the carotid bifurcation, and 24 parathyroid adenomas located deep in the neck in the para-retroesophageal/para-retrotracheal space. The mean operating time was 34 min (range, 15–60 min) and the mean hospital stay was 1.2 days (range, 1–2 days). Local anaesthesia was successfully performed in 71 patients, the majority of whom (n = 63) were elderly and with concomitant invalidating diseases contraindicating general anaesthesia. No major surgical complication (e.g., laryngeal nerve palsy, permanent hypoparathyroidism) was recorded. Transient hypocalcaemia was observed in 8% of patients. A small (1.5–2 cm) skin incision was enough to perform MIRS. It is worth noting that MIRS was successfully performed in 42/55 patients (76.4% of cases) who had previously received thyroid or unsuccessful parathyroid surgery at other centres. We did not perform a randomized study to compare MIRS with traditional bilateral neck exploration in patients with a high pre-operative probability of a solitary parathyroid adenoma. However, historical data derived from our centre showed that the operating time was approximately double when using bilateral neck exploration (60– 120 min) with a hospital stay of 3–5 days.

All patients gave their written informed consent before the performance of minimally invasive radio-guided surgery. All patients received clinical and laboratory surveys 1 month after surgery and subsequently every 2–3 months, thereafter.

The mean parathyroid adenoma-to-background (P/B) ratio was rather high (mean 2.7, range 1.3–6.8) (Fig. 1) and the lowest P/B ratio observed in our series was 1.3. The mean parathyroid adenoma-to-thyroid (P/T) ratio was also relatively high (mean 1.5, range 1.1–4.8) (Fig. 2) in the presence of a normal thyroid gland and the lowest P/T ratio in our series was 1.1. Although some patients

Data are expressed as mean ± 1 standard deviation (SD). Mean values were compared using Student’s t-test. P values lower than 0.05 were considered significant.

Fig. 1

Results Based on preoperative scintigraphy and ultrasound results, MIRS was offered to 280 patients, and in 269 of them (96.1% of cases) it was performed successfully. A conversion to BNE was required in 11 patients. In two of these, this was due to intraoperative diagnosis (frozen section) of a parathyroid carcinoma; in four, it was due to a persistently high QPTH level after removal of the preoperatively visualized parathyroid adenoma (in all four cases a second parathyroid adenoma was found during subsequent BNE); and in five, the parathyroid adenoma was situated in an unusual position, and was either large or difficult to remove with MIRS. Considering the group of 269 patients in whom MIRS was successfully preformed, the surgeon judged the gamma probe as ‘slightly useful’ in 22 cases (8.2%), ‘useful’ in 192

Parathyroid-to-background ratio

8 7 6 5 4 3 2 1 0

0

50

100 150 200 Number of patients

250

300

Distribution of parathyroid-to-background ratio intraoperatively measured by a gamma probe in a series of 280 primary hyperparathyroid patients treated by sestamibi radio-guided surgery.

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correctly suggested a deep location of parathyroid adenoma which was confirmed at subsequent surgery.

Fig. 2

Parathyroid-to-thyroid ratio

6

The mean weight of the excised solitary parathyroid adenoma by MIRS was 981 ± 323 mg (range 120– 4050 mg). The parathyroid glands removed in patients with multi-gland disease were significantly smaller (mean weight = 524 ± 411 mg, range 100–2100; P < 0.05). The lowest parathyroid adenoma visualized at preoperative scintigraphy weighed 120 mg and was found in a patient with a solitary parathyroid adenoma.

5 4 3 2 1 0

Discussion 0

50

100 150 200 Number of patients

250

300

Distribution of parathyroid-to-thyroid ratio intraoperatively measured by a gamma probe in a series of 280 primary hyperparathyroid patients treated by sestamibi radio-guided surgery.

Two main conditions clearly favoured the development of MIRS: (1) improvements achieved in preoperative localization imaging mainly related to sestamibi scintigraphy [15–20], and (2) the introduction in clinical practice of intraoperative QPTH measurements [21].

Fig. 3

6

Parathyroid-to-thyroid ratio

The first experience in limited neck exploration by adopting a unilateral approach was carried out by Tibblin in the early 1980s [9]. In their protocol, the parathyroid adenoma was removed and the ipsilateral parathyroid gland was biopsied to disclose possible glandular hyperplasia. More recently, minimally invasive endoscopic [10,11] and probe-guided [3–8,12–14] approaches were developed.

5

As previously mentioned, and in contrast with BNE, MIRS requires stringent inclusion criteria including (1) a high likelihood of a solitary parathyroid adenoma demonstrated by preoperative sestamibi/ultrasound imaging, (2) intense sestamibi uptake in the parathyroid adenoma, and (3) absence of concomitant sestamibi-avid thyroid nodules. Using these selection criteria, two thirds of PHPT patients could be offered MIRS as an alternative to BNE [3–8,12–14].

4 3 2 1 0 0

1

2 3 4 5 6 Parathyroid-to-background ratio

7

8

Relationship between the parathyroid-to-thyroid ratio and the parathyroid-to-background ratio measured by a gamma probe in a series of 280 primary hyperparathyroid patients treated by sestamibi radio-guided surgery.

showed a relatively low intraoperative P/T ratio, the corresponding P/B ratio was higher because the parathyroid adenoma was well separated from the thyroid gland and easily detected by the gamma probe (Fig. 3). Lastly, it is worth noting that the normal parathyroid glands did not show any significant uptake intraoperatively when checked by the probe. In the group of patients in whom SPECT was performed, it is worth noting that in 27 cases, SPECT analysis

The combination of sestamibi scintigraphy and neck ultrasound provides clear advantages such as (1) more accurate information about parathyroid adenoma localization (site, depth), (2) differentiation of solitary parathyroid adenoma from multi-gland disease, and (3) evaluation of presence of co-existing nodular goitre [3–8,14]. The addition of SPECT acquisition to planar sestamibi scintigraphy improves sensitivity and accuracy of the imaging procedure [22,23]. In our protocol, ultrasonography is systematically combined with sestamibi scintigraphy while SPECT is usually reserved to patients with ectopic glands or discordant sestamibi (positive)/ultrasound (negative) results. The first MIRS protocol, developed by Norman and Chheda in 1997 [12], involved a single-day imaging and surgery approach. The patient is injected with 740– 925 MBq (20–25 mCi) sestamibi, with images obtained by

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Tc-sestamibi in minimally invasive radio-guided parathyroidectomy Rubello et al. 713

dual-phase technique, followed by MIRS within 2–3 h from radiotracer administration. This protocol is attractive from a cost-analysis perspective because sestamibi scintigraphy and MIRS are performed on the same day with a single dose of sestamibi for both scintigraphic imaging and radio-guided surgery. However, it also presents some practical disadvantages given the uncertainty of the scintigraphic results and the differences between the MIRS and BNE with respect to the need for operating theatre time (BNE > MIRP) and efficient patient scheduling. This approach would be more prohibitive in areas with high prevalence of nodular goitre making a different-day protocol more appropriate [14]. The protocol we used is a different-day protocol where, on the first day, localization is obtained by means of a double-tracer [99mTc]pertechnetate/99mTc-sestamibi subtraction scintigraphy combined with neck ultrasound. MIRS is usually performed within 1 week from imaging, where a low dose (37 MBq (1 mCi)) of sestamibi is given directly in the operating theatre, a few minutes before surgery. The ‘low sestamibi dose protocol’ we used provides two main advantages: (1) less radiation exposure to the patient and operating room personnel and (2) fewer false negative results in parathyroid adenoma with rapid sestamibi washout.

2

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12

Favourable results have been reported with both Norman’s ‘high sestamibi dose’ protocol and our ‘low sestamibi dose’ protocol with a success rate in the intraoperative detection of parathyroid adenoma higher than 95%, without major intraoperative surgical complications. It appears reasonable to assume that Norman’s single-day protocol is more suitable for patients with a low likelihood of nodular goitre, while our different-day protocol is more appropriate in areas with a higher prevalence of nodular goitre.

13

Another advantage of MIRS in respect to BNE is the possibility to perform a minimally invasive neck exploration under local anaesthesia with early (mostly same-day) hospital discharge [14]. However, in elderly and medically complex patients in whom general anaesthesia would be contraindicated, we prefer the discharge to be on the second postoperative day [24,25].

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In conclusion, MIRS appears to be a safe and effective therapeutic approach for PHPT patients with a high probability of a solitary parathyroid adenoma. Preoperative accurate imaging, especially with sestamibi scintigraphy, is strongly recommended. Lastly, our data indicate that a low dose of sestamibi (37 MBq (1 mCi)) administered in the operating theatre a few minutes before surgery is adequate to successfully perform MIRS.

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Clark OH. Surgical treatment of primary hyperparathyroidism. Adv Endocrinol Metab 1995; 6:1–16.

Sackett WR, Barraclough B, Reeve TS, Delbridge LW. Worldwide trends in the surgical treatment of primary hyperparathyroidism in the era of minimally invasive parathyroidectomy. Arch Surg 2002; 137:1055–1059. Rubello D, Pelizzo MR, Casara D. Nuclear medicine and minimally invasive surgery of parathyroid adenomas: a fair marriage [Editorial]. Eur J Nucl Med 2002; 30:189–192. Rubello D, Casara D, Pelizzo MR. Symposium on parathyroid localising imaging. Optimization of peroperative procedures. Nucl Med Commun 2003; 24:133–140. Casara D, Rubello D, Piotto A, Pelizzo MR. 99mTc-MIBI radio-guided minimally invasive parathyroid surgery planned on the basis of a preoperative combined 99mTc-pertechnetate/99mTc-MIBI and ultrasound imaging protocol. Eur J Nucl Med 2000; 27:1300–1304. Casara D, Rubello D, Pelizzo MR, Shapiro B. Clinical role of 99mTcO4/MIBI scan, ultrasound and intra-operative gamma probe in the performance of unilateral and minimally invasive surgery in primary hyperparathyroidism. Eur J Nucl Med 2001; 28:1351–1359. Rubello D, Casara D, Giannini S, Piotto A, De Carlo E, Muzzio PC, et al. Importance of radio-guided minimally invasive parathyroidectomy using hand-held gamma probe and low 99mTc-MIBI dose: technical considerations and long-term clinical results. Q J Nucl Med 2003; 47:129–138. Rubello D, Piotto A, Casara D, Muzzio PC, Shapiro B, Pelizzo MR. Role of gamma probes in performing minimally invasive parathyroidectomy in patients with primary hyperparathyroidism: optimization of preoperative and intraoperative procedures. Eur J Endocrinol 2003; 149:7–15. Tibblin S, Bondeson AG, Ljubgberg O. Unilateral parathyroidectomy in hyperparathyroidism due to a single adenoma. Ann Surg 1982; 195: 245–252. Gagner M. Endoscopic parathyroidectomy [Letter]. Br J Surg 1996; 83:875. Henry JF, Iacobone M, Mirallie E, Deveze A, Pili S. Indications and results of video-assisted parathyroidectomy by a lateral approach in patients with primary hyperparathyroidism. Surgery 2001; 130:999–1004. Norman J, Chheda H. Minimally invasive parathyroidectomy facilitated by intraoperative nuclear mapping. Surgery 1997; 122:998–1004. Costello D, Norman J. Minimally invasive radioguided parathyroidectomy. Surg Oncol Clin N Am 1999; 8:555–564. Mariani G, Gulec SA, Rubello D, Boni G, Puccini M, Pelizzo MR, et al. Preoperative localization and radioguided parathyroid surgery. J Nucl Med 2003; 44:1443–1458. Coakley AJ, Kettle AG, Wells CP, O’Doherty MJ, Collings REC. 99mTcsestamibi – a new agent for parathyroid imaging. Nucl Med Commun 1989; 10:791–794. Taillefer R, Boucher Y, Potvin C, Lambert R. Detection and localization of parathyroid adenomas in patients with hyperparathyroidism using a single radionuclide imaging procedure with technetium-99m-sestamibi (doublephase study). J Nucl Med 1992; 33:1801–1807. O’Doherty MJ, Kettle AG, Wells P, Collins REC, Coakley AJ. Parathyroid imaging with technetium-99m-sestamibi: preoperative localization and tissue uptake studies. J Nucl Med 1992; 33:313–318. Hindie´ E, Melliere D, Jeanguillaume C, Perlemuter L, Chehade F, Galle P. Parathyroid imaging using simultaneous double-window recording of technetium-99m-sestamibi and iodine-123. J Nucl Med 1998; 39:1100–1105. Coakley AJ. Symposium on parathyroid localization (Editorial). Nucl Med Commun 2003; 24:111–113. Rubello D, Saladini G, Casara D, Borsato N, Toniato A, Piotto A, et al. Parathyroid imaging with pertechnetate plus perchlorate/MIBI subtraction scintigraphy. A fast and effective technique. Clin Nucl Med 2000; 25: 527–531. Irvin GL, Dembrow VD, Prudhomme DL. Clinical usefulness of an intraoperative ‘quick PTH’ assay. Surgery 1993; 114:1019–1023. Sfakianakis GN, Irvin GL 3rd, Foss J, Mallin W, Georgiou M, Deriso GT, et al. Efficient parathyroidectomy guided by SPECT-MIBI and hormonal measurements. J Nucl Med 1996; 37:798–804. Moka D, Voth E, Dietlein M, Larena-Avellaneda A, Schicha H. Technetium 99m-MIBI-SPECT: a high sensitive diagnostic tool for localization of parathyroid adenomas. Surgery 2000; 128:29–35. Rubello D, Pelizzo MR, Boni G, Schiavo R, Vaggelli L, Villa G, et al. Radioguided surgery of primary hyperparathyroidism using the low-dose 99m Tc-sestamibi protocol: multi-institutional experience from the Italian Study Group on Radioguided Surgery and Immunoscintigraphy (GISCRIS). J Nucl Med 2005; 46:220–226. Rubello D, Casara D, Giannini S, Piotto A, Dalle Carbonare L, Pagetta C, et al. Minimally invasive radioguided parathyroidectomy: an attractive therapeutic option for elderly patients with primary hyperparathyroidism. Nucl Med Commun 2004; 25:901–908.

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Original article

Accurate diagnosis of acute pyelonephritis: How helpful is procalcitonin? Ayfer G. Gu¨vena, Halis Z. Kazdala, Mustafa Koyuna, Funda Aydınb, Fırat Gu¨ngo¨rb, Sema Akmana and Yunus Emre Baysala Aim This prospective study aimed to investigate the diagnostic value of serum procalcitonin levels in children with acute pyelonephritis documented by 99m Tc-dimercaptosuccinic acid (DMSA) scintigraphy. Methods We compared the symptoms and laboratory findings of fever, vomiting, abdominal/flank pain, leukocyte count, serum C-reactive protein and procalcitonin levels with the results of the DMSA scan obtained within the first 72 h after referral in children who were diagnosed as having acute pyelonephritis. Thirty-three children (31 female and two male) aged 1–11 years (mean 4.42 years) were enrolled in this prospective study. Results Twenty-one of 33 patients (64%) had positive DMSA scans. On the scans obtained after 6 months, five of 21 patients (23.8%) had renal scars. No correlation was found between clinical and laboratory parameters, alone or combined with each other, and positive DMSA scans. Serum procalcitonin levels were 0.767 ± 0.64 and 1.23 ± 1.17 ng
ml – 1 in children with normal and positive DMSA scans, respectively. The cut-off value for procalcitonin using receiver operating characteristic analysis was 0.9605 ng
ml – 1, while sensitivity and specificity were 86.4% and 36.4%, respectively. However, if the cut-off value was chosen as 2 ng
ml – 1, the sensitivity

Introduction Commonly used clinical and laboratory parameters are not specific in the differentiation of renal parenchymal involvement from lower urinary tract infection (UTI) especially in young children. This may contribute to delay in the diagnosis and treatment of acute pyelonephritis (APN), and conversely may lead to over-diagnosis and unnecessary invasive investigations [1,2]. In these circumstances, 99mTc-dimercaptosuccinic acid (DMSA) scintigraphy is accepted as the ‘gold standard’ in the diagnosis of APN [3,4]. Differentiating APN from lower UTI is important because of the risk of renal scar formation and its subsequent complications, such as hypertension, proteinuria, hypostenuria and chronic renal failure. Development of renal scarring is closely associated with a delay in the diagnosis and appropriate treatment of APN [5]. In a recent study it was demonstrated

increased to 100% while specificity did not change markedly. Conclusion The serum procalcitonin test, like other commonly used laboratory parameters, e.g. serum C-reactive protein and white blood cell count, was inadequate in distinguishing renal parenchymal involvement in acute febrile urinary tract infections. Nucl c 2006 Lippincott Williams & Med Commun 27:715–721 Wilkins. Nuclear Medicine Communications 2006, 27:715–721 Keywords: procalcitonin, pyelonephritis, UTI, DMSA, children Departments of aPaediatrics and Division of Nephrology and bNuclear Medicine, Akdeniz University, School of Medicine, Antalya, Turkey. Correspondence to Dr Fırat Gu¨ngo¨r, Department of Nuclear Medicine, School of Medicine, Akdeniz University, Antalya 07070, Turkey. Tel: + 0090 532 446 8538; fax: + 0090 242 227 4490; e-mail: [email protected] These data were presented, in part, at the 13th Congress of the International Paediatric Nephrology Association, Adelaide, Australia, 29 August to 2 September 2004, and published in abstract form in Paediatric Nephrology 2004:19(9), C207 Received 11 April 2006 Accepted 28 June 2006

that renal scarring could be prevented if antibiotic administration was started within 24 h after the symptoms began [6]. Procalcitonin (PCT), which is a propeptide of calcitonin with 116 amino acids, was initially described as a potential marker of bacterial infections [7]. Recent reports stressed that there was a correlation between plasma PCT levels and renal parenchymal damage in UTIs [8–11]. This study prospectively evaluated a group of children diagnosed as APN on an outpatient basis. The aims of the study were to investigate the diagnostic value of serum procalcitonin levels by comparing them with conventional clinical and laboratory parameters in children with APN documented by DMSA scintigraphy, and whether PCT was able to identify children with renal parenchymal involvement soon after admission.

c 2006 Lippincott Williams & Wilkins 0143-3636

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716 Nuclear Medicine Communications 2006, Vol 27 No 9

Patients, materials and methods Thirty-three children who were referred to the paediatric department of Akdeniz University Hospital between May 2002 and June 2004, and diagnosed as APN, were consecutively enrolled in this prospective study. The prediagnosis of APN was made by paediatric residents using clinical and laboratory findings as fever (axillary, Z 38.51C), abdominal/flank pain (in older children), vomiting, leukocytosis (white blood cell (WBC) count, Z 10 000 mm – 3), high levels of C-reactive protein (CRP), and pyuria ( Z 8 leukocytes per high-power field) and bacteriuria in urinary sediment. Patients with a history of documented previous febrile UTI, with a congenital urinary tract anomaly or systemic disease or who were younger than 1 year of age were excluded from the study. The patients who fulfilled the criteria defined above were included in the study after the consent of the parents. We defined APN as UTI with renal parenchymal involvement documented by DMSA scintigraphy, and lower UTI as UTI without renal parenchymal involvement on a DMSA scan. Urinary catheterization was used to collect urine samples from children who had not completed toilet training. The mid-stream technique was used for children who were toilet trained. Urinary sediment was examined microscopically by residents. Double urine samples were obtained for urine culture. Significant bacteriuria was defined as Z 105 and Z 104 microorganisms ml – 1 of a urinary tract pathogen for mid-stream clean-void urine and for cathetherization, respectively. Blood samples were collected for the measurement of WBC count and CRP levels and stored at – 701C for analysis of PCT levels. All patients were given ceftriaxone, 75 mg
kg – 1
day – 1, by the parenteral route. Therapy was modified according to bacterial susceptibility tests and clinical pattern at the third day of treatment, which was completed to 14 days. After eradication of bacteria, all patients underwent suppression with trimethoprim–sulfamethoxazole, 2 mg
kg – 1
day – 1, for at least 6 months.

posterior oblique images were acquired in a 256  256 matrix for 10 min per image. The severity of renal lesions was determined according to the extent and the intensity of the lesions seen on two different views by two nuclear medicine physicians (F.G. and F.A.) who were unaware of the clinical status and laboratory parameters of the patients. Two physicians scored the DMSA scans without any knowledge of the other’s results. The scores for the extent of lesions were: 0, absence of lesion; 1, defect covering < 5% surface area; 2, defect covering 5–10% surface area; 3, defect covering 10–30% surface area; 4, defect covering > 30% surface area. The scores for defect intensity were: 0, absence of lesion; 1, mild hypoactivity without loss of cortical margins; 2, moderate hypoactivity with partial loss of cortical margins; 3, severe hypoactivity with loss of cortical margins. If multiple defects were present, the highest intensity score was accepted as the intensity score. Summed scores (score for extent of lesion plus score for defect intensity) were obtained for each patient and termed the DMSA inflammation score (DMSA-IS) (Fig. 1). According to DMSA-IS, the patients were divided into four groups. Group 1 included patients with normal DMSA scans, group 2 included patients with a DMSA-IS of 2–3, group 3 included patients with a DMSA-IS of 4–5 and group 4 included patients with DMSA-IS of 6–7. In groups 2, 3 and 4, repeat DMSA scans were obtained at the third month. If pathology was still present, another follow-up scan was performed at the sixth month. The study was approved by the local ethics committee. Data are presented as the mean ± 1 standard deviation (mean ± SD). The comparison between the mean values Fig. 1

WBC counts were obtained using an LH 750 Beckham Coulter complete blood counter. Serum CRP levels were measured by an immunoturbidometric method using a Roche modular P autoanalyser. Levels above 0.5 mg
dl – 1 were considered high. PCT levels were measured by an immunoluminometric method using a LUMI test PCT kit (Brahms Diagnostica, Berlin, Germany) following the manufacturer’s instructions. Levels above 0.5 ng
ml – 1 were considered elevated. Within 72 h of admission, all patients underwent renal– pelvic ultrasonography and DMSA scintigraphy. 99mTcDMSA scans were performed 3–4 h after intravenous injection of 1.5 MBq
kg – 1 of activity (minimum 20, maximum 100 MBq). A Toshiba GCA-501 S gamma camera equipped with a pinhole collimator was used for imaging. Right and left posterior, right posterior oblique and left

(a) Posterior view, left kidney in a 4-year-old girl. There is hypoactivity at the upper pole of the left kidney (arrow) in the acute phase of the disease. The DMSA inflammation score (DMSA-IS) was evaluated as 3 (extent of lesion score, 1; intensity score, 2; summed score, 3). (b) Posterior view, left kidney in a 3.5-year-old girl. There are multiple defects on the left kidney in the acute phase of the disease. DMSA-IS was evaluated as 7 (extent of lesion score, 4; intensity score, 3; summed score, 7). DMSA, dimercaptosuccinic acid.

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Procalcitonin in acute pyelonephritis Gu¨ven et al. 717

was performed by using the Mann–Whitney U test. The chi-squared test was used to compare frequencies. Pearson’s correlation test was used to compare between DMSA-IS and PCT levels. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of CRP, PCT and WBC counts of the patients were calculated. Receiver operating characteristic curve (ROC) analysis was performed to define the cut-off value, and the sensitivity and specificity of WBC, CRP and PCT measurements. A P value of less than 0.05 was considered as statistically significant. The commercial statistical software package used was SPSS/10.0.

6-month scans, five of 21 patients (23.8%) had renal scars (Fig. 3). The time between the beginning of symptoms and the start of therapy was 1–10 days (mean 2.2 days). The mean time between the onset of symptoms and performing the DMSA scans was 4.27 days (range 1–12 days). Eleven of 16 patients and 10 of 17 patients whose scans were obtained within 3 days and beyond 3 days after the symptoms began had positive DMSA scans, respectively (P = 0.721). Using the chi-squared test, no correlation was found between clinical parameters, alone or combined with each other, and DMSA uptake (Table 1).

Results Thirty-three children (31 female (94%) and two male) aged 1–11 years (mean 4.42 years) were enrolled in this prospective study. The demographic and clinical findings of the patients are outlined in Table 1. Eleven of 33 patients had no significant bacteriuria in urine cultures. Eight of these 11 children had sterile urine cultures, five of whom had a history of antibiotic use before admission.

Demographic and clinical data of 33 patients with relation to DMSA scintigraphy findings

Table 1

All patients

Gender Male Female Age 12–24 months 2–5 years 6–10 years > 10 years Fever (1C) < 38.5 38.5–38.9 > 39 Abdominal pain Yes No Flank pain Yes No Vomiting Yes No Urine culture Sterile < 100 000 CFU* Z 100 000 CFU

Twenty-one of 33 patients (64%) each had a positive DMSA scan, of which 19 showed unilateral and two showed bilateral involvement. Of the 21 patients who had positive DMSA scans, two had both negative urine cultures and had no previous antibiotic use. There were discordances in one and two of the 21 patients with pathological DMSA scans in terms of determining the intensity score and the score for the extent of the lesion, respectively, between the two nuclear medicine physicians. In these cases, consensus was reached and these cases were scored after discussion between the two physicians. According to the DMSA-IS, seven patients were in group 2, seven in group 3 and another seven were in group 4 (Table 2). On 19 follow-up scans at the third month (scans of two patients could not be obtained), 13 (68%) showed complete resolution (Fig. 2). On the Table 2

Patients with positive DMSA scan

n

%

n

%

2 31

6.0 94.0

1 20

50.0 64.5

6 14 12 1

18.1 42.4 36.3 3.0

5 9 7 0

83.3 64.2 58.3 –

3 7 23

9.0 21.2 69.6

1 7 13

33.3 100 56.5

15 12

55.5 44.5

9 7

60.0 58.3

5 22

18.5 81.5

2 14

40.0 63.6

11 22

33.3 66.6

8 13

72.7 59.0

8 3 22

24.2 9.0 66.6

5 1 15

62.5 33.3 68.1

*

All cultures were collected by mid-stream technique.

White blood cell count, and C-reactive protein and procalcitonin levels of the groups Group 1 n = 12

White blood cell count (cells
mm – 3) < 10 000 10 000–15 000 > 15 000 C-reactive protein (mg
dl – 1) < 2.0 2.0–5.0 5.1–10.0 > 10.0 Procalcitonin (ng
ml – 1) < 0.5 0.5–1.0 1.1–2.0 > 2.0 Mean ± SD of procalcitonin

Group 2 n = 7

Group 3 n = 7

Group 4 n = 7

Total n

%

1 4 7

0 2 5

0 3 4

0 2 5

1 11 21

3.0 33.3 63.6

2 2 4 4

0 1 1 5

1 2 3 1

0 1 1 5

3 6 9 15

9.1 18.2 27.3 45.4

5 4 3 0

2 2 1 2

3 1 3 0

3 0 1 3

13 7 8 5

39.3 21.2 24.2 15.3

0.77 ± 0.64

1.08 ± 0.84

0.90 ± 0.71

1.72 ± 1.71

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718 Nuclear Medicine Communications 2006, Vol 27 No 9

WBC counts were between 6900 and 29 100 mm – 3 and serum CRP levels ranged between 1.5 and 33.1 mg
dl – 1. No patient had a normal CRP level. Thirteen patients had normal PCT levels ( < 0.5 ng
ml – 1), of whom eight had positive DMSA scans. However, all five patients who had PCT levels > 2 ng
ml – 1 had positive DMSA scans. WBC counts, and serum CRP and PCT levels in relation to DMSA scintigraphy findings are outlined in Table 2. No statistically significant differences were found between each of the groups and the PCT levels. Moreover, these parameters were compared between patients with normal and positive DMSA scans (Table 3). All six patients who had CRP levels above 10 mg
dl – 1, WBC counts higher than 15 000 mm – 3 and who had PCT levels above 1 ng
ml – 1 had positive DMSA scans. The number of true positive and negative patients according to the different CRP and PCT cut-off values are shown in Fig. 4.

curve for sensitivity and specificity of PCT measurements is shown in Fig. 5. Sensitivity, specificity, PPV, NPV and the accuracy of CRP and PCT values of the patients were calculated for different cut-off values (Table 5). Sensitivity and specificity of CRP with a cutoff value of 2 mg
dl – 1 were both 66.6%, whereas sensitivity and specificity of PCT with a cut-off value of 0.5 ng
ml – 1 were 38.4% and 65%, respectively. However, if the cut-off value for PCT was chosen as 2 ng
ml – 1, the sensitivity increased to 100% while specificity did not change markedly.

Fig. 3

ROC analysis was performed for WBC, CRP and PCT. Cut-off values and area-under-the-curve values of these laboratory findings are outlined in Table 4. The ROC

Fig. 2

Posterior view, left kidney in a 9-year-old girl. Clear hypoactivity is seen (arrow) at the upper pole of the left kidney in the acute phase of the disease. (a) DMSA-IS was evaluated as 6 (extent of lesion score, 3; intensity score, 3; summed score, 6). (b) Normalization of scan appearances on the follow-up scan 3 months later.

Table 3

Posterior view, right kidney in a 5-year-old girl. (a) Acute pyelonephritis of the upper pole of the right kidney (arrow). The DMSA-IS was 6 (extent of lesion score, 3; intensity score, 3; summed score, 6). (b) Although the defect is partially improved, resulting in residual permanent damage at 3 months (arrow), the DMSA-IS was 4 (extent of lesion score, 2; intensity score, 2; summed score, 4). (c) At the 6-month (arrow) follow-up DMSA scan, the DMSA-IS was 2 (extent of lesion score, 1; intensity score, 1; summed score, 2).

White blood cell count, and C-reactive protein and procalcitonin levels in children with normal and positive DMSA scans

White blood cell count (mm C-reactive protein (mg
dl – 1) Procalcitonin (ng
ml – 1)

–3

)

Normal DMSA (n = 12)

Positive DMSA (n = 21)

P*

P**

6900–29 100 (17458 ± 6472) 1.5–26.0 (8.69 ± 7.18) 0.06–1.98 (0.766 ± 0.644)

11 900–26 300 (17933 ± 4055) 1.7–33.1 (11.8 ± 8.17) 0.19–5.04 (1.232 ± 1.172)

0.782 0.291 0.228

0.779 0.166 0.217

DMSA, dimercaptosuccinic acid. * Mann–Whitney U test. ** Multi-variant analysis.

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Procalcitonin in acute pyelonephritis Gu¨ven et al. 719

Fig. 4

Table 5 Sensitivity, specificity, PPV, NPV and accuracy of C-reactive protein and procalcitonin values for the prediction of acute pyelonephritis

DMSA (+)

C-reactive protein (mg
dl – 1)

DMSA (-)

Procalcitonin (ng
ml – 1)

20

20 18 16 14 12 10 8 6 4 2 0

16

Sensitivity (%) Specificity (%) PPV (%) NPV (%) Accuracy (%)

13 11

10

10

8

7 5

4

3 0

CRP > 2

CRP > 5

CRP >10

PCT >0.5

PCT >1

Table 4 Results of receiver operating characteristic (ROC) analysis of laboratory parameters Parameter

Cut-off value

White blood cell count C-reactive protein Procalcitonin

12 300 mm – 3

95.5

27.3

0.54

2.8 mg
dl – 1 0.9605 ng
ml – 1

86.4 50.0

36.4 81.8

0.60 0.67

Area under the ROC curve

Fig. 5

PCT 100

Z 10.0

Z 0.5

Z 1.0

Z 2.0

66.6 66.6 95.2 16.6 66.6

66.6 44.4 76.1 33.3 60.6

73.3 44.4 52.3 66.6 57.5

65.0 38.4 61.9 41.6 54.5

76.9 45.0 47.6 75.0 57.5

100 42.8 23.8 100 51.5

Cut-off values of 2, 5 or 10 mg
dl – 1 for C-reactive protein and 0.5, 1 or 2 ng
ml – 1 for procalcitonin. PPV, positive predictive value; NPV, negative predictive value.

Discussion The aim of our study was to evaluate the usefulness of the PCT test in the differentiation of renal parenchymal involvement, and to re-evaluate the effectiveness of the traditionally used clinical and laboratory parameters in the diagnosis of APN in children with a first febrile UTI. In our study, 20 of 30 patients (66%) with fever > 38.51C had positive DMSA scans. This finding is in accordance with the studies of Benador et al. [10] who demonstrated that only 71% of children aged 1 month to 16 years with febrile UTI had positive DMSA scan, and 64 of 87 (74%) children who had first episode of febrile UTI had acute parenchymal changes [12]. Majd et al. [13] showed that among 94 patients with febrile UTIs, 78% of the patients with fever > 39.41C and 55% of the patients with < 39.41C had scan evidence of APN, respectively. Biggi et al. [14] did not find a difference in maximum temperature levels between patients with positive and negative DMSA scans. These data show that high fever is not an adequate sign to distinguish renal parenchymal involvement. Our results and some other studies confirmed that blood leukocyte count do not always correlate with the severity of UTI [8,9,11]. However, Benador et al. [10] found a significant difference in WBC counts between pyelonephritic patients and patients with lower UTIs.

80

Sensitivity

Z 5.0

PCT >2

The number of true positive and negative patients according to the different cut-off values for C-reactive protein (CRP) and procalcitonin (PCT). (’), DMSA( + ); (&), DMSA( – ).

Sensitivity Specificity (%) (%)

Z 2.0

60

40

20

0 0

20

40

60

80

100

100-Specificity Receiver operating characteristic curve for sensitivity and specificity of procalcitonin (PCT) measurements.

In the present study, eight of 33 patients had sterile urine cultures, five of whom had a history of antibiotic use before admission. Two of three patients with sterile urine cultures and without previous antibiotic use had already positive DMSA scans. The reason for negative urine cultures might be an infection with unspecified microorganisms that could not be detected by routine microbiological laboratory techniques. In the remaining patient DMSA scintigraphy had been performed within 24 h after the symptoms began, which could be an explanation for the negative scan. Levtchenko et al. [15] also reported patients with positive DMSA scans and negative urine cultures. In this report, the authors suggested that a 99mTc-DMSA scan should be performed in the case of severe infection without a clear aetiology even with negative urine culture. Our study also

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720 Nuclear Medicine Communications 2006, Vol 27 No 9

supported this suggestion due to the positive DMSA scan findings with negative urine culture. These observations show how difficult it is for the clinician to diagnose APN accurately. In our study group, 30 out of 33 children had a serum CRP value greater than 2 mg
dl – 1. Neither correlation nor any difference was found between serum CRP levels and positive DMSA scans. Biggi et al. [14] reported an accuracy rate (65%) similiar to our study, whereas specificity and sensitivity rates were 68% and 64%, respectively. They also noticed that the CRP level and WBC count were able to distinguish children with severe renal parenchymal involvement from those with mild or moderate disease. In most of the recent reports there was an overlap of patients who had normal DMSA scans and who had elevated CRP values. Interestingly, when a combination of these parameters was used, correlation strength did not increase. Biggi et al. found that the accuracy of the diagnosis of APN did not increase – in fact, it decreased – when the parameters of WBC and CRP were combined instead of using each parameter alone, and 65 of 101 children received false negative diagnoses [14]. In summary, although CRP has been found to be significantly elevated in febrile UTIs, the serum CRP level, alone or in combination with an elevated WBC count, is not a good predictor of renal parenchymal involvement. In normal population of blood donors, PCT was frequently found below 0.1 ng
ml – 1 [7]. Prat et al. [8] established the range of normal PCT values in 38 healthy children without infection and showed a median PCT value of 0.363 ± 0.29 ng
ml – 1. During severe bacterial infections circulating blood PCT levels may reach 20– 2000 ng
ml – 1. In a case report, it was documented that PCT increased to a peak level around 12 h after the beginning of infection and stayed at abnormal levels for 10 days with a half-life of 22.5 h [16]. Currently, the procalcitonin test is becoming a popular marker for the diagnosis of APN and for the prediction of renal scar formation. Although a good correlation between high PCT values and APN has been reported in almost all published studies, a non-negligible number of children with positive DMSA scans had PCT levels below the cutoff values [2,8,9,11]. In our study, serum PCT levels did not show any significant difference between each group (Table 2) and overlap of the patients was prominent (Fig. 1). However, patients with markedly elevated PCT values ( > 2 mg
dl – 1) had positive DMSA scans, with a 100% sensitivity and 100% negative predictive value. In the study by Smolkin et al. [2], serum PCT levels were very variable (0.36–12.4 mg
l – 1; median 3.41) in children with APN confirmed by a DMSA scan. The sensitivity and specificity of PCT were 94.1% and 89.7%, respectively, when the PCT cut-off value was taken as 0.5 mg
l – 1 in their study group. These ratios were found to be 65%

and 38.4%, respectively, in the present study. Prat and colleagues [8] claimed that a low PCT value at the time of admission, in spite of clinical signs of APN, points to a low risk of renal scarring. In the recently published study by Pecile et al. [11], the mean PCT level was found to be significantly higher in the pyelonephritic group. Their sensitivity and specificity for prediction of APN were also higher than our results (90.7% and 70.2% with the cut-off value of 0.5 ng
ml – 1, respectively), although there were DMSA positive children with PCT levels below their cutoff value of 0.8 ng
ml – 1. In their study, the cut-off point established by ROC was 0.8 ng
ml – 1 and the area under the ROC curve was found to be 0.924. In our study, the cut-off point established by ROC was 0.96 ng
ml – 1 and the area under the ROC curve was found to be 0.661. The small area under the curve in the present study may be due to the limited number of participants in our study group. Gervaix et al. [9] similarly demonstrated that 26% of children with pyelonephritis had PCT levels below the limit of detection of 0.5 ng
ml – 1, whereas 15% of their patients with lower UTI had > 0.5 ng
ml – 1 levels of PCT. In this study, the probability of pyelonephritis increased from 34 to 92% when a cut-off value of 2.0 ng
ml – 1 was used instead of 0.5 ng
ml – 1. Benador et al. [10] compared the mean PCT levels (0.38 ± 0.19 SE) of 23 children with lower UTI with the levels (5.37 ± 1.9 SE) of 37 children with definite APN at admission (P < 0.0001). In this study, 25.6% of patients with no or grade 1 to 2 DMSA lesions and all patients with grade 3 and 4 scintigraphic lesions had PCT levels > 0.6. These findings indicate that the higher the PCT values the better is the predictive value of APN, very high PCT levels are able to identify children with severe renal involvement, and normal or low PCT values may be associated with very mild or mild lesions on DMSA scan. Markedly elevated PCT levels in children with febrile UTIs may influence the decision concerning hospitalization, the route and duration of anti-microbial therapy, and may alert the physician to rapidly investigate risk factors. On the other hand, bearing in mind that DMSA scintigraphy is more reliable, low PCT levels may protect children from time-consuming and partly invasive tests. In our study, 63% of children had parenchymal involvement verified with a DMSA scan. This result is in accord with three other studies in which 63, 67 and 64% of children with symptomatic febrile UTI had renal involvement on DMSA [9,12,17]. There are also lower ratios in the literature [18]. In the present study, 13 out of 21 children who had positive DMSA scans on admission showed complete resolution on the scans at the third month. Five of the remaining patients had scars on the 6-month DMSA scans; PCT levels of these patients ranged between 0.327 and 5.037 ng
ml – 1 (mean 1.907, median 1.017). These data show that high PCT levels may not be associated with scar formation.

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Procalcitonin in acute pyelonephritis Gu¨ven et al. 721

It was observed that the risk factors such as late admission (in one patient), breakthrough UTI (in one patient), voiding dysfunction (in three patients), urolithiasis (in one patient) and vesicoureteral reflux (in three patients) were similar in children with and without renal parenchymal involvement on consecutive DMSA scans. However, all children who had renal scarring were younger than 5 years (1–4.5 years) whereas nearly one third of the patients without renal scars were older than 5 years. The limited number of patients and the culture-negative patients may be the limitations of our study. Although febrile UTIs and renal parenchymal involvement are more common in children younger than 1 year, we preferred to exclude this age group because (1) the difficulty of obtaining consent for small infants for the invasive investigations required according to the protocol; (2) the epidemiology and risk factors of UTI in this age group have some differences compared to those in older children; and (3) it is known that serum PCT levels do not change with age [16]. So, there is no obstacle to making comparisons with study groups that included children younger than 1 year. In clinical practice, a number of patients who commenced antibiotic therapy at one hospital are referred to another (tertiary) hospital and the antibiotics may cause negative urine cultures. It was stated that high PCT levels induced by bacterial infections were not changed by administration of antibiotics [16]. It is generally difficult to precede renal parenchymal involvement in APN. Urine cultures and DMSA scan results may not be in accord with each other in situations such as previous usage of antibiotics, infection with unspecified microorganisms and possibly with DMSA scans performed during the very early stages of the infection. It is unclear how early and to what extent renal parenchymal involvement could be detected on DMSA scan. Because culture-negative patients are also under the responsibility of nephrologists, this group was not excluded in the present study. With all the data, we can conclude that the advantages of very high levels of PCT would be to estimate the severity of renal involvement from the very beginning of UTI. If further studies could be performed with adequate numbers of patients in each range of PCT levels, allowing cut-off values to be determined which could be used to define true positive cases, measurement of serum PCT levels would be useful for predicting renal parenchymal involvement in children with febrile UTIs.

Acknowledgements The authors acknowledge the contributions of the Turkish Nephrology Association for the financial support, of Akdeniz University Research Unit, and of Dr Faysal Go¨k and Dr Ugur Musabak for providing the measurements of procalcitonin.

References 1

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16 17 18

American Academy of Pediatrics, Committee on Quality Improvement. Subcommittee on Urinary Tract Infection. Practice parameter: The diagnosis, treatment, and evaluation of the initial urinary tract infection in febrile infants and young children. Pediatrics 1999; 103:843–852. Smolkin V, Koren A, Raz R, Colodner R, Sakran W, Halevy R. Procalcitonin as a marker of acute pyelonephritis in infants and children. Pediatr Nephrol 2002; 17:409–412. Majd M, Rushton HG. Renal cortical scintigraphy in the diagnosis of acute pyelonephritis. Semin Nucl Med 1992; 22:98–111. Rushton HG. The evaluation of acute pyelonephritis and renal scarring with technetium 99m-dimercaptosuccinic acid renal scintigraphy: evolving concepts and future directions. Pediatr Nephrol 1997; 11:108–120. Glauser MP, Lyons JM, Braude AI. Prevention of chronic experimental pyelonephritis by suppression of acute suppuration. J Clin Invest 1978; 61:403–407. Hiraoka M, Hashimoto G, Tsuchida S, Tsukahara H, Ohshima Y, Mayumi M. Early teatment of urinary infection prevents renal damage on cortical scintigraphy. Pediatr Nephrol 2003; 18:115–118. Gendrel D, Bohuon C. Procalcitonin as a marker of bacterial infection. Pediatr Infect Dis J 2000; 19:679–688. Prat C, Dominguez J, Rodrigo C, Gimenez M, Azuara M, Jimenez O, et al. Elevated serum procalcitonin values correlate with renal scarring in children with urinary tract infection. Pediatr Infect Dis J 2003; 22:438–442. Gervaix A, Galetto-Lacour A, Gueron T, Vadas L, Zamora S, Suter S, et al. Usefulness of procalcitonin and C-reactive protein rapid tests for the management of children with urinary tract infection. Pediatr Infect Dis J 2001; 20:507–511. Benador N, Siegrist CA, Gendrel D, Greder C, Benador D, Assicot M, et al. Procalcitonin is a marker of severity of renal lesions in pyelonephritis. Pediatrics 1998; 102:1422–1426. Pecile P, Miorin E, Romanello C, Falleti E, Valent F, Giacomuzzi F, et al. Procalcitonin: A marker of severity of acute pyelonephritis among children. Pediatrics 2004; 114:e249–e254. Benador D, Benador N, Slosman D, Nussle D, Mermillod B, Girardin E. Cortical scintigraphy in the evaluation of renal parenchymal changes in children with pyelonephritis. J Pediatr 1994; 124:17–20. Majd M, Rushton G, Jantausch B, Wiedermann B. Relationship among vesicoureteral reflux, P-fimbriated Escherichia coli, and acute pyelonephritis in children with febrile urinary tract infection. J Pediatr 1991; 119:578–585. Biggi A, Dardanelli R, Pomero G, Cussino P, Noello C, Sernia O, et al. Acute renal cortical scintigraphy in children with a first urinary tract infection. Pediatr Nephrol 2001; 16:733–738. Levtchenko EN, Lahy C, Levy J, Ham HR, Piepsz A. Role of Tc-99m DMSA scintigraphy in the diagnosis of culture negative pyelonephritis. Pediatr Nephrol 2001; 16:503–506. Brunkhorst FM, Heinz U, Forycki ZF. Kinetics of procalcitonin in iatrogenic sepsis. Intensive Care Med 1998; 24:888–889. Ilyas M, Mastin ST, Richard GA. Age-related radiological imaging in children with acute pyelonephritis. Pediatr Nephrol 2002; 17:30–34. Rosenberg AR, Rossleigh MA, Brydon MP, Bass SJ, Leighton DM, Farnsworth RH. Evaluation of acute urinary tract infection in children by dimercaptosuccinic acid scintigraphy: a prospective study. J Urol 1992; 148:1746–1749.

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Original article

Quantitative analysis of dopamine synthesis in human brain using positron emission tomography with L-[b-11C]DOPA Hiroshi Itoa, Miho Otaa, Yoko Ikomaa,b, Chie Sekia,b, Fumihiko Yasunoa, Akihiro Takanoa, Jun Maedaa, Ryuji Nakaoc, Kazutoshi Suzukic and Tetsuya Suharaa Background and objectives To estimate the presynaptic function of the central dopaminergic system, the rate of endogenous dopamine synthesis has been measured by using L-[b-11C]DOPA or 6-[18F]fluoro-L-DOPA with positron emission tomography. However, the regional kinetics of 11 L-[b- C]DOPA in human brain have not been investigated in detail. In the present study, the regional kinetics of 11 L-[b- C]DOPA in normal human brain and the accuracy of the method for quantifying L-[b-11C]DOPA kinetics, employing reference regions, were investigated. Methods After intravenous injection of L-[b-11C]DOPA, dynamic scanning was performed on ten healthy subjects for 89 min. The overall uptake rate constant K was calculated by the kinetic and graphical approaches, in which the occipital cortex was used as a reference brain region. Results Regional distribution of K was similar to those of dopamine D2 receptor. A significant negative correlation was observed between the neutral amino acid concentration in plasma and the influx rate constant through the blood–brain barrier (K1). The K values calculated by graphical approach were in good agreement with the values calculated by kinetic approach for both experimental and simulated data.

Introduction The central dopaminergic system is of interest in the pathophysiology of neuropsychiatric diseases; for example, schizophrenia and neurodegenerative diseases such as Parkinson’s disease. One way to estimate the presynaptic function of the central dopaminergic system is to measure the rate of endogenous dopamine synthesis. Endogenous dopamine is produced by enzymatic decarboxylation of L-3,4-dihydroxyphenylalanine (L-DOPA) under aromatic L-amino acid decarboxylase. Thus, 11Clabelled L-DOPA (L-[b-11C]DOPA) and 18F-labelled LDOPA (6-[18F]fluoro-L-DOPA) can be used as positron emission tomography (PET) tracers to assess the rate of dopamine synthesis, which indicates the relative activity of cerebral L-DOPA decarboxylase [1–6]. Several investigators have reported on striatal dopamine synthesis in schizophrenia using L-[b-11C]DOPA and 6-[18F]fluoro-L-

Conclusions The regional distribution of K corresponds to that of the nigrostriatal and mesolimbic dopaminergic system. Negative correlation between neutral amino acid concentration and K1 supports the suggestion that L-DOPA is transported in a competitive fashion via the same carrier system as neutral amino acids at the blood–brain barrier. Because the graphical approach can obviate the need for an arterial input function, it is useful for investigating the rate of regional dopamine synthesis in neuropsychiatric and neurodegenerative diseases. Nucl c 2006 Lippincott Williams & Med Commun 27:723–731
Wilkins. Nuclear Medicine Communications 2006, 27:723–731 Keywords: brain, DOPA, dopamine, PET a Departments of Molecular Neuroimaging, bBiophysics and cRadiochemistry, Molecular Imaging Center, National Institute of Radiological Sciences, Chiba, Japan.

Correspondence to Dr Hiroshi Ito, Clinical Neuroimaging Section, Department of Molecular Neuroimaging, Molecular Imaging Center, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555, Japan. Tel: + 81 43 206 4702; fax: + 81 43 253 0396 e-mail: [email protected]

Received 19 April 2006 Accepted 31 May 2006

DOPA with PET. Most showed an increase in the rate of dopamine synthesis [7–10], although an unaltered rate of dopamine synthesis has also been reported [11]. For quantitative analysis of L-[b-11C]DOPA kinetics in the brain, a graphical method developed by Gjedde [1] and Patlak and Blasberg [12] has been widely applied. By this method, a reference brain region with no irreversible binding is employed to preclude the need for an arterial input function. However, regional kinetics of L[b-11C]DOPA in human brain have not been investigated in detail, and therefore, the accuracy of quantitative analysis based on a reference region has also not been evaluated. In the present study, we investigated the regional kinetics of L-[b-11C]DOPA in normal human brain with consideration of L-[b-11C]DOPA transport by the carrier system at the blood–brain barrier. The

c 2006 Lippincott Williams & Wilkins 0143-3636


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724 Nuclear Medicine Communications 2006, Vol 27 No 9

accuracy of the method for quantifying L-[b-11C]DOPA kinetics employing reference regions was also evaluated by using both experimental and simulated data.

Materials and methods Subjects

The study was approved by the Ethics and Radiation Safety Committees of the National Institute of Radiological Sciences, Chiba, Japan. Ten healthy men (21–32 years of age, 23.4 ± 3.3 years (mean ± SD)) were recruited and written informed consent was obtained. The subjects were free of somatic, neurological or psychiatric disorders on the basis of their medical history and magnetic resonance imaging of the brain. They had no history of current or previous drug abuse. Procedures for positron emission tomography

All PET studies were performed with a Siemens ECAT Exact HR + system, which provides 63 sections with an axial field of view of 15.5 cm [13]. The intrinsic spatial resolution was 4.3 mm in-plane and 4.2 mm full-width at half maximum (FWHM) axially. With a Hanning filter (cut-off frequency 0.4 cycle/pixel), the reconstructed inplane resolution was 7.5 mm FWHM. Data were acquired in the three-dimensional mode. Scatter was corrected [14]. A head fixation device with thermoplastic attachments for individual fit minimized head movement during PET measurements. C]DOPA was synthesized from [11C]carbon dioxide via D,L-[3-11C]alanine as described previously [15,16]. The specific radioactivity of L-[b-11C]DOPA was 29–82 GBqmmol – 1 at the time of injection. The radioactivity injected was 320–402 MBq.

L-[b-

11

After intravenous rapid bolus injection of L-[b-11C]DOPA, data were acquired for 89 min in a consecutive series of time frames. The frame sequence consisted of seven 1min frames, five 2-min frames, four 3-min frames, and twelve 5-min. A 10-min transmission scan using a 68 Ge–68Ga line source was performed for correction of attenuation. To obtain the arterial input function, an automated blood sampling system was used during the first 12 min of each measurement [17]. Thereafter, 28 samples of arterial blood were taken 15, 25, 35, 45, 55, 65, 75, 85, 95 and 105 s, and 2, 2.5, 3, 4, 5, 6, 8, 10, 12, 15, 20, 30, 40, 50, 60, 70, 80 and 90 min after injection. The time difference between the appearance of radioactivity in the brain and automated blood sampling was experimentally defined and taken into account in the calculation. The fraction of radioactivity representing unchanged L-[b-11C]DOPA in plasma was determined by high-performance liquid chromatography (HPLC) [18,19]. For HPLC analysis,

arterial blood samples were drawn at 3.5, 18, 32, 46, 59, 75 and 89 min after injection. The 6% perchloric acid was added to each plasma sample, and samples were then centrifuged. The obtained supernatant was subjected to radio-HPLC analysis (column, Finepak SIL C18T, Jasco, Tokyo, Japan; mobile phase, 9:91 acetonitrile/ sodium phosphate buffer (100 mmoll – 1, pH 2.0) and 5 mmoll – 1 sodium octanesulfonate). Plasma protein binding was not determined in the present study. Two arterial blood samples were taken, one at the beginning and one at the end of scanning, for measurement of neutral amino acid concentration in plasma. Neutral amino acid concentrations of two samples were averaged and used for analyses. All magnetic resonance imaging studies were performed with a 1.5-T magnetic resonance scanner (Philips Medical Systems, Best, the Netherlands). Three-dimensional volumetric acquisition of a T1-weighted gradient echo sequence produced a gapless series of thin transverse sections (TE: 9.2 ms; TR: 21 ms; flip angle: 301; field of view: 256 mm; acquisition matrix: 256  256; slice thickness: 1 mm). Regions of interest

All magnetic resonance images were co-registered to the PET images with the statistical parametric mapping (SPM2) system [20]. Regions of interest (ROIs) were drawn on co-registered magnetic resonance images and transferred to the PET images. ROIs were defined for the cerebellar cortex, midbrain, thalamus, caudate head, putamen, parahippocampal gyrus, anterior part of the cingulate gyrus, frontal cortex, temporal cortex, parietal cortex and occipital cortex. Each ROI was drawn in three adjacent sections and data were pooled to obtain the average radioactivity concentration for the whole volume of interest. To obtain regional time–activity curves, regional radioactivity was calculated for each frame, corrected for decay, and plotted versus time. Quantification of L-[b-11C]DOPA kinetics

To describe the kinetics of L-[b-11C]DOPA in the brain, the standard three-compartment model with three firstorder rate constants for tracers showing irreversible binding was employed [5,21]. Three compartments are defined. Cp is the radioactivity concentration of unchanged radiotracer in plasma (arterial input function); Ce is the radioactivity concentration of unchanged radiotracer in brain tissue; Cm is the radioactivity concentration of 11C-labelled dopamine and its metabolites in brain tissue. The rate constants K1 and k2 describe the influx and efflux rates for radiotracer diffusion through the blood–brain barrier, respectively. The rate constant k3 describes utilization of L-[b-11C]DOPA including dopamine synthesis rate that indicates the relative activity of cerebral L-DOPA decarboxylase. This model can be

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Quantification of

described by the following equations:

11

C-DOPA in human brain Ito et al. 725

for t > t*, and

dCe ðt Þ ¼ K1 Cp ðt Þ  ðk2 þ k3 ÞCe ðt Þ; dt

kref ¼

ð1Þ

K Ve0 þ VP0

ð7Þ

0

dCm ðt Þ ¼ k3 Ce ðt Þ; dt

ð2Þ

C i ð t Þ ¼ C e ð t Þ þ Cm ð t Þ

ð3Þ

where Ci is the total radioactivity concentration in a brain region, which can be measured by PET. The overall uptake rate constant, K, for L-[b-11C]DOPA, which indicates the net rate of dopamine synthesis, is expressed as follows [12,22]: K1 k3 K¼ k2 þ k3

ð4Þ

For irreversible tracers, the steady-state volume of reversible compartment in a brain region, Ve, is defined as follows: Ve ¼

K1 k2 þ k3

ð5Þ

where Ci is the total radioactivity concentration in a brain 0 region with no irreversible binding; V e is the steady-state volume of reversible compartment in a brain region with 0 no irreversible binding; V p is the effective plasma volume of brain tissue in a brain region with no irreversible binding; t* is the equilibrium time of the compartment for unchanged radiotracer in brain tissue. Plotting 0 Ci(t)/Ci (t) versus Rt 0 Ci ðtÞdt 0

Ci0 ðt Þ after time t*, yields a straight line with the slope kref and intercept F. In the present study, the occipital cortex was used as a reference region with no irreversible binding, because this region is known to have the lowest dopamine concentration [24] and lowest aromatic L-amino acid decarboxylase activity [25]. The ranges of equilibrium time, t*, of 29–64 min and 29–89 min were used for experimental data, and 29–64 min, 29–89 min and 64– 89 min were used for simulated data. Simulation study

Kinetic method

The three-compartment model was used in an initial attempt to describe the regional time–activity curves for 11 L-[b- C]DOPA. The traditional strategy is to estimate the rate constants (K1, k2, and k3) by non-linear curve fitting in a least squares sense to the regional time– activity curves [23]. The model equations (Equations 1 and 2) were combined and solved in a convolution integral procedure, and then used for non-linear curve fitting analysis. In this analysis, blood volume (Vb), which depends on the first-pass extraction fraction of tracer, was also estimated using the radioactivity of whole blood in order to diminish the influence of tracer remaining in the blood. The radioactivity of unchanged L-[b-11C]DOPA in plasma was used as the arterial input function. For this analysis, the software package, PMOD (PMOD Technologies, Zurich, Switzerland) was used. Graphical method

The overall uptake rate constant K was also calculated using the graphical analysis [1,12]. This analysis has been developed for irreversible tracers and allows for the calculation of K using time–activity data in a reference brain region with no irreversible binding. K values can be estimated by using simple linear least squares fitting as follows: Rt C i ðt Þ ¼ kref  0 Ci ð t Þ

Ci0 ðtÞdt

0

Ci0 ðt Þ

þF

ð6Þ

To estimate errors in the calculation of kref with the graphical method, simulation studies were performed. Time–activity curves in brain regions were generated based on the three-compartment model using assumed values for the rate constants K1, k2 and k3. The K1, Ve and Vb values were assumed to be 0.03 mlml – 1min – 1, 0.7 mlml – 1, and 0.05 mlml – 1, respectively. A series of tissue time–activity curves were generated by varying k3 from 0.008 to 0.040 min – 1 in nine steps. These values were taken from the results obtained with the kinetic method. The average arterial input function for all subjects was used for generating the tissue time–activity curves. The tissue time–activity curve with k3 of 0.008 was considered as a reference brain region with little irreversible binding. The overall uptake rate constant kref was calculated by the graphical method for these generated tissue time–activity curves, and was compared with the assumed K values. Measurement of neutral amino acid concentration in plasma

In all subjects, natural neutral amino acid (NAA) concentration in plasma was measured by HPLC (L8500 Amino Acid Analyzer System, Hitachi Corp., Tokyo, Japan). The amino acids are phenylalanine, tryptophan, leucine, methionine, isoleucine, tyrosine, histidine, valine and threonine, which are transported via the same carrier at the blood–brain barrier, termed the L-system [26]. LDOPA is also transported from blood to brain by the same carrier system as NAAs [27], although the affinity for the carrier system of these nine NAAs and L-DOPA is

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726 Nuclear Medicine Communications 2006, Vol 27 No 9

different. When the affinity ratio of L-DOPA to the nine NAAs is defined as Km(i)/Km, where Km is the halfsaturation constant for L-DOPA and Km(i) is the halfsaturation constant for each of the NAAs, the concentration C(i) of each NAA divided by Km(i)/Km is the concentration corresponding to L-DOPA for the carrier system of NAAs. The sum of C(i)/(Km(i)/Km) of the nine NAAs, a weighted sum of the NAAs, is the L-DOPA corresponding concentration of the nine NAAs for the carrier system [1,28]. Km(i) of NAAs on rat brain was used as follows: phenylalanine, 0.011; tryptophan, 0.015; leucine, 0.029; methionine, 0.040; isoleucine, 0.056; tyrosine, 0.064; histidine, 0.100; valine, 0.21; threonine, 0.22 (in mmolml – 1) [29]. As the Km value of L-DOPA, the value determined on rat capillary cerebral endothelial cells was used (0.072 mmolml – 1) [30].

Results Typical PET images of L-[b-11C]DOPA are shown in Fig. 1. The regional time–activity curves after intravenous injection of L-[b-11C]DOPA as average of all subjects, which were normalized for body weight and injected dose, are shown in Fig. 2. The radioactivity concentrations in the striatum were higher than in the cerebral cortices 30 min after injection. Among the cerebral cortices, the occipital cortex showed lowest radioactivity 40 min after injection. Coefficients of variation of regional radioactivity concentrations for each ROI were 17–21% on average of all frames. The fractions of radioactivity representing unchanged L-[b-11C]DOPA in plasma were 88.4 ± 4.6%, 35.8 ± 8.5%, 23.0 ± 3.1%, 19.7 ± 3.8%, 16.4 ± 2.0%, 15.4 ± 4.5% and 14.8 ± 4.1% (mean ±

SD) at 3.5, 18, 32, 46, 59, 75 and 89 min after injection, respectively. The fractions of radioactivity representing the 3-O-methyl metabolite, L-[11C]3-O-methyl-DOPA, in plasma were 1.8 ± 0.7%, 13.3 ± 2.7%, 16.7 ± 6.7%, 20.5 ± 4.6%, 24.8 ± 5.6%, 25.6 ± 5.1% and 27.6 ± 5.8% (mean ± SD) at 3.5, 18, 32, 46, 59, 75 and 89 min after injection, respectively. The rate constants, overall uptake rate constant K, Ve and Vb values obtained by the kinetic method are given in Table 1. The k3 and K values were greatest in the striatum. Lowest K values were observed in the cerebellum, parietal and occipital cortex. The Ve values ranged from 0.65 to 0.76 mlml – 1 among the neocortical regions. Comparison between the weighted sum of NAAs and K1 calculated by the kinetic method is shown in Fig. 3(a). Significant negative correlations were observed between the weighted sum of NAAs and K1 in almost all brain regions (P < 0.05, correlation coefficient, r = – 0.64 to – 0.78) except the parahippocampal gyrus and caudate head, where a tendency of decrease in K1 with the weighted sum of NAAs was observed (P > 0.1, r = – 0.59 to – 0.62). Comparison between the weighted sum of NAAs and K calculated by the kinetic method is shown in Fig. 3(b). Significant negative correlations were observed between the weighted sum of NAAs and K in the putamen and temporal cortex (P < 0.05, r = – 0.64 to – 0.69). A tendency of decrease in K with the weighted sum of

Fig. 1

MRI

11

C-DOPA

Typical positron emission tomography (PET) images obtained by summation of the frames from 29 to 89 min after intravenous injection of L[b-11C]DOPA. Magnetic resonance images co-registered to the PET images using SPM2 are also shown.

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Quantification of

11

C-DOPA in human brain Ito et al. 727

Fig. 2

8000

Radioactivity (Bq.ml–1)

7000 6000 Cerebellum Midbrain Parahippocampal gyrus Thalamus Caudate head Putamen Anterior cingulate Frontal cortex Temporal cortex Parietal cortex Occipital cortex

5000 4000 3000 2000 1000 0 0

20

40

60

80

100

Time (min) Regional time–activity curves after intravenous injection of L-[b-11C]DOPA as an average of all subjects.

Table 1 Values for the rate constants, K1, k2 and k3; the overall uptake rate constant, K; the steady state volume of the reversible compartment in a brain region, Ve; and the blood volume, Vb, obtained by the kinetic method using three-compartment model with three rate constants Region Cerebellum Midbrain Parahippocampal gyrus Thalamus Caudate head Putamen Anterior cingulate Frontal cortex Temporal cortex Parietal cortex Occipital cortex

K1(mlml – 1min – 1)

k2(min – 1)

k3(min – 1)

K (mlml – 1min – 1)

Ve(mlml – 1)

Vb(mlml – 1)

0.0359 ± 0.0080 0.0219 ± 0.0051 0.0218 ± 0.0050

0.0366 ± 0.0052 0.0334 ± 0.0086 0.0287 ± 0.0121

0.0054 ± 0.0017 0.0238 ± 0.0066 0.0165 ± 0.0078

0.0045 ± 0.0017 0.0090 ± 0.0018 0.0079 ± 0.0018

0.863 ± 0.184 0.407 ± 0.136 0.524 ± 0.140

0.066 ± 0.024 0.048 ± 0.018 0.064 ± 0.028

0.0289 ± 0.0073 0.0246 ± 0.0052 0.0317 ± 0.0057 0.0269 ± 0.0051 0.0276 ± 0.0056 0.0276 ± 0.0057 0.0293 ± 0.0064 0.0357 ± 0.0069

0.0359 ± 0.0062 0.0226 ± 0.0186 0.0171 ± 0.0025 0.0257 ± 0.0047 0.0327 ± 0.0036 0.0290 ± 0.0039 0.0357 ± 0.0047 0.0475 ± 0.0066

0.0111 ± 0.0025 0.0571 ± 0.0411 0.0338 ± 0.0068 0.0080 ± 0.0036 0.0078 ± 0.0017 0.0074 ± 0.0016 0.0073 ± 0.0016 0.0080 ± 0.0017

0.0067 ± 0.0016 0.0177 ± 0.0035 0.0208 ± 0.0027 0.0059 ± 0.0024 0.0053 ± 0.0013 0.0056 ± 0.0015 0.0049 ± 0.0011 0.0051 ± 0.0012

0.620 ± 0.135 0.425 ± 0.200 0.648 ± 0.205 0.864 ± 0.344 0.687 ± 0.139 0.761 ± 0.123 0.687 ± 0.145 0.651 ± 0.133

0.051 ± 0.016 0.046 ± 0.017 0.056 ± 0.017 0.061 ± 0.021 0.053 ± 0.020 0.060 ± 0.022 0.058 ± 0.022 0.062 ± 0.023

Values are mean ± SD.

NAAs was also observed in other brain regions (P < 0.1, r = – 0.57 to – 0.63) except the cerebellum (r = – 0.50) and anterior cingulated (r = 0.03). No significant correlation was observed between the weighted sum of NAAs and k3 calculated by the kinetic method in any brain region.

depicted in Fig. 4(a). Good correlations were observed between the two values. Simulation of the relation between assumed K and kref calculated by the graphical method is shown in Fig. 4(b). The simulation study showed systemic underestimations of kref as compared with assumed K.

The kref values calculated by the graphical method are given in Table 2. By this method, the kref value in the occipital cortex was zero because this region was used as a reference region with no irreversible binding. Similar to the K values calculated by the kinetic method, the greatest kref values were observed in the striatum and the lowest kref values were observed in the cerebellum and parietal cortex.

Discussion

Comparison between the K values calculated by the kinetic method and kref values by the graphical method is

Kinetics of L-[b-11C]DOPA in the brain

After intravenous injection of L-[b-11C]DOPA, radioactivity was high in the striatum and low in the cerebellum and all neocortical regions, in particular the occipital cortex (Figs 1 and 2). The overall value of the uptake rate constant, K, was greatest in the striatum and lowest in the cerebellum, and parietal and occipital cortices (Table 2). These regional distributions were in good agreement with those of dopamine concentration measured in monkey [24] and aromatic L-amino acid

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728 Nuclear Medicine Communications 2006, Vol 27 No 9

receptor were observed in post-mortem brain [32–34] and living human brain [35,36].

Fig. 3

(a)

0.05 Putamen Temporal cortex

In the limbic system, including the anterior cingulate and parahippocampal gyrus, K values were slightly greater than in the neocortical regions (Table 2). In the anterior cingulate and parahippocampal gyrus representing the mesolimbic dopaminergic system, relatively high dopamine D2 receptor binding has also been reported [35,36]. Among the neocortical regions, the temporal cortex showed highest K values (Table 2), corresponding to the regional distribution of aromatic L-amino acid decarboxylase activity [25]. In addition, highest dopamine D2 receptor binding in the temporal cortex among the neocortical regions was also shown [35,36].

K1 (ml.ml−1.min−1)

0.04

0.03

0.02

0.01

0 1000

y=−0.000031x+ 0.072, r = 0.64 (P<0.05) y=−0.000033x+ 0.070, r = 0.69 (P<0.05) 1100

1200

1300

1400

11 L-[b- C]DOPA injected intravenously is metabolized to 11 L-[ C]-3-O-methyl-DOPA under catechol-O-methyl-

1500

Weighted sum of NAAs (nmol.ml–1) (b) 0.03

K (ml.ml−1.min−1)

Putamen Temporal cortex

0.02

0.01

y=−0.000015x+ 0.040, r= 0.64 (P < 0.05) y=−0.000009x+ 0.017, r = 0.69 (P < 0.05) 0 1000

1100

1200

1300

1400

1500

transferase (COMT) outside the brain [37–40]. The 3-O-methyl metabolite of L-DOPA is known to pass the blood–brain barrier and cause error in estimation of the overall uptake rate constant K [39,41,42]. However, it has been reported that 3-O-methylation of L-[b-11C]DOPA does not take place readily and rapidly as compared with 6-[18F]fluoro-L-DOPA in monkey and rat [39,40]. In the present study, the fraction of L-[11C]-3-O-methyl-DOPA in plasma was not as high as in the previous animal study [40]. In addition, the COMT inhibitor that inhibited 3O-methylation of L-DOPA did not change the uptake rate constant for L-[b-11C]DOPA, whereas it increased the uptake rate constant for 6-[18F]fluoro-L-DOPA in monkey [5,43]. Therefore, we did not consider the effects of metabolites crossing the blood–brain barrier [10], although they might hamper the accurate estimation of K. On the other hand, the effects of the 3-O-methyl metabolite in plasma should be considered in a study in which 6-[18F]fluoro-L-DOPA is used [2,3,44]. This might be one advantage of L-[b-11C]DOPA as compared with 6-[18F]fluoro-L-DOPA.

Weighted sum of NAAs (nmol.ml−1) (a) Comparison between the weighted sum of neutral amino acids (NAAs) and values of the rate constant K1 calculated by the kinetic method for the putamen and temporal cortex. (b) Comparison between the weighted sum of NAAs and K calculated by kinetic method for the putamen and temporal cortex.

decarboxylase activity measured in post-mortem brain [25]. In the midbrain, the K value was relatively greater than in the neocortical regions (Table 2), similar to the regional distribution of aromatic L-amino acid decarboxylase activity [25]. These regional distributions of K for 11 L-[b- C]DOPA were consistent with those of 18 6-[ F]fluoro-L-DOPA [31]. In the striatum and midbrain representing the nigrostriatal dopaminergic system, high densities of dopamine transporter [32] and dopamine D2

It has been reported that the radioactivity in rat brain after intravenous injection of L-[b-11C]DOPA consisted of the radioactivity signal of 11C-labelled dopamine and its metabolites [45,46], indicating that 11C-labelled dopamine is the main metabolite of L-[b-11C]DOPA in the brain, and the overall uptake rate constant, K, of 11 L-[b- C]DOPA can represent the net rate of dopamine synthesis. The synthesis of noradrenaline takes longer than that of dopamine [10]. Amino acid transport at the blood–brain barrier L-DOPA

is transported from blood to brain by the same carrier system as neutral amino acid at the blood–brain barrier [27]. In the present study, the influx rate constant K1 of L-[b-11C]DOPA from blood to brain was greatest in the cerebellum (Table 1), similar to the regional

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Quantification of

Fig. 4

Region

(a)

kref(graphical method) 29–64 min*

29–89 min*

0.0045 ± 0.0017 0.0090 ± 0.0018 0.0079 ± 0.0018

– 0.0007 ± 0.0007 0.0059 ± 0.0014 0.0049 ± 0.0007

– 0.0006 ± 0.0004 0.0053 ± 0.0011 0.0037 ± 0.0005

0.0067 ± 0.0016 0.0177 ± 0.0035 0.0208 ± 0.0027 0.0059 ± 0.0024

0.0027 ± 0.0008 0.0139 ± 0.0026 0.0155 ± 0.0020 0.0034 ± 0.0012

0.0019 ± 0.0006 0.0122 ± 0.0018 0.0140 ± 0.0015 0.0020 ± 0.0008

0.0053 ± 0.0013 0.0056 ± 0.0015 0.0049 ± 0.0011 0.0051 ± 0.0012

0.0015 ± 0.0006 0.0021 ± 0.0007 0.0009 ± 0.0007 0.0000 ± 0.0000

0.0008 ± 0.0004 0.0013 ± 0.0005 0.0004 ± 0.0003 0.0000 ± 0.0000

Values are mean ± SD. Units are mlml – 1min – 1 for K and min – 1 for kref * Ranges of equilibrium time t*.

distribution of L-DOPA uptake sites in rat brain [27]. Regional distribution of the neutral amino acid transport system at the blood–brain barrier has been investigated using a natural neutral amino acid tracer, L-[2-18F]fluorophenylalanine [28], and a synthetic amino acid tracer, 11 C-aminocyclohexanecarboxylate [47] with PET in humans. Those studies showed the greatest influx rate constant across the blood–brain barrier in the cerebellum, agreeing well with the present results. In addition, values of the influx rate constant, K1, are almost at the same level (0.03–0.04 mlml – 1min – 1 in cerebral cortices) among the three radiotracers, L-[b-11C]DOPA, 18 11 L-[2- F]fluorophenylalanine, and C-aminocyclohexanecarboxylate. NAAs are transported by the neutral amino acid carrier system in the blood–brain barrier in a competitive fashion [26,28,48]. It has been reported that the uptake rate of 6-[18F]fluoro-L-DOPA into the brain was inhibited by amino acid loading in human [49] and monkey [50]. In the present study, the influx rate constant, K1, of 11 L-[b- C]DOPA significantly decreased with the weighted sum of NAAs, indicating a competitive transport of L-DOPA with NAAs at the blood–brain barrier (Fig. 3). The overall uptake rate constant K of 11 L-[b- C]DOPA also decreased with the weighted sum of NAAs, but no change in dopamine synthesis rate k3 was observed. Thus, the influx rate constant K1 would be an important factor of the overall uptake rate constant K in healthy human brain.

C-DOPA in human brain Ito et al. 729

0.03

0.02

0.01

0.00

0.00

0.01

0.02

0.03

K by kinetic method (ml.ml−1.min−1) (b)

kref by graphical method (min−1)

Cerebellum Midbrain Parahippocampal gyrus Thalamus Caudate head Putamen Anterior cingulated Frontal cortex Temporal cortex Parietal cortex Occipital cortex

K (kinetic method)

kref by graphical method (min−1)

Table 2 Overall uptake rate constant K and kref, calculated by the kinetic and graphical methods, respectively

11

0.03

0.02

0.01

0.00

0.00

0.01 Assumed K

0.02

0.03

(ml.ml−1.min−1)

(a) Comparison between overall uptake rate constant, K, calculated by the kinetic method, and kref by graphical method for all regions of interest (ROIs). The respective data indicate mean and SD of each ROI. Ranges of equilibrium time, t*, of 29–64 min and 29–89 min are used in the graphical method. (*), 29–64 min; y = 0.96x – 0.0036, r = 0.99, P < 0.001. (D) 29–89 min; y = 0.89x – 0.0038, r = 0.99, P < 0.001. (b) Simulation of relation between assumed K and kref calculated by graphical method. Ranges of equilibrium time t* were set at 29–64 min (——), 29–89 min (– – – –), and 64–89 min (- - - -).

Graphical method

The kref values calculated by the graphical method were in good agreement with the K values calculated by the kinetic method for both ranges of equilibrium time t* of 29–64 min and 29–89 min as well as the simulation studies (Fig. 4, Table 2). The systemic underestimation of kref values was observed as compared with K in both

experimental and simulated data. One of the reasons for this underestimation might be the very small irreversible binding in the occipital cortex which was used as a reference region in the graphical method (Tables 1 and 2). Because aromatic L-amino acid decarboxylase activity in the occipital cortex is low but not zero [25],

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irreversible binding of L-[b-11C]DOPA in the occipital cortex is not zero. By this method, the kref value in the cerebellum was a minus quantity (Table 2). It may be difficult to estimate the rate of dopamine synthesis in the cerebellum, as it also has low aromatic L-amino acid decarboxylase activity [25]. Since this method can avoid the need for an arterial input function, it can be expected to be useful for investigating the regional dopamine synthesis rate in patients. The K1 values of L-[b-11C]DOPA were very small, indicating small values for the capillary permeability– surface area product (Table 1). Thus, K1 values of 11 L-[b- C]DOPA are independent of cerebral blood flow [51,52]. With this tracer, quantitative methods using a reference brain region can be used even if the cerebral blood flow changes in a brain region as compared to a reference brain region.
0  0 kref is theoretically expressed by kref ¼ K = V e þ V p . The effective plasma volume of brain tissue Vp can be expressed by Vp = (1 – Ht)Vb, where Ht is hematocrit. When the Ht value is assumed to be 0.4, the Ve + Vp value in the occipital cortex is about 0.7 (Table 1).

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Conclusion Regional distribution of the overall uptake rate constant for L-[b-11C]DOPA was similar to that of aromatic Lamino acid decarboxylase activity measured in postmortem brain. Negative correlation between NAA concentration and the influx rate constant was observed, indicating that L-DOPA is transported through the same carrier system as NAAs at the blood–brain barrier in a competitive fashion. The overall uptake rate constants calculated by the graphical approach, in which the occipital cortex was used as reference brain region, were in good agreement with the values calculated by the kinetic approach. Because the graphical approach does not require an arterial input function, its application for the examination of the regional dopamine synthesis rate in neuropsychiatric and neurodegenerative diseases must be considered advantageous.

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Acknowledgements This study was supported in part by a Grant-in-Aid for Molecular Imaging Program from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japanese Government. We thank Dr Jun Kosaka, Dr Yota Fujimura, Dr Shoko Nozaki, Mr Katsuyuki Tanimoto, Mr Takahiro Shiraishi and Mr Akira Ando for their assistance in performing the PET experiments at the National Institute of Radiological Sciences. We also thank Ms Yoshiko Fukushima of the National Institute of Radiological Sciences for her help as clinical research coordinator.

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Original article

Value of 99mTc-TRODAT-1 SPECT to predict clinical response to methylphenidate treatment in adults with attention deficit hyperactivity disorder Christian la Fouge`rea, Johanna Krauseb, Klaus-Henning Krausec, Franz Josef Gildehausa, Marcus Hackera, Walter Kocha, Klaus Hahna, Klaus Tatscha and Stefan Dresela,d Background In a previous study, binding of 99mTc-TRODAT-1 to the dopamine transporter (DAT) was found to be higher in patients with attention deficit hyperactivity disorder (ADHD) as compared to healthy controls. Aim To determine whether the degree of 99mTc-TRODAT-1 binding to the striatal DAT may have a predictive role on the response to methylphenidate (MPH) in patients with ADHD. Methods Twenty-two adult patients suffering from ADHD underwent a brain SPECT scan with 99mTc-TRODAT-1. After the scan patients received MPH, individually medicated up to 60 mg
day – 1. Severity of illness was estimated using the Clinical Global Impression (CGI-S) Scale before treatment. Ten weeks after the beginning of MPH treatment the improvement in global symptoms was rated by the Clinical Global Improvement Scale (CGI-I). Results Before treatment 17/22 patients with ADHD presented with higher striatal DAT binding as compared to age-matched healthy controls ( + 23.8%; P < 0.01). After treatment with MPH a significant improvement of ADHD symptoms was demonstrated by the CGI-I in 16 of these 17 patients (CGI-S before: 4.8; CGI-I after MPH: 1.9; P < 0.01). Five patients showed reduced DAT binding prior to therapy

Introduction Attention deficit hyperactivity disorder (ADHD) is the most commonly diagnosed behavioural disorder of childhood and affects between 5 and 9% of school-age population in the United States [1,2]. Although ADHD was once thought to disappear as children grow up, data suggest that one to two-thirds of children with ADHD continue to have significant symptoms throughout life [3] even if symptoms and concerns typically appear differently in adults [4]. The American Psychiatric Association differentiates three types of ADHD: the predominantly hyperactive impulsive type, the predominantly inattentive type and finally the ADHD combined type [5]. Currently, it is still unclear whether ADHD subtypes reflect a common neuropathological pathway or represent distinct disorders.

( – 14.4%; P = 0.04); these patients did not respond to MPH therapy (CGI-S before: 4.5; CGI-I after MPH: 4.2; P = 0.40). Conclusion Our findings suggest that ADHD patients with primarily elevated binding of 99mTc-TRODAT-1 to the striatal DAT responded better to therapy with MPH as compared to those with normal or low DAT binding. Consequently, our results – even if obtained on a small collective indicate that measurement of DAT may be an important prognostic predictor for therapy response to MPH. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:733–737 Nuclear Medicine Communications 2006, 27:733–737 Keywords: attention deficit hyperactivity disorder, CNS stimulants, methylphenidate, dopamine transporter, 99mTc-TRODAT-1 SPECT a Department of Nuclear Medicine, cFriedrich-Baur Institute, Ludwig-MaximiliansUniversity of Munich, bOutpatient Clinic for Psychiatry and Psychotherapy, Ottobrunn and dDepartment of Nuclear Medicine, Helios Klinikum Berlin, Germany.

Correspondence to Dr Christian la Fouge`re, Department of Nuclear Medicine, Ludwig-Maximilians-University of Munich, Marchioninistr. 15, D-81377 Munich, Germany. Tel: + 49 89 7095 4646; fax: + 49 89 7095 7646; e-mail: [email protected] Received 4 March 2006 Accepted 31 May 2006

There is considerable evidence that abnormalities of the dopaminergic system play an important role in the pathophysiology of ADHD. Molecular genetic studies have suggested that four genes are associated with ADHD (the dopamine D4 and D5 receptors as well as the dopamine and serotonin transporters) [6]. Considering an evident involvement of the DAT in ADHD it seemed to be appropriate to investigate this aspect in vivo. By now, several SPECT studies have demonstrated an increment of DAT binding with different radiopharmaceuticals, such as 123I-IPT [7,8], 123I-altropane [9] and 99mTc-TRODAT-1 [10], while in another study [11] no statistically significant difference in DAT binding was seen in eight ADHD patients compared to healthy controls when 123I-b-CIT-SPECT was used. Psychostimulants such as methylphenidate (MPH),

c 2006 Lippincott Williams & Wilkins 0143-3636

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Nuclear Medicine Communications 2006, Vol 27 No 9

which are used as first-choice drugs in treating ADHD symptoms in children as well as in adults, interact directly with the DAT due to their properties as high potent blockers of DAT. Since 20–30% of ADHD patients do not respond to stimulant therapy [12,13] and in a previous 99m Tc-TRODAT-1 study some patients presented with lower DAT binding compared to controls, it might be of interest to follow the hypothesis of whether the degree of DAT binding is related to a response to stimulants. Therefore, the aim of this study was to compare striatal DAT binding in MPH responders and non-responders with ADHD and to investigate whether DAT binding may be a useful parameter to predict clinical response to MPH treatment in adult ADHD patients. DAT binding was assessed with 99mTc-TRODAT-1 [14], which is a new ligand of a tropane derivative, with high affinity and specificity for the pre-synaptic DAT on dopamine nerve terminals in the striatum [14,15]. This was also shown by previous studies, which demonstrated the highest uptake of 99mTc-TRODAT-1 in the dopamine transporter rich areas of the caudate nucleus and putamen, whereas binding to extra-striatal regions is considered unspecific [16]. The usefulness of 99mTc-TRODAT-1 for assessment of the DAT availability in the region of the striatum has already been proven by recent publications [10,17–21].

Patients, materials and methods Twenty-two non-smoking and non-medicated patients with ADHD (11 women, 11 men; age range 21–57 years) were investigated. The diagnosis of ADHD was made by structured interviews according to DSM-IV criteria. 99m

Tc-TRODAT-1 was prepared as reported in a previous publication [22] with minor modifications. Briefly, it was prepared by reconstitution of a pre-formulated lyophilized vial (provided by the Institute of Nuclear Energy Research, Taipei, Taiwan) with 3700 MBq freshly eluted [99mTc]pertechnetate in 5 ml saline. The TRODAT-1 vial was heated at 80–1001C for 30 min. The 99mTc-TRODAT1 was obtained in a neutral solution (pH 7.0–7.5) with > 90% radiochemical purity, measured by reversed phase high-performance liquid chromatography and cartridge methods. Single photon emission computed tomography (SPECT) acquisitions were performed over a period of 50 min, 3 h after intravenous administration of 740 MBq 99mTcTRODAT-1. For acquisition a Prism 3000 triple-headed gamma camera (Philips, formerly by Picker, Cleveland, Ohio) equipped with high resolution fan beam collimators was used. The acquisition parameters included a 15% energy window centred on 140 keV, a rotational radius of 13 cm or less, 120 projection angles over 3601, and a 128  128 matrix with a pixel width of 2.26 mm in the projection domain. The projection images were reconstructed by filtered back-projection and filtered by a low

pass filter. For uniform attenuation correction, Chang’s first-order method was used. Images were uniformly resliced by drawing a line connecting the anterior-most aspect of the frontal pole to the posterior-most aspect of the occipital pole, which approximates the line connecting anterior and posterior commissures (AC–PC line). In order to assess specific tracer uptake in the striatum, we used the region of interest (ROI) approach. In each patient, data were evaluated in the two consecutive transverse slices showing the highest tracer accumulation in the basal ganglia. The arithmetic mean of these two slices was calculated. Mean specific binding in the basal ganglia regions was calculated by subtracting the mean counts per pixel in the cerebellum as background (BKG) from the basal ganglia region (STR) and dividing the results by the mean counts per pixel in the background [(STR – BKG)/BKG]. Templates were used for defining the striatal ROIs. The size and shape of the templates was established and optimized using the data of the control group. The non-specific background activity was estimated by drawing a ROI around the cerebellum. The control group utilized for this study consisted of 14 healthy, non-smoking subjects (six women, eight men, age range 21–63 years). Owing to the known agedependent decrease in DAT density [23,24], the healthy controls were divided into two age groups ( < 40 years and Z 40 years). The value of specific DAT binding in patients was expressed as specific binding [(STR – BKG)/ BKG] and as the percentage of deviation from the value obtained for the respective age-matched control group. After the SPECT investigation the patients received MPH at an initial dose of 5 mg, followed by titration in relation to the clinical improvement, up to 60 mg per day. Before treatment the severity of ADHD was initially judged by Clinical Global Impression Scale (CGI-S) for each patient. This scale is a single-item clinician rating of the clinician’s assessment of the severity of ADHD symptoms with ranges from 1 (not ill at all) to 7 (maximum profound impairment) [25]. After 10 weeks the global improvement was rated by Clinical Global Improvement Scale (CGI-I) [26]. This is an observational scale of global evaluation ranging from 1 (very much improved) to 7 (very much worse), which assesses the change in degree of illness in relation to the original assessment. Patients with a value of more than 4, which is defined as no change or worse, were determined as nonresponders. For statistical analyses, the Fisher test, regression analyses and Student’s t-test were used. Differences were considered to be statistically significant when P < 0.05 and highly significant when P < 0.01.

Results Seventeen of 22 untreated ADHD patients presented with an increased specific binding of 99mTc-TRODAT-1

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DAT binding as a predictor to MPH response la Fouge`re et al. 735

to the DAT ( + 23.8% ± 11.3%; P < 0.01) (Table 1) as compared to the mean specific binding of the corresponding control group (specific binding( < 40y), 1.27 ± 0.10; specific binding( Z 40y), 1.17 ± 0.11). In 16 of these 17 patients a significant symptom improvement could be shown under MPH treatment (CGI-S before MPH therapy: 4.8 ± 0.9; CGI-I after MPH therapy: 1.9 ± 0.7; P < 0.01). Five of 22 patients suffering from ADHD showed a DAT binding below the mean specific binding of the corresponding age group ( – 14.4% ± 8.7%; P = 0.04) (Table 1). All of these patients were non-responders (CGI-S before MPH therapy: 4.5 ± 0.8; CGI-I after MPH medication: 4.2 ± 0.4; P = 0.40). Responders and nonresponders did not differ statistically significant in CGI-S values before MPH treatment (CGI-S responders: 4.8; CGI-S non-responders: 4.5; P = 0.47) and in mean age (responders: 37.8; non-responders: 44.2; P = 0.19). Table 1 displays individual CGI-S values before and CGII values after MPH therapy and individual specific binding values to the DAT before treatment with MPH. However, the small number of patients included did not allow subgroup analysis regarding differences in ADHD subtypes (e.g., inattentive-type, combined-type and hyperactive/impulsive-type) as shown in Table 2. Our own results for non-smoking adults with persistent symptoms of ADHD revealed, though, that DAT increases in ADHD did not allow differentiation of 99m Table 1 Individual specific binding values of Tc-TRODAT-1 to the striatal dopamine transporter (DAT) and individual CGI values in ADHD patients responding and not responding respectively to methylphenidate treatment

Patient number

Specific binding to the DAT

Deviation (%) to age-matched control group

CGI-S before MPH treatment

CGI-I after MPH treatment

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16

1.90 1.39 1.50 1.34 1.61 1.39 1.54 1.51 1.58 1.75 1.49 1.27 1.63 1.62 1.51 1.63

49.6 18.8 28.2 5.5 26.8 9.4 21.3 18.9 24.4 37.8 17.3 8.5 28.3 27.6 18.9 39.3

6 3 4 5 6 5 5 6 5 4 4 5 5 4 4 5

1 2 2 3 1 2 2 1 3 3 1 2 1 2 2 2

17 18 19 20 21 22

1.01 1.08 1.56 1.12 0.92 0.88

– 13.7 – 7.7 22.8 – 4.3 – 21.4 – 24.8

5 3 5 5 5 4

4 4 4 4 5 4

CGI-S, Clinical Global Impression Scale; CGI-I, Clinical Global Improvement Scale; AHDH, attention deficit hyperactivity disorder.

Table 2 Patients’ characteristics for the entire group suffering from ADHD (n = 22) and for two subgroups different for age ( < 40 years, n = 8; Z 40 years, n = 10) Characteristic Sex (M/F) Age (n; mean ± SD) < 40 years Z 40 years CGI-S before MPH (mean ± SD) < 40 years Z 40 years CGI-I after MPH (mean ± SD) < 40 years Z 40 years ADHD subtype Combined Inattentive Hyperactive/impulsive [(STR – BKG)/BKG] (mean ± SD) < 40 years Z 40 years

Responders 7/9 16; 37.8 ± 11.0 10; 30.2 ± 5.7* 6; 47.9 ± 7.7 4.8 ± 0.9

Non-responders 4/2 6; 44.2 ± 6.1 1; 33.7 ± 0* 5; 46.2 ± 6.1 4.5 ± 0.8

4.9 ± 0.9* 4.5 ± 0.8 1.9 ± 0.7***

5 ± 0* 4.4 ± 0.9 4.2 ± 0.4***

1.6 ± 0.7* 2.3 ± 0.5***

4 ± 0* 4.2 ± 0.4***

9 6 1 1.60 ± 0.15* 1.45 ± 0.14***

2 4 0 1.56 ± 0* 1.00 ± 0.10***

P values: *not calculable; ** < 0.05; *** < 0.01. All other values showed no statistically significant difference. STR, mean counts per pixel in the basal ganglia region; BKG, mean counts per pixel in the cerebellum (the background value).

different subtypes of ADHS. Nevertheless, 99mTcTRODAT-1 may play an important role in the diagnosis of all subtypes, even if up to now the number of patients is too small to allow a definite conclusion for differentiating the subtypes [21]. No significant difference was detected between male and female patients and no right/left asymmetry was found. Regression analysis showed a significant negative correlation between DAT binding and CGI-I (r = 0.72; P < 0.01). Table 2 shows patients’ characteristics for the entire ADHD collective. Figure 1 displays the specific binding [(STR – BKG)/BKG] of 99mTc-TRODAT-1 to striatal DAT depending on age in responders, nonresponders and healthy controls.

Discussion MPH is the first-line drug for treatment of ADHD and seems to have a reasonable efficacy and safety profile. The progressive increase in prescribing psychostimulants such as MPH, however, has considerably stimulated the debate among psychiatrists, whether long-term administration to adolescents may potentially alter neural development and behaviour or could increase the risk of substance abuse [27]. In addition, there is growing evidence that a subset of ADHD patients will either fail to respond to stimulants (20–30%) [12,13] or will present with side effects that preclude their use. Therefore, it may be of great interest to identify those patients, who would benefit from MPH therapy. Despite extensive investigations the exact aetiology and the neurobiological basis of ADHD still remains

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Fig. 1

2

Controls Responder Non-responder

[(STR-BKG)/BKG]

1.8 1.6 1.4 1.2 1

The previously described decrease of DAT binding after MPH treatment [10], which is known to directly interact with the DAT and the documented improvement in clinical symptoms, correlates with the good response to MPH treatment in patients with high DAT levels. However, it still remains unclear, whether the increased DAT binding before MPH medication is either due to elevated dopamine which triggers the rise in DAT levels or whether it causes a reduction of dopamine in the synaptic cleft [30].

0.8 0.6 15

25

35

45 Age (y)

55

65

Specific binding [(STR – BKG)/BKG] of 99mTc-TRODAT-1 to striatal DAT depending on age in responders, non-responders and healthy controls. STR, mean counts per pixel in the basal ganglia region; BKG, mean counts per pixel in the cerebellum (the background value).

unresolved. However, converging evidence suggests at least an implication of the dopaminergic system in the pathogenesis of ADHD [21,27,28]. In this context, molecular genetics studies have demonstrated an overexpression and involvement of the presynaptically located DAT-1 gene (SLC3A6) in ADHD [28,29]. In addition, several studies have not only demonstrated higher DAT binding in vivo [7,9,10], but also a significant intraindividual decrease in DAT binding of about 30% after MPH treatment which was significantly correlated with an improvement of clinical symptoms [10]. In accordance with previous reports [7,9,10] the majority of our ADHD patients (17 of 22) showed higher striatal DAT concentration than non-smoking healthy volunteers without ADHD. However, a subgroup of five patients with ADHD presented with DAT levels below those of the control group, all of them without statistically significant differences in the severity of illness measured by the CGI-S. Therefore ADHD may not necessarily be associated with higher striatal DAT levels. To our knowledge no study has related DAT binding with the three subtypes of ADHD until now. This could offer one possible explanation of the discrepancy between the studies mentioned above, which either showed an increase in DAT binding or documented normal findings [11]. The major finding of our study, however, is that all except one ADHD patient (16 of 17) presenting with high striatal DAT binding responded well to MPH therapy, while none of ADHD patients with lower DAT levels showed a significant improvement of ADHD symptoms. This suggests that the DAT status in untreated ADHD patients may be used as a predictor of response to MPH therapy.

The patients with lower DAT binding in our study had no significant benefit from MPH therapy. On the basis of these findings it may be hypothesized that the reduced striatal DAT binding in these patients could be a secondary effect which results from a possible downregulation of the dopamine transporter. Furthermore, our clinical experience shows that most of the MPH nonresponders had a good benefit from therapy with amphetamines, which are known to influence the catecholamines in a more complex way than MPH. Interestingly, our results are in contrast to the most recent findings by Cheon et al. [8] who investigated the treatment response to MPH in 11 Korean children with ADHD taking into account the homozygosity for 10repeat allele at the DAT-1 gene and DAT binding. They described a good response to MPH in four children with an absence of homozygosity at the DAT-1 gene, which was associated with low DAT binding in the basal ganglia. In contrast only two of seven children with homozygosity and higher DAT binding showed an adequate response to MPH [8]. However, the authors themselves stated as limitations of their study the small number of subjects investigated and the lack of age-matched normal controls for defining ‘normal’ or ‘pathological’ DAT binding. In addition, they admitted that due to the higher DAT availability homozygotic children possibly might have needed higher doses of MPH, for an appropriate improvement of symptoms. One major limitation of this study is that, because of ethical reasons, only a small number of patients were investigated. Furthermore, recruitment of adult patients presenting with persistent symptoms of ADHD and not being biased by the influence of nicotine or previous medication is rather difficult. These are limitations that are inherent to all imaging studies in adults suffering from ADHD. Therefore, additional studies with higher number of patients are necessary to confirm these finding and may also help to further clarify discrepant findings.

Conclusion The findings of this study indicate that patients with ADHD and elevated binding levels of 99mTc-TRODAT-1 to the striatal DAT respond to therapy with MPH better

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DAT binding as a predictor to MPH response la Fouge`re et al. 737

than those with low binding values. Thus, it can be suggested that striatal DAT binding may be an important prognostic predictor for responsiveness to stimulant medication such as MPH in patients suffering from ADHD. Therefore 99mTc-TRODAT-1 SPECT might be a useful tool to support the physician in finding the most suitable treatment of ADHD symptoms in patients, being either a stimulant monotherapy or a combination with alternative drugs.

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References 1

Cantwell DP. Attention deficit disorder: a review of the past 10 years. J Am Acad Child Adolesc Psychiatry 1996; 35:978–987. 2 Goldman LS, Genel M, Bezman RJ, Slanetz PJ. Diagnosis and treatment of attention-deficit/hyperactivity disorder in children and adolescents. Council on Scientific Affairs, American Medical Association. JAMA 1998; 279:1100–1107. 3 Wender PH, Ward MF, Reimherr FW, Marchant BK. ADHD in adults. J Am Acad Child Adolesc Psychiatry 2000; 39:543. 4 Resnick RJ. Attention deficit hyperactivity disorder in teens and adults: They don’t all outgrow it. J Clin Psychol 2005; 61:529–533. 5 American Psychiatric Association. Diagnostic and Statistical Manual of Mental Disorders, 4th edition. Washington, DC: American Psychiatric Association; 1994, pp. 78–85. 6 Bobb AJ, Castellanos FX, Addington AM, Rapoport JL. Molecular genetic studies of ADHD: 1991 to 2004. Am J Med Genet B Neuropsychiatr Genet 2005; 132:109–125. 7 Cheon KA, Ryu YH, Kim YK, Namkoong K, Kim CH, Lee JD. Dopamine transporter density in the basal ganglia assessed with 123I-IPT SPET in children with attention deficit hyperactivity disorder. Eur J Nucl Med 2003; 30:306–311. 8 Cheon KA, Ryu YH, Kim JW, Cho DY. The homozygosity for 10-repeat allele at dopamine transporter gene and dopamine transporter density in Korean children with attention deficit hyperactivity disorder: relating to treatment response to methylphenidate. Eur Neuropsychopharmacol 2005; 15:95–101. 9 Dougherty DD, Bonab AA, Spencer TJ, Rauch SL, Madras BK, Fischman AJ. Dopamine transporter density in patients with attention deficit hyperactivity disorder. Lancet 1999; 354:2132–2133. 10 Dresel S, Krause J, Krause KH, la Fougere C, Brinkbaumer K, Kung HF, et al. Attention deficit hyperactivity disorder: binding of 99mTc-TRODAT-1 to the dopamine transporter before and after methylphenidate treatment. Eur J Nucl Med 2000; 27:1518–1524. 11 van Dyck CH, Quinlan DM, Cretella LM, Staley JK, Malison RT, Baldwin RM, et al. Unaltered dopamine transporter availability in adult attention deficit hyperactivity disorder. Am J Psychiatry 2002; 159:309–312. 12 Elia J, Ambrosini PJ, Rapoport JL. Treatment of attention-deficit-hyperactivity disorder. N Engl J Med 1999; 340:780–788.

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Original article

Evaluation of a new expert system for fully automated detection of the Alzheimer’s dementia pattern in FDG PET Daniel von Borczyskowski, Florian Wilke, Brigitte Martin, Winfried Brenner, Malte Clausen, Janos Mester and Ralph Buchert Objective Fluorodeoxyglucose (FDG) positron emission tomography (PET) is increasingly used to support a diagnosis of Alzheimer’s disease. The aim of the present study was to evaluate a new expert system (PALZ) for the fully automated analysis of FDG PET images for diagnosis of the disease. Methods The PALZ tool is based on the detection of the typical disease pattern in FDG PET images. Its potential for this task was evaluated in 22 consecutive patients with suspected Alzheimer’s disease who had been graded as positive for the pattern by an experienced reader (visual analysis supported by statistical parametric mapping (SPM)), and in 18 controls. Dependence on scanner performance was assessed by variation of the spatial resolution of the PET images.

transmission scan). There was only mild dependence on spatial resolution. Conclusions The results of the present study suggest that the PALZ tool provides similar performance for the detection of the typical Alzheimer’s disease pattern in FDG PET images as an experienced reader supported by SPM. The PALZ tool is fully automated, easy to use, and insensitive to the spatial resolution of the PET scanner used. Therefore, it has the potential for widespread clinical c 2006 Lippincott use. Nucl Med Commun 27:739–743 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:739–743 Keywords: Alzheimer’s disease, positron emission tomography, 2-[18F]fluoro-2-deoxy-D -glucose, statistical mapping, expert system

Results All the Alzheimer’s disease subjects were classified as pattern-positive by the PALZ tool. Fifteen controls were classified as normal. Sensitivity and specificity for differentiation of the patients with suspected Alzheimer’s disease from the controls were 100% and 83%, respectively. The false positive finding in three controls most likely was caused by differences in attenuation correction between the normal data base of the PALZ tool (cold transmission scan) and the local data sets (hot

Department of Nuclear Medicine, University Medical Center Hamburg-Eppendorf, Germany.

Introduction

In routine patient care, evaluation of a brain FDG PET scan is usually based on the visual inspection of the reconstructed tomographic slices. However, visual evaluation of FDG PET images strongly depends on the experience of the interpreter, particularly in case of modest alterations of brain metabolism in the early stages of Alzheimer’s disease. Therefore, automatic anatomical standardization and voxel-based statistical evaluation are being increasingly used for the evaluation of brain PET scans [8–12]. These new techniques allow automated and operator-independent evaluation of brain PET scans. They can substantially support conventional evaluation and, thus, lead to improved quality and stability of the interpretation of brain FDG PET in clinical routine. There are several software packages available which provide tools for automatic anatomical standardization and voxel-based statistical analysis of brain scans, e.g., Statistical Parametric Mapping (SPM, Wellcome Department of Cognitive Neurology, London, UK) [13,14],

Dementia is a widespread disease of old age and affects up to 20% of the population. The most common cause of dementia is Alzheimer’s disease. Even though there are well established criteria for clinical evaluation of the disease [1], the correct diagnosis is often established belated, and sometimes not until a histopathological examination has been undertaken (cf. Knopman et al. [2] and references therein). The need for an accurate diagnosis of the disease has increased recently because of the development of new medications. Positron emission tomography (PET) with the glucose analogue 2-[18F]fluoro-2-deoxy-D-glucose (18F-FDG) is increasingly used to aid the diagnosis of Alzheimer’s disease [3]. The FDG PET diagnosis is based on depicting the typical pattern of hypometabolic areas in the brain [4,5] which is already present early in the course of the disease [6,7].

Correspondence to Dr Ralph Buchert, Department of Nuclear Medicine, University Medical Center Hamburg-Eppendorf, Martinistr. 52, D-20246 Hamburg, Germany. Tel: + 0049 40 42803 6106; fax: + 0049 40 42803 5181; e-mail: [email protected]

Received 20 February 2006 Accepted 28 June 2006

c 2006 Lippincott Williams & Wilkins 0143-3636

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Neurological Statistical Image Analysis (NEUROSTAT, S. Minoshima, University of Washington Medical School, Seattle, Washington) [8,15], Brain Registration and Analysis (BRASS, Nuclear Diagnostics, Stockholm, Sweden) [16], Sub-volume Thresholding Analysis (SVT, Laboratory of Neuro Imaging, University of California, Los Angeles, California) [17], Scenium (Siemens CTI Molecular Imaging, Knoxville, Tennessee), and NeuroQ (Syntermed, Atlanta, Georgia). These tools are very useful for the observer-independent identification of hypometabolic areas. However, interpretation of statistical maps, i.e., recognition of the typical Alzheimer’s disease pattern, for example, remains largely the task of medical experts. Recently, a new software package, the PMOD Alzheimer’s discrimination analysis tool PALZ (PMOD Technologies Ltd., Adliswil, Switzerland) has been made available, which is designed to detect the typical Alzheimer’s disease pattern of hypometabolism from the statistical maps for the diagnosis of the disease. The purpose of the present study was to evaluate the potential of the PALZ tool for the detection of Alzheimer’s disease. In addition, its stability with respect to differences in scanner performance was assessed by variation of the spatial resolution of the PET images.

Methods The PALZ tool

The Alzheimer’s discrimination analysis software PALZ is a tool that is optionally available with the PMOD software (PMOD Technologies Ltd., Adliswil, Switzerland) [18]. The PALZ tool implements the automatic Alzheimer discrimination method developed by Herholz et al. [19], which is based on the outcome of a vast multicentre trial. In short, the static FDG PET image of the individual patient is (1) stereotactically normalized to the standard PET brain template of the SPM99 software package [13], (2) three-dimensionally smoothed by a Gaussian filter of 12 mm full width at half maximum (FWHM), (3) scaled to an average intensity of 1 within a predefined mask representing regions with preserved glucose uptake in Alzheimer’s disease (midbrain, putamen, insula, and sensorimotor and visual cortex; total volume = 177 ml), and (4) compared to a normal database of 49 healthy controls from eight different centres on a voxel-by-voxel base using a two-sample t-test. The final step is the computation of the quantity AD-t-sum, which is defined as the sum of t-values over all voxels with an age-adjusted P < 0.05 (uncorrected) within a predefined mask of Alzheimer’s disease-related brain regions (temperoparietal cortex, posterior cingulate, precuneus and frontal association cortex; total volume = 165 ml). A value of AD-t-sum > 11 090 is considered abnormal. This threshold had been determined previously as the upper

95% confidence limit in an independent normal data base of 61 healthy subjects [19]. Alzheimer’s disease subjects

To evaluate the sensitivity of the PALZ tool for detection of the Alzheimer’s disease pattern in FDG PET images, it was first applied retrospectively to 22 consecutive patients (seven women, 15 men; age, 66.8 ± 9.0 years), who had received brain FDG PET in our department because of suspected Alzheimer’s disease ( = inclusion criterion I1). Visual evaluation of the PET images by an experienced physician supported by voxel-based statistical analysis with SPM2 [13,14] had resulted in grading scale 3 (classic bilateral temporo-parietal hypometabolism) as described and validated by Hoffman et al. [20] in these subjects ( = inclusion criterion I2). Thus, all 22 subjects showed the typical Alzheimer’s disease pattern according to the SPM supported evaluation by an experienced reader. Control subjects

To assess the specificity of the PALZ tool, it was also applied to 18 control subjects in whom whole-body FDG PET including the brain had been performed because of an oncological indication (malignant melanoma) (nine women, nine men; age, 59.4 ± 5.2 years). Subject preparation, PET scanning of the brain and image processing had been performed exactly as in the Alzheimer’s disease group. Exclusion criteria were (E1) known psychiatric or neurological disorder, and (E2) reported PET abnormalities in the brain. Finally, the control subjects were without pathological findings according to voxel-based statistical evaluation (SPM) using the leave-one-out strategy (no significant clusters of either hypometabolism or hypermetabolism at the P = 0.05 level corrected for multiple comparisons). PET imaging

PET imaging was performed on a full-ring PET system ECAT EXACT 921/47 (Siemens/CTI, Knoxville, Tennessee, USA) in two-dimensional mode [21]. Patients fasted for at least 4 h prior to injection of 300–400 MBq FDG. A static emission scan of 21 min duration (composed of seven frames of 3 min duration each) was acquired 35–45 min after injection. Following the emission scan, a ‘hot’ transmission scan was performed using three rotating 68Ge rod sources. The duration of the transmission scan (4–10 min) was adjusted to the actual radioactivity of the rod sources (50–120 MBq each) to make sure that the number of trues in the transmission scan was not significantly smaller than the number of trues in the emission scan. The emission sinograms were corrected for random coincidences, radioactive decay, dead time, varying detector efficiency, scatter, attenuation and non-uniform sampling (geometric arc correction). The scatter correction was performed using the convolution subtraction method developed by Bergstrom

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Automated detection of AD pattern in FDG PET images von Borczyskowski et al. 741

No particular expedients were performed to accommodate for biases due to technical differences between the PET system used in the subjects included in the present study and the PET systems that had been used in the acquisition of the normal data base of the PALZ tool. Scanner dependence

In order to evaluate the dependence of the AD-t-sum of the PALZ tool on the spatial resolution in the reconstructed PET images, the analysis was repeated in two control subjects (including the one with highest AD-tsum) prior to smoothing the original images (FWHME6 mm), and after smoothing the images with an isotropic three-dimensional Gaussian kernel to 8, 9, 10 and 12 mm FWHM.

Fig. 2

AD-t-sum

et al. [22] using a stationary kernel as implemented in the scanner software (ECAT 6.5B). Forty-seven transaxial slices with 128  128 voxels were reconstructed using filtered back-projection with a cut-off at the Nyquist frequency and no filter. The voxel size was 1.7  1.7  3.4 mm3 and spatial resolution was about 6 mm FWHM. Finally, images were three-dimensionally smoothed with a Gaussian kernel of 3.6 mm FWHM resulting in spatial resolution of about 7 mm FWHM.

20 000 18 000 16 000 14 000 12 000 10 000 8000 6000 4000 2000 0

p05545 p05723 0

2

4

6 8 FWHM (mm)

10

12

14

Dependence of the Alzheimer’s disease score (AD-t-sum) of the PALZ tool on the spatial resolution in the reconstructed PET images in two representative control subjects. The dashed line represents the upper threshold of the normal range.

therefore, were classified as mildly abnormal. Visual inspection of the t-maps of these controls revealed reduced FDG metabolism mainly in central brain structures like striatum, thalamus and cingulate. Sensitivity and specificity for differentiation of the patients with suspected Alzheimer’s disease from the controls were 100% and 83%, respectively.

Results All 22 patients with suspected Alzheimer’s disease were classified as having the typical Alzheimer’s disease pattern by the PALZ tool (AD-t-sum = 12 026 to 122 160; median = 44 168) (Fig. 1). Fifteen of the 18 controls were classified as normal by the PALZ tool (ADt-sum = 5000 to 10 200). The remaining three controls had an AD-t-sum (13 414 to 15 704) only slightly higher than the upper threshold of the normal range and,

Fig. 1

1000 000

AD-t-sum

100 000

10 000

Patients with suspected AD (n= 22)

Controls (n =18)

1000 Alzheimer’s disease score (AD-t-sum) of the PALZ tool in patients with suspected Alzheimer’s disease and in controls. The dashed line represents the upper threshold of the normal range.

AD-t-sum showed only mild dependence on the spatial resolution in the reconstructed PET images (Fig. 2). The AD-t-sum classification, i.e., normal or abnormal, did not change when spatial resolution was varied.

Discussion Within the present sample, consisting of patients with suspected Alzheimer’s disease and control subjects with (presumably) healthy brains, the PALZ tool provided 100% sensitivity for selecting the patients with suspected Alzheimer’s disease. This result is in good agreement with numerous studies that reported excellent sensitivity of FDG PET for the detection of the disease [23,24]. It suggests that the PALZ tool provides a similar performance for the detection of the typical Alzheimer’s disease pattern in 18F-FDG PET images as does visual inspection by an experienced reader. In contrast to sensitivity, the specificity of the PALZ tool was somewhat limited in the present sample. With the value of 83%, the observed specificity was even somewhat smaller than the specificity of 90% or higher reported in the literature for the distinction of subjects with probable Alzheimer’s disease from normal controls [8,25]. This suggests a systematic ‘error’ in the present study. Three of the 18 controls had an AD-t-sum slightly above the upper threshold of the normal range and, therefore, were classified as (mildly) abnormal. The t-maps in these

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742 Nuclear Medicine Communications 2006, Vol 27 No 9

controls indicated reduced FDG uptake mainly in central brain structures, such as striatum, thalamus and cingulate. This pattern suggests an artifact caused by the fact that in the present study a post-injection (hot) transmission scan was used for attenuation correction while a preinjection (cold) transmission scan was used in the subjects of the normal database of the PALZ tool (Dr Herholz, private communication). A hot transmission scan is contaminated by emission counts so that the attenuation is underestimated leading to a corresponding underestimation of the tracer uptake [26]. The underestimation is not uniform throughout the whole brain, but is more pronounced in central brain structures [27]. Scaling the uptake images to an average intensity of 1 within a predefined mask representing regions with preserved glucose uptake in Alzheimer’s disease, as implemented in the PALZ tool, then is expected to result in underestimation of the scaled uptake in central structures (and overestimation in the cortex). This artefact might be avoided by correcting the hot transmission scan for the contamination by emission counts [26]. Nevertheless, when statistical analysis with the PALZ tool is planned, attenuation correction with a cold transmission scan is recommended. The effect of CT-based attenuation correction on the results of the PALZ tool should be investigated in a further study [28,29]. Calculated attenuation correction most likely is not appropriate for use with the PALZ tool, particularly those methods which are based on oversimplified models of the head. The calculated attenuation correction method implemented in the software (ECAT 6.5B) of the PET system used in the present study, for example, causes a systematic back-to-front gradient of the apparent tracer uptake relative to measured attenuation correction [30]. In general, scatter causes a much smaller effect in PET than attenuation. Differences in the scatter correction methods, therefore, are expected to have considerably smaller influence on the PALZ tool output than do differences in the attenuation correction method. In the present data, complete omission of scatter correction caused a decrease of the AD-t-sum of less than 10% (data not shown). This suggests that the accuracy of the PALZ tool is not significantly affected by the choice of the scatter correction method. The applicability of an external normal database might be limited not only by differences in attenuation and scatter correction, but also by differences in image characteristics because of differences in PET scanner performance and the reconstruction algorithms used [31]. The normal database of the PALZ tool is expected to be particularly widely applicable, since it is composed of images from eight different centres which used essentially four different scanner types, including both rather old

systems, with limited spatial resolution, and high-end systems that provide the best spatial resolution actually available with a whole-body PET system. In addition, PALZ applies strong smoothing prior to the statistical testing (kernel with 12 mm FWHM), which might further reduce the effects of differences in scanner performance, particularly the effects of differences in spatial resolution. In the present study, the dependence of the PALZ tool on scanner performance was tested by varying the spatial resolution in the reconstructed images between 6 and 12 mm FWHM. The main output of the PALZ tool, the AD-t-sum, was quite stable within this range of FWHM. Finally, applicability of the PALZ tool might also be limited by differences in patient preparation that affect FDG uptake in the brain, such as movement or any other task performed during the FDG uptake period. The subjects of the normal database of the PALZ tool had been examined in a resting state with eyes closed and ears unplugged. The subjects of the present study were scanned also in a resting state with ears unplugged, but with eyes open in a dimly lit room. This might cause different FDG uptake, particularly in the visual areas of the brain [32,33]. However, the present results suggest that a putative effect is rather small. Nevertheless, it is recommended that the preparation protocol used in subjects of the normal database of the PALZ tool is adopted [19]. For the computation of the AD-t-sum, the PALZ tool adjusts the confidence limits for age-related changes. This adjustment, based on age regression in the normal sample of the study by Herholz et al. [19], has been validated for Z 49 years only. The PALZ tool, therefore, requires the subject’s age to be Z 49 years. A major limitation of the PALZ tool is that, at least at present, it does not provide automatic facilities for the differential diagnosis of dementias. The automatic mode of the PALZ tool is restricted to the detection of hypometabolism in Alzheimer’s disease-related brain regions which might be present also in other types of dementia. This limits the use of the PALZ tool in an actual clinical setting. In the present study this limitation was taken into account by excluding metabolic patterns of other types of dementia from the analysis (inclusion criterion I2). However, the PALZ tool provides display of the statistical parametric map overlaid to the normalized PET image. This display can be used by an (experienced) observer for the differential diagnosis of dementias (similar as in SPM, for example). A further limitation of the PALZ tool is that it does not provide the option to create a new normal database to fit local protocols. Instead, it is recommended that the protocol is adapted to the protocol used in the normal

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Automated detection of AD pattern in FDG PET images von Borczyskowski et al. 743

database of the PALZ tool. In particular, attenuation correction should be based on a cold transmission scan.

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In conclusion, the results of the present study suggest that the PALZ tool is appropriate for the evaluation of brain FDG PET images in patients with suspected Alzheimer’s disease: it appears to provide similar sensitivity and specificity as an experienced interpreter supported by SPM. Being a fully automated expert system, i.e., independent of the experience of an interpreter, the PALZ tool might improve quality and, particularly, stability of the interpretation of brain FDG PET in patients with suspected disease. It might be particularly helpful to less expert readers, or in cases of very mild Alzheimer’s disease, when the visual evaluation is less certain. In addition, the PALZ tool appears to be rather insensitive to PET scanner performance so that there is no need to assemble an own local database of normal FDG PET images. The PALZ tool is easily installed on a recent PC under any MS Windows operating system, Linux or MacOSX. It is fast and easy to use. The PALZ tool has the potential for widespread clinical use.

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Review paper

The diabetic foot: Charcot joint and osteomyelitis Laura Giurato and Luigi Uccioli Foot problems are common causes of disability in diabetic patients with as many as 25% expected to develop severe foot or leg problems during their lifetimes. Although skin ulceration is the most frequent problem, bones may also be involved in two different clinical conditions: osteomyelitis and Charcot osteoarthropathy. Osteomyelitis causes complications in up to one third of diabetic foot infections and is due to direct contamination from a soft-tissue ulcer. Osteoarthropathy Charcot foot is a chronic and progressive disease of the bone and joints. Both osteomyelitis and Charcot joint are conditions with an increased risk of lower limb amputation, both may have a successful outcome when recognized and treated in the early stages. The major diagnostic difficulty is in distinguishing bone infection (osteomyelitis) from non-infectious neuropathic bony disorders as in osteoarthropathy Charcot foot. An additional difficulty is found when a bone infection superimposes a Charcot

Charcot neuroarthropathy in the diabetic foot A possible manifestation of the neuropathic foot is neuropathic osteoarthropathy, commonly referred to as Charcot’s joint. It is a chronic and progressive disease of the bone and joints. This pathological process culminates in bone and joint destruction and subsequent foot deformity, which predisposes to ulceration. The pathogenic mechanism(s) for the development of the Charcot foot have been the subject of a number of competing theories, but the pathogenesis remains unclear. It is certain that the cause is multifactorial [1,2]. A number of the mechanisms operate simultaneously: peripheral sensory and motor neuropathy, biomechanical factors, autonomic neuropathy are all considered potential causes in the development of the Charcot foot [3]. Motor neuropathy leads to changes in the integrity of the arches of the foot, with increased pressure loading at certain points. The concomitant sensory neuropathy leads to these pressure loads remaining unnoticed, which can induce microfractures and bone deformity. The repeated trauma in an insensitive foot leads to laxity of ligaments and instability of joints with resultant bone damage (neurotraumatic theory) [4,5]. According to neurovascular theory, sympathetic denervation is the leading cause through the induction of active bone resorption and osteopenia by osteoclasts. Autonomic neuropathy is responsible for impairment of vascular

osteopathy. This condition, which can be clinically suspected when foot ulceration appears in Charcot foot, needs to be diagnosed because it implies a different therapeutic strategy. This article aims to summarize both these two clinical conditions and give indications to make a timely and correct diagnosis. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:745–749 Nuclear Medicine Communications 2006, 27:745–749 Keywords: osteomyelitis, Charcot neuroarthropathy, foot ulcers, infection Department of Internal Medicine, ‘Tor Vergata’ University of Rome, Italy. Correspondence to Dr Luigi Uccioli, Department of Internal Medicine, ‘Tor Vergata’ University of Rome, Viale Oxford 81, 00133 Rome, Italy. Tel: + 0039 06 2090 2784; fax: + 0039 06 2090 2804; e-mail: [email protected] Received 12 July 2005 Accepted 10 May 2006

smooth muscle tone and, consequently, it produces a vasodilatory condition in the small arteries and an increase in bone blood flow. The resultant osteolysis, demineralization of bone, can predispose to the development of Charcot osteoarthropathy [4,5]. It is likely that the pathogenesis of Charcot foot is a combined effect of these theories. However, neuropathic osteoarthropathy (acute Charcot foot) is also characterized by uncontrolled inflammation. It has been suggested that the disorder arises by abnormal expression of nuclear transcription factor NF-kB in diabetic neuropathy, associated with the increased release of proinflammatory cytokines, such as tumour necrosis factor alpha and interleukin 1 beta, and this cascade results in increased osteoclastogenesis. Osteoclasts cause progressive bone lysis, leading to further fracture, which in turn potentiates the inflammatory process [6,7]. Clinical features

The Charcot foot is characterized by acute and chronic phases. In acute Charcot neuroarthropathy, the foot is warm, oedematous, markedly erythematous, and with temperature difference > 21C between the affected and non-affected foot and can be associated with a history of traumas even if under-reported [8,9]. There is always some degree of sensory neuropathy in which reflexes,

c 2006 Lippincott Williams & Wilkins 0143-3636

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vibratory sense and proprioception are either diminished or absent. Autonomic neuropathy, which co-exists with somatosensory neuropathy, can be clinically appreciated by the presence of anhydrosis with very dry skin. Pain may or may not be present. The chronic phase is characterized by deformity of the foot with abnormal pressure on the plantar surface due to the collapse of the plantar arch and the development of a rocker bottom deformity. Continued weight-bearing will cause increasing damage with progressive destruction of the foot. The skin overlying new bony prominences is associated with callus formation, which is liable to ulceration especially in the mid-foot. A concomitant ulceration will therefore raise the risk of contiguous osteomyelitis. Charcot deformities have been described for all the foot bones therefore an anatomical classification has been proposed, as given in Table 1. Investigations

The diagnosis of the Charcot foot remains primarily clinical, particularly in the early stages. Plain radiographs are not helpful for ascertaining the presence of osteoarthropathy in a warm, swollen and insensate foot. In advanced stages the typical radiology RX signs are: demineralization, bone destruction and periosteal reaction, ‘pencil and cup’ deformity at the metatarsophalangeal joints or fragmentation of the metatarsal heads, especially in the forefoot Charcot and dislocation or fracture in the mid-foot and atypical calcaneal fractures. Plain films are useful for anatomical information but are neither sensitive nor specific for separating Charcot changes from infection [10,11]. Other investigations for the diagnosis of acute Charcot neuroarthropathy are represented by computerized tomography (CT) and magnetic resonance (MR). An MR scan of a Charcot foot is extremely sensitive, having a 100% detection of abnormalities [12]. Diagnosis of underlying osteomyelitis can be difficult in chronic Charcot osteoarthropathy with foot ulcers. Scintigraphic methods such as combined 111In-leukocyte/bone or leukocyte/marrow scintigraphy are extremely useful in making differential diagnosis between a Charcot joint with and without osteomyelitis [13–15]. Table 1 Type I II III IV V

Anatomical classification* of Charcot deformities Joints involved Metatarso-phalangeal and interphalangeal Tarso-metatarsal Tarsal Sub-talar Calcaneum

* From Sanders LJ, Frykberg RG; Charcot foot in Levin ME, O’Neal LW, Bowker JH. (Eds.): The Diabetic foot. Ed 5. St Lovis Hosby Book, 1993.

In this particular condition MRI seems not to be able to distinguish the marrow oedema associated with Charcot and that associated with osteomyelitis.

Osteomyelitis in the diabetic foot Foot infections are among the most frequent and serious consequences of foot ulceration in diabetic patients and they are responsible for more hospital days than any other complications of diabetes and contribute up to 25–50% of lower-extremity amputations in diabetic patients [16,17]. An estimated 10–30% of diabetic patients with a foot ulcer will eventually require an amputation, approximately 60% of which are preceded by an infected ulcer [18,19]. Although the true prevalence of osteomyelitis in diabetic foot ulcers is not known, studies have shown that it may be present in more than 60% of infected diabetic foot ulcers [20,21]. Osteomyelitis is secondary complication of diabetic foot ulceration. It is a bone infection that induces a progressive destruction of the bone. In the diabetic ulcer the infection develops by spreading from contiguous softtissue to underlying bone. It may involve any bone of the foot, but it mostly affects forefoot bones [22]. Clinical manifestation

The first approach to diagnosis is a clinical evaluation. Skin ulceration with soft-tissue infections that have been present for more than a week or two, especially if they are located over a bony prominence, are at high risk for contiguous bone involvement. The typical signs of local infection are swelling, erythema, warm with or without discharging pus and or fragments of bone [23,24]. Systemic signs, such as fever and malaise, are unusual in any form of foot infection, including chronic osteomyelitis. The larger and deeper the skin ulceration the more likely the underlying bone will be infected [25]. All of the ulcers in which bone exposed, either visibly or by probing, have underlying osteomyelitis with a sensitivity of 66%, a specificity of 89% and a positive predictive value of 89% [26]. Moreover, it has proposed that clinically the ‘sausage toe’ deformity, due to local soft tissue infection and inflammation and underlying bony changes, is highly suggestive of osteomyelitis [27]. Ulcers of long duration are probably more often associated with underlying osteomyelitis as are those which overlie bony prominences. A systemic reaction to osteomyelitis of blood tests, including C-reactive protein (CRP) and white blood cell (WBC) count, are non-specific; in fact, both a neutrophil count and CRP were higher in soft-tissue infections than in osteomyelitis.

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Charcot joint and osteomyelitis Giurato and Uccioli 747

Fig. 1

(a)

Charcot foot Foot with swollen, warm, erythematous skin, with elevated temperature without a history or presence of ulcer in the neuropathic diabetic patient Highly suspected for acute Charcot

Plain X-ray negative

MRI foot

Compatible with Charcot (b)

Charcot Joint with ulcer in mid/hindfoot Clinical suspicion of bone infection

MRI foot

Combined leukocyte/marrow scintigraphy Osteomyelitis of the fore/midfoot

(c)

Ulcer with bone exposed

Ulcer with bone probing

Presumptive osteomyelitis

high probability of osteomyelitis Plain X-ray

Bone culture

positive

negative

Plain X-ray and/or MRI foot In 111WBC scan and /or MRI foot Follow-up with plain X-ray (d)

Ulcer with soft-tissue infection

Plain X-ray highly suspected

Compatible with osteomyelitis

Follow-up with plain X-ray

In 111WBC scan or/and MRI foot

A possible approach to the diagnostic evaluation of different clinical conditions of the diabetic foot.

In contrast, an elevated erythrocyte sedimentation rate (ESR) may be a better marker of bone infection. In fact, studies found a high ESR in 96% of cases in which bone was involved [28].

Imaging studies

Plain films are routinely used in imaging to diagnose osteomyelitis in the diabetic foot but it is difficult to interpret them in the early stages. In fact, bony

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abnormalities related to osteomyelitis are generally not evident on plain films until 10–20 days after infection. Although plain X-ray changes are not pathognomonic for infection a diagnosis of probable osteomyelitis can be made when classic findings are associated with the presence of typical clinical signs [29,30]. Plain films are diagnostically useful to monitor the treatment of osteomyelitis [30]. Radionuclide bone scanning with 99mTc-diphosphonates in three or four phases is certainly more diagnostic than X-ray in early osteomyelitis. The sensitivity is of 86% (68–100%) but its specificity of about 45% (0–79%) is poor because any type of bone disease (including neuropathic osteoarthropathy and healing infections) may be able to increase uptake of isotope [31,32]. A 111In-WBC scan gives better results in the diagnosis of infection with a sensitivity of 89% (45–100%) and specificity of 78% (29–100%). When the infection resolves, this scan normalizes. This makes it a potentially useful tool to follow the response to therapy. The major limitation of this scan in the foot is that the lower resolution may not differentiate infection of the bone from that of adjacent soft tissues [33]. Moreover, the sensitivity can be limited in the presence of peripheral ischaemia. However, a WBC scan or scans using other infectionspecific radiopharmaceuticals combined with a bone scan or 99mTc-labelled sulfur colloid (which localizes to the marrow) can increase specificity and this approach may be evaluable in excluding osteomyelitis when Charcot osteoarthropathy has been complicated by secondary neuropathic ulceration [13,14,34]. Although there have been some promising results in early studies this technique with a double scan is relatively expensive, technically demanding and requires withdrawal and re-infusion of blood [35]. Most specialists agree that MRI offers the greatest diagnostic support in clinical practice to diagnose osteomyelitis in the diabetic foot. In addition, MRI has a better resolution and better sensitivity in differentiating bone infection from soft-tissue infection than do other diagnostic techniques. The diagnostic sensitivity of MRI is 90–100%, while its specificity is 83% [35,36], although this is limited by the fact that the technique is not able to distinguish between marrow oedema, which is characteristic of osteomyelitis, and other conditions that are able to increase bone marrow oedema, such as acute Charcot osteoarthropathy. However, MRI has a positive predictive power of 93% and a negative predictive power of 100% according to a study that compared its results to those of bone biopsy, which is considered the ‘gold

standard’ technique in this context [36]. In addition, in some studies evaluating diabetic patients with a clinical suspicion of osteomyelitis secondary to an infection of soft tissues, MRI performed better in the diagnosis of osteomyelitis than did plain X-ray, bone scans and WBC scans [37,38]. Recent evidence suggests a possible role of positron emission tomography (PET) and PET/CT using 2-[18F]fluoro-2-deoxy-D-glucose (18F-FDG) for the diagnosis of diabetic foot osteomyelitis. 18F-FDG is a non-specific tracer for increased intracellular glucose metabolism and accumulates in sites of infection and inflammation [39,40]. However, bone biopsy remains the ‘gold standard’ in diagnosing osteomyelitis because it has 95% sensitivity and 99% specificity [41]. Moreover, it has the advantage of providing culture and antimicrobial susceptibility results which can guide the choice of antibiotic therapy. A possible approach in the diagnosis of different clinical conditions of the diabetic foot is outlined in Fig. 1.

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Norden CW. Acute and chronic osteomyelitis. Infect Dis 1999; 2:43–48. Wrobel JS, Connolly JE. Making the diagnosis of osteomyelitis. The role of prevalence. J Am Podiatr Med Assoc 1998; 88:337–343. Edelson GW, Armstrong DG, Lavery LA, Caicco G. The acutely infected diabetic foot is not adequately evaluated in an inpatient setting. Arch Intern Med 1996; 18:716–722. Eneroth M, Larsson J, Apelqvist J. Deep foot infections in patients with diabetes and foot ulcer: an entity with different characteristics, treatments, and prognosis. J Diabetes Complications 1999; 13: 254–263. Karchmer AW, Gibbons GW. Foot infections in diabetes: evaluation and management. Curr Clin Top Infect Dis 1994; 14:1–22. Lipsky BA. Osteomyelitis of the foot in diabetic patients. Clin Infect Dis 1997; 25:1318–1326. Rajbhandari SM, Sutton M, Davies C, Ward JD.‘Sausage toe’: a reliable sign of underlying osteomyelitis. Diabet Med 2000; 17:74–77. Grayson M, Gary W, Balogh K, Levin E, Karchmer W. Probing to bone in infected pedal ulcers. JAMA 1995; 273:721–723. Kaleta JL, Fleischli JW, Reilly CH. The diagnosis of osteomyelitis in diabetes using erythrocyte sedimentation rate: a pilot study. J Am Med Assoc 2001; 91:445–450. Mader JT, Calhon H. Staging and staging application in osteomyelitis. Clin Infect Dis 1997; 25:1303–1309. Crim JR, Seeger LL. Imaging evaluation of osteomyelitis. Crit Rev Diag Imag 1994; 35:201–256. Lipsky A. Osteomyelitis of the foot in diabetic patients. Clin Infect Dis 1997; 25:1318–1326.

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Becker W. Imaging osteomyelitis and the diabetic foot. Q J Nucl Med 1999; 43:9–20. Schauwecker DS. The scintigraphic diagnosis of osteomyelitis. AJR Am J Roentgenol 1992; 158:9–18. Tomas MB, Patel M, Marwin SE, Palestro CJ. The diabetic foot. Br J Radiol 2000; 73:443–450. Enderle MD, Coerper S, Schweizer HP, Kopp AE, Thelen MH, Meisner C et al. Correlation of imaging techniques to histopathology in patients with diabetic foot syndrome and clinical suspicion of chronic osteomyelitis. Diab Care 1999; 22:294–299. Levine SE, Neagle CE, Esterhai JL, Wright DG, Dalinka MK. Magnetic resonance imaging for diagnosis of osteomyelitis in the diabetic patient with a foot ulcer. Foot Ankle 1994; 15:151–156. Morrison WB, Schweitz M, Russel K. Osteomyelitis of the foot: relative importance of primary and secondary MR imaging signs. Radiology 1998; 207:625–632. Croll SD, Nicholas GG, Osborne MA, Wasser TE, Jones S. Role of magnetic resonance imaging in the diagnosis of osteomyelitis in diabetic foot infections. J Vasc Surg 1996; 24:266–270. Termaat MF, Raijmakers PG, Sclolten HJ, Patka P, Haarman HJ. The accuracy of diagnosis imaging for the assessment of chronic osteomyelitis: a systemic review and meta-analysis. J Bone Surg Am 2005; 87:2464–2471. Keidar Z, Militianu D, Melamed E, Ora I. The diabetic foot: Initial experience with 18F-FDG PET/CT. J Nucl Med 2005; 3:444–449. Sutton PR, Harley JD, Jacobson AF, Lipsky BA. Diagnosis of osteomyelitis with percutaneous bone biopsy in patients with diabetes and foot infections. J Gen Int Med 2000; 15:S6.

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NEWS AND VIEWS SEPTEMBER 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up-to-date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: + 44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: + 44 (0)113 206 6930, E-mail: [email protected]

Nuclear Medicine Communications, 2006, 27: 751–752 NICE Technology Appraisal Guidance 73

It is sometimes said that there is no money with NICE, but in the case of Technology Appraisal Guidance 73 (Myocardial Perfusion Scintigraphy for the Diagnosis and Management of Angina and Myocardial Infarction) this turns out to be untrue. It is not a very well known fact that the UK government has already started a phased funding stream for its implementation. Some of the money to implement it was paid to acute trusts last year and this year. Readers who are attempting to scale up their myocardial perfusion scanning in response to this guidance may wish to discuss this with their financial teams. Being phased funding the amount so far delivered is quite modest (d2M each year for the whole of England). The total bill for implementing Guidance 73 was estimated at d18M capital and d27M revenue (2003 prices). The money thus far allocated has been passed to trusts via four Payment by Results (PBR) tariffs, E11, E12, E22 and E23 (refer to the PBR technical manual). The authors are not certain if money has also been allocated in other streams (e.g., primary care trusts (PCTs)), but we are certain that the tariffs above do contain specific provision for implementing the guideline. It is possible that failure to access the funding and make a

start at implementation might compromise future instalments of the rollout.

Comments on the content and design of the new site are welcomed. Adverse events/product defects

Sentinel lymph node biopsy for breast cancer

We are now entering the final phase of introduction of this service. The New Start programme has trained a number of existing surgeons using mentored operating on five cases and prospective audit of 25 cases. As SLNB becomes the staging method of choice, it will become increasingly difficult to justify axillary clearance for auditing purposes, therefore surgeons in future will tend to develop their technique by performing 30 SLNBs without axillary clearance under the supervision of a local, New Start trained, mentor. The New Start programme has been an interesting exercise. The level of organization and the rigour applied probably sets new standards for medical technology. British Nuclear Medicine Society web site

The redesign of the BNMS web site continues under the tireless direction of Professor Alan Perkins. Expressions of interest have been received from a good handful of providers. Council and other members are reviewing examples of their past work.

The link for reporting adverse reactions and defective products has been repaired, and the community is invited to report these as before via one of two web sites: BNMS-Sub-SpecialtiesRadiopharmacy Group-Defective Products/Adverse Reactions or UK Radiopharmacy Group-Adverse Events/Product Defects Thanks to Dr Neil Hartman for bringing this to our attention.

Meeting Announcements

BNMS Autumn Meeting Dates: 4–5 September 2006 Venue: Cambridge, England Website: www.bnms.org.uk EANM Annual Meeting Dates: 30 September – 4 October 2006 Venue: Athens, Greece Website: www.eanm.org 9th World Congress of Nuclear Medicine and Biology Dates: 22–27 October 2006 Venue: Seoul, South Korea

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Website: www.wfnmb.org/congress 2006/index02.htm International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM) Dates: 13–15 November 2006 Venue: Vienna, Austria Website: www-pub.iaea.org/MTCD/ Meetings/Announcements.asp? ConfID = 146 Design of Radionuclide Facilities with Reference to Radiation Protection (Revisited) Date: 14 November 2006

Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT Website: www.bir.org.uk Education and Training

EANM Learning Courses Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php E-mail: [email protected]

EANM Distance Learning in Nuclear Cardiology This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. Website: http://www.eanm.org/edu Online/edu_start.php?navId = 332

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Editorial

Practical and clinical benefits of radioimmunotherapy lead to advantages in cost-effectiveness in the treatment of patients with non-Hodgkin’s lymphoma Andreas Ottea and Sally L. Thompsonb Nuclear Medicine Communications 2006, 27:753–756

a

Center of Clinical Trials, University Hospital Freiburg, Germany and bSchering AG, Berlin, Germany.

Introduction The recent development of radioimmunotherapy (RIT) is a significant step forward in the treatment of patients with low-grade and follicular non-Hodgkin’s lymphoma, a malignancy that is well known to be inherently radiosensitive. Highly convincing efficacy [1–3] and safety [4] data are now available to support the benefits of RIT in follicular lymphoma. In addition, RIT offers significant quality of life benefits to patients [1,2] and convenience when compared with older chemotherapy combinations. Despite the advantages of RIT, its routine use in follicular non-Hodgkin’s lymphoma currently poses a number of challenges. These include the need for hospitalization in some regions, practical safety concerns, the need for interdisciplinary cooperation and the need for adequate budgets for RIT therapy at a time when there are increasing numbers of other high-cost oncology products reaching the market. Each challenge to the widespread availability and use of RIT must be dealt with to ensure that all suitable patients have access to this new and innovative therapy.

Radioimmunotherapy Therapy with radioimmunoconjugates has been extensively tested clinically using murine anti-CD20 monoclonal antibodies (MAbs) conjugated to either 131I (131I-tositumomab [Bexxar; Corixa Corporation, Seattle, Washington, and GlaxoSmithKline, Philadelphia]) or 90 Y (90Y-ibritumomab tiuxetan [Zevalin; BiogenIdec, San Diego, USA and Schering AG, Berlin, Germany]) [5]. Studies employing one or the other radioimmunoconjugate have demonstrated significant therapeutic benefit for patients with follicular non-Hodgkin’s lymphoma and both therapies are considered to be similarly efficacious in this indication. The relative merits of both radioimmunoconjugates with respect to practical issues, including radiation exposure risk and other parameters that could affect a patient’s quality of life as well as overall costs to the health care system are important differentiators of treatment choice.

Correspondence to Professor Dr Andreas Otte, Center of Clinical Trials, University Hospital Freiburg, Elsa¨sser Str. 2, D-79110 Freiburg, Germany. Tel: + 0049 (0)761 270 7211; fax: + 0049 (0)761 270 7373; e-mail: [email protected] Received 12 April 2006 Accepted 12 April 2006

Hospitalization and time commitments for medical staff and patients The radionuclide 90Y does not produce penetrating gamma radiation, and patient isolation and lead shielding are therefore unnecessary. Accordingly, hospital-based outpatient therapy is feasible with 90Y and no isolation from family or friends is required. In contrast, the 131Ilabelled antibody requires inpatient hospitalization due to the inherent risk of exposure from gamma emissions, and patients and families need to follow detailed instructions to prevent undue exposure. A further advantage of the 90Y product and an issue that is of relevance to both patients and medical staff is the need for dosimetry to calculate effective therapeutic doses with the 131I-labelled antibody, which is not required for 90 Y. Finally, from a cost and resource point of view, the provision of hospital beds under radiation protection for inpatient radioimmunotherapy using radionuclides with a gamma component such as 131I would be a major challenge if the product was to become more widely used.

Practical safety Eschner et al. undertook an interesting radioecological calculation on radioactive excretions of patients assuming treatment with 90Y-labelled ibritumomab tiuxetan on an outpatient basis [6], indicating that, apart from 90 Y-ibritumomab tiuxetan, there is room for many more radioimmunotherapies using 90Y as the radionuclide without the necessity for implementing any inpatient infrastructure. Indeed, according to the very recent recommendation of the German Radiation Protection Commission, in 2005, an outpatient treatment with 90 Y-ibritumomab tiuxetan in Germany in the approved dosage is permissible in institutions which fulfil the necessary requirements for handling open radioactive substances [7]. The only reasons for indicating hospitalization of patients treated with 90Y-labelled ibritumomab tiuxetan would be if there was a need to monitor co-morbidities, or if there were complications arising from the treatment or

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if there was a special need for radiation protection such as small children or pregnancy in the patient member environment. Only for the latter situation would hospitalization to a nuclear medicine ward be indicated. The debate on whether the use of radioimmunotherapy is associated with an increased risk of developing treatment-related myelodysplastic syndrome (tMDS) and acute myelogenous leukaemia (AML) has sometimes led to concerns by haemato-oncologists in recommending new radioimmunotherapies to patients. However, based on new long-term data available on the development of tMDS/AML after radioimmunotherapy [8], which show no increased risk, this reservation is likely to change.

Interdisciplinary cooperation The 90Y-ibritumomab tiuxetan regimen is typically administered on an outpatient basis over approximately 1 week. The delivery of 90Y-ibritumomab tiuxetan requires the establishment of new multidisciplinary teams in many hospitals. Such teams are likely to include a haemato-oncologist, nuclear medicine physician or radiation oncologist, nuclear pharmacist, and speciality nurses. The relatively short physical half-life of 90Y (64 h) makes it essential that ibritumomab tiuxetan is radiolabelled at a radiopharmacy or similar immediately prior to use [9]. Although most information about the 90Y-ibritumomab tiuxetan therapy will have been provided by the haematooncologist, the nuclear medicine physician, who is responsible for the administration of 90Y-ibritumomab tiuxetan, should provide patients with additional information on radiation therapy and written information describing 90Y-ibritumomab tiuxetan treatment, safety precautions within the week following therapy, anticipated adverse events, and contact telephone numbers [10]. Selection of patients suitable for treatment with 90Yibritumomab tiuxetan typically remains with the haemato-oncologist and is an important step in optimizing the benefits of RIT for patients. In some countries such as Germany, where office-based haemato-oncologists are the norm, there may be concerns about ‘losing’ patients to another speciality with a consequent loss of income by the original haemato-oncologist. This can sometimes lead to haemato-oncologists trying all other therapies at his or her disposal first and only when these options have failed referring patients for treatment with 90Y-ibritumomab tiuxetan. Reassurance that the patient who has received therapy with 90Y-ibritumomab tiuxetan will be directed back to their referring haemato-oncologist should help to address this concern. Such reassurance should also encourage the referral of more patients who have received fewer prior treatments and in doing so maximize the

significant therapeutic benefits that tiuxetan is able to provide.

90

Y-ibritumomab

Reimbursement and cost-effectiveness The European reimbursement situation in relation to the approved use of radioimmunotherapies needs further improvement. In this context it is unacceptable that an approved and efficacious radioimmunotherapy in follicular non-Hodgkin’s lymphoma is sometimes reimbursed at only a small fraction of the treatment cost, as is currently the case in systems awaiting a specific RIT DRG code, such as Germany and Italy. Growing pressures on health care budgets worldwide have led to an increasing interest in the use of health economics data to support the added value of new therapies in terms of both outcomes and cost and such data is available for 90Y-ibritumomab tiuxetan demonstrating cost-effectiveness relative to rituximab. Gabriel et al. [11] compared the cost-effectiveness of 90Yibritumomab tiuxetan versus rituximab (4-dose scheme) for outpatient treatment in Germany (based on a price year of 2004) in patients with relapsed or refractory follicular NHL. In this analysis drug acquisition costs in addition to physician fees for drug application and resource utilization due to adverse events data were considered. Cost-effectiveness was determined as cost per year in remission by relating costs to the overall response rate and duration of response; cost per diseasefree year was based on complete response rate and duration of response of complete response patients. The conclusion of the analysis was that although the total cost of the 90Y-ibritumomab tiuxetan regimen was higher (h19 567 vs. h9756), the cost per year in remission (h14 862 vs. h16 967) and in particular cost per diseasefree year (h22 235 vs. h80 077) were clearly in favour of 90 Y-ibritumomab tiuxetan. This conclusion was driven largely by the superior response rates and in particular complete response rates for 90Y-ibritumomab tiuxetan over rituximab monotherapy. Also in a recent Dutch study, Thompson and van Agthoven estimated the incremental cost-effectiveness of 90Y-ibritumomab tiuxetan compared with rituximab based on either a 4-dose or an 8-dose scheme [12]. The mean total costs were estimated as follows: 90Y-ibritumomab tiuxetan h16 345, rituximab 4-dose scheme h9510 and rituximab 8-dose scheme h19 020. The expected number of months in remission per patient treated were 14.4 months for 90 Y-ibritumomab tiuxetan, 11.4 months for the rituximab 8-dose scheme and 6.2 months for the rituximab 4-dose scheme, resulting in a mean cost per month in remission for 90Y-ibritumomab tiuxetan of h1138, followed by h1544 for the rituximab 4-dose scheme and h1674 for the rituximab 8-dose scheme. The price year of this Dutch study for all resources valued, except the 90Y-ibritumomab tiuxetan product, was 2001 and, unlike for Germany,

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Editorial Otte and Thompson 755

value added tax does not need to be added to the wholesale price of the product in the Netherlands, thus the lower overall treatment cost in this market. Since the total cost of a 90Y-ibritumomab tiuxetan treatment is derived mainly from the cost of the therapy itself (rituximab pre-dosing, ibritumomab tiuxetan and 90 Y) and the product price is not subject to major differences across the world (except the USA), the cost of a treatment with 90Y-ibritumomab tiuxetan hardly varies from country to country. In contrast, the cost of therapies that 90Y-ibritumomab tiuxetan could potentially replace may vary more, so that the relative cost-effectiveness of 90 Y-ibritumomab tiuxetan could be more favourable in countries with higher wage rates for health professionals. For example, if we compare a high-income country such as Germany with a European country that is associated with a lower per capita national income such as Bulgaria, and compare the cost of 90Y-ibritumomab tiuxetan and CHOP (cyclophosphamide, adriamycin, vincristine, prednisone) in each environment, we would clearly see differences. In this example, CHOP in Germany is reasonably expensive for such an old chemotherapy (around h10 000), whereas in Bulgaria it may be about half this figure. This is because the cost of hospital and outpatient visits and health professional time required to provide all chemotherapy infusions is relatively low in Bulgaria by comparison. In addition, there are cheap generics manufactured by eastern and often state-run companies, which further reduce costs when compared to Germany. Although there are various treatment alternatives in follicular non-Hodgkin’s lymphoma, only a few studies have so far focused on their costs [13]; these include Sweetenham et al. [14], Herold et al. [15] and van Agthoven et al. [16].

h4218; chlorambucil: h2476). In this study only the mean per patient cost was assessed. The relative costeffectiveness of each therapy was not addressed. The provided data are, however, important for future costeffectiveness calculations in the context of new treatment options such as combined chemo-immunotherapy or combined chemo-radioimmunotherapy as initial therapy options.

Conclusion New clinical technologies such as radioimmunotherapy need to be further integrated into the management pathway of patients with follicular non-Hodgkin’s lymphoma. RIT and 90Y-ibritumomab tiuxetan, in particular, offer significant advantages to both the haemato-oncologist and the patient including long response durations, a favourable safety profile and the convenience of a treatment that (local radioprotection laws permitting) can be administered on an outpatient basis over two single clinic visits and within 1 week. In addition, for patients, this benefit comes without the alopoecia, mucositis, or severe nausea or vomiting often accompanying conventional chemotherapy. Furthermore, the costeffectiveness data for 90Y-ibritumomab tiuxetan reviewed in this editorial provide convincing evidence in favour of the added value of 90Y-ibritumomab tiuxetan in terms of cost per month in remission or cost per disease-free month despite higher initial product acquisition costs. Good collaboration between the haemato-oncologist and nuclear medicine physician and the referral of patients suitable for treatment are likely to maximize the outcomes RIT can provide for the patient with follicular non-Hodgkin’s lymphoma. Financial disclosure

A.O. receives a consultancy grant from Schering AG, Berlin, Germany.

References The most reliable source of data to date is from a patient level costing study by van Agthoven et al. [16]. In this study, direct health care costs associated with the most commonly prescribed treatments for indolent follicular NHL in the Netherlands were assessed. The treatments evaluated included allogeneic and autologous stem cell transplantation, chlorambucil, CVP, CHOP, fludarabine, radiotherapy, rituximab, and interferon-a maintenance treatment. The authors reported that in relation to costs only, allogeneic and autologous stem cell transplantation were the most expensive treatments identified (mean per patient overall cost impact until first discharge: h45 326 and h18 866, respectively, including the costs of the initial procedure and up to 10 days post-procedure only), compared to fludarabine costing h10 651, rituximab (4dose scheme) costing h10 628 and CHOP costing h7547. By contrast, classical NHL treatments were found to be the least expensive therapies (CVP: h5268; radiotherapy:

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Witzig TE, Gordon LI, Cabanillas F, Czuczman MS, Emmanouilides C, Joyce R, et al. Randomized controlled trial of yttrium-90-labeled ibritumomab tiuxetan radioimmunotherapy versus rituximab immunotherapy for patients with relapsed or refractory low-grade, follicular, or transformed B-cell nonHodgkin’s lymphoma. J Clin Oncol 2002; 20:2453–2463. Witzig TE, Flinn IW, Gordon LI, Emmanouilides C, Czuczman MS, Saleh MN, et al. Treatment with ibritumomab tiuxetan radioimmunotherapy in patients with rituximab-refractory follicular non-Hodgkin’s lymphoma. J Clin Oncol 2002; 20:3262–3269. Gordon LI, Witzig T, Molina A, Czuczman M, Emmanouilides C, Joyce R, et al. Yttrium 90-labeled ibritumomab tiuxetan radioimmunotherapy produces high response rates and durable remissions in patients with previously treated B-cell lymphoma. Clin Lymphoma 2004; 5:98–101. Witzig TE, White CA, Gordon LI, Wiseman GA, Emmanouilides C, Murray JL, et al. Safety of yttrium-90 ibritumomab tiuxetan radioimmunotherapy for relapsed low-grade, follicular, or transformed non-Hodgkin’s lymphoma. J Clin Oncol 2003; 21:1263–1270. Silverman DH, Delpassand ES, Torabi F, Goy A, McLaughlin P, Murray JL. Radiolabeled antibody therapy in non-Hodgkin’s lymphoma: radiation protection, isotope comparisons and quality of life issues. Cancer Treat Rev 2004; 30:165–172. Eschner W, Breustedt B, Lassmann M, Ha¨nscheid H. Erfassung der u¨ber Ausscheidungen in die Umwelt abgegebenen radioaktiven Stoffe nach ihrer Anwendung in der Nuklearmedizin. Schriftenreihe Reaktorsicherheit und

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Strahlenschutz, Bundesministerium fu¨r Umwelt, Naturschutz und Reaktorsicherheit. BMU – 2004-649. 7 Strahlenschutzkommission. Radioimmuntherapie mit Y-90-IbritumomabTiuxetan (Y-90-Zevalin), Empfehlung der Strahlenschutzkommission, 198. Sitzung, 17.02.2005. 8 Otte A, Dierckx RA. Myelosuppressive side-effects in radioimmunotherapy of non-Hodgkin’s lymphoma: Is there an increased risk? Nucl Med Commun 2005; 26:1045–1047. 9 Gordon LI. Practical considerations and radiation safety in radioimmunotherapy with yttrium 90 ibritumomab tiuxetan (Zevalin). Semin Oncol 2003; 30(6, suppl 17):23–28. 10 Hagenbeek A, Lewington V. Report of a European consensus workshop to develop recommendations for the optimal use of 90Y-ibritumomab tiuxetan (Zevalin) in lymphoma. Ann Oncol 2005; 16:786–792. 11 Gabriel A, Ha¨nel M, Wehmeyer J, Griesinger F. Advantages in cost effectiveness of Zevalin radioimmunotherapy vs. rituximab immunotherapy in patients with relapsed or refractory follicular non-Hodgkin’s lymphoma [Abstract]. Onkologie 2005; 28(suppl 3):236.

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Thompson S, van Agthoven M. Cost-effectiveness of 90Y-ibritumomab tiuxetan (90Y-Zevalin) versus rituximab monotherapy in patients with relapsed follicular lymphoma [Abstract]. Blood 2005; 106:436. Van Agthoven M, Uyl-de Groot CA, Sonneveld P, Hagenbeek A. Economic assessment in the management of non-Hodgkin’s lymphoma. Expert Opin Pharmacother 2004; 5:2529–2548. Sweetenham J, Hieke K, Kerrigan M, Howard P, Smartt PF, McIntyre AM, et al. Cost-minimization analysis of CHOP, fludarabine and rituximab for the treatment of relapsed indolent B-cell non-Hodgkin’s lymphoma in U.K. Br J Haematol 1999; 106:47–54. Herold M, Sacchi S, Hieke K. The cost of treating relapsed indolent non-Hodgkin’s lymphoma in an international setting: retrospective analysis of resource use. Haematologica 2002; 87:719–729; discussion, 729. van Agthoven M, Kramer MHH, Sonneveld P, Van der Hem KG, Huijgens PC, Wijermans PW, et al. Cost analysis of common treatment options for indolent follicular non-Hodgkin’s lymphoma. Haematologica 2005; 90:1422–1432.

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Original article

Nuclear medicine imaging of diabetic foot infection: results of meta-analysis Gabriela Capriottia, Marco Chianellib and Alberto Signorea Background and aim Osteomyelitis of the foot is the most commonly encountered complication in diabetic patients. Nuclear medicine techniques are usually complementary to radiology in the diagnosis of foot infections; they play an important role in various clinical situations. The aim of this study was to develop a practical guideline to describe the radiopharmaceuticals to be used for different clinical conditions and different aims in diabetic foot infection.

Results and conclusion We provide a guideline to assist in the selection of the optimal radiopharmaceuticals for different clinical conditions and different aims. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:757–764
Nuclear Medicine Communications 2006, 27:757–764 Keywords: diabetic foot, nuclear medicine, osteomyelitis, radiopharmaceuticals a

Nuclear Medicine, II Faculty of Medicine, University of Rome ‘La Sapienza’ and Nuclear Medicine, Regina Apostolorum Hospital, Albano, Rome, Italy.

Methods In this study, we reviewed 57 papers (published between 1982 and 2004; 50 original papers and seven reviews) that described the imaging of the diabetic foot and examined a total of 2889 lesions. We performed data analysis to establish which imaging technique could be used as a ‘gold standard’ to diagnose infection, evaluate the extent of disease and monitor the efficacy of therapy.

b

Introduction

the forefoot often develop motor, sensory and autonomic neuropathy. Motor neuropathy results first in weakness, atrophy and paresis of the muscles in the foot, followed by callus and ulcer formation on the plantar surface of the metatarsal heads [8]. Sensory neuropathy is initially painful, but then leads to a total loss of sensation. The impairment of sensation results in the foot being subjected to repeated damage and abnormal pressure distribution. Insensitive trauma leads to bony deformities and callus formation. Finally, autonomic neuropathy results in warm, dry skin, due to a lack of sweating, and, potentially, in fissure and crack formation [9]. Pressure ischaemia from callosities and microvascular damage prevents the normal response to wound healing. The resulting foot ulcers constitute a portal of entry for infection with diffusion directly to the bone surface. Most patients lack clinical signs and symptoms of infection, and the diagnosis of osteomyelitis is often overlooked. Consequently, imaging is crucial to the evaluation of these patients.

More than 190 million people in the world have diabetes mellitus and, each year, another one million adults are diagnosed with diabetes. With time, long-term complications of diabetes can occur, including vascular disease, neuropathy, retinopathy and nephropathy. The combination of peripheral neuropathy and vascular disease frequently leads to foot complications and lower limb amputations. Complications of the foot are the most common cause of non-traumatic lower extremity amputations in the industrialized world. The risk of amputation is 15–46 times higher in diabetic patients than in non-diabetic people [1,2], and 30% of diabetic patients have contralateral limb amputation within 3 years of the first amputation [3–5]. Foot ulcers are a major predictor of future lower extremity amputation in patients with diabetes. Overall, 15% of diabetic patients will develop a foot ulcer during their lifetime, and about 15–25% of these patients will require an amputation [6,7]. It is not surprising that diabetes is the leading cause of nontraumatic lower extremity amputation. Mortality is closely related to amputation, and diabetic patients with foot ulcers have a higher mortality than others [6]. The plantar surface of the forefoot is the most common location for foot ulcers. The aetiology of these ulcers is closely associated with peripheral neuropathy and vascular insufficiency. Patients with ulcers on the plantar surface of

Correspondence to Alberto Signore, MD, Nuclear Medicine, Sant’Andrea Hospital, II Faculty of Medicine, University ‘La Sapienza’, Via di Grottarossa 1035/ 1039, 00189 Rome, Italy. Tel: + 39-06-33274625; fax: + 39-06-33274621; e-mail: [email protected] Received 18 July 2005 Accepted 15 May 2006

Approximately 2.5% of diabetic patients have Charcot’s joint, usually involving the tarsal or tarsometatarsal joint [10]. This is a progressive degeneration of the musculoskeletal system characterized by joint dislocations, pathological fractures and debilitating deformities. The disorder results in the destruction of bone and soft tissue at weight-bearing joints and, in its most severe form, may cause significant damage to the bony architecture. Charcot’s joint is an underlying cause of neurological and vascular disease. Trauma is a precipitating factor to

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the neuropathy, which involves motor, sensory and autonomic fibres. Patients with advanced neuropathy lose sensation in the foot, and repetitive stress leads to bone and joint disruptions, deformities and joint instability, which cause joint degeneration, subluxation and destruction. Bone and joint destruction, incomplete healing and partial repair result in a deformed foot. Clinically, Charcot’s foot presents swelling, crepitus, palpable loose bodies and osteophytes. Tissue ischaemia and bony prominences may lead to plantar ulceration. In this foot, osteomyelitis may be indistinguishable from Charcot’s joint, and often may occur simultaneously. Infection is the most serious risk of the diabetic foot. Diabetic patients are predisposed to foot infections because of a compromised vascular supply secondary to diabetes. Local trauma and/or pressure, in addition to microvascular damage, may result in diabetic foot infection. Despite much effort directed towards amputation prevention, the incidence of lower extremity amputation in diabetic populations continues to increase. Prompt diagnosis and treatment of infection are crucial to avoiding amputation. However, the detection of diabetic foot infection can be difficult. Information obtained from physical examination and sequential imaging methods is key to identifying osteomyelitis at its initial stages. The aim of this study was to develop a practical guideline to describe the radiopharmaceutical to be used in each clinical condition and for different aims in diabetic foot infection.

The role of nuclear medicine Although bone biopsy is the definitive test for the diagnosis of both pathological and microbiological osteomyelitis, it is not always performed in patients with vascular problems. In part, this is because it is an invasive procedure, but, more importantly, because biopsy loses its reliability when the biopsy fragment is contaminated by cutaneous bacteria [11]. An early diagnosis of osteomyelitis is necessary to start antibiotic treatment in conjunction with conservative surgery. Because of the lack of signs and symptoms associated with bone infection in the diabetic foot, however, physicians often have to rely on imaging to make the diagnosis. Several techniques are available for imaging infection, and they are intended to allow specific, stage-related diagnosis. However, choosing the appropriate technique is often confusing, as factors such as previous therapy, the presence of a concomitant neuropathic joint, the time of infection and the cost of the study need to be considered. The choice of any test must always be based on the clinical presentation, the site of infection and the activity phase of the disease.

that usually requires several weeks. Prior to the development of these lesions, plain radiography may be negative. This radiographic pattern can also be seen in other diabetic conditions, such as fractures and joint deformities, which coexist in the diabetic foot. Although the accuracy of plain radiography for the early diagnosis of osteomyelitis is low, it is still considered a useful screening method to investigate the diabetic foot. Plain radiography provides anatomical information necessary for analysing any additional studies [12]. Magnetic resonance imaging (MRI) provides an excellent means for differentiation between osteomyelitis and soft tissue infection, and has recently become widely available. The typical MRI pattern of osteomyelitis is marrow replacement by oedema and infiltration, but this is not specific for bone infection. MRI specificity is enhanced by other signs, such as sequestrum formation, cortical interruption, sinus tracts and soft tissue ulceration. In the forefoot, MRI identifies osteomyelitis with a high grade of sensitivity and specificity, and makes it possible to distinguish osteomyelitis from soft tissue infection. MRI also allows the accurate delineation of the extent of infection, providing useful information for surgical planning. Bony destruction, dislocation, marrow oedema, synovial effusion and the loss of bone and joint limits characterize the neuropathic joint. These alterations may also be seen in osteomyelitis, reducing the specificity of MRI [13]. It is not always possible to distinguish the marrow oedema of neuropathy from that of osteomyelitis through MRI. From a clinical point of view, the differential diagnosis between these two conditions is important because they have a different evolution and, especially, because they each require a different therapeutic approach [14,15]. Therefore, the role of MRI in the evaluation of the neuropathic joint is still uncertain. MRI is a useful tool to exclude osteomyelitis in Charcot’s joint, when there are no marrow signal alterations [16], and it provides excellent anatomical details for surgical treatment [14]. Nuclear medicine imaging plays a key role in the diagnosis of osteomyelitis, and represents the procedure of choice for monitoring response to medical therapy. Imaging techniques include bone scintigraphy and infectious/inflammatory imaging using various agents, such as radiolabelled white blood cells (WBCs), monoclonal antibodies and their fragments, human immunoglobulin G and nanocolloid. These radiopharmaceuticals are commercially available, but they cannot be used interchangeably to study diabetic foot disorders.

Meta-analysis Demineralization, periosteal reaction and bony destruction – the typical radiographic triad of osteomyelitis – appear when 30–50% of the bone is destroyed, a process

Methods

Practice guidelines are increasingly being used as a means of reducing inappropriate care and diagnosis, and of

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Nuclear medicine imaging of diabetic foot infection Capriotti et al. 759

making more effective use of health care resources. The methods by which these guidelines are developed, as well as their purpose, dissemination, audience and ultimate use, vary considerably. There are several approaches to guideline development, such as informal and formal consensus and evidence-based and explicit development. We used the evidence-based approach, which, although rightly credited with enhancing the scientific rigor of guideline development, is often unable to produce recommendations in the absence of credible evidence. The approach included a formal assessment of scientific evidence, panel meetings and open discussion sessions that sought input from a broad constituency on relevant issues. Our goal was to establish which radiopharmaceutical could be used to diagnose infection, evaluate the extent of the disease and monitor the efficacy of therapy. To reach this goal, we used a logical organizational process (Fig. 1). The first step was to identify the diagnostic technologies that have the potential to improve the diagnosis of foot infection. Several studies have demonstrated that many radiological and nuclear medicine procedures are involved in the diagnosis of osteomyelitis, but their role is not clear. The second step was to select commercially available radiopharmaceuticals that could be used for the diagnosis of foot infection and those that could be used to monitor treatment. The principal analytical task in developing guidelines is to define which diagnostic

practices produce the best clinical outcomes. The two principal sources of information about clinical benefits are scientific evidence and expert opinion. Scientific evidence may include published and unpublished information. We performed a meta-analysis of peerreviewed articles describing the use of radiopharmaceuticals for the study of diabetic foot infections. Therefore, we conducted MEDLINE searches to capture all studies addressing the utility of different diagnostic procedures for the diagnosis of foot infection. We performed our study using different levels of search. First, we used as keywords ‘diabetic foot infection’ and ‘imaging’ to obtain papers in which different imaging modalities were described and compared. Second, we searched for ‘diabetic foot infection’ and ‘radionuclide’ and/or ‘Technetium, Indium, Iodine’. We selected the main paper regarding the diagnosis of osteomyelitis in the diabetic foot and then analysed the related articles. Case reports, letters to the editor and editorials were excluded from this study, but we considered original papers and reviews describing the sensitivity, specificity and accuracy of different imaging modalities. We reviewed and analysed 57 papers concerning the imaging of the diabetic foot, 50 original papers and seven reviews [17–73], with a total of 2889 lesions (Table 1). For each paper, the number of lesions, sensitivity, specificity, accuracy and positive and negative predictive values were recorded. In many papers, these values were not described, but we calculated the values using the available data. The data were then tabulated, expressed

Fig. 1

Assess clinical benefits

Identify the specific topics

Step 1

Assess clinical appropriateness

Step 2

Assess scientific evidence

Assess expert opinion

Determine appropriateness

Step 3

Develop a practical guideline

Step 4

Process of choosing the criteria for the development of a practical guideline.

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760

Nuclear Medicine Communications 2006, Vol 27 No 10

Table 1

Comparison of imaging modalities in diabetic foot infection: meta-analysis result Lesions

99m

Tc-MDP In-WBC Tc-WBC 111 In/99mTcWBC 99m Tc/111In-HIG 99m Tc-MoAb 99m Tc-Fab MRI Radiography Total 111

99m

Sensitivity

Specificity

Lesions

Accuracy

Lesions

PPV

NPV

719 463 283 205

90.3 86 85.8 80.7

46.4 74.4 84.5 88.5

640 428 283 147

65 77 85.9 84.6

643 398 283 53

65.1 72.4 89.9 NA

71.1 82.6 80.8 NA

97 63 151 364 544 2889

96.8 95.8 91.3 90.1 61.9

66.5 70.2 62 73.9 67.5

97 63 151 223 486 2518

83.2 84.2 81.9 80.5 65.6

97 63 151 128 332 2148

72 76.1 81.4 87.3 71.3

87.7 96.4 79.4 85.7 58.5

Fab, antibody fragment; HIG, human immunoglobulin G; MDP, methylene diphosphonate; MoAb, monoclonal antibodies against granulocyte antigens; MRI, magnetic resonance imaging; NA, data not available; NPV, negative predictive value; PPV, positive predictive value; WBC, white blood cell. All parameters are weighted for the number of lesions in each study. Numbers in bold refer to the best performing imaging tool.

as percentage values and presented as mean values. For this report, a quantitative meta-analysis was performed to reduce the total variability. A weighted value was considered for analysis. All parameters were weighted for the number of lesions according to the following formula: Xw = X/(Y/Z), where Xw is the weighted value of the analysed parameter, X is the value of the analysed parameter, Y is the number of lesions corresponding to X and Z is the number of total lesions. A qualitative revision of each paper was also performed to explain the differences between the diagnosis of osteomyelitis of the forefoot and that of the mid- and hindfoot. In many articles, nuclear medicine techniques were compared with radiological imaging. Only two articles described the use of ultrasound and colloid imaging in the differential diagnosis between osteomyelitis and soft tissue infection or marrow activity, respectively. As a result of the scarcity of published data, ultrasound and colloid imaging accuracy was not included in the analysis. We reviewed the literature and analysed the data. We then created a tabulated list, which was evaluated for appropriateness by an expert physician, and then developed a practical guideline. Results

The results are summarized in Table 1. Disorders of the forefoot are different from those of the mid- and hindfoot and require a different diagnostic approach. Data and literature analysis show that a three-phase bone scan is sensitive but not specific. Focal hyperperfusion, hyperaemia and bony uptake on delayed images are often considered confirmation of osteomyelitis on a bone scan. However, these signs may also be seen in other pathological conditions, such as fracture, neuropathic joint and chronic soft tissue infection, reducing the specificity of this method. Numerous tests to improve the specificity of the technique have been made. The addition of 24-h images, the four-phase bone scan, seems

to enhance the specificity of the technique. In contrast with normal bone, tracer uptake in woven bone continues for a longer period of time, resulting in higher lesion to background uptake on four-phase than third-phase bone scan [69]. However, although a positive test is not diagnostic for osteomyelitis, a negative image excludes infection with a high grade of certainty (good negative predictive value of bone scan). For these reasons, most nuclear physicians consider the bone scan to be a useful screening test to exclude bone infection. Extensive bony changes characterize Charcot’s joint, and therefore three-phase bone scintigraphy is often positive, even in the absence of infection. For this reason, the specificity and diagnostic accuracy of the bone scan are reduced, as for many radiological techniques. Numerous attempts have been made to enhance the specificity of the test, including dual tracer imaging, but the results have been variable [74]. In this part of the foot, radionuclide bone imaging is always positive and therefore not useful for diagnosing osteomyelitis of Charcot’s joint. In contrast with polyphosphate, radiolabelled WBCs are not taken up in healthy bone, and the presence of labelled leucocytes is associated with osteomyelitis. Scintigraphy with radiolabelled WBCs plays a key role in the diagnosis of bone infection in the diabetic foot. The mean values of sensitivity, specificity and accuracy in the detection of osteomyelitis of 99mTc- and 111In-WBC scintigraphy are reported in Table 1. Data analysis reveals that the specificity and diagnostic accuracy of 99mTc-WBC scintigraphy are higher than those of 111In-WBC scintigraphy. These results may be related to the superior physical characteristics of the 99mTc label, which shows better spatial resolution and anatomical landmarks than 111 In [75–77]. Labelled leucocyte scintigraphy can be performed in conjunction with bone scan, but this dual tracer study is

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Nuclear medicine imaging of diabetic foot infection Capriotti et al. 761

not associated with improved diagnostic accuracy [78]. The anatomical information provided by bone scan is not necessary for a better localization of radiolabelled WBC uptake in the forefoot, a region characterized by a simple anatomy compared with the mid- or hindfoot. Identifying an infected region in the forefoot is not difficult, because it is generally related to a foot ulcer. In the same region, leucocyte scintigraphy may be a useful tool for monitoring the response to medical treatment [79]. In the uninfected mid- or hindfoot, leucocyte accumulation is attributed to inflammation, fractures and reparative processes, integrating part of Charcot’s joint. The inflammatory response associated with fractures is polymorphonuclear only in its early phase, and labelled leucocyte uptake seems to be related to haematopoietically active marrow [15,80,81]. Bone marrow development is strictly involved in fracture repair, and fractures are an integral part of the neuropathic joint [15,81]. Aberrant or atypically located marrow activity results from new bone formation. From an imaging point of view, the presence of marrow decreases the specificity of WBC scintigraphy in the mid- or hindfoot, as in other parts of the skeleton [12,80]. Distinguishing osteomyelitis from

neuropathic osteoarthropathy is a common and difficult clinical problem with no highly accurate discriminatory investigation [12]. Available data suggest that leucocyte and bone marrow scintigraphies are useful for determining whether infection is present in Charcot’s joint [15]. Both leucocytes and colloids accumulate in bone marrow, but their behaviour is opposite at sites of infection. An image in which the spatial distribution of both tracers is similar indicates an absence of osteomyelitis, but, if their distribution is not congruent, the uptake of radiolabelled WBCs is related to infection [15,81]. Other radiopharmaceuticals have been proposed to identify osteomyelitis in the diabetic foot, such as 99m Tc-/111In-human immunoglobulin G (99mTc-/111InHIG), anti-granulocyte antigen monoclonal antibodies and their fragments radiolabelled with 99mTc. 99m

Tc-/111In-HIG uptake in inflamed tissue is not specific and is primarily related to vascular permeability, entrapment in the inflamed area and binding to extracellular matrix protein. For this reason, these radiopharmaceuticals have a high value of sensitivity, but they

Fig. 2

Forefoot Clinical exam

High probability of osteomyelitis

Ulcer or soft tissue infection

Medium-low probability of osteomyelitis

Osteomyelitis

Plain X-ray

Plain X-ray

Negative

Bone scintigraphy Treatment MRI

Medical Treatment

Negative

Positive

WBC scintigraphy Medical or surgical treatment

WBC scintigraphy −

+

Medical treatment curettage of the ulcer MRI

Bone biopsy

Diagnostic flow-chart of osteomyelitis in the forefoot. MRI, magnetic resonance imaging; WBC, white blood cell.

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762 Nuclear Medicine Communications 2006, Vol 27 No 10

have less specificity than radiolabelled WBCs in osteomyelitis diagnosis. In a study performed on diabetic patients with foot ulcers and neuropathic joints, it was demonstrated that HIG scintigraphy evaluation was unable to distinguish between osteomyelitis and aseptic inflammation and soft tissue infection, when compared with radiolabelled WBC scan [82]. Moreover, HIG accumulation was described in uninfected fractures of Charcot’s joint, limiting its use in the study of this region [83]. However, as with a three-phase bone scan, a negative test can exclude the diagnosis of osteomyelitis with a high grade of certainty, but a positive test does not confirm the presence of infection.

negative result excludes infection, a positive test does not confirm osteomyelitis. Radiolabelled WBC scintigraphy is necessary. This scan is useful for the diagnosis and evaluation of the extent of osteomyelitis, as well as during the follow-up of medical treatment. In the mid- and hindfoot, the diagnosis of a neuropathic joint with infection is problematic (Fig. 3). Radiolabelled WBC imaging is probably the most accurate test for determining the presence of infection. Although a negative result strongly indicates the absence of osteomyelitis, it is important to note that a positive result requires a complementary study with a marrow agent for confirmation.

The role of radiolabelled antibodies in diabetic foot infection is similar to that of HIG and, like this radiopharmaceutical, they can be used as a screening test.

Acknowledgements

The diagnosis of osteomyelitis is difficult, and the studies performed must be guided by the clinical presentation.

This work is part of a large multicentre study conducted by the Italian Study Group on Inflammation–Infection Imaging of the Italian Society of Nuclear Medicine (AIMN) coordinated by Dr Alberto Signore.

In the forefoot (Fig. 2), when the clinical suspicion of osteomyelitis is low and medical treatment is contemplated, a three-phase bone scan is the study of choice. The bone scan is sensitive but not specific as, although a

Members of the group include: Marco Agnolucci, Alessio Annovazzi, Giorgio Ascoli, Carla Augeri, Bruno Bagni, Marilena Bello`, Sergio Bissoli, Nicola Boccuni, Sergio

Clinical indication

Fig. 3

Midfoot

Clinical examination Clinical suspicion of infection without ulcer

Presence of ulcer or soft tissue infection X-ray Mid-low probability of osteomyelitis High probability of osteomyelitis

Compatible with osteomyelitis

Plain X-ray

Plain X-ray

Negative

MRI

WBC scintigraphy

Therapy MRI

Medical treatment

Neuropathic joint

No Neuropathic joint

WBC scintigraphy

WBC scintigraphy Stop

−

+

Stop Medical or surgical treatment

WBC scintigraphy

− +

Medical therapy curettage of ulcer

Marrow scintigraphy

MRI

Bone biopsy

Complementary study No osteomyelitis

No congruent distribution Osteomyelitis MRI

Diagnostic flow-chart of osteomyelitis in the mid- and hindfoot. MRI, magnetic resonance imaging; WBC, white blood cell.

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Nuclear medicine imaging of diabetic foot infection Capriotti et al. 763

Boemi, Paolo Braggio, Luca Burroni, Dario Cantalupi, Gabriela Capriotti, Giuseppe Cascini, Marco Chianelli, Arturo Chiti, Micaela D’Alberto, Diego De Palma, Giovanni D’Errico, Narcisa De Vincentis, Salvatore di Rosa, Paola Erba, Antonio Ferrarese, Guido Ferretti, Chiara Gallini, Elena Lazzeri, Lorenzo Maffioli, Giulia Manfredini, Rita Mannino, Luigi Mansi, Giuliano Mariani, Mario Marinelli, Pietro Marinelli, Luigi Martino, Federica Matteucci, Marino Mele, Angelo Mita, Monica Mori, Maria Gemma Parisella, Valentina Picardi, Carlo Poti, Michele Povolata, Napoleone Prandini, Pierfrancesco Rambaldi, Brunella Rossi, Domenico Rubello, Vittoria Rufini, Orazio Schillaci, Alberto Signore, Alberto Spina, Luca Tagliabue, Maria Cristina Tappa, Daniela Turrin, Venanzio Valenza, Anna Viglietti and Alberto Vignati. We express our gratitude and acknowledgements to all of these members for their collaboration in the preparation and progress of this study.

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Segall GM, Nino-Murcia M, Jacobs T, Chang K. The role of bone scan and radiography in the diagnostic evaluation of suspected pedal osteomyelitis. Clin Nucl Med 1989; 14:255–260. Schauwecker DS, Park HM, Burt RW, Mock BH, Wellman HN. Combined bone scintigraphy and indium-111 leukocyte scans in neuropathic foot disease. J Nucl Med 1988; 29:1651–1655. Tawn DJ, O’Hare JP, O’Brien IA, Watt I, Dieppe PA, Corrall RJ. Bone scintigraphy and radiography in the early recognition of diabetic osteopathy. Br J Radiol 1988; 61:273–279. Maurer AH, Millmond SH, Knight LC, Mesgarzadeh M, Siegel JA, Shuman CR, et al. Infection in diabetic osteoarthropathy: use of indium-labeled leukocytes for diagnosis. Radiology 1986; 161:221–225. Seldin DW, Heiken JP, Feldman F, Alderson PO. Effect of soft-tissue pathology on detection of pedal osteomyelitis in diabetics. J Nucl Med 1985; 26:988–993. Alazraki N, Dries D, Datz F, Lawrence P, Greenberg E, Taylor A Jr. Value of a 24-hour image (four-phase bone scan) in assessing osteomyelitis in patients with peripheral vascular disease. J Nucl Med 1985; 26:711–717. Sartoris DJ, Devine S, Resnick D, Golbranson F, Fierer J, Witztum K, et al. Plantar compartmental infection in the diabetic foot. The role of computed tomography. Invest Radiol 1985; 20:772–784. Edmonds ME, Clarke MB, Newton S, Barrett J, Watkins PJ. Increased uptake of bone radiopharmaceutical in diabetic neuropathy. Q J Med 1985; 57:843–855. Visser HJ, Jacobs AM, Oloff L, Drago JJ. The use of differential scintigraphy in the clinical diagnosis of osseous and soft tissue changes affecting the diabetic foot. J Foot Surg 1984; 23:74–85. Park HM, Wheat LJ, Siddiqui AR, Burt RW, Robb JA, Ransburg RC, Kernek CB. Scintigraphic evaluation of diabetic osteomyelitis: concise communication. J Nucl Med 1982; 23:569–573. Knight D, Gray HW, McKillop JH, Bessent RG. Imaging for infection: caution required with the Charcot joint. Eur J Nucl Med 1988; 13: 523–526. Fox IM, Zeiger L. Tc-99m-HMPAO leukocytes scintigraphy for the diagnosis of osteomyelitis in diabetic foot infection. J Foot Ankle Surg 1993; 32:591–594. Blume PA, Dey HM, Daley LJ, Arrighi JA, Soufer R, Gorecki GA. Diagnosis of pedal osteomyelitis with 99mTc-HMPAO labeled leukocytes. J Foot Ankle Surg 1997; 36:120–126. Devillers A, Moisan A, Hennion F, Garin E, Poirier JY, Bourguet P. Contribution of technetium-99m hexamethyl propylene amine oxime labeled leukocyte scintigraphy to the diagnosis of diabetic foot infection. Eur J Nucl Med 1998; 25:132–138. Keenan AM, Tindel NL, Alavi A. Diagnosis of pedal osteomyelitis in diabetic patients using current scintigraphic techniques. Arch Intern Med 1989; 149:2262–2266. Newman LG, Waller J, Palestro CJ, Schwartz M, Klein MJ, Hermann G, et al. Unsuspected osteomyelitis in diabetic foot ulcers: diagnosis and monitoring by leukocyte scanning with In-111 oxyquinoline. J Am Med Assoc 1991; 266:1246–1251. Palestro CJ, Roumanas P, Swyer AJ, Kim CK, Goldsmith SJ. Diagnosis of musculoskeletal infection using combined In-111 labeled leukocyte and Tc-99m SC marrow imaging. Clin Nucl Med 1992; 17:269–273. Tomas MB, Patel M, Marwin SE, Palestro CJ. The diabetic foot. Br J Radiol 2000; 73:443–450. Unal SN, Birinci H, Baktiroglu S, Cantez S. Comparison of Tc-99m methylene diphosphonate, Tc-99m human immuno globulin, and Tc-99m-labeled white blood cell scintigraphy in the diabetic foot. Clin Nucl Med 2001; 26:1016–1021. Oyen WJG, Netten PM, Lemmens AM, Claessens RA, Lutterman JA, van der Vliet JA, et al. Evaluation of infectious diabetic foot complication with In-111labelled human nonspecific immunoglobulin G. J Nucl Med 1992; 33:1330–1336.

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Original article

The influence of attenuation correction and reconstruction techniques on the detection of hypo-perfused lesions in brain SPECT images Shivani Ghooruna,c, Kristof Baeteb, Johan Nuytsb, Wilhelm Groenewaldc and Patrick Dupontb Background We evaluated the effects of attenuation correction and reconstruction techniques on the detection of hypoperfused lesions in brain SPECT imaging. Methods A software phantom was constructed using the data available on the BrainWeb database by assigning activity values to grey and white matter. The true attenuation map was generated by assigning attenuation coefficients to six different tissue classes to create a non-uniform attenuation map. The uniform attenuation map was calculated using an attenuation coefficient of 0.15 cm – 1. Hypoperfused lesions of varying intensities and sizes were added. The phantom was then projected as typical SPECT projection data, taking into account attenuation and collimator blurring with the addition of Poisson noise. The projection data were reconstructed using four different methods: filtered back-projection in combination with Chang’s first-order attenuation correction using the uniform or the true attenuation map and maximum likelihood iterative reconstruction using the uniform or the true attenuation map. Different Gaussian post-smoothing kernels were applied onto the reconstructed images and the performance of each procedure was analysed using figures of merit such as signal-to-noise ratio, bias and variance.

Introduction Functional brain imaging using single photon emission computed tomography (SPECT) has widespread applications in Alzheimer’s disease, acute stroke, transient ischaemic attacks, epilepsy, recurrent primary tumours and head trauma [1,2]. However, quantification of these images may pose difficulties due to reconstruction and attenuation correction issues. Attenuation correction in SPECT is accepted as a significant component to producing accurate quantitative data. Transmission-based methods can supply non-uniform attenuation coefficients and therefore more accurate absolute quantification [3,4]. However, transmissionbased methods require an additional scan. Radionuclide transmission scans contain noise which can propagate into the final reconstructed images [3]: there may also be patient movement between the transmission and emission scan if they are not acquired simultaneously. Zaidi

Results Uniform attenuation correction offered only slight deterioration of the signal-to-noise ratio compared to the true attenuation map. Maximum likelihood produced superior signal-to-noise ratios and lower bias at the same variance in comparison to the filtered back-projection. Conclusion Uniform attenuation correction is adequate for lesion detection while maximum likelihood provides enhanced lesion detection when compared to filtered c 2006 back-projection. Nucl Med Commun 27:765–772
Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:765–772 Keywords: filtered backprojection, maximum likelihood, OSEM Departments of aNuclear Medicine, bNuclear Medicine, UZ GasthuisbergKULeuven, Leuven, Belgium and cMedical Physics, Tygerberg Hospital and Stellenbosch University, Cape Town, South Africa. Correspondence to Professor Patrick Dupont, Nuclear Medicine, UZ Gasthuisberg, 49 Herestraat, 3000 Leuven, Belgium. Tel: + 0032 16 343715; fax: + 0032 16 343759; e-mail: [email protected] Received 17 February 2006 Accepted 4 May 2006

et al. [5] aligned the magnetic resonance images to PET reconstructed data and segmented the magnetic resonance image to identify tissues of significantly different density and composition. In this way an accurate attenuation map could be obtained. Kinahan et al. [6] segmented and scaled computed tomography (CT) data to obtain accurate attenuation coefficients. The latter method has the disadvantage of giving an additional radiation burden to the patient. Methods of non-uniform attenuation correction are, however, not always feasible in routine clinical practice. Several studies have compared uniform and non-uniform attenuation correction methods for quantification in SPECT [4,7–9]. Some studies [8,10] showed that non-uniform attenuation correction was better than uniform attenuation correction. In the study of Bai et al. [10], measurements of normal subjects were used in which no ‘gold standard’ technique was used. Licho et al. [8] used a slice independent cylindrical uniform attenuation map resulting in over-correction in

c 2006 Lippincott Williams & Wilkins 0143-3636


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the vertex. Other studies [9,11–13] demonstrated only slight improvement with non-uniform attenuation correction when comparing regional cerebral blood flow. Small differences were seen in the infra-tentorial region or the lower slices due to the airways or higher slices due to the relatively large skull. Many of these studies assessed quantification using regional cerebral blood flow measurements. In some cases, no ‘gold standard’ was available. It was still unclear, however, whether the approximation by a uniform attenuation map provides specific disadvantages in the clinical practice of lesion detection of functional brain SPECT imaging. With regard to the reconstruction method, several studies demonstrated that iterative reconstruction techniques produced superior image quality. Koch et al. [14] found that the ordered subsets expectation maximization (OSEM) algorithm provided better separation between the caudate and the putamen compared to filtered backprojection (FBP). A study by Chornoboy et al. [15] concluded that maximum likelihood reconstruction exhibit improved signal-to-noise ratios (SNRs), improved image resolution, and improved quantification compared to FBP. Kauppinen et al. [16] evaluated the quantitative accuracy of iterative reconstruction in SPECT brain perfusion imaging using a three-dimensional high resolution anthropomorphic phantom. Iterative reconstructions were individually post-filtered. The study showed that iterative reconstruction techniques increased the contrast of the image and improved separation between the different regions. Many of these comparative studies were performed as simple phantom experiments and concentrated on image quality or spatial resolution. Our study focused on the quantitative improvement on lesion detection when using iterative reconstruction techniques. We also investigated the deterioration in detection performance when a uniform attenuation map was used. The simulation experiment was designed to study the influence of both parameters in a single simulation experiment. We believe that the ability of a technique to detect lesions has direct clinical relevance in brain SPECT imaging. To our knowledge, there is limited literature related to the effects of attenuation correction and reconstruction techniques on lesion detection.

Materials and methods A simulation experiment was designed to independently evaluate the influence of attenuation correction and image reconstruction. Construction of a brain SPECT software phantom

The brain phantom was generated from the threedimensional digital phantom provided by the BrainWeb database [17,18]. Each voxel was assigned a tissue class

depending on its tissue type. The tissue types of interest were the grey matter, white matter and cerebrospinal fluid (for the activity map) and additionally the fatty tissue, skull and air (for the attenuation map). The segmented magnetic resonance imaging (MRI) volumes were assigned relative activity levels. The perfusion ratio of grey matter to white matter was taken as 4:1 to simulate 99mTc-ethyl cysteinate dimer (99mTc-ECD) studies [19]. Cerebrospinal fluid does not accumulate the radioactive isotope, and therefore these voxels were assigned a tracer uptake value of zero. All other classes were assigned a value of 0.5 except skull, and background which was set to zero. The projections were then scaled to an average count level of 16 counts per pixel. This scaling factor was determined by reviewing the total counts of the projection data of 18 normal ECD brain SPECT scans performed at our institution. The activity map used a matrix size of 83  83 with 60 planes with an isotropic voxel size of 2.6 mm. Seven spherical cold lesions with 50% and 100% reduction in intensity and radius of 5.2 mm and 10.4 mm were introduced in the phantom. The smallest lesion corresponds to a region of a similar size as the final image resolution. The larger lesion is twice this size. We took these values to avoid having results that are either influenced too much by the partial volume effect (and therefore not detectable) or to avoid reporting results of very large lesions which are most likely to be detectable by any method. Four datasets were thus generated. The lesions were positioned at the locations shown in Fig. 1. Appropriate linear attenuation coefficients, m (in cm – 1), were assigned to each voxel of the brain depending on their tissue classifications. The tissue types of interest (and the corresponding linear attenuation coefficients) used in generating the non-uniform attenuation image were [20]: grey matter (m = 0.1558 cm – 1), white matter (m = 0.1558 cm – 1), skull (m = 0.2842 cm – 1), fatty tissue (m = 0.1425 cm – 1) and air (m = 0 cm – 1). All other voxels in the brain were assigned a linear attenuation coefficient value equivalent to soft tissue (m = 0.1582 cm – 1). This image was used as the true attenuation map. Generation of the projections

The SPECT acquisition process was simulated by projecting the baseline and hypoperfused phantom for 120 angles over 3601 with a pixel size of 2.6 mm and distance between collimator and rotation axis of 12 cm. This configuration corresponds to the clinical brain SPECT acquisition protocol used in Leuven. The resolution of the system at a distance d (in mm) from the collimator was modelled as [21] FWHM2 ðd Þ ¼ FWHM2intrinsic þ ½0:04ðd þ 50Þ2 to take into account the position dependent collimator blurring (simulating a low energy high resolution collimator)

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Effect of reconstruction in detecting brain lesions on SPECT images Ghoorun et al. 767

other values were set to zero. Back-projection and normalization produced the fraction of projection lines that contained a zero for each of the pixels. Thresholding using a fraction of 0.1 yielded the object. Each voxel of the object was assigned a value of 0.15 cm – 1. A new uniform map was created for every noise realization. This approach was based on the assumption that nearly all of the projection lines intersecting the object would collect counts. The procedure was effective when sufficient tracer uptake was present near the boundary of the object.

Fig. 1

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Generation of the reconstructed data 3 5

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Each noisy sinogram was reconstructed using four different techniques (Fig. 2) namely, FBP with a uniform attenuation map (FBP-unif), FBP with a true attenuation map (FBP-true), maximum likelihood (ML) with a uniform attenuation map (ML-unif) and maximum likelihood with a true attenuation map (ML-true). In the case of FBP using Chang’s first-order attenuation correction [24], the average attenuation coefficient was computed for each pixel. Thereafter each pixel value of Fig. 2

7 A

Hypoperfused phantom showing the positions of the seven lesions.

B

and the intrinsic resolution ðFWHMintrinsic ¼ 3:7 mmÞ. The parameters were determined on the basis of line source (filled with 99mTc) measurements at different distances from the collimator using a Trionix TRIAD SPECT camera equipped with parallel ultra-high resolution, low energy collimators. We applied this model in the projection using Gaussian diffusion [22]. Attenuation was taken into account. To ensure that the data were clinically realistic, pseudo-random Poisson noise was added to the data using the pseudo-random generator of IDL 5.2. Each set of noisy projection data represented a noise realization and 100 noise realizations were generated.

C

D

Creation of a uniform attenuation map

Although on many commercial systems an ‘ellipse’ contour is taken on a selected mid-slice and the map created applied to all slices to obtain a uniform attenuation map, we followed a more accurate approach to obtain an individual slice-by-slice map. The calculated uniform attenuation map was determined as follows [23]. A median filter with a mask of 3 pixels was applied to the sinogram. Thereafter, the zero values were set to 1 and all

Reconstruction of a single noise realization using the four different methods. (A) Filtered back-projection (FBP) with the uniform attenuation map, (B) FBP with true attenuation map (C) maximum likelihood with uniform attenuation map, (D) maximum likelihood with true attenuation map.

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the regular (non-corrected) FBP image was divided by the average attenuation coefficient. In the case of the maximum likelihood methods, the attenuation coefficients were incorporated into the projection and backprojection operations for each iteration. An iterative scheme using a decreasing number of subsets was used: iterations subsets = 1 30, 2 24, 3 20, 4 15, 4 12, 4  10, 4  8, 5  6, 5  5, 5  4, 5  3, 5  2, 5  1. This was equivalent to 423 iterations. Correction for collimator blurring was not incorporated in the maximum likelihood algorithm to represent the typical clinical situation. Analysis of data

Each reconstructed image was post-smoothed, using a three-dimensional Gaussian blurring kernel. The FWHM of the kernel was varied between zero and 10 pixels (26 mm). Regions of interest, corresponding to twice the lesion size, were positioned over the lesions. It was ensured that the regions did not overlap. Three performance measures were used: (1) the nonprewhitening observer by calculating its signal-to-noise ratio (SNR), (2) the root mean squared (RMS) bias and (3) RMS variance [25–27]. For notational convenience, the following symbols were used: Bnl represents the reconstructed images of the noiseless SPECT emission data of the baseline phantom, i.e. the phantom without lesions. Hnl represents reconstructed images of the noiseless hypoperfused phantom, i.e. the phantom with lesions. and H(r) represent the values in voxel j of the reconB(r) j j structed images of noise realization r of the SPECT emission data for the baseline B and hypoperfused H phantoms respectively. Btrue and Htrue denote the ‘true’ baseline and hypoperfused phantom images respectively.

The response function snl for each reconstruction technique and for each noise realization r was measured as follows: snl ðBðrÞ ; RÞ ¼

X

ðrÞ

ð1Þ

ðrÞ

ð2Þ

nl ðBnl j  Hj ÞðBj Þ

j2R

for the baseline phantom and snl ðH ðrÞ ; RÞ ¼

X

nl ðBnl j  Hj ÞðHj Þ

j2R

for the hypoperfused phantom, with R representing a region of interest.

The SNR for R was computed using sffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi ½
sðBÞ 
sðHÞ 2 SNRðRÞ ¼ 2 2 ss ðBÞ þ s2s ðHÞ

ð3Þ

where
s(B) and
s(H) represents the mean of snl(B(r),R) and snl(H(r),R) respectively, and s2s (B) and s2s (H) are the corresponding variances over all noise realizations. Equation 3 is a measure of the performance of the nonprewhitening observer for the case of exactly known signal and background [28]. The bias and variance measurements were also determined. The bias image was calculated using bðIÞ ¼ I
 I true ð4Þ where I
is the mean of I (r) over all noise realizations and I represents B or H. Itrue represents the ‘ground truth’ image where the ‘ground truth’ image is the activity map. ~ RÞ in region R was The root mean squared bias bðI; computed using sffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi 1 X 2 ~ bðI; RÞ ¼ b ðIÞ ð5Þ nR j2R j with nR the number of voxels in R. The variance image s2 (I) was calculated using s2 ðIÞ ¼

P 1 X
2 ½ðI ðrÞ Þ  ðIÞ P  1 r¼1

ð6Þ

with P equal to the number of noise realizations. The root mean squared standard deviation was computed using sffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi 1 X 2 2 ~ ðI; RÞ ¼ s ðIÞ: ð7Þ s nR j2R j The mean image over all noise realizations using the dataset without lesions was compared to the ‘ground truth’ image. The purpose was to quantify the percentage deviation of the reconstructed image from the ‘ground truth’ image. Two-dimensional regions of interest were drawn at different locations of the brain and the mean difference, which represented the bias, was computed as meanbaseline  meantrue difference ¼ 100%: ð8Þ meantrue Two-dimensional regions were selected to represent the scenario of a normal clinical setting where, typically, regions over a particular area were drawn and the values obtained. This method provided a simple measure of the bias contained in these values.

Results Signal-to-noise ratio

Signal-to-noise ratios were determined for the different reconstruction algorithms. The higher these values, the more ‘easy’ it is to detect the lesions. These values are

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Effect of reconstruction in detecting brain lesions on SPECT images Ghoorun et al. 769

Fig. 3

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SNR vs Postsmooth (whole brain)

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FBP-unif FBP-true ML-unif ML-true

Plot of signal-to-noise ratio (SNR) versus post-smooth for different regions in the brain for dataset corresponding to 100% reduction in intensity and lesion radius of 5.2 mm. K, FBP-unif, *, FBP-true, !, ML-unif, !, ML-true.

plotted as a function of the post-smooth (expressed as the FWHM of the Gaussian kernel). Figure 3 shows the curves for a lesion size of radius 5.2 mm (2 pixels) and a 100% reduction in perfusion. The FWHM values are in pixels where 1 pixel = 2.6 mm. The SNR peaks at a smoothing of 2–4 pixels (depending on the lesion) and then decreases. Furthermore, the maximum likelihood method of reconstruction demonstrates higher SNR values when compared to the FBP method. However, very little differences in SNR can be observed between the true and the uniform attenuation maps. An attempt was made to quantify these differences using Equation 8 for each region and for each postsmooth. The maximum difference in SNR between FBP-unif and FBP-true at optimal smoothing was 1.14% and between ML-unif and ML-true was 0.95%. Comparing the SNR of ML-unif and FBP-unif, we found an increase between 0.5% (region 2) and 11.8% (region 6) for the iterative reconstruction. We also studied the SNR curves for a lesion of 50% reduction and for bigger lesions. Similar results were found although the optimal smoothing value was different for the bigger lesions (4–6 pixels). The

maximum difference in SNR between FBP-unif and FBPtrue was 1.6% and 2.2% for the maximum difference in SNR between ML-unif and ML-true showing that the effect of using a uniform attenuation map has only a moderate effect on the SNR values. Comparing the SNR values between FBP-unif and ML-unif over all lesions, sizes and decreases, we found a maximal increase of 24.1% in the SNR of the iterative reconstruction against the SNR of FBP. Variance versus bias

Figure 4 represents the region encompassing the entire brain and pertain to the baseline images, i.e., the noisy reconstructions without lesions. The root mean squared bias and standard deviation were calculated using Equations 5 and 7. The curves illustrate that the variance (expressed as the root mean squared standard deviation) decreased with post-smooth at the expense of increased bias. We observed systematic higher values in bias and variance when the uniform attenuation map was used compared to the case of the true attenuation map. However, when using maximum likelihood methods the bias and variance values were decreased irrespective of the attenuation map used.

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770 Nuclear Medicine Communications 2006, Vol 27 No 10

Fig. 4

rms std dev

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Plot of root mean squared (RMS) standard deviation versus RMS bias of the whole brain. The points of the curve indicate a post-smooth of 0, 2, 4, 6, 8 and 10 pixels. Symbols as in Fig. 3.

Mean differences between baseline and ‘ground truth’ image

In order to obtain an idea of the local bias in the different reconstructions, the mean image over all noise realizations using the data set without lesions was compared to the ‘ground truth’ image. Six two-dimensional regions of interest were drawn at different locations to simulate the clinical situation. The maximum difference between the baseline and ground truth image was found to be 9.8% with the ML-true method and 11.71% with the ML-unif method. The FBP methods showed maximum differences of 31.60% (FBP-true) and 33.53% (FBP-unif).

Discussion

Analysis of our simulation experiment necessitated comparison of a large number of images. No filtering was applied to the images as this would have introduced an additional variable in the study. It is well known that the maximum likelihood image using many iterations is subjectively a very poor one [29]. This is because the maximum likelihood algorithm strictly enforces coherence with projection data and noisy projections produce noisy reconstructions. Furthermore, FBP techniques, which mathematically include a ramp filter, result in amplification of high frequencies and thus produce noisy images. We therefore applied Gaussian post-smoothing with different kernel sizes to deal with the noise in the reconstructed data. The noise properties using a penalized likelihood method as opposed to post-smoothing the maximum likelihood reconstruction using Gaussian smoothing were compared previously [30,31]. These studies found that for applications where a shift-invariant spatial resolution was imposed, the two regularization methods performed similarly. In our study, we varied the post-smooth to obtain optimal SNR values with the objective of assessing the detection capability of the different reconstructions. In these experiments, numerical observers were used rather than human observers. It would have been impractical for the large number of images to be inspected by humans. To obtain the optimal smoothing for lesion detection, we evaluated the SNR values for a range of smoothing kernels. It was observed that SNR values initially increased, peaked between a FWHM of 2 and 4 pixels and then decreased. We found that the amount of smoothing necessary to achieve peak SNR was dependent on the size of the lesion. For bigger lesions, optimal smoothing, indicated by the peak in SNR values, was between 4 and 6 pixels (data not shown). We postulated that the phenomenon of peak SNR at a particular post-smooth was related to the theory of matched filters [32].

Accurate detection of lesions is important for the clinical evaluation of various brain disorders using perfusion SPECT. Nowadays, commercially available hybrid SPECT–CT scanners provide accurate attenuation maps. Our study is still of importance in assessing the use of the calculated uniform attenuation map because not all gamma cameras are equipped with a CT scanner. Furthermore, using a calculated attenuation map requires no additional scan and hence no extra radiation burden. Also, misregistration errors due to patient movement are obviated.

In our experiment we incorporated position dependent collimator blurring in the projection data to model the detection system. However, we did not correct for collimator blurring during the reconstructions in order to represent our standard clinical situation. This also enabled an equal comparison between FBP and the maximum likelihood technique.

In view of the lack of studies focusing on lesion detection as well as improvements advocated with iterative techniques, further evaluation of the clinical impact of attenuation correction and reconstruction methods was warranted. These were addressed by our simulation study.

Influence of attenuation correction

In the following, we first evaluated the influence of attenuation correction and then the reconstruction techniques on lesion detection.

Region 1 is located in the temporal pole of the brain and region 2 in the cerebellum. These regions were close to

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Effect of reconstruction in detecting brain lesions on SPECT images Ghoorun et al. 771

bony structures and areas where the attenuation map was clearly not uniform. Therefore they may have suffered the effects of the use of a uniform attenuation map and produced inferior reconstructed data. Regions 3 and 4 were located mostly in grey matter and regions 5 and 6 in white matter. These were at levels where we expected the attenuation coefficients could be approximated by a uniform value. Region 7 was a cranial lesion with more bony structures. We expected that the use of a uniform attenuation map may deteriorate the detection performance. However, from Fig. 3, we observed that there was very little separation of the curves when comparing the use of the uniform attenuation map to the true attenuation map, irrespective of the reconstruction method and position of the lesion. Figure 4 illustrates that, generally, smoothing decreases the variance at the expense of increased bias. When comparing the different algorithms, it was observed that at the same variance, the bias was only slightly higher when the uniform attenuation maps were used. This is in accordance with previous studies [10–12] which compared the use of uniform and non-uniform attenuation maps. From our simulation experiment, we concluded that the use of a calculated uniform attenuation correction does not markedly affect the detection of lesions. Influence of reconstruction methods

From the SNR curves (Fig. 3), a significant difference was noted between the maximum likelihood and FBP methods with the maximum likelihood methods always outperforming the FBP methods. This improvement in detection performance was seen in all lesions of the brain. This can be attributed to the fact that the maximum likelihood method takes into account the correct noise model. The bias–variance curves (Fig. 4) confirmed a systematic improvement when the maximum likelihood methods of reconstruction were used. When comparing the mean image over all noise realizations to the ‘ground truth’ image, the ML-true method performed best. The ML-true method produced a 9.8% difference whereas the ML-unif was 11.71%. This difference between ML-true and ML-unif was very small. In addition, these slight differences are probably exaggerated because a perfect attenuation map obtained from MRI data was used. In clinical practice, a density map is obtained from a transmission study using a gamma camera or by matching a CT study. The use of such a density map results in poorer resolution and lower accuracy.

We also compared the mean image over all noise realizations to the ‘ground truth’ image for the FBP methods. We obtained a maximum difference of 31.60% for the FBP-true method and 33.53% for the FBP-unif method. These large differences observed with the FBP methods further illustrated that the maximum likelihood algorithms performed superiorly and the use of the true attenuation map only added slight improvement to the quality of the image. These findings are in agreement with a study by Van Laere et al. [33] which showed improved image quality when iterative reconstruction techniques were used. The study by these authors also showed better delineation between grey and white matter, and improved contrast. Limitations of the study

The current study has a number of limitations: using lesions with a complex shape is more realistic. The exact shape of a lesion will affect detectibility, but we expect that there is almost no interaction between the shape and the method of reconstruction/attenuation correction. Therefore, we believe that the conclusions put forward in this manuscript remain valid even for lesions of different shape. The relatively large reductions in activity were chosen to avoid having a reduction which is near or below the detection limit. Furthermore, we did not include scatter in the projection data. Our study focused on attenuation and distant dependent collimator blurring, which are expected to have a much stronger effect on detectibility. Inclusion of scatter would probably have further increased the difference between MLEM and FBP, because MLEM can incorporate scatter correction in the reconstruction, which has been shown to be superior to pre-correction [34]. Extrapolation to the clinical situation

In this study we used numerical observers. The advantage is that an objective measure of image quality can be produced without human interaction, enabling the evaluation of large amounts of images. The nonprewhitening matched-filter observer (NPW) has been shown to correlate reasonably well with human observer performance for some detection tasks. However, for other tasks, humans outperform the NPW, indicating that humans have a limited pre-whitening capacity (i.e., they are able to exploit the presence of correlated noise) [35]. In this study we have used the NPW observer because it is simple to compute. The drawback is that it is not a very accurate predictor of human performance: the pixel values are actually correlated, and humans can exploit this correlation to some extent. Furthermore, in a clinical situation, lesion detection is a complex process of pattern recognition that depends on a number of factors in addition to the SNR. An experienced reader knows a

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772 Nuclear Medicine Communications 2006, Vol 27 No 10

normal perfusion pattern if he knows the structural MRI. The human observer – if experienced – can also use expected patterns of abnormality depending on the pathology which is investigated. These considerations justify (to some extent) of the ‘signal known exactly, background known exactly’ concept. But accurate modelling of the human observer is, of course, very difficult. However, in previous work [27] using both numerical and human observers, we found that the NPW observer was a better predictor for the performance of less experienced human observers. Therefore, we expect that the performance of the most experienced human readers will remain the same or only slightly improve when using an iterative reconstruction while the performance of less experienced readers may greatly improve. Based on our simulations the issue of using an uniform attenuation correction instead of the ‘true’ attenuation correction, is less important for the detection of lesions.

Conclusion Our investigation showed that, in the case of brain SPECT imaging, the use of uniform attenuation correction did not significantly affect the ability to detect lesions. On the other hand, marked improvement in detection performance was observed when iterative reconstruction algorithms were utilized. The implementation of iterative reconstruction techniques together with appropriate smoothing clearly improved detection capability in brain SPECT images.

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Acknowledgements

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This work was funded by a grant from the Research Fund Katholieke Universiteit Leuven OT/00/32 and IDO/03/010.

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References 1

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Catafau AM, Lomena FJ, Pavia J, Parellada E, Bernardo M, Setoain J, et al. Regional cerebral blood flow pattern in normal young and aged volunteers: A 99m Tc-HMPAO SPET study. Eur J Nucl Med 1996; 10:1329–1337. Holman BL, Devous Sr MD. Functional brain SPECT: the emergence of a powerful clinical method. J Nucl Med 1992; 33:1888–1904. Bailey DL. Transmission scanning in emission tomography [Review]. Eur J Nucl Med 1998; 25:774–787. Rajeevan N, Zubal G, Ramsby SQ, Zoghbi SS, Seibyl J, Innis RB. Significance of nonuniform attenuation correction in quantitative brain SPECT imaging. J Nucl Med 1998; 39:1719–1726. Zaidi H, Montandon M, Slosman DO. Magnetic resonance imaging-guided attenuation and scatter corrections in three dimensional brain positron emission tomography. Med Phys 2003; 30:937–948. Kinahan PE, Townsend DW, Beyer T, Sashin D. Attenuation correction for a combined 3D PET/CT scanner. Med Phys 1998; 25:2046–2053. Stodilka RJ, Kemp BJ, Prato FS, Kertesz A, Kuhl D, Nicholson RL. Scatter and attenuation correction for brain SPECT using attenuation distributions inferred from a head atlas. J Nucl Med 2000; 41:1569–1578. Licho R, Glick SJ, Xia W, Pan TS, Penney BC, King MA. Attenuation compensation in 99mTc SPECT brain imaging: a comparison of the use of attenuation maps derived from transmission versus emission data in normal scans. J Nucl Med 1999; 40:456–463. Iida H, Narita Y, Kado H, Kashikura A, Sugawara S, Shoji Y, et al. Effects of scatter and attenuation correction on quantitative assessment of regional cerebral blood flow with SPECT. J Nucl Med 1998; 39:181–189.

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Bai J, Hashimoto J, Ogawa K, Kubo A, Fukunaga A, Onozuka S, et al. Influence of photon scattering and attenuation on ROI analysis in brain perfusion single-photon emission tomographic imaging of normal subjects. Ann Nucl Med 2005; 19:567–572. Larsson A, Johansson L, Sundstrom T, Ahlstrom KR. A method of attenuation and scatter correction of brain SPECT based on computed tomography images. Nucl Med Commun 2003; 24:411–420. Van Laere K, Koole M, Verspijt J, Dierckx R. Non-uniform versus uniform attenuation correction in brain perfusion SPET of healthy volunteers. Eur J Nucl Med 2001; 28:90–98. Arlig A, Gustafsson A, Jacobsson L, Ljungberg M, Wikkelso C. Attenuation correction in quantitative SPECT of cerebral blood flow: a Monte Carlo study. Phys Med Biol 2000; 45:3847–3859. Koch W, Hamann C, Welsch J, Popperl G, Radau PE, Tatsch K. Is iterative reconstruction an alternative to filtered backprojection in routine processing of dopamine transporter SPECT studies? J Nucl Med 2005; 46:1804–1811. Chornoboy ES, Chen CJ, Miller MI, Miller TR, Snyder DL. An evaluation of maximum likelihood reconstruction for SPECT. IEEE Trans Med Imaging 1990; 9:99–110. Kauppinen T, Koshkinen MO, Alenius S, Vanninen E, Kuikka JT. Improvement of brain perfusion SPET using iterative reconstruction with scatter and nonuniform attenuation correction. Eur J Nucl Med 2000; 27:1380–1386. BrainWeb. http://www.bic.mni.mcgill.ca/brainweb (25 January 2006). Collins DL, Zijdenbos AP, Kollokian JG, Sled JG, Kabani NJ, Holmes CJ, et al. Design and construction of a realistic digital brain phantom. IEEE Trans Med Imaging 1998; 17:463–468. Owunwanne A, Patel M, Sadek S. The handbook of radiopharmaceuticals. London: Chapman and Hall Medical, 1995. http://physics.nist.gov/PhysRefData/XrayMassCoef/tab4.html (13 April 2006) Cherry SR, Sorenson JA, Phelps ME. Physics in nuclear medicine, 3rd edition (chapter 14), Saunders, 2003. McCarthy AW, Miller MI. Maximum likelihood SPECT in clinical computation times using mesh-connected parallel computers. IEEE Trans Med Imaging 1991; 10:426–436. Nuyts J, Dupont P, Stroobants S, Benninck R, Mortelmans L, Suetens P. Simultaneous maximum a posteriori reconstruction of attenuation and activity distributions from emission sinograms. IEEE Trans Med Imaging 1999; 18:393–403. Chang LT. A method for attenuation correction in radionuclide computed tomography. IEEE Trans Nucl Sci 1978; 25:638–643. Wagner RF, Brown DG. Unified SNR analysis of medical imaging systems. Phys Med Biol 1985; 30:489–518. Baete K, Nuyts J, Van Paesschen W, Suetens P, Dupont P. Anatomicalbased FDG-PET reconstruction for the detection of hypo-metabolic regions in epilepsy. IEEE Trans Med Imaging 2004; 23:510–519. Baete K, Nuyts J, Van Laere K, Van Paesschen W, Ceyssens S, De Ceuninck L, et al. Evaluation of anatomy based reconstruction for partial volume correction in brain FDG-PET. Neuroimage 2004; 23:305–317. Barrett HH, Yao J, Rolland JP. Model observers for the assessment of image quality [Colloquium paper]. Proc Natl Acad Sci 1993; 90: 9758–9765. Barret HH, Wilson DW, Tsui BMW. Noise properties of the EM algorithm: I. Theory. Phys Med Biol 1994; 39:833–846. Nuyts J, Fessler JA. A penalized likelihood image reconstruction method for emission tomography, compared to postsmoothed maximum-likelihood with matched spatial resolution. IEEE Trans Med Imaging 2003; 22:1042–1052. Stayman JW, Fessler JA. Compensation for nonuniform resolution using penalized-likelihood reconstruction in space-variant imaging systems. IEEE Trans Med Imaging 2004; 23:269–284. Herath KB, Sharp PF. Effects of ‘matched filter’ smoothing as measured by receiver operating characteristic curve. Phys Med Biol 1976; 21: 442–446. Van Laere K, Koole M, Kauppinen T, Monsieurs M, Bouwens L, Dierckx R. Nonuniform transmission in brain SPECT using 201Tl, 153Gd, and 99mTc static line sources: anthropomorphic dosimetry studies and influence on brain quantification. J Nucl Med 2000; 41:2051–2062. Kadrmas DJ, Frey EC, Tsui BMW. Analysis of the reconstructibility and noise properties of scattered photons in 99mTc SPECT. Phys Med Biol 1997; 42:2493–2516. Abbey CK, Barrett HH. Human- and model-observer performance in rampspectrum noise: Effects of regularization and object variability. J Opt Soc Am A 2001; 18:473–488.

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Original article

Evaluation of transcriptional activity of the oestrogen receptor with sodium iodide symporter as an imaging reporter gene Joo Hyun Kanga,b,c, June-Key Chungb,c, Yong Jin Leeb,c, Kwang Il Kimb,c, Jae Min Jeongb,c, Dong Soo Leeb and Myung Chul Leeb Background Oestrogen receptors are ligand-dependent transcription factors whose activity is modulated either by oestrogens or by an alternative signalling pathway. Oestrogen receptors interact via a specific DNA-binding domain, the oestrogen responsive element (ERE), in the promoter region of sensitive genes. This binding leads to an initiation of gene expression and hormonal effects. Objective To determine the transcriptional activity of the oestrogen receptor, we developed a molecular imaging system using sodium iodide symporter (NIS) as a reporter gene. Methods The NIS reporter gene was placed under the control of an artificial ERE derived from pERE-TA-SEAP and named as pERE-NIS. pERE-NIS was transferred to MCF-7, human breast cancer cells, which highly expressed oestrogen receptor-a with lipofectamine. Stably expressing cells were generated by selection with G418 for 14 days. After treatment of 17b-oestradiol and tamoxifen with serial doses, the 125I uptake was measured for the determination of NIS expression. The inhibition of NIS activity was performed with 50 lmol
l – 1 potassium perchlorate. Results The MCF7/pERE-NIS treated with 17b-oestradiol accumulated 125I up to 70–80% higher than did non-treated cells. NIS expression was increased according to increasing doses of 17b-oestradiol. MCF7/pERE-NIS

Introduction Oestrogen exerts profound effects on growth, differentiation and function of cardiovascular tissues and the skeletal system as well as reproductive tissues, including the genital tract and mammary gland. The highest amounts of the oestrogen receptor are found in reproductive organs such as mammary gland, ovaries, vagina, and the uterus [1,2]. Oestrogen–oestrogen receptor binding activates the receptor. With activation, the receptor homodimerizes and binds oestrogen receptor elements in the promoter region of specific genes, thereby starting the process of target gene transcription [3,4]. The co-activators and co-repressors are involved in initiation and regulation of gene transcription by the oestrogen receptor [5]. Such co-activators are believed to determine the agonistic and antagonistic properties of

treated with tamoxifen also accumulated 125I up to 50% higher than did non-treated cells. Potassium perchlorate completely inhibited 125I uptake. When MDA-MB231 cells, the oestrogen receptor-negative breast cancer cells, were transfected with pERE-NIS, 125I uptake of MDA-MB-231/pERE-NIS did not increase. Conclusion This pERE-NIS reporter system is sufficiently sensitive for monitoring transcriptional activity of the oestrogen receptor. Therefore, cis-enhancer reporter systems with ERE will be applicable to the development of a novel selective oestrogen receptor modulator with low toxicity and high efficacy. Nucl Med Commun 27:773–777

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:773–777 Keywords: breast cancer, cis-enhancer reporter system, oestrogen receptor, sodium iodide symporter (NIS) a Laboratory of Nuclear Medicine, Korea Institute of Radiological and Medical Science, bDepartment of Nuclear Medicine and cCancer Research Institute, Seoul National University College of Medicine, Korea.

Correspondence to Dr June-Key Chung, Department of Nuclear Medicine, Seoul National University College of Medicine, 28 Yongon-dong, Chongno-gu, Seoul, 110-744, Korea. Tel: + 0082 2 2062 3376; fax: + 0082 2 745 7690; e-mail: [email protected] Received 2 February 2006 Accepted 28 June 2006

compounds and to be responsible for tissue-specific action of selective oestrogen receptor modulator (SERM)-like tamoxifen or raloxifene [6]. The thyroid gland concentrates iodide by a factor of 20– 40 compared to its plasma level [7]. Thyroid follicular cells transport iodide through a specific transporter, the sodium iodide symporter (NIS), the gene of which was identified in 1996 [8]. NIS is an intrinsic membrane protein with 13 transmembrane domains [9]. Because of uptake characteristics of 99mTc as well as radioiodine by NIS expression, NIS is already proposed to imaging reporter gene [10,11]. There are many advantages of NIS as an imaging reporter gene because of the wide availability of its substrates such as radioiodine and 99m Tc and the well-understood metabolism and clearance

c 2006 Lippincott Williams & Wilkins 0143-3636

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774 Nuclear Medicine Communications 2006, Vol 27 No 10

of these substrates in the body. The complicated radiolabelling was not required compared to PET reporter genes, including HSV1-tk and the dopamine D2 receptor [12]. A cis-enhancer reporter system has been used to monitor transcriptional activity of endogenous gene expression, intracellular signal transduction and nuclear receptor [11,13–15]. In this study, we developed an evaluation system for transcriptional activity of the oestrogen receptor using a cis-enhancer reporter method and sodium iodide symporter (NIS) as a reporter gene.

Materials and methods Construction of the reporter gene, and chemicals

The hNIS gene containing FL*-hNIS/pcDNA3 was provided by Dr S. Jhiang (Ohio State University, Columbus, USA) and pERE-TA-SEAP, including artificial oestrogen responsive element (ERE) was purchased from Clontech (Palo Alto, California, USA). Cytomegalovirus promoter fragment from FL*-hNIS/pcDNA3 was removed by treatment with NruI and HindIII (New England Biolabs, Beverly, Massachusetts, USA). The ERE sequence from pERE-TA-SEAP was amplified by using the polymerase chain reaction (PCR) technique with the PCR primers ERE1-NruI(A) GTGTTCGCGACGGGAGGTA CTTGGAGCG and ERE1-HindIII(AS) CCGAAGCTTC CATTATATACCCAGATCTAG. The amplified ERE sequence was inserted into NruI and HindIII digested FL*-hNIS/pcDNA3 and this transgene was named as pERE-NIS. 17b-Oestradiol and tamoxifen were purchased from Sigma Chemical Company (St. Louis, Missouri, USA) and dissolved in DMSO. These were stored at 201C and protected from light. All chemicals, buffers and solvents were of analytical grade, and unless otherwise stated, were purchased from Sigma. 125I in the form of sodium iodide was purchased from NEN (Boston, Massachusetts, USA). Cell culture and transfections

Human breast cancer cells MCF-7 and MDA-MB-231, which are oestrogen receptor-positive and oestrogen receptor-negative cell, respectively, were obtained from the Korea Cell Line Bank, and maintained as recommended. These cells were grown as a monolayer in RPMI 1640 medium (JBI, Daegu, Korea) supplemented with 292 mg
ml – 1 glutamine, 100 000 IU
l – 1 penicillin (GIBCO, Grand Island, New York, USA), 100 mg
l – 1 streptomycin (GIBCO), and 10% fetal bovine serum (FBS). Plasmid was transfected into cells using lipofectamine plus reagent (Invitrogen, Carlsbad, California, USA) according to the manufacturer’s instructions. Stable transfectants were selected with 600 mg
ml – 1 of Geneticin (Invitrogen) in RPMI 1640 containing 10% fetal bovine serum for 3 weeks.

Radioiodine uptake assay

Cells were placed in 24-well plates and cultured with RPMI 1640 containing 10% FBS. When the cells reached confluence, they were treated with several doses of 17boestradiol or tamoxifen for 18 h. After the cells had been incubated with the drugs, 125I uptake levels were determined. Iodide uptake assays were performed using a modification of the method described by Nakamoto et al. [16]. Cells were incubated at 371C for 30 min in 500 ml of Hanks balanced salt solution (HBSS) containing 0.5% bovine serum albumin and 10 mmol
l – 1 of the sodium salt of 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid, pH 7.4, with 18.5 kBq (0.5 mCi) carrierfree Na125I and 10 mmol
l – 1 of NaI, to yield a specific activity of 740 MBq
mmol – 1 (20 mCi
mmol – 1). After incubation, the cells were then quickly washed with 3 ml of ice-cold HBSS. The cells were detached with 0.05% SDS and the radioactivity was measured with a gamma counter (Cobra II, Canberra Packard, USA). Unless otherwise stated, the iodide uptake was expressed as pmol per 106 cells. In order to modulate the iodide uptake, the cells were incubated for 30 min in either Na125I medium or Na125I medium supplemented with 50 mmol
l – 1 potassium perchlorate [17]. All data points were measured in triplicate, and are displayed as means ± SEM.

Results Oestrogen receptor activity as a transcription factor is presented by binding with oestrogen or SERMs. To determine the oestrogen receptor activation by oestrogen or SERM binding using a cis-enhancer reporter system, pERE-NIS in breast cancer cells, we used two cell lines, MDA-MB231 and MCF-7. Because MDA-MB231 cells are oestrogen receptor-negative, iodide uptake of pERENIS transfected MDA-MB231 treated with 17b-oestradiol or tamoxifen was not increased (Fig. 1). However, when MDA-MB231 cells were transfected with pCMVNIS, iodide uptake of the transfected cells was enhanced and normal functionality of NIS was shown. The iodide uptake of pERE-NIS transfected MCF-7 cells (which express the oestrogen receptor) treated with 17boestradiol was increased by up to two-fold than that of non-treated cells (Fig. 2(A)). Iodide uptake was completely inhibited by 50 mmol
l – 1 of potassium perchlorate treatment, a specific inhibitor of NIS. Tamoxifen treatment of pERE-NIS-transfected MCF-7 cells also induced iodide uptake in the same manner as 17boestradiol (Fig. 2(B)). When cells were incubated with increasing concentration of reagents, NIS activity also was increased in a dose dependent manner (Table 1). After MCF-7 cells had been transfected with pERE-NIS, four independent clones of the stable cell line were selected with treatment of G418 for 14 days. MCF7/ pERE-NIS treated with 17b-oestradiol also accumulated

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Evaluation of oestrogen receptor activity with NIS Hyun Kang et al. 775

radioiodine up to 50% higher than did non-treated cells (Fig. 3).

Discussion Oestrogen has crucial roles in the proliferation of cancer cells in reproductive organs such as breast and uterus [18,19] and oestrogen-stimulated growth of cells is mediated by the oestrogen receptor. Approximately 70% of human breast tumours express higher amounts of the oestrogen receptor than does normal breast tissue, and oestrogen receptor expression reflects the prognosis of disease. Lower oestrogen receptor expression in breast Fig. 1

125l

uptake (pmol/106 cells)

150

40 20 0 pCMV-NIS No

E-0.1

E-1

T-5

T-10

125

I uptake in pERE-NIS-transfected MDA-MB231 cells treated with either oestradiol or tamoxifen. 125I uptake by these cells was not increased. pCMV-NIS: NIS gene expression controlled by the CMV promoter; No: DMSO only; E-0.1: 0.1 nmol
l – 1 oestradiol; E-1: 1 nmol
l – 1 oestradiol; T-1: 1 nmol
l – 1 tamoxifen; T-10: 10 nmol
l – 1 tamoxifen.

tumours often reveals a more aggressive phenotype [20]. Hormone therapy with tamoxifen, a selective oestrogen receptor modulator, is recommended for oestrogen receptor-positive breast cancer in the clinic [21,22]. Therefore, measurement of oestrogen receptor status in breast cancer tissue is important when deciding upon a therapeutic strategy for patients with breast cancer. There have been several methods for demonstrating the presence and function of the oestrogen receptor in tissues. In vitro oestrogen receptor assays such as immunohistochemistry and radioreceptor assays have been used clinically to measure the existence of the oestrogen receptor, but the techniques require tissue that is of limited availability. PET images can be used to assess the in vivo functional integrity of the oestrogen receptor using 16a-[18F]fluoroestradiol at sites of primary breast carcinoma as well as distant metastates [23]. A molecular imaging method using the cis-enhancer reporter system developed in this study can demonstrate the transcriptional activity of a nuclear oestrogen receptor in vivo. This approach to molecular imaging can be used to evaluate the early stages of the oestrogen receptor and to develop new SERMs. In this study, we developed a cis-enhancer reporter system for monitoring the transcription activity of the oestrogen receptor by determining 17b-oestradiol or tamoxifen binding to the receptor using NIS as a reporter gene. We constructed a reporter gene in which the NIS expression was regulated under the control of enhancer regulatory elements that are responsive to 17b-oestradiol binding. It has been reported that the cis-enhancer reporter system was applied to monitoring of endogenous gene expression and nuclear receptor activity such as retinoic acid [11,13–15].

Fig. 2

125I

uptake (pmol/106 cells)

(a)

1000

(b)

1000

800 300 200

200 150 100

100 0 pCMV-NIS No E-0.1 E-1 E-5 KCIO4

50 0 pCMV-NIS No T-1 T-10 T-50 KCIO4

125 I uptake in pERE-NIS transfected MCF-7 cells treated with either oestradiol (a) or tamoxifen (b). 125I uptake into these cells was increased by up to 70–80% compared to that in untreated cells. 125I uptake was completely inhibited by potassium perchlorate treatment. pCMV-NIS: NIS gene expression is controlled by CMV promoter; No: DMSO only; E-0.1: 0.1 nmol
l – 1 oestradiol; E-1: 1 nmol
l – 1 oestradiol; E-5: 5 nmol
l – 1 oestradiol; T-1: 1 nmol
l – 1 tamoxifen; T-10: 10 nmol
l – 1 tamoxifen; T-50: 50 nmol
l – 1 tamoxifen; KClO4: NIS inhibitor and 5 nmol
l – 1 oestradiol or 50 nmol
l – 1 tamoxifen-treated group.

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776 Nuclear Medicine Communications 2006, Vol 27 No 10

Table 1

Sodium iodide symporter (NIS) activities in pERE-NIS transfected MCF-7 cells treated with oestradiol or tamoxifen Oestradiol-treated group 1

Conc (nmol
l ) NIS activity (pmol per 106 cells) Activity ratio vs untreated group

Tamoxifen-treated group

0

0.1

1

5

5 + KClO4

0

101.4 ± 3.8

130.3 ± 33.2

187.0 ± 0.4

202.3 ± 2.2

100.4 ± 8.9

95.3 ± 3.6

1

1.29

1.84

2.00

1.1

1

125I

uptake (pmo1/106 cells)

Fig. 3

2000

No reagent E-0.1nM

1600

E-1nM 1200 800

1

10

104.5 ± 14.4 124.1 ± 13.9 1.10

1.30

50

50 + KClO4

166.1 ± 2.0

96.2 ± 5.7

1.74

1.00

fication (TSTA) system could be applied to enhance the imaging signal. In TSTA, expression of a potent transcriptional activator such as GAL4-VP16 is controlled by weak promoter or enhancer. The expressed activator binds to several GAL4 binding sites which drive the expression of reporter or therapeutic gene [24,25]. The activity of prostate-specific antigen promoter and carcinoembryonic antigen promoter could be enhanced by the TSTA system [24,26].

400 0

Acknowledgement 1

5 6 Stable Clones

13

125

I uptake by MCF-7 cells stably expressing NIS under the control of the oestrogen responsive element (ERE). MCF-7/pERE-NIS cells treated with 17b-oestradiol also accumulated up to 50% more 125I than did untreated cells. Unfilled bars: no reagent; grey bars: 0.1 nmol
l – 1 oestradiol; filled bars: 1 nmol
l – 1 oestradiol.

This study was supported by grant number 03-2003-007-0 from the Seoul National University Hospital Research Fund (2003), Republic of Korea, and J.H. Kang, J.-K. Chung, Y.J. Lee, K.I. Kim were supported by the BK21 project for Medicine, Dentistry, and Pharmacy (2006).

References 1 2

When MCF-7 cells, human breast cancer cells that highly express oestrogen receptor-a, were transfected with pERE-NIS and treated with 17b-oestradiol or tamoxifen, 125 I uptake was increased to a greater extent than in untreated cells. After the cells had been treated with serial doses of 17b-oestradiol, the uptake of 125I was enhanced according to increasing concentrations. Although NIS activity was not determined in this study, it should be mediate the change of 125I uptake in the cells. NIS activity of MCF7/pERE was completely blocked with potassium perchlorate, an NIS specific inhibitor. MDA-MB231 cells are oestrogen receptor negative, and 125I uptake of pERE-NIS transfected MDA-MB231 did not increase in spite of treatment of 17b-oestradiol.

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Although the pERE-NIS reporter system is sufficiently sensitive for monitoring oestrogen receptor activity in vitro, NIS expression imaging with tumour-bearing mice by a gamma camera was scarcely being acquired in this study (data not shown). We suspected there was little discrepancy in NIS activity between drug-treated and untreated samples in vivo. To acquire a nuclear medicine image using a gamma camera, it is necessary to augment the transcriptional activity of the oestrogen responsive element. For example, a two-step transcriptional ampli-

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Original Article

Evaluating dopamine transporter activity with 99m Tc-TRODAT-1 SPECT in drug-naive Tourette’s adults Chin-Bin Yeha, Chiang-Hsuan Leeb, Yuan-Hwa Chouc, Chia-Jung Changd, Kuo-Hsing Mae and Wen-Sheng Huangd Objectives Findings on imaging of dopamine transporter (DAT) activity in patients with Tourette’s syndrome remain inconclusive. The present study was carried out to observe DAT activity in patients with well-controlled Tourette’s syndrome by using 99mTc-TRODAT-1 single photon emission computerized tomography (SPECT). Methods Six drug-naive patients with Tourette’s syndrome (mean age ± SD, 21.2 ± 1.5 years) were recruited. All met the criteria for Tourette’s syndrome established in the DSM-IV. Seventeen age-matched and sex-matched healthy subjects served as the controls. Brain SPECT were acquired 165–195 min after administrating 740 MBq of 99m Tc-TRODAT-1, using a double-headed camera equipped with ultra-high-resolution fan-beam collimators. The specific uptake ratio was calculated by subtracting the mean counts per pixel in the occipital cortex from the mean counts per pixel in the striatum, putamen or caudate nucleus and by dividing the result by the mean counts per pixel in the occipital cortex. Tic-severity scores were also measured and correlated with the specific uptake ratios. Results No significant difference in DAT activity between patients with Tourette’s syndrome and control subjects was found in the striatum and its sub-regions. Tic-severity

Introduction Tourette’s syndrome is a neuropsychiatric disorder of childhood onset characterized by involuntary motor and phonic tics. Substantial evidence indicates that the dopaminergic system is involved in the pathophysiology of Tourette’s syndrome [1]. This is also indicated by the fact that dopamine-blocking drugs [2] reduce tic symptoms, whereas dopaminergic agents [3] enhance them. The dopamine transporter (DAT) plays an important role in regulating dopamine neurotransmission in humans [4]. Pre-synaptic neurons are thought to control extracellular dopamine by means of the DAT. Several studies of the pre-synaptic dopaminergic function of the basal ganglia have been conducted in patients with Tourette’s syndrome. However, the results are controversial [5,6]. Post-mortem study has demonstrated increased 3Hmazindol binding to the DAT [7]. A study based on 123 I-b-CIT single photon emission computerized tomo-

scores were also not correlated with specific uptake ratios measured from the striatum and its sub-regions. Conclusions In conjunction with previous findings, our results suggested that functional abnormality of the dopamine system in patients with Tourette’s syndrome might be evident only in its early stage. Adaptation to tic symptoms might play a role in regulating the neural c 2006 Lippincott system. Nucl Med Commun 27:779–784 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:779–784 Keywords: syndrome

99m

TC-TRODAT-1 SPECT, dopamine transporter, Tourette’s

a Departments of Psychiatry, dNuclear Medicine, eAnatomy and Biology, National Defense Medical Center, Taipei, bDepartment of Nuclear Medicine, Chi-Mei Medical Center, Tainan and cDepartment of Psychiatry, Taipei Veterans General Hospital, Taiwan, ROC.

Correspondence to Dr Wen-Sheng Huang, Department of Nuclear Medicine, Tri-Service General Hospital, National Defense Medical Center, No. 325, Sec. 2, Cheng-Kung Rd., Neihu, Taipei, Taiwan, 114 ROC. Tel: + 00886 287 927374; fax: + 00886 287 927217; e-mail: [email protected] Received 5 February 2006 Accepted 28 June 2006

graphy (SPECT) of DAT binding sites in the plasma membrane showed increased binding of the DAT ligand in Tourette’s syndrome [5]. In addition, accumulation of 18 F-DOPA in the left caudate nucleus was higher in children with Tourette’s syndrome than in control subjects [8]. Using 123I-IPT SPECT, investigators have also reported an increased DAT-binding ratio in drugnaive children with Tourette’s syndrome compared with healthy children [9]. In contrast, some studies revealed no significant difference in DAT density in the basal ganglia, thalamus or midbrain of patients with Tourette’s syndrome [10,11]. Moreover, binding of the vesicular striatal monoamine transporter type-2 (VMAT2) did not significantly differ between adults with Tourette’s syndrome and healthy controls [12]. Therefore, current data from SPECT or positron emission tomography (PET) studies of DAT activity in patients with Tourette’s syndrome are still inconclusive.

c 2006 Lippincott Williams & Wilkins 0143-3636

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Cocaine analogues, such as 123I-labeled b-CIT, FP-CIT and 99mTc-labelled TRODAT-1, have been developed as SPECT radioligands for the evaluation of DAT activity. Because of the inconvenience and cost of 123I-labelled DAT ligands and the prolonged waiting time for imaging (e.g., 18–30 h after the injection of 123I-b-CIT), the use of 99mTc-labelled radioligands seems to be suitable for large-scale clinical use [13,14]. On the other hand, medical effects on the dopaminergic system confound imaging results. To improve our understanding of the relationship between DAT activity and the clinical status of patients with Tourette’s syndrome, we performed DAT SPECT by using 99mTc-TRODAT-1 in six drug-naive patients with Tourette’s syndrome.

Materials and methods Patients and control subjects

Six men with Tourette’s syndrome (mean age ± SD, 21.2 ± 1.5 years) who visited the department of psychiatry for the physical and mental examinations required for compulsory military service were recruited. All met the criteria for Tourette’s syndrome established in the Diagnostic and Statistical Manual of Mental Disorders, fourth edition (DSM-IV), and all provided informed consent. Our institutional review board for human research approved this study. None of the subjects had been exposed to psychotropic drugs or had a history of brain damage, convulsive disorder, developmental difficulties, or medication within 4 weeks before the study. They were interviewed for the history and current status of tic symptoms, and treatment history. Family members reported that patients were drug-naive to antipsychotic treatment. Their mean severity of disease was 25.2, as rated by using the Yale Global Tic Severity Scale (YGTSS) [15]. The men were interviewed and evaluated using the DSM-IV to assess for co-morbid diagnoses of attention deficit hyperactive disorder (ADHD) and obsessive–compulsive disorder (OCD). Seventeen age-matched healthy subjects (mean age ± SD, 21.7 ± 1.1 years) served as the controls. Imaging and data analysis

Brain SPECT studies were acquired 165–195 min after the intravenous injection of 740 MBq of 99mTc-TRODAT1. We used a double-headed camera equipped with ultrahigh-resolution fan-beam collimators (Helix SPX; Elscint, Haifa, Israel). Data were acquired in a 128  128 matrix with 1.4 zoom through 3601 rotation (1801 for each head) at 31 intervals for 30 s per angle step. Images were reconstructed by using linear back-projection with a modified Metz filter (power factor, 3.5; cut-off frequency, 0.28 cm – 1). Attenuation correction was performed by applying Chang’s first-order method (attenuation coefficient m = 0.12 cm – 1). SPECT images were analysed along the level of the canthomeatal line. Regions of interest

(ROIs) were marked, with reference to the corresponding magnetic resonance image, for the right and left caudate, putamen and striatum as described [14]. In brief, regions were marked on composite images of the three sections depicting the basal ganglia with the highest level of activity. The pixel size was 1.72  1.72 mm, and the section thickness was 3.4 mm. Regions of interest on one side were then fitted to the other side. The occipital cortices were drawn in the same way and served as background areas. The specific uptake ratio (SUV) was calculated by subtracting the mean counts per pixel in the occipital cortex from the mean counts per pixel in the whole striatum, putamen or caudate nucleus and by dividing the result by the mean counts per pixel in the background, as follows: (target–occipital cortex)/occipital cortex, where target represents the striatum, putamen or caudate nucleus. Statistical analysis

Analysis of variance was performed to compare groups using software (SPSS version 13.0; SPSS Inc., Chicago, Illinois) for Windows (Microsoft, Redmond, Washington). Significance was defined as P < 0.05. Results are reported as the mean ± SD.

Results Table 1 shows the demographic data of the patients and control subjects. For patients, the mean duration of Tourette’s syndrome was 12.8 ± 1.6 years, and the mean YGTSS score for total tic severity was 25.2 ± 5.9. On imaging, binding ratios on both sides of the basal ganglia were not significantly correlated with overall tic severity (right side, correlation coefficient, R = – 0.23 and P = 0.66; left side, R = – 0.17 and P = 0.75). No correlation was observed between patients’ tic-severity scores and uptake ratios, as measured either globally or different subregions of the basal ganglia. Multiple-comparison analyses of findings from different locations of basal ganglia between patients with Tourette’s syndrome and control subjects showed no significant difference (Table 2). There was no significant difference of DAT binding, i.e., SUV, between Tourette’s syndrome patient and healthy controls (Fig. 1). The representative 99mTc-TRODAT-1 SPECT at the level of basal ganglia in a Tourette’s syndrome patient and a healthy subject was shown in Fig. 2. Sub-scale scores on the YGTSS, such as scores for motor, phonic, and total tic severity, were also not correlated with specific uptake ratios for DAT binding (Table 3).

Discussion To our knowledge, we are the first to use 99mTcTRODAT-1 SPECT to compare DAT activity in patients with Tourette’s syndrome with homogeneous clinical

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Tc-TRODAT-1 in drug-naive Tourette’s adults Yeh et al. 781

Table 1 Patient number

1 2 3 4 5 6

Clinical manifestation and specific DAT uptake ratios in Tourettes syndrome patients and in controls Diagnosis

TS TS TS, ADHD TS, ADHD TS TS

Specific DAT uptake ratios

YGTSS score

RC

RP

RS

LC

LP

LS

Total

Age (years)

Illness duration

Motor tics

Phonic tics

Total tics

2.56 3.09 3.19 2.61 2.97 2.77

2.10 2.59 2.60 2.45 2.58 2.72

2.33 2.84 2.89 2.53 2.77 2.74

1.97 3.05 3.07 2.52 2.90 3.04

1.95 2.59 2.83 2.23 2.54 2.75

1.96 2.82 2.95 2.38 2.72 2.90

2.14 2.83 2.92 2.45 2.74 2.82

23 21 20 22 22 19

14 14 12 14 13 10

15 17 10 13 12 16

14 15 8 9 8 14

29 32 18 22 20 30

DAT = dopamine transporter; YGTSS = Yale Global Tic Severity Scale; RC = right caudate; RP = right putamen; RS = right striatum; LC = left caudate; LP = left putamen; LS = left striatum, Total = bilateral basal gangalia; TS = Tourette’s syndrome; ADHD = attention deficit–hyperactivity disorder.

Table 2 Comparison of specific DAT uptake ratios between patients with Tourette’s syndrome and control groups in the striatum and its sub-regions Group

Specific DAT uptake ratios

Tourette’s Syndrome Mean (SD) (n = 6) Control Mean (SD) (n = 17) ANOVA

RC

RP

RS

LC

LP

LS

Total

2.87 (0.26)

2.51 (0.22)

2.68 (0.21)

2.76 (0.44)

2.48 (0.33)

2.62 (0.38)

2.65 (0.30)

3.02 (0.64)

2.40 (0.55)

2.71 (0.58)

3.11 (0.75)

2.44 (0.51)

2.77 (0.62)

2.74 (0.59)

F = 0.31 P = 0.59

F = 0.22 P = 0.65

F = 0.01 P = 0.92

F = 1.15 P = 0.30

F = 0.04 P = 0.85

F = 0.31 P = 0.58

F = 0.13 P = 0.72

DAT = dopamine transporter; RC = right caudate; RP = right putamen; RS = right striatum, LC = left caudate, LP = left putamen; LS = left striatum; Total = bilateral striatum.

Fig. 1

Fig. 2

3

2

Normal control 1

TS group

Normal control TS group

Representative transverse images of the 99mTc-TRODAT-1 DAT SPECT showed dopamine transporter activity in an age-matched and gendermatched normal subject and a patient with Tourette’s syndrome. No apparent difference in the uptake of 99mTc-TRODAT-1 in the basal ganglia was found between the TS patient and normal control.

0 R't R't R't L't L't L't Bil Caudate Putamen Striatum Caudate Putamen Striatum Striatum

Comparison of specific dopamine transporter (DAT) uptake ratios between patients with Tourette’s syndrome and normal controls in the striatum and its different sub-regions.

characteristics to that of sex-matched and age-matched healthy subjects. The literature describes a 38-year-old woman with symptoms of both ADHD and Tourette’s

syndrome. In this patient, striatal DAT density was elevated by 24% compared with values in 14 healthy subjects, as determined by using 99mTc-TRODAT-1 imaging [16]. However, in our study, the DAT binding activity was not significantly different between patients with Tourette’s syndrome and control subjects. Two 123I-b-CIT SPECT studies showed no difference between patients and control subjects in terms of binding

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782 Nuclear Medicine Communications 2006, Vol 27 No 10

Table 3

Correlations of tic-severity sub-scores with specific DAT uptake ratios in the striatum and its sub-regions Region

R value P value

RC

RP

RS

LC

LP

LS

Total

– 0.28 0.59

– 0.13 0.81

– 0.23 0.66

– 0.15 0.78

– 0.20 0.70

– 0.17 0.75

– 0.19 0.72

DAT = dopamine transporter; RC = right caudate; RP = right putamen; RS = right striatum; LC = left caudate; LP = left putamen; LS = left striatum; Total = bilateral striatum.

ratios for striatal DAT molecules [10,11]. In another study, researchers used PET with 11C-dihydrotetrabenazine to measure binding to striatal VMAT2 and also found no evidence of increased binding in patients with Tourette’s syndrome [12]. However, others found significant increases in striatal DAT binding in patients with Tourette’s syndrome compared with healthy controls [1,5–9,17]. Additional work with well-controlled case analyses is thus needed to gain insight into the relationship between DAT activity and clinical manifestations in patients with Tourette’s syndrome. Tourette’s syndrome is recognized as a complex neuropsychiatric spectrum disorder in which tics are only one of many symptoms. The degree of homogeneity among patients may be low. Selection bias may be responsible for the association between increased striatal 123I-b-CIT uptake and self-injurious behaviour and poor impulse control, as 67% of the population with Tourette’s syndrome have these two features [18]. Various DAT activities observed in previous studies of patients with Tourette’s syndrome indicated that their characteristics were heterogeneous. In one study, five of 12 patients had DAT activities more than 2 SDs above the mean of control subjects [18]. If we consider that most neuroimaging studies performed by using radioligands in adults with Tourette’s syndrome, compensatory changes might have occurred as the disease progressed [19]. Levels of exposure to neuroleptics also vary among individuals with Tourette’s syndrome [8,13]. A 123I-IPT SPECT study of children with Tourette’s syndrome aged 6–12 years showed that their DAT activity was increased compared with that of age-matched and sex-matched control subjects [9]. In this study, although all of the patients were drug-naive, their emotional status during scanning might not be well controlled. Besides differences in the characteristics of patients with Tourette’s syndrome, inconsistencies in DAT activity might also be related to variations in imaging methods including the use of radioligands and cameras [20–24] and to techniques of data acquisition and analysis [25]. Both the occipital cortex and the cerebellum have been used as reference regions to analyse imaging data. It is of interest that DAT activity did not differ between patients and controls when the cerebellum was referenced

[13,14]. Both 123I-b-CIT and 123I-FP-CIT have higher signal-to-background ratios than that of 99mTc-TRODAT1 [1–3]. Even so, the 99mTc-labelled ligands are practical because 99mTc is the most common tracer used in clinical settings. It provides a suitable half-life and energy for imaging, at a relatively low cost and is readily available. 99m Tc-TRODAT-1 has been widely used in the clinical evaluation of patients with early Parkinson’s disease and during its progression [4], and of patients with parkinsonism, even in the subclinical stages [18,24,26], with acceptable reproducibility [27]. However, Tourette’s syndrome is a neurodevelopmental disorder that begins at a young age. In patients with Tourette’s syndrome, changes in DAT binding activity may be less obvious than those observed in neurodegenerative disorders such as Parkinson’s disease. Higher mean binding ratios with 123I-FP-CIT and 123I-bCIT than with 99mTc-TRODAT-1 were reported both in patients with Tourette’s syndrome and in control subjects [2]. Respective mean ratios for 123I-FP-CIT and 123I-bCIT were 6.5 and 10.5 in patients with Tourette’s syndrome and 6.2 and 6.3 for control subjects [5]. Another 123I-b-CIT study showed that mean binding ratios were 13.6 for the Tourette’s syndrome group and 9.4 for the control group [6]. Although DAT uptake was relatively high in the younger subjects [28], 99mTcTRODAT-1 data as shown in the present study still showed relatively lower mean binding ratios (the Tourette’s syndrome group, 2.7 and the control group, 2.7 also) than those of the aforementioned reports. Therefore, it might be not apparent enough for the subtle changes of DAT binding in Tourette’s syndrome patients measured by 99mTc-TRODAT-1 SPECT as seen in our study. Additionally, the DAT may–although not necessarily–be changed in patients with Tourette’s syndrome; and the changes appear to be dependent more on clinical presentations, such as self-injurious behaviour or a lack of impulse control than the disease per se [6]. Our patients did not present the above-mentioned clinical parameters or any that were similar. DAT activity might be investigated in young subjects because DAT density decreases by approximately 5% per decade of age [5]. Neuropathological phenomena should

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Tc-TRODAT-1 in drug-naive Tourette’s adults Yeh et al. 783

be observed at the onset of illness in order to best understand the underlying pathophysiology of developmental neuropsychiatric disorders such as Tourette’s syndrome. Measuring DAT activity in the drug-naive state is also desirable because medications used to treat the syndrome block dopaminergic interaction and thus may affect DAT activity. Accordingly, we recruited relatively young Tourette’s syndrome patients (mean age, 21 years) with a drug-naive status. Except for the possible medical interference, disease severity per se may also be responsible for changes of DAT binding seen on imaging studies. The patients had modest disease duration (mean, 12.8 years) and severity (YGTSS, 25). In view of previous studies, the mean ages of Tourette’s syndrome patients were around 30–40 years [5,6,10,11,17]. Two showed no difference in DAT binding between Tourette’s syndrome patients and controls [10,11] whereas the others showed elevated DAT bindings in Tourette’s syndrome patients [5,6,17]. Since 50% of Tourette’s syndrome patients will show a reduction in the symptoms as they grow up [1], those older than 30 years and with persistent tics may represent more severe neuropathology than those of young adults (mean age, 21 years) and thus can be easily detected by imaging studies. Two younger Tourette’s syndrome groups (mean ages 9.9 and 15.2 years, respectively) [8,9] were reported to have elevated DAT binding. It thus appears that there is an ‘imaging window’ in which TRODAT-1 might be able to distinguish subjects with Tourette’s syndrome from normal subjects. We speculated that younger children in whom tics had just begun may have more prominent image signals in the lesion area without interference from background brain regions in neuroimaging examinations. However, in the current study, drug-naive Tourette’s syndrome patients who visited us for the physical examinations might have less severe tic symptoms because they had not needed medical intervention in the previous 10 years. The results of the current study suggested that the tics symptoms might be improved in younger Tourette’s syndrome patients who showing indifferent TRODAT-1 binding compared with normal controls. Future longitudinal studies are warranted to investigate the usefulness of DAT binding with TRODAT-1 imaging in predicting the prognosis or therapeutic response of Tourette’s syndrome patients. Limitations of the study

Current findings might not be of further value for clarifying the inconsistent results of DAT binding in Tourette’s syndrome patients because of the limited number of patients or the relatively long disease course. However, our study is the first to use the novel radioligand 99mTc-TRODAT-1 to measure the DAT activity in patients with neurodevelopmental disorders such as Tourette’s syndrome. While our results need further support, the study does illustrate a clinically

feasible means of demonstrating the interplay between changes of DAT binding and neurodevelopmental disorders.

Conclusion Our findings with a limited sample size and longer disease duration implied no significant change of the striatal DAT activity in Tourette’s syndrome patients compared with control subjects, suggesting that abnormalities of the dopaminergic system in Tourette’s syndrome patients might be evident only in the early stages, i.e., in the absence of chronic compensatory change [6]. Adaptation to tic symptoms might play a role in regulating the neural system [7]. Future studies should thus be designed to observe neuropathological phenomena as early as possible after onset of the illness to determine the pathophysiology underlying developmental neuropsychiatric disorders such as Tourette’s syndrome.

Acknowledgements This work was supported by grants NSC 94-2623-7-016007, 94-2314-B-016-001 and 95-CMNDMC07 from National Science Council and National Defense Medical Center. The authors thank Dr C. Oliver Wong for his editorial comments.

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Ma KH, Huang WS, Chen CH, Lin SZ, Wey SP, Ting G, et al. Dual SPECT of dopamine system using [99mTc]TRODAT-1 and [123I]IBZM in normal and 6-OHDA-lesioned formosan rock monkeys. Nucl Med Biol 2002; 29:561–567. Asenbaum S, Brucke T, Pirker W, Podreka I, Angelberger P, Wenger S, et al. Imaging of dopamine transporters with iodine-123-beta-CIT and SPECT in Parkinson’s disease. J Nucl Med 1997; 38:1–6. Van Laere K, De Ceuninck L, Dom R, Van den Eynden J, Vanbilloen H, Cleynhens J, et al. Dopamine transporter SPECT using fast kinetic ligands: 123 I-FP-beta-CIT versus 99mTc-TRODAT-1. Eur J Nucl Med Mol Imaging 2004; 31:1119–1127. Weng YH, Yen TC, Chen MC, Kao PF, Tzen KY, Chen RS, et al. Sensitivity and specificity of 99mTc-TRODAT-1 SPECT imaging in differentiating patients with idiopathic Parkinson’s disease from healthy subjects. J Nucl Med 2004; 45:393–401. Peterson BS. Neuroimaging studies of Tourette syndrome: a decade of progress. Adv Neuro 2001; 185:179–196. Turjanski N, Sawle GV, Playford ED, Weeks R, Lammerstma AA, Lees AJ, et al. PET studies of the presynaptic and postsynaptic dopaminergic system in Tourette’s syndrome. J Neurol Neurosurg Psychiatry 1994; 57:688–692. Huang WJ, Yao WJ, Wey SP, Ting G. Reproducibility of Tc-99m TRODAT-1 SPECT measurement of dopamine transporters in Parkinson’s disease. J Nucl Med 2004; 45:207–213. Mozley PD, Acton PD, Barraclough ED, Plossl K, Gur RC, Alavi A, et al. Effects of age on dopamine transporters in healthy humans. J Nucl Med 1999; 40:1812–1817.

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Original article

Is it possible to refine the indication for sentinel node biopsy in high-risk ductal carcinoma in situ? Manel Frailea, Josep M. Gubernb, Miquel Rulla, Francisco J. Julia´na, Cristina Serrab, Mariona Llatjo´sa, Pere Culellc, Pere Puigd, Montse Sola`a, Virginia Vallejosa, Antonio Mariscala, Joan Janere, Pere Deulofeuf and Ferra`n Fuste´a Background The indication for sentinel node biopsy (SNB) has not been fully established yet for patients with ductal carcinoma in situ (DCIS).

positivity approached but did not reach statistical significance (P = 0.06); therefore a subset of further selected higher risk patients could not be defined.

Aim To relate the conversion rate to invasive carcinoma with sentinel node positivity in high risk DCIS, and to refine the clinical presentation analysis in order to better select patients for SNB. For this purpose, a risk score was devised.

Conclusion The relevance of SNB positivity cannot be overlooked in high-risk DCIS patients, however, because SNB is not free from morbidity and cost, more studies are needed to refine its final indication. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:785–789

Methods From 1998 to 2005, 151 high-risk DCIS patients from six clinical centres were included in a prospective sentinel node database. The conversion rate to invasive carcinoma was 39%. Ten of 142 (7%) successful SNBs showed a positive sentinel node (eight micrometastatic). The sentinel node was positive in 1% of pure DCIS, in 5.5% of DCIS with micro-invasion, and in 19.5% of invasive carcinoma.

Nuclear Medicine Communications 2006, 27:785–789

Results Both clinical presentation and corresponding risk score were closely related to conversion to invasive carcinoma. The association of risk score and sentinel node

Introduction Because ductal carcinoma in situ (DCIS) of the breast carries no risk of metastatic spread, either systemic or lymphatic, it would seem appropriate not to perform axillary lymph node dissection (ALND) or even sentinel node biopsy (SNB) [1]. However, some reports in the literature have shown positive SNB results in DCIS patients [2–5]. On the other hand, during the preoperative diagnostic work-up, however complete it may be, there is a well known risk of underestimating the presence of invasive carcinoma. In fact, DCIS may be regarded as an array of conditions with different invasive and metastatic potential. Therefore, indiscriminate practice of SNB in every case of DCIS should be viewed as an obvious over-treatment, leading to increased morbidity and cost, and also precluding future SNB for lesions in the same breast. However, in well-selected DCIS patients with a high-risk profile for associated invasive disease, SNB may be the optimum choice to stage the axilla. The better the selection process, the wiser the SNB indication.

Keywords: ductal carcinoma in situ, sentinel node biopsy a Hospital Germans Trias i Pujol, Badalona, Barcelona, bHospital de Mataro´, Barcelona, cAlthaia, Manresa, Barcelona, dHospital Sant Jaume, Calella, Barcelona, eHospital Esperit Sant, Santa Coloma G, Barcelona and fHospital Municipal, Badalona, Barcelona, Spain.

Correspondence to Dr Manel Fraile, Medicina Nuclear, Hospital Universitari Germans Trias i Pujol, 08916 Badalona, Barcelona, Spain. Tel: + 0034 934 978957; fax: + 0034 934 978843; e-mail: [email protected] Received 13 February 2006 Accepted 5 April 2006

The aim of our study was to assess the results of SNB in high-risk DCIS patients, and also to relate such results to the rate of conversion to invasive carcinoma. High risk at presentation was defined according to well accepted criteria, including lesions that are large, multicentric or palpable, or that show high nuclear grade or comedonecrosis [6]. Additionally, we wished to refine the analysis of preoperative diagnostic criteria to further select a subset of patients on which to establish the final indication for SNB.

Patients and method Between June 1998 and October 2005, 151 patients with a preoperative diagnosis of high-risk DCIS were included in our prospective collaborative SNB database from six clinical centres (Hospital Germans Trias i Pujol, 69 patients Hospital de Mataro´, 35 patients; Althaia Manresa, 29 patients; Hospital Sant Jaume de Calella, eight patients; Hospital Municipal de Badalona, seven patients; and Hospital Esperit Sant, three patients). The mean age was 55 years (range, 3–76 years). A diagnosis of

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DCIS had been reached by core biopsy in 128 patients. Typically, under stereotactic guidance a 14-gauge cutting needle was used to retrieve between five and seven coretissue samples. In 23 patients the diagnosis was based on the presence of pure large microcalcifications (more than 3.5 cm) and the detection of malignant ductal cells at fine needle aspiration biopsy (FNAB). Among others, the main clinical variables included in our database were: presence of a palpable lump (10 patients), comedonecrosis (116 patients), nuclear grade (high grade 139 patients), and image diagnosis: either nodular lesion, architectural distortion, associated microcalcifications, or pure microcalcifications (pure microcalcifications 110 patients). Also, a secondary variable, called risk score, was generated by adding these four primary variables, each of which had been given a value of either 0 (absence) or 1 (presence). Therefore, the risk score could range from 1 to 4 in any particular case. As for image diagnosis variables, a value of 1 was given to patients with a presentation of nodule, architectural distortion or pure microcalcification greater tha 3.5 cm. Sentinel node biopsy was done as previously described [7]. The technique was based on the use of radiocolloids, without a surgical dye. From 1998 until 2000 the peritumoral injection route was used (14 patients; 9%). From that time on, intratumoral injection was used. The intratumoral injection was achieved by sterotactic technique in 37 cases. In 100 cases, sonography was used, including those cases of radiological microcalcifications with an associated focal sonographic abnormality, usually a hypoecogenic image. In 67 patients (44%), a subdermal supplemental dose of radiocolloid was associated. From September 2002 onwards (105 cases) routine preoperative axillary sonography was added to the procedure. From each referring clinical centre, patients were sent to the Nuclear Medicine Department at the Hospital Germans Trias i Pujol 4–24 h before surgery, where the clinical indication for SNB was checked. Then, injection of the radiocolloid was followed by a preoperative limphoscintigraphy in order to map the individual sentinel nodes. A small-particle colloid (99mTc-nanocolloid human albumin) was used in 25 patients (16%), while a larger colloid (99mTc-microcolloid albumin or 99mTc-precolloidal tin) was used in 126 patients (84%). The choice of colloids was generally based on patient age: under 65 years (large size) or over 65 years (small size). The surgical procedure was done 4-24 h post-injection, with the use of a portable gamma probe (Navigator USSC; RDM, Watertown, MA, USA). The assistance of an experienced nuclear physician from the main clinical centre (Hospital Germans Trias i Pujol) was always available in the operating field. The SNB biopsy was achieved at each referring centre using exactly the same procedures.

Definitive histological work-up was done on a standard basis, with special care to describe the presence of any infiltrative focus. Microinvasive carcinoma was defined as any focus of stromal invasion less than 1 mm in size. The sentinel nodes were processed with total paraffin embedding. Serial sectioning and H&E assessment was done by at least two independent pathologists. Negative or doubtful cases were then studied with cytokeratin (CK) immunohistochemistry. Micrometastases were defined as malignant cell clusters less than 2 mm. Statistical assessment included a descriptive analysis of the main variables in our database. Qualitative variables were described using frequency tables, while quantitative variables were described as the mean and standard deviation. For the comparison of qualitative variables the chi-squared or Fisher’s exact test were used. For the comparison between qualitative and quantitative variables, the dicotomic Student t-test was used. A P value of less than 0.05 was considered statistically significant. Data analysis was accomplished using the 11.5 Windows version of the SPSS statistical software package.

Results Along with the SNB, breast-conserving surgery was achieved in 113 patients, while mastectomy was performed in 38. Of the 151 patients, definitive histology revealed pure DCIS in 92 (61%), micro-invasive carcinoma in 18 (12%), and invasive carcinoma in 41 (27%). Thus, the conversion rate to any invasive disease was 39%. There were no statistical differences in conversion rates between core-biopsy patients and FNAB patients (38.7% vs. 47.6%, P = 0.567). As for the SNB, there was a failed procedure in nine cases (6%). Of the nine cases, three refused ALND. In six cases, ALND was performed, one of them showing a single metastatic lymph node. Ten out of the 142 successful procedures showed a positive sentinel node (7%). Altogether, 11 of the 151 (7.3%) were node positive. Only one of the 10 positive SNB cases displayed pure DCIS at histology, whereas eight showed invasive carcinoma and one a micro-invasive carcinoma. The rate of sentinel node positivity was 1% for pure DCIS, 5.5% for micro-invasive carcinoma, and 19.5% for invasive carcinoma. The case of positive sentinel node in pure DCIS was a patient presenting with a cluster of isolated microcalcifications 3 cm wide and a core biopsy showing highgrade in-situ carcinoma with comedonecrosis. Out of the 10 sentinel node-positive cases, eight had micrometastases. Of these, the patient with pure DCIS refused ALND. In the other seven cases, re-operative ALND was negative. Of the two cases with macrometastasis and infiltrative disease, ALND was positive in one.

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Refining the indication for SNB in ductal carcinoma Fraile et al. 787

Relationship between clinical variables at presentation, conversion rates to invasive carcinoma at definitive histology, and positivity of sentinel node biopsy (SNB)

Table 1

Table 2 Statistical association between the risk score and conversion rates to invasive carcinoma Risk score

Variable

Converted (%)

Non-palpable Palpable

36.9 70.0

Microcalcification Other image presentation No comedonecrosis Comedonecrosis No high nuclear grade High nuclear grade

36.4 46.3

Significance

Fisher, P < 0.05. w2, NS

Positive SNB (%) 3.8 50 5.7 10.8

Significance

1–2 3–4

Not converted (%)

Converted (%)

61 (67.8) 31 (50.8)

29 (32.2) 30 (49.2)

Significance Fisher test, P = 0.027

Fisher, P < 0.05. Fisher, NS

Table 3 Statistical association between the risk score and sentinel node biopsy positivity

25.7 43.1 25.0

w2, NS

3.3 8 0

Fisher, NS

Risk score

40.3

w2, NS

7.6

Fisher, NS

1–2 3–4

Sentinel node negative Sentinel node positive (%) (%) 80 (96.4) 52 (88.1)

3 (3.6) 7 (11.9)

Significance

Fisher test, P = 0.06.

Fig. 1

scores were also prone to sentinel node positivity, between-groups difference approached but did not reach statistical significance (P = 0.06) (Table 3) and, thus a selected subset of still higher risk patients could not be defined.

80

60

Discussion In patients with DCIS, the issue of axillary lymph-node evaluation remains quite controversial. Two recent literature reviews, one from workers at the US National Cancer Institute [8] and another from the Irish National Breast Cancer Screening Program [9], as well as the ASCO guidelines for SNB [10], all conclude that ALND can lead to over-treatment for patients with DCIS, and that even SNB is not broadly indicated in such clinical context. Although some authors have strongly argued against SNB [11], it could be used in the clinical management of selected high-risk DCIS patients, many of whom can harbour occult micro-invasion [12].

40

20

0 RS1

RS2

RS3

RS4

Percent distribution of risk scores (RS) at presentation and conversion to invasive carcinoma at definitive histology. (’) Not converted; (&) converted.

Table 1 displays the relationship between clinical presentation variables, the conversion rate to invasive disease, and SNB results. Although the difference between groups reached statistical significance only for patients with palpable lesions, a rather uncommon presentation, there was an obvious trend to invasive disease conversion and SNB positivity in patients with high nuclear grade, comedonecrosis, and image diagnosis other than pure microcalcification. As for the risk score (Fig. 1, Table 2), patients with higher values also showed significantly higher rates of conversion. However, even though patients with high risk

The aim of our own investigation was to establish the yield of SNB in a selected population of patients with ductal carcinoma in situ. A high-risk profile was defined in our patients population according to well-established clinico-pathological and radiological criteria, including palpable lesions, high cytological grade, comedonecrosis, large suspicious microcalcifications and/or other image presentation such as architectural distortion or nodules. Such criteria are in keeping with those settled by the Philadelphia Consensus Conference [13] and also with those by the ASCO guidelines for SNB [10]. We did not use the need for mastectomy as a high-risk factor because there is an increasing trend to adopt oncoplastic procedures, including partial mastectomy and immediate breast reconstruction for patients with large DCIS lesions in our clinical environment. The present study shows that the usual high-risk profile actually works very well as a selection tool for DCIS

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Table 4

An update of clinical series dealing with sentinel node biopsy in patients with ductal carcinoma in situ (DCIS)

Reference Klauber-DeMore 2000 [15] Intra 2003 [16] Veronesi 2005 [17] Farkas 2004 [18] Mittendorf 2005 [19] Yen 2005 [20] Wilkie 2005 [21] Zavagno 2005 [22] Camp 2005 [23]

Number of patients

Preoperative diagnosis

Conversion rate NA

+ SN pure DCIS

+ SN DCISmic

107

DCIS, 76 DCISmic, 31

9/76 (11.8%) 3 /31 (9.7%)

41 508 46 44 141 675 102 43

Retrospective DCISmic NA – 4/41 (9.8%) Retrospective DCIS NA 9/508 (1.81%) – Retrospective DCIS NA 0/46 (0%) – DCIS, 44 7/44 (15.9%) 5/42 (11.9%) 2/2 DCIS, 141 42/141 (29.8%) 3/99 (3%) 1/3 DCIS, 613 DCISmic, 62 55/613 (9%) 27/559 (4.8%) 7/51 (13.7%) Retrospective DCIS NA 1/102 (1%) – DCIS, 30 DCISmic, 13 4/30 (13%) 1/26 (3.8%) 5/16 (31.3)

+ SN Invasive carcinoma NA

+ SN (%) Total Occult-micrometastasis/total 12/107 (11%)

9/12 (75%)

– 4/41 (10%) 2/4 (50%) – 9/508 (1.81%) 5/9 (55.5%) – 0/46 (0%) – – 7/44 (26%) 5/7 (71.4%) 10/39 (25.6%) 14/141 (10%) 11/14 (78.6%) 15/66 (22.7%) 49/675 (7.3%) 28/49 (57%) – 1/102 (1%) 1/1 1/1 7/43 (16.3%) 3/7 (42.9%)

DCISmic: DCIS with microinvasion.

patients, given the increased rate of conversion to invasive carcinoma (39%). Furthermore, the devised compiling risk score was quite good in predicting conversion. Also, our results overtly support the view that sentinel node positivity belongs almost exclusively to the subgroup of patients converted to invasive carcinoma, and that sentinel node metastasis increases proportionally from non-invasive to micro-invasive and to fully invasive carcinoma. There was only one unconverted patient displaying a single sentinel node micrometastasis in our series, which might in fact correspond to missed infiltrating carcinoma at histology, a finding that has already been reported, which could be used for upstaging purposes [3]. Nevertheless, spurious transport of benign epithelium to the axilla cannot be excluded in some cases [14]. A few reports in the literature deal with the results of SNB in patients with DCIS [15–23]. An update of such literature data is displayed in Table 4. According to these reports, the rate of sentinel node positivity varies greatly, from 0 to 26%, probably reflecting much heterogeneity in patient selection, as well as in the method used to detect sentinel node metastases. Especially remarkable is the discovery of occult sentinel node metastases, mostly by immunocytochemical analysis: roughly half of the positive cases. Interestingly, two retrospective long-term followup series before the sentinel node era show no clinical impact of such microscopic axillary disease on DCIS patient survival [24,25]. Clearly, some of the aforementioned SNB studies in DCIS are retrospective clinical series assessing the prevalence of sentinel node metastases in patients with a definite (post-operative) diagnosis of in-situ carcinoma, and do not take into account the preoperative clinical setting [18,22]. Similarly, other reports do not assess the conversion rate to invasive carcinoma [16,17]. However, what really matters from the clinical management perspective is the preoperative scenario, when the patient presents with a diagnosis of DCIS and is supposed to undergo a definitive surgical procedure,

including lymph node assessment if necessary. Such an issue is best addressed by clinical studies targeting the upstage of DCIS patients to either micro-invasive or invasive carcinoma, as well as to its relation to sentinel node metastases. It can be shown that the risk of sentinel node metastasis significantly increases with the finding of micro-invasive and truly invasive carcinoma at the definitive pathological evaluation [15,19–21,23]. In fact, the high-risk profile for invasive carcinoma may be used as the only meaningful surrogate for SNB indication in patients with DCIS. Otherwise, some believe such an indication should be disregarded [18]. Efforts to restrain and refine the indication for SNB in DCIS are not futile. On the one hand, in the low risk range, the yield of SNB is poor, precludes future sentinel node procedures in the same breast, and is not free of potential morbidity and cost. On the other hand, in well selected high-risk patients tumour excision and SNB can be acomplished in only one surgery. Therefore, it seems clinically meaningful to keep the indication for SNB well restricted to patients with high risk of invasive disease at the time of DCIS diagnosis. However, according to our results we must accept that once a standard high-risk profile had been established, as such, and used as patient study entry, further refinement in the analysis of the clinical presentation using a ‘risk score’ did not provide enough improvement in the selection of patients. The lack of statistically significant association between higher risk profile and sentinel node metastases in our study is probably due to the scant number of sentinel nodepositive cases. Should more sentinel node-positive cases have been accrued in our investigation, we believe the risk score we designed would have worked as an effective selection tool. Therefore, further studies with a similar layout are needed to settle the final indication for SNB in patients with ductal carcinoma in situ of the breast.

Acknowledgements The authors wish to express their gratitude to the following doctors for their contribution to our investigation:

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Refining the indication for SNB in ductal carcinoma Fraile et al. 789

Eva Castella` and Mireia Margelı´ at Hospital Germans Trias i Pujol. Badalona Maribel Nieto at Hospital de Mataro´ Lluis Solernou, Jordi Tarazona and Elena Martı´ at Althaia Manresa Xavier Mira, Claudia Canizzo and Mireia Recaj at Hospital Sant Jaume, Calella Enric Cristobal and Joan Ribas at Hospital Esperit Sant, Santa Coloma Anna Alcaide at Hospital Municipal, Badalona.

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14

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Silverstein MJ, Rosser RJ, Gibson ED. Axillary lymph node dissection of intraductal breast carcinoma – is it indicated? Cancer 1987; 59: 1819–1824. 2 Pendas S, Dauway E, Giuliano R, Ku NN, Cox CE, Reintgen DS. Sentinel node biopsy in ductal carcinoma in situ patients. Ann Surg Oncol 2000; 7:15–20. 3 Cox CE, Nguyen K, Gray RJ, Salud C, Ku NN, Dupont E, et al. Importance of lymphatic mapping in ductal carcinoma in situ (DCIS): why map DCIS? Am Surg 2001; 67:513–521. 4 Kelly TA, Kim JA, Patrick R, Grundfest S, Crowe JP. Axillary lymph node metastases in patients with a final diagnosis of ductal carcinoma in situ. Am J Surg 2003; 186:368–370. 5 Intra M, Veronesi P, Mazzarol G, Galimberti V, Luini A, Sachinni V, et al. Axillary sentinel lymph node biopsy in patients with pure ductal carcinoma in situ of the breast. Arch Surg 2003; 138:309–313. 6 Adamovich TL, Simmons RM. Ductal carcinoma in situ with microinvasion. Am J Surg 2003; 186:112–116. 7 Fraile M, Rull M, Julian FJ, Fuste F, Barnadas A, Llatjos M, et al. Sentinel node biopsy as a practical alternative to axillary lymph node dissection in breast cancer patients: an approach to its validity. Ann Oncol 2000; 11:701–705. Erratum in Ann Oncol 2000; 11:1619. 8 Zujewski J, Eng-Wong J. Sentinel lymph node biopsy in the management of ductal carcinoma in situ. Clin Breast Cancer 2005; 6:216–222. 9 Moran CJ, Kell MR, Kerin MJ. The role of sentinel lymph node biopsy in ductal carcinoma in situ. Eur J Surg Oncol 2005; 31:1105–1111. 10 Lyman GH, Giuliano AE, Somerfield MR, Benson III AB, Bodurka DC, Burstein HJ, et al. American Society of Clinical Oncology guideline recommendations for sentinel lymph node biopsy in early-stage breast cancer. J Clin Oncol 2005; 23:7703–7720. 11 Lagios MD, Silverstein MJ. Sentinel node biopsy for patients with DCIS: a dangerous an unwarranted direction. Ann Surg Oncol 2001; 8: 275–277.

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Sakorafas GH, Farley DR. Optimal management of ductal carcinoma in situ of the breast. Surg Oncol 2003; 12:221–240. Schwartz GF, Giuliano AE, Veronesi U. Proceedings of the consensus conference on the role of sentinel lymph node biopsy in carcinoma of the breast, April 19–22, 2001, Philadelphia, Pennsylvania. Cancer 2002; 94:2542–2551. Diaz NM, Cox CE, Ebert M, Clark JD, Vrcel V, Stowell N, et al. Benign mechanical transport of breast epithelial cells to sentinel lymph nodes. Am J Surg Pathol 2004; 28:1641–1645. Klauber-DeMore N, Tan LK, Liberman L, Kaptain S, Fey J, Borgen P, et al. Sentinel lymph node biopsy: Is it indicated in patients with high-risk ductal carcinoma-in-situ and ductal carcinoma-in-situ with microinvasion? Ann Surg Oncol 2000; 7:636–642. Intra M, Zurrida S, Maffini F, Sonzogni A, Trifiro G , Gennari R, et al. Sentinel lymph node metastasis in microinvasive breast cancer. Ann Surg Oncol 2003; 10:1160–1165. Veronesi P, Intra M, Vento AR, Naninato P, Caldarella P, Raganelli G, et al. Sentinel lymph node biopsy for localised ductal carcinoma in situ? Breast 2005; 14:520–522. Farkas EA, Stolier AJ, Teng SC, Bolton JS, Fuhram GM. An argument against routine sentinel node mapping for DCIS. Am Surg 2004; 70: 13–17. Mittendorf ME, Arciero CA, Gutchell V, Hooke J, Schriver CD. Core biopsy diagnosis of ductal carcinoma in situ: an indication for sentinel lymph node biopsy. Curr Surg 2005; 62:253–257. Yen TW, Hunt KK, Ross MI, Mirza NQ, Baviera GV, Meric-Bernstam F, et al. Predictors of invasive breast cancer in patients with an initial diagnosis of ductal carcinoma in situ: a guide to selective use of sentinel lymph node biopsy in management of ductal carcinoma in situ. J Am Coll Surg 2005; 200:516–526. Wilkie C, White L, Dupont E, Cantor A, Cox CE. An update of sentinel lymph node mapping in patients with ductal carcinoma in situ. Am J Surg 2005; 190:563–566. Zavagno G, Carcoforo P, Marconato R, Franchini Z, Scalco G, Burelli P, et al. Role of axillary sentinel lymph node biopsy in patients with pure ductal carcinoma in situ of the breast. BMC Cancer 2005; 5:18. Camp R, Feezor R, Kasraeian A, Cendan J, Schell S, Wilkinson E, et al. Sentinel lymph node biopsy for ductal carcinoma in situ: an evolving approach at the University of Florida. Breast J 2005; 11: 394–397. Tamhane R, Dahlstrom JE, McCallum DD, Buckingham JM. The clinical significance of cytokeratin-positive cells in lymph nodes at the time of mastectomy from patients with ductal carcinoma-in-situ. Ann Surg Oncol 2002; 9:999–1003. Lara JF, Young SM, Velilla RE, Santoro EJ, Templeton SF. The relevance of occult axillary micrometastasis in ductal carcinoma in situ. A clinicopathologic study with long-term follow-up. Cancer 2003; 98: 2105–2113.

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Original article

Myocardial perfusion quantification in patients suspected of cardiac syndrome X with positive and negative exercise testing: A [13N]ammonia positron emission tomography study Jessica de Vriesa, Mike J.L. DeJongsteb, Gillian A.J. Jessurunb, Pieter L. Jagerc, Michiel J. Staald and Riemer H.J.A. Slartc Background The combination of angina pectoris, angiographically normal coronary arteries, and a positive exercise stress test (EST) is referred to as cardiac syndrome X. However, a large group of patients suspected of syndrome X reveals a normal exercise stress test and weakens the diagnosis of syndrome X. Previous studies demonstrated an impaired coronary flow reserve on ammonia positron emission tomography (PET) in patients with syndrome X. Aim To evaluate the coronary flow reserve in patients suspected of syndrome X with positive and negative EST findings, using [13N]ammonia PET as the diagnostic aid. Methods Forty-two patients with chest pain and a normal coronary angiography, were analysed by exercise stress testing (EST) and the dypyridamole stress test (DST) on [13N]ammonia PET. Two subgroups were predefined, based on outcome of EST: an EST positive and negative group. A normal control group was used as the reference method. Results A total of 24 (57%) out of 42 patients had significant ST-T changes (EST positive). [13N]ammonia PET showed a significantly lower rest flow in the EST positive

Introduction The proportion of patients with angina pectoris who are found to have normal coronary arteriographic findings ranges between 10 and 20%, depending on the characteristics of the patient group studied [1]. The term ‘syndrome X’, first used by Kemp [2] in 1973, is now frequently used as a diagnostic label for patients who have exertional angina, a positive response to exercise testing and angiographically normal coronary arteries [3]. For the last 30 years this terminology has been coined for the heterogeneous group of patients with the triad of exertional angina, ST-T segment changes during exercise testing and angiographically normal coronary arteries [3].

and EST negative group compared to controls (P < 0.001 and P = 0.0028, respectively). DST [13N]ammonia PET perfusion was significantly reduced in flow in both the EST positive and EST negative groups (P < 0.001 both), as was the DST/rest [13N]ammonia perfusion reserve (P < 0.001 for both), compared to normal controls. Conclusion PET demonstrates a reduced coronary flow reserve in patients suspected of syndrome X, irrespective c of the EST findings. Nucl Med Commun 27:791–794 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:791–794 Keywords: cardiac syndrome X, exercise testing, positron emission tomography a

Faculty of Medicine, Departments of bCardiology, Thoraxcenter, cNuclear Medicine and Molecular Imaging and dNeurosurgery, University Medical Center Groningen and University of Groningen, the Netherlands. Correspondence to Dr Riemer H.J.A. Slart, Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, Hanzeplein 1, P.O. Box 30001, 9700 RB, Groningen, the Netherlands. Tel: + 0031 50 361 3541; fax: + 0031 50 361 1712; e-mail: [email protected] Received 7 February 2006 Accepted 28 June 2006

combination of angina pectoris, ST-segment depression during exercise and a reduced CFR. However, a large group of patients suspected of cardiac syndrome X lacks ST-segment changes during exercise testing. It is unclear whether the patients without ST-segment depression also have impaired CFR, as has been demonstrated in patients with cardiac syndrome X and ST-segment exercise testing. We therefore sought to study, irrespective of the exercise test outcome, the CFR of patients with typical chest pain and normal coronary arteries, by means of a rest and dipyridamole (DST) stress [13N]ammonia PET.

Patients and methods In a significant proportion of patients with cardiac syndrome X a reduced coronary flow reserve (CFR) has been observed [4]. Cannon and Epstein [5] applied the term ‘microvascular angina’ to describe patients with the

Selection of patients

Patients with normal coronary findings on angiography, typical chest pain (without suffering from other diseases responsible for chest complaints) were consecutively

c 2006 Lippincott Williams & Wilkins 0143-3636

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792 Nuclear Medicine Communications 2006, Vol 27 No 10

included in our study between 1999 and 2005. In contrast to many other studies we also included subjects without ST-segment changes during exercise (ST negative group). Seven patients were excluded, five of them because of left bundle branch block, one patient because of first degree AV block and one diabetic patient. All subjects gave informed consent. Chest pain

To score whether a patient’s chest pain was typical of angina pectoris, we used the classification proposed by Diamond [6]. This classification includes three elements: substernal location, precipitation by exertion and relief by rest or nitroglycerine. When all three elements are present angina pectoris is defined as typical, otherwise as atypical. Also, the number of sublingual nitrate tablets used or the number of inhalations undertaken was noted. Exercise testing

Twelve-lead ECGs were recorded and blood pressure was taken at rest, at 1-min intervals during exercise and following exercise, for 5 min. Baseline heart rate and systolic blood pressure were measured and subsequently the so-called ‘double product’ (i.e., rate pressure product) was calculated. A standardized symptom-limited exercise test was performed according to the Weber– Janicki protocol [7]. ST-T segment depression was considered to be significant when ST-T segment depression exceeded 0.1 mV, 60 ms after the J-point at maximal exercise in more than two leads. The patient should at least stress until 85% of their maximum calculated heart rate to consider the exercise test as conclusive. The calculated maximum heart rate was corrected for age, weight and gender. Exercise test end-points were progressive angina, ST-T segment depression > 0.3 mV, systolic blood pressure > 250 mmHg and fatigue. Based on the criteria for ST-T segment depression all subjects were divided into the following two groups: (1) group ST positive – with significant ST-T segment depression on the exercise ECG; and (2) group ST negative – without ST-T segment depression on the exercise ECG. PET imaging

All anti-anginal medication and caffeine-containing beverages were withdrawn at least 24 h prior to the PET studies. Myocardial perfusion was studied according to the methods of Schelbert et al. [8], using [13N]ammonia as the tracer. Subjects were positioned in a ECAT-951/31 PET system (Siemens/CTI, Knoxville, Tennessee, USA), imaging 31 planes simultaneously over 10.8 cm. The measured spatial resolution of the system was 6 mm full width at half maximum. Positioning was performed with

the aid of a rectilinear scan. Photon attenuation was corrected using a retractable external ring source filled with 68Ge/68Ga. Studies were conducted with subjects in the supine position. Room temperature was set at 231C. Measurements were done in a quiet environment after an acclimatization period of 30 min. Dynamic imaging of the rest was started at the time of [13N]ammonia injection (370 MBq) and continued for 8 min (frames: 12  10 s, 1  2 min, and 1  4 min). Following the rest study, dypiridamole infusion (0.56 mg kg – 1 body weight in 6 min) was started. The dynamic acquisition procedure was started immediately after injection of 400 MBq of [13N]ammonia, 6 min after start of dipyridamole infusion, and continued for 15 min. The electrocardiogram was monitored continuously throughout the procedure, using a three-channel electrocardiographic monitor. Blood pressure was monitored every 3 min at rest and every 60 s during stress testing, using an electronic sphygmomanometer (Dynamap, Critikon Inc., Tampa, USA). For constructing parametric polar maps, data of each investigation were reoriented to ten short-axis images, with reference to a manually drawn long axis in the left ventricle. The myocardium in the different slices was divided into 48 segments (7.51 each). Time–activity curves were established in all segments of all slices. To avoid spillover effects, the blood pool was defined into three slices close to the base of the heart. The average activity of these three regions of interest was calculated to give a single blood pool time–activity curve. For all frames polar maps were calculated, using the same set of reorientation data. Thus, for all segments of all slices dynamic tissue data were obtained yielding 480 separate segments. From these data dynamic parametric polar maps were constructed. These polar maps reflect threedimensional imaging of myocardial perfusion of 480 segments of the left ventricle. All data were compared with a database of healthy volunteers [9]. To compensate for the possible influence of rate pressure product on myocardial blood perfusion, perfusion was corrected for rate pressure product, defined as the multiplication of heart rate (bpm) and systolic blood pressure (mmHg). The calculation of normalized perfusion = myocardial perfusion/individual rate pressure product  10 000. These criteria were analysed separately as the coronary flow per segment and as the total coronary flow. Statistical analysis

To compare the PET data of the two groups with the controls, Student’s t-test was used. To compare the PET data of the two groups with each other, the outcomes were first evaluated for normality using the Shapiro–Wilk test. Differences with a P value < 0.05 were considered to be statistically significant.

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PET and exercise testing in syndrome X de Vries et al. 793

Results The baseline characteristics of the 42 patients are shown in Table 1. Twenty-one individuals without chest pain or coronary artery disease were used as the reference [9] (healthy volunteers).

Discussion

Exercise ECG (Table 1)

Based on ST-T segment depression patients were divided in two groups: (1) group ST positive – with ST-T segment depression: 24 patients, 16 females; and (2) group ST negative – without ST-T segment depression: 18 patients, 10 females. Patients with a negative EST had more attacks of chest pain per week and also used more nitroglycerine per attack. On the other hand, patients with a postive EST were treated with a higher dose of beta blockers and Ca2 + channel blockers (Table 1). Positron emission tomography

Table 2 shows the average, normalized myocardial perfusion at rest, the perfusion during DST and flow reserve measured by DST. Data were found to be normally distributed. For the entire group (ST-positive and ST-negative subgroups) the normalized rest flow was significantly lower as compared to controls (P < 0.001 and P = 0.0028, respectively). During DST, the ST-positive and ST-negative groups showed both had reduced flow Table 1

Baseline clinical features of 42 patients

Characteristic Female/male ratio Age (years) Duration of complaint (years) Average number of attacks per week Average nitroglycerine per attack Medication (%) Ca2 + channel blockers Beta blockers Lipid-lowering drugs Alpha1 antagonists Long-acting nitrates ACE inhibitors Diuretics

Total (n = 42)

EST positive (n = 24)

EST negative (n = 18)

26/16 58 ± 12 8.6 ± 8.7 8.23

16/8 59 ± 11 8.7 ± 10.3 7.46

10/8 57 ± 13 8.4 ± 6.4 9.49

0.34

0.22

0.47

43 40 36 21 19 10 10

54 50 38 25 13 4 13

28 28 33 17 28 17 6

Female/male ratio, ST depression are expressed in number of patients; age and duration of complaints are expressed in mean ± SD; medication are expressed in percentage; EST, exercise stress testing; ACE, angiotensin-converting enzyme.

Table 2

compared to controls (P < 0.001 both). Also, the perfusion reserve during DST was reduced in the ST-positive and ST-negative groups in comparison with the controls (P < 0.001 both).

The main finding of this study is that the reduced coronary flow reserve, following pharmacological induced cardiac stress in patients suspected of cardiac syndrome X, is not necessarily associated with ST-segment changes during bicycle exercise stress testing. According to the findings of Cannon et al. [4] and Cannon [10] the absence of ST-segment depression during exercise does not rule out microvascular angina (i.e., cardiac syndrome X). The most plausible explanation for the low sensitivity of the electrocardiogram in detecting exercise-induced ischaemia may be related to the manifestation of microvascularinduced ischaemia. Since the myocardial ischaemia in patients with cardiac syndrome X has been found to be spread both diffuse and ‘patchy’ over the ventricular wall, the ischaemic changes may not be distinguished by the standard electrocardiogram [11]. Furthermore, left ventricular hypercontractility in patients with syndrome X is more likely to be associated with a positive exercise test than in cardiac syndrome X patients without left ventricular hypercontractility [12]. Cannon et al. also observed that CFR was significantly lower in patients with microvascular angina who had ST-T segment depression in response to adenosine than in those who did not have ST-T segment depression. In our study CRF in positive EST group was also lower than the EST negative group, although not significantly. This may be due to the different vasodilatation methods, because in our study DST was used instead of adenosine. We chose to achieve coronary vasodilation with DST because pharmacologically induced hyperaemia is reduced specifically with DST, whereas adenosine may yield preserved CFR in patients with cardiac syndrome X [13]. The cause of the reduced hyperaemic CFR is unknown, but it has previously been related to structural alternations of arterioles as observed in small endomyocardial biopsy samples taken from cardiac syndrome X patients [14]. Biopsy of the ventricle of the heart showed pathological small coronary arteries with fibromuscular hyperplasia, hypertrophy of the media, intima proliferation,

Perfusion dynamics of 42 patients Total (n = 42)

Normalized rest perfusion DST perfusion Perfusion reserve (DST)

*

124 ± 34 213 ± 51** 2.04 ± 0.63**

EST positive (n = 24) **

122 ± 31 215 ± 48** 1.99 ± 0.70**

EST negative (n = 18) ***

126 ± 38 207 ± 56** 2.05 ± 0.56**

Controls (n = 21) 148 ± 35 292 ± 95 2.91 ± 1.04

Perfusion in ml
min – 1  100 g tissue; normalized perfusion = rate pressure product corrected perfusion in ml
min – 1  100 g tissue; perfusion reserve = ratio perfusion during DST/perfusion at rest. * P < 0.002 vs. normal controls; ** P < 0.001 vs. normal controls; *** P < 0.003 vs. normal controls. EST, exercise stress test; DST, dipyridamole stress test.

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endothelial degeneration, and capillaries with swollen endothelial cells and patchy fibrosis [14]. Clinical implications

To diagnose cardiac syndrome X, PET is not necessary in the work-up of patients with cardiac syndrome X. However, we recommend low threshold referral to centres experienced in the management of these complex patient categories. In these clinical settings, multidisciplinary teams are able to tailor the most appropriate treatment to the individual patient. The use of PET in the work-up of cardiac syndrome X should be preserved at specialized centres and preferably implemented when patients have typical chest pain without significant STsegment depression. To date, PET is the most reliable tool to assess the myocardial microvascular state.

Conclusion PET demonstrates a reduced coronary flow reserve in patients suspected of cardiac syndrome X, independently of the exercise stress test findings. PET may therefore be considered as an additional tool to quantify the diagnosis of cardiac syndrome X.

References 1

2 3

4 5

The underlying pathogenetic mechanism in this syndrome appears to be multifactorial and in this heterogeneous group of patients is thought to be related to an altered sympathetic state, to endothelial dysfunction, an abnormal pain perception and abnormal metabolism inducing microvascular ischaemia [15]. Patients with cardiac syndrome X may benefit from cardiovascular drug therapy or electrical neurostimulation [4,16]. With respect to cardiovascular drugs, the supposed underlying endothelial dysfunction can be treated by several agents, such as angiotensin-converting enzyme (ACE) inhibitors and cholesterol-lowering drugs. However, since conventional therapies often show dissatisfying results, new therapeutic modalities, such as neuromodulation should be considered [17]. Electrical neurostimulation appears to be of benefit in refractory chest pain and concomitant myocardial perfusion dynamics in these patients [16]. It should be pointed out that the exact underlying cause in subjects with cardiac syndrome X is unknown, and that subsequently the therapeutic approach will be palliative. Future perspectives

A study by Lanza et al. [18] observed decreased meta[123I]iodobenzylguanidine (123I-MIBG) uptake in the left ventricle, indicating an abnormal cardiac adrenergic nerve function [18]. These authors concluded that chest pain in syndrome X is induced by cardiac nerve dysfunction, although the mechanism and the pathological meaning of the abnormal cardiac 123I-MIBG uptake deserves further investigation. This adrenergic function may increase both, microvascular tone and sensitive small coronary arteries to vasoconstriction [19]. So, an abnormal cardiac adrenergic tone may play a key role in cardiac syndrome X [20,21]. To study this hypothesis further, quantification of regional sympathetic nerve disturbances in patients with cardiac syndrome X, the use of the PET tracer 11Chydroxyephedrine (11C-HED) is worth considering [22]. However, so far, no data concerning 11C-HED for the diagnosis of cardiac syndrome X are available.

6 7

8

9

10

11

12

13

14

15 16

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20

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Kemp HG, Kronmal RA, Vlietstra RE, Frye RL. Seven year survival of patients with normal or near normal coronary arteriograms: a CASS registry study. J Am Coll Cardiol 1986; 7:479–483. Kemp Jr HG. Left ventricular function in patients with the anginal syndrome and normal coronary arteriograms. Am J Cardiol 1973; 32:375–376. Kaski JC, Crea F, Nihoyannopoulos P, Hackett D, Maseri A. Transient myocardial ischemia during daily life in patients with syndrome X. Am J Cardiol 1986; 58:1242–1247. Cannon III RO, Camici PG, Epstein SE. Pathophysiological dilemma of syndrome X. Circulation 1992; 85:883–892. Cannon III RO, Epstein SE. ‘Microvascular angina’ as a cause of chest pain with angiographically normal coronary arteries. Am J Cardiol 1988; 61:1338–1343. Diamond GA. A clinically relevant classification of chest discomfort. J Am Coll Cardiol 1983; 1:574–575. Weber KT, Janicki JS. Equipment and protocols to evaluate the exercise response. In: Weber KT, Janicki JS (editors): Cardiopulmonary exercise testing. Physiologic principles and clinical applications. Philadelphia: WB Saunders Company; 1986, pp. 138–150. Schelbert HR, Phelps ME, Huang SC, MacDonald NS, Hansen H, Selin C, et al. N-13 ammonia as an indicator of myocardial blood flow. Circulation 1981; 63:1259–1272. Blanksma PK, Willemsen AT, Meeder JG, de Jong RM, Anthonio RL, Pruim J, et al. Quantitative myocardial mapping of perfusion and metabolism using parametric polar map displays in cardiac PET. J Nucl Med 1995; 36:153–158. Cannon III RO. Association of abnormal left-ventricular reponse to exercise with dynamic limitation in coronary flow reserve in patients with chest pain, angiographically normal coronary-arteries. Circulation 1992; 86:588. Taggart P, Sutton P, Opthof T, Coronel R, Kallis P. Electrotonic cancellation of transmural electrical gradients in the left ventricle in man. Prog Biophys Mol Biol 2003; 82:243–254. Tousoulis D, Crake T, Lefroy DC, Galassi AR, Maseri A. Left-ventricular hypercontractility and ST segment depression in patients with syndrome-X. J Am Coll Cardiol 1993; 22:1607–1613. Holdright DR, Lindsay DC, Clarke D, Fox K, Poole Wilson PA, Collins P. Coronary flow reserve in patients with chest pain and normal coronary arteries [Abstract]. Br Heart J 1993; 70:513–519. Mosseri M, Yarom R, Gotsman MS, Hasin Y. Histologic evidence for smallvessel coronary-artery disease in patients with angina-pectoris and patent large coronary-arteries. Circulation 1986; 74:964–972. Kaski JC, Aldama G, Cosin-Sales J. Cardiac syndrome X. Diagnosis, pathogenesis and management. Am J Cardiovasc Drugs 2004; 4:179–194. Jessurun GA, Hautvast RW, Tio RA, DeJongste MJ. Electrical neuromodulation improves myocardial perfusion and ameliorates refractory angina pectoris in patients with syndrome X: fad or future? Eur J Pain 2003; 7:507–512. Lanza GA, Sestito A, Sgueglia GA, Infusino F, Papacci F, Visocchi M, et al. Effect of spinal cord stimulation on spontaneous and stress-induced angina and ‘ischemia-like’ ST-segment depression in patients with cardiac syndrome X. Eur Heart J 2005; 26:983–989. Lanza GA, Giordano A, Pristipino C, Calcagni ML, Meduri G, Trani C, et al. Relationship between myocardial 123I-metaiodobenzylguanidine scintigraphic uptake and heart rate variability in patients with syndrome X. Ital Heart J 2000; 1:221–225. Camici PG, Marraccini P, Gistri R, Salvadori PA, Sorace O, Labbate A. Adrenergically mediated coronary vasoconstriction in patients with syndrome-X. Cardiovasc Drugs Ther 1994; 8:221–226. Montorsi P, Fabbiocchi F, Loaldi A, Annoni L, Polese A, De Cesare N, et al. Coronary adrenergic hyperreactivity in patients with syndrome X and abnormal electrocardiogram at rest. Am J Cardiol 1991; 68:1698–1703. Rosano GMC, Ponikowski P, Adamopoulos S, Collins P, Poole-Wilson PA, Coats AJS, et al. Abnormal autonomic control of the cardiovascular-system in syndrome-X. Am J Cardiol 1994; 73:1174–1179. Knuuti J, Sipola P. Is it time for cardiac innervation imaging? Q J Nucl Med Mol Imaging 2005; 49:97–105.

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Original article 18

F-FDG PET and PET/CT for detection of pulmonary metastases from musculoskeletal sarcomas

Andrei Iagarua, Sant Chawlab, Lawrence Menendezc and Peter S. Contia Objective Sarcomas represent a significant therapeutic challenge and their potential for distant pulmonary metastases is well known. [18F]Fluorodeoxyglucose (18F-FDG) positron emission tomography (PET) has a role in differentiating sarcomas from benign tumours and assessing the response to therapy in advanced sarcomas. However, PET appears to be less accurate in detection of pulmonary metastases. We were therefore prompted to review our experience with PET and PET/computed tomography (CT) in osseous and soft tissue sarcomas (OSTSs). Methods This is a retrospective study (January 1995 to December 2004) of 106 patients with histological diagnosis of OSTS, who had PET and PET/CT at our institution. The group included 52 men and 54 women, aged 12–92 years (average, 45 ± 20 years). Results For all the patients in the analysis, the sensitivity and specificity were 68.3% (95% CI: 53–80.4) and 98.4% (95% CI: 91.8–99.7) for PET, with 95.1% sensitivity (95% CI: 83.8–98.6) and 92.3% specificity (95% CI: 83.2–96.7) for CT. Pulmonary metastases were seen in 40 patients. CT

Introduction Osseous and soft-tissue sarcomas (OSTSs) represent a histological heterogeneous group of malignant tumours. Bone sarcomas account for 0.2% of all primary cancers in adults and approximately 5% of childhood malignancies. Soft-tissue sarcomas are extremely rare tumours, accounting for 0.7% of adult malignancies. In children younger than 15 years of age they represent 6.5% of cancers. According to American Cancer Society estimates, approximately 10 700 new cases were expected in 2002, comprising 8300 soft tissue and 2400 bone and joints cancers. In 2002 about 5200 patients died from these cancers (3900 from soft tissue and 1300 from bone and joints malignancies) [1]. OSTSs are classified according to their grade, which represents the most important prognostic factor. The histopathological grade of a sarcoma is based on the degree of differentiation, cellularity, number of mitoses, pleomorphism and amount of necrosis. OSTSs present significant diagnostic and therapeutic challenges. Treatment generally includes a combination of surgery, radiation therapy and chemotherapy. Patients with high-grade soft tissue sarcomas are at high risk of developing distant pulmonary metastases.

identified 17 lesions larger than 1.0 cm, while PET identified 13 of them (76.5%). Conclusions Chest CT is more sensitive than PET in detecting pulmonary metastases from OSTS. A significant portion of known pulmonary metastases greater than 1.0 cm on CT, are PET negative. Sub-centimetre CT lesions should not be considered false positive if inactive on PET. A negative PET scan in the presence of suspicious CT findings in the chest cannot reliably exclude pulmonary metastases from OSTS. Nucl Med Commun 27:795–802

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:795–802 Keywords: sarcoma, lung, metastases,

18

F-FDG, PET/CT

a PET Imaging Science Center, cDepartment of Orthopaedic Surgery, Keck School of Medicine of USC, Los Angeles and bThe Cancer Center of Midway Hospital, Los Angeles, USA.

Correspondence to Dr Peter S. Conti, PET Imaging Science Center, Keck School of Medicine of USC, 1510 San Pablo St, Suite 350, Los Angeles, CA 90033, USA. Tel: + 001 323 442 5940; fax: + 001 323 442 5778; e-mail: [email protected] Received 14 February 2006 Accepted 4 May 2006

Diagnostic imaging plays a major role in the evaluation of patients with OSTS. Anatomical imaging includes radiography, computed tomography, magnetic resonance imaging and bone scintigraphy. Molecular imaging with positron emission tomography (PET) has an important role in the imaging evaluation of patients with OSTS, including detecting local recurrence and metastatic disease, guiding biopsy, predicting and monitoring response to therapy and assessing prognosis. An extensive meta-analysis of the published data regarding osseous and soft tissue sarcomas found 29 studies with subject numbers ranging from five to 202. However, the authors describe these studies as having poor methodological quality [2]. In addition, the accuracy of [18F]fluorodeoxyglucose (18F-FDG) positron emission tomography (PET) for detection of pulmonary metastases from OSTS has not been studied extensively: we found only one study that included 71 patients investigated with a dedicated PET unit, without hardware-fused PET/CT images available [3]. Therefore, we were prompted to review our experience with 18F-FDG PET and PET/computed tomography (CT) in OSTS, specifically with respect to understanding the pattern of FDG uptake in pulmonary metastases.

c 2006 Lippincott Williams & Wilkins 0143-3636

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Materials and methods This is a retrospective study (1 January 1995 to 31 December 2004) of patients with histological diagnosis of OSTS, who had 18F-FDG PET and CT or 18F-FDG PET/ CT at our institution. The study was performed with approval of the institutional review board. The inclusion criteria were proven diagnosis of primary musculoskeletal and soft tissue malignancy, imaging studies for review and access to the patients’ clinical charts. A total of 295 cases were identified initially in the Imaging Center database under the subgroup of ‘musculoskeletal cancers’. However, 189 patients were excluded because their musculoskeletal lesions were metastases from other malignancies, rather than primary OSTS tumours. The final analysis group consisted of 106 patients, 52 men and 54 women, 12–92 years of age (average, 45 ± 20 years). The tumour types were identified as rhabdomyosarcoma (two patients), chondrosarcoma (six patients), leiomyosarcoma (seven patients), Ewing’s sarcoma (eight patients), liposarcoma (10 patients), malignant fibrous histiocytoma (20 patients), osteosarcoma (21 patients), and other sarcomas (32 patients). Twenty-two patients had the PET performed for initial staging of disease, while the remaining 84 patients had the PET scan for restaging after therapy. Therapeutic options included surgery (19 patients), chemotherapy (21 patients), surgery/chemotherapy (14 patients), chemo/ radiation therapy (nine patients) and surgery/chemo/ radiation therapy (21 patients). The reports of PET, CT and PET/CT were reviewed and their results were recorded. Re-interpretation of the studies was performed for accuracy. Whenever the imaging study report indicated pulmonary metastases, follow-up clinical outcome and pathology reports were reviewed for consistency in diagnosis. Multiple pulmonary nodules larger than 5 mm on CT were considered to be metastases by the referring oncologists and treated as such, without biopsy confirmation prior to therapy. Pulmonary findings on PET were recorded as suspicious for metastases when maximum standardized values (SUVmax) were higher than 2. Seventy-six patients were studied on a dedicated PET scanner and included in group A, while 30 of the patients had the studies acquired on a dedicated PET/CT scanner and included in group B. Group A included 34 men and 42 women, 12–92 years of age (average 46 ± 21 years). Group B consisted of 18 men and 12 women, 12–73 years (average 41 ± 15 years). The PET scans for group A (76 patients) were acquired on an ECAT Exact 953A PET scanner (Siemens/CTI, Knoxville, Tennessee). Data were registered in the twodimensional mode, with attenuation correction calculated

from a 3-min transmission scan. Images were acquired 60 min after i.v. administration of an average dose of 550 MBq of 18F-FDG. Each data set consisted of 31 contiguous, 3.375-mm thick, tomographic sections, for a total field of view of 10 cm. For whole-body examinations, data were collected at each bed position for 4 min and then the bed was advanced until the whole body was imaged. Images were interpreted on a Sun-based Sparc 10 workstation, with software provided by CTI. Dedicated PET scans were visually correlated with recent chest CT examinations performed prior to PET (at intervals of 1–30 days). The PET/CT studies (30 patients) were acquired with a Biograph LSO PET/CT scanner (Siemens/CTI, Knoxville, Tennessee). The system consists of a dual-slice, spiral CT (Siemens Somatom Emotion) in tandem with an ACCEL PET and is optimized for use in whole-body oncology. Data were obtained in the three-dimensional mode, with attenuation correction calculated from coregistered CT images. Images were acquired 60 min after i.v. injection of an average dose of 550 MBq of 18F-FDG. The images were interpreted on a Windows NT-based computer system, with a Siemens/Syngo user interface. Specificities and sensitivities for pulmonary metastases detection using PET and CT were calculated using the pathology results of the excised pulmonary lesions or follow-up evidence of pulmonary disease progression (increase in size or number of pulmonary lesions) as the gold standard. Multiple PET and CT specificities and sensitivities were calculated for all the patients included in the cohort, for the groups with dedicated PET and PET/CT, as well as based on the indication for the imaging studies (initial staging or restaging following therapy). Confidence interval (CI) estimations were performed using the Wilson score method.

Results Pulmonary metastases were detected in 40 patients (38%). PET identified pulmonary metastases in 29 (73%) of these patients, while chest CT demonstrated pulmonary metastases in 39 (98%) patients. In one of the positive PET exams, the CT report did not describe pulmonary lesions. On the other hand, for 11 of the pulmonary metastases seen on CT of the chest, the lesions were not seen on PET. However, all these latter CT findings were multiple sub-centimetre nodules. CT identified 17 lesions larger than 1.0 cm, while PET identified 13 of them (76.5%). Overall, concordant PET and CT detection of pulmonary metastases was noted in 27 patients (67.5%). The four lesions larger than 1.0 cm seen on CT and not detected by PET originated from primary tumours with the histological diagnosis of liposarcoma (two), osteosarcoma (one), and angiosarcoma (one). The size of the 17 lesions larger than 1.0 cm

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PET/CT detection of pulmonary metastases from sarcomas Iagaru et al. 797

identified on chest CT was 1.1–2.5 cm (average, 1.72 ± 0.37 cm), while the four missed by PET measured 2.2 cm, 1.5 cm, 1.5 cm and 1.7 cm, respectively. All four were biopsy proven metastases. FDG uptake was noted in the primary tumour in 99 (93.4%) of the 106 patients included in the study. All four pulmonary lesions missed by FDG PET originated from FDG-avid primary tumours.

The performance of 18F-FDG PET for the detection of pulmonary metastases was similar in groups A and B: 68.7% vs. 66.7% for sensitivity and 97.9% vs. 100% for specificity. However, when comparing the chest CT performance in the two groups the results were better in group A: 96.8% vs. 90% for sensitivity and 93.9% vs. 87.5% for specificity.

For all the patients included in the analysis, the sensitivity and specificity for PET were 68.3% (95% CI: 53–80.4) and 98.4% (95% CI: 91.8–99.7). CT had a sensitivity of 95.1% (95% CI: 83.8–98.6) and specificity of 92.3% (95% CI: 83.2–96.7). For the patients referred for initial staging, the sensitivity and specificity for PET were 50% (95% CI: 18.7–81.2) and 93.8% (95% CI: 71.7–98.9), with 100% sensitivity (95% CI: 60.9–100) and 93.7% specificity (95% CI: 71.7–98.9) for CT. In the group of patients referred for restaging, the sensitivity and specificity for PET were 71.4% (95% CI: 54.9–83.7) and 100% (95% CI: 92.7–100), 94.3% (95% CI: 81.4–98.4) and 91.8% (95% CI: 80.8–96.8) for CT.

The sensitivities and specificities of PET and CT for pulmonary metastases detection in the studied population are presented in Fig. 1. Figure 2 demonstrates multiple pulmonary metastases seen on both PET and CT in a 46-year-old woman with right femur osteosarcoma. As expected because of the spatial resolution of PET, pulmonary lesions less than 1 cm were not seen, as is the case in Fig. 3. It presents a 0.8 cm pulmonary nodule seen on CT, but not on PET in a 56-year-old man with right calf malignant fibrous histiocytoma. Figure 4 illustrates pulmonary metastases larger than 1 cm (in this case measuring 1.7 cm) that were seen on CT, but not on PET in a 46-year-old man with right humeral osteosarcoma.

In group A (dedicated PET and visual fusion with CT), the sensitivity and specificity were 68.7% (95% CI: 51.4– 82) and 97.9% (95% CI: 89.1–99.6) for PET, 96.8% (95% CI: 83.8–99.4) and 93.9% (95% CI: 83.5–97.9) for CT, respectively. Sub-analysis for group A revealed that the dedicated PET scan identified pulmonary metastases in 23 patients (30%), while pulmonary lesions were described on concurrent (within 1 month) CT in 29 cases (38%). The results of PET and CT were concordant in 21 patients, PET identifying one patient with pulmonary metastases not seen on CT. However, the chest CT scan demonstrated pulmonary metastases in eight patients who had negative PET studies. In one of these patients the lesion missed by PET was larger than 1 cm (1.5 cm) on CT, while the other missed metastases were sub-centimetre in diameter. CT identified nine lesions larger than 1 cm, while PET identified eight of them. The lesion larger than 1 cm missed by PET was in a case of angiosarcoma.

Discussion

In group B (hardware fusion of PET and CT), the sensitivity and specificity for PET were 66.7% (95% CI: 35.4–87.9) and 100% (95% CI: 81.6–100) and 90% (95% CI: 59.6–98.2) and 87.5% (95% CI: 64–96.5) for CT. Acquisition of PET and CT was done simultaneously. However, analysing the results of PET and CT independently, PET identified pulmonary disease in six patients (20%) and chest CT demonstrated lesions in 10 patients (33%). The findings of PET and CT were concordant in six cases, with chest CT demonstrating pulmonary disease in four cases not seen on PET. CT identified eight lesions larger than 1 cm, while PET identified five and missed three of them (37.5%). These three missed lesions originated from primary osteosarcoma and liposarcoma.

Diagnostic imaging plays a major role in the evaluation of patients with OSTS. Plain radiography, computed tomography, magnetic resonance imaging and bone scintigraphy are involved in the initial staging of these tumours. Molecular imaging with 18F-FDG PET emerged in recent years as a powerful modality not only for initial assessment, but for evaluation of response to therapy and detection of recurrences as well. In 1996, the role of 18 F-FDG PET in differentiation of benign from malignant primary bone and soft tissue lesions was discussed by Conti et al. [4]. In a more recent literature review, the same group concluded that 18F-FDG PET may have an important role in the imaging evaluation of patients with bone and soft tissue sarcoma [5]. 18F-FDG PET was evaluated in relation to traditional imaging modalities in various studies. A meta-analysis of these clinical studies on PET and sarcomas was undertaken by Bastiaannet et al. This systematic approach resulted in the finding that 18FFDG PET can discriminate between sarcomas and benign tumours, as well as low and high grade sarcomas. The investigators recommended that PET should be directed to the clinical applications of detection and grading of sarcomas and the treatment evaluation of locally advanced sarcomas [2]. During the initial work-up of OSTS, MRI provides better description and definition of the extent of the tumour [6]. But high FDG uptake in the primary tumour usually predicts a poor prognosis, as is the reduced decrease in FDG uptake in the tumour after therapy. 18F-FDG PET is useful in predicting outcome response in neoadjuvant therapy, allowing the recognition of postoperative changes from residual sarcoma/local recurrence and as a wholebody approach to diagnostic imaging [7,8]. In this

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Fig. 1

Sensitivity Specificity

Sensitivities and specificities for PET and CT in the studied population 98.4

95.1 92.3

97.8

100

96.7 93.5

90.9 89.5

70.8

68.3 %

60

PET (overall)

CT (overall)

PET (group A) CT (group A) PET (group B) CT (group B)

The sensitivities and specificities of postiron emission tomography (PET) and computed tomography (CT) for pulmonary metastases detection in the studied population.

Fig. 2

A 46-year-old woman with right femur osteosarcoma. Lung metastases are demonstrated on PET (arrows) and CT (arrow heads). Postchemotherapy bone marrow hyperplasia is seen as well in the sternum and vertebral bodies.

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PET/CT detection of pulmonary metastases from sarcomas Iagaru et al. 799

Fig. 3

A 56-year-old man with right calf malignant fibrous histiocytoma. No activity is noted on fluorodeoxyglucose PET in the RLL nodule seen on CT (arrows) and measuring 0.8 cm.

context, SUVmax [9] uptake in the sarcoma is an independent predictor of survival and disease progression, as well as a marker of the preoperative neoadjuvant chemotherapy efficacy and the risk for metastases. 18FFDG PET can identify the patients with sarcoma who are most likely to benefit from neoadjuvant chemotherapy [10–17]. Bredella et al. compared 18F-FDG PET with magnetic resonance imaging (MRI) in a study that suggests that PET is a useful adjunct to MRI in distinguishing viable tumour from post-therapeutic changes in patients with OSTS [18]. Additional tracers have been investigated to describe OSTS metabolic activity over the last several decades. For example, in 1987, Schmall et al. describe their experience with 11C-labelled alpha-aminoisobutyric acid in malignant fibrous histiocytoma [19]. Recent data with [18F]fluoride for PET bone scanning shows promising results in evaluation of osseous neoplasms, both for primary staging and for evaluation of response to therapy [20]. Advances in molecular biology also will likely lead to new, more specific radiotracers for sarcoma imaging.

The introduction of adjuvant chemotherapy changed the overall survival in OSTS. Still, the presence of pulmonary metastases is consistently associated with a poor prognosis. Early detection of pulmonary metastases allows for surgical removal, thus being an important step in the management of OSTS. Traditionally, chest CT screening was employed to achieve high detection rates in OSTS pulmonary metastases. However, Fleming et al. demonstrated that less than 1% of patients with T1 primary extremity soft tissue sarcomas were found to have pulmonary metastases that were detectable using a staging algorithm that employs routine chest radiography with the selective use of chest CT. Thus, chest CT as part of the staging evaluation of patients with T1 disease is not recommended [21]. Even in patients with T2 soft tissue sarcomas, the findings of Porter et al. do not support the routine use of chest CT scanning in all patients, except in patients with high-grade lesions or extremity lesions [22]. The technological advances of CT scanning allow high rates of detection of pulmonary nodules, but these are non-specific findings in most instances. Chest CT has a limited positive predictive value for OSTS

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Fig. 4

A 46-year-old man with right humerus osteosarcoma. No activity is noted on PET in the 1.7 cm RLL pleural-based nodule identified on CT (arrows).

pulmonary metastases, but as surgery is the only way to cure metastatic patients, CT is still used as the reference technique until a more specific approach can be found [23]. These results with chest CT prompted the evaluation of 18 F-FDG PET as a better modality to identify pulmonary metastases from OSTS. An initial report using sodium iodide gamma camera based PET scanning was promising, with sensitivity of 91% and specificity of 88%, concluding that 18F-FDG PET is an accurate tool for the characterization of indeterminate pulmonary masses or nodules [24]. False negative results in this paper included a 1.3 cm sarcoma metastasis. Lucas et al. found that FDG PET is inferior to chest CT in detecting lung metastases [25]. These results were considered to be secondary to the scanning methodology (images acquired too soon after FDG injection), soft tissue photon attenuation in the chest in the absence of a transmission scan, and the observation that lung metastases do not appear to have as good vascular supply as the original tumour until they reach a specific size. In a comparison of spiral chest CT with 18F-FDG PET, Franzius et al. noted that 18F-FDG PET had a sensitivity of 50%, a specificity of 98%, and an

accuracy of 87% on a patient based analysis, while comparable values for spiral CT were 75%, 100% and 94% [3]. These values are similar to our results. We expected in our study that chest CT would be superior to PET in the detection of multiple subcentimetre pulmonary lesions, since the lesions would be under the spatial resolution of most current PET and PET/CT scanners. This was confirmed, as chest CT scanning documented sub-centimetre pulmonary metastases in 33 patients (31%), while PET identified subcentimetre pulmonary metastases in 25 patients (24%). When the lesions larger than 1 cm were analysed, chest CT identified 17 lesions, while PET identified 13 metastases (76.5%). All of the four lung biopsy proven metastases larger than 1 cm not seen on PET, but visualized on CT, originated from primary tumours that demonstrated intense 18F-FDG avidity. They measured 1.5, 1.5, 1.7 and 2.2 cm on the CT. Overall, chest CT demonstrated pulmonary metastases in 39 patients (37%) and 18F-FDG PET identified them in 29 patients (27%). While the inferior performance of PET in detection of sub-centimetre pulmonary metastases can be explained

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PET/CT detection of pulmonary metastases from sarcomas Iagaru et al. 801

by the physical limitations of this technology, its poor performance when compared to chest CT in the identification of lesions larger than 1 cm may have other causes. An FDG-avid primary OSTS does not imply equally intense FDG uptake in the pulmonary metastasis. This could be explained by a reduced number of glucose transporter proteins in the metastases or otherwise altered glucose metabolism. As suggested by other investigators above, the blood flow to the metastases may also be a factor. As mentioned in the results section, there were no significant differences in sensitivities and specificities of 18F-FDG PET for detection of pulmonary metastases in groups A and B: 68.7% vs. 66.7% for sensitivity and 97.9% vs. 100% for specificity. We can conclude that the hardware fused studies are not adding power to the diagnostic value of 18F-FDG PET in this cohort. The analysis of the sensitivities (90.9–100%, average: 95.4 ± 3.3) and specificities (89.5–93.7%, average: 92.1 ± 1.7) for CT in the subgroups presented in this study suggests an important role for CT in the evaluation for pulmonary metastases of all patients presenting with OSTS, both for initial staging and restaging of the disease. However, these results are affected by the fact that this is a selected population, with known diagnosis of OSTS prior to the study and high pretest probability for pulmonary metastases in the course of disease. There was a difference in the sensitivity and specificity of chest CT between group A and group B (96.8% and 93.9% vs. 90% and 87.5%, respectively). This is probably due to the fact that, opposed to the dedicated chest CT in group A, the CT scans from fused PET/CT imaging were acquired without i.v. contrast or breath-hold technique. Thus, obtaining a dedicated chest CT in patients with OSTS and suspected pulmonary metastases appears to be a reasonable approach. In the case of 18F-FDG PET, sensitivities (50–71.4%, average: 64.1 ± 9.1) for pulmonary metastases detection across the subgroups of this cohort suggest a poor performance, despite excellent specificities (93.8–100%, average: 98 ± 2.5). The role of 18F-FDG PET is probably most important to serve as a baseline and for monitoring response to therapy in the primary tumour, since 93.4% of the patients demonstrated FDG avid primary lesions. FDG PET might have a role in evaluation of distant soft tissue and/or osseous metastases in the patients referred for restaging of OSTS, but further studies are needed to evaluate this hypothesis.

Findings were similar using dedicated PET with a recent CT scan reference versus hybrid PET/CT. Sub-centimetre CT lesions on concurrent or recent scans should not be considered false positive if inactive on PET, in this clinical setting. A negative PET scan in the presence of suspicious CT findings in the chest therefore cannot reliably exclude pulmonary metastases from OSTS. Further prospective evaluation of pulmonary disease in this patient population using PET/CT assessment is warranted. Factors explaining the difference in uptake patterns in primary OSTS and pulmonary metastases should be investigated.

References 1 2

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Conclusions As expected, CT of the chest is more sensitive than PET in detecting pulmonary metastases from OSTS. However, a significant portion of known pulmonary metastases greater than 1.0 cm on CT, are also PET negative.

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Jemal A, Murray T, Ward E, Samuels A, Tiwari RC, Ghafoor A, et al. Cancer statistics, 2005. CA Cancer J Clin 2005; 55:10–30. Bastiaannet E, Groen H, Jager PL, Cobben DC, van der Graaf WT, Vaalburg W, et al. The value of FDG-PET in the detection, grading and response to therapy of soft tissue and bone sarcomas; a systematic review and meta-analysis. Cancer Treat Rev 2004; 30:83–101. Franzius C, Daldrup-Link HE, Sciuk J, Rummeny EJ, Bielack S, Jurgens H, et al. FDG-PET for detection of pulmonary metastases from malignant primary bone tumors: comparison with spiral CT. Ann Oncol 2001; 12: 479–486. Conti PS, Feske WI, Grafton ST, Hayles SO, Menendez LR. Differentiation of benign from malignant primary bone and soft tissue lesions with PET. J Nucl Med 1996; 37:140P. Jadvar H, Gamie S, Ramanna L, Conti PS. Musculoskeletal system. Semin Nucl Med 2004; 34:254–261. Brisse H, Ollivier L, Edeline V, Pacquement H, Michon J, Glorion C, et al. Imaging of malignant tumors of the long bones in children: monitoring response to neoadjuvant chemotherapy and preoperative assessment. Pediatr Radiol 2004; 34:595–605. Brenner W, Bohuslavizki KH, Eary JF. PET imaging of osteosarcoma. J Nucl Med 2003; 44:930–942. Conti PS, Menendez LR, Ramanna L, Henderson RW. Use of positron emission tomography in the evaluation of cryosurgical treatment of musculoskeletal tumors. Radiology 1999; 213(P):118. Sugawara Y, Zasadny KR, Neuhoff AW, Wahl RL. Reevaluation of the standardized uptake value for FDG: variations with body weight and methods for correction. Radiology 1999; 213:521–525. Eary JF, O’Sullivan F, Powitan Y, Chandhury KR, Vernon C, Bruckner JD, et al. Sarcoma tumor FDG uptake measured by PET and patient outcome: a retrospective analysis. Eur J Nucl Med Mol Imaging 2002; 29: 1149–1154. Conrad 3rd EU, Morgan HD, Vernon C, Schuetze SM, Eary JF. Fluorodeoxyglucose positron emission tomography scanning: basic principles and imaging of adult soft-tissue sarcomas. J Bone Joint Surg Am 2004; 86-A(suppl 2):98–104. Schulte M, Brecht-Krauss D, Werner M, Hartwig E, Sarkar MR, Keppler P, et al. Evaluation of neoadjuvant therapy response of osteogenic sarcoma using FDG PET. J Nucl Med 1999; 40:1637–1643. Schuetze SM, Rubin BP, Vernon C, Hawkins DS, Bruckner JD, Conrad 3rd EU, et al. Use of positron emission tomography in localized extremity soft tissue sarcoma treated with neoadjuvant chemotherapy. Cancer 2005; 103:339–348. Stokkel MP, Draisma A, Pauwels EK. Positron emission tomography with 2-[18F]-fluoro-2-deoxy-D-glucose in oncology. Part IIIb: Therapy response monitoring in colorectal and lung tumours, head and neck cancer, hepatocellular carcinoma and sarcoma. J Cancer Res Clin Oncol 2001; 127:278–285. Franzius C, Sciuk J, Brinkschmidt C, Jurgens H, Schober O. Evaluation of chemotherapy response in primary bone tumors with F-18 FDG positron emission tomography compared with histologically assessed tumor necrosis. Clin Nucl Med 2000; 25:874–881. Gamie S, Ramanna L, Cham D, Conti PS. Clinical utility of 18F-FDG-PET in the evaluation of bone and soft tissue sarcoma. J Nucl Med 2003; 44:75P. Schwarzbach MH, Hinz U, Dimitrakopoulou-Strauss A, Willeke F, Cardona S, Mechtersheimer G, et al. Prognostic significance of preoperative [18-F] fluorodeoxyglucose (FDG) positron emission tomography (PET) imaging in

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patients with resectable soft tissue sarcomas. Ann Surg 2005; 241: 286–294. Bredella MA, Caputo GR, Steinbach LS. Value of FDG positron emission tomography in conjunction with MR imaging for evaluating therapy response in patients with musculoskeletal sarcomas. Am J Roentgenol 2002; 179:1145–1150. Schmall B, Conti PS, Bigler RE, Zanzonico PB, Reiman RE, Benua RS, et al. Imaging studies of patients with malignant fibrous histiocytoma using C-11-alpha-aminoisobutyric acid (AIB). Clin Nucl Med 1987; 12:22–26. Hoegerle S, Juengling F, Otte A, Altehoefer C, Moser EA, Nitzche EU. Combined FDG and F-18 fluoride whole-body PET: a feasible two-in-one approach to cancer imaging? Radiology 1998; 209:253–258. Fleming JB, Cantor SB, Varma DG, Holst D, Feig BW, Hunt KK, et al. Utility of chest computed tomography for staging in patients with T1 extremity soft tissue sarcomas. Cancer 2001; 92:863–868.

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Porter GA, Cantor SB, Ahmad SA, Lenert JT, Ballo MT, Hunt KK, et al. Cost-effectiveness of staging computed tomography of the chest in patients with T2 soft tissue sarcomas. Cancer 2002; 94:197–204. Picci P, Vanel D, Briccoli A, Talle K, Haakenaasen U, Malaguti C, et al. Computed tomography of pulmonary metastases from osteosarcoma: the less poor technique. A study of 51 patients with histological correlation. Ann Oncol 2001; 12:1601–1604. Pitman AG, Hicks RJ, Binns DS, Ware RE, Kalff V, McKenzie AF, et al. Performance of sodium iodide based (18)F-fluorodeoxyglucose positron emission tomography in the characterization of indeterminate pulmonary nodules or masses. Br J Radiol 2002; 75:114–121. Lucas JD, O’Doherty MJ, Wong JC, Bingham JB, McKee PH, Fletcher CD, et al. Evaluation of fluorodeoxyglucose positron emission tomography in the management of soft-tissue sarcomas. J Bone Joint Surg Br 1998; 80:441–447.

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Original article

The ratio of the apex/anterior wall: A marker of breast attenuation artifact in women Christopher L. Hansen and Senthil Sundaram Background Breast attenuation artifact is well known for reducing the accuracy of myocardial perfusion imaging in women. We have noticed the particular pattern of relative preservation of apical activity in women with breast attenuation and decreased anterior wall counts. This study was undertaken to see if this finding could be used to improve the accuracy of perfusion imaging in women. Methods We identified 295 women referred for exercise stress testing using 201Tl single photon emission computed tomography (SPECT) of whom 193 had less than 5% probability of coronary disease and 102 had coronary artery disease documented by catheterization within 60 days of stress testing (mean of 1.8 ± 0.8 vessels with = 50% stenosis). Patients with documented myocardial infarction, pathologic Q waves, left bundle branch block, non-ischaemic cardiomyopathy or prior bypass grafting were excluded. Volume-weighted bullseye plots were generated and normalized to 100; next, regions of interest were drawn over the anterior wall and apex in all patients and the ratio of the mean counts of each region was calculated. The normals were further divided into those with breast attenuation (defined as mean anterior counts < 70% maximum) and those without. Defect scores of all patients were calculated; a formula to adjust the score for patients with breast attenuation was developed. Accuracy was assessed by calculating the area under the receiver operating curve.

Introduction Coronary artery disease (CAD) is the leading cause of death in women in industrialized nations. Noninvasive evaluation of women for CAD is hampered by the lower accuracy of EKG stress testing in women [1]. We have previously shown that the accuracy of single photon emission computed tomography (SPECT) 201Tl perfusion imaging in women is lower than in men [2]. Although in this previous study much of the lower accuracy appeared to be due to differences in chamber size, it appears that other factors, such as breast attenuation, can reduce accuracy [3–5]. Genderspecific normal databases have been introduced to help compensate for this. More recently, attenuation correction has been introduced [6,7]. However, attenuation correction is not performed routinely in all laboratories and its clinical utility has been challenged [8].

Results The normals, overall, had a mean ratio of 1.0 ± 0.08 vs. 0.9 ± 0.16 for those with coronary disease (P < 0.0001). In normals with breast attenuation the ratio was 1.1 ± 0.08 compared to 0.99 ± 0.07 (P < 0.0001) without. By adjusting the anterior wall defect score in patients with apex/anterior ratio > 1 we were able to improve the accuracy from 0.808 ± 0.028 to 0.826 ± 0.027 (P < 0.01). Conclusions A ratio of the apex to the anterior wall > 1 is not physiological and suggests the presence of significant breast attenuation artifact. This finding can be used to produce a small but statistically significant improvement in the accuracy of quantitative thallium SPECT in women who have not undergone coronary bypass grafting. c 2006 Lippincott Williams Nucl Med Commun 27:803–806 & Wilkins. Nuclear Medicine Communications 2006, 27:803–806 Keywords:

201

Tl, CAD, women, SPECT artifacts

Temple University Hospital, Philadelphia, USA. Correspondence to Dr Christopher L. Hansen, Thomas Jefferson University, 925 Chestnut St, Mezzanine Level, Philadelphia, PA 19107, USA. Tel: + 1 215 955 5050; fax: + 1 215 955 7855; e-mail: [email protected] Received 27 April 2006 Accepted 30 June 2006

We have noted a particular pattern in women with significant breast attenuation: breast attenuation artifact frequently causes a greater decrease in anterior wall counts than the apex. The causes for this can often be seen on review of the raw data; the breast frequently overlaps the anterior wall but spares the apex resulting in relatively well preserved apical counts in the reconstructed image. This pattern is in contrast to that seen in patients with coronary disease: ischaemia of the anterior wall is almost always due to disease of the left anterior descending coronary artery which, almost invariably, involves the apex as well. Thus, this finding is not physiological and is suggestive of artifact. To further explore this finding, we focused on two goals: firstly to see if the ratio of apex to anterior wall counts was significantly different in patients with coronary disease, normals and normals with breast attenuation

c 2006 Lippincott Williams & Wilkins 0143-3636

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Fig. 1

Normal

Apex/Anterior = 1.23

70% LAD Stenosis

Apex/Anterior = 0.96 Corresponding examples of a normal (top) subject and a patient with ischaemia in the LAD territory (bottom) are shown. The apical, mid and basal short axis are shown, with the vertical long axis and corresponding bullseye plots and regions. The ratio of the apex to anterior wall was 1.23 in the normal subject and 0.96 in the patient with LAD ischaemia.

artifact; and, secondly, to see if this finding could be exploited to improve the diagnostic accuracy of myocardial perfusion imaging in women.

Methods We identified two groups of women in the general population referred for exercise 201Tl SPECT. The first was 193 women (age 48 ± 11 years) who had < 5% probability of coronary disease [9] (normals) and the second was 102 women (age 61 ± 10 years) who had catheterization documented coronary disease within 60 days of stress testing (patients). Subjects with previously documented myocardial infarction, pathological Q waves or left bundle branch block on EKG, non-ischaemic cardiomyopathy or prior coronary artery bypass graft were not included. The patients had a mean of 1.8 ± 0.8 vessels with 50% stenosis (patients). Seventy-six (75%) of the patients had lesions affecting the left anterior descending artery distribution (left main lesions were included as affecting the LAD). Patients were exercised on a Bruce or modified Bruce protocol according to their reported exercise tolerance. At peak exercise 110–130 MBq (3–3.5 mCi) of 201Tl were injected and the subjects were encouraged to continue exercising for at least another minute. SPECT recon-

struction was performed using filtered back-projection, volume-weighted bullseye plots were generated and normalized to 100. Quantitative analysis was performed by generating a corresponding volume-weighted bullseye plot identifying where thallium uptake fell more than 2 standard deviations below the mean of a gender-based normal database. Constant regions of interest representing the apex and anterior walls were used for all patients (Fig. 1). The ratio of apex to anterior wall for each patient was calculated by dividing the mean counts in the apical region by the mean counts in the anterior wall region. Women with breast attenuation artifact were identified as normals who had average mean anterior wall counts < 70% of maximal counts. A defect score for each study was calculated as the sum of the contributions from the entire anterior wall (including the apex) and the remainder of the left ventricle. A quantitative method to adjust the defect score based on the apex/anterior wall ratio was introduced by using two parameters. The first was a threshold, if the apex/anterior wall ratio was above the threshold, a reduction of the anterior contribution to the defect score was made. The second was a weighting factor that could vary from 0 to 1.

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Ratio apex/anterior wall Hansen and Sundaram 805

Statistical methods

Differences between groups were evaluated with the Student’s t-test using an appropriately adjusted P value. Accuracy was determined by performing receiver operating characteristics (ROC) analysis using the method of Metz et al. [10]. A P value = 0.05 was considered significant.

Results The ratio of apex to anterior wall was significantly higher in normals compared to patients (1.0 ± 0.8 vs. 0.9 ± 0.16; P < 0.0001). Nineteen of the normals had mean anterior wall counts less than 70% which classified them as having significant breast attenuation artifact. These women had an even greater difference in the apex to anterior wall ratio compared to those without (1.1 ± 0.8 breast attenuation, 0.99 ± 0.07 normals without attenuation; P < 0.0001). Box-and-whisker plots comparing normals with and without breast attenuation and patients with coronary disease are shown in Fig. 2.

Fig. 2

1.3 1.2 1.1 Ratio apex/anterior wall

If the ratio was above the threshold (implying the presence of breast attenuation), the contribution of the anterior wall to the total defect score was decreased multiplying it by the weighting factor. Finally, two post hoc methods were then attempted to try to improve accuracy, the first restricted the anterior region to the anteroseptal wall by splitting the region in Fig. 1 vertically and performing the analysis with only the anteroseptal half of the region. This was employed to exclude possible confounding by diagonal lesions affecting the anterolateral wall and sparing the apex. The second explored the use of a minimum threshold that the apical counts needed to be above before decreasing the contribution of the anterior wall counts to see if this could more correctly identify actual ischaemia in patients with breast attenuation.

1 0.9 0.8 0.7 0.6 All P < 0.001 0.5 CAD

NI No breast

NI breast

Box-and-whisker plots of the ratio of the apex to anterior wall in patients with coronary artery disease, normal subjects without and with breast attenuation artifact are shown. There was a highly significant difference between the groups. The plots show the mean (dot), median (horizontal line) 25th and 75th percentile (ends of box) and 10th and 90th percentiles (ends of whiskers), respectively.

chambers by reducing blurring by means of the use of deconvolving filters and by size-based normal databases, unfortunately, were disappointing [11].

Discussion

The difficulties created by breast attenuation have been appreciated for some time. Review of the raw tomographic images is sometimes helpful, but in our experience, this does not reliably predict the amount of attenuation artifact encountered on the reconstructed images. Initially, normal databases were introduced as a device to give readers greater comfort with the range of normal in women. The advent of gated SPECT has allowed readers to identify mild fixed defects as artifact when the wall motion was normal [12]. However, confidence in this technique is less when there are differences in breast positioning between the rest and stress images. Clearly, the ideal approach would be accurate attenuation correction. Although initial reports are encouraging, it requires hardware that is not available for all cameras and clinical experience has not always been consistent [8].

Improving the accuracy of myocardial perfusion imaging in all patients is an important goal; the factors reducing accuracy in women present special challenges. The two main causes that decrease accuracy in women appear to be smaller chamber size and the highly variable effects of breast attenuation [2–5]. The results of our previous attempts to address the problems found in smaller

There are several points about our technique that deserve mention. The first is that breast attenuation artifact offers no protection against coronary disease; obviously women can have both. For this reason, we decided to decrease but not eliminate the contribution from the anterior wall in women with an increased apex-

We found that by empirically choosing a threshold of 1 and a weighting factor of 0.3 we could produce a small, but statistically significant, increase in accuracy from 0.808 ± 0.028 to 0.825 ± 0.027 (P < 0.01). Evaluating the subgroup of patients with involvement of the LAD territory produced comparable results (0.796 ± 0.029 to 0.814 ± 0.029; P < 0.05). Neither of the post hoc modifications to the correction technique produced any further increase in accuracy.

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to-anterior wall ratio. We found that accuracy actually decreased if the weighting factor was decreased below 0.3. The second point is that there are sure to be women with milder degrees of LAD stenosis who have normal perfusion of the anterior wall but, because of coincidental breast attenuation artifact, appear to have anterior perfusion defects. In these patients the artifact coincidentally facilitates the correct interpretation of the scan. Both attenuation correction and the method we propose here will tend to transform the scans of these women into false negatives. However, we feel that it is best for clinicians to have an accurate measure of the actual perfusion instead of depending on compensating errors. Third, we excluded patients with prior bypass grafting in the current study and would advise caution in using it in patients who have had a prior bypass graft to the LAD. It is possible for these patients to have preserved perfusion to the apex and still have ischaemia in a more proximal portion of the anterior wall due to disease proximal to the insertion of the graft. However, this finding may still help in clinical management of these patients in that it would suggest either attenuation artifact or ischaemia in the presence of a patent graft and lead to the conclusion that it is less likely that the woman would benefit from repeat surgery. Also, it should be understood that isolated diagonal or ramus intermedius lesions will generally not involve the apex. In the authors’ experience these lesions tend to cause very characteristic antero-lateral perfusion defects somewhat different than what is seen with breast attenuation. Although our post-hoc analysis suggested that there was no confounding by diagonal lesions, the reader should be aware of this and consider such lesions before attributing them to breast attenuation. Finally, the correction algorithm we used was developed empirically to prove that this observation actually can increase accuracy. If it was deemed worthwhile to employ this routinely for the small increase in accuracy, the values for the parameters would first need to be validated in a much larger group of patients. After noting the differences between the groups as shown in Fig. 2, we anticipated a larger improvement in accuracy and were disappointed by the relatively small improve-

ment this technique provided. It may be that we correctly identified breast attenuation using conventional methods and that this technique may be more helpful for less experienced observers. It has been reported that the diagnostic benefit of gated SPECT becomes more in the increased confidence it gives the reader as opposed to an actual increase in accuracy [13]; our experience has been that this observation increases our confidence in identifying defects as breast attenuation artifact.

Conclusion Breast attenuation artifact tends to produce a pattern of decreased anterior uptake with relatively preserved uptake in the apex. Recognition of this pattern allows for a small but statistically significant improvement in accuracy of myocardial perfusion imaging in women.

References 1

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Kwok Y, Kim C, Grady D, Segal M, Redberg R. Meta-analysis of exercise testing to detect coronary artery disease in women. Am J Cardiol 1999; 83:660–666. Hansen C, Crabbe D, Rubin S. Lower diagnostic accuracy of thallium-201 SPECT myocardial perfusion imaging in women: an effect of smaller chamber size. J Am Coll Cardiol 1996; 28:1214–1219. Dunn R, Wolff L, Wagner S, Botvinick E. The inconsistent pattern of thallium defects: a clue to the false positive perfusion scintigram. Am J Cardiol 1981; 48:224–232. Goodgold H, Rehder J, Samuels L, Chaitman B. Improved interpretation of exercise Tl-201 myocardial perfusion scintigraphy in women: characterization of breast attenuation artifacts. Radiology 1987; 165:361–366. Beller G. Diagnostic accuracy of thallium-201 myocardial perfusion imaging. Circulation 1991; 84:I1–I6. Ficaro E, Fessler J, Ackermann R, Rogers W, Corbett J, Schwaiger M. Simultaneous transmission–emission thallium-201 cardiac SPECT: effect of attenuation correction on myocardial tracer distribution. J Nucl Med 1995; 36:921–931. Miles J, Cullom S, Case J. An introduction to attenuation correction. J Nucl Cardiol 1999; 6:449–457. Th Wackers F. Attenuation compensation of cardiac SPECT: a critical look at a confusing world. J Nucl Cardiol 2002; 9:438–440. Diamond G, Forrester J. Analysis of probability as an aid in the clinical diagnosis of coronary-artery disease. N Engl J Med 1979; 300:1350–1358. Metz C, Wang P-L, Kronman H. A new approach for testing the significance of differences between ROC curves measured from correlated data. In: Deconinck F (editor). Information processing in medical imaging. The Hague: Martinus Nijhoff; 1984, pp. 432–445. Hansen C, Kramer M, Rastogi A. Lower accuracy of Tl-201 SPECT in women is not improved by size-based normal databases or Wiener filtering. J Nucl Cardiol 1999; 6:177–182. DePuey E, Rozanski A. Using gated technetium-99m-sestamibi SPECT to characterize fixed myocardial defects as infarct or artifact. J Nucl Med 1995; 36:952–955. Smanio P, Watson D, Segalla D, Vinson E, Smith W, Beller G. Value of gating of technetium-99m sestamibi single-photon emission computed tomographic imaging. J Am Coll Cardiol 1997; 30:1687–1692.

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Original article

A method for quantitative cell tracking using SPECT for the evaluation of myocardial stem cell therapy Robert Z. Stodilkaa,b,c, Kimberley J. Blackwooda,b, Huafu Konga and Frank S. Pratoa,b,c Purpose A promising SPECT-based method for evaluating stem cells therapy uses 111In-labelled cells, transfected with a reporter gene. Cells are first transplanted to the infarct, and subsequently interrogated for transgenic expression using a systemic injection of an 131I-labelled reporter probe. The method is impeded by the physical effects of scatter, 131I/111In cross-talk, and attenuation. We hypothesize that correcting for physical effects improves detection of transgenic expression in transplanted cells when 111In localization is available. Methods Canine bone marrow mesenchymal cells (BMMCs), radiolabelled and transfected, were injected into infarcted myocardium. Next, a reporter probe was injected systemically, and 22 SPECT scans were acquired over 20 h. Finally, 99mTc-sestamibi was injected and imaged. The animal was killed, the heart sectioned, and counted for 131 I and 111In in a well-counter (‘gold standard’). Canine SPECTs were reconstructed in two ways: with corrections for physical effects and without corrections. The first 111In reconstruction and the 99mTc reconstruction were used to define volumes-of-interest over the transplanted BMMC (VBMMC) and normal myocardium (VNM), respectively. Results 131I reconstructions without corrections for physical effects had negligible differential uptake. With corrections, VBMMC was consistently higher than VNM,

Introduction Work performed primarily in small animals suggests that transplantation of stem cells, either embryonic or adult, may have the potential to reduce the incidence of heart failure through the development of new myocardial tissue [1,2]. Currently, there are significant issues given that recent studies have refuted some of the original findings [3,4]. Preliminary clinical trials have indicated benefit [5] although the mechanism governing this benefit is not understood. Thus, external non-invasive monitoring would be useful in understanding the underlying mechanisms of myocardial stem cell therapy. Three aspects of evaluating cardiac stem cell therapy using an imaging modality are: (1) tracking transplanted cells and quantifying cell number; (2) interrogating the transplanted cells for function, including differentiation and engraftment; and (3) monitoring the status of underlying

demonstrating transgene expression. 131I had the following VBMMC:VNM activity ratio: without correction for physical effects = 0.869; with corrections = 1.23; and well-counter = 1.21. VNM showed the following 131I:111In activity ratio: without corrections = 3.07; with corrections = 1.38; and well-counter = 1.58. Conclusions In dual-isotope SPECT, corrections for physical effects were required to detect transgene expression in cells transplanted into an infarction when localization information was available. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:807–813
Nuclear Medicine Communications 2006, 27:807–813 Keywords: stem cell, SPECT cardiac imaging, reporter gene imaging, image reconstruction, cardiology, molecular imaging a Imaging Program, Lawson Health Research Institute, bDepartment of Medical Biophysics, University of Western Ontario and cDepartment of Nuclear Medicine, St Joseph’s Health Care, London, Ontario, Canada.

Correspondence to Dr Robert Z. Stodilka, Department of Nuclear Medicine, St Joseph’s Health Care–London, 268 Grosvenor Street, London, Ontario, Canada N6A 4V2. Tel: + 001 (519) 646 6000 ext. 64657; fax: + 001 (519) 646 6135; e-mail: [email protected] Received 5 February 2006 Accepted 28 June 2006

tissue. The use of nuclear medicine has been of value in the evaluation of these parameters. Single photon emission computed tomography (SPECT) has been used to track 111In-labelled transplanted progenitor cells in murine and porcine models of myocardial infarction for as long as 14 days [6,7]. Regarding functional assessment, cardiac transgene expression using herpes simplex virus thymidine kinase (HSV1-tk) was first demonstrated in a porcine model by Bengel et al. [8] using PET and the systemically administered reporter probe 20 -fluoro-20 deoxy-5-[124I]iodo-1-b-D-arabino-furanosyluracil (FIAU). Finally, both SPECT and positron emission tomography (PET) are widely acknowledged for their ability to measure myocardial perfusion and viability. Blackwood et al. [9] have recently demonstrated the potential for using multi-isotope SPECT to measure these three parameters simultaneously.

c 2006 Lippincott Williams & Wilkins 0143-3636


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808 Nulear Medicine Communications 2006, Vol 27 No 10

The purpose of this work is to evaluate the importance of scatter, attenuation and cross-talk correction for the application of tracking transplanted cells. In evaluating these parameters, monitoring of autologous bone marrow cell therapy is accomplished in a canine model of myocardial infarction. In our experiment, we use dualisotope SPECT to simultaneously track radioactively labelled cells transplanted into an infarct and monitor transgene expression in those cells using a reporter probe.

Methods Scatter and cross-talk correction

Scatter was corrected using deconvolution scatter subtraction [22,23]. Scatter subtraction kernels, which also include geometric collimator response, were based on experimental measurements of line spread functions, and modelled as circularly symmetric decaying single exponentials, and were applied separately for 131I and 111In, and 99mTc. The effects of cross-talk (XT) were corrected by extending the deconvolution scatter subtraction methodology as follows. Consider the simultaneous 131 I and 111In imaging situation where projections are recorded in energy windows, WI and WIn, corresponding, respectively, to the emission energies of the two radioisotopes. In what follows, WI was defined as an energy window centred at 364 keV with 20% width; and WIn was defined as two summed energy windows centred at 171 keV and 245 keV, each with 20% width. The projection in WI, TWI, including the effects of resolution and scatter, can be expressed as a function of the spatial coordinate x: TWI ðxÞ ¼ PWI ðxÞ  QWI ðxÞ;

ð1Þ

Fig. 1

1

0.8 Intensity (arbitrary units)

Multi-isotope SPECT [10–12] provides the benefits of decreased scan time, true simultaneous acquisition, and perfect registration of images to maximize the benefit of data fusion, but it is more demanding on hardware and complicates post-acquisition processing. Images can be misinterpreted without adequate correction for Compton scatter, attenuation and radionuclide cross-talk [11,13]. For example, in the case of simultaneous 99mTc/201Tl SPECT, 99mTc gamma rays with an energy of 140 keV can be detected in a 201Tl energy window centred at 72 keV, due to energy loss to Compton scattering [14]. This cross-talk can lead to significant overestimation of activity and loss of contrast in small lesions [13,15–17]. Many authors discuss the need for correcting for physical degradation processes [10,11,15,18–20], yet implementation remains controversial and inconsistent. For example, O’Connor et al. [21] highlight the importance of attenuation correction, but demonstrate variability in quantitative accuracy, especially in non-uniform attenuation correction, depending upon hardware design and reconstruction technique.

0.6

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Position (mm) The line spread function of 131I in WI: measured data (  ) and corresponding analytical fit (thick solid) from which the circularly symmetric QWI is derived; the line spread function of 131I in WIn: measured data (o) and corresponding analytical fit (thin solid) from which the circularly symmetric QXTWIn is derived; and the transfer function mapping QWI to QXTWIn, QTFWI!WIn (dotted line). The ratio of the areas under (QXTWIn): (QWI) is 1.16, which is also the area under QTFWI!WIn . Note that QTFWI!WIn is rescaled for display purposes. (See text for definitions of the variables.)

where PWI is the 131I projection that is free of resolution and scatter effects, # is a two-dimensional convolution operator, and QWI is a model of the summation of these degrading effects and is circularly symmetric. The projection in WIn is expressed similarly with an additional term describing 131I cross-talk into WIn [24]: TWIn ðxÞ ¼ PWIn ðxÞ  QWIn ðxÞ þ PWI ðxÞ  QXTWIn ðxÞ;

ð2Þ

131

where QXTWIn is the projection of a I point source measured in WIn (Fig. 1). Using the Fourier transform operator, F, we now define QTFWI!WIn as a transfer function mapping the projection of a 131I point source from WI to WIn:  QTFWI!WIn ðxÞ ¼ F1 F½QXTWIn ðxÞ=F½QWI ðxÞ : ð3Þ This transfer function, together with Equation 1, allows us to re-write the second term of Equation 2 in terms of the measurable quantity TWI: PWI ðxÞ  QXTWIn ðxÞ ¼ TWI ðxÞ  QTFWI!WIn :

ð4Þ

131

Thus, the I cross-talk in the measured projection TWIn is estimated by convolving the measured projection TWI with QTFWI!WIn . The effects of cross-talk in TWIn are then removed by subtracting the quantity of Equation 4 from TWIn.

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Tracking stem cells in SPECT Stodilka et al. 809

Phantom imaging

In order to calibrate the cross-talk correction, two 1 mm  100 mm line sources were prepared: a 0.89 MBq 131 I line source, and a 1.2 MBq 111In line source. The line sources were placed separately along the axis of a cylindrical water-filled phantom with dimensions of 150 mm diameter and 200 mm length, and imaged using a gamma camera. The gamma camera was a dual-head Millennium MG (General Electric, Milwaukee, Wisconsin, USA) equipped with medium energy, parallel hole collimators. For each line source, a single projection was acquired with a 230 mm radius of rotation (centre-ofrotation to imaging plane) and an acquisition time of 600 s. The projection field-of-view had dimensions of 343  343 mm and events were digitized into a 128  128 pixel matrix. The WIn energy window was used to image the 111In line source, while the 131I line source was acquired using both the WI and WIn energy windows. In order to obtain QWI and QXTWIn from the experimental data and to then stabilize the division operation in Equation 3, we fitted each line spread function to the sum of a Gaussian which models resolution effects, and an exponential which models scatter [23]. Creation of infarct, cell preparation and transplantation

Our protocol for creating a stable infarction in canine myocardium has been described previously in detail [25]. General anaesthesia was induced in a 23-kg female mongrel dog using propofol and maintained using 2.5% isofluorane via endotracheal intubation. A left thoracotomy was created to access the left anterior descending artery, around which a stenosing device was placed and then the thoracotomy was closed. An infarct was created by a 2-h occlusion using the stenosing device, followed by reperfusion. The presence and extent of the infarct was confirmed using Gd-DTPA enhanced T1-weighted MRI (not shown). Seven months after the creation of the infarct, general anaesthesia was induced in the dog, and 10 ml of bone marrow was aspirated by syringe from the marrow cavity of the sternum and mixed with heparinized saline. Mononuclear cells were isolated from the aspiration by centrifugation, plated at a density of 2 105 cellscm – 2 in growth medium (Dulbecco’s Modified Eagle Medium) with antibiotics and incubated. Non-adherent cells were washed away; remaining bone marrow mesenchymal cells (BMMCs) were incubated and culture expanded for another 10 days. BMMCs were then lipid-transfected with HSV1-tk which also contained a gene conferring antibiotic resistance. Two days after transfection, thymidine kinase positive cells were selected through a 10-day incubation in a growth medium containing the antibiotic G418 (200 mg). Surviving cells were further incubated for 10 days in a maintenance dose of G418 (50 mg).

Prior to transplantation, transfected BMMCs were incubated with 0.3 MBq 111In-tropolone, resulting in a labelling of approximately 0.048 Bq per cell (80% labelling efficiency), which does not affect cell proliferation potential for at least 2 months [26]. Transfected radiolabelled BMMCs (5  106) were finally suspended in 1.5 ml saline and transplanted by multiple injections into the myocardium of the same dog. BMMC transplantation took place during a second thoracotomy where infarcted myocardium was identified by its discoloration relative to the rest of the myocardium. Canine SPECT

Thirty minutes after the second thoracotomy was closed, the dog was moved to the SPECT suite and imaging commenced. The imaging parameters were identical to those previously described, except SPECT was acquired: 64 projections over 1801 of rotation for each gamma camera. In total, 22 SPECT scans were acquired over 20 h. Our nominal acquisition time was 55 s per projection, allowing one complete scan every hour. However, at the beginning of the experiment, a shorter scan time (20 s per projection) was used to capture possible higher-order kinetics of cell migration from the site of transplantation. For the initial WIn SPECT acquisition (20 s per projection), no 131I was present, and count rate was approximately 0.5  103 counts per second (cps) in WIn. After the first SPECT acquisition, 43.6 MBq 131I-FIAU was injected in a peripheral vein. For the next SPECT acquisition, count rate in WI was approximately 2.7  103 cps, and count rate in WIn had increased to 3.3  103 cps due to cross-talk contamination. Acquisition time was maintained at 20 s per projection. After 5 h, the count rate in WI dropped to approximately 2.3  103 cps, and the acquisition time was increased to 55 s per projection. Data in this manner was acquired for an additional 15 h. For the last scan, 80.0 MBq 99mTc-sestamibi was injected, resulting in approximately 20  103 cps in the 140 keV 20% 99mTc energy window (WTc). All projections were corrected for gamma camera uniformity, linearity and radioisotope decay and normalized to the same acquisition time. Projection data was processed both with and without correction for the physical effects of scatter, cross-talk and attenuation. When corrections were applied, WI projection data were corrected for scatter, attenuation and distance-dependent resolution recovery, and WIn projection data were also corrected for cross-talk. Projections were reconstructed using iterative reconstruction [27]. The uniform attenuation map was derived by an edge detection algorithm that delineated the outer boundaries on the WTc reconstruction and assigned a radionuclide-specific narrow-beam uniform attenuation coefficient to all pixels within that boundary: 0.15 cm – 1

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810 Nulear Medicine Communications 2006, Vol 27 No 10

for 99mTc [28], 0.135 cm – 1 for 111In (which considered summing of the 171 keV and 245 keV energy peaks) [29], and 0.11 cm – 1 for 131I [30]. When corrections were not applied, reconstruction involved only distance-dependent resolution recovery. Ex-vivo counting

At the conclusion of SPECT imaging, the dog was killed and the heart excised. The left ventricle was segmented from the heart, and cut into two groups of small sections: a group of sections comprising discolored myocardium, and a group of sections comprising normal myocardium. Each section of myocardium was counted for 111In and 131 I in a high-purity germanium (HPGe) well-counter (GWL Ortech; Oak Ridge, Tennessee, USA). Of primary interest was the 131I activity ratio of discolored myocardium:normal myocardium. The HPGe well-counter measurement of this ratio was considered to be the ‘gold standard’. Image quantification and time–activity curves

The WTc reconstructions were used as a guide to draw manually a volume-of-interest around the region of the heart with normal 99mTc uptake, which was assumed to be normal myocardium. This was done on the WTc reconstruction without corrections for physical effects, and the volume-of-interest labelled VNM; and on the WTc reconstruction with corrections for physical effects, where the volume-of-interest was labelled VNMCORR. On the WIn data at the first time point (i.e., prior to 131I injection), a single focus of activity was visualized, and this was assumed to be the location of the BMMCs. Beginning with the WIn reconstruction without correction for physical effects, the 10 voxels with the highest activity where identified. A volume-of-interest was drawn automatically to enclose all voxels with more than 10% of the average activity of these 10 hottest voxels. This volume was labelled VBMMC. This procedure was repeated on the WIn reconstruction that was corrected for physical effects, yielding another region,

VBMMCCORR. Voxels from VBMMC or VBMMCCORR that overlapped with the normal myocardium volumes-ofinterest (VNM and VNMCORR) were deleted from the latter. To create time–activity curves, VBMMC and VNM were applied to the reconstructions without correction for physical effects, and VBMMCCORR and VNMCORR were applied to reconstructions with corrections. Quantitative accuracy of the SPECT reconstructions was assessed by calculating activity ratios, such as VBMMC:VNM, at the end of the time–activity curves and comparing these ratios with the corresponding ratios obtained from the measurements of the HPGe well-counter. Biological half-life of activity was estimated for VBMMC, VNM and their corrected counterparts by fitting each time–activity curve to an analytical mono-exponential equation. For WIn data, analytical equations were fit to the entire time–activity curve. For WI data, to minimize the influence of 131I-FIAU blood pool kinetics, the analytical equation was fit to only the last 10 h of data (i.e., the second half of the experiment).

Results Figure 1 shows profiles through the experimental measurements of the 131I line-spread functions, together with analytical fits of QWI, QXTWIn, and QTFWI-WIn. Sample images from the WTc, WIn, and WI reconstructions are shown in Fig. 2. The myocardium was visualized well on the WTc reconstruction, and demonstrated a region of reduced activity corresponding to the infarct identified on the MRI. The WIn reconstructions were characterized by a single focus of increased activity that remained on all WIn reconstructions throughout the course of the experiment. This focus corresponded to the location of reduced activity in the WTc reconstructions, which could be verified since WTc and WIn were inherently registered. WI reconstructions were diffuse and lumpy, and activity was also noticed in the stomach and esophagus (not shown).

Fig. 2

(a)

WIn +WTc

(b) WI

(c)

WIn + WTc

(d)

BMMC

WI BMMC

NM Corr

Corr

Reconstructed transaxial slices. (a) Two-colour composite, where WIn = green scale (10.5 h), and WTc = red scale. (b) Blue scale of WI (10.5 h). a and b are without corrections for physical effects; c and d are with corrections for physical effects. Labels: NM = normal myocardium, BMMC = site of transplanted bone marrow mesenchymal cells. Note contrast improvement due to correction for physical effects. WIn images were used to draw the BMMC volume-of-interest, the delineation of which is improved by corrections. The dog was kept stationary throughout imaging. All images are registered.

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Tracking stem cells in SPECT Stodilka et al. 811

Correction for physical effects was found to improve visual contrast in the WI reconstructions, making uptake of 131I in the location corresponding to the transplanted BMMC apparent, while reducing background in surrounding regions. However, despite these corrections, WI reconstructions remained diffuse and lumpy. In the WIn reconstruction, corrections were found to improve contrast of the single focus, while again reducing reconstructed activity in the surrounding region. Correction for physical effects improved the accuracy of quantifying, in a relative sense, reconstructed activity in the WI window,

Fig. 3

Activity (counts/voxel)

(a)

Monoexponentials were fitted to time–activity curves (see Fig. 3) for WI and WIn data and the volumes of interest corresponding to the bone-marrow mesenchymal cells (VBMMC) and normal myocardium (VNM), without and with correction for physical effects (CORR subscript). The table shows the equations, where the t variable denotes time in hours, and (inc) denotes increasing activity

Table 1

Window

Volume of interest

Equation

Uncorrected curves WI VBMMC 0.135e – 0.035t WI VNM 0.0980e – 0.0169t WIn VBMMC 0.253e – 0.0518t WIn VNM 0.0201e + 0.0139t Curves corrected for physical effects 0.0990e – 0.053t WI VBMMCCORR 0.0262e – 0.0137t WI VNMCORR WIn VBMMCCORR 0.571e – 0.0502t 0.0148e + 0.0102t WIn VNMCORR

R2

Half-life (h)

0.784 0.768 0.844 0.0459

19.8 41.0 13.4 49.9 (inc)

0.633 0.144 0.873 0.292

13.1 50.6 13.3 67.9 (inc)

WIn

1

and as well as relating activity between the WI and WIn reconstructions. For the endpoints of the WI time activity curves, the ratio of VBMMC:VNM (i.e., without correction for physical effects) activity was 0.869. When corrections were applied, the ratio was 1.23, which compared favourably with the HPGe well-counter ratio of 1.21. Of parenthetical interest were activity ratios between WI and WIn. For VNM, WI:WIn = 3.07 and VNMCORR WI:WIn = 1.38, compared with the goldstandard ratio of 1.58.

0.1

0.01 0

5

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Time (hours)

Activity (counts/voxel)

(b)

WI

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Time (hours) (a) Time–activity curves for WIn from volumes-of-interest around normal myocardium (VNM = *) and bone marrow mesenchymal cells (VBMMC = ~) without corrections for physical effects. Filled symbols represent curves corrected for physical effects (VNMCORR = K and VBMMCCORR = ~). Single exponential fits are superimposed, where solid lines are for VNM and VNMCORR and dotted lines are for VBMMC and VBMMCCORR. (b) Corresponding curves for WI. Note that for WI, only the last 10 h of data is used in curve fitting. Curves represent average activity per voxel from their respective volumes-of-interest, are shown on a log scale, and have been corrected for physical decay.

Time–activity curves are shown in Fig. 3. For WIn, the curves confirmed the high contrast visualized in the WIn reconstructions. Throughout the course of the experiment, the volume-of-interest corresponding to BMMC transplantation had activity that was substantially increased over the background of normal myocardium. In the region of BMMC transplantation, WIn activity had a rapid washout during the very initial hours of imaging, after which washout declined. Compared with the WIn time–activity curves, the WI curves feature substantially less contrast between volumes-of-interest corresponding to BMMC transplantation and the background of normal myocardium. As with the WIn time–activity curves, the WI curves also demonstrate rapid washout during the very initial hours of imaging which declines with time. Time–activity curves were fit to mono-exponentials, and the results are presented in Table 1. Activity was found to clear from the BMMC volumes-of-interest (VBMMC and VBMMCCORR) much more rapidly than the normal myocardium volumes-of-interest. In the case of WIn for the normal myocardium volumes-of-interest (VNM and VNMCORR), activity was found to be increasing, albeit very slowly. Correction for physical effects was found to impact calculated half-lives in most cases. The half-lives for WI and WIn in the normal myocardium were increased after corrections were applied. The half-life of WI in the BMMC volume-of-interest was found to decrease from

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812 Nulear Medicine Communications 2006, Vol 27 No 10

19.8 to 13.1 h after applying corrections. The half-life of WIn in the BMMC volume-of-interest was approximately 13 h, and was not substantially altered by correction for physical effects.

Discussion Two mechanisms that have been proposed for cell tracking are direct loading of cells with a contrast agent [6,7,31,32], and enzymatic conversion and retention of a contrast substrate [33]. In this experiment, we used both mechanisms, but relied on the first method for cell tracking. Direct loading is simpler experimentally but is compromised by both dilution effects following cell division, and a tradeoff between radioisotope half-life and long-term exposure to ionizing radiation [26,34]. However, direct loading was found to provide a specific signal, and in conjunction with a short-lived radiotracer, may be useful in the earliest stages of monitoring transplanted cells. The second mechanism, enzymatic conversion of a contrast substrate, does not suffer from dilution with cellular division. It could be used to follow cell populations indefinitely assuming stable transgene integration, and can be used to interrogate various aspects of cellular function [34]. In our study, we found that the reporter probe 131I-FIAU accumulated within the location of the transplanted cells, but accumulation was not specific to that location. Without a priori localization information from the 111In-tropolone direct loading mechanism, in-vivo quantification of 131I accumulation would have been questionable. Time–activity curves show that both WI and WIn activity decreases from the region corresponding to BMMCs with a biological half-life of between 13 and 19 h, depending on whether physical effects are corrected. We have previously determined 111In-tropolone washout from viable canine BMMC in vitro to have a half-life of 6.78 days [26]. Our faster 111In in-vivo kinetics may be due to BMMCs leaving the site of transplantation or cell death [35]. Kinetic analysis of WI data was complicated both by the potential of blood pool activity, since 131IFIAU was administered intravenously; and by systemic degradation of labelled FIAU [8,36]. We assumed these effects were transitory, and that after 10 h no additional FIAU would enter transfected BMMC or normal myocardium. Thus, we used only the last 10 h of data in the kinetic analysis of the WI time–activity curves. Correction for the effects of scatter, cross-talk and attenuation improved relative quantification of data as verified by the HPGe well-counter, and had a marked impact on time–activity curves by making 131I and 111In efflux from the BMMC site more consistent. In general, applying scatter and cross-talk corrections has the effect of decreasing count density. Deconvolution scatter subtraction also has the effect of correcting for the

geometric response of the collimator [37], which further decreases count density. Geometric response includes septal penetration, which increases with energy and, in the case of 131I imaged with a medium-energy collimator, can lead to a 10–40% overestimation of reconstructed activity [38]. Conversely, attenuation correction increased count density, especially for lower energy gamma rays due to the energy dependence of attenuation coefficients. Taken together, these corrections decreased the average count density in WI and improved image contrast in general, allowing for improved delineation of the myocardium and the transplanted cell colony. Improved region-of-interest delineation, together with improved reconstruction accuracy, resulted in further separation between corrected time–activity curves (VNMCORR and VBMMCCORR for WI and WIn) compared with uncorrected time–activity curves. The purpose of the present work was to demonstrate a method for detecting radiolabelled cells that were transplanted for the purpose of providing myocardial stem cell therapy, and to determine whether correction for physical effects would impact kinetic analysis of this imaging situation. We refrain from making biological conclusions. As such, this work could have been performed in a realistic canine phantom that modelled the kinetics of cell migration from the myocardium, although such a phantom is not available currently. Future work could include the modelling of degradation kinetics of iodine labelled FIAU in the blood pool of a canine model, and imaging more than one dog, which would allow for a biological analysis.

Conclusion In the application of stem cell tracking, dual-isotope SPECT requires corrections for the physical effects that degrade the imaging process. With these corrections in hand, SPECT can be used to track in vivo the location of radiolabelled cells transplanted into a canine heart as well as interrogate the functional capacity of these cells using a peripherally injected contrast agent for at least 20 h. Many imaging modalities are being evaluated for utility in cardiac stem cell therapy and all offer unique strengths and weaknesses in tackling such a multi-faceted challenge. In this application, the strength of SPECT lies neither with resolution nor sensitivity. Rather, we have shown that SPECT can image, simultaneously, multiple parameters associated with monitoring regenerative therapy, and it is in this multi-parameter imaging that the value of SPECT can be maximized.

Acknowledgements Funding was provided, in part, by a grant to Dr Prato from the Canadian Institutes of Health Research, and by additional funding from the Ontario Research and

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Tracking stem cells in SPECT Stodilka et al. 813

Development Challenge Fund to Drs Stodilka and Prato. The authors would like to thank Dr Savita Dhanvantari for assistance with molecular biology techniques, Dr Peter Merrifield for assistance in cell population selection, Ms Jane Sykes for the canine surgery, and Dr Glenn Wells for assistance with SPECT imaging. All procedures in this experiment were done in accordance with the University of Western Ontario Animal Care Committee of the Council on Animal Care Guidelines, and comply with the current laws of Canada.

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Cao Z, Chen CC, Manoury C, Holder LE, Abraham TC, Tehan A. Phantom evaluation of simultaneous thallium-201/technetium-99m acquisition in single photon emission tomography. Eur J Nucl Med 1996; 23:1514–1520. Hannequin P, Mas J, Germano G. Photon energy recovery for crosstalk correction in simultaneous 99mTc/201Tl imaging. J Nucl Med 2000; 41:728–736. Constantinesco A, Mertz L, Brunot B. Myocardial perfusion and function imaging at rest with simultaneous thallium-201 and technetium-99m bloodpool dual-isotope gated SPECT. J Nucl Med 1997; 38:432–437. de Jong HWAM, Beekman FJ, Viergever MA, van Rijk PP. Simultaneous 99m Tc/201Tl dual-isotope SPET with Monte Carlo-based down-scatter correction. Eur J Nucl Med 2002; 29:1063–1071. O’Connor MK, Kemp B, Anstett F, Christian P, Ficaro EP, Frey E, et al. A multicenter evaluation of commercial attenuation compensation techniques in cardiac SPECT using phantom models. J Nucl Cardiol 2002; 9:361–376. Floyd CE, Jaszczak RJ, Greer KL, Coleman RE. Deconvolution of Compton scatter in SPECT. J Nucl Med 1985; 26:406–408. Msaki P, Axelsson B, Larsson SA. Some physical parameters influencing the accuracy of convolution scatter correction in SPECT. Phys Med Biol 1989; 34:283–298. Stodilka RZ, Blackwood KJ, Kong H, Wells GR, Prato FS. Quantitative tracking and characterization of transplanted cells in an autologous canine model of myocardial infarction [Abstract]. J Nucl Med 2005; 46:339. Pereira RS, Prato FS, Sykes J, Wisenberg G. Assessment of myocardial viability using MRI during a constant infusion of Gd-DTPA: further studies at early and late periods of reperfusion. Magn Reson Med 1999; 42:60–68. Kong H, Blackwood KJ, Stodilka RZ, Wells RG, Wisenberg G, Merrifield P, et al. 111In-Tropolone labeling of canine bone marrow mesenchymal cells: in vitro viability and washout kinetics [Abstract]. J Nucl Med 2005; 46(suppl 2):119P. Byrne CL. Accelerating the EMML algorithm and related iterative algorithms by rescaled block-iterative methods. IEEE Trans Image Processing 1998; 7:100–109. NIST 1988 X-ray and Gamma Ray Attenuation Coefficient and Crosssection Database version 2.0. Gaithersburg, MD: US Dept of Commerce, National Institute of Standards and Technology. Gilland DR, Jaszczak RJ, Turkington TG, Greer KL, Coleman RE. Quantitative SPECT imaging with indium-111. IEEE Trans Nucl Sci 1991; 38:761–766. Ljungberg M, Sjogreen K, Liu X, Frey E, Dewaraja Y, Strand SE. A 3dimensional absorbed dose calculation method based on quantitative SPECT for radionuclide therapy: evaluation for 131I using Monte Carlo simulation. J Nucl Med 2002; 43:1101–1109. Gao J, Dennis JE, Muzic RF, Lundberg M, Caplan AI. The dynamic in vivo distribution of bone marrow-derived mesenchymal stem cells after infusion. Cells Tissues Organs 2001; 169:12–20. Barbash IM, Chouraqui P, Baron J, Feinberg MS, Etzion S, Tessone A, et al. Systemic delivery of bone marrow-derived mesenchymal stem cells to the infarcted myocardium: feasibility, cell migration, and body distribution. Circulation 2003; 108:863–868. Gambhir SS, Herschman HR, Cherry SR, Barrio JR, Satyamurthy N, Toyokuni T, et al. Imaging transgene expression with radionuclide imaging technologies. Neoplasia 2000; 2:118–138. Frangioni JV, Hajjar RJ. In vivo tracking of stem cells for clinical trials in cardiovascular disease. Circulation 2004; 110:3378–3384. van Reenan PC, Lotter MG, Heyns AD, de Kock F, Herbst C, Kotze H, et al. Quantification of the distribution of 111In-labeled platelets in organs. Eur J Nucl Med 1982; 7:80–84. Jacobs A, Braunlich I, Graf R, Lercher M, Sakaki T, Voges J, et al. Quantitative kinetics of [124I]FIAU in cat and man. J Nucl Med 2001; 42:467–475. Yanch JC, Flower MA, Webb S. A comparison of deconvolution and windowed subtraction techniques for scatter compensation in SPECT. IEEE Trans Med Imag 1988; 7:13–20. Gonzalez Trotter DE, Bowsher JE, Jaszczak RJ. Absolute quantification of a spherical I-131 activity distribution using a high-resolution rotating collimator: A phantom study. IEEE Trans Nucl Sci 2001; 48:65–73.

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Original article

Attenuation-corrected 99mTc-MIBI SPECT in overweight patients with chronic ischaemic dysfunction: a comparison to NH3 PET and implications for the diagnosis of myocardial viability Ve´ronique Roelantsa, Xavier Bernardb, Stephan Walranda, Anne Bolc, Ann Coppensc, Jacques Jamartd, Jacques Melina and Jean-Louis Vanoverscheldeb Objective We determined the value of attenuation correction (AC) of myocardial perfusion estimation with 99m Tc-MIBI SPECT in overweight patients by comparison of uncorrected (filtered back-projection (FBP) and corrected (an iterative algorithm with a measured attenuation coefficients map (FL-AC)) 99mTc-MIBI relative uptake to perfusion data obtained in the same patients with NH3 PET. In addition, the impact of attenuation correction for the assessment of myocardial viability with 99mTc-MIBI SPECT was determined using FDG PET as the reference method. Methods Thirty consecutive overweight patients (BMI = 28 ± 4) with left ventricular dysfunction underwent a resting 99mTc-MIBI SPECT and a PET study (NH3 and FDG). 99m Tc-MIBI SPECT scans were reconstructed without attenuation correction (FBP) and with attenuation correction (FL-AC). The left ventricle was divided into 16 segments, in which the relative uptake was quantified using circumferential profiles. A relative uptake Z 60% was considered consistent with viable myocardium for FDG and MIBI. Results The absolute difference between 99mTc-MIBI SPECT and NH3 PET uptakes was less pronounced in the inferior (12 ± 10% vs. 17 ± 12%, P < 0.001), anteroseptal (12 ± 11% vs. 16 ± 12%, P = 0.009) and septal (15 ± 12% vs.

Introduction The diagnostic accuracy of single photon emission computed tomography (SPECT) myocardial perfusion imaging is profoundly influenced by the presence of tissue attenuation. Several techniques such as additional right-side left lateral decubitus images [1], acquisition in supine–prone position [2] or acquisition in gating mode [3] have been shown to be useful in minimizing the impact of attenuation. However, since attenuation artifacts may vary from one imaging session to another and since patients with demonstrable attenuation artifacts also may have coronary artery disease in the same territories, none of these methods is able to provide a diagnosis of certainty with regards to the presence of such artifact. Specific hardware (external radionuclide source-

18 ± 14%, P = 0.003) regions (FL-AC vs. FBP, respectively). The sensitivity of MIBI for diagnosing myocardial viability increased from 83 to 100% (P = 0.034), without loss in specificity. Conclusion Attenuation correction improves myocardial perfusion estimation by 99mTc-MIBI SPECT in the inferior, anteroseptal and septal regions and increases its sensitivity for the diagnosis of myocardial viability. Nucl c 2006 Lippincott Williams & Med Commun 27:815–821 Wilkins. Nuclear Medicine Communications 2006, 27:815–821 Keywords: myocardial perfusion SPECT, attenuation correction, myocardial PET, myocardial viability Departments of aNuclear Medicine, bCardiology, Cliniques Universitaires St-Luc, Universite´ Catholique de Louvain, Brussels, cIMRE, Universite´ Catholique de Louvain, Louvain-la-Neuve and dDepartment of Statistics, Cliniques Universitaires de Mont-Godinne, Universite´ Catholique de Louvain, Mont-Godinne, Belgium. Correspondence to Dr Ve´ronique Roelants, Department of Nuclear Medicine, Cliniques Universitaires St-Luc, Avenue Hippocrate, 10, B-1200 Brussels, Belgium. Tel: + 32 2 764 25 81; fax: + 32 2 764 54 08; e-mail: [email protected] Received 8 May 2006 Accepted 1 July 2006

based systems or computed tomography-based systems) designed to acquire and reconstruct transmission data to create each patient specific attenuation map are now commercially available. The importance of an effective quality control of the system used was emphasized in the recent position statement of the American Society of Nuclear Cardiology and the Society of Nuclear Medicine on the use of attenuation correction in nuclear cardiology [4]. Generally, cardiac perfusion SPECT images obtained after non-uniform photon attenuation correction demonstrate more uniform regional activity distribution than the corresponding uncorrected images in patients with a low likelihood of coronary artery disease [5,6] and increase significantly the diagnostic performances of cardiac perfusion single photon emission computed tomography

c 2006 Lippincott Williams & Wilkins 0143-3636

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816 Nuclear Medicine Communications 2006, Vol 27 No 10

(SPECT) in various circumstances [7–10]. In addition, attenuation correction is particularly relevant to the assessment of myocardial viability with perfusion SPECT since it requires relative quantitation of tracer uptake after generation of circumferential profiles. Using fluorodeoxyglucose (FDG) positron emission tomography (PET) or FDG SPECT as the ‘gold standard’, attenuation correction was found to improve the ability of perfusion SPECT for the diagnosis of myocardial viability [11–13]. However, in all these studies uncorrected emission images were reconstructed using conventional filtered back-projection (FBP) while attenuation corrected images were reconstructed using iterative algorithms that incorporate a map of measured attenuation coefficients. Moreover, the impact of attenuation correction with an external radionuclide source-based system on the perfusion estimation by comparison of uncorrected and corrected 99mTc-MIBI SPECT data to the well admitted clinical gold standard for perfusion, i.e., [13N]ammonia PET (NH3 PET) has not yet been reported. Accordingly, the aim of the present study was to evaluate the effect of non-uniform attenuation correction on myocardial perfusion estimation with rest 99mTc-MIBI SPECT by comparison of values obtained after data reconstruction (1) by conventional FBP, (2) by an iterative algorithm without attenuation correction (FL) and (3) by an iterative algorithm that incorporates each patient measured attenuation map (FL-AC) to the values obtained in the same patients with NH3 PET. Additionally, using FDG PET as the gold standard, we determined the impact of attenuation correction for the assessment of myocardial viability by rest 99mTc-MIBI SPECT.

Materials and methods

camera. Studies were performed in a 1-week period and no clinical event or change in medication occurred between them. All subjects signed an informed consent form before participation in the study. The study was approved by our local ethics committee. Data acquisition Single photon emission computed tomography

Data were acquired 90& min after injection of 1110 MBq (30 mCi) of MIBI at rest using a three-head camera (Triad XLT 20, Trionix, Twinsburg, USA) equipped with low energy ultra-high resolution collimators. For the emission scan, 60 projections in a 64  64 matrix over 3601 were performed in gating mode (8 bins/ cardiac cycle, 25–35 beats/projection (10% tolerance/normal period)), 8.43 mm pixel size. The transmission scan was performed with a linear source of 153Gd, moving opposite to the parallel collimator. Thirty projections of 30 s duration each were acquired in a 64  64 matrix over 1801, immediately after the emission scan. At each acquisition angle, the source moved parallel to the long axis of the patient. Two images were acquired simultaneously: the first one corresponds to the events occurring inside a 9 mm crystal strip in front of the translating linear source (153Gd + 99mTc events), and the second one corresponds to the events occurring outside this strip (99mTc events only). The outside image was normalized to correspond to the same acquisition time per pixel than the inside image. Finally, a 153Gd image free of downscatter was obtained by subtracting the normalized outside image to the inside image. The transverse linear source was collimated along the longitudinal direction to minimize patient exposure. Immediately after the transmission scan, a 5 min static image blank scan was acquired to compute patient attenuation coefficients from the transmission data.

Patient population

The study group included 30 consecutive patients with chronic left ventricular ischaemic dysfunction who were referred for evaluation of myocardial viability. There were 28 men and two women with a mean age of 63 ± 10 years (range, 44–79 years) and with a mean body mass index (BMI) of 28 ± 4 (range, 18–35; BMI < 25: n = 5; BMI between 25 and 29: n = 18; and BMI Z 30: n = 7). Twenty-five of them had history of prior myocardial infarction. Their mean global left ventricular ejection fraction (LVEF) obtained by radionuclide ventriculography was 32 ± 11%, ranging from 13 to 58%. Fifteen patients had a LVEF Z 30%, 12 had an LVEF between 31 and 49% and the remaining three patients had a normal LVEF. The regional left ventricular function was assessed by two-dimensional echocardiography. Coronary angiography revealed significant stenosis of three vessels in 20, significant stenosis of two vessels in nine, and one vessel disease in the remaining patient. All patients underwent a resting 99mTc-MIBI SPECT and a NH3/FDG PET study in random order, according to the availability of each

Positron emission tomography

Patients were studied during a hyperinsulinaemic–euglicaemic glucose clamp, as previously described by De Fronzo et al. [14] and by Knuuti et al. [15]. Briefly, insulin was infused intravenously at an initial rate of 160 mU
min – 1
m – 2 for 4 min, then reduced to 80 mU
min – 1
m – 2 for 3 min and maintained thereafter at a constant rate of 40 mU
m – 1
m – 2 until the end of the study. Glucose (20%) was co-administered, starting 4 min after the beginning of the insulin infusion. The glucose infusion rate was adapted to maintain glucose plasma levels between 75 and 95 mg
dl – 1 throughout the study. Arterial blood samples were withdrawn from the radial artery every 10–15 min and analysed using a Beckman Glucose Analyzer Type II. Myocardial perfusion and glucose metabolism were assessed with NH3 and FDG, respectively. Tracers were injected intravenously over 30 s by means of an infusion pump (model 351, Sage Instruments). Data were acquired on an ECAT Exact HR multi-slice tomograph (CTI/Siemens, Knoxville,

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Attenuation-corrected

99m

Tc-MIBI SPECT and myocardial viability Roelants et al. 817

Tennessee). A 15 min transmission scan using 68Ga was first acquired and was followed by the emission scans of NH3 and FDG. Dynamic images were acquired for 16 min with NH3 (13 frames) and for 52 min with FDG (16 frames).

feature of the reconstructed activity. Attenuation correction and reduction of scattering contribution was further obtained by incorporating in FL the patient attenuation map rescaled to the effective attenuation (FL-AC). No filter was applied.

Two-dimensional echocardiography

Each reconstructed attenuation map was qualitatively reviewed to ascertain the absence of truncation artifact. The absence of patient movement between the emission and the transmission scans was checked by drawing a heart boundary on an axial slice of the middle region of the left ventricle of the emission scan that was translated to the corresponding slice of the attenuation scan. The method accuracy was previously evaluated by comparing images obtained from a phantom (clinical count rate of 99m Tc) reconstructed without and with FL-AC to the corresponding images obtained from the same phantom (clinical count rate FDG) acquired on a PET camera (data not published).

Echocardiograms were obtained using a 2.5–3.5 MHz wide-angle phased-array transducer with 64 or 96 channels. Images from the parasternal long-axis and short-axis and apical four-chamber and two-chamber views were digitized on-line (ImageVue, Nova Microsonics, Rochester, New York, USA) in basal conditions. Radionuclide ventriculography

Red blood cells were labeled in vivo 20 min after pyrophosphate injection with 740 MBq (20 mCi) of free [99mTc]pertechnetate. Images from the ‘best septal’ left anterior oblique projection were obtained at rest with a small-field-of-view camera (Apex 215, Elscint, Haifa, Israel) before and after revascularization. Data were acquired for 250 000 counts per frame at 32 frames per cardiac cycle, with a 5% rate window tolerance. Image processing and data analysis 99m Tc-MIBI SPECT

In order to take into account the difference of gamma ray energy between 153Gd (99 keV) and 99mTc (140 keV), the attenuation map was obtained by rescaling the FBP reconstruction of the 153Gd transmission data to obtain the effective 99mTc coefficient (0.08 cm – 1) in soft tissues (mediastinum). The value of 0.08 cm – 1 gave a flat profile for the reconstruction of an elliptical cylinder filled with a solution of 99mTc (large axis 40 cm, small axis 20 cm). As our goal was to measure the relative ratio of activity along the myocardium wall, this value was preferred to the 0.12 cm – 1 value usually used to obtain an absolute uptake quantification, but which resulted in an overestimation of the activity deeply located in the body. A Shepp–Logan Hanning window filter was used to reduce the noise of the transmission data. After summation of each angular projection data of each head and the eight gated bins, three reconstruction procedures were simultaneously performed, i.e., filtered back-projection (FBP), a simple iterative algorithm (FL) and an iterative algorithm with a measured attenuation coefficients map (FL-AC). FBP was performed on a half sinogram (1801 around the left side of the patient) with a Shepp–Logan Hanning window filter. Non-attenuation-corrected iterative reconstruction (FL) was performed using a multiplicative algorithm applied on the 3601 sinogram [16]. This algorithm is an acceleration of the image space reconstruction algorithm [17,18] and preserves the positive

NH3 PET and FDG PET

After correction for dead time, decay, scatter and nonuniform attenuation, raw data were reconstructed using a Hanning filter with a cut-off frequency of 0.3 resulting in an effective in-plane resolution of 8 mm full width at half maximum. Comparison of PET and SPECT images

The three reconstructed SPECT images (FBP, FL and FLAC) were transferred to the PET processing workstation. SPECT images were first manually aligned to the NH3 PET images by translation in the three dimensions using dedicated alignment software [19]. Then, the NH3 PET images were reoriented and resliced with a program that divides the left ventricle in one apical, one mid-ventricular and one basal short-axis plane. Each short-axis plane was divided into six segments and 16 of the 18 created segments were retained for the final analysis, according to recommendations of the American Society of Echocardiography [20]. The three SPECT and the FDG PET images were then reoriented and resliced with the parameters of the NH3 PET study. The relative uptake of each segment was calculated using operator-interactive software that created circumferential profiles [21]. Within a given myocardial segment, the mean counts were normalized to the maximal activity of the corresponding short axis plane and the tracer uptake was expressed in relative terms as percentage of maximal activity. A segment was considered as viable by FDG PET or by 99mTc-MIBI SPECT if the relative uptake was Z 60%. Two-dimensional echocardiography

Images were analysed qualitatively by one experienced observer, blinded to the scintigraphic, angiographic and clinical data. Regional function was graded as normal, hypokinetic or akinetic. Normal wall motion was defined

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818 Nuclear Medicine Communications 2006, Vol 27 No 10

as Z 5 mm of endocardial excursion and obvious systolic wall thickening, hypokinesia as < 5 mm of endocardial excursion and reduced wall thickening, and akinesia as near absence of endocardial excursion or thickening.

Table 1 Absolute difference (%, mean ± SD) between NH3 and MIBI uptakes in the apical, middle and basal regions of the left ventricle FBP

FL

FL-AC FL vs. FBP

Statistical analysis

Data are expressed as mean ± standard deviation. The statistical analysis was performed on the absolute difference obtained for each segment between the relative uptake of MIBI (of each reconstruction procedure) and that of NH3. Regression analysis of repeated measures with generalized estimating equations was used to take into account the non-independence of the various segments within a given patient [22]. Comparisons in each segment were corrected for regions as co-variate and vice versa. This analysis was performed using the RMGEE program [23]. Patients were used as the unit of analysis for the detection of myocardial viability. For each method we calculated the percentage of concordant segments with FDG for the diagnosis of viable and nonviable segments and derived estimations of sensitivity and specificity for each patient [12]. The sets of estimations were then compared between the three methods by using the Friedman test, followed in the case of heterogeneity by the Wilcoxon signed rank test for two-by-two comparisons. All statistical tests were twotailed. A P value < 0.05 was considered as statistically significant.

Results The basal segments of a single patient could not be analysed for technical reasons. Therefore, the final analysis was performed on 474 segments amongst which 291 (61%) were dysfunctional. One patient did not undergo FDG PET. Comparison between NH3 PET and 99mTc-MIBI SPECT for myocardial perfusion estimation in the apical, middle and basal regions of the left ventricle

Table 1 shows the absolute difference (as a percent, mean ± SD) between the relative uptake of NH3 and the one of MIBI derived from the three SPECT reconstruction algorithms (FBP, FL and FL-AC), in the apical, middle and basal regions of the left ventricle and, according to the regional function. As expected, the underestimation of myocardial perfusion estimation by using FBP as a reconstruction algorithm is more obvious in the basal regions than in the apical regions (18 ± 14% vs. 10 ± 8%). In our population and with a 16-segment model matching the two-dimensional echographic regions, the FL-AC reconstruction algorithm allows an improvement of the myocardial perfusion estimation in the middle and basal regions of the left ventricle whereas the perfusion estimation is not improved when the iterative algorithm FL is used, whatever the region being studied.

Comparisons

Apical All Normal Dysf. Middle All Normal Dysf. Basal All Normal Dysf.

FL-AC vs. FL-AC vs. FL FBP

12 ± 9 10 ± 8 12 ± 10

11 ± 9 8±7 11 ± 9

12 ± 10 10 ± 10 13 ± 10

NS NS NS

NS NS NS

NS NS NS

13 ± 10 12 ± 11 13 ± 10

12 ± 9 12 ± 9 12 ± 9

9±8 8±7 10 ± 8

NS NS NS

< 0.001 < 0.001 0.038

< 0.001 < 0.001 0.003

16 ± 14 14 ± 13 18 ± 14

16 ± 14 15 ± 13 18 ± 15

14 ± 12 13 ± 10 15 ± 14

NS NS NS

< 0.001 NS 0.047

< 0.001 NS 0.047

FBP, filtered back-projection; FL, an iterative algorithm without attenuation correction; FL-AC, an iterative algorithm that incorporates each patient measured attenuation map; dysf. = dysfunctional. Table 2 Absolute difference (%, mean ± SD) between NH3 and MIBI uptakes in the different regions of the left ventricle FBP

FL

FL-AC

Comparisons FL vs. FBP

Inferior Posterior Lateral Anterior Anteroseptal Septal

FL-AC vs. FL-AC vs. FL FBP

17 ± 12 9±8 9±9 11 ± 10 16 ± 12

13 ± 12 10 ± 8 10 ± 9 11 ± 9 15 ± 12

12 ± 10 10 ± 8 10 ± 10 10 ± 9 12 ± 11

< 0.001 NS NS NS NS

NS NS NS NS < 0.001

< 0.001 NS NS NS 0.009

18 ± 14

18 ± 14

15 ± 12

NS

< 0.001

0.003

Abbreviations as in the footnote to Table 1.

Comparison between NH3 PET and 99mTc-MIBI SPECT for myocardial perfusion estimation in the walls of the left ventricle

Table 2 shows the absolute difference (mean ± SD) between the relative uptake of NH3 and the one of the three SPECT reconstruction procedures, according to the different walls of the left ventricle (inferior, posterior, lateral, anterior, anteroseptal and septal). FL-AC significantly improves tracer uptake estimation in the inferior, anteroseptal and septal regions without impairing uptake estimation in the posterior, lateral and anterior regions. Tracer uptake estimation is already greatly improved by using the algorithm FL in the inferior region. The impact of attenuation correction in normal and in dysfunctional regions of the anterior, anteroseptal and inferior regions is shown in Table 3. Compensation for photon attenuation improves uptake estimation in normal as well as in dysfunctional segments of the inferior region. However, the use of an iterative algorithm plays an important role in such improvement. The effect of FLAC is statistically significant in the dysfunctional segments of the anteroseptal region and in the normally contracting segments of the septal region. Looking at the middle and basal regions of the left ventricle (where the effect of FL-AC was significant), an improvement of

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Attenuation-corrected

uptake estimation was observed in the inferior, anteroseptal and septal regions. No statistical difference was found in the other regions (Table 4). Diagnosis of myocardial viability

The overall performance of each reconstruction method for the diagnosis of myocardial viability is shown in Table 5. The use of an iterative algorithm and compensation for photon attenuation significantly improve the sensitivity of 99mTc-MIBI SPECT for the diagnosis of myocardial viability without any statistically significant loss in specificity. Table 3 Absolute difference (%, mean ± SD) between NH3 and MIBI uptakes in the inferior, anteroseptal and septal regions according to the regional function FBP

FL

FL-AC

Comparisons FL vs. FBP

Inferior Normal (n = 22) Dysf. (n = 67) Anteroseptal Normal (n = 24) Dysf. (n = 35) Septal Normal (n = 39) Dysf. (n = 50)

FL-AC vs. FL-AC vs. FL FBP

18 ± 13

14 ± 11

12 ± 9

0.002

NS

0.045

17 ± 11

13 ± 12

12 ± 10

0.002

NS

< 0.001

13 ± 11

14 ± 12

10 ± 10

NS

0.001

NS

18 ± 12

16 ± 12

14 ± 12

NS

NS

0.007

22 ± 14

20 ± 13

14 ± 12

NS

< 0.001

< 0.001

16 ± 13

17 ± 14

16 ± 13

NS

NS

NS

Abbreviations as in the footnote to Table 1.

Table 4 Absolute difference (%, mean ± SD) between NH3 and MIBI uptakes in the different regions of the middle and basal regions of the left ventricle FBP

FL

FL-AC

Comparisons FL vs. FBP

Inferior Posterior Lateral Anterior Anteroseptal Septal

FL-AC vs. FL-AC vs. FL FBP

20 ± 12 9±8 9±8 10 ± 10 16 ± 12

16 ± 12 10 ± 8 10 ± 9 10 ± 9 15 ± 12

13 ± 10 10 ± 8 9±7 9±9 12 ± 11

0.003 NS NS NS NS

0.041 NS NS NS < 0.001

< 0.001 NS NS NS 0.009

22 ± 15

22 ± 15

16 ± 13

NS

< 0.001

< 0.001

Abbreviations as in the footnote to Table 1.

99m

Tc-MIBI SPECT and myocardial viability Roelants et al. 819

Discussion The present study provides a direct comparison of myocardial perfusion evaluations using conventional and attenuation corrected 99mTc-MIBI SPECT to NH3 PET. The most important findings were that, on overweight patients suffering from chronic ischaemic left ventricular dysfunction, compensation for non-uniform photon attenuation with a system based on an external radionuclide source improves tracer uptake estimation in the inferior, antero-septal and septal regions without impairing the performance of this technique in the other regions of the left ventricle. Data reconstruction with the iterative algorithm FL instead of the conventional FBP improves tracer uptake estimation only in the inferior region. This can be explained by the fact that FL preserves the positive feature of the reconstructed activity in a natural way, avoiding the presence of wide artifacts which can occur in FBP reconstruction, especially for large patients where the attenuation dramatically reduces the count rate. In addition to that, it is suggested that the sensitivity of rest 99mTc-MIBI SPECT for the diagnosis of myocardial viability is improved without any statistically significant loss in specificity. Effect of attenuation correction on tracer uptake estimation

The clinical utility of compensation for non-uniform photon attenuation in nuclear cardiology has been reported by different groups, using various tracers and various techniques [5,11,24–30]. However, in those papers, no direct comparison of iterative algorithms with and without attenuation correction has been performed. In accordance with our previous experience [16], our study shows that using the iterative algorithm FL significantly improves myocardial perfusion estimation with 99mTc-MIBI SPECT, but only in the inferior region. In our population of overweight patients suffering from chronic left ventricular ischaemic dysfunction, compensation for non-uniform photon attenuation further improves tracer uptake estimation in the inferior, anteroseptal and septal regions of the left ventricule, without impairing tracer uptake estimation in the other regions of the left ventricle. These results are completely in agreement with the data of the literature. Generally, the apex of attenuation corrected image shows a decrease in activity that is consistent with wall thinning at this site [5,31], which we also experienced in the clinical routine use of attenuation corrected images. In the present study, we

Table 5 Sensitivity and specificity of uncorrected and attenuation corrected MIBI data for the diagnosis of myocardial viability with relative FDG uptake Z 60% as the ‘gold standard’ (medians with interquartile range into brackets) FBP

Sensitivity Specificity

83 (65–96) 63 (26–95)

FL

100 (75–100) 42 (4–100)

FL-AC

100 (82–100) 46 (21–100)

Comparisons FL vs. FBP

FL-AC vs. FL

FL-AC vs. FBP

0.026 NS

NS NS

0.034 NS

Abbreviations as in the footnote to Table 1.

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820 Nuclear Medicine Communications 2006, Vol 27 No 10

did not observe any decreased uptake in the apical regions of the FL-AC images as compared to the non corrected images. A possible explanation probably lies in the model we have used for the data analysis and comparison to two-dimensional echocardiography where the named ‘apical region’ does not strictly correspond to the real extreme anatomic apical region of the left ventricle. Comparisons between myocardial perfusion imaging with NH3 PET and with attenuation corrected 99m Tc-MIBI SPECT have recently been reported by Fricke et al. [32]. These authors concluded that attenuation correction derived from computed tomography leads to SPECT images that represent myocardial perfusion more accurately than non-attenuation-corrected SPECT images, particularly in the inferior wall. Our results, which were obtained with a correction based on an external radionuclide source system, are completely in agreement with their finding. Resting

99m

Tc-MIBI SPECT and myocardial viability

the mortality rate of medically treated patients suffering from chronic ischaemic left ventricular dysfunction [38], it is necessary to precisely quantify the perfusion to accurately estimate the time course of functional recovery and the prognosis of patients suffering from ischaemic dysfunction. Since attenuation correction allows a better estimation of tracer uptake, our data suggest that attenuation correction should be systematically performed when myocardial viability is evaluated with 99m Tc-MIBI SPECT/FDG PET.

References 1 2

3

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99m

Compared with FDG PET, non-corrected Tc-MIBI SPECT has been shown to underestimate the myocardial viability in a substantial proportion of patients [33]. The lack of attenuation correction may partially explain that finding. In our study, we found an improvement in sensitivity without any significant loss in specificity. The low specificity was obtained with a cut-off of 60% to define viability by resting attenuation corrected 99mTcMIBI SPECT, classically used for data reconstructed with FBP. The optimal cut-off for the FL algorithm (allowing a better specificity to be obtained) needs to be determined by using a more adequate gold standard, i.e., functional improvement after revascularization. Nevertheless, our results are in agreement with the ones obtained by Matsunari et al. [12] and Slart et al. [34]. These authors found that attenuation correction of 99mTc-tetrofosmin SPECT studies increases the percentage of concordant segments regarding viability between 99mTc-tetrofosmin SPECT and FDG PET or FDG SPECT, mainly by decreasing the underestimation of viability in the inferior septal region. Clinical implications

Myocardial viability assessment is an important clinical issue for patients with coronary artery disease and severe left ventricular dysfunction. Combination of FDG PET and 99mTc-MIBI SPECT reconstructed without attenuation correction was found to be a valuable method for the diagnostic of myocardial viability [35]. However, as compared to NH3 PET, rest 99mTc-MIBI SPECT tends to overestimate perfusion defects in the septal and inferior regions [36]. Moreover, since dysfunctional segments with reduced perfusion/preserved glucose utilization may take longer to recover function after revascularization than dysfunctional segments with normal perfusion/glucose utilization [37], and, since the extent of perfusion 18F-FDG PET mismatch determines

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Technical note

Comparison of multi-ray and point-spread function based resolution recovery methods in pinhole SPECT reconstruction Antti Sohlberga,b, Hiroshi Watabea, Tsutomu Zeniyaa and Hidehiro Iidaa Background and objectives Statistical reconstruction methods allow resolution recovery in tomographic reconstruction. Even though resolution recovery has the potential to improve overall image quality, pinhole SPECT images are still often reconstructed using simplified models of the acquisition geometry in order to reduce reconstruction time. This paper investigates the benefits of two resolution recovery methods, multi-ray and point-spread function based, in pinhole SPECT by comparing them to uncorrected reconstruction. Methods Resolution recovery was incorporated into ordered subsets expectation maximization reconstruction algorithm. The first of the correction methods used a simple but very fast multiple projection ray approach, whereas the second, much slower, method modelled the acquisition geometry more accurately using the analytical point-spread function of the pinhole collimator. Line source, Jaszczak and contrast phantom studies were performed and used for comparison. Results Resolution recovery improved resolution, contrast and visual quality of the images when compared to

Introduction The use of pinhole single photon emission computed tomography (SPECT) in clinical practice has been limited to small and superficial targets such as the thyroid [1] and joints [2] due to the reduced field of view. Recently, there has been renewed interest in pinhole SPECT, because it enables small animal imaging, where a small field of view is not a serious problem [3–7]. The attractiveness of small animal pinhole SPECT arises from the fact that it can be performed without any dedicated hardware using only a conventional gamma camera, whereas small animal PET, for example, requires an imaging device suitable only for laboratory animals [8]. The quality of SPECT is degraded by three main factors: attenuation, scatter and collimator blurring of which attenuation and scatter are less pronounced in small animal pinhole SPECT [9]. The collimator blurring reduces spatial resolution and forces the use of small diameter pinhole apertures at the cost of severely reduced sensitivity, which is the most important drawback of small animal pinhole SPECT. The sensitivity of pinhole SPECT can be increased by using multi-pinhole

reconstructions without it. The method based on the point-spread function performed slightly better, but was almost 50 times slower than the much simpler multi-ray approach. Conclusion The multiple projection ray approach is a promising method for very fast and easy resolution recovery in pinhole SPECT. It has a profound effect on image quality and can markedly improve the resolution–sensitivity trade-off. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:823–827
Nuclear Medicine Communications 2006, 27:823–827 Keywords: pinhole SPECT, resolution recovery, statistical reconstruction a

National Cardiovascular Center Research Institute, Suita City, Osaka, Japan and Department of Clinical Physiology & Nuclear Medicine, Kuopio University Hospital, Finland.

b

Correspondence to Dr Antti Sohlberg, National Cardiovascular Center Research Institute, 5-7-1 Fujishiro-dai, Suita City, Osaka, Japan 565-8565. Tel: + 0081 6833 5012 (ext. 2559); fax: + 0081 6 6835 5429; e-mail: [email protected] Received 13 February 2006 Accepted 4 May 2006

collimators [10], but this requires modifications to the standard clinical imaging equipment and is not widely applied. One solution to the poor sensitivity problem might be the use of statistical reconstruction methods such as the maximum likelihood expectation maximization (ML-EM) [11] or the ordered subsets expectation maximization (OS-EM) algorithms [12]. ML-EM and OS-EM can partly recover the loss in resolution caused by collimator blurring by incorporating a model of the acquisition geometry into the algorithm and might therefore allow the use of larger diameter pinhole apertures. Recovery of resolution has been shown to improve the quality of conventional SPECT imaging [13,14], but is not yet commonly used in small animal pinhole SPECT. The biggest problem in incorporating resolution recovery in pinhole SPECT reconstruction is the large increase in computational burden. The calculation of point-spread function (PSF) look-up tables can take hours and might require several gigabytes of storage space. The fast resolution recovery methods such as the slice-to-slice blurring [15] often used in parallel-beam SPECT are not

c 2006 Lippincott Williams & Wilkins 0143-3636


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824 Nuclear Medicine Communications 2006, Vol 27 No 10

very straightforward to extend for pinhole SPECT either due to converging nature of pinhole collimation. To overcome computational problems a relatively simple resolution recovery method for pinhole SPECT was recently presented [16]. This method is based on the use of multi-ray projections, where 7 or 21 projection rays, which intersect the pinhole aperture in a predetermined pattern, are used instead of a single ray going through the pinhole centre. The aim of this work is to compare the multi-ray projection approach to a method with a more accurate model of the acquisition geometry.

Materials and methods

was implemented using ray-driven forward projection with seven projection rays and voxel-driven back-projector with a single ray. The seven projection rays intersected the pinhole aperture in a hexagonal pattern modelling the effect of the finite pinhole aperture diameter by inverse cone of rays [16]. The third method (OS-EMpsfRR) incorporated analytical pinhole collimator point-spread functions into the reconstruction algorithm. The PSFs were calculated according to Metzler et al. [17] and stored into hard disk prior to reconstruction in contrast to approach used by the first and second method where the forward projection and back-projection were calculated on-the-fly during the reconstruction.

Implementation of the algorithms

The ML-EM algorithm can be presented as lk ðbÞ X n ðd Þ pðb; d Þ P lkþ1 ðbÞ ¼ P pðb; d Þ d pðb0 ; d Þlk ðb0 Þ d

Phantom studies

ð1Þ

b0

where l(b) is the number of counts emitted from image voxel b, k is the number of iteration, p(b,d) is the probability that the emission in voxel b is detected in detector bin d, and n*(d) is the measured projection count in detector bin d. The ML-EM algorithm updates the current image estimate using forward projection, X pðb0 ; d Þlk ðb0 Þ; b0

and back-projection, X pðb; d Þ P d

n ðd Þ ; pðb0 ; d Þlk ðb0 Þ

b0

operations. In this study three different reconstruction algorithms based on ML-EM and accelerated using the ordered subsets approach were implemented. The first method (OS-EMnoRR) used a simple ray-driven forward projector/voxel-driven back-projector pair assuming zero pinhole diameter. The second algorithm (OS-EMrayRR)

The phantom studies were performed using a Toshiba GCA-7200A (Toshiba, Japan) gamma camera equipped with a 251 mm focal length pinhole collimator (0.5, 1.0 and 2.0 mm pinhole apertures specially fabricated for small animal studies). Three phantoms were imaged. The line source phantom consisted of a line source holder and single line source filled with 5 MBq of 99mTc and placed accurately on the axis of rotation. The resolution was measured as full width at half maximum of horizontal profile taken at the central slice of the phantom. The Jaszczak phantom had four sectors with 1.0, 1.5, 2.0 and 2.5 mm diameter rods separated by a distance twice the rod diameter (Fig. 1). The Jaszczak phantom was filled with 52 MBq of 99mTc and used to assess the image quality visually. The contrast phantom consisted of eight rods (2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0 and 9.0 mm diameter) and a large background compartment (Fig. 1). The rods were filled with an approximately five times higher concentration of 99mTc than the background. The total activity in the phantom was 780 MBq. The contrast for each rod was calculated as C¼

Irod  Ibg Irod þ Ibg

ð2Þ

Fig. 1

2.5 mm

2.0 mm

2.0 mm 3.0 mm

9.0 mm

4.0 mm

8.0 mm

5.0 mm 1.5 mm

7.0 mm

1.0 mm 6.0 mm

One slice of the Jaszczak phantom (on the left) and contrast phantom (on the right). The phantoms had outer diameter and height of 50 mm. The diameters of the rods are marked on the phantoms.

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Comparison of resolution recovery methods in pinhole SPECT Sohlberg et al. 825

Resolution (measured as full width at half maximum) for 0.5, 1.0 and 2.0 mm pinhole aperture diameters obtained from the line source phantom study, which was reconstructed using OS-EM (five iterations) with the three different resolution recovery methods: no resolution recovery (noRR), multi-ray based resolution recovery (rayRR) and point-spread function-based resolution recovery (psfRR)

Table 1

Aperture diameter (mm) Algorithm

0.5

1.0

2.0

noRR rayRR psfRR

1.4 1.3 1.2

1.6 1.4 1.4

2.4 1.3 1.3

where Irod is the average count in the region of interest (ROI) of the rod and Ibg the average count in background ROI. The rod ROIs were circular and had the same diameter as the corresponding rod, whereas each background ROI was annular with inner diameter equal to the diameter of the rod and outer diameter was inner diameter plus 2 mm.

All the acquisitions were performed using 0.5, 1.0 and 2.0 mm pinhole aperture (601 opening angle), 55 mm radius of rotation, 4.3 mm pixel size (128  128 matrix), 3601 circular orbit and 120 projection angles. In order to avoid centre of rotation shifts the projection data was acquired by rotating the target instead of the detector using the system presented by Zeniya et al. [18]. The imaging time per projection angle was selected so that the number of total projection counts was similar in the 0.5, 1.0 and 2.0 mm pinhole aperture studies. The average total projection counts in the line source, Jaszczak and contrast phantom study were 0.9 MCts, 5.2 MCts and 51.8 MCts. The images were reconstructed with the three OS-EM algorithms using eight subsets and five iterations with 0.94 mm voxel size.

Results Table 1 presents the resolution values after five iterations for the three pinhole apertures and three algorithms. The resolution recovery clearly improves resolution and its effect is more pronounced at larger

Fig. 2

Transverse slices of the Jaszczak phantom study (five slices summed into one); from top to bottom 0.5, 1.0 and 2.0 mm pinhole aperture diameter and from left to right OS-EM with no resolution recovery, multi-ray based resolution recovery and point-spread function based resolution recovery. Air bubbles are marked with arrows on one of the images.

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826 Nuclear Medicine Communications 2006, Vol 27 No 10

Contrast for 0.5, 1.0 and 2.0 mm pinhole aperture diameters obtained from the contrast phantom study reconstructed using OS-EM (five iterations) with the three different resolution recovery methods: no resolution recovery (noRR), multi-ray based resolution recovery (rayRR) and point-spread function based resolution recovery (psfRR)

Table 2

Rod diameter (mm)

Aperture diameter and resolution recovery method 0.5 mm

2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0

1.0 mm

2.0 mm

noRR

rayRR

psfRR

noRR

rayRR

psfRR

noRR

rayRR

psfRR

0.34 0.46 0.48 0.48 0.50 0.50 0.49 0.51

0.35 0.48 0.49 0.49 0.51 0.50 0.50 0.52

0.35 0.50 0.51 0.51 0.53 0.52 0.52 0.53

0.29 0.38 0.51 0.49 0.49 0.50 0.48 0.53

0.30 0.39 0.54 0.51 0.51 0.53 0.50 0.55

0.31 0.41 0.55 0.52 0.53 0.54 0.52 0.57

0.13 0.28 0.34 0.37 0.38 0.41 0.39 0.47

0.14 0.32 0.40 0.43 0.43 0.46 0.43 0.51

0.13 0.28 0.38 0.42 0.41 0.44 0.43 0.49

True contrast of the rods versus background according to Equation 2 is 0.67.

pinhole diameters. On the other hand, the results of the multi-ray and the point-spread function based methods do not differ much. The images of the Jaszczak phantom (Fig. 2) confirm the above findings. The resolution recovery has the biggest effect at the 2 mm pinhole diameter case and the quality of the OS-EMrayRR and OSEMpsfRR images is very similar. The results of the contrast phantom experiment after five iterations are illustrated in Table 2. The methods based on the point-spread function performs slightly better, overall, than does the multi-ray based method.

Discussion This study compared fast multiple projection ray and analytical point-spread function based resolution recovery methods to uncorrected reconstruction in pinhole SPECT. The multi-ray and point-spread function approaches improved resolution and contrast. The greatest improvement in resolution and contrast was noticed at large pinhole diameters (Tables 1 and 2, Fig. 2). The poor performance of uncorrected reconstruction at larger pinhole diameters is due to the fact that at larger diameters the zero pinhole diameter assumption made in uncorrected reconstruction is violated more than at smaller aperture diameters. As mentioned in the introduction the biggest problem in incorporating resolution recovery in pinhole SPECT reconstruction is the large computational burden. The calculation time for one iteration of the uncorrected pinhole OS-EM (1.7 GHz Intel processor with 1.0 GB RAM) was 4 min, whereas the computation time for OSEMrayRR and OS-EMpsfRR was 15 min and 10.5 h, respectively, using the acquisition parameters mentioned in the methods section. The reconstruction time of the PSFbased method includes the calculation of the system model, which dominates the calculation time. If the system model for the PSF-based resolution recovery is already stored in a hard disk prior to reconstruction the reconstruction procedure is very fast. Therefore the

point-spread function based resolution recovery is suitable for systems where the imaging geometry is fixed [19], because every time the imaging geometry changes, a new system model needs to be generated. The OSEMnoRR and OS-EMrayRR, on the other hand, model the geometry during the reconstruction and can offer clinically acceptable reconstruction times in every case. The resolution improvement achieved with resolution recovery is important because it allows the use of larger pinhole diameters, which provide higher sensitivity. The poor sensitivity of pinhole collimators has been the biggest problem in small animal pinhole SPECT. Several methods, such as multiple detector heads [3,20] and multiple pinholes [21], have been proposed to overcome this problem, but these approaches require either a multi-headed gamma camera and/or special hardware and are not yet widely applied. Larger injected activities or longer acquisition times are not an optimal approach either, because the radiation burden to the animal or the difficulty of maintaining proper anaesthesia can cause problems. Resolution recovery with a fast resolution recovery algorithm, on the other hand, is almost free of side effects. The only drawback of the multi-ray based approach method is a small increase in noise when compared to reconstruction without resolution recovery, which has been illustrated by Beque et al. [16]. The coefficient of variation (COV = standard deviation/ mean  100%) of a large ROI drawn on the uniform background of the contrast phantom also showed this small noise increase. After five iterations the COVs for the 0.5 mm pinhole aperture diameter were 16.1%, 15.4 and 13.7%, when reconstructed with OS-EMnoRR, OSEMrayRR and OS-EMpsfRR. The respective values for the 1.0 mm aperture were 16.7%, 17.0% and 14.0%, and 14.6%, 15.9% and 8.4% for the 2.0 mm aperture. The noise properties of the resolution recovery methods were not investigated in detail in this study, because they depended on the implementation of the algorithms. For instance, the noise level of the multi-ray based method

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Comparison of resolution recovery methods in pinhole SPECT Sohlberg et al.

could be decreased by implementing resolution recovery also in the back-projector, but unfortunately this happens at the expense of increased computation time. Noise can also be decreased by using Bayesian reconstruction methods [7], which are almost as fast to execute as the common OS-EM. Therefore from the point of view of the computational burden the best alternative for noise reduction when multi-ray resolution recovery is applied might be to use Bayesian reconstruction methods and model collimator blurring only into the forward projector.

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Even though the results presented in this paper were obtained from phantom experiments with activity levels higher than normal can be found in target organs in small animal studies, we are confident that the resolution recovery offers similar improvement in resolution and contrast in small animal studies. However, it is difficult to predict what kind of impact these improvements have, e.g. in brain receptor quantification or measurement of myocardial infarct size in mice and rats. Therefore we plan to further investigate the effect of resolution recovery on (semi-) quantitative values in most frequently used small animal models.

Conclusion The fast multi-ray resolution recovery method performs almost as well as resolution recovery with accurate pointspread functions and therefore shows promise in improving the resolution–sensitivity trade-off in small animal pinhole SPECT.

Acknowledgement This work was supported by Japan Society for the Promotion of Science and Nuclear Diagnostics AB.

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NEWS AND VIEWS OCTOBER 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 2066930, E-mail [email protected] Nuclear Medicine Communications, 2006, 27, 829–830 Why can’t we have statistical parametric mapping (SPM) as an option on all processing workstations?

It’s the same problem we face at the supermarket: eight brands of identical baked beans but no flageolet beans. Most camera manufacturers will offer you three or more varieties of quantitative myocardial SPECT analysis, but no SPM. In fact, the diversity of myocardial software (from LA, Atlanta and Michigan) creates a problem: lack of inter-institutional standardization. For brain perfusion we face the opposite problem: virtually no off-the-peg quantification software. It is difficult to surmise why the camera manufacturers do not offer SPM; it probably has a wider validation in published literature than did the original myocardial software when it was first introduced. The most recent IPEM/BNMS software audit was on quantitative brain SPECT and it is not surprising that the number of returns was disappointing. The software has been downloadable for many years now but the fact that it is not integrated into commercial nuclear medicine systems has obviously been a major disincentive for its widespread use. Assessment of gamma cameras and PET scanners

Gamma cameras used to be assessed in a very thorough and

comprehensive way by the UK Department of Health and later by the Medical Devices Agency. Sadly, these activities ceased a few years ago. The tests were quite different from the National Electrical Manufacturer’s Association (NEMA) tests, since they were performed independently, they were designed to be slightly more clinically orientated, and the authors kept performance charts for each test to which each new camera’s results were added. BNMS and IPEM feel resumption of these tests would be welcome and have proposed this to the NHS Purchasing and Supplies Agency. In our application we have also included PET scanners, both devices to be tested together with CT options. A decision is keenly awaited. NICE guidance for the use of trastuxumab (Herceptin) in HER2 positive breast cancer: a comment on monitoring of left ventricular function

Readers will probably be aware that NICE have recently issued draft guidance on Herceptin, following licensing by the regulatory authorities for use in early breast cancer. The draft guidance recommends the drug for women with early stage HER2-positive breast cancer, except where there are concerns about the woman’s cardiac function. While BNMS agree with the recommendation of the recent

NICE review that cardiac monitoring is essential in the follow-up of patients, some reservations were expressed about the use of a single cut-off value of 55% for all centres and for all techniques. The various bodies, including BNMS, IPEM and BNCS have written to NICE to explain these reservations and also to make suggestions. Meeting Announcements 9th World Congress of Nuclear Medicine and Biology

Dates: 22–27 October 2006 Venue: Seoul, South Korea Website: www.wfnmb.org/ congress2006/index02.htm 2006 IEEE Nuclear Science Symposium and Medical Imaging Conference

Date: 29 October to 4 November 2006 Venue: San Diego, California, USA Website: http://www.nss-mic.org Radiation, Research and Risk

Date: 9 November 2006 Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT, UK Website: www.bir.org.uk International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM)

Dates: 13–15 November 2006 Venue: Vienna, Austria

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Website: www-pub.iaea.org/MTCD/ Meetings/ Announcements.asp?ConfID = 146 Design of Radionuclide Facilities with Reference to Radiation Protection (Revisited)

Date: 14 November 2006 Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT, UK Website: www.bir.org.uk Nuclear Medicine Technologists Meeting

Date: 16 November 2006 Venue: Queen Elizabeth Hospital Postgraduate Centre, Birmingham, UK Website: www.ipem.ac.uk British Nuclear Cardiology Society – Annual Conference

Date: 4 December 2006

Venue: National Heart and Lung Institute, London, UK Website: www.bncs.org.uk British Nuclear Medicine Society – Annual Conference

Dates: 19–21 March 2007 Venue: Manchester, UK Website: www.bnms.org.uk ICNC 2007: 8th International Conference of Nuclear Cardiology

Dates: 29 April to 2 May 2007 Venue: Prague, Czech Republic Website: http://www.icnc8.org

Education and Training EANM Learning Courses

Dates: Weekend courses throughout 2006

Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM Executive Secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected] EANM distance learning in nuclear cardiology

This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. Website: http://www.eanm.org/ eduOnline/edu_start.php?navId = 332

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Erratum 831

Erratum Evaluation of left ventricular ejection fraction by the quantitative algorithms QGS, ECTb, LMC and LVGTF using gated myocardial perfusion SPECT: investigation of relative accuracy Magdy Mohamed Khalila,b, Abdelhamid Elgazzara,b and Wafaa Khalilc Nuclear Medicine Communications 2006, 27:321–332

The publisher would like to apologize for the following error in the Original article above. The legend symbols for the columns in Fig 5 were not shown. The missed figure symbols are shown below.

The author would like to apologize for the following error in the Original article above. Paragraph 4, page 325 ‘‘Both revealed a significant difference among the methods in calculating the ejection fraction (P < 0.0001).’’ To read ‘‘ANOVA analysis revealed a significant difference among the methods in calculating the ejection fraction (P < 0.0001)’’

Fig. 5

Mean EF (%)

70 60

Reference

50

Khalil M. Evaluation of the left ventricular ejection fraction by the quantitative algorithms QGS, ECTb, LMC and LVGTF using gated myocardial perfusion SPECT: Investigation of relative accuracy. Nuclear Medicine Communications 2006; 27:321–332.

40 30 20 10 0 SSS = 4−8

SSS = 9− 13

SSS > 13

Histogram of the mean ejection fraction by using four algorithms – QGS , ECTb , LVGTF , and LMC – versus the summed stress score.

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Editorial

Sentinel lymph node biopsy in differentiated thyroid cancer: Standard of care or experimental tool? Domenico Rubelloa and Michael J. O’Dohertyb Nuclear Medicine Communications 2006, 27:833–835 a Nuclear Medicine Service, ‘S. Maria della Misericordia’ Hospital, Istituto Oncologico Veneto (IOV), Rovigo, Italy and bDepartment of Nuclear Medicine, St. Thomas’ Hospital, London, UK.

The sentinel lymph node (SLN) procedure is considered the standard of care for small breast tumours and melanomas in most of the world [1]. It is based upon the concept that the sentinel node is the first lymph node in the regional nodal basin that drains the primary tumour, thus reflecting the tumour status of the remaining lymph nodes of that basin [2]. In the case of breast cancer patients, it is assumed that all of the breast (both skin and glandular components) drain to the same SLN, usually located in the axilla. This assumption cannot be made for all tumours in the body since lymphatic drainage may be complex and variable. Thyroid tumours is one such tumour with highly variable lymphatic drainage patterns, although levels III, IV and VI lymph nodes are at high risk for harbouring metastases, but drainage can be to other sites. A number of questions are raised with regard to thyroid cancer and nodal disease. Does nodal disease matter, can it be identified using sentinel node techniques and if known about should conventional management be changed? The reported incidence of nodal metastasis in differentiated thyroid cancer (DTC) depends on the size of involved nodes, histological variety, tumour aggressiveness, age of the patient, extent of lymph-node surgery and on the number of sections examined by the pathologist [3]. Macroscopic or clinically apparent nodal involvement varies between 15 and 50% [3], whereas microscopic metastases have been found up to 70–80% of adult patients with DTC [3]. The highest incidence of nodal metastasis is noted in young individuals (up to 80% of children), who on the whole have an excellent prognosis demonstrating that the age at initial diagnosis is one of the most important prognostic factors in the multivariate analysis [3]. The central compartment (level VI) is involved in around 90% of node positive (N + ) patients [4]. There is also a high frequency of lateral node involvement among patients with positive nodes, ranging from 51 to 100% of the reported series [4,5], with the caudal portions of the cervicolateral compartments (level III,IV) involved more frequently than the cranial portions [6]. Supraclavicular node involvement is the third site

Correspondence to: Dr Domenico Rubello, PET Unit, ‘S. Maria della Misericordia’ Hospital, Istituto Oncologico Veneto (IOV), Viale Tre Martiri, 140, 45100, Rovigo, Italy. E-mail: [email protected] Received 18 September 2006 Accepted 18 September 2006

involved, with a reported rate involvement of 10–52% of N + patients [4–6]. Mediastinal nodal involvement is infrequent and in most cases localized to the anterosuperior mediastinum [4–6]. Whilst metastatic spread to distant site(s) is associated with a worse prognosis, the prognostic relevance of metastatic deposits into locoregional lymph nodes is a matter of debate in literature. Most authors consider that nodal involvement has little influence on long-term survival of DTC patients and extensive literature review shows that lymph node metastases at presentation is not associated with a poorer patient survival [3,5,6]. These results are reflected in the commonly utilized thyroid cancer patient risk assessment systems (AMES, AGES, MACIS, etc.) [7]. However, the presence of cervical lymph node metastases has been associated with an increased incidence of locoregional recurrence [8] and the presence of extracapsular invasion of lymph node metastases has been reported to be an indicator of distant metastases and poor prognosis in patients with thyroid papillary carcinoma (PTC) [9]. The surgical management of lymph node metastases at first operation in patients with DTC has been an ongoing source of debate because the benefit of node dissection on recurrence and survival is still controversial [3]. Appropriate application and extension of lymph node dissection is based predominantly on histological type, stage of the primary tumour, and the extent of nodal involvement and often depends more on the attitude of the surgeon than on the above-mentioned clinical characteristics [10]. In cases with gross clinical or ultrasound lymph node involvement, there is no debate about the need and prognostic benefit of a neck dissection in addition to the total thyroidectomy [11]. The management of apparently normal lymph nodes is generally much more conservative. However, some surgeons use prophylactic node dissection in patients with proved PTC in the absence of clinically suspicious

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nodal involvement (cN0) and base this approach on the high rate of occult lymph node metastasis and the favourable clinical results. Noguchi et al. found metastatic nodal involvement on histological examination in 82% of cN0 patients [5], and Hamming et al. [12] reported 70% of cN0 patients, having metastatic nodal disease after prophylactic modified neck dissection. Hamming et al. [12] did find more loco-regional recurrences in the group of patients that did not have prophylactic modified neck dissections (MND). Other groups have reported that compartment-oriented nodal dissection does tend to decrease locoregional recurrence and improve survival [13]. Other surgical strategies in the presence of apparently normal cervical lymph nodes include ‘node picking’, prophylactic routine neck dissection of the ipsilateral central and jugular compartments or prophylactic lymphadenectomy of the ipsilateral central compartments [14]. To date there is no evidence in the literature proving a general benefit of routine prophylactic neck dissection for PTC. With regard to the lack of data, the costs, the inconvenience for the patients and the potential risks of the neck dissection, most surgeons currently favour a conservative policy. En-bloc dissection of regional nodes in patients with DTC either through MND or, more likely, SLN dissection (SLND) of indicated nodal levels represents appropriate treatment in two categories of patients. These include patients with clinically positive preoperative regional nodal disease and patients with metastatic disease demonstrated at thyroidectomy in the central neck compartment (level VI) or the mid/lower jugular nodes (levels III, IV) [10]. An aggressive surgical approach is indicated in patients with medullary thyroid carcinoma (MTC) because of the predilection to metastasize through nodal and extranodal tissues and lack of response to adjuvant therapy exhibited by this tumour [14]. Since the sentinel node identification may influence management in niche areas we should explore how it can be performed. SLND during thyroidectomy can be performed using vital dye technique, lymphoscintigraphy and gamma probe, and combined technique using both vital dye and radiotracer. Ozaki et al. [15] first used thyroid cromolymphography by chlorophyll and Lipiodol UF for diagnosis of cervical lymph node metastases in DTC patients. Kelemen et al. [16] first reported the use of vital dye in a group of 17 patients, this has been performed by others from the same institute [2] and from other institutes [17] with a reported sensitivity ranging from 71 to 100%, a specificity of 100%, and a diagnostic accuracy ranging from 75 to 100%. This technique is associated with the disadvantage that both parathyroid tissue and fat tissue may take up blue dye: recognition of blue dye in a node is not always easy

and finding nodes that are outside the central compartment after a collar incision for thyroidectomy is not always easy. To overcome the above-mentioned drawbacks of the vital dye technique, some authors proposed the use of the combination of lymphoscintigraphy and the intraoperative gamma probe instead of the vital dye technique to localize the SLN [18,19]. It offers significant advantages over the vital dye technique: (1) the injection of the radiopharmaceutical is done preoperatively, therefore eliminating disruption of the lymphatics during the initial dissection; (2) the use of the radiolabelled material permits the disclosure of SLNs that lie outside the central compartment; and (3) there is no ‘false positive’ uptake in the parathyroid glands. The injection technique should be intratumoural rather than peritumoural since the rich vascularity of the thyroid gland is likely to result in distribution of tracer to other parts of the body and not localize flow to lymph nodes [15,19]. After thyroidectomy a hand-held gamma probe is used through the cervical incision to search the central compartment to verify the presence and location of hot spots suggestive of SLNs. After the central compartment has been examined, the lateral compartments of the neck must be bilaterally scanned with the gamma probe through the intact skin. With the lymphoscintigraphy and probe procedure the reported sensitivity in literature ranges from 80 to 100%, with a specificity of 100% and a diagnostic accuracy ranges from 91 to 100% [18–20]. Catarci et al. [20] have performed the only combined study using the combination of preoperative lymphoscintigraphy, intra-operative vital dye and intra-operative hand-gamma probe for SLND in DTC. It is clear that the concept of sentinel node identification can be performed in patients with DTC but should it be performed. Clinical, ultrasonography and intraoperative findings have a low predictive value for metastatic nodal involvement in DTC. Surgical exploration and palpation have been demonstrated to predict nodal spread of disease inaccurately because lymph-node metastases may not be palpable when they are soft or small, and when they are located in the central compartment of the neck or behind the vessels. Occult nodal disease was found in 23 to 69% of patients with apparently negative nodes [21], and 17% of 159 unselected patients with node dissection showed occult disease in the presence of normal macroscopic nodal appearance [22]. Nodal size increase is also an inaccurate marker of metastatic nodal involvement and absence of lymph-node metastases has been reported in 6% of patients with enlarged nodes [2,4,10]. For these reasons SLND in patients with DTC has been proposed to avoid the morbidity of routine and unnecessary node dissection, allowing the identification

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Editorial Rubello and O’Doherty 835

of the first draining lymph node from the tumour and the advantage of identifying early, occult metastasis.

3

4

The advantages that could be achieved through SLND are the following: 1.

2.

3. 4.

Selection of patients who would benefit from compartment oriented nodal dissection, so reducing any unnecessary surgery and possible morbidity; Selection of patients with metastatic disease would permit a more accurate indication to ablative 131I treatment with high doses useful to kill nodal occult micrometastases; Avoidance of 131I ablation in patients with low-risk thyroid tumours and negative SLNs; Identification of lymph node drainage outside the central compartment, allowing a more selective approach and the extension of routine central node dissection in nodes other than the central neck.

In the previously untreated neck, the lymph node drainage of thyroid although variable follows a sufficiently predictable pattern that the concept of selective treatment and SLND has a legitimate rationale. However, unlike breast cancer and melanoma, the role for SLND in the surgical management of patients with DTC has yet to be delineated because of the questionable impact that lymph node metastases have on prognosis. It is also unlikely that the identification of micrometastatic disease will change management since the majority of surgeons only remove clinically involved nodes at surgery.

5 6 7

8 9

10

11 12

13

14 15

16 17

18

It is likely therefore that although SLN biopsy is possible and is accurate it will never achieve a major clinical relevance as a standard of care for all patients with DTC, but might play a role in some sub-groups.

19

20

References 1

2

Gipponi M, Solari N, Di Somma FC, Bertoglio S, Cafiero F. New fields of application of the sentinel lymph node biopsy in the pathologic staging of solid neoplasms: review of literature and surgical perspectives. J Surg Oncol 2004; 85:171–179. Haigh PI, Giuliano AE. Sentinel lymph node dissection for thyroid malignancy. Recent Results Cancer Research 2000; 157:201–205.

21

22

Shaha AR, Shah JP, Loree TR. Patterns of nodal and distant metastasis based on histologic varieties in differentiated carcinoma of the thyroid. Am J Surg 1996; 172:692–694. Mirallie E, Visset J, Sagan C, Hamy A, Le Bodic MF, Paineau J. Localization of cervical node metastasis of papillary thyroid carcinoma. World J Surg 1999; 23:970–974. Noguchi M, Kinami S, Kinoshita K, et al. Risk of bilateral cervical lymph node metastases in papillary thyroid cancer. J Surg Oncol 1993; 52:155–159. Machens A, Heinze R, Thomusch O, Dralle H. Pattern of nodal metastasis for primary and reoperative thyroid cancer. World J Surg 2002; 26:22–25. Rubello D, Pelizzo MR, Al-Nahhas A, Salvatori M, O’Doherty MJ, Giuliano AE, et al. The role of sentinel lymph node biopsy in patients with differentiated thyroid carcinoma. Eur J Surg Oncol 2006: (in press). American Joint Committee on Cancer. Thyroid. In: AJCC cancer staging handbook, 6th edition. Springer: New York; 2002, pp. 89–98. Yamashita H, Noguchi S, Murakami N, Kawamoto H, Watanabe S. Extracapsular invasion of lymph node metastasis is an indicator of distant metastasis and poor prognosis in patients with thyroid papillary carcinoma. Cancer 1997; 80:2268–2272. Robbins KT, Atkinson JLD, Byers RM, Cohen JI, Lavertu P, Pellitteri P. The use and misuse of neck dissection for head and neck cancer. J Am Coll Surg 2001; 193:91–102. British Thyroid Association. Guidelines for the management of thyroid cancer in adults. London: Royal College of Physicians, 2002. Hamming JF, van de Velde CJH, Goslings BM, Fleuren GJ, Hermans J, Delemarre JF, van Slooten EA. Pereoperative diagnosis and treatment of metastases to the regional lymph nodes in papillary carcinoma of the thyroid gland. Surg Gynecol Obstet 1989; 169:107–112. Coburn MC, Wanebo HJ. Prognostic factors and management considerations in patients with cervical metastases of thyroid cancer. Am J Surg 1992; 164:671–675. Robbins KT. Classification of neck dissection. Current concepts and future considerations. Otorhinolaryngl Clin North Am 1998; 31:639–655. Ozaki O, Hirai K, Maruyama S, Sageshima M, Mori T. Experimental and clinical studies on the thyroid chromo-lymphography. Part II: clinical study. Jpn J Surg 1982; 83:53–59. Kelemen PR, Van Herle AJ, Giuliano AE. Sentinel lymphadenectomy in thyroid malignant neoplasms. Arch Surg 1998; 133:288–292. Pelizzo MR, Merante Boschin I, Toniato A, Bernante P, Piotto A, Rinaldo A, Ferlito A. The sentinel node procedure with patent blue V dye in the surgical treatment of papillary thyroid carcinoma. Acta Otolaryngol 2001; 121: 421–424. Rettenbacher L, Sungler P, Gmeiner D, Kassman H, Galvan G. Detecting the sentinel lymph node in patients with differentiated thyroid carcinoma. Eur J Nucl Med 2000; 27:1399–1401. Stoeckli SJ, Pfaltz M, Steinert H, Schmid S. Sentinel lymph node biopsy in thyroid tumors: a pilot study. Eur Arch Otorhinolaryngol 2003; 260: 364–368. Catarci M, Zaraca F, Angeloni R, Mancini B, De Filippo MG, Massa R, et al. Preoperative lymphoscintigraphy and sentinel lymph node biopsy in papillary thyroid cancer. A pilot study. J Surg Oncol 2001; 77:21–24. Wiseman SM, Hicks Jr WL, Chu QD, Rigual NR. Sentinel lymph node biopsy in staging of differentiated thyroid cancer: a critical review. Surg Oncol 2002; 11:137–142. Noguchi S, Yamashita H, Murakami N, Nakayama I, Toda M, Kawamoto H. Small carcinomas of the thyroid: a long-term follow-up of 867 patients. Arch Surg 1996; 131:187–191.

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Original article

Efficacy of milk versus water to reduce interfering infracardiac activity in 99mTc-sestamibi myocardial perfusion scintigraphy Michael Hofman, John McKay and Dee Nandurkar Objective Interference from infra-cardiac radionuclide activity prevents accurate interpretation of true myocardial perfusion. The study aim was to compare the efficacy of milk versus water in reducing infra-cardiac activity in myocardial perfusion scintigraphy. Methods We prospectively randomized 198 patients undergoing stress–rest 99mTc-sestamibi SPECT with exercise or pharmacological stress to drink 300 ml of water or milk prior to imaging. A semi-quantitative grading of the relative intensity of infra-cardiac activity compared to the myocardial activity and a qualitative assessment of the effect on the overall interpretation was performed. Results For stress images, there was no infra-cardiac activity in 37.9%, less intense infra-cardiac activity in 40.8%, equal in 11.7% and greater than infra-cardiac activity in 9.7% with milk, compared with 20.0%, 49.5%, 20.0% and 10.5%, respectively, with water (P = 0.038). For rest images, there was also less intense infra-cardiac activity with milk

Introduction The use of SPECT myocardial perfusion scintigraphy with 99m Tc-methoxyisobutyl isonitrile (sestamibi) and similar compounds is an established diagnostic tool for both the diagnosis and prognostication of patients with ischaemic heart disease. Sestamibi is cleared by the liver and excreted by the hepatobiliary system into the duodenum [1]. Through a variety of mechanisms including attenuation artifact, Compton scatter and tracer washout, the presence of infra-cardiac activity can lead to artifact, reducing the desired target to background ratio. This creates difficulty in both visual and quantitative interpretation of perfusion particularly of the inferior and inferoseptal walls and also hinders image normalization. Infra-cardiac activity is uncommon with exercise but common with pharmacological stress and rest images [2]. Using phantom models of the liver and heart it has been clearly demonstrated [3] that increasing tracer intensity in the liver and decreasing distance between the two organs, results in negative counts being introduced by filtered back-projection, leading to underestimation of

compared to water (P = 0.014). However, no change in subsequent image interpretation was seen. Conclusion Administration of milk resulted in a significant decrease in the intensity of infra-cardiac activity compared to water. However, this did not translate into an improvement in image interpretation. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:837–842 Nuclear Medicine Communications 2006, 27:837–842 Keywords: sestamibi, myocardial perfusion imaging, SPECT, image artifact, image quality Department of Nuclear Medicine, Monash Medical Centre, Clayton, Australia. Correspondence to Dr Dee Nandurkar, Department of Medical Imaging, Monash Medical Centre, Locked Bag 29, Clayton, VIC 3168 Australia. Tel: + 0061 3 9594 2232; fax: + 0061 3 9594 6693; e-mail: [email protected] Part of this paper was presented orally at the 18th Annual Congress of the European Association of Nuclear Medicine in Istanbul, 2005. Received 20 May 2006 Accepted 3 July 2006

actual tracer uptake. The effect is greatest with 1801 reconstruction [4] and can be reduced by using higher frequency cut-offs [5]. Conversely, intense activity near the myocardium can lead to apparent increased counts via the so-called spillover effect. This is not uncommonly seen with duodenogastric reflux in which adjacent gastric activity can be more intense than left ventricular activity [6–8] and is also noted with high activity in the transverse colon [6]. Iterative reconstruction techniques do offer some relief from reconstruction artifacts caused by hepatic and infra-cardiac activity [5]. This technique is often not applied to the gated portion of the study, however. Since the introduction of sestamibi it has been a recommendation to give a fatty meal to reduce interference from liver and gallbladder activity [1]. The recent procedure guidelines adopted by the British Cardiac Society [9], however, state that the value of a fatty meal is uncertain and may be counter-productive if there is gastroduodenal reflux or if the tracer reaches the transverse colon. The aim of this study was to evaluate

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the efficacy of milk administration compared to water to decrease infra-cardiac activity and also to assess any resultant effect on image interpretation of myocardial perfusion.

Methods This randomized control trial was conducted at a tertiary referral hospital. The regional Human Research Ethics Committee approved the study. Study population

The study population consisted of 198 consecutive patients referred to the nuclear medicine department, between September and October 2004, and included 93 females and 105 males with a mean age of 63 years. All of the patients underwent same day stress–rest 99mTcsestamibi gated SPECT imaging. Patients were excluded if they had an allergy/intolerance to, or refused to drink milk, or were unable to drink 600 ml of fluids secondary to medically essential fluid restriction. Patients undergoing a 2-day protocol were also excluded from the study (typically males over 120 kg). Randomization

A computer-generated random-number list was used to allocate patients to either milk or water groups. Stress images were obtained 30 min after 99mTc-sestamibi injection in patients undergoing exercise, and 45 min after 99mTc-sestamibi injection in patients undergoing pharmacological stress. One hundred and fifty millilitres of either chilled water, or chilled full-fat milk (4%) was administered 5 min after completion of exercise or pharmacological stress, and again 5 min before image acquisition. Resting images were obtained 45 min after 99m Tc-sestamibi injection. Patients also received 150 ml of chilled water, or milk, 5 min after the rest injection, and again 5 min before rest image acquisition (total 600 ml of fluids for stress and rest images). Stress testing protocol

Exercise and pharmacological testing were performed as per standardized protocols and were physician-supervised. Patients were fasted for at least 4 h prior to stress testing (usually overnight) and asked to abstain from caffeine for at least 24 h. Beta-blockers, calcium antagonists, nitrates and other drugs were withheld where appropriate. Patients were permitted to eat and drink during the interval between stress imaging and rest 99m Tc-sestamibi injection. Table 1

SPECT protocol 99m

Tc-sestamibi (400 MBq and 1200 MBq) was injected at stress and rest respectively, in an average weight adult, with doses adjusted for weight in heavier individuals. SPECT images of the heart were obtained using a dualhead gamma camera (ADAC Forte, California, USA) fitted with a low-energy, high-resolution collimator. For stress images, 64 frames of 35 s using a 64  64 matrix were acquired, and stress images were gated where possible. A roving zoom of 38 cm was used. In male subjects, a further image with the patient in the prone position was routinely obtained using the same matrix size and 32 frames for 15 s each. In our institute supine images are routinely acquired in both male and female subjects and were thus analysed in this study for both genders. The prone images were acquired to differentiate between diaphragmatic attenuation and inferior ischaemia [9]. As this is a greater concern in males, the prone images were restricted to male patients. Both supine and prone images were acquired and analysed in keeping with the British Nuclear Medicine Society guidelines, which recommends acquisition of both prone and supine images and not prone images in isolation. For rest images, the frame length was 20 s and slices were reconstructed by filtered back-projection using a Butterworth filter and processed using AutoQUANT software. No attenuation correction was used. Image interpretation

Each study was interpreted by consensus agreement between two experienced nuclear medicine physicians. The physicians were blinded to the patients’ randomization and to the patients’ clinical histories. The save screen consisting of the short axis, vertical long axis and horizontal long axis in grey scale was used. A semiquantitative scale similar to that used by Rehm et al. [2] was utilized. Firstly, the relative intensity of any infracardiac activity compared to myocardial activity was graded as 0, no infra-cardiac activity; 1, infra-cardiac activity less than myocardial activity; 2, infra-cardiac activity equals myocardial activity; 3, infra-cardiac activity greater than myocardial activity (Table 1, Fig. 1). Secondly, the effect on interpretation was graded. This was a qualitative assessment of the degree to which infracardiac activity affected interpretation by resulting in either an overestimation or an underestimation of uptake in the myocardium. The grading ‘mild’ was assigned if infra-cardiac was thought to have minimal or no effect on image interpretation. A ‘moderate’ grading was assigned if

Grading of infra-cardiac activity

Relative intensity of infra-cardiac activity compared to myocardial activity

Effect of infra-cardiac activity on the interpretation of inferior cardiac wall imaging

0 1 2 3

None Mild Moderate Severe

Absent Less than myocardium Equal to myocardium Greater than myocardium

No effect on image interpretation Minimal effect on image interpretation Potential compromise of image interpretation Inferior cardiac wall activity not assessable due to infra-cardiac interference

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Reduction of infra-cardiac activity by milk versus water Hofman et al. 839

Fig. 1

(a)

None

(b)

Mild

(c)

Moderate

(d)

Severe

Example of grading of relative intensity of infra-cardiac activity compared to myocardial activity.

infra-cardiac activity potentially compromised interpretation and ‘severe’ if the inferior wall uptake could not be assessed because of interference from the infra-cardiac activity. Statistical analysis

Comparison of milk versus water was completed using a chi-squared statistic. Ordinal logistic regression was used to assess the influence of exercise or pharmacological stress on the results. A value of P < 0.05 was considered statistically significant and P < 0.01 highly significant. To determine the effect of stress imaging in the prone position compared to the supine position, scores of 0–4 were assigned for none, less, equal or greater intensity of infra-cardiac compared to myocardial activity and the difference between the prone and stress images were calculated. A Wilcoxon test of the median difference being zero was used to assess statistical significance.

Table 2

Characteristic

Table 3

One hundred and ninety-eight patients were randomized to receive either milk or water (Table 2). There were no statistically significant differences in the two groups in terms of age, sex or risk factors (P > 0.2 for each) except for previous percutaneous coronary intervention, which was greater in the milk arm. The method of stress was exercise in 47.5%, dipyridamole in 46.0% and dobutamine in 6.5% patients.

Milk (n = 103)

Water (n = 95)

53 50 55 48 64.0 90 72 24 8 7 30 27

52 43 39 56 63.6 83 68 24 8 10 24 21

25

14

24

7

18

19

Male Female Exercise stress Pharmacological stress Age Hypertension Hyperlipidaemia Diabetes mellitus Current smoker Ex-smoker ( < 10 years) Ex-smoker ( > 10 years) Family history of cardiovascular disease Previous acute myocardial infarction Previous percutaneous coronary intervention Previous coronary artery bypass graft

Results Patient characteristics

Patient characteristics

Intensity of infra-cardiac activity versus myocardial activity None

Less

Equal

Greater

Stress Milk Water

37.9% (39) 40.8% (42) 11.7% (12) 9.7% (10) 20.0% (19) 49.5% (47) 20.0% (19) 10.5% (10)

Rest Milk Water

22.3% (23) 57.3% (59) 13.6% (14) 6.8% (7) 9.5% (9) 52.6% (50) 27.4% (26) 10.5% (10)

P value* 0.038

0.014

*

Pearson chi-squared.

Effect on interpretation Intensity of infra-cardiac activity

There was an overall statistically significant reduction in the intensity of infra-cardiac activity with milk compared to water, for both stress and rest images (P = 0.038 and 0.014 respectively) (Table 3, Fig. 2). The frequency of grade 0 and grade 1 intensity of infra-cardiac activity was 37.9% and 40.8%, respectively, for stress images with milk ingestion, compared to 20.0% and 49.5%, respectively, for the water group. However, there was no significant difference in the frequency of grade 3 intensity infracardiac activity. The effect after milk ingestion in the rest imaging group was also most apparent in the frequency of grade 0 intensity.

Reduction in the intensity of infra-cardiac activity in the milk group did not translate into a statistically significant benefit in image interpretation (Table 4, Fig. 3). For stress images, comparison of milk versus water revealed that the effect of milk on the interpretation was graded as none in 69.9%, mild in 23.3%, moderate in 6.8% and severe in 0%, vs. 71.6%, 17.9%, 9.5% and 1.1%, respectively, with water (P = 0.56). For the rest images again no significant difference was noted (P = 0.50). Sub-group analysis

Using ordinal logistic regression, pharmacological or exercise stress was not found to be significant variable

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840 Nuclear Medicine Communications 2006, Vol 27 No 11

Effect of prone imaging

Table 5

Fig. 2

P value

*

0: None

Intensity of infra-cardiac activity

Effect on interpretation

0.004 2 5 70 24 4

< 0.001

1: Less than myocardium 2: Equal to myocardium

–2 –1 0 2 3

3: Greater than myocardium Stress images 37.9%

Milk n=103

11.7% 9.7%

40.8%

5 56 36 8

*

Wilcoxon test of the median difference being 0.

20%

Water n=95

20%

49.5%

10.5%

Rest images 22.3%

Milk n=103

9.5%

Water n=95

13.6%

57.3%

52.6%

0%

20%

40%

Effects of prone imaging

10.5%

27.4%

60%

6.8%

80%

100%

The percentages of studies with absent, less than, equal to or greater infra-cardiac activity compared to myocardial activity, for both milk and water groups, and stress and rest images.

Table 4

Effect of infra-cardiac activity on interpretation None

Mild

Moderate

Severe

Stress Milk Water

69.9 (72) 71.6 (68)

23.3 (24) 17.9 (17)

6.8 (7) 9.5 (9)

0 (0) 1.1 (1)

Rest Milk Water

59.2 (61) 56.8 (54)

31.1 (32) 36.8 (35)

8.7 (9) 5.3 (5)

1 (1) 1.1 (1)

P value* 0.563

0.502

*

Pearson chi-squared.

Fig. 3 None Mild Moderate Severe Stress images 69.9%

Milk n=103

23.3%

71.6%

Water n=95

17.9%

0%

6.8%

9.5%

1.1%

Rest images 59.2%

Milk n=103

56.8%

Water n=95

0%

20%

40%

60%

31.1%

8.7%

36.8%

5.3

80%

Stress imaging in the prone position was performed routinely in male patients (n = 105). Prone imaging resulted in a highly significant increase in infra-cardiac activity (P = 0.004) and a highly significant worsening (P < 0.001) in the effect on interpretation (Table 5). The prone images were obtained after the supine stress image acquisition and often acquired at least one hour after the stress injection. At this time, the clearance of the hepatic activity to the duodeno-jejunal area together with an empty stomach and presence of gastroduodenal reflux may have, at least in part, contributed to the increased activity seen on the prone images.

Discussion Infra-cardiac activity arises predominantly from the liver, hepatobiliary system, bowel or gastro-duodenal reflux and can result in either apparent increase or reduction in radionuclide uptake in the myocardium, particularly in the inferior and inferoseptal walls. Various techniques have been employed to reduce infra-cardiac sestamibi activity, including oral administration of various fluids or solid meals and use of pharmacological agents. The proposed mechanism of action is to fill the stomach, increasing the distance between the left ventricle and interfering infra-cardiac activity, or to increase liver clearance of isotope via gallbladder contraction. Several methods have failed to demonstrate significant benefit, including administration of metoclopramide [10] or carbonated water [11].

1.0%

1.1%

100%

The percentages of studies in which infra-cardiac activity was assessed to have no, mild, moderate or a severe effect on interpretation, for both milk and water groups, and stress and rest images.

(P = 0.75 and 0.90 for stress and rest images respectively) indicating that the effect of milk was not more beneficial for a particular stress modality. Overall, pharmacological stress caused more intense infra-cardiac activity compared with exercise (P = 0.001).

Hurwitz et al. [12] randomized 32 patients to receive a 235 ml milkshake or nothing combined with either standing or supine position. The milkshake reduced gallbladder activity by 40% (P = 0.01) but had no effect on liver parenchymal activity. Of interest, the milkshake reduced infra-cardiac activity on early imaging but not on late imaging, suggesting that the decrease in activity was not due to lipid per se but due to volume in the stomach. There was also no difference between standing or sitting. On the contrary, an effect on hepatic parenchymal activity was noted by Fig et al. [13] who studied 166 consecutive patients randomized to three groups: 250 ml of water alone 45 min post-adenosine compared with a protocol consisting of a fatty meal (15 g) 5 min

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Reduction of infra-cardiac activity by milk versus water Hofman et al. 841

post-adenosine along with water, or a protocol with no intervention. The fatty meal group demonstrated quantitatively decreased gallbladder activity, decreased liver activity and lower left epigastric activity. Van Dongen et al. [14] divided 97 patients into three groups consisting of no action, 150 ml milk 10 min after tetrofosmin administration or 450 ml water 10 min prior to image acquisition, and found that milk significantly decreased the presence of interfering activity. Interfering activity was seen in 33% of patient in the milk group vs. 83% with no action and 74% with water ingestion. A subsequent comparison utilizing attenuation correction found that administration of milk and water resulted in less interfering activity compared to water or milk alone. In a study of 60 patients, Boz et al. [15] investigated the effect of a standardized meal of solid food and liquid (sandwich and 200 ml water) in patients undergoing exercise stress and found a reduction in frequency of intestinal activity by visual assessment from 63% to 10% (P < 0.0001) in the meal group and an increase in the inferior wall to abdomen count ratio of 1.48 to 2.09 (P < 0.0001) compared with no change in the control group. In a recent study of 260 patients, Peace et al. [16] imaged patients at 0.5, 1 or 2 h post-injection and found that infra-cardiac activity significantly decreased with time (P < 0.05). The level of extra-cardiac activity was quantified using an automated extra-cardiac region of interest which was observer validated. The study also examined the effect on a subgroup given 150 ml of full fat milk immediately after injection with a group given 150 ml of milk and 450 ml of water between injection and imaging. There was no statistically significant difference in either group. Iqbal et al. [17] proposed that iodinated oral contrast absorbs gamma rays and would thereby reduce scatter and randomized 30 patients undergoing adenosine stress 99m Tc-sestamibi to receive either one litre of iodinated oral contrast, one litre of water or no intervention. There was a reduction in infra-cardiac counts and image contrast in two of the intervention groups, but no statistically significant difference between the two interventions. In summary, previous studies have provided strong evidence that administration of food or fluid results in reduction in infra-cardiac activity. This occurs by pushing the sub-diaphragmatic activity caudally and increasing the distance between the inferior myocardial wall and liver or bowel. There is disagreement, however, between studies in the efficacy of a fatty meal versus water. The studies discussed are generally limited by their small

numbers, and failure to compare the same volume of fluids in each arm, or failure to randomize. In this study of 198 patients, we found that administration of milk prior to imaging significantly reduces infracardiac activity compared to water alone in patients undergoing either exercise or pharmacological stress. The mechanism of action is thought to be multifactorial. Firstly, administration of a fatty meal delays gastric emptying resulting in increased volume in the stomach [18]. Furthermore, milk stimulates gallbladder contraction [19] resulting in movement of tracer from the liver to the duodenum. The relative contribution of these two mechanisms in reducing infra-cardiac activity is uncertain and it may result from a combination of the two activities. The reduction in infra-cardiac activity by milk compared with water, however, did not result in a significant change in image interpretation. This may be because the reduction in infra-cardiac activity with milk was primarily in the groups with ‘no’ and ‘equal’ infra-cardiac activity compared with myocardial activity, rather than those with infra-cardiac activity greater than myocardial uptake. This implies that despite the presence of infra-cardiac activity in many patients, the target-to-background ratio remained sufficient to allow accurate interpretation. Alternatively, it is possible that the deleterious effects of infracardiac activity were underestimated even by the experienced observers. To accurately assess any resultant error in interpretation, correlation with subsequent angiographic findings or other outcome measurement would be required. Imaging in the prone position which is routinely performed in the male population in our department significantly increased both the intensity of infra-cardiac activity (P = 0.004) and had a deleterious effect on interpretation (P < 0.001). Therefore, in the presence of infra-cardiac sestamibi activity additional prone image may be of limited value because it will accentuate artifact. Study limitations

Our study uses 99mTc-sestamibi and therefore the results are not necessarily applicable to similar isotopes such as 99m Tc-tetrofosmin. 99mTc-tetrofosmin has faster liver clearance but shorter myocardial clearance half-time compared with sestamibi resulting in similar heart-tolung contrasts [20]. Previous studies have not shown any difference in infra-cardiac activity of the two isotopes [2], although in a prospective trial of 686 patients by Ravizzini et al. [21] there was a significantly larger number of patients in the sestamibi group requiring a repeat scan because of liver, or liver and bowel activity – 19.7% vs. 7.9% (P = 0.01).

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842

Nuclear Medicine Communications 2006, Vol 27 No 11

The use of attenuation correction is increasing and our study did not assess the effects of milk in this setting. Attenuation correction and scatter correction are both required to overcome the major sources of apparent count changes in the heart associated with hepatic uptake [22].

Conclusion Infra-cardiac activity causes significant artifacts with filtered back-projection and can lead to error in both visual and quantitative assessment of myocardial perfusion, particularly involving the inferior and inferoseptal walls. Our study demonstrates that administration of milk rather than water results in a significant decrease in intensity of infra-cardiac activity. Nevertheless, no difference was seen in subsequent qualitative effect on interpretation.

8

9

10 11

12

13

14 15

16

References 1

2

3

4 5

6

7

Wackers FJ, Berman DS, Maddahi J, Watson DD, Beller GA, Strauss HW, et al. Technetium-99m hexakis 2-methoxyisobutyl isonitrile: human biodistribution, dosimetry, safety, and preliminary comparison to thallium-201 for myocardial perfusion imaging. J Nucl Med 1989; 30:301–311. Rehm PK, Atkins FB, Ziessman HA, Green SE, Akin EA, Fox LM, et al. Frequency of extra-cardiac activity and its effect on 99Tcm-MIBI cardiac SPET interpretation. Nucl Med Commun 1996; 17:851–856. Nuyts J, Dupont P, Van den Maegdenbergh V, Vleugels S, Suetens P, Mortelmans L. A study of the liver–heart artifact in emission tomography. J Nucl Med 1995; 36:133–139. Hillier D, Wallis J, Miller T. Cardiac artifacts due to hepatobiliary uptake in Tc-99m myocardial SPECT. J Nucl Med 1997; 38(suppl):75. Germano G, Chua T, Kiat H, Areeda JS, Berman DS. A quantitative phantom analysis of artifacts due to hepatic activity in technetium-99m myocardial perfusion SPECT studies. J Nucl Med 1994; 35:356–359. Shih WJ, McFarland KA, Kiefer V, Wierzbinski B. Illustrations of abdominal abnormalities on 99mTc tetrofosmin gated cardiac SPECT. Nucl Med Commun 2005; 26:119–127. Middleton GW, Williams JH. Significant gastric reflux of technetium-99mMIBI in SPECT myocardial imaging. J Nucl Med 1994; 35:619–620.

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Donovan P, Kieffer V, Shih WJ. Duodenogastric reflux on (99m)Tctetrofosmin myocardial SPECT mimics left ventricle inferior wall reverse redistribution and falsely decreases ejection fraction: a case report. J Nucl Med Technol 2001; 29:193–196. Anagnostopoulos C, Harbinson M, Kelion A, Kundley K, Loong CY, Notghi A, et al. Procedure guidelines for radionuclide myocardial perfusion imaging. Nucl Med Commun 2003; 24:1105–1119. Weinmann P, Moretti JL. Metoclopramide has no effect on abdominal activity of sestamibi in myocardial SPET. Nucl Med Commun 1999; 20:623–625. Chambers AA, Mondo CK, Nadarajah J, Myers A, Al-Nahhas A. Does the use of fizzy water improve inferior wall clarity in MPI using 99mTc sestamibi [Abstract]? Nucl Med Commun 2005; 26:75–76. Hurwitz GA, Clark EM, Slomka PJ, Siddiq SK. Investigation of measures to reduce interfering abdominal activity on rest myocardial images with Tc-99m sestamibi. Clin Nucl Med 1993; 18:735–741. Fig L, Chalam G, Steventon R, Deters E, BA D, Gross M, et al. Strategies to minimize abdominal activity on adenosine 99mTc-sestamibi myocardial perfusion imaging [Abstract]. J Nucl Med 2002; 43(suppl):195P. van Dongen AJ, van Rijk PP. Minimizing liver, bowel, and gastric activity in myocardial perfusion SPECT. J Nucl Med 2000; 41:1315–1317. Boz A, Gungor F, Karayalcin B, Yildiz A. The effects of solid food in prevention of intestinal activity in Tc-99m tetrofosmin myocardial perfusion scintigraphy. J Nucl Cardiol 2003; 10:161–167. Peace RA, Lloyd JJ. The effect of imaging time, radiopharmaceutical, full fat milk and water on interfering extra-cardiac activity in myocardial perfusion single photon emission computed tomography. Nucl Med Commun 2005; 26:17–24. Iqbal SM, Khalil ME, Lone BA, Gorski R, Blum S, Heller EN. Simple techniques to reduce bowel activity in cardiac SPECT imaging. Nucl Med Commun 2004; 25:355–359. Boulby P, Moore R, Gowland P, Spiller RC. Fat delays emptying but increases forward and backward antral flow as assessed by flow-sensitive magnetic resonance imaging. Neurogastroenterol Motil 1999; 11:27–36. Fisher RS, Rock E, Malmud LS. Effects of meal composition on gallbladder and gastric emptying in man. Dig Dis Sci 1987; 32:1337–1344. Munch G, Neverve J, Matsunari I, Schroter G, Schwaiger M. Myocardial technetium-99m-tetrofosmin and technetium-99m-sestamibi kinetics in normal subjects and patients with coronary artery disease. J Nucl Med 1997; 38:428–432. Ravizzini GC, Hanson MW, Shaw LK, Wong TZ, Hagge RJ, Pagnanelli RA, et al. Efficiency comparison between 99mTc-tetrofosmin and 99mTc-sestamibi myocardial perfusion studies. Nucl Med Commun 2002; 23:203–208. King MA, Xia W, deVries DJ, Pan TS, Villegas BJ, Dahlberg S, et al. A Monte Carlo investigation of artifacts caused by liver uptake in single-photon emission computed tomography perfusion imaging with technetium 99mlabeled agents. J Nucl Cardiol 1996; 3:18–29.

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Original article

The value of registration correction in the attenuation correction of myocardial SPECT studies using low resolution computed tomography images Christine M. Tonge, Gordon Ellul, Manish Pandit, Richard S. Lawson, Robert A. Shields, Parthiban Arumugam and Mary C. Prescott Background Artifacts caused by tissue attenuation create problems in the interpretation of myocardial perfusion studies. In a previous study we evaluated attenuation correction using ‘Hawkeye’ and noted that the incidence of anterior/apical defects increased after attenuation correction. This increased incidence appeared to be associated with mis-registration between emission and transmission images. The main aim of this study was to determine whether correction of mis-registration between emission and transmission scans reduced the incidence of these anterior/apical defects. Methods Ninety-four patients (64 men, 30 women) underwent stress/rest myocardial perfusion imaging using 99mTc-tetrofosmin (188 studies). Bull’s-eye perfusion plots were created using proprietary software (QPS).

the defect size. After registration correction fewer anterior/ apical defects were created. Conclusion Attenuation correction using ‘Hawkeye’ reduces the incidence of inferior myocardial perfusion defects but can create anterior and/or apical artifacts. It is essential to evaluate registration carefully in three dimensions before reporting the images. Correction of mis-registration reduces the incidence of anterior/apical defects and can restore the appearance of the anterior/ apical area to pre-correction levels. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:843–852 Nuclear Medicine Communications 2006, 27:843–852 Keywords: myocardial perfusion imaging (SPECT), attenuation correction, image fusion, image artifacts Department of Nuclear Medicine, Manchester Royal Infirmary, UK.

Results The marked reduction in defect size, particularly obvious in male patients, in the inferior wall after attenuation correction was not significantly changed by the addition of registration correction. In the anterior and apical walls attenuation correction produced a confusing pattern particularly in females with an overall tendency to increase

Introduction For many years, tissue attenuation has caused problems in the correct interpretation of myocardial perfusion studies [1,2]. Methods of correcting for tissue attenuation using radionuclide transmission sources have been shown to be of variable usefulness in clinical studies [3–7]. We have previously shown that attenuation correction using an X-ray transmission source (Millennium VG gamma camera with Hawkeye, GE Medical Systems) is effective in reducing the incidence of perfusion defects in the inferior wall of the myocardium [8]. The method has also recently been clinically validated for perfectly registered studies against angiographic data in a group of 118 patients [9]. The Hawkeye system uses a low resolution computed tomography (CT) scan carried out at the end of the emission scan to create a map for attenuation correction and it has been demonstrated that systems like this using a high quality attenuation map generate the best results [10].

Correspondence to Christine Tonge, Department of Nuclear Medicine, Manchester Royal Infirmary, Oxford Road, Manchester, M13 9WL, UK. Tel: + 0044 161 276 4820 or 4786; fax: + 0044 161 276 8023; e-mail: [email protected] Received 13 February 2006 Accepted 12 July 2006

However, as emission and transmission imaging are carried out sequentially, rather than simultaneously, it is possible for the patient to move between the two images. It has been shown, by deliberately mis-registering the transmission images, that further artifacts can be introduced if the emission and transmission images are not correctly registered [11]. In our previous work we confirmed that, in clinical studies, defects could be created in the anterior wall of the myocardium following attenuation correction using Hawkeye [8]. These defects were more likely to be created if there was misregistration between the transmission and emission images. Similar results have also been reported by Fricke et al. [12]. The work reported here is a continuation of our previous work and used the same patient group, but is now focussed on further investigating the changes in the anterior/apical segments. Since our previous work, our processing systems have been changed. We now use QPS

c 2006 Lippincott Williams & Wilkins 0143-3636

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844 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 1

Mis-registered study

Same study with registration correction applied Study showing poor registration before and after registration correction.

(Cedars Sinai Medical Centre, Los Angeles, USA) instead of ECToolbox to produce bull’s-eye plots, and Xeleris processing workstations (GE Healthcare Technologies, Milwaukee, USA) instead of Entegra. This change has given us the opportunity to evaluate new commercial software for quality control of Hawkeye attenuation correction. This software (ACQC, GE Healthcare Technologies, Milwaukee, USA) is an integral part of the QC section of the myocardial perfusion processing protocol for Xeleris workstations. It is an interactive rigid-body registration procedure, applied to realign the transmission data to the emission data, by modifying the three shift and three rotation parameters. In our previous study we were able to detect mis-registration by looking at fused transaxial images, but no correction of any detected misregistration was possible. ACQC is designed to make it easier to identify mis-registration by displaying all three orthogonal planes. More importantly, it also includes the ability to correct any mis-registration detected.

Our aim in the current work was therefore to determine if the use of ACQC before reconstruction could reduce the incidence of the apparently artifactual anterior defects that we have previously seen.

Methods Study population

The study group was the same as in our previous work [8]. Data from four patients were excluded from the current analysis – in one case because the patient had moved during the emission image and in the other three cases because of problems during the X-ray acquisition. A further study was excluded because of an artifact on the X-ray image. Hence, data from 179 studies were analysed. 99m

Tc-Myoview SPECT myocardial imaging

Data acquisition was as described in our previous study [8]. Results were transferred to an Xeleris workstation for analysis.

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The value of registration correction Tonge et al. 845

Registration correction

In our previous analysis mis-registration was judged from fused emission and transmission images in the trans-axial plane only and correction of any mis-registration identified was not possible. For the current study the same group of patients was re-examined for mis-registration using the ACQC software. This displayed the CT transmission image, the emission image and the CT image overlaid with a thresholded outline of the left ventricular myocardium. The threshold was adjusted to best represent the myocardial edge. This allowed registration to be easily assessed in three dimensions on one screen. If mis-registration was seen, the ACQC software was then used to interactively shift the emission images in all three dimensions until they visually aligned with the transmission images. Rotational shifts could also be applied, but were not judged necessary for any of the studies in this group. All the studies were examined for any mis-registration between emission and transmission images by fusing the two images using ACQC. An example of the fusion screen for a study judged to be mis-registered is shown in Fig. 1. In this case the transmission image is so badly aligned that myocardial tissue on the emission image overlies lung tissue on the transmission image, although only to a small degree. The thresholded outline of the myocardium from the emission data is superimposed on the black and white transmission data. The lower half of the figure shows the same study after registration correction has been applied. The robustness of the technique was checked by asking three different operators to repeat the re-alignment process. This check used a subset of seven studies showing an obvious anterior artifact after attenuation correction without registration correction. All studies deemed to be mis-registered, even those showing a shift of less than 1 pixel (7.5 mm) were realigned. These re-aligned images were then reconstructed again. The maximum shift required to re-align a study was 23 mm (3 pixels). SPECT analysis

Iterative reconstruction using the ordered subsets expectation maximization algorithm (OSEM) was carried out using 10 subsets and two iterations. A Butterworth post-filter of order 5, cut-off 0.3 cycle/cm was applied to the three-dimensional reconstructed volume as in our previous study. Bull’s-eye images (perfusion polar plots) were produced using QPS software for stress and rest, registration and attenuation corrected (RC), attenuation corrected only (AC) and non-corrected (NC) images. Each bull’s-eye plot was divided into five segments: anterior, inferior, septal, lateral and a central region over

the apex (Fig. 2). These segments were then graded by two experienced observers for the extent of perfusion defects using a subjective scale ranging from 0 for no defect to 6 for an intense defect extending over the whole of a segment (Table 1). Our previous study used a scale ranging from 0 to 5, but during this study we had found a problem in distinguishing the small changes in the anterior wall which are of particular interest in our current study. Therefore an additional grade was added for the current study in order to distinguish between a small defect with a slight reduction in blood flow and a small defect with a moderate reduction. The observers were shown all three sets of images (RC, AC and NC) as a set for each study. The studies were presented in random order and the observers were encouraged to grade such that any perceived difference, however minor, between NC, AC and RC studies was identified in their scoring. The scores of each observer were then averaged to give the overall score for each segment. A Wilcoxon signed ranks test was used to determine the significance of any changes.

Results Table 2 shows the degree of mis-registration as evaluated in three dimensions with ACQC. These results show significantly more mis-registration than in our previous analysis of this patient group. However, the previous method of checking only allowed for evaluation in one plane, whereas ACQC allows for checking in all three dimensions. The ACQC software was felt to be easier to use, particularly in identifying upward shifts, and more confidence was felt in the results. It allowed more accurate detection of small amounts of mis-registration and also significantly increased the number identified as severely mis-registered from 21% in our previous study [8] to 35%. Registration correction was carried out on all studies showing any degree of mis-registration. After registration correction all studies were judged to be satisfactorily aligned and the program was able to correct all the studies in this sample. Although the criteria for acceptable registration have to be subjective, the reproducibility of the technique was checked by using multiple operators for a small subset of seven mis-registered studies in which artifacts were detected. Careful scrutiny of the appearance of the reregistered images revealed no significant differences between operators. Analysis of the results for the group as a whole

Figures 3 and 4 show the results for all patients (179 studies) with registration and attenuation correction applied (RC) and with attenuation correction only (AC). The horizontal axis of the histogram is the change

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846 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 2

Non corrected Atten. corrected

ANT

SEP

LAT

Stress

INF

Rest

Segment definition in a typical study.

Table 1

Grading of defect severity

Table 2

Distribution of studies according to registration grading

Grade

Description

Registration

0 1 2

No defect Small defect with slight reduction in blood flow Small defect with moderate reduction, or moderate defect with small reduction in blood flow Moderate defect with moderate reduction, or small defect with severe reduction in blood flow Moderate defect with severe reduction, or large defect with moderate reduction Large defect with severe reduction Intense defect extending to whole quadrant

Perfect registration Slight (less than 1 pixel) mis-registration Mis-registered by more than 1 pixel, but only within heart tissue Mis-registered so that myocardium appears to overlap chest wall Mis-registered so that myocardium appears to overlap lung

3 4 5 6

in defect score from registration and attenuation corrected to non-corrected (RC – NC) or attenuation corrected to non-corrected (AC – NC). Therefore a negative value indicates a decrease in defect intensity and/or extent and would suggest that attenuation artifacts were being removed.

Number of studies

Percent of studies

27 64

15 36

7

4

19

10

62

35

Figure 3 shows the results for the inferior wall and Fig. 4 for the anterior wall. An alternative display of the same data is shown in Tables 3 and 4. Here the number of patients with the specified defect scores in the inferior

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The value of registration correction Tonge et al. 847

AC using Hawkeye produced a large significant reduction of more than one grade (median change – 1.75 with P < 0.0001) in defect score in the inferior wall. Smaller but still significant reductions (median changes – 0.25 and – 0.5 with P = 0.0002 and P = 0.0001 respectively) in defect score were seen in the septal and lateral walls. There was no significant difference in the appearances of the septal and lateral walls with the additional application of registration correction.

Fig. 3

50 AC - NC

45

RC - NC

Percentage of studies

40

N = 179

35 30

Surprisingly, in the inferior segment, RC caused a small increase in defect score (median change 0.25 with P < 0.0001) as compared with AC.

25 20

In the anterior and apical segments AC produced a confusing picture with both increases and decreases in defect score. RC produced a small but significant reduction (median change – 0.25 with P < 0.0001) in the overall defect score in the anterior and apical segments when compared with AC.

15 10 5 0 −4

−3

−2

−1

0

1

2

3

4

Analysis of the results according to gender

Change in defect size Defect score change with registration and attenuation correction (RC) and attenuation correction only (AC) in the inferior segment.

Fig. 4

The defect score change in the anterior segment analysed according to sex is shown in Figs 5 and 6. For men there was no significant trend in the anterior or apical walls, with as many defects created as removed.

50 AC - NC

45

RC - NC

Percentage of studies

40

As expected, men showed a much higher incidence of inferior defects, but for both genders there was a highly significant reduction in the defect score with AC (median value – 2.0 with P < 0.0001 for men and median value – 0.75 with P < 0.0001 for women). There was no significant difference in the effects of RC as opposed to AC in the inferior wall between the sexes.

N = 179

35

In women, in the anterior segment, the overall tendency was a significant, though small, overall increase (median value 0.25 with P = 0.0174) in defect score, signifying that more defects were being created than removed.

30 25 20

Discussion

15 10 5 0 −4

−3

−2

−1

0

1

2

3

4

Change in defect size Defect score change with registration and attenuation correction (RC) and attenuation correction only (AC) in the anterior segment.

and anterior segments after AC and RC are tabulated. Studies which deviate from the 451 line (shaded grey) represent changes from AC to RC.

The large, significant reduction in defect score over the group as a whole in the inferior wall with AC clearly shows that the technique is successful in removing inferior attenuation artifacts. From a first glance at Fig. 3 it might be thought that RC causes no significant change from AC in the inferior wall. However, after a Wilcoxon signed ranks statistical analysis was performed and from examining Table 3 it was noted that over the whole study group RC caused a small increase in defect score as compared with AC. As mis-registration of the inferior wall would be most likely to result in this area being corrected with either the attenuation coefficient of blood or liver tissue, rather than heart tissue, it would be expected that mis-registration would have little effect on the image and few artifacts would be created in the inferior wall by mis-

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848 Nuclear Medicine Communications 2006, Vol 27 No 11

Table 3

Number of patients showing specified defect scores in the inferior segment before and after registration correction

RC defect score 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 5

AC defect score 0

0.5

35 22 4 1

11 37 18 3

1

1.5

3 11 5 2 1

2 4 1

2

2.5

3

3.5

1

1 1 1 1

1 2

1

2 2

4

4.5

2 2

1

5

1

RC: Registration and attenuation correction; AC: attenuation correction only.

Table 4

Number of patients showing specified defect scores before and after registration correction in the anterior segment

RC defect score 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 5

AC defect score 0

0.5

1

1.5

2

2.5

16 8 2

10 25 9 1 1

12 16 22 5

10 9 5

5 4

1 1 2

1

3

3.5

2 1

1

4

4.5

5

1

1 1 1

2 1 1

1 1

Abbreviations as in the footnote to Table 3.

registration. Previous attempts to create an artifact by deliberate mis-registration were unable to create a defect in the inferior wall [8]. Therefore to see any change with registration correction was unexpected. It is possible that reporting variation could explain variations of one grade, particularly as the observers were attempting to identify all changes, however small. However, it was concerning that further investigation showed that four studies had been graded as showing an increase in defect score of Z 1.5 grade. Although the great majority of studies were unchanged by registration correction in the inferior wall and the changes seen in these four patients were unlikely to affect any clinical report, it is concerning that the reasons for these inferior changes still remain unclear. Mis-registration resulting in heart tissue being corrected by the attenuation coefficient of lung tissue is most likely to create an artifact. This is most likely to occur in the anterior and apical segments. We have previously demonstrated [8] that deliberate mis-registration of a study by as little as 1 pixel (7.5 mm) can create a defect in the anterior or apical wall. As the deliberate misregistration increases, the artifact created becomes more obvious. Therefore it was not unexpected that the changes in the anterior and apical walls were difficult to interpret, with AC appearing to create defects in some patients while removing them in others. As the anterior

wall is the vascular territory most commonly involved in coronary artery disease, it is particularly important to obtain accurate information on the perfusion of this area. Figure 7(a and b) shows two examples from our study group in which a significant anterior defect was created after attenuation correction by mis-registration. Note that in Fig. 7(a) a substantial inferior defect has also been removed by AC. In Fig. 7(b) there is no evidence of an anterior defect before attenuation correction. In Fig. 7(a) there is a minor anterior defect before correction. This is deepened substantially by attenuation correction. In both these patients RC restores the appearance of the area to pre-correction levels. In the group of patients as a whole RC produced a small but significant reduction in the overall defect score in these walls and it can be seen from Fig. 4 that registration correction reduces the percentage of studies with an increase in defect score of greater than one grade from 19% (34 studies) to 7% (13 studies). The results in the apical wall are similar, although the distribution of changes is somewhat more widespread reflecting the greater uncertainty of reporting defects in this area. Attenuation correction is known to change the appearance of the normal heart. Removing attenuation artifacts from one area of the heart may result in the apparent enhancing of small perfusion deficits elsewhere. In

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The value of registration correction Tonge et al. 849

Fig. 5

Fig. 6

50

50 AC - NC RC - NC

45

40

N = 120

35

Percentage of studies

Percentage of studies

40

30 25 20

N = 59

35 30 25 20

15

15

10

10

5

5

0

AC - NC RC - NC

45

0 −4

−3

−2

−1

0

1

2

3

−4

4

−3

−2

Change in defect size

−1

0

1

2

3

Defect score change for men with AC and RC in the anterior segment. (Abbreviations as in the legend to Fig. 4.)

Defect score change for women with AC and RC in the anterior segment. (Abbreviations as in the legend to Fig. 4.)

particular, attenuation correction enhances the appearance of apical thinning and normalizes the gradient between septal and lateral walls [13,14].

K

However, many of the defects seen in the anterior or apical wall after attenuation correction were extensive and severe. In the majority of cases no defect was visible in these areas prior to correction. Also the defects created in the anterior/apical walls did not correspond with other clinical evidence. This suggested that at least some of the defects created in the anterior/apical walls were artifactual, rather than that existing defects were being enhanced by attenuation correction using the Hawkeye system. Although we have no ‘gold standard’ evidence to prove which is the correct appearance, we feel that the factors listed below provide compelling evidence that misregistration between emission and transmission images can produce anterior/apical artifacts: It can be expected that mis-registration is likely to affect the anterior and apical walls rather than the inferior walls because of the sudden change in attenuation from heart to lung tissue. K Similar defects to those seen clinically can be produced by deliberate mis-registration. K

4

Change in defect size

Correction of the mis-registration using ACQC results in a reduction in the number of anterior defects seen.

Gender differences

In men, attenuation is created by the muscular diaphragm leading to inferior wall attenuation artifacts [15,16], while in women, attenuation is more likely to be caused by breast tissue attenuation and to affect the anterior wall [1,15,17]. Therefore we would expect to see a clear difference between the genders in our results. As expected, men showed a much higher incidence of inferior defects, but for both genders there was a highly significant reduction in the defect score with AC. In men, 97% of studies showed a reduction in defect score after both AC and RC. This suggests a very high incidence of attenuation artifacts in men. It might be expected that the incidence of attenuation artifacts would increase with body mass, but although the group as a whole consisted of overweight individuals there were also several of normal weight who showed attenuation artifacts. This has significant implications for practice as it suggests that it is not possible to pre-select patients who would benefit from attenuation correction on the criteria of body weight.

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Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 7

(a)

No correction

Attenuation correction only

Registration and attenuation correction

No correction

Attenuation correction only

Registration and attenuation correction

No correction

Attenuation correction only

Registration and attenuation correction

(b)

(c)

Examples of the effects of attenuation correction and registration correction. (a) and (b) Studies in which attenuation correction creates an anterior defect which is removed by the application of registration correction. (c) An example of the application of registration correction causing a change in the appearance of the inferior wall.

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The value of registration correction Tonge et al. 851

For both men and women there was a confusing pattern in the anterior wall with AC reducing defects in some patients and creating defects in others. In women the overall tendency was a significant, though small, increase in defect score, signifying that more defects were being created than removed. This fact implies that attenuation correction using Hawkeye is not a useful technique in women unless registration correction is applied. Attenuation correction with registration correction reduced the number of studies in which anterior defects increased in size in both sexes. After RC there were no studies with an increase of greater than two grades in the anterior wall. The changes with RC were more obvious in women, with 29% of studies showing a decrease of greater than 0.5 grade in the anterior wall with RC as opposed to AC alone. In men the corresponding figure was 16%. These results demonstrate that correct registration (and therefore the need for a registration correction program) is more important in women than men because of the increased likelihood of an anterior attenuation artifact due to breast tissue in women. The application of a registration correction program is likely to increase the diagnostic accuracy of attenuation correction using Hawkeye in women. Registration effects

Because artifacts can be caused by even a small amount of mis-registration, an accurate method of assessing misregistration is essential. The use of the ACQC program allowing simultaneous display of all three dimensions was found to allow more accurate assessment of mis-registration than the previous single-plane method and the registration gradings changed significantly. Only 15% of the studies were assessed as being perfectly aligned using ACQC although a further 36% were only slightly misregistered. This study group was imaged soon after the installation of the Hawkeye camera. There was a strong learning curve in our implementation of attenuation correction using Hawkeye, not only in the reporting of the data and the re-learning of normal appearances, but also in the optimization of technique. Changes in acquisition technique applied subsequent to this study have considerably reduced the incidence and severity of mis-registered studies. The registration correction module of ACQC was very successful, with all studies in this group being judged visually as having acceptable registration after correction. Reproducibility between operators was good even in those more difficult studies in which an obvious artifact was produced after attenuation correction alone. An increasing incidence of anterior/apical defects was observed as the degree of mis-registration increased. Only one study out of 27 (3.7%) judged to be perfectly

registered without correction showed an increase in defect score of greater than or equal to 1.5 grades in the anterior or apical walls. On reviewing the fusion screen display of this study there was in fact a very small degree of movement of heart tissue into the chest wall so possibly it had been wrongly classified. In contrast such an increase was observed in 27% of those studies where heart tissue had moved into lung. Thus artifacts become more common as the degree of mis-registration increases, but severe mis-registration does not necessarily produce an artifact. In some severely mis-registered studies, where the creation of an artifact would appear likely, no obvious changes were observed. The group of studies observed to be only slightly misregistered was of particular interest as a deliberate misregistration of only 1 pixel was observed to create an obvious defect. Our results confirmed that even these small mis-registrations could also create artifacts in the clinical situation. An increased rate (8%) of anterior/apical artifacts was observed.

Conclusions We have demonstrated that attenuation correction with Hawkeye reduces the incidence of inferior myocardial defects, but can create anterior and/or apical artifacts. Attenuation correction without evaluation and correction of registration errors is likely to give erroneous results, particularly in the anterior/apical regions. Therefore it is essential to evaluate carefully the registration in three dimensions before reporting images. The ACQC software achieves this. Correction of mis-registration using ACQC reduces the incidence of anterior/apical defects and can restore the appearance of the anterior/apical area to pre-correction levels. With the application of registration correction, attenuation correction using Hawkeye is a useful technique in both men and women.

References 1 2 3

4

5

6

7

DePuey EG, Garcia EV. Optimal specificity of thallium-201 SPECT through recognition of imaging artifacts. J Nucl Med 1989; 30:441–449. DePuey EG. How to detect and avoid myocardial perfusion SPECT artifacts. J Nucl Med 1994; 35:699–702. Ficaro EP, Fessler JA, Shreve PD, Kritzman JN, Rose PA, Corbett JR. Simultaneous transmission/emission myocardial perfusion tomography. Diagnostic accuracy of attenuation-corrected 99mTc sestamibi single-photon emission computed tomography. Circulation 1996; 93:463–473. Kjaer A, Cortsen A, Rahbek B, Hasseldam H, Hesse B. Attenuation and scatter correction in myocardial SPET: improved diagnostic accuracy in patients with suspected coronary artery disease. Eur J Nucl Med 2002; 29:1438–1442. Shotwell M, Singh BM, Fortman C, Bauman Bd, Lukes J, Gerson MC. Improved coronary disease detection with quantitative attenuation-corrected Tl-201 images. J Nucl Cardiol 2002; 9:52–61. Vidal R, Irene B, Darcourt J, Mignico O, Desvignes P, Baudouy M, et al. Impact of attenuation correction by simultaneous emission/transmission tomography on visual assessment of 201Tl myocardial perfusion images. J Nucl Med 1999; 40:1301–1309. Slart RHJA, Tjin HQ, Van Veldhuisen DJ, Poot L, Blanksma PK, Piers DA, et al. Effect of attenuation correction on the interpretation of 99mTc-sestamibi

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myocardial perfusion scintigraphy: the impact of 1 year’s experience. Eur J Nucl Med 2003; 30:1505–1509. 8 Tonge CM, Manoharan M, Lawson RS, Shields RA, Prescott MC. Attenuation correction of myocardial SPECT studies using low resolution computed tomography images. Nucl Med Commun 2005; 26:231–237. 9 Masood Y, Liu Y, DePuey G, Taillefer R, Araujo L, Allen S, et al. Clinical validation of SPECT attenuation correction using x-ray computed tomography-derived attenuation maps: Multicenter clinical trial with angiographic correlation. J Nucl Cardiol 2005; 12:676–686. 10 O’Connor MK, Kemp B, Anstett F, Christian P, Ficaro EP, Frey E, et al. A multicenter evaluation of commercial attenuation compensation techniques in cardiac SPECT using phantom models. J Nucl Cardiol 2002; 9:361–376. 11 Takahashi Y, Murase K, Higashino H, Mochizuki T, Motomura N. Attenuation correction of myocardial SPECT images with X-ray CT: Effects of registration errors between X-ray CT and SPECT. Ann Nucl Med 2002; 16:431–435. 12 Fricke H, Fricke E, Weise R, Kammeier A, Lindner O, Burchert W. A method to remove artefacts in attenuation-corrected myocardial perfusion SPECT

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introduced by misalignment between emission scan and CT-derived attenuation maps. J Nucl Med 2004; 45:1619–1625. Links JM, Becker LC, Anstett F. Clinical significance of apical thinning after attenuation correction. J Nucl Cardiol 2004; 11:26–31. Prvulovich EM, Lonn AHR, Bomanji JB, Jarritt PH, Ell PJ. Effect of attenuation correction on myocardial thallium-201 distribution in patients with a low likelihood of coronary artery disease. Eur J Nucl Med 1997; 24:266–275. Tsui BMW, Zhao XD, Gregoriou GK, Lalushl DS, Frey EC, Johnston RE, et al. Quantitative cardiac SPECT reconstruction with reduced image degradation due to patient anatomy. IEEE Trans Nucl Sci 1994; 41:2838–2848. Eisner RL, Tamas MJ, Cloninger K, Shonkoff D, Oates JA, Gober AM, et al. Normal SPECT thallium-201 bull’s-eye display: Gender differences. J Nucl Med 1988; 29:1901–1909. Manglos SH, Thomas FD, Gagne GM, Hellwig BJ. Phantom study of breast tissue attenuation in myocardial imaging. J Nucl Med 1993; 34:992–996.

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Original article

X-ray-based attenuation correction of myocardial perfusion scans: practical feasibility and diagnostic impact Thomas Gru¨ninga,b, Claudia Brogsitterc, Mehrdad Khonsaria, Ivor W. Jonesa, Paul L. Ormsbya and Wolfgang Burchertd Objectives This study describes the practical implementation of X-ray-based attenuation correction (AC) of myocardial perfusion scans in a large teaching hospital, characterizes the impact of AC on the diagnostic confidence of the interpreter and tries to predict which patients are likely to benefit from the technique. Methods One hundred and seven consecutive patients underwent a 2 day 99mTc-tetrofosmin protocol with adenosine stress using GE Millennium VG with AC and ECG-gated acquisition (ECG-g). The diagnostic impact of AC/ECG-g was judged by a panel of three observers. Results AC was not achieved in 46 patients. Individual observers rated AC ‘essential’ in 37 scans and ‘helpful’ in 68 scans. For ECG-g, this applied to 12 and 78 scans, respectively. The rating for AC was better than that for ECG-g in 57 scans, and vice versa in 31 scans. Equal ratings were recorded in 41 scans, and neither technique was needed in 54 scans. Diagnostic interpretation of abnormal scans was significantly more likely to benefit from either AC or ECG-g than interpretation of normal scans. Patients in whom AC was considered useful had a

Introduction Myocardial perfusion scintigraphy is a widely available, non-invasive diagnostic method for assessing patients with ischaemic heart disease [1–3]. It is used to rule out or confirm haemodynamically significant ischaemic heart disease, characterize disease extent and severity, predict patients’ prognosis and assess response to treatment. 201 Tl or 99mTc tracers sestamibi and tetrofosmin can be used, and stress can be applied either mechanically or pharmacologically with a number of agents. Scans produced with any of these protocols can have a number of well-recognized artefacts, including those related to biliary activity, extra-myocardial activity close to the inferior left ventricular wall, contraction anomalies in the septum in patients with left bundle branch block and attenuation artefacts in the anterolateral wall (caused by the left breast) and inferior wall (caused by the left hemidiaphragm) of the left ventricle. Attenuation correction (AC) is a technique developed to overcome the latter artefact, and is now commercially

significantly higher body mass and chest circumference, but the overlap was large. Conclusions In practice, AC was not feasible in a significant proportion of our patients. AC received better ratings from observers more often than ECG-g. Interpreter confidence with AC was significantly greater in scans with perfusion defects than in normal scans. Body mass and chest circumference cannot be used to predict which patients will benefit from AC. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:853–858 Nuclear Medicine Communications 2006, 27:853–858 Keywords: myocardial perfusion scan, rection, X-rays

99m

Tc-tetrofosmin, attenuation cor-

a Department of Nuclear Medicine, Derriford Hospital, Plymouth, UK, bPeninsula Medical School, Universities of Exeter and Plymouth, UK, cDepartment of Nuclear Medicine, University of Dresden, Germany and dInstitute of Molecular Biophysics, Radiopharmacy and Nuclear Medicine, Heart and Diabetes Center, Bad Oeynhausen, Germany.

Correspondence to Dr Thomas Gru¨ning, Department of Nuclear Medicine, Derriford Hospital, Plymouth PL6 8DH, UK. Tel: + 0044 1752 792280; fax: + 0044 1752 517587; e-mail: [email protected] Received 6 June 2006 Accepted 10 July 2006

available from a number of manufacturers [4]. All these implementations make use of an external radiation source, using either long-lived radioisotope sources in varying configurations or X-rays, to acquire a transmission scan. From this, an attenuation map is produced that can be used for correction, depending on tissue density and depth. Using this technique requires resources over and above those for a non-AC myocardial perfusion scintigraphy, either to pay for replacing radioactive sources or X-ray tubes, or extra time for acquiring the transmission scan, depending on the particular implementation. There is now a good body of evidence documenting the extra diagnostic information provided in return [5–10], but whether there is a good balance between the former and the latter remains a subject of debate [11–13]. This study describes the practical feasibility and impact on the diagnostic confidence of the interpreter of X-raybased AC of myocardial perfusion scans using a 99mTc tracer in a typical clinical setting, and investigates whether a patient’s weight or chest circumference can

c 2006 Lippincott Williams & Wilkins 0143-3636

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be used to predict which patients are likely to benefit from the technique.

Methods

a high-quality attenuation map [4]. Regular checks using an appropriate phantom were carried out on the Millennium VG camera to ensure good alignment between the emission and transmission scans.

Patients

One hundred and seven patients (63 men, 44 women, mean age 65 ± 11 years) routinely referred for a myocardial perfusion scan were consecutively included in this study. Pharmacological stress was applied using a 6 min adenosine infusion (AdenoScan; Sanofi Winthrop, Guildford, UK) at 140 mg
kg – 1
min – 1 with injection of 600 MBq 99mTc-tetrofosmin (Myoview; Amersham, Little Chalfont, UK) after 4 min. Seven patients with contraindications to adenosine were excluded. The rest scan was performed on a separate day using 400 MBq 99mTctetrofosmin. A rest scan was only performed if the stress scan was judged to be visually abnormal. Patients were weighed and their chest circumference was measured from a single transaxial slice of the X-ray attenuation map at the level of the left ventricle.

ECG gating

R-wave triggered ECG-g with 8 bins per cardiac cycle was used on both cameras. Rejected beats are automatically discarded during acquisition with the Millennium VG camera. On the Infinia camera, rejected beats are stored separately and contribute to the summed (non-gated) dataset to improve count statistics. Acquisition

Identical parameters were used on both cameras: LEHR collimators, detectors at 901, 1801 rotation at 31 steps with automatic body contouring, 20 s (no ECG-g)/30 s (ECG-g) acquisition per step, 64 64 matrix, zoom 1.28 , pixel size 6.9 mm. Processing

Cameras and acquisition protocol

Scans were acquired 60 min after tracer injection using two double-headed gamma cameras, Millennium VG and Infinia (both General Electric Medical Systems, Slough, UK). With low-energy high-resolution (LEHR) collimators and 99mTc, comparable system sensitivities (Millennium VG 76 s – 1
MBq – 1 at 100 mm, Infinia 72 s – 1
MBq – 1 at 100 mm) and identical resolutions (7.4 mm full width half maximum at 100 mm) are stated by the manufacturer. Both cameras are equipped for ECG gating and the Millennium VG has Hawkeye AC. An ‘optimal’ imaging protocol was required to include both AC and ECG-g for the stress scan, and AC for the rest scan. This means that the ‘ideal’ patient would have both scans on the Millennium VG camera. Due to limited capacity on this camera, any repeat acquisitions needed were performed on the Infinia camera without AC. Patients in whom AC was not feasible were scanned on the Infinia camera as well and the reason for this was documented by the technologist. Attenuation correction

After completion of the emission scan, the imaging table is transferred further into the gantry and the transmission scan commences. The Hawkeye AC system is mounted on the slip-ring gantry such that it rotates together with the detector heads at a speed of 2.6 rotations per min. It consists of an X-ray tube (tungsten target, 0.5 mm copper beam filter, fixed 140 keVp/2.5 mA) directing a collimated fan beam across the patient aperture at a rotating CT detector of 384 elements. CT slices are acquired along a 2161 arc, followed by a 1 cm table translation giving a fixed slice thickness of 1 cm which covers the limited field of view needed for myocardial perfusion scans in about 5 min. The resulting images contain motion artefacts in the abdomen, but are sufficient to produce

Non-AC acquisition data were transferred to a Hermes workstation (Nuclear Diagnostics, Stockholm, Sweden) for processing. Filtered back-projection with a Wiener filter (order 20) was used for reconstruction, and reorientation was performed manually, with the angles recorded in a database. Final processing steps and display of the results were automatic using the QGS software for ECG-g and QPS software for non-ECG-g and summed ECG-g studies (both Cedars-Sinai Medical Center, Los Angeles, USA; supplied by Nuclear Diagnostics). AC acquisition data were processed on a Xeleris workstation (GE Medical Systems). The built-in ordered subset expectation maximum algorithm was used for reconstruction, and reorientation was performed using the same angles that were used on the Hermes workstation. Scatter correction was not available. A Metz filter with a power factor of 5 was applied to the transaxial slices. The main risk with sequential emission–transmission scanning is misalignment, which even if fairly minor can cause artificial perfusion defects. A visual check for misalignment of emission scan and attenuation map was performed if a perfusion defect that was not present in the non-AC scan was seen in the AC study. AC was applied to summed (non-gated) data. Clinical interpretation

Screenshots of the QPS splash and results pages (containing all available slices and stress/rest polar plots) and QGS results pages (containing left ventricular ejection fraction and polar plots of end systolic perfusion, wall motion and thickening) were reviewed independently by three observers with prior experience of the Millennium VG/Hawkeye system (C.B., M.K., T.G.) who were given each patient’s gender, but were blinded to further clinical information and to the interpretations of the other observers. AC and non-AC studies were

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X-ray-based attenuation correction of myocardial perfusion scans Gru¨ning et al. 855

reviewed side by side. Comparison with the normal template built into QPS was not performed as this is based on non-AC studies only. For each patient, observers were required to give a brief description of type (ischaemia, scar, both, none) and location of perfusion defects and then asked to judge whether AC and ECG-g were ‘essential’, ‘helpful’ or ‘diagnostically irrelevant’. Individual observers’ judgments were combined as follows. On a patient-by-patient basis, AC and ECG-g were considered overall ‘very useful’ if any two observers rated a technique as ‘essential’, regardless of the opinion of the third observer. An overall ‘useful’ rating required a rating of at least ‘helpful’ from all observers, and ‘of some use’ meant at least two ‘helpful’ ratings. The remainder of the scans were termed ‘of no benefit’. Observers were also asked to note in which patients they required AC for the rest scan. Quantitative analysis

Normal stress scans (as determined by the three observers, see below) acquired with and without AC were analysed with QPS. This uses a 20-segment model to quantify myocardial perfusion on an arbitrary scale of 0–100. We analysed two apical segments and two segments each for the anterior and posterior wall, disregarding the most basal segments to avoid errors related to contour finding. The built-in normal database was not used as it is only applicable to non-AC scans. Statistical tests

A one-way ANOVA with post-test for linear trend was used for analysing the influence of body mass and chest circumference. A chi-squared test was employed to look at differences in observers’ ratings. Perfusion obtained with QPS was analysed with paired t-tests. All calculations were carried out using Prism 3.03 (Graphpad Software, San Diego, USA).

Results Practical feasibility

AC could not be achieved in 46 patients (43%). In 25 patients (23%), this was due to problems not specific to our imaging protocol: patients unable to position arms in the narrower X-ray gantry (n = 13), claustrophobia (n = 4), patients exceeding the camera’s weight limit (n = 1), camera downtime for both routine maintenance and breakdown (n = 7). In the remaining 21 patients (20%), the limitations of our imaging protocol precluded the use of AC to facilitate patient throughput: repeat acquisition (n = 6), patients with cardiac arrhythmias who were scanned on the Infinia camera (n = 15). This left 41 patients with both a stress and rest scan, and 20 patients in whom no rest scan was needed following a normal stress scan. Clinical interpretation

Thirty-nine patients had normal findings. There was concordance for this between all observers in 36 patients.

In the remaining three patients, two observers agreed that these scans were normal. The normal group comprised all 20 patients who had no rest scan as well as 19 patients with a stress and rest scan. Twenty-two patients had perfusion defects, either reversible or irreversible. There was concordance for this between all observers in 19 patients. In the remaining three patients (one with a scar and two with ischaemia in the posterior wall), one observer rated their scans as normal. Individual observers rated AC ‘essential’ in 37 scans (20%) and ‘helpful’ in a further 68 scans (37%). For ECGg, these figures were 12 scans (6.6%) and 78 scans (43%), respectively. The rating for AC was better than that for ECG-g in 57 scans (31%), and vice versa in 31 scans (17%). Both techniques were found to be ‘diagnostically irrelevant’ in 54 scans (30%), and equal ratings were recorded in 41 scans (22%). Individual observers’ ratings were then combined as described earlier to achieve an overall judgment for each patient (Table 1). AC was found to provide a diagnostic benefit (rated ‘very useful’, ‘useful’ or ‘of some use’) in 22 patients (56%) with normal scans and in 16 patients (73%) with scans that contained perfusion defects. For ECG-g, these figures were 10 patients (26%) and 15 patients (68%), respectively. Diagnostic interpretation of abnormal scans was significantly more likely to benefit from either AC (P < 0.05) or ECG-g (P < 0.01) than interpretation of normal scans. However, the number of patients who derived a benefit was not significantly different between AC and ECG-g. Observers required AC for the rest scan in 13 scans, but there was only one patient in whom all observers agreed that rest AC was needed and a further patient where two observers felt that this was necessary. Patients with different observer ratings for AC showed significant differences in mean body mass (P = 0.0013) and chest circumference (P = 0.0498) with a linear trend, although the overlap was large (Fig. 1). No such differences were found for ECG-g ratings (data not shown). There was also no difference in body weight between those patients in whom AC was performed and those in whom it could not be performed. Quantitative myocardial perfusion of normal scans was analysed with QPS (scale 0–100) and showed mean values of 72 ± 7.1 (non-AC) and 70 ± 6.9 (AC) in the anterior wall, 65 ± 8.3 (non-AC) and 70 ± 4.8 (AC) in the inferior wall, 69 ± 8.2 (non-AC) and 63 ± 6.4 (AC) in the apex (Fig. 2). Applying AC to normal scans led to a significantly higher myocardial perfusion in the inferior

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Nuclear Medicine Communications 2006, Vol 27 No 11

Table 1 Observers’ combined judgement on the usefulness of attenuation correction and ECG gating

3 14 17 1 3 6 30

8 6 6 0 5 10 7

Results are given as the number of patients.

60

40

(a)

150

Body mass (kg)

70

50

Fig. 1

125

Quantitative myocardial perfusion of normal scans measured with QPS (scale 0–100) for anterior, posterior wall and apex in scans with and without attenuation correction.

100

75

technique designed to overcome artificial perfusion defects in the inferior wall of the left ventricle, related to attenuation caused by the left hemidiaphragm.

50 (very) useful

of some use Attenuation correction

not useful

of some use Attenuation correction

not useful

(b) 125

Chest circumference (cm)

80

Apex AC

ECG gating

2

Apex non-AC

Useful Of some use No benefit Very useful Useful Of some use No benefit

5

90

Post AC

Very useful

Scans with perfusion defects

Post non-AC

Attenuation correction

Normal scans

Ant AC

Observers’ judgement

Ant non-AC

Technique

Fig. 2

Perfusion

856

100

75 (very) useful

Patient body mass (a) and chest circumference (b), depending on observers’ combined judgment on the usefulness of attenuation correction.

wall (P = 0.0009), but resulted in a significantly lower perfusion in the anterior wall (P = 0.042) and apex (P < 0.0001).

Discussion Myocardial perfusion scans are an established tool for assessing patients with ischaemic heart disease. AC is a

AC is far from standardized and technical implementation varies with different gamma camera manufacturers. A comparative review of eight different systems found marked differences in performance, even though only looking at phantoms [4]. All of these systems, however, fit into two categories, depending on the nature of the transmission source used. The majority of solutions use radioactive sources with long-lived isotopes (e.g., 133Ba, 153 Gd, 241Am) in varying configurations (e.g., scanning point or line sources, array of line sources) [11]. Alternatively, an X-ray transmission source in the form of a low-resolution, non-diagnostic CT-like apparatus (Hawkeye) has been available from GE Medical Systems on a number of their recent gamma cameras. This technique may be used more widely with the introduction of gamma cameras fused to a full diagnostic CT scanner (SPECT/CT) by other manufacturers. It has long been recognized that AC is capable of introducing new artificial perfusion defects, mainly in the anterior wall, by over-correction. Our study confirmed that this also applies to the GE Hawkeye system: normal scans with AC had significantly lower perfusion in the anterior wall than non-AC scans. We also identified a significant decrease in perfusion of the apex with AC, consistent with artificial perfusion defects. This phenomenon has been extensively studied by Fricke et al. [14] using both patients and phantoms on the GE Hawkeye system. They found apical or anterior perfusion defects in

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X-ray-based attenuation correction of myocardial perfusion scans Gru¨ning et al. 857

the AC scans in 27 of 140 patients (19%) who had a normal non-AC scan. In 21 of these patients, defects disappeared after correcting for misalignment of the emission scans and the transmission map, caused by a slight dip in the imaging table when the patient was transferred from the emission to the transmission position. Apical, anterior and septal defects occurred with a misalignment of as little as 0.5 pixel when using a phantom. The effect is only partially corrected for following the introduction of larger supporting rollers by the manufacturer. Similar findings have been reported by Coffey [15] and Tonge et al. [16].

in abnormal and 26% in normal scans, respectively). This may to some extent demonstrate the limitations of using human observers as judges of the ‘best’ technique: their approach will always, at least partially, be guided by personal preference, previous practice and training. That AC was only rarely required for the rest scan is not surprising: the distinction between true and artificial perfusion defects is largely being made in the stress scan, with the rest scan only being used to assess reversibility. To do away with AC for the rest scan has the benefit of significant time savings, in the order of 5 min per scan, and also lower radiation exposure.

From a diagnostic point of view, there now seems to be consensus that AC can deliver a benefit [1,2,11], and there is a substantial number of studies in support of this [5,7,8,10,17]. Some studies, however, found no improvement over ECG-g [18] or even a worse performance of AC compared to conventional acquisition [19]. Some of these differences may be explained by the different technical performance of various commercial systems [4] or by the impact of growing experience in reading AC scans [20].

From a practical point of view, the fact that we failed to achieve AC in a significant proportion of our patients for a wide variety of reasons (in 23% of patients for reasons that could occur in other departments as well, and in 20% of patients due to our particular imaging algorithm) may seem disappointing. This should, however, be seen in light of the fact that this study was conducted soon after the system had been installed and that lower downtime and better operator experience might be expected in the future. Some of the limitations we encountered in larger numbers of patients (narrow X-ray gantry) may be resolved with future implementations. Perhaps, contrary to expectations, we found body mass and chest circumference not to be useful predictors for benefit from AC; although patients who did benefit from AC had a significantly higher body mass and chest circumference than those who did not, the overlap was too large to make this of any practical value.

The design of our study is less ambitious than some of the previous work in that our observers did not read AC and non-AC scans independently. We are, therefore, unable to determine whether AC led to normal scans becoming abnormal and/or vice versa, and whether it influenced the number of equivocal reports. Whilst we realize that such an independent reading design is superior in a strictly methodical sense, we feel that a completely blind reading of AC and non-AC scans with experienced observers is impossible to achieve in practice (at least with the Hawkeye system), because the AC slices are significantly noisier which makes them easy to recognize, thereby introducing a significant bias in the observers reports. Instead, we have opted to read under clinical conditions when AC and non-AC studies should always be interpreted side by side [2,11]. This precludes the unbiased creation of separate AC and non-AC reports, but still enables the observers to state whether there was a benefit from AC. In our study, observers found AC essential in 20% of scans, compared to 6.6% for ECG-g. Observers’ rating for AC was higher than that for ECG-g in 31% of scans, and vice versa in 17% of scans, indicating superior performance of AC in 14% of scans. Both techniques received an equal rating in 22% of scans. Neither technique was needed for interpretation of 30% of scans. This is not surprising as an experienced observer will be able to recognize typical attenuation artefacts and will, therefore, be able to correctly identify normal scans with a very high accuracy. Indeed, when differentiating between normal scans and those that contained perfusion defects, we found that AC provided a diagnostic benefit significantly more often in abnormal (73%) than normal (56%) scans. The benefit provided by AC was not significantly different from that derived by ECG-g (68%

Conclusions In day-to-day practice, X-ray-based AC acquisition was not feasible in 23% of patients for reasons that are applicable to other departments as well. K For clinical interpretation, AC was found to be essential more often than ECG-g, and also received higher ratings from observers more often, but these differences were not significant. The benefit from both AC and ECG-g was significantly greater in scans with perfusion defects than in normal scans. K The diagnostic impact of AC for the rest scan is minimal. K The GE Hawkeye system causes AC-related artificial perfusion defects in the apex and anterior wall. K Easily available parameters such as body mass and chest circumference cannot be used to predict which patients will benefit from AC. K

References 1

Strauss HW, Miller DD, Wittry MD, Cerqueira MD, Garcia EV, Iskandrian AS, et al. Procedure guideline for myocardial perfusion imaging. Society of Nuclear Medicine. J Nucl Med 1998; 39:918–923.

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Anagnostopoulos C, Harbinson M, Kelion A, Kundley K, Loong CY, Notghi A, et al. Procedure guidelines for radionuclide myocardial perfusion imaging. Nucl Med Commun 2003; 24:1105–1119. 3 Scha¨fers M. Methods and clinical applications in nuclear cardiology: a position statement. Nuklearmedizin 2002; 41:3–13. 4 O’Connor MK, Kemp B, Anstett F, Christian P, Ficaro EP, Frey E, et al. A multicenter evaluation of commercial attenuation compensation techniques in cardiac SPECT using phantom models. J Nucl Cardiol 2002; 9:361–376. 5 Duvernoy CS, Ficaro EP, Karabajakian MZ, Rose PA, Corbett JR. Improved detection of left main coronary artery disease with attenuation-corrected SPECT. J Nucl Cardiol 2000; 7:639–648. 6 Gallowitsch HJ, Unterweger O, Mikosch P, Kresnik E, Sykora J, Grimm G, et al. Attenuation correction improves the detection of viable myocardium by thallium-201 cardiac tomography in patients with previous myocardial infarction and left ventricular dysfunction. Eur J Nucl Med 1999; 26:459–466. 7 Kjaer A, Cortsen A, Rahbek B, Hasseldam H, Hesse B. Attenuation and scatter correction in myocardial SPET: improved diagnostic accuracy in patients with suspected coronary artery disease. Eur J Nucl Med 2002; 29:1438–1442. 8 Kluge R, Sattler B, Seese A, Knapp WH. Attenuation correction by simultaneous emission–transmission myocardial single-photon emission tomography using a technetium-99m-labelled radiotracer: impact on diagnostic accuracy. Eur J Nucl Med 1997; 24:1107–1114. 9 Links JM, Becker LC, Rigo P, Taillefer R, Hanelin L, Anstett F, et al. Combined corrections for attenuation, depth-dependent blur, and motion in cardiac SPECT: A multicenter trial. J Nucl Cardiol 2000; 7:414–425. 10 Links JM, DePuey G, Taillefer R, Becker LC. Attenuation correction and gating synergistically improve the diagnostic accuracy of myocardial perfusion SPECT. J Nucl Cardiol 2002; 9:183–187. 11 Hendel RC, Corbett JR, Cullom SJ, DePuey EG, Garcia EV, Baterman TM. The value and practice of attenuation correction for myocardial perfusion SPECT imaging: A joint position statement from the American Society of

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Nuclear Cardiology and the Society of Nuclear Medicine. J Nucl Med 2002; 43:273–280. Ficaro EP. Should SPET attenuation correction be more widely employed in routine clinical practice? For. Eur J Nucl Med 2002; 29:409–412. Wackers FJT. Should SPET attenuation correction be more widely employed in routine clinical practice? Against. Eur J Nucl Med 2004; 29:412–415. Fricke H, Fricke E, Weise R, Kammeier A, Lindner O, Burchert W. A method to remove artifacts in attenuation-corrected myocardial perfusion SPECT Introduced by misalignment between emission scan and CT-derived attenuation maps. J Nucl Med 2004; 45:1619–1625. Coffey JP. Attenuation correction of myocardial SPECT studies. Nucl Med Commun 2005; 26:929–930. Tonge CM, Manoharan M, Lawson RS, Shields RA, Prescott MC. Attenuation correction of myocardial SPECT studies using low resolution computed tomography images. Nucl Med Commun 2005; 26:231–237. Narayanan MV, King MA, Pretorius PH, Dahlberg ST, Spencer F, Simon E, et al. Human-observer receiver-operating-characteristic evaluation of attenuation, scatter, and resolution compensation strategies for 99mTc myocardial perfusion imaging. J Nucl Med 2003; 44:1725–1734. Lee DS, So Y, Cheon GJ, Kim KM, Lee MM, Chung JK, et al. Limited incremental diagnostic values of attenuation-noncorrected gating and ungated attenuation correction to rest/stress myocardial perfusion SPECT in patients with an intermediate likelihood of coronary artery disease. J Nucl Med 2000; 41:852–859. Banzo I, Pena FJ, Allende RH, Quirce R, Carril JM. Prospective clinical comparison of non-corrected and attenuation- and scatter-corrected myocardial perfusion SPECT in patients with suspicion of coronary artery disease. Nucl Med Commun 2003; 24:995–1002. Slart RH, Que TH, van Veldhuisen DJ, Poot L, Blanksma PK, Piers DA, et al. Effect of attenuation correction on the interpretation of 99mTc-sestamibi myocardial perfusion scintigraphy: the impact of 1 year’s experience. Eur J Nucl Med Mol Imaging 2003; 30:1505–1509.

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Original article

Acceleration of hepatobiliary excretion by lemon juice on 99m Tc-tetrofosmin cardiac SPECT Shiou-Chi Chernga, Yeong H. Chenb, Meei S. Leec, Shih P. Yangd, Wen S. Huanga and Cheng Y. Chenga Background We sought to determine whether drinking lemon juice reduces extra-cardiac activity and improves image quality on 99mTc-tetrafosmin myocardial single photon emission computed tomography (SPECT). Methods Eighty male patients were enrolled in this study and divided into four groups with 20 patients in each group. Each patient received 259–333 MBq tetrofosmin. Ten minutes after injection no action was taken for group 1 (G1), patients in group 2 (G2) each drank 250 ml of water, patients in group 3 (G3) each drank 250 ml of whole milk, and patients in group 4 (G4) each drank 250 ml diluted lemon juice. Myocardial perfusion imaging without attenuation correction was performed after a 1 day rest–stress protocol. Both rest and stress images were aligned at corresponding slices for comparison. Interfering activity was determined visually on reconstructed images, and the heart-to-liver (H/L) ratios were calculated with planar images at 25–30 min and at 45–50 min. Results Interfering activity was seen in 80% of G1, 70% of G2, 60% of G3, and 35% of G4 (G4 vs. G1, P = 0.006) on rest images, and in 70% of G1, 60% of G2, 55% of G3, and 30% of G4 (G4 vs. G1, P = 0.014) on stress images at 25–30 min. It was also observed in 60% of G1, 50% of G2, 45% of G3, and 15% of G4 (G4 vs. G1, P = 0.006) on rest images, and in 50% of G1, 45% of G2, 40% of G3, and 10% of G4 (G4 vs. G1, P = 0.011) on stress images at 45–50 min. The mean

Introduction When using 99mTc-tetrofosmin or 99mTc-sestamibi (MIBI) cardiac single photon emission computed tomography (SPECT) imaging in the evaluation of myocardial perfusion, extra-cardiac activity from the liver and bowel may lead to false negative or false positive results because the heart lies on the diaphragm just above the left lobe of the liver and near the bowel [1–3]. Several different protocols, including eating a fatty meal [4], drinking milk [4], drinking milk and water [5], drinking milkshakes [6], drinking iodinated oral contrast [7], intravenous injection of cholecystokinin (CCK) [4], and administration of metoclopramide [8], have been studied as methods in an attempt to reduce the artifacts arising from abdominal activity. The conclusive benefits, however, are controversial [5,7–10]. Physiologically, acid-rich food or drink has the potential to facilitate hepatobiliary clearance of the bile by increasing

H/L ratios of rest images were 0.47 ± 0.13 for G1, 0.71 ± 0.17 for G2, 0.65 ± 0.12 for G3, and 0.93 ± 0.23 for G4 at 25–30 min, and 0.63 ± 0.14 for G1, 0.73 ± 0.14 for G2, 0.85 ± 0.25 for G3, and 1.15 ± 0.25 for G4 at 45–50 min. On stress images, they were 0.49 ± 0.11 for G1, 0.74 ± 0.16 for G2, 0.69 ± 0.11 for G3, and 0.98 ± 0.22 for G4 at 25–30 min, and 0.66 ± 0.15 for G1, 0.77 ± 0.11 for G2, 0.89 ± 0.26 for G3, and 1.21 ± 0.19 for G4 at 45–50 min. Conclusion Drinking 250 ml of diluted lemon juice accelerates the transit of tetrofosmin through the liver parenchyma and improves image quality on 99mTc-tetrafosmin myocardial SPECT. Nucl Med Commun 27:859–864

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:859–864 Keywords: lemon juice, extra-cardiac radioactivity, myocardial SPECT, heart-to-liver ratio

99m

Tc-tetrofosmin

Departments of aNuclear Medicine, bFamily Medicine, cSchool of Public Health and dDivision of Cardiology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan, R.O.C. Correspondence to Dr Shiou-Chi Cherng, Department of Nuclear Medicine, Tri-Service General Hospital, 325, Section 2, Cheng-Kung Road, Taipei 114, Taiwan, R.O.C. Tel: + 00886 2 8792 7374; fax: + 00886 2 8792 7217; e-mail: [email protected] Received 16 May 2006 Accepted 4 August 2006

the secretion of secretin [11]. To our knowledge, the effect of acid on interfering extra-cardiac activity in myocardial perfusion SPECT has not been described. In the present study, we used diluted lemon juice, an acidrich drink, as an alimentary cholekinetic stimulus to assess the influence of drinking lemon juice on tetrofosmin transit through the liver. The aim of this study was to determine whether this protocol could reduce extracardiac activity and result in an improvement of image quality.

Methods Study population

A total of 80 male patients (aged 42–74 years; mean 60 years) referred for myocardial perfusion imaging (MPI) for coronary artery disease (CAD) or suspicion of CAD were enrolled in this study. Subjects were randomized to four groups of 20 patients in each group. There was no

c 2006 Lippincott Williams & Wilkins 0143-3636

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860 Nuclear Medicine Communications 2006, Vol 27 No 11

Table 1

Patients’ characteristics shown as mean numbers and standard deviations

Variable Number of patients Age (years) Weight (kg) BMI (kg
m – 2)

Total

Group 1

Group 2

Group 3

Group 4

P value*

80 60 ± 8.8 74 ± 4.8 26.7 ± 3.0

20 59 ± 8.2 75 ± 4.7 26.9 ± 2.3

20 62 ± 8.9 73 ± 5.1 26.4 ± 3.6

20 60 ± 9.1 74 ± 4.2 26.5 ± 3.1

20 58 ± 8.7 74 ± 4.9 27.2 ± 2.9

– > 0.05 > 0.05 > 0.05

Group 1: no action taken after injection; group 2: patients drank 250 ml water, group 3: patients drank 250 ml whole milk, and group 4: patients drank 250 ml diluted lemon juice 10 min after injection. BMI: body mass index. * One-way ANOVA.

intervention in group 1 (G1), water was given to group 2 (G2), milk to group 3 (G3), and diluted lemon juice to group 4 (G4). All patients met the following criteria: no previous cholecystectomy, no liver or biliary system disease, and no peptic ulcer within the last 6 months. Patients with a history of diabetes, previous myocardial infarction within the last 2 months, unstable angina, severe primary valvular disease, left ventricular aneurysm, primary cardiomegaly, left ventricle hypertrophy or severe conduction disturbances were also excluded from the study. Patient characteristics are listed in Table 1. Protocol

After a 1 day rest–stress imaging protocol, which was approved by the Tri-Service General Hospital Institutional Review Board, all groups underwent 99mTctetrofosmin gated cardiac SPECT. Rest images were first obtained from all patients. After injection of 259– 333 MBq of tetrofosmin for 10 min, patients in G1 did nothing; each patient in G2 drank 250 ml water; G3 drank 250 ml whole milk (nutrient content per 30 ml was: total fat, 1.2 g; protein, 0.9 g; carbohydrate, 1.3 g; cholesterol, 10 mg; and sodium, 13 mg; pH = 6.8), and G4 drank 250 ml diluted lemon juice (150 ml juice + 100 ml water; nutrient content per 30 ml was: vitamin C, 7 mg; carbohydrate, 0.7 g; and sodium, 10 mg; pH = 2.0); all groups had their first rest MPI carried out at 25–30 min and the second MPI at 45–50 min post-injection. Later, a standard 4 min infusion of 0.56 mg
kg – 1
min – 1 of dipyridamole was administrated to stress the heart, and 740– 925 MBq of tetrofosmin was injected 3 min after the end of the dipyridamole infusion. Then, patients underwent light walking exercise for 5 min and were treated following rest image for all groups; they had their two stress MPIs carried out at the same time points as rest MPI. Imaging acquisition

Imaging was performed using a double-head rotating large field of view gamma camera (Hawkeye, Millennium VG; GE Company, Wisconsin, USA) equipped with a lowenergy, all-purpose, parallel-hole collimator and connected to a dedicated computer system. One planar image was first obtained in the true anterior position with an acquisition time of 90 s; then SPECT images were acquired on a 64  64 matrix. Sixty images (25 s for rest, 20 s for stress) were obtained over a semicircular 1801 arc.

Filtered back-projection was performed with a lowresolution Butterworth filter; no attenuation or scatter correction was applied. Transaxial tomograms were reconstructed, and the images were re-oriented into three sets of orthogonal slices, including short-axis, horizontal, and vertical long axis for each study. Data analysis

Both rest and stress images were aligned at corresponding slices for comparison. The existence of radioactivity in the liver, biliary tract, gut or stomach in all patients was determined visually on filtered back-projection images by three experienced nuclear medicine physicians. Activity was classified as interfering when it could result in either an overestimation or underestimation of uptake in the myocardium and when the readers required additional information from the stress SPECT to confidently assess the perfusion in the inferior, posterior or septal walls of the left ventricle. In a few instances in which there was no initial agreement, the data were reviewed simultaneously by the three physicians at a later date, and a consensus was reached. In addition, the heart-to-liver (H/ L) ratio, another parameter for comparing image quality, was calculated for each patient. Individual regions of interest (ROIs) were manually drawn over the heart and the right lobe of the liver in the image, just as was performed in the study of Munch et al. [12]. The same ROIs were used for the two time points for each patient, and similar ROIs were used for all patients. Statistical analysis

Patient characteristics and the H/L ratios were expressed as mean ± standard deviation. The differences of mean H/L ratios of different time points among groups were assessed by one-way ANOVA. Post-hoc comparisons between groups were done by Tukey tests. Chi-squared tests and logistic regression analyses were used to determine the effect of different treatments on interfering activity. A P value of less than 0.05 was considered to be statistically significant.

Results Table 1 shows that differences in ages, body weights, and body mass indexes among different study groups were not statistically significant (P > 0.05). Livers, gallbladders, common bile ducts and splanchnic activities were apparently seen in all patients’ myocardial images. Gastric

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Effect of lemon juice on cardiac SPECT Cherng et al. 861

activity was seen in some of them. Figures 1 and 2 demonstrate a subject’s rest and stress images in each group at 25–30 min and at 45–50 min after injection, respectively. The myocardial activities caused marked interference with the hepatic activities in the images obtained at 25–30 min, and the interference was more severe in G1 than in G2, G3 and G4. Comparing the interference at 25–30 min and at 45–50 min, the latter interferences were milder in G2, G3 and G4 groups.

The percentages of interfering activity that existed at the two moments of rest image are shown in Table 2, and of stress image in Table 3. At rest, there were significant differences in interfering activity among these four groups at 25–30 min (one-way ANOVA, P = 0.024) and at 45–50 min (P = 0.027). Further comparison by post-hoc tests all showed that G4 had significantly lower interfering activity than G1 at the two time points of imaging (P = 0.006). On stress, there was significant difference in

Fig. 1

A subject’s planar imaging in each group obtained at 25–30 min (upper panel) and at 45–50 min (lower panel) after tetrofosmin injection at rest.

Fig. 2

The same subject’s planar imaging in each group obtained at 25–30 min (upper panel) and at 45–50 min (lower panel) after tetrofosmin injection on stress.

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Table 2

Percentages of interfering activity observed at the two moments of the rest image

Measurement

At 25–30 min At 45–50 min

Group 1

2

3

4

80% 60%

70% 50%

60% 45%

35% 15%

P value*

OR (95% CI)

0.006 0.006

0.032–0.562 0.026–0.537

P value*

OR (95% CI)

0.014 0.011

0.047–0.710 0.020–0.610

*

Chi-squared tests and logistic regression analyses, OR: odds ratio.

Table 3

Percentages of interfering activity observed at the two moments of the stress image

Measurement

At 25–30 min At 45–50 min

Group 1

2

3

4

70% 50%

60% 45%

55% 40%

30% 10%

*

Chi-squared tests and logistic regression analyses, OR: odds ratio.

Table 4 Mean heart-to-liver (H/L) ratios of rest image at the two time points

Table 5 Mean heart-to-liver (H/L) ratios of stress image at the two time points

Group

Group

1 2 3 4

Measurement At 25–30 min

At 45–50 min

0.47 ± 0.13** 0.71 ± 0.17** 0.65 ± 0.12 0.73 ± 0.23*

0.63 ± 0.14** 0.73 ± 0.14 0.85 ± 0.25** 1.15 ± 0.25*

1 2 3 4

Measurement At 25–30 min

At 45–50 min

0.49 ± 0.11** 0.74 ± 0.16** 0.69 ± 0.11 0.98 ± 0.22*

0.66 ± 0.15** 0.77 ± 0.11 0.85 ± 0.26** 1.21 ± 0.19*

*

Group 4 vs. group 1, P < 0.01; Group 2 vs. group 1 at 25–30 min and group 3 vs. group 1 at 45–50 min, P < 0.05.

*

**

**

interfering activity among these four groups at 45–50 min (P = 0.039), but not at 25–30 min (P = 0.072). Post-hoc tests also showed that G4 had significantly lower interfering activity than G1 at 25–30 min (P = 0.014) and at 45–50 min (P = 0.011).

ratios can be increased in patients who drink 250 ml of water, milk or diluted lemon juice 10 min after injection on both rest and stress images. The effects were more pronounced in the group that drank diluted lemon juice (Fig. 1 and Tables 2–5).

The mean H/L ratios of rest image at 25–30 min and at 45–50 min are shown in Table 4. The ratios of G4 patients were significantly higher than those of G1, G2 and G3 (G4 vs. G1, P < 0.01). Comparing G1, G2, and G3, we found: G1 vs. G2, P < 0.05; G1 vs. G3 and G2 vs. G3, P > 0.05 at 25 to 30 min, and G1 vs. G3, P < 0.05; G1 vs.G2 and G2 vs. G3, P > 0.05 at 45 to 50 min. Table 5 shows the H/L ratios of stress image at the two time points. The ratios of G4 patients were significantly higher than those of G1, G2, and G3 (G4 vs. G1, P < 0.01). Although the standard deviation of the ratios of G3 patients was large, there was still a significant difference between the ratios of G3 and those of G1 at 45–50 min (P < 0.05). Comparing other groups, we found: G1 vs. G2, P < 0.05; G1 vs. G3 and G2 vs. G3, P > 0.05 at 25–30 min, and G1 vs. G2 and G2 vs. G3, P > 0.05 at 45–50 min.

Several reports [7,9,10] have mentioned that the combination of exercise and pharmacological stress, just as in our protocol, will improve image quality and enhance the H/L ratio. In our study, the percentages of interfering activity of stress image were slightly lower than those of rest image, and the H/L ratios were mildly higher, but there were no significant differences in both between the two images. Thus, it becomes difficult to determine the effects of drinking water, milk and lemon juice on stress imaging.

Discussion Our study demonstrated that the passage of tetrofosmin through the hepatobiliary system can be accelerated, the extra-cardiac activities can be decreased, and the H/L

Group 4 vs. group 1, P < 0.01; Group 2 vs. group 1 at 25–30 min and group 3 vs. group 1 at 45–50 min, P < 0.05.

The modality widely used to decrease extra-cardiac activity for tetrofosmin or MIBI cardiac imaging is to prolong the injection-to-imaging time [5,7,10]. Peace and Lloyd [10] thought that delayed imaging was the only way to significantly reduce the interference of extracardiac activity with observer interpretation. Essentially, our results are similar to the previous findings and indicate that longer injection-to-imaging times improve image quality. However, the effects are limited if the delayed time is less than 1 h.

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Effect of lemon juice on cardiac SPECT Cherng et al. 863

Drinking water to eliminate the extra-cardiac activity of 99m Tc-labelled cardiac SPECT has been studied by several investigators. van Dongen and van Rijk [5] reported that drinking 450 ml of water 10 min before imaging did not reduce extra-cardiac activity. Peace and Lloyd [10] also showed that drinking 150 ml of full-fat milk and 450 ml of water made no improvement versus control groups. In contrast, Boz and Karayalcin [13] found that administrating 300 ml of water before imaging did increase diagnostic accuracy. Our study was different in that we gave 250 ml of water 10 min after tetrofosmin injection. The mean H/L ratios significantly increased at 25–30 min although they did not at 45–50 min, comparing G2 and G1 groups (Tables 4 and 5). These results suggested that the effect of accelerating hepatic tetrofosmin clearance after drinking water is better if patients drink water at a time closer to the time of imaging acquisition. A possible explanation is that gastric dilatation after drinking water pushes the extra-cardiac activity away from the inferior wall of the myocardium only, but does not enhance bile secretion and decrease extracardiac activity. Lipid-rich foods in the duodenum enhance the secretion of CCK, accelerating bile secretion and gallbladder emptying [14]. Many researchers thus design protocols for tetrofosmin or MIBI MPI with a lipid-rich drink (milk) as a factor to observe influences on the images. However, these studies are not in agreement with each other [3,5]. From our results, we observe that drinking milk decreases interference and improves image quality of tetrofosmin MPI, but only to a suboptimal degree. The possible causes of these discrepancies include different compositions of various milk products, the different times of drinking the milk and the different amounts drunk, different patient characteristics, and the possibility of hepatobiliary diseases and gastroesophageal reflux [1]. Lemon juice is rich in vitamin C, with a pH as low as 2.0. It is acidic enough to enhance bile secretion by stimulating intestinal release of secretin [11]. Unlike CCK, secretin significantly accelerates bile secretion, but plays little part in emptying the gallbladder [11]. Therefore, the hepatic clearance of tetrofosmin is increased, but splanchnic activities are not enhanced by gallbladder emptying.

of G2 at 45–50 min, however, and the difference between ratios for G3 and G1 patients became significant (Table 4). It is most likely that 6–26 min is required for milk to enhance CCK release, and bile secretion [15,16] limits the effect on the images that are taken at 25–30 min, only 15 min after milk intake. Conversely, lemon juice can more quickly enhance bile secretion and hepatic clearance of tetrofosmin than can milk. We designed this study so that four groups were included and tried to take only one parameter for comparison for tetrofosmin myocardial SPECT. We did our best to enroll patients having similar characteristics, including gender, body mass indexes and normal serum levels of liver enzymes. Thus, case collection was not easy, and the study population was not large. Additionally, a major limitation of our study is that we did not compare the percentages of interfering activities and the H/L ratios between different protocols (none, water, milk, lemon juice) within each patient. Also, it is challenging to measure secretin increases after patients drink lemon juice; such measurement was not done in this study because of the short half-life of secretin, which is 3–8 min [14].

Conclusion Drinking 250 ml diluted lemon juice 10 min after tetrofosmin injection on rest and stress MPI can improve image quality by accelerating tetrofosmin passage through the liver, and decreasing extra-cardiac, notably hepatic, interfering activities. According to these results, a drink of lemon juice may be recommended as a simple technique to decrease extra-cardiac activity on 99mTctetrofosmin cardiac SPECT. 99mTc-MIBI, which is another myocardial perfusion tracer, has a longer hepatic clearance time than tetrofosmin. Further investigation is required to determine whether drinking lemon juice has similar effects on 99mTc-MIBI and 99mTc-tetrofosmin MPI.

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Our results also demonstrated that drinking water, milk and lemon juice and delayed imaging were useful for improving the MPI quality because the mean H/L ratios of G2, G3, and G4 patients at 25–30 min were higher than those of G1, and even much higher than those at 45– 50 min. Interestingly, the H/L ratios of G3 patients were lower than those of G2 at 25–30 min, and there was no significant difference between ratios for G3 and G1 groups. The ratios of G3 patients were higher than those

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Pinkert J, Kutzner H, Lindner C, Lopez J, Kropp J, Franke WG. Reduction of artifacts related to intestinal activity in myocardial perfusion SPECT with sestamibi. J Nucl Med 1997; 38(suppl):75P. 10 Peace RA, Lloyd JJ. The effect of imaging time, radiopharmaceutical, full fat milk and water on interfering extra-cardiac activity in myocardial perfusion single photon emission computed tomography. Nucl Med Commun 2005; 26:17–24. 11 Chen WY, Li P, Jin H, Lee KY, Chang TM. Mechanisms on the release and physiological actions of secretin. Biomed Res 1994; 15(suppl):151–159. 12 Munch G, Neverve J, Matsunari I, Schroter G, Schwaiger M. Myocardial technetium-99m-tetrofosmin and technetium-99m-sestamibi kinetics in normal subjects and patients with coronary artery disease. J Nucl Med 1997; 38:428–432.

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Boz A, Karayalcin B. Which is better for inferior wall evaluation: a full or empty stomach?. J Nucl Med 1996; 37:1916–1917. Kutchai HC. Gastrointestinal secretions. In: Berne RM, Levy MN (editors): Physiology, third edition. Missouri: Mosby Year Book; 1993, pp. 684–685. Bobba VR, Krishnamurthy GT, Kingston E, Turner FE, Brown PH, Langrell K. Gallbladder dynamics induced by a fatty meal in normal subjects and patients with gallstone: concise communication. J Nucl Med 1984; 25: 21–24. Krishnamurthy GT, Brown PH. Comparison of fatty meal and intravenous cholecystokinin infusion for gallbladder ejection fraction. J Nucl Med 2002; 43:1603–1610.

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Original article

Clinical value of planar and tomographic dual-isotope scintigraphy using 99mTc-methylene diphosphonate and 131I in patients with thyroid cancer Johann Schoenberger, Ernst Fuchs, Victoria Fertig, Attila Szikszai, Peter Maenner and Christoph Eilles Purpose 131I whole-body scintigraphy is a highly sensitive method for the detection of differentiated thyroid tumours and metastases. However, a lack of anatomical landmarks and the physiological excretion of the tracer complicates the evaluation of the images. Therefore, we determined whether additional bone scintigraphy in combination with 131 I scintigraphy, simultaneously acquired via planar and tomographic techniques, positively contributes to the treatment plan in patients with non-conclusive 131I images. Methods Twenty-one patients with differentiated thyroid cancer and known metastases or unclear findings in the 131 I whole-body scan underwent dual-isotope scintigraphy (DIS) within 2–7 days after application of 5000–8000 MBq 131 I. Dual-energy planar and tomographic data were acquired simultaneously and the results compared with other imaging modalities.

(24%), DIS did not add any new data regarding the extent of the disease. Conclusions The simultaneous acquisition of 131I and Tc-methylene diphosphonate provides clear landmarks and facilitates the localization of functioning metastases from differentiated thyroid cancer as well as improves the fusion with morphological images. It can be performed easily and also transferred to other isotope combinac 2006 Lippincott tions. Nucl Med Commun 27:865–871 Williams & Wilkins. 99m

Nuclear Medicine Communications 2006, 27:865–871 Keywords: thyroid cancer, radionuclide bone scanning, whole-body scintigraphy, dual-isotope scintigraphy

131

I

Department of Nuclear Medicine, University of Regensburg, Germany.

Results In 48% of the cases (10 of 21), DIS supplied important additional information that either altered the treatment plan or staging of the patients. In 28% (six of 21), DIS provided new information that was not known before, but did not change the staging of the patients. In five cases

Correspondence to Dr Johann Schoenberger, Department of Nuclear Medicine, University of Regensburg, 93042 Regensburg, Germany. Tel: + 0049 941 944 7524; fax: + 0049 941 944 7502; e-mail: [email protected]

Introduction

ability of thyroid-derived malignant cells to concentrate iodine is the basis for the diagnostic and therapeutic use of 131I [3]. The demonstration of abnormal foci of radioiodine accumulation seen on the post-therapy scintigraphy is the strongest evidence of metastases or recurrence [4].

Patients with papillary and follicular thyroid cancer show an excellent long-term outcome due to the high grade of differentiation and the use of 131I for treatment [1]. Nevertheless, there are also patients who already show a metastatic spread of the disease at the time of initial diagnosis, or other patients with a high risk of tumour recurrence and progression including the development of distant metastases. Most distant metastases from differentiated thyroid cancer are regional lymph node metastases and metastases located in the lungs and bones, or both. Finally, the major causes of death include respiratory failure from pulmonary metastases, airway obstruction, or bleeding from local vascular invasion, as well as neurological complications from brain and spinal cord involvement [2]. Therefore, the early detection of distant metastases is a very important issue and is based on the use of several different imaging modalities. Among these, scintigraphy using 131I represents the most relevant technique. The

Received 12 April 2006 Accepted 12 July 2006

However, there are also certain factors that weaken the clinical value of this imaging technique. Due to the lack of anatomical landmarks, a precise localization of the foci of 131 I accumulation can be difficult. In addition, the physiological excretion and the regular bio-distribution of 131 I in the salivary glands, gut, stomach and urinary bladder complicates the evaluation of 131I whole-body images [5]. In patients suspected of having metastatic spread of the disease, bone scintigraphy using 99mTc-methylene diphosphonate (99mTc-MDP) is one of the most commonly used screening methods for the detection of bone metastases [6–8]. However, bone metastases are often purely osteolytic, causing bone scintigraphy to show

c 2006 Lippincott Williams & Wilkins 0143-3636

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866 Nuclear Medicine Communications 2006, Vol 27 No 11

decreased, or only moderately increased, uptake which leads to a low sensitivity and poor specificity. On the other hand, bone scintigraphy represents an excellent method for visualizing the whole skeleton and significantly improves the anatomical orientation. In a preliminary report, Tenenbaum and colleagues described the usefulness of the combination of bone scintigraphy and 131I whole-body scintigraphy in localizing bone metastases of differentiated thyroid carcinoma [9]. However, the acquisition of only planar images in their study posed a limitation to this technique. Today, modern gamma cameras partially equipped with thicker crystals provide the ability to obtain 131 I single photon emission computed tomography (SPECT) images of a much higher quality.

October 2005) with known metastases from thyroid cancer or unclear results of the 131I scintigraphy (13 females, eight males; age, 23–78 years; mean 58 years; nine patients had a follicular tumour, nine a papillary tumour and in three patients the primary tumour was not found in the thyroid (Table 1)). All patients had been previously treated by thyroid surgery and had received their first or second therapy with 131I. Diagnosis of detectable lesions was proven by spiral computer tomography (CT), magnetic resonance imaging (MRI) or follow-up (median follow-up period was 21.5 months). The study was approved by a local ethics committee and all patients gave their informed written consent.

Therefore, the purpose of this study was to evaluate the combination of bone and 131I whole-body scanning for the detection of functioning metastases of thyroid cancer and to identify its potential role in treatment planning. We determined whether the addition of dual-isotope scintigraphy (DIS) to the standard evaluation process, using planar and tomographic techniques, contributes to the management plan in patients with functioning metastases from differentiated thyroid carcinoma. For this purpose, we present the data of 21 patients whose clinical assessment included a DIS in addition to standard imaging techniques.

Gamma camera and acquisition parameters

Patients and methods Patients

For this retrospective analysis we evaluated the data of 21 patients (131I whole-body scan between March 2002 and Table 1

Images were acquired using a double-head gamma camera (E.CAM duet; Siemens, Erlangen, Germany) equipped with a 1-inch NaI crystal and high-energy collimators. Scans were usually performed after a dose rate lower than 12 mSv
h – 1 at 1 m distance was reached. In all patients, a planar whole-body scan as well as tomographic images were taken. SPECT was performed with a 3601 rotation (1801 per head) in 64 steps (128 projections) of 45 s in two independent energy windows: 140 keV ± 15% for 99m Tc-MPD and 360 keV ± 15% for 131I. Data sets of window 1 and window 2 were processed using a Butterworth filter and iterative reconstruction (cycles/cm: 0.8, order: 5; iteration: 16 subsets, 8 iterations; matrix 128  128).

Characteristics of the patients

Patient number

Thyroglobulin (ng
ml – 1)

Age (years)

Sex

Tumour histology

Stage (UICC 2002)

Metastases

Category 1 1 2 3 4 5 6 7 8 9 10

69 53 60 61 65 43 67 47 60 64

Male Male Male Male Female Male Male Female Female Female

Follicular Papillary Follicular Papillary Follicular Papillary Follicular Papillary Papillary Papillary

pT4TxG1NxM1Rx TxNxM1 T4G4N1aMxRx T1mN0MxR0 T4N1M1R1 T1NxMxL0V0R0 T3NxMx T2aN0M0 Primary not found T3N1M1

Bone Lung Bone Lung Lung, cutan Lymph node Lymph node Bone Bone Lung

<1 <1 22.2 <1 50,2 <1 <1 22,1 71.8 <1

Category 2 11

60

Female

Papillary

T4N1bM1

8558.0

12 13 14 15 16

78 63 23 36 27

Male Female Female Female Female

Follicular Follicular Papillary Follicular Papillary

TxNxM1 Primary not found T3mN1R0M0 primary not found pT4bN1bM0R0

Lung, bone, lymph nodes Bone, lung Bone Lymph nodes Lung, lymph nodes Lymph nodes

Category 3 17 18 19 20 21

51 68 72 72 74

Male Female Female Female Female

Follicular Follicular Follicular Follicular Follicular

T3N1bM1 T1N0V1L0R0M1 T3G1NxM1 T3N0R0L0V0M1 T2NxMx

Lung Bone Liver Bone Lung

467.4 672.2 2.3 1.2 1

293.5 2702 125.7 13 240 <1

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Dual-isotope scintigraphy in thyroid cancer Schoenberger et al. 867

131

I and bone scintigraphy

An activity of 5000–8000 MBq 131I was given after a period of 4–6 weeks during which any thyroid hormone treatment was withheld and iodine contamination was carefully avoided. Two to seven days after application, data acquisition (planar whole-body scan and tomography) was done. To reduce the rate of false positive findings due to the physiological excretion of the tracer, additional procedures were applied. In patients with high tracer accumulation in the salivary glands, for example, lemon juice was given to increase clearance of the iodine. In patients with high intestinal uptake, laxatives were administered and images were acquired again up to 4 days later. If these procedures did not lead to a reliable statement, lesions were defined as suspicious for metastatic disease when abnormal or unclear 131I accumulation on whole-body iodine scintigraphy was present. In these patients an additional bone scintigraphy was performed in which 500 MBq 99mTc-MPD were injected and scans were started 3 h post-injection. First, a wholebody scan was performed, followed by a tomography of at least one, in most cases two, positions with a length of 36.2 cm for each position. Computed tomography and magnetic resonance imaging

Computed tomography data were acquired with a 16-slice CT scanner (Sensation 16; Siemens AG, Erlangen, Germany). For an optimal assessment of the gastrointestinal tract, oral administration of 1500 ml diluted diatrizoate meglumine (Gastrografin 1.5%, Schering AG, Berlin, Germany) was performed 45 min before starting the examination. Scanning parameters were 120 kV, 160 mAs, CARE Dose, 16  0.75 mm section collimation, 500 ms rotation time, and 15 mm table feed per rotation. Axial CT images were reconstructed using an increment of 1.5 mm with a 500 mm field of view and a kernel of B30f medium smooth. Matrix size was 512  512. For further radioiodine administration, CT was performed without contrast medium. MRI was performed using a 1.5-T system (Magnetom Symphony, Siemens Erlangen, Germany). T1-weighted and T2-weighted images with and without fat saturation in the coronal and axial planes were obtained in 5 mm slices. Ten millilitres of gadolinium was injected intravenously as a contrast medium. Thyroglobulin

Thyroglobulin (TG) measurements for each patient were performed using a demeditec (Kiel-Wellsee, Germany) radioimmunoassay with an analytic sensitivity of 0.1 ng
ml – 1. To guarantee optimal results of the test, all

patients showed at least a thyroid stimulating hormone (TSH) level of 30 mU
l – 1 or higher. A TG level < 1 ng
ml – 1 was defined as normal and indicated no evidence for the existence of thyroid tissue. Additionally, autoantibodies against TG were checked in all patients either during the initial examination or when TG recovery of the TG test was not valid (radioimmunoassay by demeditec). Data processing and image analysis

Initially, all reviewers were unaware of the results of other imaging modalities. First, experienced nuclear medicine physicians independently interpreted 131I and bone scans separately. Next, SPECT images without image fusion were also evaluated. Finally, all cases were re-evaluated on the basis of the fusion images including knowledge of the results of other imaging modalities. To calculate the usefulness of DIS each clinical case was categorized as (1) DIS provided additional information that altered the treatment plan or current staging of the patient, (2) DIS provided additional information that was not known before and changed the previous staging of the patient, but ultimately did not alter the treatment plan, or (3) DIS did not provide any additional information.

Results The study included 21 patients with suspected metastatic disease of differentiated thyroid cancer or unclear findings of the 131I scintigraphy who had undergone standard treatment. In 10 cases (48%), DIS generated additional information that was not known before and altered the treatment plan and staging of the patient (category 1). In six cases (28%), DIS clarified findings from other imaging modalities and/or changed the current staging of the patient (category 2). In five cases, DIS did not contribute to the further treatment of the patient or alter the staging (category 3). The following data provide details for each category. Category 1

DIS provided new information that was not known before and altered the treatment plan or previous staging of the patient. Ten of 21 cases were placed in category 1. In four of these patients, DIS demonstrated clearly osseous metastases that were not known before and not identifiable on CT or conventional X-ray (Fig. 1). In only one of these patients, bone scintigraphy alone showed an increased tracer uptake in all osseous metastases (three lesions); in one case one of seven osseous lesions was visible on bone scintigraphy; in the other two patients, bone scintigraphy showed no evidence of bone metastases (one and five lesions within one patient, respectively). Three of these patients showed an increased TG

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868 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 1

(a)

(b)

BS

IS

DIS

BS

IS

DIS

BS

IS

DIS

(a) A 60-year-old man (no. 3) with follicular thyroid cancer. First whole-body scan 4 days after application of 5 GBq 131I and corresponding bone scintigraphy. None of the osseous metastases demonstrate increased metabolic activity on the bone scan or are visible on CT or conventional X-ray. (b) Serial coronal cross-sections give an excellent overview of the correct anatomical localisation of all metastases of the patient (BS = bone SPECT, IS = 131I SPECT, DIS = image fusion).

level (71.8, 22.2 and 22.1 ng
ml – 1) and one patient was TG negative (0.74 ng
ml – 1). In three patients DIS identified pulmonary metastases where 131I planar and tomographic images could not delineate a skin contamination from a metastatic uptake of the tracer. The combination of bone scintigraphy and 131 I images demonstrated a clear intrapulmonary position of the lesions. CT and TG were negative in these patients, but follow-up confirmed the existence of lung metastases.

In two patients 131I showed an unclear focal uptake in the mediastinum. The DIS images with a clear illustration of osseous structures facilitated a correct diagnosis after fusion with the CT images, such that both foci could be identified as affected lymph nodes. Finally, in one patient with a history of thyroid cancer and hypernephroma and newly detected lung lesions (two nodules close to each other), DIS images after fusion with CT showed that only one focus was a metastatic lesion of

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Dual-isotope scintigraphy in thyroid cancer Schoenberger et al. 869

thyroid cancer, the other one belonged to the hypernephroma. The follow-up confirmed these findings. After 131 I therapy, one lesion disappeared whereas the other increased and finally the patient died from metastatic hypernephroma. Figure 1 presents an example from this category where DIS rendered additional information that led to a change in the treatment plan and staging of the patient. Category 2

DIS provided new information that was not known before. Six patients were placed in category 2. In two patients, after fusion with CT images, DIS demonstrated that lung metastases as well as lymph nodes in the mediastinum were involved, which led to a change of the staging from a former N negative status to N positive. In two other patients DIS showed the correct localization of increased foci. 131I images alone did not allow a clear statement of whether only lymph nodes were involved or also osseous structures in the neck and upper mediastinum. In one patient DIS showed additional osseous lesions that were previously not correctly identified as osseous metastases (Fig. 2), and in another patient DIS demonstrated the presence of a pulmonary lesion in addition to already known osseous metastases. Category 3

DIS did not generate any new additional information. Five of 21 cases were placed in category 3. In these patients DIS did not demonstrate any new pathological findings that would change further treatment. In all cases, 131I whole-body scan and SPECT images clearly showed the correct extent of the metastatic disease; in cases where an exact anatomical correlation by 131I images alone was not possible, an additional CT or MRI scan gave the correct information without making use of the DIS. Other imaging modalities

Nearly every patient received additional imaging techniques, preferably CT or MRI. However, the results of these techniques were disappointing. The sensitivity of CT for the detection of osseous or pulmonary metastases was 50%. In only one of six cases were lymph node metastases correctly identified; in the other cases where no enlarged lymph nodes could be seen, it was not possible to determine whether lymph nodes were involved or not. Furthermore, the results of bone scintigraphy alone for the detection of osseous metastases were also unsatisfactory. The sensitivity for this method was 35% with a high rate of unclear findings due to the advanced age of the patients and thus the increased level of degenerative changes in the skeleton. Thyroglobulin

As shown in Table 1, only 13 of 20 patients (65%) showed an increased thyroglobulin level, even under high TSH

stimulation. In one patient with osseous metastases and three patients with lung or lymph node metastases, respectively, the thyroglobulin level was below or equal to the limit of 1 ng
ml – 1. One reason for false negative TG levels might be the presence of TG autoantibodies, however in none of the patients presented in this study was the TG–AB concentration above the value of 60 U
ml – 1 (test cut-off point for healthy subjects).

Discussion Metastatic differentiated thyroid carcinoma remains a diagnostic and therapeutic challenge. Early and exact detection of tumour recurrence and metastases is essential for further treatment and avoidance of serious complications. 131I whole-body scintigraphy plays a central role in the detection, localization and therapy of functioning metastases from differentiated thyroid cancer. Distant metastases are usually located in the lungs and bones, with other less common sites being the skin, liver, and brain where metastases may occur during the course of dissemination. The presence of pathological foci of 131I accumulation in the neck, chest, or osseous structures is strong evidence for metastases or recurrence. Although the specificity of 131I scanning is extremely high, several factors can influence the reliability of this technique [5]. Mainly, the physiological secretion and excretion of 131I from the nasopharynx, salivary glands, stomach, bowel and genitourinary system may lead to false positive as well as false negative results (e.g. Fig. 2 presents a patient with an intense focus along the descending colon that could be caused by colon diverticula, particularly since bone scintigraphy shows a highly increased tracer uptake within only one lesion in the centre of the sacrum). Even images acquired several days after application, where a better background clearance should have taken place, are sometimes difficult to interpret. In contrast, some metastases may possess a capacity for increased radioiodine clearance, such that images acquired some days later no longer display pathological foci. Consequently, misinterpretation of 131I scans is an important source of potential errors that should be avoided whenever possible. In a preliminary study in 1993, Tenenbaum et al. reported on the effectiveness of using a combination of 99mTcdiphosphonate and 131I scanning for localizing bone metastases of differentiated thyroid cancer [9]. Technical limitations allowed only planar images but no tomographic acquisition. However, recent refinements in the technical equipment and the usage of a 1-inch crystal have led to a significant improvement in gamma cameras, particularly for the detection of isotopes with higher energy levels such as 131I.

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870 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 2

(a)

(b)

(c) Tc-99m MDP

I-131

DIS

(d)

(e)

BS

IS

DIS

BS

IS

DIS

(a) A 60-year-old woman (no. 9) with a history of nearly total thyroid resection 8 years ago with no evidence of malignancy. Five weeks before therapy with radioiodine, a follow-up resection was performed due to the presence of cervical metastases. A whole-body 131I scan performed 5 days after application of 5 GBq 131I shows, in addition to residual thyroid tissue, pathological foci in the abdomen (one along the descending colon) and along the femur. (b) Corresponding bone scan demonstrates an intense pathological focus in the sacrum and a poor focus along the left femur. All other foci show no increased tracer-uptake on the bone scan and remain unclear, especially since CT gives no evidence for further osseous or lymph node metastases. (c) Transverse cross-sections of DIS allow a clear localisation of the lesions. All pelvic foci appear to be related to osseous structures, whereas the abdominal focus represents a lymph node metastasis. (d) Serial coronal cross-sections of the abdomen and pelvic area. Bone-SPECT (BS), 131I SPECT (IS), and fusion images (DIS). (e) Corresponding volume-rendered images of the same patient (grey = osseous structures, red = metastases).

In many cases, SPECT can improve the detectability of metastases and provide more information about their correct anatomical localization, but many foci still may remain unclear. Lesions located near the body surface are especially difficult to distinguish from skin contamination. Also, foci located in the centre of the thorax or abdomen are hard to assign to the correct anatomical structure by using tomographic techniques alone. In cases with unclear 131I accumulation on the wholebody scan, a variety of other imaging methods are available for clarification [10]. The differentiation of scar tissue from local recurrence and the discrimination of unspecific lymph node enlargement from lymph node

metastases using CT or MRI is often very difficult [11]. Additionally, the use of CT is limited because this method can usually only be applied without contrast enhancement as a result of the further radioiodine administration. Moreover, all morphological imaging techniques lack sensitivity to some extent due to their limited field of view. In particular, the correct detection of whether lymph nodes are involved is a major problem. In fact, the size of the lymph nodes still remains the most important criterion for assessment [12]. In our study, most of the lymph nodes involved did not show a size greater than 1 cm in diameter, which led to an inconspicuous result from the technique.

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Dual-isotope scintigraphy in thyroid cancer Schoenberger et al. 871

The clinical usefulness of 131I SPECT and CT fusion has been previously described by Yamamoto and colleagues [13]. They used external markers on the skin of the patients to adjust the sections of CT and 131I SPECT images. In this way, they gained important additional information for image interpretation in 15 of 17 patients. In our study, the presence of osseous structures from bone scintigraphy facilitates the correct fusion of CT with 131 I images, and allows the proper diagnosis of lymph nodes involved, the identification of distant metastases, and the delineation of residual thyroid tissue from lymph node metastases in patients receiving their first therapy after thyroidectomy without the need for external markers.

positive metastases in patients with thyroid cancer. Dualisotope scintigraphy allowed precise localization of 131I uptake in patients with differentiated thyroid carcinoma. In addition, the presence of osseous structures facilitates the fusion of functional and morphological images generated by CT or MRI. The technique does not significantly increase the radiation dosage and can be easily performed in any nuclear medicine department.

References 1

2

Bone scintigraphy represents another frequently used method for diagnostic imaging in differentiated thyroid carcinomas. This method is widely accepted as the best screening technique for assessing the whole skeleton for osseous metastases. In differentiated thyroid cancers, however, bone scintigraphy alone is considered to lack the adequate sensitivity for detecting bone metastases [14]. As seen in our study, the sensitivity for the detection of osseous metastases by bone scintigraphy alone was 35%. An unsatisfactory result but, nonetheless, in agreement with other published data. Therefore, in selected cases, the method described above may be an excellent technique for the localization of pathological tracer accumulation. An image that simultaneously shows osseous structures and areas with high 131I uptake allows a clear delineation of intra- or extra-osseous foci. Furthermore, due to the simultaneous acquisition of both nuclides, artefacts caused by movement of the patient are excluded. A recent, newly developed technique is the combination of CT and a gamma camera (SPECT/CT). The combination of functional imaging with the anatomical detail of multislice CT represents the best solution in cases involving metastatic thyroid cancer [15]. Although the availability of this advanced technique is growing, it is currently limited to certain medical centres. Therefore, the technique described in our study offers an effective and beneficial existing alternative for many other departments.

Conclusion The purpose of this study was to establish a more effective method for the anatomical localization of iodine-

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Mazzaferri EL. Radioiodine and other treatments and outcomes. In: Braverman LE, Utiger RD (editors): Werner and Ingbar’s the thyroid: a fundamental and clinical text, 7th edition. Philadelphia: Lippincott-Raven; 1996, pp. 922–943. Kitamura Y, Shimizu K, Nagahama M, Sugino K, Ozaki O, Mimura T, et al. Immediate cases of death in thyroid carcinoma: clinicopathological analysis of 161 fatal cases. J Clin Endocrinol Metab 1999; 84:4043–4049. Spitzweg C, Morris JC. The sodium iodide symporter: its pathophysiological and therapeutic implications. Clin Endocrinol 2002; 57:559–574. Freitas JE, Gross MD, Ripley SD, Shapiro B. Radionuclide diagnosis and therapy of thyroid cancer: current status report. In: Freeman L (editor): Freeman and Johnson’s clinical radionuclide imaging. Orlando: Grune and Stratton; 1986, pp. 1994–2027. Shapiro B, Ruffin V, Jarwan A, Geatti O, Kearfott KJ, Fig LM, et al. Artefacts, anatomical and physiological variants, and unrelated diseases that might cause false-positive whole-body 131-I scans in patients with thyroid cancer. Semin Nucl Med 2000; 30:115–132. McVeil B. Value of bone scanning in neoplastic diseases. Semin Nucl Med 1984; 14:277–286. Franks JA, Ling A, Patronas NJ, Carrasquillo JA, Horvath K, Hickey AM, et al. Detection of malignant bone tumours: MR imaging vs. scintigraphy. AJR Am J Roentgenol 1990; 155:1043–1048. Haubold-Reuter BG, Duewell S, Schlicher BR, Marincek B, von-Schulthess GK. The value of bone scintigraphy, bone marrow scintigraphy and fast spin-echo magnetic resonance imaging in staging of patients with malignant solid tumours: a prospective study. Eur J Nucl Med 1993; 20: 1063–1069. Tenenbaum F, Schlumberger M, Bonnin F, Lumbroso J, Aubert B, Benali H, et al. Usefulness of technetium-99m hydroxymethylene diphosphonate scans in localising bone metastases of differentiated thyroid carcinoma. Eur J Nucl Med 1993; 20:1168–1174. ¨ zbey N, Alago¨l F, et al. Thallium-201, U¨nal S, Menda Y, Adalet I, Boztepe H, O technetium-99-tetrofosmin, and iodine-131 in detecting differentiated thyroid carcinoma metastases. J Nucl Med 1998; 39:1897–1902. Steinkamp HJ, Heim T, Schubeus P, Schorner W, Felix R. The magnetic resonance differential diagnosis between reactively enlarged lymph nodes and cervical lymph node metastases. Ro¨Fo 1992; 157:406–413. van den Berkel M, Stel H, Castelijns J, Nauta J, van der Waal I, Valk I, et al. Cervical lymph node metastasis: assessment of radiologic criteria. Radiology 1990; 177:379–384. Yamamoto Y, Nishiyama Y, Monden T, Matsumura Y, Satoh K, Ohkawa M. Clinical usefulness of fusion of 131I SPECT and CT images in patients with differentiated thyroid carcinoma. J Nucl Med 2003; 44: 1905–1910. Alam MS, Takeuchi R, Kasagi K, Misaki T, Miyamoto S, Iida Y, et al. Value of combined technetium-99m hydroxy methylene diphosphonate and thallium201 imaging in detecting bone metastases from thyroid carcinoma. Thyroid 1997; 7:705–712. Ruf J, Lehmkuhl L, Bertram H, Sandrock D, Amthauer H, Humplik B, et al. Impact of SPECT and integrated low-dose CT after radioiodine therapy on the management of patients with thyroid carcinoma. Nucl Med Commun 2004; 25:1177–1182.

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Original article

Clinical importance of anti-thyroglobulin auto-antibodies in patients with differentiated thyroid carcinoma: Comparison with 99mTc-MIBI scans Ozlem N. Ku¨c¸u¨k, Gu¨lseren Aras, Hulya A. Kulak and Erkan ´Ibis¸ Aims (1) To investigate whether elevated serum anti-thyroglobulin antibody (ATG) reflects the recurrence of cancer in patients with differentiated thyroid carcinoma (DTC) in whom thyroglobulin was undetectable after radioiodine ablation. (2) To assess the sensitivity of disease detection for 99mTc-MIBI whole-body scans (WBSs) in these patients and investigate the correlation between MIBI WBS results and high serum ATG levels. Materials and methods In this retrospective study, we evaluated 14 patients (13 women and 1 man; mean age 44 ± 19 years) with DTC who underwent total or near-total thyroidectomy followed by an ablative dose of 131I at various time intervals. According to histopathological findings, 10 patients (71.4%) who were diagnosed as having papillary carcinoma and four patients (28.6%) as having follicular cell carcinoma, had high serum ATG concentrations ( > 40 IU
ml – 1; range, 62–2000 IU
ml – 1), but low serum thyroglobulin concentrations ( < 1.6 ng
ml – 1). Post-therapeutic and diagnostic 131I WBSs and 99mTc-MIBI WBSs were performed. Scans were visually evaluated for detecting recurrence. If necessary, bone scans, chest X-rays, computerized tomography, ultrasonography and histopathological evaluation were performed. Results Recurrent and/or persistent disease was found in 12 of the patients. This was confirmed pathologically in four patients and by using other imaging methods in eight (bone scans, computerized tomography, ultrasonography). The sensitivity and specificity of disease detection for MIBI WBSs was 66.7% and 100%, respectively. For 131I WBSs,

Introduction Periodic serum thyroglobulin (Tg) measurements and radioiodine whole-body scans (WBSs) are recommended during the follow-up of patients with differentiated thyroid carcinoma (DTC) [1,2]. The major clinical application for serum Tg measurement is the postoperative and post-ablative follow-up of patients with DTC, and this is the most sensitive modality for the detection of recurrent or persistent disease during followup [3,4]. However, measurement of serum Tg is one of the most difficult biochemical tests in which to maintain high reliability [5]. A particularly problematic issue is the detection of Tg in the serum of patients with high antithyroglobulin antibody (ATG). The underestimation of

the sensitivity of disease detection was 55.6%. Among these 12 patients, 10 responded to treatment (three underwent surgery, seven received radioiodine therapy, and two had surgery + radioiodine therapy). ATG levels decreased in eight of the 10 patients, but remained persistently elevated in two despite treatment. Conclusions (1) Persistently elevated ATG levels appear to serve as a useful marker for recurrent or persistent DTC in patients with undetectable serum tyroglobulin levels. Thus, the routine measurement of ATG antibody in such patients is of great value. (2) In these patients, 99mTc-MIBI has a relatively high sensitivity in the diagnosis of a recurrence of thyroid cancer or metastases. So, in patients with elevated ATG but undetectable serum thyroglobulin levels, 99mTc-MIBI can be used to determine whether there is a recurrence of DTC or metastases. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:873–876 Nuclear Medicine Communications 2006, 27:873–876 Keywords: anti-thyroglobulin antibody, carcinoma, recurrence

99m

Tc-MIBI, differentiated thyroid

Department of Nuclear Medicine, Ankara University Faculty of Medicine, Turkey. Correspondence to Dr Ozlem N. Ku¨c¸u¨k, Ankara University Medical School, Nuclear Medicine Department, 06100, Cebeci, Ankara, Turkey. Tel: + 0090 312 562 01 98; fax: + 0090 312 362 08 97; e-mail: [email protected] Received 20 June 2006 Accepted 8 August 2006

serum Tg in ATG-positive patients has the potential to mask the presence of recurrent or persistent thyroid carcinoma [6]. ATGs are present in 10–25% of the patients treated for DTC, invalidating thyroglobulin determination [7–9]. Those patients who undergo total thyroidectomy and ablation treatment should show reduced production of these antibodies, due to a lack of the antigenic stimulus. Thus, it can be suggested that the presence of antithyroglobulin antibodies may be used to detect early relapses and recurrent disease. A few reports have shown that among ATG-positive patients, about 20–30% have proven recurrent DTC [10,11]. This study investigates

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874 Nuclear Medicine Communications 2006, Vol 27 No 11

whether elevated levels of serum ATG reflect the recurrence of cancer in patients with DTC in whom Tg is undetectable after radioiodine ablation. A 99mTc-sestamibi (99mTc-MIBI) scan is commonly used to evaluate DTC patients and is especially useful in patients with high Tg levels and normal 131I WBS [12–14]. This study assesses the sensitivity of disease detection for 99mTc-MIBI WBSs in these patients and investigates the correlation between MIBI WBS results and high serum ATG levels.

graphy, histopathological evaluation and evolution of disease during follow-up. Findings were classified as true positive (TP) or false positive (FP) when the positive 99m Tc-MIBI scan was confirmed or not confirmed by another diagnostic method (bone scans, chest X-rays, CT, ultrasonography and histopathological evaluation). A true negative (TN) or false negative (FN) scan was defined when the negative 99mTc-MIBI scan was accompanied by the absence or presence, respectively, of active thyroid cancer evaluated by other diagnostic modes.

Results Patients, materials and methods Patients

One hundred and ninety patients with histologically confirmed DTC were admitted to the Nuclear Medicine Department of Ankara University Faculty of Medicine between 2000 and 2005. All patients were investigated retrospectively and had initially undergone total or subtotal thyroidectomy, followed by ablation of the residual thyroid remnant by 131I therapy at 1–3 months after the operation. Fourteen patients (13 women and 1 man; mean age 44 ± 19 years, range, 28–71 years) with DTC, had undetectable Tg levels ( < 1.6 ng
ml – 1) and high serum ATG concentrations ( > 40 IU
ml – 1) even with stimulated TSH levels. According to histopathological findings, 10 patients were diagnosed as having papillary and four patients as having follicular cell carcinoma. The mean follow-up period was 13 months (range, 6–26 months) after the Tg and ATG levels had been measured. Laboratory tests

Serum Tg and ATG concentrations were determined by using an immunoradiometric assay that is available commercially in kit form (IMMULITE 2000 TG kit/ L2KTY2; IMMULITE 2000 Anti-TG Ab kit/ L2KTG:Euro/DPC United Kingdom). Scintigraphic techniques

For the scintigraphic examination all patients undergoing thyroid hormone treatment were examined for metastatic sites using 99mTc-MIBI scans, which were performed 20–30 min and 2 h after intravenous injection of 500 MBq 99m Tc-sestamibi (anterior/posterior and static 10 min images were acquired). A Siemens E.CAM SPECT gamma camera with a low-energy, high-resolution, parallel-hole collimator (20% window, 140 keV peak energy) was used. Scans were visually evaluated for detecting lymph node metastases and/or local recurrence, lung metastases and skeletal metastases. The final evaluation for the presence or absence of recurrence was made with other diagnostic methods, including 131I WBS, post-ablative 131I WBS, bone scans, chest X-rays, computerized tomography (CT), ultrasono-

The clinical, pathological and treatment-related characteristics are listed in Table 1. In all patients, serum thyroglobulin concentrations were undetectable ( < 1.6 ng
ml – 1) while ATG levels were > 40 IU
ml – 1 (range, 62–2000 IU
ml – 1) at the beginning of therapy. In 12 of these 14 patients (86%) with high serum ATG levels, recurrent and/or persistent disease was detected, which was confirmed pathologically in four patients and with other diagnostic methods (bone scans, computerized tomography, ultrasonography) and clinical findings in the other eight. Results of 99mTc-MIBI scanning and 131I WBSs were evaluated for three disease categories: neck (local tumour and/or local lymph node metastases), chest (pulmonary metastases) and bone metastases. Table 2 summarizes the scintigraphic results for the 12 patients in whom disease was confirmed by imaging methods, surgery or adequate follow-up. Eighteen lesions were detected in these 12 patients. Twelve (66.7%) of these lesions (detected in 12 patients) were demonstrated on a 99m Tc-MIBI scan. For the six lesions that were negative on a 99mTc-MIBI scan (FN results), a bone-scan showed bone metastases in one patient (in the sacral region), and neck CT showed small cervical lymph nodes in five patients. 131

I WBS demonstrated 10 of the 18 lesions in 12 patients (55.6%). In eight lesions with negative 131I WBS findings (FN results), a bone scan showed bone metastases in one patient (in the sacral region), chest CT showed lung metastases in two patients, and neck CT showed small cervical lymph nodes in five patients. The sensitivity of disease detection for MIBI WBS was calculated as 66.7%; the specificity was 100%. For 131I WBS, the sensitivity of disease detection was 55.6%. Among 12 patients with recurrent and/or persistent disease, 10 showed responses to treatment: three patients underwent surgery, seven received radioiodine treatment, and two had surgery + radioiodine treatment. Five patients had surgery and four had positive and one had negative pathology. ATG levels decreased in eight of

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Importance of anti-Tg auto-antibodies in thyroid carcinoma Ku¨c¸u¨k et al. 875

Clinical, histopathological and treatment-related characteristics of 14 patients with differentiated thyroid carcinoma and high serum anti-thyroglobulin antibody levels

Table 1

Patient number

ATG level (IU
ml – 1) Site of recurrence

Age (years)/ gender

Histology

1

45/F

Fol

1695

2 3

68/F 29/F

Pap Pap

175 62

4 5 6

28/F 30/F 49/F

Pap Pap Pap

2000 91 150

7 8 9 10 11 12

43/M 49/F 30/F 38/F 59/F 71/F

Pap Pap Fol Pap Pap Fol

654 939 63 640 195 725

13 14

58/F 28/F

Fol Pap

86 69

Neck Lung Bone Neck Neck Lung Neck Neck Neck Bone Neck Neck – Neck Neck Neck Lung – Neck Lung

131

I WBS

MIBI scan

N N N N N N N P N P P P – P P P P N P P

P P P P P P P N N N N N N P P N P N P P

Treatment

ATG level after therapy (IU
ml – 1)

131

Decreased

131

Decreased Decreased

I Tx

I Tx I Tx

131

Surgery 131 I Tx 131 I Tx

Decreased Decreased Decreased

Surgery + 131I Tx Surgery Follow-up Surgery + 131I Tx Surgery 131 I Tx

Persistent Decreased Persistent Decreased Decreased Persistent

Follow-up 131 I Tx

Persistent Decreased

F: female; M: male; Pap: papillary; Fol: follicular thyroid carcinoma; ATG: anti-thyroglobulin antibody; WBS: whole-body scan; 131I Tx: radioiodine treatment; P: positive; N: negative. Table 2

Results of MIBI and

Result True positive True negative False positive False negative

131

I whole-body scanning in respect of local disease, pulmonary and bone metastases

Neck MIBI

Lung MIBI

Bone MIBI

7 1 0 5

4 0 0 0

1 0 0 1

10 patients but remained persistently elevated in two patients despite treatment.

Discussion Serum Tg is the most sensitive modality for the detection of recurrent or persistent DTC and is routinely used during the follow-up of patients with the disease [1–4]. However, the presence of ATG may interfere with Tg concentration and cause false negative results [5,6]. Previous studies showed that ATG was higher in patients with DTC than in the normal population [8–11,15]. In such patients ATG may interfere with Tg and cause an underestimation of serum Tg levels and might mask the presence of recurrence or persistence of disease [8]. A few reports have also shown that 20–30% of patients with high serum ATG levels had recurrence [10,11,15]. It was concluded that ATG could be used as a marker of the presence of recurrence of disease in DTC patients. However, there is no general concensus for using ATG measurements to complement Tg recovery test. In our department we routinely perform ATG measurements for 5 years because we think it may be an important marker for follow-up of these patients. We observed high ATG levels in 14/190 patients (7.4%). This is a finding that is comparable with recent studies. Elevated ATG levels may be a sign of recurrence: in these cases the Tg level is low in spite of the presence of

Neck 7 0 1 5

131

I

Lung 2 0 0 2

131

I

Bone

131

I

1 0 0 1

recurrence of DTC. On the other hand, ATG levels may be persistently elevated in some patients without evidence of recurrence, as shown by other diagnostic modalities. A possible explanation is that if very small remnants are present ATG levels may be associated with autoimmune thyroid disease, and lymphocytic memory cells continue to produce antibody for a long time [8–11]. Spencer et al. [8] reported that most surgically cured patients lost their ATG positivity during follow-up. Powell and Harmer [16] considered that there might be microfoci or recurrent disease that was not shown with other diagnostic modalities and therefore the patients were considered to be free of disease. In our retrospective analysis, recurrence was detected in 12/14 patients: two patients had no pathological findings with other diagnostic modalities and 12 with recurrence were treated. Ten of those 12 patients showed response to therapy and ATG levels decreased, while two patients did not respond to therapy. Two patients with high ATG levels had no pathological findings with other diagnostic modalities. They had no autoimmune thyroiditis before the total thyroidectomy and we thought that the cause of these elevated ATG levels would be diminished in the follow-up period. It might be explained by the presence of lymphocytic memory cells. Spencer et al. [8] also reported most patients lost their ATG positivity during the

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876 Nuclear Medicine Communications 2006, Vol 27 No 11

follow-up. Although the follow-up period was short, we accepted these patients as disease-free they are in follow-up.

References 1

2 99m

Tc-MIBI is commonly used to evaluate DTC patients. The plasma and mitochondrial membrane potential of tumour cells and the cellular mitochondrial content are important factors in tumoral uptake of this agent [12–14]. It is especially useful in patients with high Tg levels and normal 131I WBS. Obviously, in these cases, 131I cannot accumulate in tumour tissue but the metabolic cellular activity remains elevated and retains the ability to synthesize and secrete Tg [13,14]. In this analysis, we thought that 99mTc-MIBI scans could be more helpful than 131I WBSs for detecting foci secreting Tg and we compared the results. The sensitivity of disease detection for the MIBI scan was calculated as 66.7%, specificity 100%, positive predictive value 100% and negative predictive value 14.3%. On the other hand, the sensitivity of 131I WBS was calculated as 55.6%, and positive predictive value 90.9%.

3 4

5 6

7

8

9

10 11

Conclusion Persistently elevated anti-thyroglobulin levels appear to serve as a useful marker for recurrent or persistent DTC in patients with undetectable serum Tg results.Thus, the routine measurement of ATG in such a patient population is of great value. Also, in these patients, 99mTc-MIBI has a relatively high sensitivity in the diagnosis of thyroid cancer recurrence or metastases. It may be an additive effect to 131I WBS. So, 99m Tc-MIBI can be used in cases of elevated ATG levels but undetectable serum Tg levels for examination of recurrence or metastases, in addition to other diagnostic methods.

12

13

14

15 16

Schlumberg MJ. Diagnostic follow-up of well differentiated thyroid carcinoma: historical perspective and current status. J Endocrinol Invest 1999; 22:3–7. Mazzaferri EL, Kloos RT. Using recombinant human TSH in the management of well differentiated thyroid cancer: current strategies and future directions. Thyroid 2000; 10:767–778. Herle AJ, Uller RP. Elevated serum thyroglobulin. A marker of metastases in differentiated thyroid carcinomas. J Clin Invest 1975; 56:272–277. Black EG, Cassoni A, Gimlette TM, Harmer CL, Maisey MN, Oafer GD, Hoffenberg R. Serum thyroglobulin in thyroid cancer. Lancet 1981; 2: 443–445. Spencer CA, Takeuchi M, Kazarosyan M. Current status and performance goals for serum thyroglobulin assays. Clin Chem 1996; 42:164–173. Ligabue A, Poggioli MC, Zacchini A. Interference of specific autoantibodies in the assessment of serum thyroglobulin. J Nucl Biol Med 1993; 37: 273–279. Kumar A, Shah DH, Shrihari U, Dandekar SR, Viyojan U, Sharma SM. Significance of anti-Tg autoantibodies in differentiated thyroid carcinoma. Thyroid 1994; 4:199–202. Spencer CA, Takeuchi M, Kazarosyan M, Wang CC, Guttler RB, Singer PA, et al. Serum thyroglobulin autoantibodies: Prevalence, influence on serum thyroglobulin measurement, and prognostic significance in patients with differentiated thyroid carcinoma. J Clin Endocrinol Metab 1998; 83: 1121–1127. Pacini F, Mariotti S. Thyroid autoantibodies in thyroid cancer: incidence and relationship with tumour outcome. Acta Endocrinol 1988; 119: 373–380. Rubello D, Casara D. Clinical meaning of circulating ATG in differentiated thyroid cancer: a prospective study. J Nucl Med 1992; 33:1478–1480. Rubello D, Girelli ME, Casaro D, Piccolo M, Perin A, Busnardo B. Usefulness of the combined antithyroglobulin antibodies and thyroglobulin assay in the follow-up of patients with differentiated thyroid cancer. J Endocrinol Invest 1990; 13:737–742. Larbre H, Schvartz C, Schneider N, Delcourt AC, Maes B, Pochart JM, Vaudrey C. Positive antithyroglobulin antibodies in patients with differentiated thyroid carcinoma. What significance? Ann Endocrinol (Paris) 2000; 61:422–427. Iwata M, Kasagi K, Misaki T, Matsumoto K, Lida Y, Ishimori T, et al. Comparison of whole-body 18F-FDG PET, 99mTc-MIBI SPET, and posttherapeutic 131I-Na scintigraphy in the detection of metastatic thyroid cancer. Eur J Nucl Med Mol Imaging 2004; 31:491–498. Rubello D, Mazzarotto R, Casara D. The role of technetium-99m methoxyisobutylisonitrile scintigraphy in the planning of therapy and followup of patients with differentiated thyroid carcinoma after surgery. Eur J Nucl Med 2000; 27:431–440. Hjiyiannakis P, Mundy J, Harmer C. Thyroglobulin antibodies in differentiated thyroid cancer. Clin Oncol 1999; 11:240–244. Powell S, Harmer C. Thyroid cancer causing death after 40 years: rationale for initial intensive treatment. Eur J Surg Oncol 1990; 19:457–461.

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Original article

A report on the incidence of intestinal 99mTc-methylene diphosphonate uptake of bone scans and a review of the literature ¨ . Kiratli, Emel C. Gu¨nay and Belkis Erbas¸ Eser L. Ergu¨n, Pinar O Background In addition to well-known specific conditions for soft-tissue uptake of bone-seeking radiotracers, there is a limited number of reports on intestinal uptake of 99mTc-methylene diphosphonate (99mTc-MDP) on bone scans. Aim To describe the incidence of intestinal accumulation of 99m Tc-MDP on bone scans in adult patients, define the patterns of this unusual finding and review the literature on its causes. Methods Two thousand, one hundred and forty-four consecutive patients have been evaluated for intestinal 99m Tc-MDP uptake on bone scans. Intestinal uptake was observed visually 3–4 h after the administration of the radiopharmaceutical. A whole-body bone scan and various spot views of the abdomino-pelvic region were obtained with a dual-headed gamma camera to evaluate the intestinal uptake. Delayed scans were also obtained as well as co-relative imaging and/or colonoscopic studies in some of intestinal uptake patients. Six patients had delayed scans of the abdomino-pelvic region. Fourteen patients had comparable scans either a year before or a year later. The positive intestinal uptake scans were further grouped according to the localization and intensity (mild uptake: lower than iliac bone; moderate uptake: equal to iliac bone; significant uptake: higher than iliac bone). Results Twenty-two (17 female, five male) patients out of 2144 with a mean age of 57 years showed intestinal 99m Tc-MDP uptake. The localization was mainly (20/22) in the right abdomino-pelvic region projecting on and in the configuration of ascending colon while one patient showed intestinal uptake all over the abdomen and one displayed

Introduction A bone scan is an important imaging modality to evaluate skeletal pathological conditions. Normally, uptake of bone-seeking tracers is seen in the osseous structures and to a lesser degree in the kidneys and bladder. However, extra-skeletal uptake in soft-tissue structures on bone scans have been reported occasionally [1–3]. The pathogenesis of this finding is usually multifactorial and in some instances the exact mechanism of soft-tissue uptake is not apparent. One of the primary underlying factors is excess calcium in the soft tissue and uptake of

diffuse intestinal radioactivity in his right hemithorax. The majority of the cases showed moderate to intense intestinal uptake (18/22). Six patients showed a decrease, disappearance or alteration in the intestinal uptake on the delayed images. Re-evaluation bone scans in five patients 1 year later showed no intestinal uptake this time. Among nine patients with prior bone scans 1 year before, intestinal uptake was negative in seven at that time. No significant pathology was obtained on the correlative images. Conclusion 99mTc-MDP uptake can be observed in the intestines in 1% of bone scans with a prominent localization in the ascending colon and rarely all over the intestines or in thorax due to Chilaiditi’s syndrome, as well. The mechanism of intestinal uptake is still unclear in some of the patients. Delayed imaging, additional spot views and SPECT studies help in the differentiation of this finding from possible misinterpretation. Intestinal 99mTc-MDP uptake on bone scan could be an intermittent process and should be included among other well-known reasons of soft-tissue uptake. Nucl Med Commun 27:877–885

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:877–885 Keywords: intestinal MDP uptake, bone scan,

99m

Tc-MDP

Department of Nuclear Medicine, School of Medicine, Hacettepe University, Ankara, Turkey. Correspondence to Dr Eser Lay Ergu¨n, Department of Nuclear Medicine, School of Medicine, Hacettepe University, 06100 Sihhiye, Ankara, Turkey. Tel: 0090 312 3051336; fax: 0090 312 3093508; e-mail: [email protected] Received 18 April 2006 Accepted 6 July 2006

bone-seeking agents in soft tissue is believed to be due to chemisorption of 99mTc-methylene diphosphonate (99mTc-MDP) on the surface of calcium salts, including hydroxyapatite crystals [4,5]. Mechanisms of increased soft-tissue calcium deposition [6–8] may be listed as follows: 1. Dystrophic calcification in which loss of intracellular calcium and phosphate from injured cells to the extracellular compartment, precipitation of calcium phosphate salts and subsequent 99mTc-bisphosphonate

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878 Nuclear Medicine Communications 2006, Vol 27 No 11

2.

3. 4. 5.

binding occurs (e.g., necrosis, hypoxia, tumour, amyloid) Metastatic calcification because of increased calcium phosphate produced systemically or locally (lung, stomach, kidney) Increased ectopic osteoblastic activity Metastases from osteoid-forming primary tumours (e.g., osteogenic sarcoma) Increase of calcium binding tissue cations (e.g., iron, magnesium)

Table 1

Causes of soft-tissue uptake

Organ or parameter

Cause of uptake

Brain

Infarcts Metastases Haematomas Abscess Meningiomas Hypercalcaemia Malignant effusions Metastases Primary lung tumours Infarct Ischaemia Pericarditis Amyloidosis Electrical injury Contusion Normal Carcinoma Fibrocystic disease Metastases Colon, neuroblastoma, breast, lung carcinoma Colloid formation because of high aluminium/problems in 99m Tc-bisphosphonate preparation Amyloidosis Necrosis Hypercalcaemia Free pertechnetate Gastric carcinoma Hypercalcaemia Hydronephrosis Obstuction Nephrotoxic agents Radiation therapy Tumours Sickle cell disease Haemosiderosis Myositis ossificans Rhabdomyolysis Muscular dystrophies Thermal or electrical injury Ischaemia/infarct Polymyositis Heterotopic bone formation Osteosarcoma Neuroblastoma Colon carcinoma Breast carcinoma Lung carcinoma etc. Pseudoxanthoma elasticum Misinterpretation Injection site Urine contamination Uptake from a previous study Improper compounded radiopharmaceuticals/abnormal distribitution in the body Errors in instrument calibration

Lungs, pleura

Heart

Breast

Liver

Stomach

Kidneys

Spleen Muscles

Tumours

Skin Scan artifacts

A detailed list of the well-known causes in soft-tissue uptake [1–8] is given in Table 1. Therefore, the recognition of specific conditions with extra-skeletal accumulation of bone-seeking tracers is important because it greatly enhances the diagnostic value of the study. An additional unusual extra-skeletal uptake in bone scintigraphy is intestinal accumulation of bone-seeking tracers and a few cases have been reported [9–14]. This study was undertaken to investigate the frequency of intestinal accumulation of 99mTc-MDP on bone scans, describe the patterns of this unexpected extra-skeletal 99m Tc-MDP uptake and discuss the possible mechanisms of this unusual finding.

Materials and methods During a period of 18 months, 2144 consecutive patients in our department have been prospectively evaluated for intestinal 99mTc-MDP uptake on bone scans. The patients were diagnosed with various types of cancer and referred to the nuclear medicine department for the evaluation of skeletal metastases. After quality control studies of the 99mTc-MDP (Medrocis, CIS, France) to be used, each patient received 740 MBq of the radiopharmaceutical intravenously. The patients were told to drink sufficient water to maintain proper hydration and were advised to void frequently until the scanning time to reduce the radiation dose to the bladder wall. Three to 4 h after the administration of the radiopharmaceutical, each patient voided immediately prior to the scintigraphic study for the elimination of bladder radioactivity for a perfect abdomino-pelvic image. Bone scans were obtained with a dual-headed gamma camera equipped with a high resolution, parallelhole collimator. The imaging protocol utilized a wholebody survey (256  1024 matrix; scan speed: 10 cm
min – 1) followed by high-resolution, high count (a 256  256 matrix, 750 kcounts/view) supplementary spot views of suspicious or symptomatic areas. Anterior, posterior and oblique spot views of the abdomino-pelvic region were also obtained in each patient to examine the intestinal uptake. Only one patient had an additional single photon emission computed tomography (SPECT) study of the abdomino-pelvic region. Besides the evaluation for skeletal metastases, each bone scan was visually assessed for the intestinal 99mTc-MDP uptake. The positive intestinal uptake scans were further grouped according to localization and intensity of the intestinal radiopharmaceutical uptake. The intensity of the intestinal uptake was visually scored as mild uptake (lower than iliac bone), moderate uptake (equal to iliac bone), and significant uptake (higher than iliac bone). Six

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Intestinal

99m

Tc-MDP uptake and literature review Ergu¨n et al. 879

patients underwent delayed abdomino-pelvic imaging (four patients at 24 h and two patients at 6 h). Additionally, five patients had re-evaluation with a bone scan 1 year later and nine patients had prior bone scans 1 year before.

Results

Apart from bone scans, abdominal ultrasonography (USG), computed tomography (CT) and colonoscopy results of the patients were also added to the investigation.

In 20 of 22 patients (91%) intestinal 99mTc-MDP uptake was found in the right abdomino-pelvic region projecting on and in the configuration of ascending colon (Figs 1–3). The abdominal spot views and the SPECT study (patient

Table 2 Patient number

Twenty-two (17 female, five male; mean age: 57 years; range: 43–74) of 2144 patients (1%) had intestinal 99mTcMDP uptake. The features of the scintigraphic studies and diagnosis of these patients are given in Table 2.

Summary of scan results and diagnosis of the patients Age (years)

Gender

Localization and intensity of the intestinal uptake

Prior bone scan

Follow-up bone scan

Primary diagnosis

1

65

Male

No intestinal uptake

NP

Prostate carcinoma

2

53

Female

Right abdomino-pelvic–anterior sacro-iliac region Mild uptake Right sacro-iliac region

NP

1 year later

Nasopharyngeal carcinoma

3

57

Female

4

66

Male

5

52

Female

6

60

Female

7

67

Female

8

49

Female

9

61

Female

10

50

Female

11

48

Female

12

51

Female

13

72

Female

14

52

Male

15

58

Female

16

62

Female

17

58

Female

18

55

Male

19

53

Female

20

43

Female

21

53

Female

22

74

Male

Significant uptake Right abdomino-pelvic region Moderate uptake Right abdomino-pelvic–anterior sacro-iliac region Moderate uptake Right abdomino-pelvic region Mild uptake Right abdomino-pelvic region Moderate uptake Right abdomino-pelvic region Significant uptake

NP

No intestinal uptake 1 year later No intestinal uptake NP

Bladder carcinoma

NP

NP

Cervical carcinoma

NP

24-h late image No intestinal uptake 24-h late image Decreased intestinal uptake 1 year later No intestinal uptake 1 year later

Ovarian carcinoma

NP

NP

Right abdomino-pelvic region Positive intestinal uptake Moderate uptake Right abdomino-pelvic–anterior sacro-iliac NP region Moderate uptake Right abdomino-pelvic region Positive intestinal uptake Significant uptake Right abdomino-pelvic region Moderate uptake Right abdomino-pelvic–anterior sacro-iliac region Significant uptake Right abdomino-pelvic region Moderate uptake Right abdomino-pelvic region Mild uptake Right abdomino-pelvic–anterior sacro-iliac region Moderate uptake Abdomino-pelvic region Significant uptake Right abdomino-pelvic–anterior sacro-iliac region Significant uptake Right abdomino-pelvic region Moderate uptake Right abdomino-pelvic region Significant uptake Right abdomino-pelvic–anterior sacro-iliac region Significant uptake Right abdomino-pelvic region Moderate uptake Right hemithorax Mild uptake

Breast carcinoma

Breast carcinoma

Breast carcinoma Breast carcinoma

No intestinal uptake

No intestinal uptake 2-h late image Change in uptake pattern 24-h late image No intestinal uptake 24-h late image

NP

No intestinal uptake NP

Uterus carcinoma

NP

NP

Gastric carcinoma

No

1 year later

Breast carcinoma

NP

No intestinal uptake NP

Breast carcinoma

NP

2-h late image

Gastric carcinoma

NP

Change in uptake pattern NP

Prostate carcinoma

No intestinal uptake

NP

Breast carcinoma

No intestinal uptake

NP

Breast carcinoma

No intestinal uptake

NP

Breast carcinoma

NP

NP

Prostate carcinoma

No intestinal uptake

Breast carcinoma

Breast carcinoma Breast carcinoma

NP: Not performed

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880 Nuclear Medicine Communications 2006, Vol 27 No 11

number 11) in these 20 patients confirmed that 99mTcMDP uptake was in the ascending colon (Figs 4–6).

Fig. 2

In seven of these patients (numbers 1, 4, 9, 12, 15, 17 and 20) intestinal 99mTc-MDP uptake on the right abdominopelvic region extended through the anterior sacroiliac region but the posterior pelvic spot images showed bilaterally symmetric sacroiliac uptake (Fig. 7). Intestinal uptake either disappeared or decreased in four patients (numbers 6, 7, 11 and 12) (Fig. 8) and changed its pattern in two patients (numbers 10 and 17) (Fig. 9) on the delayed images. Among 22 patients, only one showed significant intestinal uptake all over the intestinal abdomino-pelvic region (patient number 16) (Fig. 10) and one patient (number 22) revealed intestinal uptake in his right hemithorax which was confirmed with a consecutive thorax CT (Fig. 11). The intensity of the intestinal 99mTc-MDP uptake was significant in eight patients, moderate in 10 and mild in four (Figs 1–3).

Anterior

Posterior

Moderate intestinal 99mTc-MDP uptake is shown on right abdominopelvic anterior sacroiliac region in patient 9.

Fig. 1

Fig. 3

Anterior

Posterior

Significant intestinal 99mTc-MDP uptake is seen on right abdominopelvic anterior sacroiliac region in patient 17.

Anterior pelvic spot 99mTc-MDP image of patient 1. Mild intestinal 99m Tc-MDP uptake is displayed on right abdomino-pelvic anterior sacroiliac region.

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Intestinal

Fig. 4

99m

Tc-MDP uptake and literature review Ergu¨n et al. 881

Fig. 5

L

Anterior

L

Right lateral

Anterior and right lateral 99mTc-MDP images of patient 10. Right lateral view confirms that intestinal 99mTc-MDP uptake is located anteriorly and in the ascending colon.

Five patients (numbers 2, 3, 8, 9 and 15) had reevaluation with bone scintigraphy 1 year later and no intestinal 99mTc-MDP uptake was noted this time in these patients (Fig. 12). Nine patients (numbers 1, 8, 10, 11, 12, 15, 19, 20 and 21) had prior bone scans 1 year before. No intestinal uptake was observed on the prior bone scans of seven of these patients. On the prior bone scans of two patients (numbers 8 and 10), intestinal uptake was positive. Fourteen of the patients (numbers 1–3, 5, 7, 9–11, 14–16, 18, 19 and 21) had normal abdominal USG at the time of the bone scanning and seven (numbers 2, 3, 5, 9–11 and 15) who had a follow-up abdominal USG at 6 months to 1 year revealed no pathology. Abdominal CT was obtained in seven patients. Two patients (numbers 4 and 6) had abnormal findings (one had left abdominal mass and one had right ovarian mass) on abdominal CT at the time of the bone scan. Five patients had normal abdominal CT findings (numbers 1, 5, 13, 16 and 20) while a follow-up abdominal CT, which was performed in two patients (numbers 5 and 13) after 8 months and 1 year, respectively, also displayed no abnormality. Colonoscopy was performed in four patients. Two of them (numbers 7 and 18) had internal haemorrhoid while the other two had normal findings (numbers 8 and 14). The laboratory findings available for some of the patients who were investigated retrospectively from the electronic database did not show any abnormality that suggested hypercalcaemia and/or hyperthyroidism. All the patients were asked whether they had the diagnosis of hypercalcaemia/hyperthyroidism or inflammatory bowel disease and none reported inflammatory intestinal disease or

Right anterior oblique

Anterior and right anterior oblique 99mTc-MDP images of patient 12. Right anterior oblique view indicates that intestinal 99mTc-MDP uptake is in the ascending colon.

hyperparathyroidism, and no gastrointestinal symptoms or signs were noted.

Discussion Extra-skeletal uptake of 99mTc-MDP in soft-tissue structures has been reported occasionally [1–3]. The pathogenesis of this finding is various and sometimes the exact mechanism remains unknown. The present study has shown that intestinal uptake of Tc-MDP can be observed in 1% of the bone scans and the majority of this intestinal uptake is seen in the

99m

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882

Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 6

54

56

84

85

Coronal

Sagittal SPECT images of patient 11 clearly display the intestinal

99m

Tc-MDP uptake in the ascending colon.

ascending colon. On very rare occasions it may be observed all over the intestinal abdomino-pelvic region or in hemithorax. In seven of our intestinal uptake patients (numbers 1, 4, 9, 12, 15, 17 and 20) intestinal 99mTc-MDP uptake was observed in the right abdomino-pelvic region extended through the anterior sacro-iliac region. The importance of localization is to distinguish this finding from right sacroiliac metastatic bone uptake especially in oncology patients. In our patients, the increased 99mTc-MDP uptake patterns in the right sacro-iliac region were prominent only on the anterior views while the posterior views were bilaterally symmetric and showed normal uptake (Fig. 7). Additionally, intestinal uptake disappeared or showed a change of pattern on delayed images, which helped us to discriminate the intestinal uptake from metastatic sacroiliac bone uptake. So we suggest performing delayed imaging and evaluating the posterior bone scan views carefully for symmetry in patients who show increased 99mTc-MDP uptake on the right sacroiliac region on anterior projection.

According to our observation, the intensity of intestinal 99m Tc-MDP uptake was significant or moderate in most of the patients (82%) and this helps the interpreter to delineate the pattern of the radioactivity and realize that the uptake is intestinal. Clues for intestinal 99mTc-MDP uptake in our study population were disappearance or decreased uptake and change in the pattern of the radioactivity in the intestine on delayed imaging and also depiction of intestinal configuration especially on lateral, oblique spot abdomino-pelvic views and SPECT study (Figs 4–6 and 12). Additionally, the SPECT study shown in Fig. 6 suggests that activity was more likely in the lumen. Additionally, intestinal 99mTc-MDP uptake seems to be an intermittent process because we did not observe any intestinal uptake on the 1 year prior or 1 year later bone scans in 11 of our patients. One of the reasons of this unusal uptake pattern is oral radioactive urine intake and a few cases have been

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Intestinal

Fig. 7

99m

Tc-MDP uptake and literature review Ergu¨n et al. 883

Fig. 9

4th hour image

6th hour late image

Intestinal 99mTc-MDP uptake on the 4-h image changes its pattern on the 6-h late image in patient 17.

Fig. 10

Anterior

Posterior

On the anterior pelvic image, intestinal uptake extends through the right anterior sacroiliac region (thick arrow). However, on the posterior image 99m Tc-MDP uptake is bilaterally symmetrical and normal (thin arrows).

Fig. 8

4th hour image

24th hour late image

Intestinal 99mTc-MDP uptake on the 4-h image (arrow) totally disappears on the 24-h late image.

reported on in this issue [9,12,15]. In these reports, the radioactivity has been reported to be depicted in the stomach and gastrointestinal tract. In our study, we did not observe any accumulation of radioactivity in the stomach and we found 99mTc-MDP uptake in the whole gastrointestinal tract in only one patient (number 16). None of our patients told us that they drank their own urine. In certain countries, ingestion of one’s own urine is thought to be a helpful folk remedy to improve health and treat some diseases. However, there is no such

Arrows show intestinal 99mTc-MDP uptake all over the abdomino-pelvic intestinal region in patient 16.

tradition in Turkey. So we think that the cause of intestinal uptake in our study population is not oral radioactive urine intake. Intestinal activity in two neuroblastoma patients has been reported by McCarthy and Heyman [10]. These authors could not explain the exact mechanism of the intestinal 99m Tc-MDP uptake in their patients but proposed that microscopic tumour involvement at the site of the

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884 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 11

Anterior bone scan

Posterior bone scan

Thorax CT 99m

Black arrows reveal intestinal Tc-MDP uptake in the right hemithorax on anterior and posteror thorax views of bone scan in patient 22. Thorax CT of the same patient show (white arrow) the colon in right hemithorax which is due to Chilaiditi’s syndrome.

Fig. 12

Lee and co-workers [11] have reported a patient with primary intestinal lymphangiectasia who demonstrated abnormal intestinal accumulation of tracer during 99mTcMDP skeletal scintigraphy. They have proposed that a significant portion of MDP binds to plasma proteins in the early stage and hypothesized that interstitally extravasated tracer bound proteins are reabsorbed by local lymphatics, transported retrograde to the intestinal lymphatic system where they are excreted intraluminally via dilated lymphatics. Protein-losing enteropathy has been reported as a cause of bowel visualization on bone scan [13] because of some protein binding of 99m Tc-MDP. Urinary diversion surgical procedures [16], enterovesical fistula [17], gastrointestinal bleeding [18], increased softtissue calcium deposition due to intestinal infarction [19], neonatal necrotizing enterocolitis [20], systemic amyloidosis [21] are reported causes of intestinal accumulation of bone-seeking tracers. Normal intestinal uptake in children has also been displayed [22]. In our study group, USG, CT and colonoscopy studies in our patients did not display any significant intestinal pathology as urinary diversion surgical procedures or that they had protein-losing enteropathy, enterovesical fistula, gastrointestinal bleeding, intestinal infarction, neonatal necrotizing enterocolitis or systemic amyloidosis. Besides, none of the patients were diagnosed with neuroblastoma and involvement of the intestines with metastasis had not been reported with other techniques. In patient 22, we noticed diffuse increased radioactivity in his right hemithorax both on the whole-body and spot thoracic view. Thorax CT of the patient showed that he had Chilaiditi’s syndrome, which is a relatively rare disease and has been reported only sporadically [23,24]. It is a variant of rotation of the colon resulting in interposition of the colon between diaphragm and the liver, with an estimated incidence of less than 3% of the general population.

Causes of 99mTc-MDP uptake in the intestines (with the reference, where appropriate) Table 3

Follow-up bone scan Positive intestinal 99mTc-MDP uptake (arrow) in patient 8 is not observed on the follow-up bone scan 1 year later.

activity or local tumour invasion with extravasation by the neuroblastoma into nearby bowel with normal transit of the isotope through the bowel might be the reason.

Oral radioactive urine intake [9,12,15] Neuroblastoma [10] Primary intestinal lymphangiectasia [11] Protein-losing enteropathy [13] Urinary diversion surgical procedures [16] Enterovesical fistula [17] Gastrointestinal bleeding [18] Increased soft tissue calcium deposition due to intestinal infarction [19] Neonatal necrotizing enterocolitis [20] Systemic amyloidosis [21] Paediatric patients Chilaiditi’s syndrome Possible improper radiopharmaceutical compound Unknown

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Intestinal

Excess biliary excretion due to improper radiopharmaceutical preparation or formation of technetium–aluminium particles because of excess aluminium or an unknown 99mTc complex leading to hepatic uptake and excretion eventually to the intestine may be other possible causes of the intestinal 99mTc-MDP uptake. However, we excluded this possibility in our patients because there was no visualization of the liver in the patients and other bone scans performed on the same day with pertechnetate from the same generator, using the same MDP kit, showed a normal distribution of tracers. In our department, thin-layer chromatography is used to determine the labelling efficiency of the MDP kit, and labelling efficiency is always higher than 98%. Possible causes of intestinal uptake on bone scans including our data can be summarized in a detailed list given in Table 3. In conclusion, intestinal 99mTc-MDP uptake can be observed in 1% of bone scans and usually localizes in the ascending colon and, on rare occasions, all over the intestinal abdomino-pelvic region. It might also be observed in the thorax as a result of Chilaiditi’s syndrome. Although several causes have been reported for this unusual uptake pattern, the mechanism is still unclear in some of the patients. Delayed imaging (2 h or 24 h) and depiction of disappearance or change in intestinal uptake pattern during a period of time, taking lateral and oblique spot views, performing a SPECT study, evaluating the posterior symmetry of the 99mTc-MDP uptake especially in the sacro-iliac regions, determining the pattern and configuration of the uptake during the interpretation help to recognize the intestinal uptake and avoid false positives. Intestinal 99mTc-MDP uptake on a bone scan could be an intermittent process in a patient and should be included among other well-known reasons of softtissue uptake.

4

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6 7

8

9

10 11

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13 14

15 16

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References 1

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Silberstein EB, Elgazzar AH, Ulloa MF, Nishiyama H. Skeletal scintigraphy in non-osseous disorders. In: Henkin RE, Boles MA, Karesh SM, et al. (editors): Nuclear Medicine. St Louis: Mosby; 1996, pp. 1141–1197. Palmer ED, Scott JA, Strauss HW. Bone imaging. In: Bralow L, Scott JA, Strauss HW (editors): Practical Nuclear Medicine. Philadelphia: WB Saunders; 1992, pp. 121–183. Mettler FA, Guiberteau MJ. Skelatal system. In: Mettler FA (editor): Essentials of Nuclear Medicine. Philadelphia: WB Saunders; 1991, pp. 209–236.

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Freed JH, Hahn H, Menter R, Dillon T. The use of the three-phase bone scan in the early diagnosis of heterotopic ossification (HO) and in the evaluation of Didronel therapy. Paraplegia 1982; 20:208–216. Freed JH, Hahn H, Menter R. The use of three phase bone scan in the early diagnosis heterotopic ossification (HO) and in the evaluation of Didronel therapy. Proceedings of the Seventh Annual Scientific Meeting of the American Spinal Injury Association. XXXXX 1981:63–64. Gentili A, Miron SD, Bellon EM. Nonosseous accumulation of bone-seeking radiopharmaceuticals. Radiographics 1990; 10:871–881. Silberstein EB, DeLong S, Cline J. Tc-99m diphosphonate and sulfur colloid uptake by the spleen in sickle disease: interrelationship and clinical correlates: concise communication. J Nucl Med 1984; 25:1300–1303. Prakash V, Lin MS, Perkash I. Detection of heterotopic calcification with 99m Tc-pyrophosphate in spinal cord injury patients. Clin Nucl Med 1978; 3:167–169. Weismuller S, Baum RP, Zoller A, Hor G. Intestinal accumulation of 99mTcHMDP: an unusual finding in bone scintigraphy. Nuklearmedizin 1994; 33:142–144. McCarthy KE, Heyman S. Diphosphonate intestinal activity seen on two bone images in neuroblastoma. Clin Nucl Med 1986; 11:501–502. Lee KH, Chung JK, Lee DS, Lee MC, Song IS, Koh CS. Intestinal leakage of technetium-99m-MDP in primary intestinal lymphangiectasia. J Nucl Med 1996; 37:639–641. Kosuda S, Katagiri S, Ka WJ, Tominaga S, Kusano S. Demonstration of the ascending colon on Tc-99m MDP skeletal imaging: pitfall in bone scanning by a faith cure of drinking urine. Clin Nucl Med 2000; 25:1040–1041. Roach PJ, Itrato D, Treves ST. Bowel visualization on bone scan because of protein losing enteropathy. Clin Nucl Med 1994; 19:1114–1116. Wang YF, Cherng SC, Cheng CY, Shen HY, Huang WS. Colon visualization of bone scan: a special and interesting case. Clin Nucl Med 1998; 23:723–724. Tsai SC, Kao CH, Lin WY, Wang SJ. Intestinal accumulation of Tc 99m MDP on bone scan. Semin Nucl Med 1999; 29:80–81. Mariani G, Levorato D, Tuoni M, Giannotti P. Incidental imaging of the large bowel in patients with uretero-sigmoidostomy during bone scintigraphy with 99m Tc-pyrophosphate. J Nucl Med Allied Sci 1978; 22:153–157. Suga K, Ohono Y, Yoneshiro S, Fujita T, Nakanishi T, Utsumi H, et al. A case of squamous cell carcinoma of urinary bladder with an ileo-vesical fistula detected on bone scintigraphy. Kaku Igaku 1992; 29:105–110. Lee VW, Leiter BE, Weitzman F, Shapiro JH. Occult gastric bleeding demonstrated by bone scan and Tc-99m-DTPA renal scan. Clin Nucl Med 1981; 6:470–473. Barth KH, Alderson PO, Strandberg JD, Strauss HW, White Jr RI. 99mTcpyrophosphate imaging in experimental mesenteric infarction: relationship of tracer uptake to the degree of ischemic injury. Radiology 1978; 129:491–495. Safakianakis G, Ortiz VN, Haase GM, et al. 99m Tc-diphosphonate abdominal imaging in necrotizing enterocolitis: a prospective study. J Nucl Med 1978; 19:691–692. Janssen S, van Rijswijk MH, Piers DA, de Jong GM. Soft-tissue uptake of 99m Tc-diphosphonate in systemic AL amyloidosis. Eur J Nucl Med 1984; 9:538–541. Conway JJ, Weiss SC, Khentigan J, et al. Gall bladder and bowel localization of bone imaging radiopharmaceuticals [Abstract]. J Nucl Med 1979; 20:622. Chilaiditi D. Zur frage der hepatoptose und ptose im allgemeinen in anschlus an drei falle von temporarer, partieller leberverlgerung. Fortschr Geb Rontgenstr 1911; 16:173–178. Campana L. What is your roentgen diagnosis? Hepatodiaphragmatic interposition of the right colonic flexure, Chilaiditi’s syndrome in dolichocolon. Shweiz Rundsch Med Prax 1992; 16:813–814.

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Original Article

Usefulness of 99mTc-Technegas and ventilatory impairment

133

Xe dynamic SPECT in

Koiku Yokoea, Katashi Satoha, Yuka Yamamotoa, Yoshihiro Nishiyamaa, Hirofumi Asakuraa, Reiji Habab and Motoomi Ohkawaa Aim To assess the usefulness of SPECT images using 99m Tc-Technegas (Technegas) and 133Xe dynamic single photon emission computed tomography (SPECT) (Xe gas) and high-resolution computed tomography (HRCT), as compared with pathological assessment in the detection of small-airway disease including pulmonary emphysema. Methods Seventeen patients with lung cancer were studied. All patients who had undergone both Technegas and Xe gas and CT prior to surgery were examined. SPECT and HRCT results were compared with the results of pathological findings. Histopathological analysis was performed in an area distant from cancer in lobectomy specimens obtained at surgery. Pathological analysis was performed in relation to bronchitis, bronchiolitis, fibrosis of the alveoli and disruption in walls of the alveoli. Results Pathological abnormality (mild-to-moderate abnormal change) was seen in all 17 cases. Three patients showed low attenuation areas on CT, and abnormal patterns in SPECT images. In 11 of 14 patients who showed normal findings on CT, SPECT imaging depicted

abnormal findings. The remaining three patients had no abnormal findings on CT and both SPECT imaging. Conclusion Technegas and 133Xe SPECT imaging is useful for evaluating small-airway disease including pulmonary emphysema. Furthermore, SPECT imaging is more useful than morphological HRCT imaging in the evaluation of small-airway disease including pulmonary emphysema. c 2006 Lippincott Williams Nucl Med Commun 27:887–892 & Wilkins. Nuclear Medicine Communications 2006, 27:887–892 Keywords:

99m

Tc, Technegas,

Correspondence to Dr Koiku Yokoe, Department of Radiology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793, Japan. Tel: + 0081 87 891 2219; fax: + 0081 87 891 2220; e-mail: [email protected] Received 4 June 2006 Accepted 21 July 2006

Methods

Morphologically, computed tomography (CT) of the chest is a routine investigation [1]. Moreover, highresolution CT (HRCT) has been used for the evaluation of detailed structure information such as pulmonary emphysema as low attenuation areas (LAAs).

Patient population

The objective of the present study is to assess the usefulness of SPECT images using Technegas and Xe gas and HRCT, as compared with pathological assessment in the detection of small-airway disease including pulmonary emphysema.

Xe, SPECT, emphysema, ventilation

Departments of aRadiology and bPathology, Faculty of Medicine, Kagawa University, Japan.

Introduction

On the other hand, as functional imaging, recently ultrafine 99mTc-labelled carbon particles are being used for ventilation scintigraphy including single photon emission computed tomography (SPECT) [2–12]. Burch et al. [2] reported that 99mTc-Technegas (Technegas) has a considerably small particle size and can reach the peripheral parts of the lung. 133Xe gas (Xe gas) is also used for dynamic SPECT in addition to planar images [13].

133

Seventeen patients with lung cancer were studied (14 men and three women) with an age range of 51–81 years and mean age of 67.6 years. There are 13 smokers and four non-smokers. All patients who had undergone both Technegas and Xe gas and CT prior to surgery were examined. Imaging acquisition and interpretation Technegas SPECT

Technegas was generated in a proprietary generator (Technegas Generator, Tetley Manufacturing Ltd, Sydney, Australia) by the resistive heating of a graphite crucible up to 25001C in which a saline solution of 505 MBq of [99mTc]pertechnetate had been placed and dried. After generation of the aerosol, it was dispersed in a lead-lined chamber in an atmosphere of 100% argon. Following inhalation of 100% oxygen at 5 l
min – 1 for 3 min, all patients were given 505 MBq 99mTc-Technegas by inhalation in several tidal volume breaths without breath holding through a mouthpiece while wearing a

c 2006 Lippincott Williams & Wilkins 0143-3636

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888 Nuclear Medicine Communications 2006, Vol 27 No 11

nose clip and lying in a supine position. The SPECT system comprised a gamma camera with dual detectors (Picker model Prism 2000, Northford, Connecticut, USA) equipped with a low-energy, high-resolution collimator. Seventy-two images were acquired for 40 s each at 51 intervals. Each image was stored in a 128  128 pixel matrix. Reconstruction of images was performed with no correction for attenuation, using low-pass and ramp filters [10]. 133

Xe dynamic SPECT

Following inhalation of 100% oxygen at 5 l
min – 1 for 3 min, the patients also inhaled 370 MBq 133Xe gas through a mask over the nose and mouth connected to the 133Xe gas ventilation system (Xe-VSS set, Nippon Mediphysics, Osaka, Japan), as a single breath of planar image and held their breath for 15 s. 133Xe dynamic SPECT was performed in the equilibrium phase for the last 1 min of the 3 min breathing in closed circuit, and in the washout phase for 6 min of breathing in semi-closed circuit, using a Picker model Prism 2000 equipped with a low-energy, general, all-purpose parallel collimator. Dynamic SPECT data acquisition by each detector was performed for 1.5 s per projection with 20 views encompassing a 1801 arc. Each detector was rotated in clockwise and counter-clockwise directions across the same projection arc for 30 s. Each image was stored in a 64  64 pixel matrix. Reconstruction of images was performed with no correction for attenuation, using Butterworth and ramp filters. Two sequential 30 s images were added to obtain one 1 min image.

Interpretation of SPECT images

All SPECT images were interpreted by two radiologists. Abnormal findings on Technegas imaging were the extent of heterogeneity, defects and hot spot formation. Abnormal findings on Xe gas imaging were heterogeneity and defect on single breath images and retention images 3 min after washout [14–16]. Computed tomography

HRCT examination was performed with Aquilion (Toshiba, Tokyo, Japan) or HiSpeed Advantage (GE, Wisconsin USA). HRCT images were assessed for LAAs by two radiologists. Interpretation and quantification of pathology findings

Interpretation of pathology findings was performed by one pathologist. Histopathological analysis was performed in an area distant from cancer in lobectomy specimens obtained at surgery. The major site of obstruction is actually found in the smaller conduction airways [17]. Small-airway obstruction was characterized as bronchitis, bronchiolitis and fibrosis of the alveoli. Bronchitis and bronchiolitis were evaluated by infiltration of inflammatory cells. Fibrosis of the alveoli was evaluated by connective tissue deposition. Additionally, disruption in walls of the alveoli was evaluated as emphysematous change. Disruption in walls of the alveoli was classified as normal, mild, moderate and severe based on examination of entire tissue specimen. Representative data are shown in Fig. 1.

Fig. 1

Bronchitis

Bronchiolitis

Fibrotic changes in the alveoli

Disruption in walls of the alveoli

Representative data of pathological findings are shown. The upper row shows the normal findings and the lower row shows the abnormal findings (arrows).

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Usefulness of Technegas and SPECT in ventilatory impairment Yokoe et al. 889

Table 1

Patients’ characteristics, and findings of radiography and pathology

Case number

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17

Age (years)

52 73 81 64 51 74 75 67 78 56 77 70 84 72 72 70 58

Gender

M M M F F M M M M M M M M M M M F

Cigarette index (packs/year)

30 66.3 64 0 25 84 35 40.5 58 12.5 0 50 52 25 30 0 0

Radiographic findings

Pathological findings

CT

Technegas

Xe gas

Bronchitis

Bronchiolitis

Fibrotic change in the alveoli

Disruption in walls of the alveoli

+ – + – – – – – – – – – – + – – –

+ + + – + + + + + + – + + + + + –

– – + – + – + – – + – + – + – – –

+ + + + + + + + + + – + + + + + +

+ + + + + + + + + + + + + + + – +

– + + – – – + – + + – – – + – – –

+++ + +++ + ++ ++ ++ + ++ ++ ++ ++ ++ +++ +++ ++ ++

Radiographic findings and pathological findings (bronchitis, bronchiolitis and fibrotic change in the alveoli) are expressed as follows: + ; abnormal, – ; normal. Disruption in walls of the alveoli is classified as follows: – ; no disruption, + ; mild disruption, + + ; moderate disruption, + + + ; severe disruption.

This analysis was approved by the institutional review board.

Results The results are shown in Table 1. Three of all 17 patients showed LAAs on CT, and abnormal patterns in both SPECT imaging. They had bronchitis, bronchiolitis, fibrosis of the alveoli and disruption in walls of the alveoli. These pathological abnormal findings were more severe than in the remaining 14 patients. In 11 of 14 patients who showed normal findings on CT, SPECT imaging depicted abnormal findings. All had mild-tomoderate bronchitis or bronchiolitis. One of 11 had severe disruption in walls of the alveoli, while the other 10 patients had mild-to-moderate disruption. The remaining three patients had no abnormal findings in CT, Technegas or Xe gas. Mild bronchitis and bronchiolitis were detected in these three patients, and they had mild-to-moderate disruption in walls of the alveoli. Fibrotic changes in the alveoli were detected in only six of all 17 patients. In two of six patients, abnormal findings were depicted on CT and scintigraphy. Three patients, who had no abnormality in radiographic findings, showed no fibrotic changes, and they were also non-smokers. Representative cases are shown in Figs 2 and 3.

Discussion Structural abnormalities, such as emphysema and fibrosis, have been reported in the small airways of smokers [18]. Thin-section CT is currently the best method for assessing morphological changes in emphysema and has been shown to correlate well with pathological grading. However, thin-section CT has limited sensitivity in the detection of early emphysematous changes [19]. Although the spatial resolution of the scintigram in

nuclear medicine is inferior to that of CT, nuclear medicine can depict physiological kinetic movement and assess regional pulmonary function in contrast with overall function [16,20]. We have previously reported the usefulness of Technegas scintigraphy for the diagnosis of pulmonary emphysema [6,21–23]. In mild-to-moderate pulmonary emphysema demonstrated by CT, more detailed findings were shown by Technegas than by CT, and abnormalities were demonstrated by Technegas even in cases without abnormal findings on CT [22]. With respect to recent ventilation studies, some advantages of clinical use of Technegas have been reported and have appropriate energy level [4,5,10]. Previously, the ideal size for aerosol droplets to obtain good uniformity in the lungs during deep tidal volume breathing was considered to be between 0.1 and 0.5 mm [24]. Because particles larger than 2 mm are likely to be deposited in the proximal bronchial trunci, 99mTc-phytate aerosol images were limited due to intense bronchial foci or hot spot formation in cases with severe chronic obstructive pulmonary disease [8]. Technegas is ultra-fine carbon particles of the order of 0.005 mm [2]. Technegas imaging also provides similar or better diagnostic information on lung ventilation imaging in quantitative comparison with Xe gas and 81mKr due to the necessity for advance preparation of the latter two gases and their high cost [4,5]. The fact that no ventilation system is needed in the case of Technegas is another advantage over Xe gas scintigraphy. Technegas SPECT images could better assess pulmonary diseases than planar images [10,11]. A washout study using 133Xe gas is widely used to detect regional ventilation impairment in airway diseases

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890 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 2

(a)

(d)

(b)

Technegas (c)

Xe gas

A 67-year-old man with adenocarcinoma. The findings appeared almost normal on HRCT (a) and Xe gas (c), but there are heterogeneous distributions of Technegas (b; arrow). There are mild disruptions in walls of the alveoli (d; arrow).

Fig. 3

(a)

(d)

(b)

Technegas (c)

Xe gas A 75-year-old man with squamous cell carcinoma. There are no abnormal findings in CT (a). Heterogeneous distribution of Technegas (b; arrow) and retention of 133Xe gas (c; arrow) are detected on the scintigram. Moderate disruptions in walls of the alveoli (d; arrow) and mild fibrotic change (d; arrowhead) are shown pathologically.

[14–16,20,25,26]. The late phases of these regional ventilation studies are helpful in evaluating cases of mild-to-moderate airway obstruction, which would other-

wise be misdiagnosed by radiographs [16,20]. Although Xe gas scintigraphy is sensitive in detecting emphysematous regions due to its retention during washout, as we

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Usefulness of Technegas and SPECT in ventilatory impairment Yokoe et al. 891

previously reported, Technegas SPECT could detect more detailed findings in areas where retention was not seen on 133Xe dynamic SPECT [6].

References 1

2

In the present study, CT could depict emphysematous change as LAAs in a few cases, which have more severe disruption in the walls of the alveoli. On the other hand, Technegas could depict abnormalities in almost all cases. In addition to emphysematous change, bronchitis and bronchiolitis were also evaluated. Although these changes were found to some extent in all cases, there were more severe findings in the cases in which emphysematous changes were shown by CT. Technegas may reflect mild pathological changes that could not be detected by CT. Fibrotic changes in the alveoli were detected in some patients in whom abnormal findings were depicted on CT or scintigraphy. The patients in whom no abnormal findings were depicted on CT or scintigraphy did not have fibrotic changes in the alveoli. Scintigraphy may represent the differences in fibrotic changes.

3

4

5

6

7 8

9

10

In five of 17 cases in the present study, there were abnormalities on Xe gas, and all also presented abnormality on Technegas. On the other hand, there were eight cases with abnormal findings on Technegas, and no abnormality on Xe gas. Technegas is an aerosol consisting of ultra-fine particles, and thus it differs from Xe gas in this respect. This may account for the differences in findings between Technegas SPECT and Xe gas, which may reflect the pathological findings. However, there were no obvious differences in the pathological findings, regardless of whether abnormalities were present on Xe gas or not. In our patient population, three of four non-smokers had no abnormal findings on both CT and scintigraphy. In contrast, pathological abnormal findings were detected in all of the four non-smokers. For reasons of ventilatory impairment, age-related changes [27] and environmental influence including passive smoking exposure, occupational exposures, living environment were reported [28,29]. However, we have no detailed data about the environment to which these four non-smokers were exposed; their changes may reflect various factors such as age-related changes and environmental influence.

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Conclusion Technegas and Xe gas are useful for evaluating the smallairway disease including pulmonary emphysema. Furthermore, SPECT imaging is more useful than morphological HRCT imaging in the evaluation of small-airway disease including pulmonary emphysema.

23

Acknowledgement

24

We thank Dr M. Yoden for her cooperation in some cases.

22

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Original article

Pre-surgical identification of epileptogenic areas in temporal lobe epilepsy by 123I-iomazenil SPECT: A comparison with IMP SPECT and FDG PET Koichiro Kanekoa, Masayuki Sasakib, Takato Moriokac, Hirofumi Kogaa, Koichiro Abea, Hirofumi Sawamotoa, Nobuyoshi Ohyad, Takashi Yoshiuraa, Futoshi Miharaa and Hiroshi Hondaa Objectives The aim of this study was to evaluate the usefulness of 123I-iomazenil (IMZ) single photon emission computed tomography (SPECT) for the pre-surgical identification of epileptogenic areas in patients with temporal lobe epilepsy and to compare the results with those of 123I-IMP SPECT and 18Fluorodeoxyglucose positron emission tomography (FDG PET). Methods We examined seven patients with medically refractory temporal lobe epilepsy (five men and two women; mean age, 28 years) with no remarkable findings on magnetic resonance imaging. Before surgery, IMZ SPECT, IMP SPECT and FDG PET were all performed in the interictal state. Then, visual assessment and region-of-interest (ROI) analysis were performed on each image. Final definitions of the epileptogenic areas were made by electrocorticography and histopathology. Results By IMZ SPECT, a decreased IMZ uptake in the ipsilateral temporal lobe was found in all patients, while a similar decrease in the contralateral temporal lobe was also found in one patient. In comparison to IMP SPECT, the extent of the abnormal area on IMZ SPECT was equal to that on IMP SPECT in one patient while it was more restricted to the epileptogenic area in five patients. In comparison to FDG PET, the extent of the abnormal area on IMZ SPECT was equal to that on FDG PET in three patients

Introduction Patients with medically refractory epileptic seizures can be successfully treated with surgical resection of the epileptogenic area, especially if the zone can be localized to one of the temporal lobes [1]. For this purpose, a careful pre-surgical evaluation to precisely define the zone of ictal onset is necessary in patients with temporal lobe epilepsy (TLE). Scalp electroencephalography (EEG) and video-surface monitoring EEG are the primary laboratory methods of searching for ictal activity, but the success rate of EEG methods ranges from 60 to 90% [2]. Some patients may require an invasive intracranial EEG to determine both the location and the extent of the epileptogenic area. Invasive intracranial EEG has an accompanying risk and is a burden on the patient. Being able to forego these invasive

while it was more restricted in the epileptogenic area in four patients. In ROI analysis, decreases of IMZ, IMP and FDG uptake were observed in the epileptogenic area, although they were not statistically significant. Conclusions IMZ SPECT was considered to be useful for pre-surgical determination of the epileptogenic areas in temporal lobe epilepsy with no remarkable MRI findings, and it was also found to be superior to IMP SPECT and FDG PET for this purpose. Nucl Med Commun 27:893–899

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:893–899 Keywords: IMZ SPECT, CBF SPECT, FDG PET, temporal lobe epilepsy, pre-surgical identification Departments of aClinical Radiology, cNeurosurgery, Graduate School of Medical Sciences, Kyushu University, bLaboratory of Molecular Imaging Sciences, School of Medicine, Kyushu University and dDivision of Radiology, Department of Medical Technology, Kyushu University Hospital, Japan.

Correspondence to Dr Koichiro Kaneko, Department of Clinical Radiology, Graduate School of Medical Sciences, Kyushu University, Maidashi 3-1-1, Higashi-ku, Fukuoka 812-8582, Japan. Tel: + 0081 92 642 5695; fax: + 0081 92 642 5708; e-mail: [email protected]

Received 19 April 2006 Accepted 25 July 2006

examinations would thus be of great benefit for such patients. Among the neuroimaging methods, magnetic resonance imaging (MRI) is a widely available and useful method for identifying structural abnormalities such as hippocampal sclerosis [3], and plays a key role in the evaluation of patients with epilepsy [4]. However, MRI findings have been reported to be normal in 26–30% of patients with TLE [5,6]. Nuclear medicine imaging has been used in non-invasive examinations to determine the epileptogenic area. Evaluation of the cerebral blood flow (CBF) by single photon emission tomography (SPECT) shows the hypoperfused area when performed in the interictal state.

c 2006 Lippincott Williams & Wilkins 0143-3636

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894 Nuclear Medicine Communications 2006, Vol 27 No 11

However, the sensitivity of interictal CBF SPECT is not satisfactory [7]. Ictal and post-ictal CBF SPECT appear to significantly improve the sensitivity [8,9], and recently, a computer-aided subtraction method called subtraction ictal SPECT co-registered to MRI (SISCOM) has been reported to be useful for evaluating the epileptogenic zone [10,11]. However, this method requires two different CBF SPECT images (i.e., ictal and interictal) in addition to MRI, and it is thought to be too cumbersome for routine clinical usage. [18F]Fluorodeoxyglucose (FDG) positron emission tomography (PET) in the interictal state shows decreased glucose metabolism including those in the epileptogenic area and its sensitivity is higher than that of interictal CBF SPECT [12]. However, because the abnormal area it reveals is usually wider and more extended than the epileptogenic area, it is not possible to determine the exact location of the epileptogenic area. Furthermore, the clinical utility of FDG PET is limited by its restricted availability and high cost. A recent autoradiographic study demonstrated that the central-type benzodiazepine receptor (BZR) density was decreased in the resected epileptogenic tissue of patients with focal epilepsies [13]. In another study, 11 C-flumazenil (FMZ) PET was able to detect reduced BZR binding in the epileptogenic focus and thus is expected to be a more specific imaging probe [14]. Decreased FMZ binding in the hippocampus was seen even in TLE patients with normal MRI [15–17]. 123 I-Iomazenil (IMZ) is a radioligand for the central-type BZR, and can be used with SPECT. IMZ SPECT is expected to be useful in the planning for surgical resection by reducing the number of invasive intracranial EEGs. However, there have been only a few studies comparing the efficacy of imaging with IMZ SPECT, CBF SPECT and FDG PET in the same patients [18,19]. The aim of this study was thus to evaluate the usefulness of IMZ SPECT for the pre-surgical identification of epileptogenic areas in patients with temporal lobe epilepsy (TLE) and to compare the results with those of N-isopropyl-p-[123I]iodamphetamine (IMP) SPECT and FDG PET.

Material and methods Patients

This study was approved by the Committee for the Clinical Application of Cyclotron Producing Radionuclides in Kyushu University Hospital, and written informed consent was obtained from all patients before starting the PET study. Between September 1999 and August 2001, we examined seven patients with medically refractory TLE (five men and two women; mean age, 28.0 ± 8.3 years; range, 18–39 years). The clinical data of the patients are summarized in Table 1. Brain MRI was performed with a 1.5 T

Magnetom Vision system (Siemens, Erlangen, Germany) with a standard head coil. Axial T1-weighted, T2weighted and FLAIR images parallel to the long axis of the hippocampus and coronal FLAIR images with 5 mm slice thickness were obtained. On the brain MRI, no remarkable abnormalities were found in any patients upon visual inspection by three radiologists. Before surgery, IMZ SPECT, IMP SPECT and FDG PET were all performed with the patient in the interictal state within 3 weeks. All surgery was guided by electrocorticography. Finally, an anterior temporal lobectomy with hippocampectomy was performed in all seven patients. In patient 3, a resection of the middle temporal gyrus and a multiple subpial transection for lateral cortex was also performed. The histopathological diagnosis was mesial temporal sclerosis in five of the seven patients, mesial temporal sclerosis with a gliosis in the lateral cortex in one patient, and a small focal cortical dysplasia in the lateral cortex in one patient. Postoperative outcome was accessed by using the classification of Engel et al. [20]. All patients except number 3 became seizure free (class I). Patient 3 has residual seizure (class III) because the epileptogenic area on the left lateral cortex of language area was not completely resected. The final definition of the epileptogenic area was medial TLE in five patients, lateral TLE in one patient, and both medial and lateral TLE in one patient, based on ictal or interictal scalp EEG, chronic subdural EEG, electrocorticographic findings and histopathological findings. Protocol for SPECT

IMZ SPECT and IMP SPECT images were obtained by 20 min acquisition beginning at 3 h after the administration of 16 MBq of IMZ and at 20 min after the administration of 222 MBq of IMP. All tracers were injected intravenously with the patients with supine position with their eyes closed. SPECT studies were performed using a three-detector system GCA9300A/HG (Toshiba Corp., Tokyo, Japan) with a fanbeam collimator (128  128 matrix). Each detector was set to rotate continuously through 1201 in 3 min, alternating in both the clockwise and counter-clockwise directions. A Butterworth filter with a cut-off of 0.24 cycle/cm and order of 8, and a ramp filter were used for image reconstruction. Attenuation correction was performed using Sorenson’s method with an attenuation coefficient of 0.12 cm – 1. The spatial resolution in the plane was 7.4 mm of full width at half maximum (FWHM). Protocol for positron emission tomography

PET studies were performed on an ECAT EXACT HR + device (Siemens, Knoxville, Tennessee). FDG (370 MBq) was given intravenously to each patient in the supine

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Clinical data for the subjects in this study

Pt.

Age/gender Type/frequency

Onset

Medication

1

18/F

CPS: 5/D SGS: 3/Y

11Y

2

22/M

CPS: 3/D SGS: 3/D

3

22/M

4

Seizure

Surgery resected area

Seizure outcome*

Pathology

FDG PET

IMP SPECT

IMZ SPECT

PHT CZP CLB

Te, l, med

ATL, l

Class I

HS

Te, l, med + lat

Te, l, med + lat

Te, l, med

14Y

PHT CBZ

Te, l, med

ATL, l

Class I

HS

Te, l, med

Te, b, med + lat

Te, l, med

SPS: 3/Mo CPS: 3/Mo SGS: 0.5/Mo

8Y

CBZ ZSM PHT CLB

Te, l, med + lat

ATL, l CR, l MST, ;l

Class III LG

HS

Te, b, med + lat

Te, b, med

Te, l, med

27/M

SPS: 4/Mo CPS: 2/Mo SGS: 2/Y

18Y

PHT CBZ CLB

Te, l, med

ATL, l

Class I

HS

Te, l, med + lat

Te, l, med + lat

Te, l, med

5

29/M

CPS: 5/Mo SGS: 1/Mo

14Y

CBZ

Te, l, lat

ATL, l

Class I

FCD

Te, l, med + lat

Te, l, med + lat

Te, l, med + lat

6

39/F

CPS: 10/Mo SGS: 2/Mo

16Y

PHT

Te, l, med

ATL, l

Class I

HS

Te, l, med + lat

Te, l, med + lat

Te, l, med + lat

7

39/M

SPS: 6/D CPS: 6/D SGS: 4/Y

10Mo

VPA PHT CZP

Te, l, med

ATL, l

Class I

HS

Te, r, med Te, l, med + lat Fr, l,

Te, r, med Te, l, med + lat Fr, l, Occ, l

Te, r, med Te, l, med + lat Fr, l, Occ, l

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I-iomazenil SPECT Kaneko et al. 895

SPS: simple partial seizure; CPS: complex patial seizure; SGS: secondary generalized seizure; PHT: phenytoin; PB: phenobarbital; CBZ: carbamazepine; AZA: acetazolamide; CZP: clonazepam; DZP: diazepam; ESM: ethosuximide; PRM: primidone; ST: sulthiame; VPA: sodium valproate; ZSM: zonisamide; CLB: clobazam; Te: temporal lobe; Fr: frontal lobe; Occ: occipital lobe; l: left; b: bilateral; r: right; med: medial; lat: lateral; ATL: anterior temporal lobectomy and hippocampectomy; CR: cortical resection of the lateral temporal lobe; MST: multiple subpial transection for lateral temporal lobe; HS: hippocampal sclerosis; FCD: focal cortical dysplasia; LG: lateral temporal gliosis. * Accessed by Engel’s classification.

123

Finally diagnosed epileptogenic area

Identification of epileptogenic areas by

Table 1

896 Nuclear Medicine Communications 2006, Vol 27 No 11

position with their eyes closed. Thirty-five minutes after the completion of FDG administration, a post-injection transmission scan was performed for 5 min. The threedimensional mode emission scan (10 min) was performed immediately after the post-injection transmission scan. Attenuation correction was applied to the post-injection transmission-based segmented attenuation correction method, which has been well correlated with the conventional pre-injection transmission-based attenuation correction method [21]. The images were reconstructed with a filtered back-projection by means of a Hanning filter (cut-off = 0.4 cycle/pixel). The spatial resolution in the plane was 4.2 mm of FWHM. Data analysis

Transaxial, coronal, and longitudinal images of the hippocampus of IMZ SPECT, IMP SPECT and FDG PET were used for the analysis. In IMZ SPECT, we analysed only the late scan images (3 h after IMZ i.v.) that reflect binding potential, which have higher sensitivity and specificity than early images [19]. The abnormalities of IMZ SPECT, IMP SPECT and FDG PET were visually evaluated by a consensus of three nuclear medicine physicians concerning (1) lateralization of the abnormal area, (2) localization of the abnormal area, and (3) explicitness of the abnormal findings. Then, square regions of interest (ROIs) of 6.8  6.8 mm were carefully placed on the medial and lateral temporal lobes on coronal images and bilateral cerebellar cortex on transaxial images with reference to the corresponding MRI. The mean SPECT count of the bilateral cerebellar cortex was calculated and used as the value of the cerebellum. The values of the region-to-cerebellar cortex were used to compare the epileptogenic area with the non-epileptogenic area in both the medial and lateral cortices. Statistical significance was determined by the paired Student’s t-test.

Results Lateralization of the abnormal area

Table 2 shows the lateralization of the abnormal area on each image. By IMZ SPECT, a decreased IMZ uptake in the ipsilateral temporal lobe was found in all patients, and a similar decrease in the contralateral temporal lobe was found in one patient. Abnormalities in the ipsilateral temporal lobe were also observed in all seven patients on both IMP SPECT and FDG PET. However, abnormalities in the contralateral temporal lobe were observed in three Table 2

Lateralization of abnormal area

Ipsilateral Ipsi- and contralateral Contralateral

Localization of the abnormal area

Table 3 shows the localization of the abnormal area on each image. The abnormal region on IMZ SPECT was more restricted to the epileptogenic area than that on IMP SPECT in five patients, and was equal to that on IMP SPECT in one patient. A comparatively more extensive abnormality was observed in one patient on IMZ SPECT. The extent of the abnormal area on IMZ SPECT was equal to that on FDG PET in three patients, and more restricted to the epileptogenic area in four patients. Finally, abnormal findings localized in the epileptogenic area were observed in three patients by IMZ SPECT, zero patients by IMP SPECT, and one patient by FDG PET. On the other hand, abnormal findings located outside the epileptogenic lobe were observed in one patient by IMZ SPECT, three patients by IMP SPECT and two patients by FDG PET. Figure 1 shows results for a patient with left hippocampal sclerosis (patient 6 in Table 1). The epileptogenic area was finally determined to be localized in the left medial temporal lobe, and the patient underwent anterior temporal lobectomy with hippocampectomy. MRI demonstrated minimal atrophy of the left hippocampus. Although FDG PET and IMP SPECT showed widespread decreased uptake in the entire left temporal lobe, the extent of the decreased uptake area on IMZ SPECT was restricted to the left medial temporal lobe. Explicitness of the abnormal findings

Table 4 shows the explicitness of the abnormal findings on each image. The explicitness was determined by overall evaluation of the clinical usefulness of the findings for the pre-surgical determination of the epileptogenic area in comparison with IMP SPECT and FDG PET. The abnormal findings on IMZ SPECT were more clear than those on IMP SPECT in five patients, the two imaging modalities were equally effective in one patient, and the abnormal findings were more clear on IMP SPECT than on IMZ SPECT in one patient. The abnormal findings on IMZ SPECT were more clear than

Table 3

IMZ SPECT

IMP SPECT

FDG PET

6 1

4 3

5 2

0

0

0

Results are given as the number of patients.

patients on IMP SPECT and two patients on FDG PET. Finally, lateralization of the epileptogenic area was definitively diagnosed in six patients by IMZ SPECT, four patients by IMP SPECT and five patients by FDG PET.

Localization of the abnormal area

Restricted to focus Include focus but extensive Outside the focus

IMZ SPECT

IMP SPECT

FDG PET

3 3

0 6

1 6

1

3

2

Results are given as the number of patients.

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Identification of epileptogenic areas by

Table 5

Fig. 1

(b)

(a)

I-iomazenil SPECT Kaneko et al. 897

Comparison of the region-to-cerebellar cortex ratios

Medial Epileptogenic Non-epileptogenic Lateral Epileptogenic Non-epileptogenic

IMZ SPECT

IMP SPECT

FDG PET

0.845 ± 0.090 1.008 ± 0.249

0.660 ± 0.181 0.892 ± 0.316

0.761 ± 0.182 0.945 ± 0.151

1.024 ± 0.238 1.245 ± 0.208

1.125 ± 0.150 0.932 ± 0.300

0.779 ± 0.095 1.135 ± 1.176

(d)

(c)

A 39-year-old female patient with partial seizure due to hippocampal sclerosis in the left medial temporal lobe (patient 6 in Table 1). (a) MRI demonstrated minimal atrophy of the left hippocampus. Although FDG PET (b) and IMP SPECT (c) showed widespread decreased uptake in the entire left temporal lobe, the extent of the decreased uptake area on IMZ SPECT was restricted to the left medial temporal lobe (d).

Table 4

123

Explicitness of abnormal findings IMZ SPECT

vs IMP vs FDG

Superior

Equal

Inferior

5 4

1 3

1 0

Results are given as the number of patients.

those on FDG PET in four patients, while IMZ SPECT and FDG PET yielded similar results in three patients. Analysis of regions of interest

The values of the region-to-cerebellar cortex ratio are shown in Table 5. The ratios in the most epileptogenic area were lower than those in the non-epileptogenic area, although the difference was not statistically significant.

Discussion In this study, IMZ SPECT, IMP SPECT and FDG PET all detected an abnormal area in the ipsilateral temporal lobe, though an additional abnormal area in the contralateral temporal lobe was more frequently seen on both IMP SPECT and FDG PET than on IMZ SPECT. Further, the extent of the abnormal area was more restricted on IMZ SPECT than on IMP SPECT or FDG PET. Overall evaluation of the explicitness of abnormal findings suggested that IMZ SPECT was superior.

In past studies, the sensitivity of interictal CBF SPECT has varied from 48 to 79% [9,22,23]. The meta-analysis sensitivity was reported to be 0.43 for interictal CBF SPECT, and it was not dependent on the tracer used [11]. Although post-ictal and ictal CBF SPECT appeared to significantly improve the sensitivity (0.77 and 1.00, respectively), these modalities are too cumbersome for routine clinical usage. FDG PET in the interictal state is able to visualize an area of decreased glucose metabolism that includes the epileptogenic focus. The sensitivity of FDG PET is higher than that of interictal CBF SPECT, but the abnormal area it visualizes is somewhat larger than that of the epileptogenic focus [12]. FDG PET shows decreased uptake in a wide area including the surrounding brain region, and is not highly accurate for locating the lesion. This is because hypometabolism can also be caused by disturbances other than neural loss [24]. Interictal CBF SPECT provides results similar to those of FDG PET, but with a lower sensitivity and lower specificity [12]. These findings suggest that the hypoperfused or hypometabolic area does not always represent the epileptogenic area; it may also include the surrounding area. In-vitro and in-vivo studies have demonstrated a reduction of BZR in epileptic areas [25,26], and it has been reported that BZR density is well correlated with cell density both in rats [27] and in humans with TLE [28]. Thus the area of BZR abnormality is considered to be smaller than that of decreased metabolism. These findings suggest that IMZ SPECT may be able to distinguish the pathologically affected area from the surrounding brain region. There have been two studies comparing the efficacy of imaging with IMZ SPECT, interictal CBF SPECT, and FDG PET in the same patients, but these studies showed conflicting results [18,19]. Tanaka et al. reported that IMZ SPECT has not only high sensitivity (90%) but also high specificity (80%), while FDG PET has much lower specificity (30%) and much higher sensitivity (100%) [18]. It has also been reported that FMZ PET was more sensitive than FDG PET in patients with TLE with normal MRI, although it was less reliable than FDG PET because of false lateralization [14]. Conversely, Lamusuo et al. reported that FDG PET had higher agreement (77%) with the

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898 Nuclear Medicine Communications 2006, Vol 27 No 11

subdural EEG and operative findings than did IMZ SPECT (55%) [19]. Interictal CBF SPECT had lower sensitivity than IMZ SPECT in these two studies [18,19], and the sensitivity was particularly low in the study by Lamusuo et al. [19]. In our study, IMZ SPECT had the best agreement with the operative findings (86%), followed by FDG PET (71%) and IMP SPECT (57%). Our results suggest that IMZ SPECT was more useful for pre-surgical identification of epileptogenic area than FDG PET and IMP SPECT. These results are in agreement with the study by Tanaka et al. [18]. Two possible explanations for this discrepancy in the detectability of the epileptogenic area by IMZ SPECT and FDG PET are as follows. First, there was a difference in the performance of the SPECT device between our study and that of Lamusuo et al., with our device having a higher spatial resolution than that of Lamusuo et al. (FWMH = 16 mm). This difference of spatial resolution could have resulted in the higher detectability for epileptic focus in our study. Second, it is thought that loss of hippocampal volume on MRI is well correlated with neural loss [24], and thus we suspected that hippocampal neural loss would be mild in TLE patients with normal MRI. In fact, our ROI analysis data showed a decreased IMZ binding potential in the epileptogenic focus, but the difference was not statistically significant (Table 5). These mild abnormal findings might be missed in imaging modalities with low spatial resolution, such as SPECT. Computed statistical imaging analyses, such as those using statistical parametric mapping or three-dimensional stereotactic surface projection, may have the potential to detect decreased IMZ binding more objectively, and further studies will be needed to investigate this possibility. The principal limitation of our study was the small number of subjects, which limited the statistical power of the results. Further examinations with a larger number of patients are required to clarify the usefulness of IMZ SPECT.

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

Conclusion IMZ SPECT was found to be useful for pre-surgical determination of the location of epileptogenic areas in cases of TLE with no remarkable MRI findings, and it was also found to be superior to IMP SPECT and FDG PET for this purpose.

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2

Wiebe S, Blume WT, Girvin JP, Eliasziw M. Effectiveness and Efficiency of Surgery for Temporal Lobe Epilepsy Study Group. A randomized, controlled trial of surgery for temporal lobe epilepsy. N Engl J Med 2001; 345:311–318. Schomer DL. Current concepts in neurology. Partial epilepsy. N Engl J Med 1983; 309:536–539.

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Original article

Characterization of septal penetration in 511 keV SPECT Charles M. Laymon and Timothy G. Turkington Background Single photon emission computed tomography (SPECT) with 511 keV photons is a challenging modality and collimators for this purpose require trade-offs among resolution, sensitivity and septal penetration. While PET is the modality of choice for imaging at 511 keV, there are some procedures, e.g., dual-isotope imaging, in which 511 keV SPECT has a role. Aim To measure the imaging properties of a VPC-93 SPECT collimator designed for imaging at 511 keV and to isolate the effects of septal penetration. Methods NaI gamma camera projection images of 18 F (511 keV) and 99mTc (140 keV) point sources were measured and the corresponding modulation transfer functions calculated. The projection images were reconstructed via filtered back-projection to obtain the tomographic three-dimensional (3-D) point spread function. Differences between the 511 and 140 keV results were attributed mainly to septal penetration. Contrast measurements were made separately using 18F and 99mTc of a 20 cm phantom containing hot spheres and a warm background. Both isotopes were also used in imaging studies of a 3-D Hoffman brain phantom. Results Reconstructed 511 keV point source images were spatially extended with more than half of the total reconstructed counts appearing away from the point source region. The number of false counts contained in the image as a function of distance from the true

Introduction The quality of images acquired in single photon emission computed tomography (SPECT) mode with a modern gamma camera is largely determined by the properties of the collimator used in the acquisition. Both system resolution and sensitivity depend mainly on collimator specifications. An important consideration in collimator design is the degree to which radiation penetrates through the collimator septa. Septal penetration results in the assignment of detected radiation to incorrect lines of response, degrading resolution and causing distortion of images. Muehllehner and Huig [1] discuss aspects of collimator design related to septal penetration. Their work describes calculations of collimator line-spread functions that are most appropriate for energies below 511 keV. They also discuss some earlier work on the subject.

source location remains approximately constant for large distances out to at least 14 cm. Septal penetration results in a rapid roll-off with spatial frequency of collimator response. The response of the collimator to 511 keV photons falls to half of its 0-frequency response at 0.03 cm – 1. For 140 keV photons this value is 0.20 cm – 1. A result is reduced image contrast as measured in the phantom sphere studies. Septal penetration causes image degradation through large-scale blurring. Image noise characteristics are modified and correlations are extended into many transaxial planes. Conclusions Both 2-D and 3-D point spread functions for 511 and 140 keV photons using the VPC-93 collimator have been measured. Septal penetration unfavourably affects image resolution and changes image noise characteristics. Without compensation, the effects of septal penetration are readily apparent in images of real objects. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:901–909
Nuclear Medicine Communications 2006, 27:901–909 Keywords: SPECT, 511 keV imaging, SPECT collimator, image reconstruction Correspondence to Dr Charles M. Laymon, Department of Radiology, University of Pittsburgh, UPMC, Presbyterian B-938 200 Lothrop St., Pittsburgh, PA 15213, USA. Tel: + 001 412 647 0730; fax: + 001 412 647 0700; e-mail: [email protected] Received 4 April 2006 Accepted 25 July 2006

Examples of experimental work relevant to this topic include that of Bolmsjo¨ et al. [2] who measured the response of five collimators to 440 and 529 keV gamma rays from 123I. Their work included a medium-energy, round-holed collimator with septa of 1.8 mm thickness and holes of 7.5 cm length, as well as a pinhole collimator. Their results showed the best frequency response for the pinhole collimator, presumably because of less septal penetration. Clarke et al. [3] compared septal penetration effects at 364 keV (131I) in several collimator designs. They found an appreciable penetration effect on the recovery coefficient measured as a function of object size using a high sensitivity, medium-energy collimator. Little penetration effect was seen in tests using a lowsensitivity medium-energy collimator. Positron-emitting radiotracers, particularly 18F-labelled 2fluoro-2-deoxy-D-glucose (FDG), coupled with positron

c 2006 Lippincott Williams & Wilkins 0143-3636


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emission tomography (PET) have proven valuable for metabolic imaging. The recent increase in the availability of FDG, through commercial ventures, offers previously unavailable opportunities to many nuclear medicine facilities for positron imaging. Because of its high sensitivity and favourable resolution properties, PET, using a dedicated scanner, is generally the preferred method for imaging these tracers. Additionally, dedicated PET scanners are becoming increasingly available. Nevertheless, 511 keV SPECT is a valuable modality in protocols for simultaneous multi-isotope (particularly 99m Tc and 18F) imaging. Such protocols are typically used for the simultaneous measurement of blood flow and metabolic rate and have applications in cardiology [4,5] and neurology [6]. Septal penetration becomes a bigger consideration with increasing photon energy because of the rapidly falling electromagnetic interaction cross sections. To set the scale, the thickness of lead necessary to achieve a particular reduction in gamma flux increases by a factor of about 16 between 140 and 511 keV [7]. Thus, given the practical limits on collimator weights, 511 keV SPECT is a particularly challenging modality. Several authors have discussed aspects of 511 keV SPECT imaging. An introduction to the subject of gamma-camera imaging using positron emitters and a general comparison of the clinical applicability of single and coincidence imaging has been given by Smith and O’Mara [8]. Various studies can be cited which represent a sample of the literature that focuses on the clinical aspects of 511 keV SPECT [9–15]. Patton et al. [16] report on the modifications necessary to transform a traditional SPECT scanner into one capable of imaging at 511 keV. The authors present resolution, sensitivity and contrast data for parallel beam (similar to the one used in this study) and fan-beam collimators. They also discuss oncological, neurological and cardiac applications of 511 keV SPECT. The purpose of this work is to present a quantitative evaluation of septal penetration and its effects on reconstructed images. Because of the interest in FDG imaging coupled with the difficulty in collimating 511 keV photons, we performed these studies using collimators designed for 511 keV imaging. Additional motivation for this study is the possibility that, for a coincidence capable gamma-camera system, attenuation properties can be deduced without a transmission scan from the combination of coincidence and collimated singles data [17]. There are three main parts to this work. Firstly, in the point source experiments and analysis we deduce the fundamental imaging and noise properties of a high-

energy collimator starting with a measurement of its point spread function (PSF). Secondly, in the sphere phantom studies we measure contrast and present images that, in a direct way, show the effects of blurring and modified noise propagation on image quality. Thirdly, we present phantom brain images in which septal penetration effects are more complicated but also of greater clinical interest. Each study reported in this paper was performed at 140 keV and 511 keV using a collimator designed for 511 keV SPECT. This is not the optimal way to image at 140 keV: a comparison of the results serves to isolate the effects of septal penetration. In this respect this work is similar to that of Clarke et al. [3] in which penetration effects at 364 keV were identified by comparing results obtained with 131I (364 keV) and 99mTc (140 keV). Additionally, the present results are directly applicable to protocols in which 99mTc and 18F are simultaneously imaged.

Methods We employed several methods to demonstrate and quantify the effects of septal penetration of 511 keV photons through high-energy collimators. In all cases data were acquired with a two-headed gamma camera (Helix or Varicam, Elscint, Ltd, Haifa, Israel) with 1.3 cm (0.5") thick NaI(Tl) crystals using collimators (Elscint VPC-93) designed specifically for 511 keV imaging. The collimators are of standard hexagonal-hole design. The holes are approximately 4 mm in diameter and 80 mm long. The septal thickness is 2.5 mm. The source of 511 keV photons was either [18F]fluoride or 18F-FDG. Pertechnetate labelled with 99mTc served as the source of 140 keV photons. Point source studies

A goal of this work was to determine the point spread function (PSF) of the collimator to 511 keV photons. Measurements were performed using a point source formed by placing several drops containing 18F into a small cone vial. The vial was suspended in the field of view of the gamma camera via a jig constructed for these experiments that allowed accurate repositioning of the source. Static acquisitions (60 s) were done at source-tocollimator distances of 12 cm and 30 cm, relevant for brain and body imaging. Data were acquired into a 128  128 array with a pixel size of 0.44 cm. The spatial-frequency components of each PSF were determined by taking its Fourier transform. No attempt was made to investigate the dependence of the PSF on position within the plane parallel to the collimator face. The point source activity was about 74 MBq and a 20% energy window centred at 511 keV was used resulting in processed count rates of 9000 and 7000 cps for distances of 12 and 30 cm. The measurements were repeated with a 140 keV point source

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Characterization of septal penetration in 511 keV SPECT Laymon and Turkington

under similar count rate conditions. Data at both energies were acquired with the same high-energy collimator and we assume that the difference between the 511 keV and 140 keV images is dominated by septal penetration effects. Three-dimensional (3-D) PSFs for 18F and 99mTc were produced from the 2-D PSFs. The 2-D PSF was first smoothed with the kernel 2 3 1 1 1 1 1 61 2 2 2 17 6 7 7 K ¼6 61 2 3 2 17 41 2 2 2 15 1 1 1 1 1 and shifted to form a low-noise ‘master’ projection that was centred in the projection frame. This was replicated to form a 120-element set of (identical) projection images, as would be obtained for a source at the centre of the field of view. The master projection set was ‘reconstructed’ via filtered back-projection with ramp filter only. For a particular collimator, the hexagonal holes, and therefore the septal penetration pattern has a specific orientation with respect to the axis of rotation. The process described above for generating 3-D PSFs was applied to rotated versions of the master images corresponding to two different collimator orientations. The 511 and 140 keV master projections were also used to study effects of septal penetration on the propagation and distribution of noise in the 3-D PSFs. For a given master projection, a series of projection images was produced in which each pixel had a value randomly chosen [18] from a Poisson distribution based on the value of the corresponding pixel in a version of the master image normalized to contain 124 000 counts in a circular central area of diameter 5 cm. The noisy projection images were reconstructed via filtered back-projection. For each case studied, 10 such noise variate images were independently produced and used to generate a standard deviation image. Analyses of the 3-D images and standard deviation images were performed using a set of regions of interest (ROIs) in the form of spherical shells of 4.4 mm (1 voxel) thickness. The radii of the ROIs ranged from 1 to 32 voxel in 1 voxel increments. Thus, this set contains ROIs of increasing volume, proportional to the square of the ROI radius. Extended phantom studies Sphere phantom

Measurements were conducted using a 20 cm cylindrical phantom containing spheres with nominal inner diameters of 3.1, 2.5, 2.0, 1.6 and 1.2 cm and some additional spheres with diameters of l cm or less to measure the cumulative effect of noise and PSF distortion on an image

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of a real but simple object. The ratio of activity concentration contained in the spheres to the background activity was 10:1 in both the 140 keV and the 511 keV measurements. Data were acquired at 120 projection angles spanning 3601 at radii of 14 and 26 cm. Projections were binned into a 128 by 128 array with a pixel size of 0.35 cm. Scans of varying duration were acquired so that results spanning a range of count densities could be produced. For each of the four energy and radius combinations, the total number of counts acquired ranged from 1.2  107 to 4.7  107. Reconstruction was via filtered back-projection with a ramp filter only. A multiplicative Chang [19] correction, generated from attenuation coefficients for water [7], was applied to the data. Brain phantom

To demonstrate the effect of septal penetration on a clinically relevant subject where contrast is important, images of a Hoffman 3-D brain phantom [20] (Data Spectrum Corp., Hillsborough, North Carolina) were acquired using the 511 keV collimator. To produce the 511 keV images the phantom was loaded with 89 MBq of activity and scans of various durations were performed over a 5 h period resulting in data spanning a range of count densities. The 140 keV data were obtained similarly. The phantom was loaded with 30 MBq of 99m Tc and scanned for a total acquisition of 106 counts. For both studies the radius of rotation was 12.3 cm. Other acquisition and reconstruction parameters were the same as outlined in the preceding section except that the pixel size was 0.30 cm. Additionally, a 3-D Hann filter with a 1.0 cm – 1 cut-off was applied to the reconstructed image.

Results Point spread functions

The main result of this study is displayed in Fig. 1. Part (a) shows the two-dimensional point spread function from the 140 keV source obtained with a source to collimator distance of 30 cm. This pattern is largely isotropic and has an average full width at half maximum (FWHM) of about 2.1 cm. Part (b) is the 2-D PSF for a 511 keV source at the same distance. The well-known sixpointed starburst pattern [21] seen in the 511 keV image as well as the isotropic background is the result of septal penetration. Although the anisotropy in this distribution is readily apparent, the FWHM values taken in different directions vary by only about 2 mm with an average value of about 3.2 cm. The lower section of each figure shows the collimator modulation transfer function (MTF) from the Fourier transform of the collimator PSF. In the case of an ideal response the MTF is a constant, independent of the spatial frequency.

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Fig. 1

Top sections are two-dimensional images of point sources acquired at a source-to-camera distance of 30 cm. Part (a) was obtained with 140 keV source and part (b) with a 511 keV source. The vertical axis represents intensity while the x and y values represent position on the camera face. The bottom sections show the modulation transfer function from the same data.

Figures 2 and 3 show reconstructed images (3-D PSFs) of the 30 cm, 511 keV master projection obtained from the 2-D PSF shown in Fig. 1. The orientation of the master projection (section a) with respect to the vertical reconstruction rotation axis is different in each figure to simulate possible collimator hole orientations indicated by the inset figure. Sections b, c and d show transaxial slices from the 3-D PSFs obtained from reconstruction of the 2-D images. The PSFs acquired with a collimator-tosource distance of 12 cm are qualitatively similar. For display purposes, the profile scales shown in parts b, c and d of each figure are different. For Fig. 2 the relative fullscale profile values in sections b, c and d are 1:0.022:0.011. In Fig. 3 the values are 1:0.045:0.032. The reconstructed 3-D PSFs for 140 keV are not shown. In this case, as can be seen in Fig. 1. there is no measurable septal penetration and the master projection images do not show a starburst pattern. Likewise, the reconstructed 3-D PSFs are unremarkable and fall off

isotropically with increasing distance from the PSF centre. Figure 4 is a histogram of the number of 3-D PSF counts integrated over a 1 voxel (4.4 mm) thick shell as a function of radius (R) measured from the centre of each PSF. Plots are shown for 511 keV data acquired with source-to-collimator distances of 12 and 30 cm. These can be compared with the equivalent histogram from the 140 keV data. The plots have been normalized such that the total number of counts in the central region, taken to extend to a radius of 1.8 cm, is the same for each PSF. A similar plot for a uniform spatial distribution would be proportional to R2, i.e., proportional to the volume of each shell used in the histogram. The long-dashed line in the figure is a plot of such a function. The results show that averaged (not integrated) over angle, the 511 keV 3-D PSFs fall approximately as 1/R2. A plot for a system with perfect collimation would drop to zero away from the R = 0.

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Characterization of septal penetration in 511 keV SPECT Laymon and Turkington

Fig. 2

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Fig. 4

Reconstructed counts

35000 30000 25000 20000 15000 10000 5000 Part (a) shows a starburst pattern resulting from the penetration of 511 keV photons through the collimator septa. The camera rotation axis is taken to be vertical with the collimator-hole orientation as indicated in the inset. The remaining panels (b–d) are transaxial slices of 4.4 mm thickness of a reconstruction based on the projection shown in part (a). The slices shown are located 0 cm, 4.4 cm and 8.8 cm from the centre of the point spread function. The profiles shown are for a horizontal taken through the centre of each image.

0 0

2

4

6 8 R (cm)

10

12

14

Distribution of voxel intensity from the reconstruction of a 140 keV point source at a camera to source distance of 30 cm (solid line) and of a 511 keV point source at distances of 30 cm (dotted line) and 12 cm (short–dashed line). The plots show the total intensity contained in a 1voxel thick spherical shell as a function of radius. Also shown is a function proportional to R2 (long–dashed line) and thus to the volume of each radial shell.

Fig. 3

Fig. 5

1 0.9 0.8 0.7 MTF

0.6 0.5 0.4 Similar to Fig. 2 except that the penetration pattern has been rotated to correspond to the collimator orientation shown in the inset.

0.3 0.2 0.1 0

Details of the Fourier decompositions of the 140 keV and 511 keV 2-D PSFs for a source-to-collimator distance of 30 cm are shown in Fig. 5 in which we plot MTF value versus spatial frequency. These plots are profiles through the centre of the functions shown in Fig. 1. In the 511 keV case the profile is aligned with two of the small ribs visible on the MTF in Fig. 1 (dashed line). This corresponds to the MTF passing halfway between the tails in the point source projection. Another profile taken at a 301 angle with respect to this that passes between the ribs is also shown dotted line. The difference between these two profiles represents the anisotropy of the MTF. The results of the 3-D PSF noise studies (not shown) indicate that, averaged over angle, the standard deviation per voxel at large distances from the centre of the

0

0.1

0.2 0.3 0.4 Spatial frequency [cm-1]

0.5

0.6

Modulation transfer functions from the 140 keV (solid line) and 511 keV (dashed and dotted lines) point-source acquisitions. Because of the anisotropic septal penetration pattern, the 511 keV modulation transfer function (MTF) is also anisotropic. The dashed and dotted lines represent the MTF along different orientations of the spatial-frequency axis as explained in the text.

distribution falls approximately as 1/R in the 511 keV studies. The 140 keV standard deviation results fall significantly faster than this. Extended phantom studies Sphere phantom

Figure 6 shows 17 mm thick transaxial slices from the reconstructed sphere-phantom data. Parts (a) and (b) of the figure were acquired at camera rotation radii of 14 cm and 26 cm at 511 keV. Parts (c) and (d) are similar but

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906 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 6

Fig. 8

Images of a cylindrical phantom containing spheres with a sphere-tobackground activity concentration ratio of 10:1. Results are shown for 511 keV at radii of rotation of 14 cm (a) and 26 cm (b). Parts c and d are similar but were acquired at 140 keV. Sizes of visible spheres were 3.1, 2.5, 2.0, 1.6 and 1.2 cm. The phantom also contained spheres with diameters of 1 cm and less that were not visible in any study.

Transaxial slice and profile from brain phantom studies at 140 keV (a) and at 511 keV. The total number of counts acquired in the first 511 keV study (b) and the 140 keV study were approximately equal. The additional 511 keV images shown came from studies containing about four times (c) and 18 times (d) the number of counts as in the 140 keV study.

Brain phantom

Results from the studies of the brain phantom are presented in Fig. 8, which shows transaxial slices from the 140 keV study (a) as well as slices from three studies at 511 keV. The first 511 keV slice (b) came from a set of data containing 106 counts which is equal to the total number of counts acquired in the 140 keV study from which the slice in section A was taken. The second (c) and third (d) 511 keV slices were produced from data containing totals of four times and 18 times the number of counts in the 140 keV data.

Fig. 7

4.5 4.0 3.5 Contrast

3.0 2.5 2.0 1.5 1.0

Discussion

0.5

Figure 4 illustrates the fundamental difficulty in obtaining high quality images in cases where there is significant septal penetration. While the radial histogram from the 3-D PSF for 140 keV photons falls almost to zero as a function of radius, those produced with 511 keV photons do not. At 511 keV the number of counts integrated over angle as a function of radius stays approximately constant and thus, the PSF intensity (i.e. count density) decreases approximately as 1/R2.

0.0 1

1.5

2 2.5 Sphere diameter (cm)

3

3.5

Plots of contrast as a function of sphere diameter for images acquired at 140 keV with radii of rotation of 14 cm (solid line) and 26 cm (dashed line) and for images acquired at 511 keV with radii of rotation of 14 cm (dot–dashed line) and 26 cm (dotted line).

were acquired at 140 keV. To allow visual comparison, each of the four images shown was reconstructed from subsets of the projection data containing similar numbers of counts (107). For improved statistical accuracy, all available data were used to determine contrast. Figure 7 shows contrast values as a function of sphere size. We define the contrast, C, to be C¼

F B B

where F is the mean number of counts per pixel within a region of interest and B is the mean number of background counts per pixel. With this definition, the actual contrast value for these studies is 9.

Because we do not observe a significant fall-off in the tails of the 511 keV histograms out to the maximum radius in our analysis (about 14 cm), we cannot state a value for the fraction of the counts in the 511 keV PSFs that result from septal penetration. However, the number of counts outside the central PSF region up to the maximum radius shown in the figure is greater than the number within the central region, even if we consider the central region to extend to a radius of 4 cm. From an imaging standpoint this means that most of the activity in any given volume element in an object will be misplaced in its 3-D image. Furthermore, in the case of a uniform object, misplaced image activity is as likely to have its origin in a distant object volume element as in a nearby one. The magnitude of the blurring effect of

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Characterization of septal penetration in 511 keV SPECT Laymon and Turkington 907

septal penetration becomes apparent by comparing the plots of the 2-D MTFs (which are closely related to the 3-D MTFs by the central section theorem) in Fig. 5. The fall-off with frequency, f, of the 511 keV function is much faster than in the 140 keV case. For example, the 511 keV MTFs drops to 0.5 at f = 0.03 cm – 1 whereas the 140 keV MTF drops to 0.5 at f = 0.20 cm – 1. The 3-D PSFs were constructed from a set of identical projections. The result is an image possessing rotational symmetry about the z-axis but not spherical symmetry. Coupling this with the starburst form of the septal penetration pattern results in a 3-D image consisting of cone-like structures. The PSF contains both positive and negative regions. We do not expect this quantity, averaged over direction (Fig. 4) to be particularly sensitive to different master projection orientations. However, as illustrated by Figs 2 and 3, the orientation of the collimator hole pattern has an effect on the details of the 3-D PSF. For example, if the collimator orientation is such that the septal penetration pattern has tails perpendicular to the camera rotation axis (‘transverse’ orientation), as in Fig. 2, then a significant fraction of misplaced image activity will be spread over the transaxial slice containing the centre of the PSF. If the penetration pattern is rotated by 301 so that there are tails parallel to the camera rotation axis (‘axial’ orientation) then many of the misplaced image counts will fall along the rotation axis. Figure 3 illustrates this. A persistent hot spot can be seen at the centre of each transaxial slice throughout the image. The same effects can be seen in the transfer function of Fig. 5. There is an anisotropy of the dependence on spatial frequency of the 511 keV MTF. The collimator response to spatial frequencies above about 0.03 cm – 1 is noticeably smaller when measured along one of the starburst spires than when measured at an angle that passes between the spires. This is a small effect. Additionally, the axial orientation results in a 3-D PSF that has a region possessing a greater density of misplaced counts, i.e., the persistent hot spot mentioned previously, than can be found in images produced using other orientations. The VPC-93 collimators used for this study, when used as designed, are oriented halfway between transverse and axial. Also, the collimator orientation on one camera is rotated by 301 with respect to the other so that the effective septal penetration pattern is a 12-point starburst (for a 3601 acquisition). Both of these features tend to decrease the anisotropy of the transfer function. In addition to blurring, an extended PSF affects image quality by modifying noise properties. Noise characterization is complicated by the correlations among imagevoxel values introduced during the reconstruction process. For example, in filtered back-projection any active object voxel contributes noise to all image voxels within

the same transaxial slice. The size of the contribution to an image voxel falls with distance from the contributing object voxel. An aspect of septal penetration is that the 3D PSF, and therefore image-noise correlation, extends into many transaxial slices. In this work we found that noise associated with the 511 keV 3-D PSF exhibited a relatively slow 1/R fall-off at long distances from the centre of the distribution. This is consistent with the 1/ R2 average fall-off of the 3-D PSF itself. An effect of septal penetration, summarized in Fig. 7, is a marked decrease in contrast. Separate from penetration effects, there is an increase in contrast with sphere diameter as well as a decrease in contrast as the distance between the camera and the object is increased. These phenomena are related and reflect the fact that contrast degradation depends on the relative sizes of the object and the collimator-induced blurring scale. The effect of blurring and noise can be seen in the figures showing images from phantom studies. In Fig. 6 each image was reconstructed from projections with about the same number of counts. Related to the effect of increased blurring on contrast, one can see decreased edge sharpness between the foreground and background regions. Also, increased noise can be seen throughout the 511 keV images. The same concepts are illustrated by the more complicated and clinically more interesting brain phantom images in Fig. 8. The first of the 511 keV images (b) was produced from raw projection data containing about the same number of counts as the data from which the 140 keV image (a) was produced. The difference in image quality is obvious. The second and third 511 keV images (c and d) were produced from data containing four times and 18 times the number of counts used to produce the 140 keV image. The improvement in image quality from b to d is due to decreased image noise. Image blurring caused by septal penetration, of course, does not improve with increasing count densities. The quality of the image that contains an unrealistically large number of counts (d) is limited by blurring. Collimator design entails trade-offs between sensitivity, blurring due to hole size and spacing (geometric resolution) [22], blurring due to septal penetration, and, in the case of 511 keV imaging, collimator weight. Design choices also affect the noise properties of an image. For example, one can compare the results of this work with that of Leichner et al. [23] who examined some of the imaging characteristics of 511 keV collimator (Picker International, Inc., Cleveland, Ohio) designed for decreased septal penetration. The collimator has larger holes (5.08 mm diameter) and hole spacing (3.43 mm septal thickness) than in the present work.

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908 Nuclear Medicine Communications 2006, Vol 27 No 11

In 511 keV SPECT imaging, as in other imaging modalities, final image quality is affected by the sophistication of the reconstruction algorithm. For example, Zeng et al. [24] have included a distancedependent Gaussian-based model of collimator response in an ordered subsets expectation maximization reconstruction algorithm. The algorithm was tested for 511 keV imaging using the reduced penetration collimator described above. The authors found that including the collimator response produced significant resolution recovery. The results of Zeng et al. suggest that modelling septal penetration effects when appropriate, although computationally expensive, could provide a method for improving image quality. While septal penetration represents a serious impediment to SPECT imaging with 511 keV tracers, there are some situations in which imaging requirements are modest. Several authors [8,12,25] have pointed out that 511 keV SPECT is well suited for dual nuclide imaging (specifically, 99mTc and 18F-FDG) in studies of myocardial perfusion and metabolism, where spatial resolution requirements are not high, and where there is high contrast in tracer uptake.

increase both of these effects and to extend them into many reconstruction planes.

Acknowledgement Support for this research was provided by Elscint, Ltd, Haifa Israel.

References 1 2

3

4

5

6

7

Conclusion We have quantified several aspects of septal penetration using a collimator designed for 511 keV imaging by comparing results of studies at 140 keV with identical studies acquired at 511 keV. Using similar techniques, we have shown the effects of septal penetration on reconstructed images. The results of this work have applications in the planning, processing and interpretation of simultaneous, multi-isotope SPECT procedures. They also provide the collimator response function required for implementation of attenuation correction based on the combination of singles and coincidence data. The degree of septal penetration observed in these studies resulted in significant image blurring. It was found that more than half the reconstructed counts in the 3-D image of a 511 keV point source appeared at distances greater than 4 cm from the image centre. Additionally, there is anisotropy in the blurring associated with septal penetration, the details of which depend on the orientation of the collimator-hole pattern with respect to the camera rotation axis. This effect is unlikely to be of importance in most imaging situations, unless the tail is aligned with the axis of rotation.

8 9

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18

In addition to blurring, septal penetration unfavourably changes image noise characteristics. In general, even in the case of an ideal delta-function PSF, image noise is highly correlated and non-local in nature. The effect of an extended PSF arising from septal penetration is to

19 20

Muehllehner G, Huig L. Septal penetration in scintillation camera collimators. Phys Med Biol 1973; 18:855–862. Bolmsjo¨ MS, Persson BR, Strand SE. Imaging 123I with a scintillation camera. A study of detection performance and quality factor concepts. Phys Med Biol 1977; 22:266–277. Clarke LP, Leong LL, Serafini AN, Tyson IB, Silbiger ML. Quantitative SPECT imaging: influence of object size. Nucl Med Commun 1986; 7: 363–372. Sandler MP, Patton JA. Fluorine 18-labeled fluorodeoxyglucose myocardial single-photon emission computed tomography: an alternative for determining myocardial viability. J Nucl Cardiol 1996; 3:342–349. Sandler MP, Videlefsky S, Delbeke D, Patton JA, Meyerowitz C, Martin WH, Ohana I. Evaluation of myocardial ischemia using a rest metabolism/stress perfusion protocol with fluorine-18 deoxyglucose/technetium-99m MIBI and dual-isotope simultaneous-acquisition single-photon emission computed tomography. J Am Coll Cardiol 1995; 26:870–878. Peschina W, Conca A, Konig P, Fritzsche H, Beraus W. Low frequency rTMS as an add-on antidepressive strategy: heterogeneous impact on 99mTcHMPAO and 18F-FDG uptake as measured simultaneously with the double isotope SPECT technique. Pilot study. Nucl Med Commun 2001; 22: 867–873. Hubbell JH. Photon cross-sections, attenuation coefficients, and energy absorption coefficients from 10 keV to 100 GeV. National Bureau of Standards Reference Series 1969; 29:62–66. Smith EM, O’Mara RE. 511 keV imaging: SPECT, coincidence, or both. Appl Radiol 1996; 25:6–27. Martin WH, Delbeke D, Patton JA, Hendrix B, Weinfeld Z, Ohana I, et al. FDG-SPECT: correlation with FDG-PET. J Nucl Med 1995; 36: 988–995. Mukherji SK, Drane WE, Tart RP, Landau S, Mancuso AA. Comparison of thallium-201 and F-18 FDG SPECT uptake in squamous cell carcinoma of the head and neck. Am J Neuroradiol 1994; 15:1837–1842. Worsley DF, Celler A, Adam MJ, Kwong JS, Muller NL, Coupland DB, Champion P, Finley RJ, Evans KG, Lyster DM. Pulmonary nodules: differential diagnosis using 18F-fluorodeoxyglucose single-photon emission computed tomography. Am J Roentgenol 1997; 168:771–774. Bax JJ, Cornel JH, Visser FC, Fioretti PM, Huitink JM, van Lingen A, Sloof GW, Visser CA. F18-fluorodeoxyglucose single-photon emission computed tomography predicts functional outcome of dyssynergic myocardium after surgical revascularization. J Nucl Cardiol 1997; 4: 302–308. Chen EQ, MacIntyre WJ, Go RT, Brunken RC, Saha GB, Wong CYO, Neumann DR, Cook SA, Khandekar SP. Myocardial viability studies using fluorine-18-FDG SPECT: a comparison with fluorine-18-FDG PET. J Nucl Med 1997; 38:582–586. Martin WH, Delbeke D, Patton JA, Sandler MP. Detection of malignancies with SPECT versus PET, with 2[fluorine-18]fluoro-2-deoxy-D-glucose. Radiology 1996; 198:225–231. Drane WE, Abbott FD, Nicole MW, Mastin ST, Kuperus JH. Technology for FDG SPECT with a relatively inexpensive gamma camera. Work in progress. Radiology 1994; 191:461–465. Patton JA, Sandler MP, Ohana I, Weinfeld Z. High-Energy (511-keV) imaging with the scintillation camera. Radiographics 1996; 16:1183–1194. Laymon CM, Turkington TG. Calculation of attenuation factors from combined singles and coincidence emission projections. IEEE Trans Med Imaging 1999; 18:1194–1200. Press WH, Teukolsky SA, Vetterling WT, Flannery BP. Numerical recipes in C,2nd edition. Cambridge: Cambridge University Press; 1992. Chang LT. A method for attenuation correction in radionuclide computed tomography. IEEE Trans Nucl Sci 1978; 25:638–642. Hoffman EJ, Cutler PD, Guerrero TM, Digby WM, Mazziotta JC. Assessment of accuracy of PET utilizing a 3-D phantom to simulate the activity distribution of [18F]fluorodeoxyglucose uptake in the human brain. J Cereb Blood Metab 1991; 11:A17–A25.

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Characterization of septal penetration in 511 keV SPECT Laymon and Turkington 909

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Harris CC, Bell PR, Francis JE, Satterfield MM, Jordan JC, Murray JP. Collimators for radioisotope scanning. In: Knisely RM, Gould AA, Harris CC (editors): Progress in Medical Radioisotope Scanning. Proceedings of a Symposium at the Medical Division of the Oak Ridge Institute of Nuclear Studies, 22–26 October 1962. U.S. Atomic Energy Commission, Division of Technical Information, 1963. Metz CE, Atkins FB, Beck RN. The geometric transfer function component for scintillation camera collimators with straight parallel holes. Phys Med Biol 1980; 25:1059–1070.

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Original article

Scintimammographic detection of usual ductal breast hyperplasia with increased proliferation rate at risk for malignancy Vassilios Papantonioua, Spyridon Tsiourisa, John Koutsikosa, Maria Sotiropouloub, Ekaterini Maintaa, Dimitrios Lazarisc, Pipitsa Valsamakia, Maria Melissinoud, Cherry Zervaa and Aris Antsaklisc Objectives To estimate whether breast uptake of 99m Tc-(V)DMSA and 99mTc-sestamibi in usual ductal epithelial breast hyperplasia (UDH) and apocrine metaplasia is related to cell proliferation rate (Ki-67) and oestrogen receptor (ER) expression, both of which are associated with the potential risk of evolving to malignancy. Methods Among patients referred for suspicious breast findings on palpation and/or mammography and evaluated preoperatively with both radiopharmaceuticals, we retrospectively studied 17 (10 with UDH: group I; and seven with apocrine metaplasia: group II). Lesion-to-background (L/B) ratios in early and late acquisitions were calculated for both radiotracers in both groups, as well as their retention ratios. Ki-67 and oestrogen receptor expression were determined immunohistochemically. The late L/B ratios between the two tracers were compared, as were the late ratios for each tracer between Ki-67 r 3% and > 3%, and between ER r 15% and > 15%. Linear regression analysis was also performed between L/B and retention ratios and Ki-67 expression.

group (P = 0.156 and 0.274, respectively). Significant coefficient correlation was found between the 99m Tc-(V)DMSA L/Blate ratios and retention ratios and Ki-67 levels only for group I (r = 0.889, P < 0.001 and r = 0.802, P < 0.01, respectively). The 99mTc-(V)DMSA L/Blate ratios in group I were significantly higher when Ki-67 > 3% than when Ki-67 r 3% (P = 0.016) but did not differ considerably between ER > 15% and r 15% (P = 0.732). Conclusion 99mTc-(V)DMSA uptake in UDH correlates with Ki-67 expression. This could prove useful in identifying women with benign but high-risk breast pathologies who might benefit from chemoprophylaxis. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:911–917 Nuclear Medicine Communications 2006, 27:911–917 Keywords: scintimammography, 99mTc-(V)DMSA, 99mTc-sestamibi, usual ductal hyperplasia, apocrine metaplasia, Ki-67, oestrogen receptor Departments of aNuclear Medicine, bPathology, cObstetrics & Gynaecology, ‘‘Alexandra’’ University Hospital, Athens and dInternal Medicine, ‘‘Metropolitan’’ Hospital, Athens, Greece.

Results There was a significant increase of the 99m Tc-(V)DMSA L/B ratio in late images as compared to the early images in group I (P < 0.05), while in group II it was not significantly increased (P = 0.084). 99mTc-sestamibi ratios did not demonstrate variability over time in either

Correspondence to Dr John Koutsikos, ‘‘Alexandra’’ University Hospital, Department of Nuclear Medicine, 80, Vas. Sophia’s Avenue & 2, K. Lourou Street, 115 28, Athens, Greece. Tel: + 0030 210 33 81 785; fax: + 0030 210 77 07 404; e-mail: [email protected]

Introduction

during puberty, the early post-menopausal period and the menopausal period [3]. The presence of estrogen receptor type A (ERA) in benign lesions could be a risk factor for progression to breast malignancy, by rendering cells sensitive to the proliferatory stimulus of oestrogens [4].

The multi-step model for breast carcinogenesis proposes that invasive cancer arises through a series of hyperplastic lesions followed by various grades of atypia, from epithelial hyperplasia to in-situ and invasive carcinoma. Usual hyperplasia of the ductal epithelium (UDH) is a common lesion representing an, as yet, not committed precursor phase of carcinogenesis, but is associated with an increased risk of developing cancer [1,2]. It consists of a variety of changes, from minimal stratification of intraluminal cells to proliferation that could be considered as atypical ductal hyperplasia (ADH). Epidemiological and experimental evidence suggest that breast cancer risk is associated with oestrogen exposure

Received 10 December 2005 Accepted 24 January 2006

Scintimammography is an established method for imaging invasive lesions greater than 1 cm in size, particularly where conventional X-ray mammography faces inherent limitations (e.g., dense breasts, augmentation implants, scars from previous surgical intervention or radiotherapy) with very good sensitivity (88%), specificity (93%) and accuracy (90%) [5,6]. Various tumour-seeking compounds labelled with 99mTc have been used, including 99m Tc-2-methoxy isobutyl isonitrile (99mTc-sestamibi),

c 2006 Lippincott Williams & Wilkins 0143-3636

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912 Nuclear Medicine Communications 2006, Vol 27 No 11

99m

Tc-6,9-bis(2-ethoxyethyl)-3,12-dioxa-6,9-diphosphate tradecane (99mTc-tetrofosmin) and 99mTc(V)-dimercaptosuccinic acid (99mTc-(V)DMSA). Moreover, 99mTc(V)DMSA exhibits favourable features in imaging preinvasive breast pathologies (reported sensitivity of B95% and negative predictive value of B98% for ductal carcinoma in situ (DCIS) and lobular carcinoma in situ (LCIS)), particularly those comprising highly proliferative epithelial cell populations, this trait being probably due to the close relationship between 99mTc-(V)DMSA cellular uptake and cell proliferation rate (Ki-67). This has been verified both in vivo [7,8] and in vitro in several cancer cell lines [9]. A surprising finding in one our previous series [8] was that a diffuse pattern of radiotracer distribution occurred in the benign lesion of ductal breast hyperplasia, in both the usual and the atypical type, but never in the benign lesions of fibrosis, adenosis and ductal cystic dilatation. This diffuse uptake was observed predominantly with 99mTc-(V)DMSA and to a considerably lesser extent with 99mTc-sestamibi, varying from faint (mainly with 99mTc-sestamibi) to intense (mainly with 99mTc-(V)DMSA). With the latter agent, uptake also displayed a tendency to increase over time. We assumed this could probably be attributed to the different degree of proliferative activity in epithelial cells, given the proven correlation between this radiotracer and cell proliferation. This prompted us to investigate, retrospectively, this phenomenon in epithelial breast hyperplasia, a lesion that theoretically might represent a premature stage in the process of breast carcinogenesis, and to correlate the intensity of the diffuse uptake pattern with the degree of cellular proliferative activity and with other potentially crucial parameters in the process of breast carcinogenesis, such as oestrogen receptor expression. This study aimed to assess, retrospectively, whether Tc-(V)DMSA and 99mTc-sestamibi uptake in the benign breast lesions of UDH is related to Ki-67 and oestrogen receptor expression, both of which are parameters associated with an increased risk of evolving into ADH, DCIS and invasive carcinoma.

Among these 45 women, we retrospectively studied 10 in whom histology had eventually confirmed UDH (group I: mean age 52.7 years (95% confidence interval (CI) = 45.9– 59.5 years); in three it was the sole histological finding, while in seven patients UDH co-existed with an apocrine metaplasia component, in accordance with literature reports estimating concurrent apocrine metaplasia in 80–85% of UDH cases [10]. Due to this co-existence, it would not be prudent to attribute a priori any increased radiotracer uptake to hyperplasia, to the apocrine component, or both. To overcome this, a second group of seven women (from the 45 patients) with isolated apocrine metaplasia was studied (group II: mean age 52.0 years (95% CI = 41.7–62.3 years)). The same immunohistological and scintigraphic parameters were analysed in group II, in order to have a net estimate of the behaviour of the apocrine component (where present) in group I and its possible interference in the interpretation of the results. Scintimammography

The patients underwent a 99mTc-sestamibi–99mTc-(V) DMSA double-phase study, with a 48 h time interval. Informed consent was obtained in every case. 99m

Tc-(V)DMSA was prepared using a commercially available kit (DMS(V)/Demoscan, National Center of Physical Sciences, Institute of Radioisotopes and Radiodiagnostics ‘DEMOKRITOS’, Athens, Greece). The kit for the preparation of 99mTc-sestamibi (Cardiolites) was obtained from Bristol–Myers Squibb GmbH (Regensburg, Germany). Both compounds were labelled with 99mTc. After intravenous administration of the radiotracer (925–1100 MBq), early (at 10–20 min) and late (at 60–70 min) lateral and anterior planar images were acquired; the lateral images with the patient prone and the anterior ones in the supine position.

99m

Methods Study population

One hundred and two women who were referred with suspicious breast lesions on physical examination and/or an abnormal mammogram, underwent 99mTc-MIBI and/or 99m Tc-(V)DMSA scintimammography prior to any surgical intervention. All patients were intended to have studies performed with both agents. The first agent to be used was selected on a random basis. Not all patients managed to have the second study performed preoperatively, for various non-clinical reasons. Forty-five of them underwent both 99mTc-(V)DMSA and 99mTc-sestamibi breast scintigraphy, at separate sessions, in a head-tohead, double-phase study.

Acquisitions were carried out using a special positioning pad (PBI-2 Scintimammography Pad Sets, Pinestar Technology Inc., Greenville, Pennsylvania, USA). A single-head tomographic gamma camera was used (Sophycamera DS7s, Sopha Medical Vision International, Buc Cedex, France), equipped with a high-resolution parallel-hole collimator. The matrix was 256  256 pixels and the photopeak was centred at 140 keV, with a symmetrical 10% window. Image analysis

Radiotracer accumulation in the breast was evaluated qualitatively in early and late images, regarding the site, shape, pattern, extent, homogeneity and dispersion of activity. A characteristic pattern of diffusely increased uptake corresponding to the lesion was observed in most 99m Tc-(V)DMSA scans (Figs 1 and 2(a and b), and in some 99m Tc- sestamibi studies. Semi-quantitative analysis was also performed, by calculating the lesion-to-background (L/B) ratio in early and late acquisitions, as follows: regions

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Breast epithelial hyperplasia at risk for cancer Papantoniou et al. 913

Fig. 1

Fig. 2

Increased diffuse uptake of the radiotracer into the breast and definition of the region of interest (ROI). (a) Usual ductal hyperplasia of the right breast. 99mTc-(V)DMSA scintimammography, right lateral projection at 60 min. Increased diffuse tracer uptake corresponding to the lesion (curved arrow). (b) The same image as (a), with defined ROIs. The 60 min lesion-to-background (L/Blate) ratio was 1.92.

of interest (ROIs) of standardized size and shape were defined over the site of the greatest radioactivity within the areas of diffusely increased tracer uptake, and over the surrounding normal parenchyma (Fig. 1(b)). When no uptake was observed during the early acquisition but only in the late images, the early ROIs were defined in the corresponding areas where the late ROIs were drawn. If no uptake was apparent throughout the study (Fig. 2(c and d)), both breast ROIs were randomly selected and, on the assumption that the L/B ratio should be at least 1.00, the ROI encompassing the higher average counts was used as the numerator, while the other was set as the denominator.

Dual-tracer study in usual breast hyperplasia. Usual ductal hyperplasia of the right breast in a 68-year-old woman. Scintimammography, right lateral projection; 99mTc-(V)DMSA at (a) 10 min and (b) 70 min. Diffusely increased tracer uptake in the area corresponding to the lesion is observed, barely visible during early acquisition (arrowheads), but clearly perceptible at 70 min (lesion-to-background ratios: L/ Bearly = 1.32 and L/Blate = 1.83). 99mTc-sestamibi at (c) 10 min and (d) 65 min. No increased tracer activity is identified in the corresponding area throughout the study (L/Bearly = 1.12 and L/Blate = 1.14).

Statistical analysis Histopathology and immunostaining

The diagnosis of UDH and apocrine metaplasia was based on histopathology of the specimens obtained surgically. The avidin–biotin–alkaline phosphatase immunohistochemical method was carried out on paraffin-embedded tissue sections, for the demonstration of Ki-67 and oestrogen receptor expression. For oestrogen receptor proteins the 1D5 monoclonal antibody (DAKO, Glostrup, Denmark) was used at a dilution of 1:400, and a polyclonal antibody (DAKO), at a dilution of 1:350, was used for Ki-67. The quantitative estimation of Ki-67 and oestrogen receptor expression was based on the percentage of positively staining cells. The Ki-67 and oestrogen receptor levels for normal breast epithelium varies in several literature reports, ranging up to 3% for Ki-67 [11], and between 10 and 20% for oestrogen receptor [1]. Using a 3% cut-off level for Ki-67 as the demarcation line between normal and abnormal and 15% oestrogen receptor expression in a similar manner, an attempt was made to see whether there was a relationship between the in-vivo radiotracer uptake and the levels of Ki-67 and/or oestrogen receptor in UDH. The same determination was performed on group II.

In order to assess any significant differences concerning the uptake of each radiotracer, comparisons were conducted between the L/B ratios of early and late acquisitions (hereafter referred to as L/Bearly and L/Blate, respectively) and the difference between them (retention ratio), within groups and for each radiotracer separately and were compared between patients with Ki-67 r 3% and Ki-67 > 3%, and with ER r 15% and ER > 15%. The two-sided, paired, two-sample t-test for means was used for the comparison between L/Bearly and L/Blate, and the two-sided Mann–Whitney non-parametric U-test was applied for all other analyses. The ratios were correlated (linear regression analysis) with Ki-67 and oestrogen receptor levels in each group. P < 0.05 was considered statistically significant.

Results Expression of Ki-67 and oestrogen receptor in both groups is shown in Table 1. With regard to the qualitative scan interpretation, a diffuse 99mTc-(V)DMSA uptake pattern (in early and/or late images) was observed in 8/10

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914 Nuclear Medicine Communications 2006, Vol 27 No 11

These qualitative features of image reading were verified semiquantitatively (Table 2). There was a significant increase of 99mTc-(V)DMSA L/B ratio in late images as compared to early in group I (L/Bearly vs. L/Blate, P < 0.05) while in group II it was not significantly increased (P = 0.084). 99mTc-sestamibi ratios did not demonstrate variability over time in either group (P = 0.156 and 0.274, respectively). Statistical differences between the two groups for both radiotracers are also shown in Table 2. Significant statistical differences were found only for L/B ratios for 99m Tc-(V)DMSA. Within group I the 99mTc-(V)DMSA L/B ratios were significantly higher, in comparison to those for 99mTc-sestamibi (P < 0.05 for L/Bearly and P < 0.005 for L/Blate). This was not found in group II (P = 0.86 and 0.9, respectively). With regard to the correlation between L/B ratios and Ki-67 and oestrogen receptor levels (separately assessed Table 1 Expression of the cell proliferation index (Ki-67) and the oestrogen receptor in patients with usual hyperplasia of the ductal epithelium (group I) and in patients with apocrine metaplasia (group II) Ki-67 or oestrogen receptor Ki-67 (%) Mean (95% CI) Range Oestrogen receptor (%) Mean (95% CI) Range

Table 2

Group I

Group II

6.10 (2.09 ± 10.11) 1–20

3.86 (0 ± 8.04) P = 0.279 1–10

45.5 (23.88 ± 67.12) 5–90

0 0

for each group), statistical significance was found only between the L/Blate and retention ratios for 99mTc(V)DMSA uptake and Ki-67 levels for group I (r = 0.889, P < 0.001 and r = 0.802, P < 0.01, respectively) (Figs 3 and 4). All other correlations were found to be non-significant. Finally, within each group the L/Blate ratios and P values for both radiotracers, with respect to Ki-67 levels and to oestrogen receptor levels, are summarized in Table 3. The only significantly increased uptake ratio observed concerned 99mTc-(V)DMSA in subjects with UDH lesions over-expressing Ki-67 (P = 0.016).

Discussion According to reports on breast cancer screening, if the disease was detected at an earlier stage, the absolute mortality could be reduced [12–14]. The majority of invasive breast malignancies are considered to develop from certain pre-existing non-malignant lesions over long time periods. Only a few types of benign lesions have significant malignant potential, UDH being one example [2,3]. Two such markers are cell proliferation index (Ki67) and oestrogen receptor expression. Ki-67 is an indicator of UDH potential to evolve to malignancy, as

Fig. 3

99mTc − (V)DMSA L/B LATE ratio

patients in group I and in 0/7 in group II. With 99mTcsestamibi, such a pattern was found in 3/10 and 0/7 women, respectively. The visual intensity of the uptake ranged from rather faint (mainly with 99mTc-sestamibi or in early acquisitions) to distinct (mostly with 99mTc(V)DMSA or in late acquisitions). As a general point, an L/B threshold around 1.30 corresponded to lesions that were scarcely visible (Fig. 2(a)). The relative intensity of 99m Tc-(V)DMSA uptake in group I – as compared to the surrounding normal breast uptake – demonstrated an obvious tendency to increase over time (Fig. 2(b)).

3 2.5 r =0.889, P < 0.001

2 1.5 1 0.5 0

0

5

10 Ki − 67 levels (%)

15

20

Linear correlation of the L/Blate ratios for 99mTc-(V)DMSA uptake and Ki67 levels (%) in group I (r = 0.889, P < 0.001).

Lesion-to-background (L/B) uptake ratios and retention ratios (L/Blate – L/Bearly) for both radiotracers in the two groups

Radiotracer and uptake ratios

Group I

Statistical significances

Group II

99m

Tc(V)-DMSA L/Bearly L/Blate Retention ratio (L/Blate – L/Bearly) 99m Tc-sestamibi L/Bearly L/Blate Retention ratio (L/Blate – L/Bearly)

1.41 (1.22–1.60) 1.73 (1.40–2.05) 0.316 (0.112–0.52)

P < 0.01 P< P < 0.005

1.10 (1.02–1.19) 1.15 (1.11–1.19) 0.046 (0.003–0.088)

1.18 (1.09–1.26) 1.20 (1.13–1.27) 0.025 ( – 0.007–0.057)

P = 0.2 P = 0.26 P = 0.64

1.11 (1.05–1.17) 1.15 (1.11–1.19) 0.044 ( – 0.028–0.116)

Group I consisted of patients with usual hyperplasia of the ductal epithelium. Group II consisted of patients with apocrine metaplasia. Results are given as the mean with the 95% confidence interval.

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Breast epithelial hyperplasia at risk for cancer Papantoniou et al. 915

well as an indicator of the aggressiveness of cancer cells [15,16]. Shaaban and colleagues [1] have demonstrated correlation between both Ki-67 status and oestrogen receptor A expression in a subset of UDH that definitely progressed to breast cancer. The cellular biological pathways of 99mTc-(V)DMSA are less distinct. Its incorporation in cancer cells has been related to glucose metabolism mediated acidosis [17] and to phosphate transport by specific NaPi co-transporters [18]. Our previous scintimammographic studies involving invasive and pre-invasive breast cancer populations recognized for the first time a significant correlation between 99mTc-(V)DMSA uptake and cell proliferation activity, as measured by Ki-67 expression [7,8]. These invivo observations have very recently been verified in vitro, not only for breast, but in six different human cancer cell lines, where 99mTc-(V)DMSA cell incorporation was found linked to cellular proliferation [9]. In the present study, UDH lesions with Ki-67 > 3% had significantly increased 99mTc-(V)DMSA uptake, in comparison with lesions expressing Ki-67 r 3%. As described for DCIS/LCIS imaging with 99mTc-(V)DMSA, the diffuse uptake of this tracer tends to characteristically increase in intensity over time, being more prominent in

late imaging [7]. This was observed in the present study also, while, in contrast, no significant L/B ratio fluctuation with time was found for 99mTc-sestamibi. This is concordant with several reports which concluded that the relationship between 99mTc-sestamibi uptake and cell proliferation activity in rapidly proliferating cancerous and precancerous breast tissues is, at most, rather poor [19,20]. Late, rather than early, 99mTc-(V)DMSA uptake has been verified as representing the cellular proliferative activity [9]. The incremental uptake of this tracer over time, as well as its relation to cell proliferation, are indicative of a distinct cellular pathway for its incorporation in breast lesions, possibly reflecting phosphate uptake and/or phosphorylation–activation of the focal adhesion kinase enzyme, which is implicated in the cascade leading to cellular proliferation and progression of carcinogenesis [9,18,21]. The possibility that this finding is due to locally increased vascularity, angiogenesis or vessel permeability is therefore highly unlikely; if that was the case, then this diffuse pattern would have been equally clearly visible in the early 10 min images as well. Actually, the only correlation observed in our study was between L/ Blate ratios and retention ratios for 99mTc-(V)DMSA uptake and Ki-67 levels in group I. We therefore deemed it most proper to use the late L/B ratios for the analyses rather than the ratios derived from early imaging.

99mTc − (V)DMSA retention ratio

Fig. 4

1 0.8 r = 0.802, P < 0.01

0.6 0.4 0.2 0 0

2

4

6

8

10

12

14

16

18

20

Ki − 67 levels (%) Linear correlation of the retention ratios (L/Blate – L/Bearly) for 99mTc(V)DMSA uptake and Ki-67 (%) levels in group I (r = 0.802, P < 0.01).

In the same way with Ki-67, the mean oestrogen receptor expression in UDH lesions was above the threshold value for normal epithelium (15%). Nevertheless, lesions with ER > 15% did not demonstrate significantly different 99m Tc-(V)DMSA uptake, as compared to those with ER r 15%. This implies that oestrogen receptor positivity in UDH does not interfere with 99mTc-(V)DMSA uptake. With regards to 99mTc-sestamibi, no correlation has been recognized between its uptake and oestrogen receptor status in breast cancer, something that was also found by our first group. However, the lack of relationship between 99m Tc-(V)DMSA uptake and oestrogen receptor expression does not seem to be a real obstacle with regard to possible anti-oestrogen chemoprophylaxis, taking into account that in the vast majority of UDH cases the lesions are positive for the oestrogen receptor.

Table 3 Lesion-to-background uptake ratios for both radiotracers at late images (L/Blate) in both groups, with respect to Ki-67 levels r 3% and > 3%, and to oestrogen receptor (ER) levels r 15% and > 15% Group I*

Radionuclide Ki-67 r 3% 99m

Tc(V)-DMSA

99m

Tc-sestamibi

Ki-67 > 3%

1.20 (0.77–1.62) 1.95 (1.66–2.25) P = 0.016 1.18 (0.77–1.58) 1.21 (1.14–1.28) P = 0.908

Group II** ER r 15%

ER > 15%

1.67 (0.23–3.10) 1.75 (1.34–2.16) P = 0.732 1.20 (0.77–1.62) 1.20 (1.14–1.27) P = 0.730

Ki-67 r 3%

Ki-67 > 3%

1.14 (1.07–1.21) 1.16 (1.01–1.31) P = 0.858 1.15 (1.09–1.22) 1.15 (1.01–1.29) P = 0.858

Results in brackets are the 95% confidence intervals. * In group I, patients showed usual ductal epithelial breast hyperplasia (UDH) with or without apocrine metaplasia. The results for this group refer to the UDH component. ** In group II, patients showed isolated apocrine metaplasia. All ER values in this group were zero.

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916 Nuclear Medicine Communications 2006, Vol 27 No 11

An apocrine histology is observed in a spectrum of epithelial breast lesions, from benign metaplasias to apocrine carcinoma. Apocrine metaplasia with UDH constitutes a common finding in fibrocystic breast disease and they very often co-exist [10,22]. The mean Ki-67 expression in group II was slightly above 3%; oestrogen receptor expression was absent, in agreement with the literature [23]. The fact that no diffusely increased uptake of either tracer was observed in group II subjects and, furthermore, that the L/Blate ratios for both radiotracers in this group did not differ significantly between Ki-67 above and below 3%, indicate that the apocrine component alone (and its Ki-67/ER profile) does not appear to influence the breast uptake of either compound. This irrelevancy between 99mTc-(V)DMSA uptake and cell proliferation in apocrine metaplasia – dissimilar to UDH, DCIS/LCIS, and invasive carcinoma – is an indeterminate interesting issue that remains to be investigated. It could be attributed to the fact that cellular proliferation in apocrine metaplastic epithelium is seldom increased to the levels observed in UDH and apocrine metaplasia very rarely progresses to apocrine carcinoma. Similarly, in our apocrine metaplasia population the mean Ki-67 value was only slightly increased above 3%, nearly the half of that found in UDH. As a result, the higher 99mTc-(V)DMSA uptake in UDH lesions overexpressing Ki-67, in comparison with these displaying Ki-67 r 3%, cannot be attributed to the coexistent apocrine component (where present). In that sense, the concurrence of apocrine metaplasia in the majority of group I patients should not be considered as a significant overlapping factor affecting 99mTc-(V)DMSA uptake and interfering with the interpretation of the results.

invasive cancer. There is indeed a need to verify these observations in a larger series and to estimate the ability of the method in affecting treatment decisions.

Acknowledgements The authors wish to thank Drs Nikolaos Ptohis and Eva Sotiropoulou for their co-operation in mammographic screening of the patients and Mr Dimitrios Dristiliaris for help with statistical analysis.

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Similar studies involving pre-malignant hyperplastic lesions such as UDH, suggest that the increased coexpression of both ERA and Ki-67 defines a population subset at increased risk of subsequent cancer development, with Ki-67 being the major predictor of this aspect [1]. This subgroup could benefit from prophylactic antioestrogen therapy, by diminishing the proliferative activity of these benign but high-risk lesions [24]. To the best of our knowledge, no imaging modality, to date, can recognize these breast lesions and provide an estimate of their risk for malignant transformation. Scintigraphy with 99mTc-(V)DMSA could therefore be used as a screening tool together with standard mammography for selection of patients for biopsy and the determination of Ki-67 expression.

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Original article

Incidence and radio-uptake patterns of femoral head avascular osteonecrosis at 1 year after renal transplantation: a prospective study with planar bone scintigraphy Eun Jeong Leea, Kyung-Han Leea, Woo Seong Huhb, Joon-Kee Yoonc, Hyun Woo Chunga, Joon Young Choia, Yearn Seong Choea, Yong Choia, Ha Young Ohb and Byung-Tae Kima Objectives We prospectively investigated the incidence of femoral head avascular osteonecrosis (AVN) and radio-uptake patterns of femoral heads on bone scintigraphy at 1 year after renal transplantation. Methods A total of 237 subjects (473 femoral heads) were included. A bone scintigraphy was performed at 12 ± 1.1 months after renal transplantation, and 17 hips were painful at the time of bone scintigraphy. We graded the radioactivity in each femoral head as normal (grade 0), mildly increased (grade I), and definitely increased (grade II). Typical photon defects in the upper lateral femoral heads were evaluated separately. AVN was confirmed with clinical follow-up of more than 1 year and MRI and/or plain radiography findings. Results Femoral head AVN was detected in 15 of the 237 patients and 23 of the 473 femoral heads. When grade I and II activities were used as positive criteria, bone scintigraphy had a sensitivity of 91.3% (100% with pain) and specificity of 74.0% (100% with pain) for AVN diagnosis. When only grade II activity was considered positive, the rates were 56.5% (80.0% with pain) and 99.5% (100% with pain), respectively. The presence of a typical

Introduction Avascular osteonecrosis (AVN) is a well known musculoskeletal complication after renal transplantation with an incidence of 4–41% [1–4]. The most common site of involvement is the femoral head. Early detection of femoral head AVN is important because early surgical core decompression may arrest progress of the disease and prevent collapse of the femoral head [5,6]. The true prevalence of femoral head AVN following renal transplantation and the time to its development have not been clearly elucidated. In several prior prospective studies with longer follow-up periods, nearly all occurrences of AVN were identified on magnetic resonance imaging (MRI) by the first 3 months after transplantation [7–9]. However, a more recent report suggests a delayed presentation of AVN in two of seven patients, occurring at 8 and 10 months after transplantation [10].

photon defect had a low sensitivity of 47.8%, although the specificity was high (99.1%). Conclusions The incidence of femoral head AVN was low among a prospective cohort of renal transplantation recipients at the time of 1 year after engraftment. Planar bone scintigraphy is sufficient to diagnose AVN in symptomatic patients at risk for femoral head AVN using grade I and II activities as positive criteria. Nucl Med c 2006 Lippincott Williams & Wilkins. Commun 27:919–924 Nuclear Medicine Communications 2006, 27:919–924 Keywords: AVN, femoral head, renal transplantation, bone scintigraphy Departments of aNuclear Medicine, bMedicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul and cNuclear Medicine, Ajou University School of Medicine, Suwon, Korea. Correspondence to Dr Kyung-Han Lee, Department of Nuclear Medicine, Samsung Medical Center, 50 Ilwondong, Kangnamgu, Seoul, Korea Tel: + 0082 2 3410 2630; fax: + 0082 2 3410 2639; e-mail: [email protected] This work was presented, in part, at the 50th annual meeting of the Society of Nuclear Medicine, New Orleans, USA, 23 June 2003. Received 21 April 2006 Accepted 28 July 2006

For diagnosing femoral head AVN, MRI is sensitive and specific and has become the tool of choice for early and accurate detection [11]. Several studies reported that the sensitivity of MRI is 10–20% higher than that of bone scintigraphy [12–15]. This latter technique, however, has several merits such as that it provides whole-body scanning, is widely accessible, and may be competitive in cost-effectiveness. We therefore prospectively investigated the incidence of femoral head AVN and radio-uptake patterns of femoral heads on bone scintigraphy 1 year after renal transplantation.

Methods Subjects

A total of 333 patients received a renal transplant at our institution between February 1995 and June 2000. Among

c 2006 Lippincott Williams & Wilkins 0143-3636

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920 Nuclear Medicine Communications 2006, Vol 27 No 11

these patients, the renal graft was removed (range, 9 days to 3.4 months) because of acute rejection in 10 patients and surgical complications in two patients. One patient died 25 days after transplantation because of a surgical complication. Seven patients were lost to follow-up study after being referred to other hospitals. Excluding the above subjects, a total of 313 consecutive patients were prospectively referred for bone scintigraphy to screen for femoral head AVN, approximately 1 year after transplantation. Of these subjects, 44 refused the bone scintigraphy. In addition, five were excluded because they had undergone bilateral femoral head core decompression due to AVN prior to the bone scintigraphy , and 13 were also excluded because the interval between 1 year posttransplantation and the bone scintigraphy was greater than 4 months. Fourteen cases were later excluded from analysis because of image data loss. One femoral head was excluded because the patient received total replacement arthroplasty of the right hip due to a traffic accident before the transplantation. As a result, a total of 237 subjects and their 473 femoral heads were finally included in the study. The bone scintigraphy was performed at 12 ± 1.1 months after transplantation (range, 8–16 months). There were 127 male and 110 female patients with mean age at the time of bone scintigraphy of 40 years (range, 21–75 years). Seventeen hips were painful at the time of bone scintigraphy , and the time interval between transplantation and the development of hip pain was 12.3 ± 1.8 months (range, 9–15 months). The time interval between the pain onset and bone scintigraphy was 1.9 ± 2.0 months (range, 10 days to 6.7 months). The remaining 456 hips were asymptomatic (Table 1). All patients were treated with steroids, azathioprine and cyclosporin after transplantation. All received a mean dose of 1.85 g of intravenous methylprednisolone and 1.34 g of oral prednisolone during the first 90 days after transplantation. Mean cumulative doses of prednisolone given during the first 90 days and 1 year after transplantation in all patients were prednisolone equivalent doses of 3.65 g and 4.5 g, respectively.

Review of medical records, plain radiography of hips, MRI of hips, post-operative pathology, and/or clinical follow-up of more than 1 year after bone scintigraphy were used to determine the presence of AVN. The mean duration of clinical follow-up after bone scintigraphy was 3.1 ± 1.3 years (range, 1.1–6.2 years). This study was approved by our institutional review board. Bone scintigraphy

Bone scintigraphy was performed 4 h after intravenous injection of 740 MBq (20 mCi) 99mTc-methylene diphosphonate (MDP). Anterior and posterior whole-body images were simultaneously obtained by a large-field-ofview dual headed gamma camera with a general purpose, parallel-hole collimator (Biad XLT; Trionix Research Laboratory, Ohio, USA) for 106 counts (scan speed of 18 cm
min – 1) with zoom factor of 1.0, and stored on a 1024  256 pixel matrix. Bone scintigraphy images were visually assessed by two experienced nuclear medicine physicians independently, unaware of the results of plain radiography, MRI and clinical information. The radioactivity in each femoral head was graded as normal (grade 0, similar to that of proximal femoral shaft; Fig. 1), mildly increased (grade I, more intense than that of proximal femoral shaft and similar to that of acetabulum; Fig. 2 ), or definitely increased (grade II, more intense than that of acetabulum; Fig. 3). This visual scoring analysis has been used in our nuclear medicine department for interpretation and research of bone scintigraphy to analyse foci of increased radio-uptake accurately. There was agreement in scores for most of the cases, and a consensus reading was derived for the few cases in which the initial score was not identical. A photon defect in the upper lateral portion of the femoral head was evaluated separately (Fig. 3(a)).

Fig. 1

Table 1 Characteristics of patients grouped according to the presence or absence of avascular osteonecrosis (AVN) Characteristic Age (years) Men/women Duration between transplantation and bone scintigraphy (months) Hip pain Bilateral Unilateral None

Total (n = 237)

AVN ( + ) (n = 15)

AVN ( – ) (n = 222)

40 ± 12 127:110 12.0 ± 1.1

42 ± 9 8:7 12.3 ± 1.5

40 ± 10 119:103 12.7 ± 1.1

5 7 225

5 5 5

0 2 220

Data are as mean ± SD. AVN ( + ): patients with AVN; AVN ( – ): patients without AVN.

A patient with grade 0 bone scintigraphy and no avascular osteonecrosis. (a) Bone scintigraphy shows the normal range of radioactivities in bilateral femoral heads. (b) A T1-weighted magnetic resonance image of both hips shows normal findings.

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Bone scan of hip AVN 1 year after renal transplantation Lee et al. 921

Fig. 2

Table 2 Incidence of avascular osteonecrosis (AVN) and information for the ‘gold standards’ for a diagnosis of AVN Incidence of AVN Patient-based Hip-based Bilateral AVN (patient-based) Gold standards of AVN diagnosis (hipbased) Histopathological examination Core decompression Total hip replacement Imaging modalities Simple radiography MRI

A patient with grade I bone scintigraphy and avascular osteonecrosis of bilateral hips. (a) Bone scintigraphy shows mildly increased radioactivities in bilateral femoral heads. (b) T1-weighted magnetic resonance image demonstrates irregular hypo-intense lesions with dark signal intensity rims involving the anterosuperior aspects with associated flattening of bilateral femoral heads. (c) Plain X-ray images demonstrate a suspicious irregular radiolucent focus with marginal sclerosis in the right femoral head, without definite evidence suggesting avascular osteonecrosis in either hips.

Fig. 3

6.3% (15/237) 4.9% (23/473) 53.3% (8/15)

12 3 9 11 2 9

diagnosed histopathologically through core decompression in three femoral heads and total hip replacement arthroplasty in nine. In the remaining 11 hips, imaging studies including plain radiography (n = 2) and MRI (n = 9) confirmed the diagnosis of AVN (Table 2). Among the remaining 450 femoral heads, 73 showed normal appearance in radiologic studies (plain radiography only in 31, MRI only in 30, both in 12). Another 377 femoral heads did not have any radiological study at the time of bone scintigraphy. However, all of the 450 femoral heads did not show any clinical and/or radiological evidence of AVN during the follow-up period. Of the 473 femoral heads, 17 were associated with hip pain at the time of bone scintigraphy whereas the remaining 456 were asymptomatic. AVN was confirmed in 15 of the 17 painful hips and eight of the 456 asymptomatic hips (Fig. 4). Bone scintigraphy

Bone scintigraphy demonstrated the level of radioactivity in the 473 femoral heads to be grade 0 in 335, grade I in 123, and grade II in 15. A photon defect in the femoral head was seen in six of grade I and nine of grade II cases.

A patient with grade II bone scintigraphy and avascular osteonecrosis of bilateral hips. (a) Bone scintigraphy shows definitely increased radioactivity with a typical photon defect in the right hip, and mildly increased radioactivity in the left hip. (b) T1-weighted magnetic resonance images shows geographic areas of dark signal intensity in both femoral heads. (c) Plain X-ray images do not show any definite evidence of avascular osteonecrosis or joint space narrowing in bilateral hips.

Results Incidence of avascular osteonecrosis

Femoral head AVN was detected in 15 of 237 patients (6.3%), and 23 of 473 femoral heads (4.9%). Disease was bilateral in eight of the 15 patients (53.3%). AVN was

Of the 23 femoral heads confirmed to have AVN, two showed grade 0 radioactivity, eight (three painful and five painless hips) were seen as grade I radioactivity, and 13 had grade II radioactivity (12 painful and one painless hip; Fig. 4). A photon defect was observed in 2/8 and 9/13 femoral heads with AVN showing grade I and II radioactivities, respectively. Of a total of 450 femoral heads without AVN, 333 (two painful and 331 painless hips) had grade 0 radioactivity, 115 had grade I radioactivity (all painless), and two (all painless) had grade II radioactivity. A photon defect was seen in 4/115 femoral heads with grade I radioactivity (Fig. 4).

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922 Nuclear Medicine Communications 2006, Vol 27 No 11

Fig. 4

A total of 473 femoral heads Grade 0: 335 (0) Normal: 333 (0)

Grade I: 123 (6)

AVN: 2 (0)

Normal: 115 (4)

Grade II: 15 (9)

AVN: 8 (2)

Normal: 2 (0)

AVN: 13 (9)

No pain: 331

No pain: 2

No pain: 115(4)

No pain: 5(1)

No pain: 2

No pain: 1(0)

Pain: 2

Pain: 0

Pain: 0

Pain: 3(1)

Pain: 0

Pain: 12(9)

Breakdown of bone scintigraphy results. Numbers in parenthesis denote number of femoral heads demonstrating a typical photon defect in the upper lateral portion.

Test sensitivity, specificity and accuracy

When grade I and grade II activities were used as positive criteria, bone scintigraphy had a sensitivity of 91.3% (100% with hip pain and 75% without), specificity of 74% (100% with hip pain and 73.9% without), and accuracy of 74.8% (100% with hip pain and 73.9% without) for the AVN diagnosis.

Fig. 5

(a) 100

100 75.0

75

100

100

91.3 74.0

73.9

74.8

73.9

50

When only grade II activity was considered positive, the rates were 56.5% (80% with hip pain and 12.5% without), 99.5% (100% with hip pain and 99.6% without), and 97.5% (82.6% with hip pain and 98% without), respectively (Fig. 5). The presence of a typical photon defect had a low sensitivity of 47.8%, although the specificity and accuracy were high (99.1%, and 96.6%, respectively).

Discussion AVN of the femoral head is a serious post-transplant musculoskeletal complication with incidence rates that vary in reported series. A prior retrospective study in 1976 reported AVN as frequently as 41% post-transplant [4]. The incidence of femoral head AVN was decreased as much as 20% in more recent prospective studies using MRI. In 1991, Kopecky and associates [9] discovered osteonecrosis in 22% of 64 patients at 12 months after transplantation. In 1997, Kubo et al. [8] documented femoral head MRI abnormalities of osteonecrosis in 25% of 51 renal allograft recipients. Marston et al. [10] reported 20% of 52 patients and 11% of 103 hips developed AVN of the femoral head within 1 year after transplantation. Most recently, however, Lopez-Ben et al. [16] reported 4% of 48 patients and three of 96 hips had AVN of the femoral head within 6 months after renal transplantation. In our study, AVN of the femoral head developed in 6.3% of the 237 patients and 4.9% of the 473 femoral heads from 8 months to 16 months after renal transplantation. These rates are apparently similar to

25 0 Sensitivity(%)

Specificity(%)

Accuracy(%)

Total

Hip pain(+)

No hip pain

(b) 99.5 100 99.6

100

98.0

97.5 82.6

80.0 75 56.5 50 25

12.5

0 Sensitivity(%)

Specificity(%)

Accuracy(%)

Total

Hip pain(+)

No hip pain

Hip-based sensitivity, specificity and accuracy of bone scintigraphy. (a) Grade I and grade II activities were used as positive criteria. (b) Only grade II activity was considered as positive for avascular osteonecrosis.

those in the most recent study by Lopez-Ben et al. [16] and lower than those in other previously reported studies [4,8–10]. If we include the five bilateral AVN patients who were excluded in this study because AVN developed

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Bone scan of hip AVN 1 year after renal transplantation Lee et al. 923

before the bone scintigraphy, the rates increase as 8.3% of the 242 patients and 6.8% of the 483 femoral heads. Of the total of 20 patients who developed AVN within 16 months after transplantation in our study, AVN was detected more than 9 months after transplantation in 75% of the cases. Although the exact mechanism remains controversial, corticosteroid therapy appears to occupy a central role in the pathogenesis of AVN and many studies in renal transplant patients have shown a relationship between steroid dosage and the prevalence of AVN of the femoral head. Lausten et al. [17], in a retrospective study of the medical records of 750 patients, showed an 11.2% incidence of symptomatic osteonecrosis with high-dose glucocorticoids (cumulative mean dose of prednisolone 12.5 g at 1 year post-transplant) and 5.1% with low-dose (cumulative mean dose of prednisolone average dose 6.5 g). The cohort of Kopecky et al. [9], with an osteonecrosis incidence of 22%, had a prednisolone equivalent dose of 7 ± 3.25 g prednisolone during the first 90 days after transplantation. The cohort of Lopez-Ben et al. [16], with an osteonecrosis incidence of 4%, had a slightly lower cumulative steroid dosage (2.1 g prednisolone at 3 months after transplantation) compared to previously reported cohorts. Although the immunosuppressive regimens are difficult to compare due to different steroids used and different follow-up periods reported, these previously reported cohorts with higher incidences of AVN appear to have had a slightly higher cumulative steroid dosage than our current cohort. And in our study, there was no difference of corticosteroids dosage between the patients with and without AVN. On bone scintigraphy images, the earliest and most evident finding of AVN is a photon defect in the femoral head due to impaired vascular supply of the femoral head [18,19]. This occurs prior to the radiographic evidence of AVN [18]. Recently, Ryu et al. [20] reported that bone SPECT of the hip region was more sensitive than MRI in the detection of early AVN of the femoral heads after renal transplantation and should be used to diagnose AVN, particularly in patients with hip pain but normal radiographic findings after renal transplantation. On planar scintigraphic images, a photon defect in the femoral head may be obscured by the acetabular and other surrounding bone activity. Using SPECT, it is possible to separate the femoral head from other overlying bony structures. However, it takes more time and cost to acquire SPECT images than planar images. In our study, photon defects were seen in 11 of 23 hips (47.8%) with AVN and all of them were of normal appearance on plain hip radiography at that time of bone scintigraphy. After 3–4 months, four of these hips became abnormal on X-ray images. Of a total of 17 painful hips, 15

with AVN showed mildly (grade I, n = 3) or definitely (grade II, n = 12) increased radioactivities. The remaining two painful hips without AVN showed normal radioactivity and were also free from any clinical and radiological evidence of AVN during the follow-up period of 3.5 and 4.7 years, respectively. Thus, for the diagnosis of AVN in symptomatic patients at risk for the disease, planar bone scintigraphy is sufficient, since in our experience all symptomatic hips were correctly diagnosed when grade I and grade II activities were used as positive criteria regardless of the presence or absence of photon defects. In reported series, the interval between transplantation and the onset of symptoms of femoral head AVN, such as hip pain, varies between 5 and 126 months, with a mean of 9–19 months [3,21–24]. In our study, only eight of the 23 hips (35%) with femoral head AVN were asymptomatic at the time of diagnosis. The time of onset of hip pain in out patients ranged from 9 to 15 months post-transplantation (mean interval, 12.3 ± 1.8 months). In a prospective study of patients with renal transplant followed with 6-monthly bone scintigraphies scans for 5 years, the most common sites of AVN involvement were femoral heads, often bilaterally, followed by the knees, humeral heads and elbows [25], and LaPorte et al. [26] reported multifocal AVN in 32 of 1056 (3%) patients. In their study, renal transplantation was one of the associated factors. In our study, however, AVN in sites other than femoral head was not found on bone scintigraphy. In addition, none of the patients showed any clinical or radiological abnormalities in other sites during clinical follow-up. In this study, radionuclide angiography, pin-hole collimator images, SPECT images were not acquired because of the time pressure of gamma cameras in our institute. Quantitative analysis was not used in this study because visual analysis is the usual method of bone scintigraphy analysis in our nuclear medicine department. It should be noted that the use of such techniques may influence the sensitivity, specificity and accuracy of bone scintigraphy for diagnosing femoral head AVN. In summary, we found a low incidence of femoral head AVN in renal transplantation patients at the time of 1 year post-transplantation, which seems to be related with low cumulative steroid dosage of our current cohort. Of the femoral head AVN identified on bone scintigraphy, 75% appeared more than 9 months after transplantation. Planar bone scintigraphy can adequately diagnose femoral head AVN in symptomatic patients at risk using mildly or definitely increased radioactivities of the femoral head regardless of photon defects as positive criteria.

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924 Nuclear Medicine Communications 2006, Vol 27 No 11

Acknowledgement

13

This work was supported, in part, by the Samsung grant SBRI C-A6-419-1, Korea.

14

References

15

1

Mankin HJ. Nontraumatic necrosis of bone (osteonecrosis). N Engl J Med 1992; 326:1473–1479. 2 Farge D, Parfrey PS, St Andre C, Kuo YL, Guttmann RD. Aseptic necrosis following renal transplantation: a 25-year experience. Transplant Proc 1985; 17:1947–1950. 3 Elmstedt E. Avascular bone necrosis in the renal transplant patient: a discriminant analysis of 144 cases. Clin Orthop 1981; 158:149–157. 4 Hawking KM. Avascular necrosis of bone after renal transplantation [Letter]. N Engl J Med 1976; 294:397. 5 Gillespy 3rd T, Genant HK, Helms CA. Magnetic resonance imaging of osteonecrosis. Radiol Clin North Am 1986; 24:193–208. 6 Castro Jr FP, Barrack RL. Core decompression and conservative treatment for avascular necrosis of the femoral head: a meta-analysis. Am J Orthop 2000; 29:187–194. 7 Fink B, Degenhardt S, Paselk C, Schneider T, Modder U, Ruther W. Early detection of avascular necrosis of the femoral head following renal transplantation. Arch Orthop Trauma Surg 1997; 116:151–156. 8 Kubo T, Yamazoe S, Sugano N, Fujioka M, Naruse S, Yoshimura N, et al. Initial MRI findings of non-traumatic osteonecrosis of the femoral head in renal allograft recipients. Magn Reson Imaging 1997; 15:1017–1023. 9 Kopecky KK, Braunstein EM, Brandt KD, Filo RS, Leapman SB, Capello WN, et al. Apparent avascular necrosis of the hip: appearance and spontaneous resolution of MR findings in renal allograft recipients. Radiology 1991; 179:523–527. 10 Marston SB, Gillingham K, Bailey RF, Cheng EY. Osteonecrosis of the femoral head after solid organ transplantation: a prospective study. J Bone Joint Surg Am 2002; 84A:2145–2151. 11 Lavernia CJ, Sierra RJ, Grieco FR. Osteonecrosis of the femoral head. J Am Acad Orthop Surg 1999; 7:250–261. 12 Nakamura T, Matsumoto T, Nishino M, Tomita K, Kadoya M. Early magnetic resonance imaging and histologic findings in a model of femoral head necrosis. Clin Orthop Relat Res 1997; 334:68–72.

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Conway WF, Totty WG, McEnery KW. CT and MR imaging of the hip. Radiology 1996; 198:297–307. Kokubo T, Takatori Y, Ninomiya S, Nakamura T, Kamogawa M. Magnetic resonance imaging and scintigraphy of avascular necrosis of the femoral head. Prediction of subsequent segmental collapse. Clin Orthop Relat Res 1992; 277:54–60. Markisz JA, Knowles RJ, Altchek DW, Schneider R, Whalen JP, Cahill PT. Segmental patterns of avascular necrosis of the femoral heads: early detection with MR imaging. Radiology 1987; 162:717–720. Lopez-Ben R, Mikuls TR, Moore DS, Julian BA, Bernreuter WK, Elkins M, et al. Incidence of hip osteonecrosis among renal transplantation recipients: a prospective study. Clin Radiol 2004; 59:431–438. Lausten GS, Lemser T, Jensen PK, Egfjord M. Necrosis of the femoral head after kidney transplantation. Clin Transplant 1998; 12:572–574. Collier Jr BD, Hellman RS, Krasnow AZ. Bone SPECT. Semin Nucl Med 1987; 17:247–266. Collier BD, Carrera GF, Johnson RP, Isitman AT, Hellman RS, Knobel J, et al. Detection of femoral head avascular necrosis in adults by SPECT. J Nucl Med 1985; 26:979–987. Ryu JS, Kim JS, Moon DH, Kim SM, Shin MJ, Chang JS, et al. Bone SPECT is more sensitive than MRI in the detection of early osteonecrosis of the femoral head after renal transplantation. J Nucl Med 2002; 43:1006–1011. Susan LP, Braun WE, Banowsky LH, Straffon RA, Bergfeld JA. Avascular necrosis following renal transplantation. Experience with 449 allografts with and without high-dose steroid therapy. Urology 1978; 11:225–229. Ibels LS, Alfrey AC, Huffer WE, Weil 3rd R. Aseptic necrosis of bone following renal transplantation: experience in 194 transplant recipients and review of the literature. Medicine (Baltimore) 1978; 57:25–45. Parfrey PS, Farge D, Parfrey NA, Hanley JA, Guttman RD. The decreased incidence of aseptic necrosis in renal transplant recipients–a case control study. Transplantation 1986; 41:182–187. Gottlieb MN, Stephens MK, Lowrie EG, Griffiths HJ, Kenzora J, Strom TB, et al. A longitudinal study of bone disease after successful renal transplantation. Nephron 1978; 22:239–248. Spencer JD, Maisey M. A prospective scintigraphic study of avascular necrosis of bone in renal transplant patients. Clin Orthop 1985; 201: 125–135. LaPorte DM, Mont MA, Mohan V, Jones LC, Hungerford DS. Multifocal osteonecrosis. J Rheumatol 1998; 25:1968–1974.

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NEWS AND VIEWS NOVEMBER 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + )44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF Tel: ( + )44 (0)113 2066930, E-mail: [email protected] This month’s article is kindly contributed by Mrs Beth Gawthorpe, medical physicist at St James’ Hospital, Leeds. Nuclear Medicine Communications, 2006, 27, 925–926 Radioiodine treatments and homeland defence

The phone rings at 2 a.m. on a Sunday morning. Having successfully (on the second attempt) positioned the receiver near both your mouth and ear simultaneously you find yourself speaking to a slightly panicked American police officer. He suspects that a recent patient of yours is smuggling parts for a dirty bomb; she has produced a tattered yellow card from the depths of her handbag and is quietly trembling with thoughts of Guanta`namo Bay. According to the International Atomic Energy Agency, between 1993 and 2004, there were 650 confirmed cases of illicit trafficking in nuclear and radiological materials worldwide. Many cases involved material that could be used to produce a nuclear weapon or a device using conventional explosives with radioactive material, known as a ‘dirty bomb’. Ports and airports around the UK use radiation detectors to combat this threat (http://security.homeoffice. gov.uk/counter-terrorism-strategy/ border-security/cylclamen/) and the United States is ‘leading the way’ with detectors also fitted to buildings, subway stations and road tunnels. Security services are understandably reluctant to publish the sensitivity of their devices, but

anecdotal evidence collected as part of HSE Research Report 416 suggests that patients retaining less than 30 MBq may still be detected. Dr L Zuckier of New Jersey Medical School has constructed a radiation calculator to help in counselling patients on their risks of setting off ‘Homeland Security’ detectors (http: //njmstss.umdnj.edu/radalarm/). At a recent conference he claimed that a patient receiving 131I for an overactive thyroid could still trigger alarms up to 95 days after treatment. So your hapless patient hadn’t lied to you when you asked if she was travelling soon after her thyroid treatment, she hadn’t thought that 3 months was ‘soon’. Most therapy patients are given a card explaining their treatment and restrictions, which seems to be gradually evolving into a ‘passport’ for their radioactivity. However, according to Dr Zuckier and further anecdotal evidence, patients who undergo imaging are also at risk of triggering alarms. So should we be handing out warning cards on the door to every patient? Maybe. Do the cards actually mean anything? Unfortunately, probably not, because any terrorist (or 8-year-old child for that matter) could forge one. 24-hour phone numbers are now included on many patient cards (especially in the US) to confirm that they are real, but

are we really to believe that radiation smugglers don’t have a friend who can do a passable impression of a medical physicist? Perhaps most worryingly, does having an over-active thyroid gland preclude a person from being a terrorist? If not, should we really be persuading immigration staff to admit them without checking that the level and distribution of activity is consistent with their treatment? So, in our world of escalating threats, paranoia and detection technology, what are we going to say when the phones start ringing? As we go to press there are indications that the anti-terrorist police unit and ARSAC are working on some recommendations which might lead to an answer in due course. Meeting Announcements Radiation, Research and Risk

Date: 9 November 2006 Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT, UK Website: www.bir.org.uk International Conference on Quality Assurance and New Techniques in Radiation Medicine (QANTRM)

Dates: 13–15 November 2006 Venue: Vienna, Austria Website: www.pub.iaea.org/MTCD/ Meetings/ Announcements.asp?ConfID = 146

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926 Nuclear Medicine Communications 2006, Vol 27 No 11

Design of Radionuclide Facilities with Reference to Radiation Protection (Revisited)

British Nuclear Medicine Society – Annual Conference

Date: 14 November 2006 Venue: British Institute of Radiology, 36 Portland Place, London, W1B 1AT, UK Website: www.bir.org.uk

Dates: 19–21 March 2007 Venue: Manchester, UK Website: www.bnms.org.uk

Nuclear Medicine Technologists Meeting

Dates: 29 April to 2 May 2007 Venue: Prague, Czech Republic Website: http://www.icnc8.org

Date: 16 November 2006 Venue: Queen Elizabeth Hospital Postgraduate Centre, Birmingham, UK Website: www.ipem.ac.uk British Nuclear Cardiology Society – Annual Conference

Date: Monday 4 December 2006 Venue: National Heart and Lung Institute, London, UK Website: www.bncs.org.uk

ICNC 2007: 8th International Conference of Nuclear Cardiology

Education and Training EANM Learning Courses

Dates: Weekend courses throughout 2006 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on + 43 1 2128030,

fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected]

EANM distance learning in nuclear cardiology

This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. Website: http://www.eanm.org/edu Online/edu_start.php?navId = 332

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Editorial 123

I- Metaiodobenzylguanidine in chronic heart failure: is there a clinical use?

Duncan W. Motherwell, Mark C. Petrie, William Martin and Stuart M. Cobbe Nuclear Medicine Communications 2006, 27:927–931 Department of Medical Cardiology, Glasgow Royal Infirmary, Glasgow, UK.

Sponsorship: D.W.M. and S.M.C. are supported by the British Heart Foundation. Received 29 September 2006 Accepted 20 October 2006

Correspondence and requests for reprints to Duncan W. Motherwell, Department of Medical Cardiology, Glasgow Royal Infirmary, 16 Alexandra Parade, Glasgow G31 2ER, UK. Tel: + 44 (0)141 2114592; fax: + 44 (0)141 2320492; e-mail: [email protected]

Introduction 123

I-Metaiodobenzylguanidine (MIBG), a metabolically inactive structural analogue of noradrenaline, has been used extensively in the investigation of cardiac disease subsequent to its introduction 25 years ago [1]. Despite numerous studies yielding further insight into the role of the sympathetic nervous system in the regulation of cardiac function, the agent has not achieved routine use in clinical cardiology.

line, but is not metabolised by monoamine oxidase or catechol-O-methyl transferase, which would normally break down noradrenaline [1,6]. This presynaptic uptake allows the distribution of the myocardial sympathetic nervous system to be established scintigraphically in the heart. A strong correlation between MIBG myocardial uptake and noradrenaline content in endomyocardial biopsy samples has been reported [11].

Myocardial kinetics and imaging

The correct timing of MIBG imaging after injection is unclear. MIBG can be taken up by neuronal and nonneuronal tissue, so to image the sympathetic nervous system using MIBG, non-neuronal uptake must be excluded. Nakajo et al. [6], in a rat model using reserpine to pharmacologically block uptake-1 function, identified that early uptake at 10–15 min is predominantly via the non-neuronal uptake mechanism (uptake-2). The authors suggested that late uptake (4 h or later) would better determine neuronal uptake of MIBG, when neuronal and non-neuronal uptake is approximately equal. This will then represent noradrenaline reuptake and hence the integrity of the sympathetic nerve [6]. Dae et al. [12] performed MIBG imaging in cardiac transplant patients expecting to find normal early non-neuronal uptake but no evidence of neuronal uptake. Surprisingly, there was no uptake of MIBG in the transplanted heart, suggesting that uptake-2 function into non-neuronal tissue is negligible in man. All of these patients were receiving steroid medication, which is known to interfere with uptake-2 function, making these data difficult to interpret. The above discordance has led to a difference in timing of MIBG imaging in the literature, with the majority of centres measuring both early and late parameters.

MIBG, given intravenously, leaves the blood and is taken up by the sympathetic nerve terminal by the uptake-1 mechanism, which clears the synapse of noradrenaline following stimulation of the sympathetic nerve. MIBG is then stored presynaptically in a similar way to noradrena-

Measuring early and late MIBG uptake allows the measurement of myocardial washout or washout rate (WR). This may provide functional information about the sympathetic nervous system, although it has not been

Chronic heart failure (CHF), of both ischaemic and nonischaemic aetiology, is an increasing health burden to the Western world, particularly with increased survival rates following myocardial infarction. Medical and device management of CHF has improved the quality of life and survival for many patients, with transplantation reserved for more severe cases. MIBG has been proposed as a tool to assess prognosis [2]. However, the subsequently published literature has been largely overtaken by advances in medical therapy with the introduction of angiotensin-converting enzyme inhibitors (ACEI) and b-blockers [3–5]. The majority of original work using MIBG originates from Japan [6–10] and, although many areas of cardiology have been investigated, the prognostic studies in CHF have involved small numbers of patients (Table 1). The lack of a standard imaging protocol in terms of timing of acquisition, collimator selection, use of scatter correction and method of image quantification, makes inter-study comparison difficult. To date, no large multicentre trial has been conducted.

c 2006 Lippincott Williams & Wilkins 0143-3636

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928 Nuclear Medicine Communications 2006, Vol 27 No 12

Table 1

Prognostic studies comparing MIBG indices with other parameters n

Non-ischaemic (%)

Merlet et al. [2] (1992)

90

73

22

Cohen-Solal et al. [29] (1999) Momose et al. [10] (1999) Ogita et al. [30] (2001)

93 59 79

74 100 43

Ebina et al. [31] (2002)

62

Gerson et al. [32] 2003 Yamada et al. [33] (2003)

37 65

100 37

De Milliano et al. [23]; 2002

58

47

26.5

Anastasiou-Nana et al. [34] (2005)

52

48

31

64.5

Mean LVEF

BB (%)

Loop diuretic (%)

Endpoint

Follow-up, mean ± SD (months)

Multivariate predictor of endpoint

9

91

Death

27

25 28 29

13 34 25

92 N/A N/A

10 ± 8 24.9 ± 13.2 31

42

29

58

Death ± transplant Cardiac death Death or hospitalization with CHF Cardiac death or hospitalization with CHF

HMR (late)a LVEF CTR LVEDD VO2a WRa WRa

27 28

24 40

92 76

48.8 ± 8.6 34 ± 19

100

94

Cardiac death or transplant Death or hospitalization with CHF Cardiac death or transplant

29

100

Death

24

16.2

37

HMRa ANP BNP HMR (early)a WRa VO2a Plasma NA Exercise time Walk distance LVEDD HMR (early)a LVEF

a Best predictor of endpoint; N/A, not available; HMR, heart to mediastinum ratio; LVEF, left ventricular ejection fraction; CTR, cardiothoracic ratio; LVEDD, left ventricular end diastolic dimension; VO2, oxygen capacity at maximal exercise; WR, washout rate; ANP, atrial natriuretic peptide; BNP, B type natriuretic peptide; CHF, chronic heart failure; NA, noradrenaline.

shown to correlate with noradrenaline spillover, which is considered to be the gold standard for assessing sympathetic function. The exact mechanism of the release of MIBG is not well understood. MIBG uptake can be measured both globally and regionally. Heart to mediastinal ratio (HMR), measured early or late, is the global measure of MIBG uptake. Using tomography, it is possible to obtain information about regional sympathetic function, although there is often a relative lack of counts making analysis difficult. Both of these methods are limited by extra-cardiac uptake of MIBG by the liver and lungs, resulting in scatter that interferes with cardiac assessment. This is particularly relevant for the HMR, where estimates of myocardial background should be employed, because it may be comparable with myocardial activity in the low uptake state of patients with CHF. This would lead to significant overestimates of HMR in these patients. A further problem with HMR is the debate regarding which collimator to employ due to scatter of high energy ( > 300 keV) photons falling in the 159-keV imaging window. This makes the results obtained from low energy and medium energy collimators not directly comparable, although Kobayashi et al. [13] have suggested that imaging in an adjacent window immediately above the principal window can compensate for this scatter. This has been confirmed by Small et al. [14] who recommend the use of scatter correction in all quantitative work with 123I. New methods of analysis are required to provide an accurate assessment of MIBG uptake by the heart. A

better understanding of the kinetics of MIBG would allow standardization of the correct timing and imaging protocol.

MIBG parameters following therapeutic intervention The majority of previous cardiology research with MIBG has been in the field of CHF. Patients with non-ischaemic CHF have globally and regionally reduced uptake with increased washout of MIBG [11,15]. This is thought to represent a dysfunction in uptake-1 function, denervation and sympathetic nerve over-activity. Although there are a large number of studies in this area, small numbers of patients are examined, often in comparison with a small number of ‘normals’. Patients with non-ischaemic CHF have heterogeneous uptake, and HMR correlates with left ventricular ejection fraction (LVEF), New York Heart Association (NYHA) class, plasma noradrenaline, and endomyocardial biopsy 123 I-MIBG activity [11,15,16]. CHF due to coronary artery disease has been less well characterized using 123I-MIBG imaging. Increasing WR correlates with worsening NYHA functional class and increased plasma noradrenaline [7]. Cardiac sympathetic nerve dysfunction in CHF has been assessed after therapeutic intervention. There is evidence of an improvement in MIBG parameters illustrating a reversible component, and a recovery of sympathetic nerve integrity and function. Patients with non-ischaemic CHF have been examined before and during medical

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123

I-MIBG in chronic heart failure: is there a clinical use? Motherwell et al. 929

therapy, but those with ischaemic CHF have not been investigated. Although there are some inconsistencies in the literature, ACEI b-blockers and spironolactone have all been reported to improve global MIBG uptake and can reduce WR [17–20]. No studies demonstrate that the change in MIBG correlates with the change in LVEF. Whether the recovery of MIBG parameters following the introduction of medical therapy is related to an improvement in haemodynamics or a primary recovery in sympathetic function is unknown. Cardiac resynchronization therapy (CRT), which improves haemodynamics and can induce reverse remodelling, may provide the answers. ErolYilmaz et al. [21] reported an improvement in MIBG indices and LVEF after 6 months of CRT in 13 patients. However, changes in regional MIBG uptake have not been correlated with corresponding changes in regional wall motion. There is some evidence that MIBG can be used to predict the response to medical therapy in CHF. In nonischaemic dilated cardiomyopathy, Suwa et al. [22] demonstrated that MIBG uptake could predict those patients who had a clinical response to b-blocker therapy. There is so far no evidence that MIBG uptake can predict those patients who will sustain an improvement in LVEF after b-blocker in CHF of any aetiology. Certainly, no patient should be excluded from prognostically beneficial medication on the basis of MIBG uptake.

Prognostic value of MIBG A clinical application of MIBG could be to determine the prognosis of patients with chronic heart failure and assist in the selection of patients for cardiac transplantation. MIBG must improve on the powerful prognostic information gained from the current investigations employed. In 1992, Merlet et al. [2] reported MIBG as the most powerful independent prognostic marker in heart failure patients of both ischaemic and non-ischaemic aetiology. Late MIBG uptake was able to predict survival more powerfully than LVEF. However, only 9% of these patients received bblocker medication, which is now standard treatment of heart failure, and has been demonstrated to improve survival in several large scale trials [3,4]. By contrast, several subsequent studies have shown that MIBG uptake, although useful, is inferior on multivariate analysis to other parameters such as VO2 max and B type natriuretic peptide, particularly in dilated cardiomyopathy. As shown in Table 1, the prognostic studies comparing MIBG with other indices predominantly examine small numbers of patients with non-ischaemic heart failure, with few patients on regular diuretic therapy or b-blockers. When an assessment of MIBG is followed by at least 6 months of b-blockade, the prognostic value of MIBG is lost, whereas VO2 remains predictive of events [23]. This is not surprising given the improvement in MIBG parameters which is seen after b-blockade. This would suggest there is

little to be gained by performing MIBG imaging in patients with CHF prior to the introduction of this disease modifying medication. Fujimoto et al. [24] have performed the first study of MIBG imaging on patients established on b-blocker for 6 months. MIBG parameters of WR, extent score and severity score were predictive of cardiac death or admission to hospital. LVEF was not predictive of outcome in this study. There may be a prognostic role for MIBG imaging in patients who are established on b-blocker medication, although further studies with larger patient numbers and ischaemic heart failure are required. This would obviously require comparison with the current available tools.

New directions The indications for inserting an implantable cardioverter defibrillator (ICD) have expanded with the publication of MADIT-II and SCD-HeFT [25,26]. ICDs improve mortality when used following a myocardial infarction with associated reduction in LVEF and in patients with chronic heart failure and a reduced ejection fraction (LVEF < 35%). This expansion in the criteria for ICD insertion has placed an enormous burden on the health care systems of many countries, to a level that cannot be sustained. An additional problem, yet to be addressed, is how to identify those patients who will never require defibrillation and thus avoid the potential complications, expense and follow-up. A pathophysiological relationship between arrhythmogenesis and the sympathetic nervous system has been postulated, and MIBG imaging [27] has been viewed as a possible route to screen patients who might benefit from ICD insertion. Arora et al. [28] performed myocardial 123I-imaging and heart rate variability (HRV) analysis on patients with an ICD previously inserted. Patients in this study had both ischaemic and non-ischaemic aetiology, with the majority having had at least one ICD discharge. Patients who received a subsequent discharge had a reduced global early uptake of 123I-MIBG and more extensive defects on tomographic imaging. Heart rate variability analysis was also useful in determining those patients who received a discharge, although the greatest predictive benefit was seen when the two were combined. This study looked at a small patient sample of 17, and the individual groups were not directly comparable. Higher left ventricular ejection fractions and a higher rate of ACEI and b-blocker use were seen in those patients who did not have an ICD discharge, suggesting a lower risk group. A larger prospective study is required to assess the potential use of 123I-MIBG as a screen for ICD insertion possibly with the addition of heart rate variability measurements and QRS duration.

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Nuclear Medicine Communications 2006, Vol 27 No 12

Conclusion Despite significant research in the area of CHF, MIBG cardiac imaging remains a research tool looking for an application. A better understanding of the kinetics of MIBG is required before universal agreement is possible on the details of imaging. Debate about the timing of imaging, which collimator to use or whether to image with planar or tomography has diluted and confused the currently available literature. As in many areas, progress requires standardization of imaging techniques and analysis.

10

11

12

13

There are two possible areas where MIBG may be used clinically. Studies have suggested that global analysis of noradrenaline re-uptake may provide prognostic information, although these have been overtaken by advances in medical therapy. However, when patients are taking currently available medical therapy including b-blockers, MIBG may assist in decision making, and this is the most likely area for potential use. The identification of patients with sympathetic nervous system dysfunction who benefit from an ICD in CHF is a second area in which MIBG could be used. The one study in this area has suggested that MIBG may be able to identify patients who would not require this expensive treatment.

14

15

16

17

18

19

A large multi-centre trial with a standardized imaging protocol is required, examining MIBG uptake in CHF, including patients with ischaemic CHF. By examining large numbers of patients, it may be possible to finally ascertain if anything useful is to be gained from MIBG and, if not, it may be consigned to history.

20

21

22

References 1

2

3 4

5

6

7

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9

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Original article

Two-year follow-up in 150 consecutive cases with normal dopamine transporter imaging Vicky L. Marshalla, Jim Pattersonb, Donald M. Hadleyc, Katherine A. Grosseta and Donald G. Grosseta Background and aims Functional pre-synaptic dopamine brain imaging is generally abnormal when parkinsonism is degenerative (such as in idiopathic Parkinson’s disease) and normal in patients with non-degenerative movement disorder (such as essential tremor). However, some patients diagnosed as early Parkinson’s disease have normal presynaptic dopamine imaging. Follow-up of patients with normal imaging should help determine whether such patients truly have degenerative parkinsonism (and therefore represent false negative imaging results), or emerge as cases of non-degenerative parkinsonism (and therefore represent initial clinical over-diagnosis of Parkinson’s disease). Methods and results One hundred and fifty cases with normal 123I-FP-CIT SPECT undertaken during routine care over a 3-year period were reviewed 2.4 years (interquartile range, 2.2–3.1 years) after SPECT. Diagnosis after follow-up was non-degenerative parkinsonism or tremor in 146 (97%), who did not progress clinically, and degenerative parkinsonism in four (3%), in whom clinicial progression was noted. Anti-Parkinson therapy was used in 36, and withdrawn in 27 with no deterioration in 25. Patients strictly fulfilling Brain Bank criteria (part 1) were more likely to

Introduction Misdiagnosis of Parkinson’s disease (PD) occurs both early [1] and late [2,3] during the clinical course. It is documented both in community [4] and specialist settings [2]. Mimics of Parkinson’s disease include other types of degenerative parkinsonism (e.g., multiple system atrophy, progressive supranuclear palsy) in which presynaptic dopaminergic neurones are abnormal, and nondegenerative disorders (e.g., essential tremor, vascular parkinsonism, drug-induced parkinsonism) in which presynaptic dopaminergic neurones are normal. Diagnostic application in patients with clinically uncertain parkinsonism is widely reported [5–7] using radioligands which bind to the dopamine transporter such as N-ofluoropropyl-2b-carboxymethoxy-3b-(4-[123I]iodophenyl) nortropane (123I-FP-CIT) and single photon emission computed tomography (SPECT). Such imaging differentiates patients with established degenerative parkinsonism from cases of essential tremor [8,9] and blinded prospective assessment in early disease shows increased congruence over time to the baseline image result [10,11].

undergo a trial of anti-Parkinson therapy (P < 0.05) but were no more likely to maintain or respond to anti-Parkinson therapy than those not fulfilling criteria. Conclusion The clinical profile and therapy response during follow-up of patients with normal presynaptic dopamine imaging supports the diagnosis of a non-degenerative movement disorder in nearly all c 2006 Lippincott cases. Nucl Med Commun 27:933–937 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:933–937 Keywords: Parkinson’s disease, essential tremor, drug-induced parkinsonism, vascular parkinsonism, parkinsonism, diagnosis, dopamine transporter, 123I-FP-CIT SPECT, levodopa Departments of aNeurology, bClinical Physics and cNeuroradiology, Institute of Neurological Sciences, Southern General Hospital, Glasgow, UK. Correspondence to: Dr Vicky Marshall, Institute of Neurological Sciences, Southern General Hospital, 1345 Govan Road, G51 4TF, UK. Tel: + 0044 141 201 1100; fax: + 0044 141 201 2510; e-mail: [email protected] Received 16 May 2006 Accepted 22 July 2006

Theoretically, however, SPECT may test normal in early Parkinson’s disease [12]. Between 4 and 15% of patients diagnosed as early Parkinson’s disease in clinical trials of anti-Parkinson medication have normal presynaptic dopamine imaging [13–15]. Limited follow-up data leaves the question open as to whether these are a form of Parkinson’s disease [13], other degenerative parkinsonian syndrome or have a benign tremor disorder with some parkinsonian features [15]. This study assessed clinical features, the use of antiParkinson therapy, and the response during follow-up of patients with normal 123I-FP-CIT SPECT undertaken during routine clinical care.

Methods Consecutive patients with normal 123I-FP-CIT SPECT undertaken in a regional neuroradiology unit from October 2000 to December 2003 were reviewed. Assessment during a minimum follow-up of 2 years after imaging noted clinical features, progression, working clinical diagnosis and use and response to anti-Parkinson

c 2006 Lippincott Williams & Wilkins 0143-3636

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medication. A review of case records was supplemented whenever necessary by contacting the treating neurologist and/or general practitioner. SPECT was applied because of clinical uncertainty about the presence or absence of underlying presynaptic dopaminergic deficiency. 123I-FP-CIT SPECT was acquired using a multidetector system (NeuroFocus, NeuroPhysics Corp, Shirley, Massachusetts) which acquires 10–12 sequential single transaxial sections 5 mm apart through the basal ganglia. Semiquantitative assessments were performed using predetermined region of interest (ROI) templates from Talairach Daemon [16,17]. Specific:non-specific ratios were calculated from three adjacent axial slices through the ROIs. The occipital region was used as reference. Visual grading was described as previously [8]. Focal reductions suggesting a structural lesion were considered non-degenerative since they did not show the characteristic pattern of asymmetrical putamen greater than caudate loss [18]. Data are presented as mean (standard deviation, SD) when normally distributed, otherwise median (interquartile range, IQ). Chisquared analysis was used to test differences of antiParkinson therapy use between groups.

Results Of 462 123I-FP-CIT SPECT scans undertaken, 150 fulfilled the study criteria and had available follow-up records (a further 30 with normal scans had incomplete follow-up and are not included). SPECT was requested by 16 hospital specialists and visual assessment was normal in 146 (97%) and showed focal reduction in four (3%). Semiquantitative analysis showed radioligand uptake within 2 SDs of age-matched normals in all striatal subregions, apart from the four scans with a focal reduction. The mean age was 63 years (SD 12); 84 were female (56%) and 66 male (44%). Patients had movement disorder symptoms for 2.7 years (IQ 1.3–5.9) at imaging. Final case record review was 2.4 years (IQ 2.2–3.1) after SPECT and the median follow-up duration from first clinic attendance was 3.2 years (IQ 2.5–4.3). Whilst uncertainty occurred in all cases, the favoured documented diagnosis pre-scan was non-degenerative disorder in 73 (49%) and degenerative parkinsonism in 44 (29%), while in 33 (22%) no favoured diagnosis was documented. Clinical features

The predominant clinical feature was tremor in 112 (75%) and gait abnormality in 18 (12%); the remaining cases had mixed features. Documented clinical features are outlined in Table 1. Twenty-five patients fulfilled part 1 of the United Kingdom Brain Bank criteria, while an additional 37 had incomplete fulfilment through the presence of only possible features. Considering more specific criteria for Parkinson’s disease [19], seven

Table 1 Presence and certainty of clinical features in 150 patients with normal 123I-FP-CIT SPECT Definite

Possible

Bradykinesia

29

41

Rigidity

36

31

10 43 62

0 0 1

16

7

Tremor

Postural instability

Rest only Postural only Rest and postural

One site only, e.g., reduced arm swing n = 31 An additional two had spasticity and one had myotonia

patients fulfilled these through the presence of cardinal features, asymmetrical onset, and absence of any other possible causes of a parkinsonian syndrome. Anti-Parkinson therapy was used in 36 (levodopa in 30, of whom seven also had used dopamine agonists; combinations of anticholinergics, amantadine, dopamine agonists and selegiline without levodopa were used in six). This figure of 36 included 23 of the 44 subjects (51%) who had a pre-scan diagnosis of degenerative parkinsonism. Medication was used prior to SPECT in 34 cases and afterwards in two. Anti-Parkinson therapy was later withdrawn in 27 of 36 (75%), who had either no therapy response (10) or an uncertain or unsustained response (17) but restarted in two of those 27 with clinical worsening (worse Unified Parkinson’s Disease Rating Scale motor score in 1; during hospital admission with terminal acute-on-chronic liver failure and uncertain benefit in one). In the remaining 25 patients there was no deterioration off anti-Parkinson therapy. Anti-Parkinson therapy withdrawal is planned in two of the remaining cases, not planned in six, and not documented in one. Table 2 presents anti-Parkinson therapy use and effect of any withdrawal according to criteria fulfilment. Patients strictly fulfilling Brain Bank criteria (part 1) were more likely to undergo a trial of antiParkinson therapy (P < 0.05) but were no more likely to maintain or respond to anti-Parkinson therapy than those not fulfilling criteria (P = 0.95). Structural brain imaging was performed in 99 (computed tomography, 53; magnetic resonance, 57; both, 11) which showed atrophy alone in nine, vascular changes in 40 and miscellaneous abnormalities in five. Of the four patients with focal striatal reduction in 123I-FP-CIT uptake, only one had a matching vascular putamenal lesion on computed tomography (CT) (diagnosis: VP and unclassified degenerative parkinsonism), repeat SPECT in the second patient became normal after 2.5 years (diagnosis: VP and DIP), the third had no structural disease on CT (diagnosis: non-degenerative parkinsonism) and the last

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Follow-up of patients with normal DAT imaging Marshall et al. 935

[18,21,22] and in anosmic relatives of patients with Parkinson’s disease [23]. Theoretically, a small proportion of scans may initially be normal in patients with early idiopathic Parkinson’s disease [12], or have a normal or near-normal scan due to unmasking of Parkinson’s disease by dopamine depleting drugs [24]. In our series, 3% with normal SPECT retained a diagnosis of degenerative parkinsonism and those cases had a 3-year symptom duration when SPECT was performed; none was on dopamine depleting medication. Repeat scanning after an interval is a possible approach in such cases. At least some scans should become abnormal if there is degenerative parkinsonism, since the progressive loss of striatal uptake in early Parkinson’s disease is 3–13% per annum [25–29]. Since the time period of this study we found all of 22 normal SPECT scans which were repeated (15 through participation in a study protocol and seven because of remaining clinical uncertainty) to remain normal in 21, at an interval of 18 months (IQ 17–19) (Marshall et al., unpublished observations). The other subject had repeat imaging at 3.3 years when new reduced focal putamenal uptake was noted to match a vascular lesion on CT. Two subjects with a remaining diagnosis of degenerative parkinsonism were within this group even though repeat imaging showing no evidence of emerging parkinsonian dopaminergic loss.

patient with a left striatal focal reduction had evidence of widespread vascular disease including the right striatum (diagnosis: VP). The diagnosis after follow-up was nondegenerative parkinsonism or tremor in 146 cases (Fig. 1). In four patients, the clinician considered that clinical progression was in keeping with a diagnosis was degenerative parkinsonism (one, Parkinson’s disease; one, corticobasal degeneration; and two, unclassified).

Discussion Presynaptic dopaminergic imaging is a potential adjunct to differentiate degenerative parkinsonism from its mimics in cases of clinical uncertainty [6,7,11,20]. It is abnormal in early and preclinical disease, e.g., in the asymptomatic striatum in unilateral Parkinson’s disease Use of anti-Parkinson medication in 150 patients with normal 123I-FP-CIT SPECT, according to presence of definite clinical features Table 2

Criteria (total number fulfilling)

Brain Bank criteria part 1 (25) Brain Bank part 1 plus asymmetry and no exclusions (7) Bradykinesia plus rigidity and rest tremor (11)

Use of anti-Parkinson therapy

Used and maintained

Used and worse on withdrawal

Used and stopped without deterioration

Never used

2

0

9

14

1

0

0

6

2

0

4

5

Repeat normal imaging at up to 4 years was identified in three trials of anti-Parkinson therapy in between 4 and 15% of patients enrolled as early Parkinson’s disease [13–15]. These have been termed subjects with scans without evidence of dopamine deficiency (SWEDDs). The progressive loss of striatal uptake seen in Parkinson’s disease is not seen in SWEDDs [30], and expert video

Fig. 1

Number of patients

75

50

25

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O

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n-

ta

ou

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pd

ia no

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at

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g

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m

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0

Final diagnosis in 150 patients with normal

123

I-FP-CIT SPECT.

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Table 3 Summary of available data on subjects entered as Parkinson’s disease into neuroprotective trials with repeat normal dopaminergic imaging Trial

Normal scan/PD diagnosis (SWEDDs)

Interval of repeat normal imaging (months)

Outcome/interpretation of SWEDDs

Not stated Considered ‘These patients may receive an alternative diagnosis such as essential tremor’ [15] Levodopa (150–600 mg
day – 1) response was poor [13] Blinded video review: 7 of 13 (54%); SWEDD’s thought unlikely to have PD versus 3 of 72 (14%) with abnormal baseline scans [31] Progression: SWEDDs showed no change in putamen uptake (unlike cases with abnormal scans) [30]

%

n

CALM-PD-CIT [14] REAL-PET [15]

4 11

3 of 78 21 of 183

46 24

ELLDOPA [13]

15

21 of 142

48

SWEDDs: subjects with scans without evidence of dopaminergic degeneration.

review blind to scan results challenged the diagnosis of Parkinson’s disease in most cases [31] (Table 3). Misdiagnosis also occurred in earlier clinical trials with 8.1% of 800 patients, after follow-up for 8 years, undergoing diagnostic change [1] which included nondegenerative disorders and was supported by postmortem in selected cases. In 100 patients diagnosed clinically as Parkinson’s disease, post-mortem examination showed misdiagnosis in 25 [2] of whom 12 would be expected to show normal presynaptic dopaminergic imaging. Stricter clinical criteria would reduce the misdiagnosis rate in those with normal SPECT to 1%, but at the expense of increased diagnostic uncertainty in patients with Parkinson’s disease [3]. There is limited post-mortem verification of functional dopaminergic imaging results. An 80% correlation of the 123 I-FP-CIT SPECT diagnosis with post-mortem was found in 10 patients with dementia [32]. In one of the mismatch cases, a unilateral putamen vascular lesion was misinterpreted as early degenerative parkinsonism. In the other, a patient with post-mortem verified corticobasal degeneration had radioligand uptake within the study defined boundaries for normality although on retrospective visual assessment, the image was felt to show symmetrical reduction of uptake [32]. Different patterns of abnormalities are reported in ‘Parkinson’s disease plus’ disorders, for example a diffuse [33] rather than asymmetrical reduction, and caudate loss preceding putamen loss. Combining visual with semiquantitative assessments may help in difficult cases, but a small proportion of cases will have a non-diagnostic scan appearance. Dopamine transporter imaging results have a strong diagnostic influence [6], and for this reason clinical follow-up is important to determine whether degenerative parkinsonism emerged clinically and/or anti-Parkinson therapy is introduced. Our results agree with blinded studies in uncertain parkinsonism, in which there is increased congruence with a baseline scan result over

3 [10] to 6 [11] months. In our series, eight (5%) (six with a working diagnosis of a benign disorders, and two with a working diagnosis of degenerative parkinsonism) did not obtain a definitive diagnosis during follow-up. Unclassifiable parkinsonism at a rate of 4–5% in specialist clinics is already reported [34], and in that study a poor or equivocal response to anti-Parkinson medication was the main reason for uncertainty. The therapeutic approach in patients with normal 123 I-FP-CIT SPECT and clinical features of a movement disorder is worth review. Firstly, to identify clinical features of patients who may have a false negative scan, i.e., where idiopathic or degenerative parkinsonism is present, and benefit might accrue from anti-Parkinson therapy. Secondly, to assess what proportion of cases are commenced on anti-Parkinson therapy, the clinical response, and effects of any withdrawal. Patients strictly fulfilling Brain Bank criteria (part 1) were more likely to undergo a trial of anti-Parkinson therapy but were no more likely to remain on therapy compared to those who did not fulfil criteria. The results suggest a closer correlation between 123I-FP-CIT SPECT and therapy response than with criteria fulfilment, but require comparative data from patients with abnormal scans.

Conclusion The emerging clinical profile of lack of clinical progression in line with degenerative parkinsonism and lack of response to anti-Parkinson medication or repeat normal 123 I-FP-CIT SPECT imaging where this has been applied suggests that nearly all patients with normal baseline 123 I-FP-CIT SPECT scan have a benign movement disorder rather than degenerative parkinsonism. A subset of patients with normal 123I-FP-CIT SPECT fulfils clinical criteria for Parkinson’s disease, but even in these cases there is negligible response to anti-Parkinson therapy, which is therefore seldom maintained.

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Gibb WRG, Lees AJ. The significance of the Lewy body in the diagnosis of idiopathic Parkinson’s disease. Neuropath Appl Neurobiol 1989; 15:27–44. Booij J, Speelman JD, Horstink MW, Wolters EC. The clinical benefit of imaging striatal dopamine transporters with [123I]FP-CIT SPET in differentiating patients with presynaptic parkinsonism from those with other forms of parkinsonism. Eur J Nucl Med 2001; 28:266–272. Filippi L, Manni C, Pierantozzi M, Brusa L, Danieli R, Stanzione P, et al. 123 I-FP-CIT semi-quantitative SPECT detects preclinical bilateral dopaminergic deficit in early Parkinson’s disease with unilateral symptoms. Nucl Med Commun 2005; 26:421–426. Marek K, Seibyl J, Zoghbi SS, Zea-Ponce Y, Baldwin RM, Fussell B, et al. [123I]beta-CIT/SPECT imaging demonstrates bilateral loss of dopamine transporters in hemi-Parkinson’s disease. Neurology 1996; 46:231–237. Berendse HW, Booij J, Francot CM, Bergmans PL, Hijman R, Stoof JC, et al. Subclinical dopaminergic dysfunction in asymptomatic Parkinson’s disease patients’ relatives with a decreased sense of smell. Ann Neurol 2001; 50:34–41. Kemp PM. Imaging the dopaminergic system in suspected parkinsonism, drug induced movement disorders, and Lewy body dementia. Nucl Med Commun 2005; 26:87–96. Marek K, Innis RB, Van Dyck CH, Fussell B, Early M, Eberly S, et al. [123I]beta-CIT SPECT imaging assessment of the rate of Parkinson’s disease progression. Neurology 2001; 57:2089–2094. Winogrodzka A, Bergmans P, Booij J, van Royen EA, Janssen AG, Wolters EC. [123I]FP-CIT SPECT is a useful method to monitor the rate of dopaminergic degeneration in early-stage Parkinson’s disease. J Neural Transm 2001; 108:1011–1019. Winogrodzka A, Bergmans P, Booij J, van Royen EA, Stoof JC, Wolters EC. [123I]beta-CIT SPECT is a useful method for monitoring dopaminergic degeneration in early stage Parkinson’s disease. J Neurol Neurosurg Psychiatry 2003; 74:294–298. Bruck A, Aalto S, Nurmi E, Vahlberg T, Bergman J, Rinne JO. Striatal subregional 6-[18F]fluoro-L-dopa uptake in early Parkinson’s disease: A two-year follow-up study. Mov Disord (published online) 2006. Nurmi E, Ruottinen HM, Kaasinen V, Bergman J, Haaparanta M, Solin O, et al. Progression in Parkinson’s disease: A positron emission tomography study with a dopamine transporter ligand [18F]CFT. Ann Neurol 2000; 47:804–808. Marek K, Jennings DL, Seibyl JP. Long-term follow-up of patients with scans without evidence of dopaminergic deficit (SWEDD) in the ELLDOPA study. Neurology 2005; 64(suppl 1):A274. Marek KL. Dopamine transporter imaging–challenging the evidence (platform presentation at the International Congress on Parkinson’s Disease and Related Disorders, Berlin, Germany). 2005. Walker Z, Costa DC, Walker RW, Shaw K, Gacinovic S, Stevens T, et al. Differentiation of dementia with Lewy bodies from Alzheimer’s disease using a dopaminergic presynaptic ligand. J Neurol Neurosurg Psychiatry 2002; 73:134–140. Filippi L, Manni C, Pierantozzi M, Brusa L, Danieli R, Stanzione P, et al. 123 I-FP-CIT in progressive supranuclear palsy and in Parkinson’s disease: a SPECT semiquantitative study. Nucl Med Commun 2006; 27:381–386. Katzenschlager R, Cardozo A, Cobo MRA, Tolosa E, Lees AJ. Unclassifiable parkinsonism in two European tertiary referral centres for movement disorders. Mov Disord 2003; 18:1123–1131.

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Original article

Discrimination between parkinsonian syndrome and essential tremor using artificial neural network classification of quantified DaTSCAN data David Hamiltona, Adrian Lista, Timothy Butlera, Stephen Hoggb and Martin Cawleya Background In the semi-quantitative assessment of DaTSCAN images, it has been suggested that the ratio of tracer accumulation in the putamen to that in the caudate nucleus may be helpful and could allow parkinsonian syndromes progression to be assessed. Separation of ratio values has been reported when early Parkinson’s disease is compared with essential tremor. The separation is lost, however, when the Parkinson’s disease is not early stage. Objectives To evaluate whether a two-stage analysis can differentiate between parkinsonian syndromes, of various stages, and essential tremor, and whether such a two-stage analysis can be undertaken in a single step using artificial neural networks (ANNs).

remaining six patients were then successfully classified into early parkinsonian syndromes and essential tremor. The ANN analysis successfully discriminated parkinsonian syndromes from essential tremor, in all patients, in a single step. Conclusions The two-stage process provides a method for classifying early disease without being compromised by the noise from non-early disease. The results of the single stage ANN analysis were very definite and it may be considered to have potential in the quantification of DaTSCAN images for clinical use. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:939–944 Nuclear Medicine Communications 2006, 27:939–944

Methods Data from 18 patients were analysed. Quantification was undertaken by manually drawing irregular regions of interest (ROIs): over each caudate nucleus and putamen and over an occipital cortex area near the posterior edge of the brain. A two-stage analysis was undertaken and was repeated, in a single step, using an ANN. Results The first stage, of the two-stage analysis, identified 12 patients with non-early parkinsonian syndromes. The

Introduction DaTSCAN (123I-FP-CIT, GE Healthcare) images are interpreted, visually, by assessing the shape of the striatum on the transverse image [1]. In essential tremor, tracer accumulation is similar in both the putamen and caudate nucleus, resulting in a comma shape [2]. In parkinsonian syndromes, loss of function is more pronounced in the putamen than in the caudate nucleus [3]. With increasing severity of parkinsonism, the striatal pattern shrinks like a ‘retreating comma’ [1], the tails of the comma shape become less pronounced and a circular full stop is eventually seen [2]. With further disease progression, the tracer accumulation in the caudate nucleus becomes evidently less [3,4]. Visual assessment is also particularly informative about asymmetrical tracer accumulation [5], important because of parkinsonism being typically unilateral in onset [1,6,7].

Keywords: DaTSCAN, quantification, artificial neural network, classification, essential tremor, parkinsonian syndromes Departments of aMedical Physics and bRadiology, County Hospital, Lincoln, UK. Correspondence to David Hamilton, Department of Medical Physics, Lincoln County Hospital, Greetwell Road, Lincoln LN2 5QY, UK. Tel: + 44 (0)1522 573674; fax: + 44 (0)1522 529858; e-mail: [email protected] Received 17 May 2006 Accepted 22 July 2006

Semi-quantitative assessment reinforces visual interpretation [3,4] and is essential to objectively assess striatal dopamine transporter binding [5]. It is most often based on the ratio of the activity in the striatum, caudate nucleus or putamen to that in the occipital cortex [2–6, 8–17]; the occipital cortex being used because it is devoid of dopaminergic receptors [18] but receives an equal supply of tracer [1]. Discrimination between parkinsonian syndromes and essential tremor has been documented [3,8,18], as has statistically significant putamen uptake asymmetry [2]. It has been suggested that consideration of the ratio of tracer accumulation in the putamen to that in the caudate nucleus may be helpful [5] and could allow disease progression to be assessed [3,4]. Separation of ratio values has been reported when early Parkinson’s disease is compared with normal control subjects [2,19],

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essential tremor [20] or progressive supranuclear palsy [14]. The separation is lost, however, when the Parkinson’s disease is not early-stage [2,16].

Fig. 1

A two-stage analysis may be a sensitive technique for both differentiating between parkinsonian syndromes and essential tremor, and for following the disease progression of parkinsonian syndromes. The first stage would be to determine non-early parkinsonian syndromes by assessing the striatum-to-occipital cortex tracer accumulation ratio (SOR). Patients with a low ratio would not be progressed to the second stage. The second stage would be to evaluate early parkinsonian syndromes by calculating the putamen-to-caudate tracer accumulation ratio (PCR). Such a two-stage analysis can be undertaken in a single step using artificial neural networks (ANNs). This is possible because ANNs use internal feature detection to discover multiple non-linear interactions among a set of variables, which permits a complex partitioning of the pattern space [21]. They have the advantage over traditional pattern recognition procedures in that they are taught by example, the decision rules are acquired rather than specified a priori and no features need be identified [22].

Method Data from 18 patients, referred for routine evaluation of possible parkinsonian syndromes and essential tremor, were analysed. Six were female, 12 were male and their ages ranged from 35 to 87 years. Each was administered 185 MBq DaTSCAN (123I-FP-CIT) and imaged using a Philips Solus dual-head gamma camera loaded with lowenergy, high-resolution collimators. The planar spatial resolution at 10 cm, through Perspex as scatter medium, averaged 8.5 mm full width at half maximum. The single photon emission tomography (SPET) acquisition and processing parameters were those established as being optimal, for this camera, by GE Healthcare during an onsite phantom study. They comprised a matrix size of 128  128 and a zoom of 1.0 giving a pixel size of 4.7 mm, a circular orbit with as small a radius of rotation as possible and typically < 17 cm, and a 3601 step-and-shoot acquisition over 128 azimuths at 30 s per stop. Reconstruction was undertaken using ADAC Pegasys version 4.20 clinical software running on a SUN Ultra 1 workstation and filtered back-projection with a Butterworth filter of order 9 and cut-off frequency of 0.5 (proportion of Nyquist frequency). This gave a transverse slice thickness of 4.7 mm. Analytical attenuation correction was not applied, as specified by GE Healthcare. Scatter correction was also not applied [17]. The single transverse slice that demonstrated the highest counts per pixel (c/p) in either striatum was identified and reorientation of the reconstructed data set, to obtain

Transverse image of reconstructed SPET DaTSCAN data, from one of the 18 patients, showing manually constructed regions of interest around the caudate nuclei, putamen and occipital cortex area.

optimum visualization of the striatum, was undertaken in both transverse and sagittal planes. An experienced observer visually classified each set of reconstructed SPET images as parkinsonian syndromes or essential tremor. This classification was taken as the ‘gold standard’ against which the results of quantification analyses were compared. Quantification was undertaken by manually drawing five irregular regions of interest (ROIs) on the single reoriented transverse slice that demonstrated the highest c/p in either striatum. The ROIs were constructed over each caudate nucleus and putamen. These were bilaterally symmetrical and divided approximately one third of the distance from the anterior to posterior of the striatum [23]. An ROI was also constructed over an occipital cortex area near the posterior edge of the brain (Fig. 1). This method is similar to the crescent analysis [18] provided in the QuantiSPECT (GE Healthcare) computer program [15]. The simplest method of ROI construction, i.e., completely freehand, was used. The ROIs were then

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ANN classification of parkinsonian syndrome and essential tremor Hamilton et al. 941

Fig. 2

Output from network

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Input into network Connection diagram of the artificial neural network (ANN), showing the four inputs, representing the counts per pixel (c/p) from the putamen and caudate nucleus regions of interest (ROIs) normalized to the c/p in the occipital cortex ROI. The ANN had one hidden layer with two neurons and one neuron in the output layer and, therefore, comprised 10 connections. The two hidden and one output neurons also had an additional bias value thus giving 13 adjustable variables for the network. W1 to W10: weights on neuron inputs; I1 to I4: input neurons; H1 to H2: hidden neurons; O: output neuron; WB: neuron biases.

translated to the two adjacent transverse slices. The mean c/p in each ROI over the three slices was recorded. Previously reported analyses [15,16] were undertaken. SOR involved calculating the ratio of the c/p in each striatum to the c/p in the occipital cortex area. PCR involved calculating the ratio of the c/p in each putamen to the c/p in the corresponding caudate nucleus. Both ratios were compared with the visual interpretation. A two-stage analysis was then undertaken. The SOR was used to identify non-early parkinsonian syndromes patients presenting low striatum c/p to occipital cortex c/p tracer accumulation ratios. The remaining patient data, those with high SOR ratios, were subject to PCR comparison with the visual interpretation. The two-stage analysis was then undertaken, in a single step, using a feed-forward back-propagation multilayer perception ANN. This software was developed in-house, using compiled C ++ , to run on Windows-based or Unixbased computers. It was validated by training and

testing with the XOR mathematical function. The ANN had four inputs, representing the c/p from the putamen and caudate nucleus ROIs normalized to the c/p in the occipital cortex ROI (Fig. 2). This normalization was applied in order to reduce variation amongst patients that would complicate ANN training. The ANN had one hidden layer with two neurons and an output layer comprising one neuron. This gave 10 weighted connections. Each hidden and output neuron also had an additional bias value thus giving the network 13 adjustable variables (10 weights + 3 biases). The ANN was trained using the gradient descent algorithm where output errors are fed back through the ANN to update all the relevant weights and biases. During training a target output of 0.01 was set for parkinsonian syndromes inputs, and a target of 0.99 was set for essential tremor inputs. Training continued until the network mean squared error (MSE) was < 0.01%. Because data were available from only 18 patients, the ANN was trained and tested using the leave-k-out, or

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Fig. 3

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Values of the striatum-to-cortex ratio (SOR) for each patient. These values considered the ratios of the c/p in each striatum to the c/p in the occipital cortex area, and discriminated from essential tremor in 14 patients but experienced overlapping values in 4 patients. The patient values, which overlap, are shown as rectangular icons for visual emphasis.

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The PCR values of the six patients with high SOR values. These values successfully discriminated between early parkinsonian syndromes and essential tremor. (Abbreviations as in the legends to Figs 4 and 5.)

Table 1

Results of the artificial neural network analysis for each

patient Fig. 4

0.95 0.9 0.85 0.8 0.75 0.7 0.65 0.6

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Values of the putamen-to-caudate ratio (PCR) for each patient. These values considered the ratios of the c/p in each putamen to the c/p in the corresponding caudate nucleus, and did not discriminate between parkinsonian syndromes and essential tremor. The patient values, which overlap, are shown as rectangular icons for visual emphasis.

‘round robin’, technique. In sequence, 17 patients were used to train the ANN and then the excluded patient was used to test the trained ANN. In each case a new ANN was created using randomly generated weights and biases. This was repeated 18 times so that all the data were considered.

Results The values of SOR, for each patient, are presented in Fig. 3. These values, which considered the ratios of the c/p in each striatum to the c/p in the occipital cortex area, discriminated parkinsonian syndromes from essential

Patient

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0.00 0.00 0.00 0.94 0.01 0.03 0.99 0.01 0.97 0.06 0.01 0.99 0.99 0.00 0.03 0.04 0.00 0.00

The table shows the number of training cycles required to achieve convergence, the results of visual interpretation of the images and the output of the trained network when it was tested. PS: parkinsonian syndromes; ET: essential tremor.

tremor in 14 patients but there was overlap of the values in four patients. The values of PCR, for each patient, are presented in Fig. 4. These values, which considered the ratios of the c/p in each putamen to the c/p in the corresponding caudate nucleus, did not discriminate between the two patient groups. The two-stage analysis, which involved using the SOR to identify non-early parkinsonian syndromes patients, identified 12 patients with ratios below 3. This level was chosen arbitrarily to include parkinsonian syndromes patients and to exclude essential tremor patients. The remaining six patients with high SORs, were then successfully classified into early parkinsonian syndromes and essential tremor using the PCR (Fig. 5).

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ANN classification of parkinsonian syndrome and essential tremor Hamilton et al. 943

The ANN analysis, which undertook the two-stage analysis in a single step, successfully discriminated parkinsonian syndromes from essential tremor in all patients (Table 1).

Discussion Before visual interpretation and quantification, reorientation of the reconstructed data set was undertaken in both transverse and sagittal planes. This is important as rotation in the sagittal plane can significantly affect basal ganglia/occipital ratios. The simplest form of ROI construction, i.e., completely freehand, was used in order to test the robustness of the ANN analysis. Using more advanced ROI construction techniques, such as thresholded levels, mirrored constructions or voxel-based analysis methods which can define regions of abnormal tracer retention more objectively, better operator reproducibility of SOR and PCR values would be expected and even better results may be obtained from the ANN analysis. The constructed ROIs were then translated to the adjacent two slices. This was done to improve the image statistics and to accommodate any residual uncorrected patient rotation. However, construction over a summed three-slice slab will be evaluated in future, as it may reduce construction noise and partial volume effects. Quantification of DaTSCAN images is widely used to reinforce visual interpretation but its effectiveness is not comprehensive across all disease severity states. Although SORs can discriminate between parkinsonian syndromes and essential tremor, it is difficult to classify disease severity in individual patients [3,7]. Visual interpretation relies more on the shape of the striatum image [1] than on the total tracer accumulation. However, the PCR is able to discriminate between parkinsonian syndromes and essential tremor or normal subjects only in early parkinsonian syndromes [2,19,20] and not over a range of disease severity states [2,16]. In early parkinsonian syndromes, tracer accumulation in the putamen is reduced whilst that in the caudate nucleus is maintained, hence PCR should be a sensitive indicator of disease progression. In more severe disease, however, tracer accumulation in the caudate nucleus also starts to reduce [3,4], which compromises the relationship between PCR and disease severity. A two-stage strategy, whereby non-early parkinsonian syndromes is detected initially by the SOR and only then is the PCR used to detect early parkinsonian syndromes, has been shown to be able to classify parkinsonian syndromes of various severities. SORs discriminated between parkinsonian syndromes and essential tremor in most patients but failed in four. The PCR could not effect discrimination in the whole

population, which demonstrates that the data comprised various disease states. However, when the patients demonstrating low SORs were removed from the population before PCR analysis, PCR successfully discriminated between parkinsonian syndromes and essential tremor in the remaining patients. This two-stage process should provide a method for classifying early disease without being compromised by the noise from non-early disease. A robust single-step process is clinically attractive and has less risk of data transcription errors. In this case, the ANN analysis clearly discriminated between parkinsonian syndromes and essential tremor in all patients with no equivocal results (Table 1). The results are so definite, on this small scale, that ANN analysis may be considered to have potential in the quantification of DaTSCAN images for clinical use. The technique could be of value in clinical trials where numerical data are required to correlate with clinical or other outcome measures. The strength of the technique lies in the fact that ANNs form a non-linear representation of the relationships between the input values and the expected output values. This internal representation does not necessarily match that of the experts and more analysis will be required to determine whether it takes into account, for example: SORs, PCRs and any asymmetry between left and right activity accumulations. Two limitations of the ANN training were identified. The ‘gold standard’ outcome was the judgement of a single experienced observer and the patient numbers in each disease category were not well balanced, with 13 parkinsonian syndromes patients and only five essential tremor patients. The latter would have presented a biased training population to the ANN and it would be expected that the trained ANN would then preferentially predict parkinsonian syndromes results. Even so, extremely good results were obtained. However, it would be expected that, by using more observers and with a more numerous and well balanced patient population, consistency in the results could be better ensured. An ANN with better training could be made into a webbased application for the nuclear medicine community to use, possibly as a learning aid to help those training to read the scans achieve ‘experienced’ standard. Development work along this route is now under way. More complicated ANNs can be imagined, comprising additional inputs and, possibly, outputs. Since striatal uptake is age dependent, uptake reducing with age, an input related to age may be able to reduce the bias of age dependency. An input to document the side of the tremor, and outputs to document the stage of the disease may also be considered. There is a possibility of variation in the results of the quantitative analysis of data obtained

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944 Nuclear Medicine Communications 2006, Vol 27 No 12

from different gamma camera types, although this has not been shown to be statistically significant in the type of analysis used here [15]. There may also be the possibility of variation among different centres with the same camera type. If necessary, an additional input to represent camera type, and possibly investigation site, could be implemented. With data from a number of sites, once trained, an ANN could possibly be used in any centre. It may be preferable to use one ANN, rather than an ANN for each site and camera type, because more patient data would then be available for training and there should be only subtle variations among sites and camera types.

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Baker MG, Brooks DJ, Burn DJ, Chaudhuri KR, Findley LJ, Grosset DG, et al. DaTSCAN–SPECT diagnostic imaging in the management of Parkinson’s disease. A consensus report. London: Parkinson’s Disease Society, 2001. Booij J, Tissingh G, Boer GJ, Speelman JD, Stoof JC, Janssen AG, et al. 123 I-FP-CIT SPECT shows a pronounced decline of striatal dopamine transporter labelling in early and advanced Parkinson’s disease. J Neurol Neurosurg Psychiatry 1997; 62:133–140. Staffen W, Mair A, Unterrainer J, Trinka E, Ladurner G. Measuring the progression of idiopathic Parkinson’s disease with 123I-FP-CIT SPECT. J Neural Transm 2000; 107:543–552. Habraken JB, Booji J, Slomka P, Sokole Busemann E, van Royen EA. Quantification and visualization of defects of the functional dopaminergic system using an automatic algorithm. J Nucl Med 1999; 40:1091–1097. Tatsch K, Asenbaum S, Bartenstein P, Catafau A, Halldin C, Pilowsky LS, Pupi A. European Association of Nuclear Medicine guidelines for brain neurotransmission SPET using 123I-labelled D2 transporter ligands. Eur J Nucl Med Mol Imaging 2002; 26:BP30–35. Asenbaum S, Brucke T, Pirker W, Podreka I, Angelberger P, Wenger S, et al. Imaging of dopamine transporters with [123I] b-CIT and SPECT in Parkinson’s disease. J Nucl Med 1997; 38:1–6. Staffen W, Mair J, Unterrainer J, Trinka E, Bsteh C, Ladurner G. [123I] b-CIT binding and SPET compared with clinical diagnosis in parkinsonism. Nucl Med Commun 2000; 21:417–424. Verhoeff NP, Kapucu O, Sokole-Busemann E, van Royen EA, Janssen AG. Estimation of dopamine D2 receptor binding potential in the striatum with iodine-123-IBZM SPECT: technical and interobserver variability. J Nucl Med 1993; 34:2076–2084. Kim SE, Lee WY, Choe YS, Kim JH. SPECT measurement of iodine-123beta-CIT binding to dopamine and serotonin transporters in Parkinson’s disease: correlation with symptom severity. Neurol Res 1999; 21:255–261.

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Benamer HT, Patterson J, Wyper DJ, Hadley DM, Macphee GJ, Grosset DG. Correlation of Parkinson’s disease severity and duration with 123I-FP-CIT SPECT striatal uptake. Mov Disord 2000; 15:692–698. Benamer TS, Patterson J, Grosset DG, Booji J, de Bruin K, van Royen E, et al. Accurate differentiation of parkinsonism and essential tremor using visual assessment of 123I-FP-CIT SPECT imaging: the 123I-FP-CIT study group. Mov Disord 2000; 15:503–510. Winogrodzka A, Bergmans P, Booij J, van Royen EA, Janssen AG, Wolters EC. [I123]FP-CIT SPECT is a useful method to monitor the rate of dopaminergic degeneration in early-stage Parkinson’s disease. J Neural Transm 2001; 108:1011–1019. Walker Z, Costa DC, Walker RW, Shaw K, Gacinovic S, Stevens T, et al. Differentiation of dementia with Lewy bodies from Alzheimer’s disease using a dopaminergic presynaptic ligand. J Neurol Neurosurgery Psychiatry 2002; 73:134–140. Antonini A, Benti R, De Notaris R, Tesei S, Zecchinelli A, Sacilotto G, et al. 123 I-Ioflupane/SPECT binding to striatal dopamine transporter (DAT) uptake in patients with Parkinson’s disease, multiple system atrophy and progressive supranuclear palsy. Neurol Sci 2003; 24:149–150. Morton RJ, Guy MJ, Marshall CA, Clarke EA, Hinton PJ. Variation of DaTSCAN quantification between different gamma camera types. Nucl Med Commun 2005; 26:1131–1137. Morton RJ, Guy MJ, Claus R, Hinton PJ, Marshall CA, Clarke EA. Comparison of different methods of DaTSCAN quantification. Nucl Med Commun 2005; 26:1139–1146. Popperl G, Radau P, Linke R, Hahn K, Tatsch K. Diagnostic performance of a 3-D automated quantification method of dopamine D2 receptor SPECT studies in the differential diagnosis of parkinsonism. Nucl Med Commun 2005; 26:39–43. Lokkegaard A, Werdelin LM, Friberg L. Clinical impact of diagnostic SPET investigations with a dopamine re-uptake ligand. Eur J Nucl Med 2002; 29:1623–1629. Seibyl JP, Marek KL, Quinlan D, Sheff K, Zoghbi S, Zea-Ponce Y, et al. Decreased single photon emission computed tomographic [123I] b-CIT striatal uptake correlates with symptom severity in Parkinson’s disease. Ann Neurol 1995; 38:589–598. Asenbaum S, Pirker W, Angelberger P, Bencsits G, Pruckmayer M, Brucke T. 123I-FP-CIT and SPECT in essential tremor and Parkinson’s disease. J Neural Transm 1998; 105:1213–1228. Dawson MRW, Dobbs A, Hooper HR, McEwan AJB, Triscott J, Cooney J. Artificial neural networks that use single-photon emission tomography to identify patients with probably Alzheimer’s disease. Eur J Nucl Med 1994; 21:1303–1311. Chan KH, Johnson KA, Becker JA, Satlin A, Mendelson J, Grarada B, Holman BL. A neural network classifier for cerebral perfusion imaging. J Nucl Med 1994; 35:771–774. Radau PE, Linke R, Slomka PJ, Tatsch K. Optimization of automated quantification of I123-IBZM uptake in the striatum applied to parkinsonism. J Nucl Med 2000; 41:220–227.

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Original article

Regional cerebral blood flow in Parkinson’s disease as an indicator of cognitive impairment Mirosława Derejkoa, Jarosław Sławekb, Dariusz Wieczorekc, Bogna Brockhuisd, Mirosława Dubaniewiczd and Piotr Lassd Objective To investigate the pattern of regional cerebral blood flow (rCBF) deficits in Parkinson’s disease patients in relation to cognitive decline and to assess the clinical usefulness of single photon emission tomography (SPET) scanning in differentiation between Parkinson’s disease patients with dementia and those without cognitive deficits. Methods We performed 99mTc-ECD SPET in 60 patients with idiopathic Parkinson’s disease (F: 25, M: 35), with average age of 68.4 years (SD ± 7.3, range 51–81 years). All patients were examined neurologically with the assessment of stage and severity of Parkinson’s disease (Hoehn–Yahr scale, UPDRS, Schwab–England scale). Detailed neuropsychological examination was performed in each Parkinson’s disease patient. On the basis of DSM-IV criteria of dementia and the results obtained in psychological examination, the whole group was divided into three subgroups: I, with no cognitive changes (n = 17); II, with mild cognitive impairment (n = 25); and III, with dementia (n = 18). Results There was noticeable significant decrease of perfusion in all areas in Parkinson’s disease patients when compared to the age-matched control group of healthy volunteers (n = 20). In group III, perfusion was significantly decreased (when compared to groups I and II), particularly in parietal and temporal areas with the predominance of the left side. Regression analysis revealed two

Introduction Dementia, one of the most common neuropsychiatric presentations of Parkinson’s disease, affects 20–40% of this patient population [1–3]. Although dementia as a feature of Parkinson’s disease is well recognized, its aetiology still remains unclear. More recently, it has become known that cognitive decline in Parkinson’s disease is attributable not only to sub-cortical pathway dysfunction, but also to cortical pathology. Diffuse Lewy bodies as well as Alzheimer-like abnormalities have been reported at neuropathological examination of Parkinson’s disease patients [4–6]. However, cognitive deficits in Parkinson’s disease have also been described in the absence of any cortical involvement, with ‘purely’ parkinsonian pathology [4,7]. The relative contribution of cortical and sub-cortical pathological changes to the development of dementia in Parkinson’s disease is

independent factors related to dementia: decrease of perfusion within left temporal lobe and its increase within left thalamus. Conclusion Parkinson’s disease patients with dementia showed left temporo-parietal hypoperfusion as compared to a group of patients without dementia, which resembles perfusion deficits described in Alzheimer’s disease. The hypoperfusion of the left temporal lobe with increase of rCBF within the left thalamus might be clinically useful in discrimination of Parkinson’s disease patients with dementia against those without cognitive impairment. c 2006 Lippincott Williams Nucl Med Commun 27:945–951 & Wilkins. Nuclear Medicine Communications 2006, 27:945–951 Keywords: Parkinson’s disease, cognitive impairment, dementia, regional cerebral blood flow, single photon emission tomography a Neurophysiology Department, Institute of Psychiatry and Neurology, Warsaw, Poland, bMovement Disorders and Functional Neurosurgery Department and Department of Neurological and Psychiatric Nursing, Medical University of Gdansk, Poland, cRehabilitation Department and dNuclear Medicine and Radiology Department, Medical University of Gdansk, Poland.

Correspondence to Dr Mirosława Derejko, Neurophysiology Department, Institute of Psychiatry and Neurology, Sobieskiego Street 9, 02-957 Warsaw, Poland. Tel: + 0048 224582713; fax: + 0048 226425375; e-mail: [email protected] Received 25 June 2006 Accepted 25 August 2006

uncertain. It is also unclear whether cognitive impairment in Parkinson’s disease represents a pathological process specific for the disease or if it is a result of the overlapping of other concomitant pathologies characteristic for advanced age, such as Alzheimer’s disease or diffuse Lewy bodies (DLBs). Multifactorial pathophysiology of dementia in Parkinson’s disease is also reflected by functional studies using single photon emission tomography (SPET) with the tracer 99m Tc-ethyl cysteinate dimer (99mTc-ECD). Previously reported studies have shown inconsistent patterns of regional cerebral blood flow (rCBF) deficits in patients with Parkinson’s disease and dementia (PDD). Bilateral hypoperfusion in frontal areas, parieto-temporal areas (a similar pattern to that described in Alzheimer’s disease), as well as global cortical hypoperfusion including occipital

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946 Nuclear Medicine Communications 2006, Vol 27 No 12

cortex (resembling the changes seen in DLB) were noticed [8–11]. Functional neuroimaging studies of the brain perfusion may help to characterize the topography and degree of rCBF deficits, providing adjunct support to the clinician in the diagnosis and management of PDD patients, but it also may give insight into the underlying pathology. The aim of this study was to investigate the pattern of rCBF deficits in Parkinson’s disease patients in relation to four cognitive decline (both in mild cognitive impairment and dementia) and to assess the clinical usefulness of SPET scanning in discrimination of PDD patients against those without cognitive deficits.

Methods Patient selection

We studied 60 patients (25 female and 35 male) with idiopathic Parkinson’s disease, whose intellectual function ranged from normal status to dementia. The average age of patients was 68.4 years (SD 7.3, range 51–81 years). The Parkinson’s disease patients were recruited from a community population of patients referred to regional outpatient movement disorder clinic by neurologists and general practitioners from the area of Gdansk, Poland. Parkinson’s disease was diagnosed according to the Parkinson’s Disease United Kingdom Brain Bank criteria [12]. Magnetic resonance imaging (MRI) was performed in all Parkinson’s disease patients. Patients were included only if cognitive abnormalities were not attributable to medication side-effects or causes other than idiopathic Parkinson’s disease, and when MRI did not suggest an alternative cause of parkinsonism (e.g. tumour or hydrocephalus). Normal, age-matched controls (12 females and 8 males) with average age of 66.9 ± 7.20 years were recruited among friends and spouses of patients included in this and other research studies. Neuropsychological assessment

Detailed neuropsychological examination was performed in the whole group of Parkinson’s disease patients with a test battery that included the Mini Mental State Examination (MMSE) [13], the Wechsler Adult Intelligence Scale – Revised (Vocabulary and Information subtests), Word Fluency Test, Rey Auditory Verbal Learning Test (AVLT) [14], Visual Learning and Memory Test [15], Tower of Toronto [16], dynamic apraxia tests (according to Luria: learning two motor sequences of three conflicting hand movements – alternating flexion, extension and rotation) [17–20]. The presence of depression was assessed with the Beck Depression Inventory [21]. On the basis of DSM-IV criteria of dementia and the results of the neuropsychological assessment, patients were divided into three groups: group I (17 patients), no cognitive deficits (MMSE score = 28–30, normal results of AVLT, visual learning and

verbal fluency tests); group II (25 patients), mild cognitive impairment (MMSE score = 24–27, and deficit in one of tests: AVLT, visual learning, verbal fluency); group III (18 patients), demented (MMSE score < 24 and deficit in at least one of the tests: AVLT, visual learning, verbal fluency or MMSE score > 23, but deficit in at least two other tests). Clinical assessment

The examination of motor functions was performed in the ‘on’ phase, when patients were able to cooperate. The assessment of motor functions included the Hoehn and Yahr disease stage (H–Y), the Unified Parkinson’s Disease rating Scale (UPDRS) with three sub-scales: activities of daily living (ADL–UPDRS part II), motor examination (UPDRS part III), complications of therapy (UPDRS part IV) and the Schwab and England Activities of Daily Living Scale (SE) [22–24]. The average duration of the disease in the whole group of Parkinson’s disease patients was 8.38 years (range, 1–27 years). The mean H–Y stage of patients was 2.73 (range, 1.5–5). The mean total UPDRS score obtained in the ‘on’ phase of examination was 48.61 ± 24.12. All demographic parameters of three groups of Parkinson’s disease patients are shown in Table 1. Single photon emission tomography

Regional CBF SPET scanning was performed 30 min following intravenous injection of 740 MBq 99mTc-ECD (FAM, Ło´dz´, Poland). Injection was performed in a quiet, dimly lit room with the patient seated facing a blank cream-coloured wall. Imaging was performed using a triple-headed gamma camera (Multi-SPET-3; Siemens, Erlangen, Germany) using a low-energy, ultra-highresolution, parallel-hole collimator. The data were collected into a 128  128 matrix, 4.3 mm per pixel, with a rotation angle of 1201, step-and-shoot acquisition, 128 steps, 20 000 counts/step. Uniform attenuation correction was not applied because of the limitations of the computer system (ICON, Siemens). The raw data were smoothed with a Butterworth filter with a cut-off frequency of 0.35 cycle/cm. The final data were displayed on a 10-grade colour scale. A semi-quantitative method of assessing regional tracer uptake on transverse slices was used. Regions of interest (ROIs) consisted of bilateral: prefrontal, frontal, parietal, temporal, occipital and subcortical (thalamus and striatum) regions (Fig. 1). The reference region used in this study was the cerebellum. Diffuse areas of decreased radiotracer uptake, usually seen in both hemispheres and detected by comparison to cerebellar uptake were considered as rCBF decrease. Statistical analysis

Statistical comparison of relative tracer activities between groups, as well as comparisons of some of the demographical data was made using one-way ANOVA F-test

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Regional CBF in Parkinson’s disease with cognitive impairment Derejko et al. 947

Comparison of clinical data of Parkinson’s disease patients in three subgroups: I, without cognitive impairment; II, with mild cognitive impairment; III, with dementia

Table 1

Characteristic

Group I, No cognitive impairment (n = 17)

Age (years) Disease duration (years) Age at onset (years) Hoehn–Yahr scale Schwab–Ehngland scale (%) UPDRS–II UPDRS–III UPDRS–IV UPDRS (II + II + IV) Mean dose of L-dopa (mg)

64.47 6.82 57.65 2.32 82.35 12.12 20.88 2.47 35.47 623.53

Group II, Mild cognitive impairment (n = 25)

(9.2)I–III (5.6) (9.2) (0.3)I–III (8.3)I–III (5.1)I–III (8.1)I–III (2.3)I–II, I–III (11.8)I–III (264)I–III

68.64 9.00 59.64 2.72 74.40 15.84 25.12 5.60 46.56 773.00

(6.8) (5.9) (9.4) (0.8) (17.1)II–III (9.2)II–III (16.2)II–III (3.9)II–I (26.3)II–III (433)II–III

Group III, Dementia (n = 18) 71.67 9.00 62.67 3.14 63.33 21.78 36.72 5.39 63.89 1027.78

F(2.57) or H(2,N = 60) values

P

4.90 3.78* 1.67 13.29* 16.80* 13.15* 11.54* 10.71* 15.07* 9.55*

< 0.05 NS NS < 0.05 > 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05

(3.3)III–I (4.2) (4.4) (0.7)III–I (17.2)III–I, III–II (7.4)III–I, III–II (13.8)III–I, III–II (3.1)III–I (21.9)III–I, III–II (397)III–I, III–II

*

Kruskal–Wallis (H) statistics were used to compare groups data which were not normally distributed. ANOVA (F) was used when analysing data with normal distribution (with the Scheffe test, for post hoc comparisons). Parkinson’s disease patients are divided into three groups: group I, without cognitive changes; group II, with mild cognitive impairment; and group III, with dementia. Indexes: I–II, I–III indicate pairs of subgroups that showed statistically significant (P < 0.05) differences. Note: values in columns I, II, III are means; SD within parentheses. NS, not significant.

Table 2 Regional cerebral blood flow (percent of cerebellar perfusion) in controls and Parkinson’s disease patients

Fig. 1

2

1

2

Area

2 1

2

2

1 2 2

6 5 4

3

3 3

3 3

3 3 4

7

3 7

3

1 - Prefrontal 2 - Frontal 3 - Parietal 4 -Temporal 5 -Thalamus 6 - Striatum 7- Occipital

3

The detailed composition of particular regions of interest (for Tables 2 and 3).

(Tables 2 and 3, and data with normal distribution in Table 1). Other group differences (data which were not normally distributed in Table 1) were assessed with the Kruskall–Wallis test. Kendall rank correlation coefficients were calculated to assess the magnitude of association between variables. Stepwise logistic regression analysis was used to identify the perfusion pattern that would allow classifying patients as demented (group III) or cognitively unimpaired (group I). The criterion of statistical significance was P < 0.05. The study was approved by the regional ethics committee at the Medical University of Gdansk, but due to the observational (not interventional) mode of the study, informed consent was not signed by patients.

Results The distribution of rCBF in the Parkinson’s disease patients showed significant differences when compared with controls in all ROIs (Table 2). In PDD patients (group III) the rCBF of the bilateral temporal and left parietal lobes was significantly reduced when compared

Right prefrontal Right frontal Right parietal Right temporal Right thalamus Right striatum Right occipital Left prefrontal Left frontal Left parietal Left temporal Left thalamus Left striatum Left occipital

Controls (n = 20) 92.75 94.94 96.11 104.93 86.75 88.15 115.40 90.98 94.74 95.13 104.65 85.80 89.55 115.40

(5.63) (5.67) (6.23) (6.37) (5.47) (7.25) (8.71) (5.07) (5.37) (5.94) (5.30) (5.86) (6.73) (7.35)

Patients (n = 60) 82.10 85.73 87.15 93.43 80.78 81.25 102.03 81.34 85.48 85.20 93.19 81.58 81.38 103.59

(7.20) (7.25) (6.92) (7.79) (8.48) (8.87) (8.06) (6.80) (7.35) (7.20) (7.88) (7.02) (10.08) (8.12)

F (1,78)

P

36.22 26.75 26.34 35.49 8.66 9.88 39.66 33.76 26.81 30.95 36.60 5.84 11.38 33.20

< 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05

Note: values for controls and Parkinson’s disease patients are means. SD within parentheses.

with Parkinson’s disease patients without cognitive deficits (group I) and patients with mild cognitive impairment (group II). The rCBF in the other ROIs did not show significant differences between subgroups of patients. Comparisons of rCBF are presented in Table 3. Despite significant differences between groups presented in Table 3, there was a partial overlapping of perfusion results, which could limit the potential suitability of our results for the individual diagnosis. In order to solve this problem, and find out what perfusion patterns are the most important in the identification of dementia patients, we used additional statistical analysis to differentiate between patients with dementia (group III) and patients without cognitive deficits (group I). The intermediate group (II), in which the cognitive status could not be classified either as entirely normal or demented, was excluded. We introduced the forward stepwise logistic regression model (probability of F to enter = 0.05 and probability of F to remove = 0.10). All the ROIs listed in Table 3 served

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948 Nuclear Medicine Communications 2006, Vol 27 No 12

Table 3

Comparison of regional cerebral blood flow (percent of cerebellar perfusion) in controls and subgroups of Parkinson’s disease

patients Area Right prefrontal Right frontal Right parietal Right temporal Right thalamus Right striatum Right occipital Left prefrontal Left frontal Left parietal Left temporal Left thalamus Left striatum Left occipital

Group 0 Controls (n = 20) 92.7 94.9 96.1 104.9 86.7 88.1 115.4 90.9 94.7 95.1 104.6 85.8 89.5 115.4

(5.6)0–I, 0–II, 0–III (5.6)0–I, 0–II, 0–III (6.2)0–I, 0–II, 0–III (6.3)0–I, 0–II, 0–III (5.4)0–I (7.2)0–I (8.7)0–I, 0–II, 0–III (5.1)0–I, 0–II, 0–III (5.3)0–I, 0–II, 0–III (5.9)0–I, 0–II, 0–III (5.3)0–I, 0–II, 0–III (5.8)0–I (6.7)0–III (7.3)0–I, 0–II, 0–III

Group I No cognitive Group II Mild cognitive impairment (n = 17) impairment (n = 25) 82.8 87.1 88.8 95.8 77.29 79.1 102.6 83.9 86.9 87.8 96.2 79.0 81.3 104.1

(6.3)I–0 (7.7)I–0 (5.8)I–0 (6.1)I–0, (9.3)I–0 (9.6)I–0 (8.3)I–0 (6.3)I–0 (6.3)I–0 (5.7)I–0, (5.2)I–0, (8.3)I–0 (10.3) (7.2)I–0

I–III

I–III I–III

82.38 86.64 87.23 95.24 81.80 82.44 104.10 80.84 86.05 87.12 94.96 82.84 84.36 103.92

Group III Dementia (n = 18)

(8.1)II–0 81.0 (7.3)II–0 83.1 II–0 (7.9) 85.4 II–0, II–III (7.9) 88.6 (7.74) 82.6 (7.18) 81.6 II–0 (7.78) 98.5 (7.59)II–0 79.5 (7.93)II–0 83.1 (6.90)II–0, II–III 80.0 (8.40)II–0, II–III 87.8 (6.47) 82.3 (8.10) 77.3 II–0 (9.66) 102.6

(7)III–0 (6.24)III–0 (6.3)III–0 (7.1)III–0, III–I, III–II (8.1) (10.2) (7.4)III–0 (5.5)III–0 (7.3)III–0 (6.4)III–0, III–I, III–II (6.8)III–0, III–I, III–II (6.1) (11.3)III–0 (6.8)III–0

F(3, 76)

P

12.1 10.4 9.5 17.4 4.6 3.8 15.4 13.08 9.96 18.21 19.95 3.19 6.06 10.96

< 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05 < 0.05

Control group is marked as ‘0’. Other details as in the footnote to Table 1.

Table 4 Results of stepwise logistic regression analysis conducted to determine regions of interest essential for the identification of dementia Variable Perfusion in the left temporal lobe Perfusion in the left thalamus Constants

B

Wald test

df

P

R

exp B

– 0.369

9.503

1

0.002

– 0.393

0.691

0.206

5.450

1

0.019

0.266

1.229

17.694

3.951

1

0.046

The cognitive status, estimated on the basis of the neuropsychological assessment results (group I (without cognitive impairment) versus group III (with dementia)) served as a dependent variable. All regions of interest (see Table 3) were included as independent variables. B, partial regression coefficient; R, regression correlation coefficient.

as independent variables. Classification of patients as demented or not demented obtained in neuropsychological examination served as the dependent variable. The results of regression analysis with the information necessary to calculate classifications for individual cases are presented in Table 4. Two out of all ROIs entered remained in the regression equation. Hypoperfusion of left temporal lobe (R = – 0.393) with additional hyperperfusion of left thalamus (R = 0.266) formed the pattern that enabled us to properly differentiate the greatest number of demented from not demented patients. The observed and predicted (in statistical sense) group membership, calculated from regression equation is shown in Table 5. For group I (Parkinson’s disease without cognitive changes), 16 (94.1%) patients were classified correctly; one patient was misclassified. In group III (patients with dementia), 16 out of 18 (88.9%) patients were classified properly. Analysis of the relationship between rCBF and the clinical factors showed the following significant (P < 0.05) correlations:

1. The duration of the disease had a significant negative correlation with rCBF in right parietal cortex (t Kendall = – 0.182); left parietal cortex (t Kendall = – 0.203); left prefrontal cortex (t Kendall = – 0.22); right occipital cortex (t Kendall = – 0.221);

Predictive value of regional cerebral blood flow SPET in terms of identification of dementia

Table 5

Prediction

Group membership determined by neuropsychological examination Group I Group III

Group membership determined by regression equation Group I

Group III

16 2

1 16

Correct classifications (%)

94.12 88.89

The observed and predicted group membership, as calculated from regression equation. Group 1: no cognitive impairment; group III: dementia.

2. The stage of disease measured according to the Hoehn–Yahr scale had a negative correlation with rCBF in right prefrontal cortex (t Kendall = – 0.237), left prefrontal cortex (t Kendall = – 0.274), right frontal cortex (t Kendall = – 0.228), left frontal cortex (t Kendall = – 0.255), left parietal cortex (t Kendall = – 0.341), left temporal cortex (t Kendall = – 0.221), right striatum (t Kendall = – 0.197).

Discussion Our study showed diffuse cortex reduction of ECD uptake in patients with Parkinson’s disease, when compared with age-matched controls. We also observed significantly greater bilateral temporal and left parietal

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Regional CBF in Parkinson’s disease with cognitive impairment Derejko et al. 949

decrease of rCBF in PDD patients as compared to the group without cognitive impairment. This pattern of perfusion deficits is called a ‘posterior’ type of hypoperfusion and resembles the one that is mostly described in Alzheimer’s disease patients. A clear distinction between demented and not demented Parkinson’s disease patients was established with the use of the stepwise logistic regression analysis. Hypoperfusion within left temporal cortex with additional hyperperfusion in the ipsilateral thalamus were the most significant factors attributable to cognitive decline and dementia. Changes in rCBF, observed in Parkinson’s disease patients, were associated with the stage according to H–Y scale and the duration of the disease. It is still difficult to interpret the rCBF data obtained in our study because of the low specificity of SPET results and insufficient knowledge of the mechanisms underlying them. The major determinant of cerebral hypoperfusion is thought to be a structural change with the loss of synapses due to the decreased numbers of cortical neurons, degeneration of axons or reduced dendritic branching [25]. Alternatively, cortical hypoperfusion could be the result of functional abnormalities unaccompanied by structural changes, e.g. loss of excitatory or inhibitory input from the subcortical structures. This phenomenon is described as deafferentiation or diaschisis. Cerebral regions receiving poor afferent signals become hypofunctioning, decrease their metabolism, and appear as low-uptake area in SPET images. In Parkinson’s disease several subcortical neurotransmitter systems, such as dopaminergic, noradrenergic, cholinergic, serotonergic, g-aminobutric acid (GABA-ergic), and peptidergic that project widely to the cortex are involved and might account for the diffuse changes in metabolism found in our study. The results of 11 previous studies on rCBF in Parkinson’s disease patients are inconsistent. Frontal, parietal, temporal, global cortical hypoperfusion or unchanged blood flow have been reported [8,11,25– 29]. The predilection of hypoperfusion to the left temporo-parietal region in PDD patients, observed in our study, is consistent with an observation made by Peppard et al. [25] and Tachibana et al. [27]. A ‘posterior’ type of hypoperfusion in PDD patients is also reported in other studies [26,28,29]. The most striking of our findings was the hyperperfusion of the left thalamus in PDD group. This may reflect the relative hyperfunction of that structure, which is known also from the studies on neurosurgical effects of thalamotomy or thalamic deep brain stimulation. To our knowledge, in previous studies this phenomenon has never been reported. Several investigators have described no significant differences in perfusion or glucose metabolism of thalamus between Parkinson’s disease patients with and without dementia [25–29]. However, a few previous studies have reported increased glucose metabolism in basal ganglia of Parkinson’s disease patients contralateral to the more affected

body side, and involved also patients with more advanced disease taking high doses of levodopa [26,30]. Eidelberg et al. suggested that dopaminergic denervation of the basal ganglia was related to an increased regional glucose metabolism [31]. We cannot also exclude the possibility of direct effect of pharmacological therapy on the regional cerebral responsiveness, which may be related to the motor complications in advanced Parkinson’s disease. The hypoperfusion differed significantly between group I (no cognitive impairment) and group III (dementia), but ‘transitional’ patterns were seen in group II, which may suggest that it is a group in which dementia might occur in the following years, but it requires a longer observational study. Our results of SPET study in PDD patients resemble those described in Alzheimer’s disease, except that defects were often unilateral, whereas predominantly bilateral defects are seen in Alzheimer’s disease. Although left-sided deficits might be more closely related to intellectual degradation and might have greatest predictive value for increasing cognitive impairment even in Alzheimer’s disease. The similar SPET patterns in PDD and Alzheimer’s disease might suggest that dementia in Parkinson’s disease may be the result of cortical pathology similar to Alzheimer’s disease. In a study by Jellinger et al. [32] a combination of the pathological changes seen in both Parkinson’s disease and Alzheimer’s disease has been observed in about 94% of PDD patients and in 30% with Parkinson’s disease without dementia. The degeneration of nucleus basalis of Meynert, which is the origin of the cholinergic fibres that project diffusely to the cortex, was initially found in Alzheimer’s disease, but has also been described in PDD, and cholinergic deficit is even more pronounced in PDD than in Alzheimer’s disease [33,34]. However, it is not clear if a chronic change in cortical metabolism is a secondary effect of nucleus basalis degeneration. A ‘posterior’ pattern of functional imaging abnormalities on SPET has been also observed in DLB [35]. In many cases the clinical picture of PDD resembles DLB, but according to current criteria, the diagnosis of DLB cannot be made in those who have had Parkinson’s disease symptoms preceding the cognitive decline for more than 12 months. Similarities have been described not only in motor phenotype, neuropsychological profile, positive response to cholinergic therapy, but also in neuropathological changes [36–38]. Apaydin et al. [6] in a clinicopathological study of 13 cases with PDD found pathological features of DLB in 12 of them [6]. Those results are consistent with observation made by other authors [5,39]. However, in contrast to Alzheimer’s disease and PDD, a very consistent finding from functional imaging studies in DLB is a profound occipital hypoperfusion [40]. On the other hand, Hughes et al. did not find any cortical pathology except that seen in

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Parkinson’s disease in 55% of examined cases with PDD [41]. These data may prove that dementia in Parkinson’s disease could be associated with natural progression of pathological changes subsequently from brain stem and subcortical areas to limbic and other cortical regions [42]. Schapiro et al. [43] made similar observations. Authors described a patient with parkinsonism and dementia who showed temporo-parietal hypometabolism on a PET scan. At autopsy the patient was confirmed to have Parkinson’s disease with no evidence of Alzheimer’s disease. These data suggest that reduced cortical uptake within temporo-parietal regions in PDD patients might have rather non-specific character [43]. Additionally, correlations detected in our study between cortical and subcortical rCBF deficits and stage of the disease in H–Y scale may indicate the reduced cortical metabolism in PDD patients as a result of disease progression and, thus, they are consistent with known data indicating that the duration of the disease and age are known risk factors for cognitive decline in Parkinson’s disease [3].

12 13

14 15

16 17 18

19

20

21 22

Conclusions

23

Data obtained in our study show the characteristic pattern of rCBF SPET in PDD patients. Left temporoparietal hypoperfusion and left thalamic hyperperfusion were found to be the most attributable to PDD regional blood flow pattern.

24

25

Our data suggest that perfusion changes observed PDD share a pattern of rCBF deficits similar Alzheimer’s disease and, thus, may also contribute our understanding of the pathogenesis of dementia Parkinson’s disease.

in to to in

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Original article

Can administration of metoclopramide reduce artefacts related to abdominal activity in myocardial perfusion SPECT? Thomas Gru¨ninga,b, Claudia Brogsitterc, Mehrdad Khonsaria, Ivor W. Jonesa, Sue M. Nevina and Wolfgang Burchertd Objectives Myocardial perfusion SPECT is frequently affected by artefacts related to abdominal activity. Metoclopramide has been suggested to relieve this, but two previous studies have shown conflicting results.

administration of metoclopramide, irrespective of whether it was given before the stress or rest scan, made no significant difference to inferior wall-to-abdomen count ratio.

Methods Ninety-five patients received 10 mg metoclopramide orally after injection of 99mTc-tetrofosmin for the stress scan and 86 patients had metoclopramide after their rest injection. A control group of 82 patients did not receive metoclopramide. Scans were evaluated visually by three readers.

Conclusion Neither qualitative nor quantitative analysis showed an effect of metoclopramide on abdominal activity in myocardial perfusion SPECT. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:953–957

Results Metoclopramide given before the stress scan led to abdominal activity being visually better in 16 scans, worse in 10, and unchanged in 67 scans, compared to the same patient’s rest scan without metoclopramide administration. Metoclopramide administered before the rest scan resulted in abdominal activity in 11 scans being visually better, in 19 worse, and 53 scans were deemed unchanged. These differences were not significant. The number of repeat stress or rest scans was not significantly different between patients who had received metoclopramide and those who had not. The

Introduction Myocardial perfusion scintigraphy is a widely available, non-invasive diagnostic method for assessing patients with ischaemic heart disease [1,2]. It is used to rule out or confirm haemodynamically significant ischaemic heart disease, characterize disease extent and severity, predict patients’ prognosis and assess response to treatment. The tracers 201T1, 99mTc-Tetrofosmin and 99mTc-Sestamibi, can be used, and stress can be applied either mechanically or pharmacologically with a number of agents. Scans produced with any of these protocols can have a number of well-recognized artefacts, including those related to biliary activity and abdominal activity close to the inferior left ventricular wall. A number of proposals have been made to overcome these artefacts, including: K Prone imaging to reduce or recognize inferior attenuation [3–5] K Strapping of breasts to reduce anteroseptal attenuation [6] K A fatty meal given between injection of the radiopharmaceutical and scanning to help clear the tracer from liver and gallbladder [7,8]

Nuclear Medicine Communications 2006, 27:953–957 Keywords: myocardial perfusion imaging, SPECT, artefact, metoclopramide

99m

Tc-tetrofosmin,

a Department of Nuclear Medicine, Derriford Hospital, Plymouth, UK, bPeninsula Medical School, Universities of Exeter and Plymouth, UK, cDepartment of Nuclear Medicine, University of Dresden, Germany and dInstitute of Molecular Biophysics, Radiopharmacy and Nuclear Medicine, Heart and Diabetes Center Bad Oeynhausen, Germany.

Correspondence to Dr Thomas Gru¨ning, Department of Nuclear Medicine, Derriford Hospital, Plymouth PL6 8DH, UK. Tel: + 0044 1752 792280; fax: + 0044 1752 517587; e-mail: [email protected] Received 6 June 2006 Accepted 21 July 2006

A substantial meal to increase stomach volume being given just before scanning [9,10] or the use of water [8] or iodinated oral contrast [11] to improve myocardialto-abdominal activity ratio K ECG-gated acquisition to help distinguish artefacts from true perfusion defects [4,12–15] K Pixel truncation to reduce the visual impact of activity in the gallbladder [16] K Supplemental exercise with pharmacological stress [17,18]. K

Pinkert et al. [19] reported in an observer-based study of 261 patients that the use of metoclopramide had a beneficial effect on abdominal activity. A later study by Weinmann and Moretti et al. [20] using a semiquantitative technique in 12 patients could not confirm this. Our study combines the methodologies employed for both previous papers and uses an improved study protocol in a large number of patients. It aims to conclusively determine whether metoclopramide is useful in reducing artefacts related to abdominal activity in myocardial perfusion SPECT.

c 2006 Lippincott Williams & Wilkins 0143-3636

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Methods Study design

This study is designed as a prospective, randomized trial in which each patient serves as their own control, as the study drug was given either before the stress or the rest scan. Allocation to either group was alternated on a weekly basis for reasons of logistical ease. Allocation of an individual patient to either group was, therefore, random. The study was open-label for patients, but readers were blinded. Approval from an ethics committee was obtained. Patients

Two-hundred and ninety-five patients routinely referred for a myocardial perfusion scan and judged to be suitable for adenosine stress were approached. A small number of asthmatic patients in whom dobutamine was used for stress were not included. Fifty-one patients declined to participate. The remaining 244 patients received 10 ml of 1 mg
ml – 1 metoclopramide solution orally immediately after tracer injection for either stress or rest scan. Forty patients who received metoclopramide after their stress injection did not require a rest scan due to a normal stress scan and were excluded for lack of a control scan. Twentythree patients who were scheduled to receive metoclopramide after their rest injection did not require a rest scan and were also excluded. One-hundred and eightyone patients received metoclopramide and completed a stress and rest scan. Of these, 95 patients received metoclopramide after their stress injection, and 86 after their rest injection. A control group comprised 82 patients who did not receive metoclopramide. They were routinely referred for a myocardial perfusion scan just prior to commencement of this study, and completed a stress and rest scan. Further details on all of these patients can be found in Table 1. Imaging protocol

A 6 min adenosine infusion (AdenoScan; Sanofi Winthrop, Guildford, UK) at 140 mg
kg – 1
min – 1 was used for the stress test, with 600 MBq 99mTc-tetrofosmin (Myoview; Amersham, Little Chalfont, UK) injected after the 4th minute of the adenosine infusion. Additional bicycle exercise of up to 20 W was used whenever possible, and this could be achieved by approximately 90% of patients. Table 1

Scanning could commence as early as 30 min after tracer injection, but was usually started 60 min post-injection (Table 1, right). A rest scan was only performed if the stress scan was judged to be visually abnormal, and was always performed on a separate day. Patients who were not already on nitrate medication and who had a blood pressure > 120/95 mmHg received a puff of 400 mg glyceryl trinitrate spray immediately before injection of 400 MBq 99mTc-tetrofosmin for the rest scan. Scanning was commenced as for the stress scan. A Varicam doubleheaded gamma camera equipped with low-energy highresolution collimators was used (General Electric Medical Systems, Slough, UK). Acquisition parameters were as follows: detectors at 901, 1801 rotation at 3 degree; steps with automatic body contouring, 20 s (ECG-gated stress scans)/30 s (non-gated rest scans) acquisition per step, 64 64 matrix, zoom 1.28 , pixel size 6.9 mm, 8 bins per cardiac cycle. For processing, data were transferred to a Hermes workstation (Nuclear Diagnostics, Stockholm, Sweden) and reconstructed using filtered back-projection with a Wiener filter (order 20). Reorientation was performed manually. Final processing steps and display of the results were automatic using the QGS software for ECG-g and QPS software for non-gated and summed ECG-gated studies (both Cedars-Sinai Medical Center, Los Angeles, USA; supplied by Nuclear Diagnostics). Qualitative analysis

Screenshots of the QPS splash pages containing all available slices were reviewed independently by three readers with prior experience of the Varicam system and QPS software (C.B., M.K., T.G.) who were given patients’ gender, but were blinded to further clinical information. For each patient, they were asked to assign the stress and rest scan to one of four categories with regard to the presence of artefacts related to abdominal activity: 1, no artefact; 2, abdominal activity just touching myocardium or not touching, but at same level as inferior wall (not interfering with interpretation); 3, abdominal activity covering some myocardium (interpretation still meaningful); and 4, non-diagnostic scan for large parts of myocardium. If stress and rest scans of a particular patient received a different rating, one of the scans was regarded as ‘better’.

Left: Patient demographics*. Right: Injection to first scan delay, in minutes (mean ± standard deviation)**

Metoclopramide administration

Patient demographics

Injection to first scan

Male

Stress scan Rest scan No metoclopramide (controls)

Female

Number of patients

Age (years)

Number of patients

Age (years)

Stress

Rest

62 56 41

65 ± 10 62 ± 10 62 ± 13

33 30 41

64 ± 10 67 ± 8.7 67 ± 9.3

71 ± 23 69 ± 23 79 ± 28

65 ± 26 61 ± 20 66 ± 29

*

Age was not significantly different between patients who received metoclopramide for their stress and rest scans (t-test, P = 0.59), nor between patients who received metoclopramide and the control group (t-test, P = 0.92). There were no significant differences.

**

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Metoclopramide to reduce artefacts in myocardial perfusion SPECT? Gru¨ning et al. 955

‘No change’ means that both scans received an equal rating. Individual readers’ ratings were then grouped as follows: ‘complete agreement’, i.e., all readers agreed on whether the stress or rest scan was the better one; ‘majority decision’, i.e., two readers came to such an agreement; ‘discordance’, i.e., stress and rest scans were regarded ‘better’ by at least one reader each. Quantitative analysis

The number of repeat acquisitions necessitated by abdominal activity was recorded. The following analysis based on Weinmann and Moretti [20] was performed on each stress and rest scan. Patients who showed reversible perfusion defects in the inferior wall were excluded from this analysis, and this applied to three patients who had

Fig. 1

metoclopramide for their stress scan, seven patients who had it for their rest scan, and five patients in the control group. The anterior raw projection and the two raw projections 31 either side of this were summed to obtain a substitute anterior planar image containing sufficient counts. Three adjacent 5  10 pixel rectangular regions of interest were placed in this image such that one covered the inferior left ventricular wall, and the other two contained adjacent abdominal activity (Fig. 1). An inferior wall-to-abdomen count ratio was calculated for both regions 2 and 3. Statistical tests

Patients who received metoclopramide for the stress and rest scan, respectively, were compared using a paired t-test for age (Table 1, left), and a one-way analysis of variance for injection to first scan delay (Table 1, right). t-tests were used to compare the study population with the control group in these respects. Data in Fig. 2 and Table 2 (top rows) were re-arranged and analysed as contingency tables using a chi-squared test. Data in Table 2 (bottom rows) were analysed with one-way analysis of variance and Bonferroni’s post-test. All calculations were carried out using Prism 3.03 (Graphpad Software, San Diego, USA).

Results Homogeneity of groups

Patients who received metoclopramide for the stress or rest scan, respectively, showed no significant differences with regard to age (Table 1, left) or injection to first scan delay (Table 1, right). There were no significant differences in these respects between the study population and the control group either. Qualitative analysis

Quantitative evaluation: position of the three regions of interest.

Table 2

The results of the readers’ ratings are shown in Fig. 2. To summarize, metoclopramide given before the stress scan led to abdominal activity being better in 16 scans, worse in 10 and unchanged in 67 scans, compared to the same patient’s rest scan without metoclopramide administration. Metoclopramide administered before the rest scan resulted in abdominal activity in 11 scans being better,

Top rows: Number of repeat acquisitions necessitated by abdominal activity*. Bottom rows: Quantification of abdominal activity** Metoclopramide administration

Stress scan

Repeat acquisitions Abdominal activity

Rest scan

Region 2 Region 3

Repeat acquisitions Abdominal activity

Region 2 Region 3

Stress scan

Rest scan

None (controls)

19/95 (20%)

17/86 (20%)

14/82 (17%)

1.49 ± 0.43 1.28 ± 0.48

1.44 ± 0.39 1.27 ± 0.44

2.05 ± 0.58 1.77 ± 0.53

11/95 (12%)

12/86 (14%)

6/82 (7%)

1.42 ± 0.35 1.19 ± 0.40

1.35 ± 0.34 1.10 ± 0.35

1.61 ± 0.37 1.37 ± 0.39

*

There were no significant differences. Expressed as an inferior wall-to-abdomen count ratio (mean ± standard deviation), for two regions of interest (see Fig. 1): one immediately adjacent to the inferior wall (region 2 in Fig. 1), and one further below (region 3 in Fig. 1). There were no significant differences. **

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Fig. 2

100%

Discordance Worse-majority decision Worse-complete agreement No change-majority decision No change-complete agreement Better-majority decision Better-complete agreement

80%

60%

40%

20%

0% Stress (n = 95) Rest (n = 86) Stress (n = 71) Rest (n = 70) Metoclopramide

Control

Quantitative rating of the influence of metoclopramide on artefacts related to abdominal activity, depending on whether the drug was given before the stress or rest scan. These differences were not significant (P = 0.10). Patients in the control group in whom either scan was better are shown for comparison, with the remaining patients showing no difference between stress and rest scans.

in 19 worse and 53 scans were deemed unchanged, compared to the same patient’s stress scan without metoclopramide administration. These differences were not significant (P = 0.10). In the control group, 12 patients had less abdominal activity in their stress than in their rest scan, with 11 patients vice versa, and there was no difference between stress and rest scans in the remaining 58 patients. Quantitative analysis

The number of repeat acquisitions due to abdominal activity is shown in Table 2 (top rows). The number of repeat stress or rest scans was not significantly different between patients who had received metoclopramide and those who had not. The number of repeat acquisitions in the control group is shown for comparison only. It seems to be somewhat lower, but this difference was not significant. The results of the quantification of abdominal activity can be found in Table 2 (bottom rows). The administration of metoclopramide, irrespective of whether it was given before the stress or rest scan, made no significant difference to abdominal activity. There was, however, significantly less abdominal activity in both stress and rest scans of the control group than in the study population.

Discussion Two of the authors (T.G., C.B.) were trained in the department from which the study by Pinkert et al. [19] originated. The use of 10 mg metoclopramide orally after both the stress and rest scan had become part of the

routine protocol for myocardial perfusion scans there. The subsequent publication by Weinmann and Moretti [20], stimulated by Pinkert’s earlier work, was unable to confirm these findings, but used a different methodology in a substantially smaller group of patients. The aim of our study was to combine the methodologies employed by both previous papers and to use an improved study protocol in a large number of patients to conclusively determine whether metoclopramide is useful in reducing artefacts related to abdominal activity in myocardial perfusion SPECT. An important improvement of this over the two previous studies is that we gave metoclopramide either before the stress or rest scan, but not both. This enabled us to use each patient as their own control (Table 2 and Fig. 2), thereby avoiding the influence of inter-individual variations in abdominal activity. A drawback of this approach is that abdominal activity has been thought to be less in the stress than in the rest scan, particularly with the use of additional exercise for pharmacological stress tests [17,18]. We do not believe that this had a marked influence on our results as, firstly, approximately 90% of patients were able to perform additional exercise. Secondly, to overcome this problem, we used a control group without administration of metoclopramide. Analysis of this group showed that readers’ ratings for stress and rest scans in this group were comparable to those in the study group (Fig. 2). An unexpected finding was a significant difference between the study and control populations with regard to quantification of abdominal activity (Table 2, bottom row). We believe, but are unable to prove, that this may be due to subtle, unintentional variations of the imaging protocol at the time this study was commenced, possibly with regard to injection to first scan timing. This finding does not invalidate the integrity of the control group for those purposes for which it is needed most, i.e., data contained in Fig. 2. An alternative explanation is provided by the timing of metoclopramide administration (immediately after tracer administration) which would lead to its peak action being observed 30–60 min later [21], i.e., around the time of scanning when it may have led to increased, rather than decreased uptake in the small bowel. Our results demonstrate that neither qualitative nor quantitative analysis showed an effect of metoclopramide on abdominal activity in myocardial perfusion SPECT. Whilst many suggestions on how to overcome this problem have been made (some previous studies are detailed in the introduction), it seems fair to say that no solution has been universally accepted to date. This may be due, in part, to the fact that protocols found to be working elsewhere cannot always be successfully transferred to another department. For now, abdominal activity will continue to be a frequent feature in myocardial perfusion scans: the quest continues.

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Metoclopramide to reduce artefacts in myocardial perfusion SPECT? Gru¨ning et al. 957

References Anagnostopoulos C, Harbinson M, Kelion A, Kundley K, Loong CY, Notghi A, et al. Procedure guidelines for radionuclide myocardial perfusion imaging. Nucl Med Commun 2003; 24:1105–1119. 2 Strauss HW, Miller DD, Wittry MD, Cerqueira MD, Garcia EV, Iskandrian AS, et al. Procedure guideline for myocardial perfusion imaging. Society of Nuclear Medicine. J Nucl Med 1998; 39:918–923. 3 Segall GM, Davis MJ. Prone versus supine thallium myocardial SPECT: a method to decrease artifactual inferior wall defects. J Nucl Med 1989; 30:548–555. 4 Dogruca Z, Kabasakal L, Yapar F, Nisil C, Vural VA, Onsel Q. A comparison of 201 Tl stress–reinjection–prone SPECT and 99mTc-sestamibi gated SPECT in the differentiation of inferior wall defects from artifacts. Nucl Med Commun 2000; 21:719–727. 5 Hayes SW, De LA, Hachamovitch R, Dhar SC, Hsu P, Cohen I, et al. Prognostic implications of combined prone and supine acquisitions in patients with equivocal or abnormal supine myocardial perfusion SPECT. J Nucl Med 2003; 44:1633–1640. 6 Germano G. Technical aspects of myocardial SPECT imaging. J Nucl Med 2001; 42:1499–1507. 7 Hurwitz GA, Clark EM, Slomka PJ, Siddiq SK. Investigation of measures to reduce interfering abdominal activity on rest myocardial images with Tc-99m sestamibi. Clin Nucl Med 1993; 18:735–741. 8 van Dongen AJ, van Rijk PP. Minimizing liver, bowel, and gastric activity in myocardial perfusion SPECT. J Nucl Med 2000; 41:1315–1317. 9 Boz A, Yildiz A, Gungor F, Karayalcin B, Erkilic M. The volume effect of the stomach on intestinal activity on same-day exercise–rest Tc-99m tetrofosmin myocardial imaging. Clin Nucl Med 2001; 26:622–625. 10 Boz A, Gungor F, Karayalcin B, Yildiz A. The effects of solid food in prevention of intestinal activity in Tc-99m tetrofosmin myocardial perfusion scintigraphy. J Nucl Cardiol 2003; 10:161–167. 11 Iqbal SM, Khalil ME, Lone BA, Gorski R, Blum S, Heller EN. Simple techniques to reduce bowel activity in cardiac SPECT imaging. Nucl Med Commun 2004; 25:355–359.

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Original article 51

Cr-EDTA measurements of the glomerular filtration rate in patients with sickle cell anaemia and minor renal damage

Fabiana B. Barrosa, Carmen S.P. Limab, Allan O. Santosa, Mariana F.C. Mazo-Ruiza, Mariana C.L. Limaa, Elba C.S.C. Etchebeherea, Fernando F. Costab, Sara T.O. Saadb, Edwaldo E. Camargoa and Celso D. Ramosa Background Creatinine clearance has been reported to be inaccurate for the estimation of glomerular filtration rate (GFR) in patients with sickle cell anaemia (SCA). Inulin clearance, the reference method for GFR estimation, is impractical for routine use in these patients, and 51Cr-EDTA measurements of the GFR have been rarely reported in this disease. Methods In order to obtain reference 51Cr-EDTA values in this disease, we studied 70 patients (40 females; 13–59 years of age, mean: 31.6 years) with homozygous SCA, normal serum creatinine and urinary albumin excretion < or = 200 lg
min – 1. All patients were submitted to single-injection 51Cr-EDTA GFR, urinary albumin and haematocrit measurements. 51Cr-EDTA clearances were calculated in different age groups ( < 20, 20–29, 30–39, 40–49 and > 50 years). Results The mean GFR ( ± standard deviation) obtained for the 70 patients was 111.5 ± 23.1 ml
min – 1. Analysis of variance for evaluation of the possible interaction effect between 51Cr-EDTA clearance and sex, age, urinary albumin and haematocrit demonstrated patient age as the only factor influencing 51Cr-EDTA clearance (P < 0.001). The Spearman correlation coefficient showed a significant relationship between 51Cr-EDTA clearance and patient age

Introduction The improved medical care for patients with sickle cell anaemia (SCA) has considerably prolonged their life expectancy in recent years [1]. As a consequence, the SCA patient population is also exhibiting more end-stage organ damage, a good example of which is sickle cell glomerulopathy [2]. Many functional and structural abnormalities of the kidney are observed in patients with homozygous SCA. Creatinine clearance has been reported to be inaccurate for the estimation of glomerular filtration rate (GFR) in these patients, leading to an overestimation [3] or underestimation [4] of the GFR. Inulin clearance, considered as the reference method for GFR estimation, is impractical for routine use or if a large number of patients needs to be studied. Although single injection of 51Cr-EDTA with subsequent plasma sampling

(r = – 0.44, P = 0.0001), but not between 51Cr-EDTA and urinary albumin (r = – 0.17, P = 0.1546) or haematocrit (r = 0.079, P = 0.5121). The group aged 20–29 years presented the highest 51Cr-EDTA clearance mean value (126.7 ± 20.4 ml
min – 1), with a progressive reduction in the older groups. Conclusion Young adults with homozygous SCA, normal serum creatinine and micro-albuminuria or normo-albuminuria present supranormal 51Cr-EDTA GFR values. These values rapidly decrease after 30 years of age. We did not find association between urinary albumin and GFR in these patients. Nucl Med Commun 27:959–962

c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:959–962 Keywords: 51Cr- EDTA, glomerular filtration rate, sickle cell anaemia, urinary albumin a Nuclear Medicine Division, Department of Radiology and bHematology Division, Department of Internal Medicine, State University of Campinas, Brazil.

Correspondence to: Dr Celso D. Ramos, Rua Murici 207, Bairro Alphaville, Campinas SP, Brazil 13098-315. Tel: + 55 19 3521 7772; fax: + 55 19 3521 7821; e-mail:[email protected] Received 8 November 2005 Accepted 30 January 2006

has become the most popular method for accurate assessment of GFR in the routine clinical practice [5], its use in SCA has been rarely reported and only in a limited number of patients and age range [4]. In order to obtain reference values to facilitate the outcome evaluation of the renal function of SCA patients, we measured 51 Cr-EDTA clearance in a wide age range of patients with homozygous SCA, normal serum creatinine and microalbuminuria or normo-albuminuria. We also evaluated the relationship between 51Cr-EDTA clearance and urinary albumin in these patients.

Patients and methods Patients

Seventy patients (40 females and 30 males, mean age of 31.6 ± 9.8 years, range 13–59 years) with homozygous

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960 Nuclear Medicine Communications 2006, Vol 27 No 12

SCA were investigated after receiving approval from the local ethics committee and obtaining informed consent from all patients. Exclusion criteria were abnormal serum creatinine (serum creatinine > 1.1 mg
dl – 1), clinical albuminuria (> 200 mg
min – 1), hypertension, diabetes and pregnancy. All patients were free of pain at the time of the study and had not been hospitalized or transfused for at least 3 months before the study. The patients were not receiving any medication other than folic acid. All patients had the haematocrit measured within 1 week of the 51Cr-EDTA procedure. 51

haematocrit. The ANOVA test (analysis of variance) was used to evaluate the possible interaction effect between 51 Cr-EDTA clearance and sex, age, urinary albumin and haematocrit. Significance level was set at 5% [9,10]. A scatter plot was drawn of the 51Cr-EDTA GFR values against age, which demonstrated a progressive reduction of GFR in patients older than 30 years. Linear regression analyses were then used to separately study the groups of patients aged below 30 years and patients aged 30 years or above. The Mann–Whitney test also was used to compare the 51Cr-EDTA clearances of patients aged below 30 years and 30 years or above.

Cr-EDTA clearance and urinary albumin

The patients were instructed to drink normally on demand during the test and to take 300–500 ml of water 1 h prior to the administration of 3 MBq of 51Cr-EDTA, injected into an antecubital vein. Accurately timed 10 ml venous blood samples were subsequently taken from the contralateral arm, into a heparinized collection tube, at 2, 3 and 4 h post-administration. The samples were then centrifuged and 2 ml of plasma were transferred into counting tubes (each plasma sample in duplicate). These tubes were counted together with two 2 ml standards taken as aliquots from 3 MBq 51Cr-EDTA diluted to 1000 ml. The injected doses were determined by assessing the difference in weight of the syringes before and after injection using a high precision balance. GFR was calculated by using the slope–intercept method, corrected for body surface area using the Haycock formula [6] and for the fast exponential using the BrochnerMortensen equation [7,8].

Results The descriptive analysis of the numerical variables of the patients is shown in Table 1. The mean GFR ( ± standard deviation) obtained for the 70 patients was 111.5 (23.1) ml
min – 1. GFR was higher in male patients (116.8 ± 23.1 ml
min – 1) than in female patients (107.5 ± 22.5 ml
min – 1), although the difference was not significant (P = 0.132) (Table 2). Urinary albumin excretion was normal (0–20 mg
min – 1) in 47 patients (67%) and 23 (33%) had microalbuminuria (> 20– 200 mg
min – 1). The ANOVA for the evaluation of the possible interaction effect between 51Cr-EDTA clearance and sex, age, urinary albumin and haematocrit demonstrated patient age as being the only factor influencing 51Cr-EDTA clearance (Table 3). The Spearman correlation coefficient demonstrated a significant relationship between 51Cr-EDTA clearance and patient age (r = – 0.44, P = 0.0001), but not between 51 Cr-EDTA and urinary albumin (r = – 0.17, P = 0.1546) or haematocrit (r = 0.079, P = 0.5121). The analysis of the different age groups showed the 20–29 years group as having the highest 51Cr-EDTA clearance mean value, with a progressive reduction in the older groups (Table 4).

Urinary albumin excretion was assessed by nephelometry using rabbit antiserum to human albumin (Dade Behring, Marburg, Germany). The urinary albumin and the 51 Cr-EDTA GFR measurements were performed within a maximum time interval of 2 months.

The scatter plot of the 51Cr-EDTA measurements of GFR of the 70 patients are shown in Fig. 1. The mean GFR ( ± standard deviation) of patients aged below 30 years was 123.1 ± 20.2 ml
min – 1 (median: 121.2 ml
min – 1) and the linear regression analysis of this group of patients demonstrated a non-significant increase in GFR at a rate of 0.67 ml
min – 1 per year (P = 0.434). The mean GFR (± standard deviation) of patients aged 30 years and above was 102.3 ± 24.2 ml
min – 1 (median: 98.5 ml
min – 1). The linear regression analysis of this group of patients

Statistical analysis

Descriptive analysis with determination of position and dispersion values for continuous variables was performed. The position and dispersion values were also analysed in different age groups (less than 20, 20–29, 30–39, 40–49 and greater than 50 years of age). The association between 51Cr-EDTA clearance and sex was analysed by the Mann–Whitney test. The Spearman correlation coefficient was used to verify the relationship between 51 Cr-EDTA clearance and age, urinary albumin and Table 1

Descriptive analysis of the numerical variables of the patients

Variable Age (years) Cr-EDTA (ml
min – 1) Albuminuria (mg
min – 1) Serum creatinine (mg
dl – 1) Haematocrit (%) 51

n

Mean

Standard deviation

Minimum

Median

Maximum

70 70 70 70 70

31.6 111.5 26.1 0.61 25.0

9.8 23.1 35.9 0.19 5.7

13 48.2 0.0 0.22 13.4

31 110.2 12.3 0.57 23.7

59 162.5 193.0 1.04 40.8

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GFR in sickle cell anaemia Ramos et al. 961

Comparison of 51Cr-EDTA clearance (ml
min – 1) between the sex subgroups of patients with sickle cell disease Table 2 Sex Female Male

n

Mean

Standard deviation

Minimum

Median

Maximum

40 30

107.5 116.8

22.5 23.1

48.2 73.8

103.4 115.4

162.5 157.1

Analysis of variance (ANOVA) for the evaluation of the possible interaction effect between 51Cr-EDTA clearance and sex, age, urinary albumin and haematocrit Table 3

Variable

Interaction with

Sex Age* Urinary albumin* Haematocrit*

51

Cr-EDTA clearance

F = 1.02; P = 0.317 F = 14.43; P < 0.001 F = 1.63; P = 0.206 F = 0.16; P = 0.693

*

Variables converted in ranks.

Table 4 Total plasma clearance of ized in age subgroups Age (years) < 20 20–29 30–39 40–49 > 50

51

Cr-EDTA (ml
min – 1) categor-

n

Mean

Standard deviation

Minimum

Median

Maximum

7 24 23 14 2

110.7 126.7 106.3 97.2 91.2

13.5 20.4 23.3 18.0 10.3

95.3 90.6 48.2 73.8 83.8

111.5 124.2 105.5 95.6 91.2

125.5 162.5 153.3 144.2 98.5

Fig. 1

51

Cr-EDTA Clearance (ml/min/1.73m2)

180

Women Men

160 140

Discussion Although 51Cr-EDTA measurements of the GFR have been in use for more than 30 years, there are limited data in the literature reporting its use in SCA [4]. This is particularly surprising in SCA, a disease in which glomerular dysfunction is common, frequently progressing to end-stage renal disease [2], and in which the creatinine clearance is inaccurate [3]. Patients with SCA have supra-normal proximal tubular function, as manifested by their increased ability to reabsorb phosphate and a2-microglobulins and to secrete uric acid and creatinine [3,11,12]. This last is clinically important in that it may result in significant overestimation of GFR by creatinine clearance [3,12]. On the other hand, creatinine clearance underestimated GFR in a group of SCA patients aged over 40 years, possibly due to advanced renal dysfunction or to a methodological error of urine collection [4]. All these factors do not affect the 51CrEDTA clearance unless the ‘distribution volume’ method is inadvertently used in a SCA patient with a low clearance value. Recent studies have provided important reference normal values for both adults [13,14] and children [15]. Such as in the present study, these authors have also used the ‘slope–intercept’ method for 51Cr-EDTA clearance measurement and the Brochner-Mortensen equation for correction of the fast exponential, a more sophisticated and probably more precise technique than the ‘distribution volume’ method [8]. This aspect may be critical in patients with SCA, a disease in which substantial deterioration in renal function may occur before it can be detected by clinical measurements, such as creatinine clearance [3].

120 100 80 60 40 10

20

30

40

50

60

Age (years) Scatter plot of the 51Cr-EDTA clearance against age in patients with sickle cell anaemia. The straight-line fits to the data show the results of modelling the association of 51Cr-EDTA clearance with age for patients younger than and older than 30 years. The central line represents the mean 51Cr-EDTA clearance and the upper and lower lines are ± 2 standard errors of the predicted mean clearance.

demonstrated a significant decline in GFR at a rate of 2.11 ml
min – 1 per year (P < 0.001). 51Cr-EDTA GFR values obtained for patients aged below 30 years were statistically different from the values obtained for patients aged 30 years and above (P < 0.001, Mann–Whitney test).

In our study, the mean ( ± standard deviation) 51Cr-EDTA clearance in the entire population was 111.5 ± 23.1 ml
min – 1, which is only slightly higher than the normal mean values reported for adults below 40 years (103.4 ± 15.5 ml
min – 1) [14] and for children (107 ± 17 ml
min – 1) [15]. On the other hand, we found a statistically significant difference between the mean ( ± standard deviation) values obtained in patients aged below 30 years (123.1 ± 20.2 ml
min – 1) and the values obtained in patients aged 30 years and above (102.3 ± 24.2 ml
min – 1) (P < 0.001). We did not find a significant difference between 51Cr-EDTA clearance in male and female patients, as reported for normal individuals [5,13,14]. We found a mean ( ± standard deviation) 51Cr-EDTA clearance of 110.7 ± 13.5 ml
min – 1 in our small group of patients below 20 years of age, a value that is close to the normal value reported by Blake et al. [15] in children and adolescents below 18 years (107 ± 17 ml
min – 1). Patients aged 20–29 years presented an elevated mean ( ± standard deviation) 51Cr-EDTA clearance (126.7 ± 20.4 ml
min – 1),

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962

Nuclear Medicine Communications 2006, Vol 27 No 12

which seems to be clearly supra-normal when compared to the normal value reported for adults (103.4 ± 15.5 ml
min – 1) [14]. The mean ( ± standard deviation) GFR of these patients is below the value described by Schimitt et al. [16] using inulin clearance measured in 14 SCA patients with normal serum creatinine and a mean age of 26.6 ± 1.3 years (146 ± 9 ml
min – 1). On the other hand, these values cannot be considered very different from each other since renal clearance of 51Cr-EDTA underestimates that of inulin by around 10% [17]. In our patient population, 51Cr-EDTA mean clearance rapidly decreased to normal values in 30–39-year-old patients (106.3 ± 23.3 ml
min – 1), and to more reduced values in the subsequent decades: 97.2 ± 18.0 and 91.2 ± 10.3 ml
min – 1 for 40–49 and > 50 years, respectively; although the oldest group should be carefully analysed since it included only two subjects. As expected, these values are higher than the 75.9 ± 30.2 ml
min – 1 51 Cr-EDTA mean clearance reported by Aparicio et al. [4] in SCA patients aged over 40 years, but with a wide range of normal and abnormal serum creatinine levels.

It is difficult to define a normal GFR value for patients with SCA since it is a congenital disease which can potentially affect the kidneys in the early phases of life. In this manuscript, we described the 51Cr-EDTA measurements of the GFR in a wide age range of patients with homozygous SCA and minor renal damage and we found supranormal 51Cr-EDTA GFR levels in the third decade of life followed by a fast reduction of these levels in the subsequent decades of life. We did not find association between urinary albumin and GFR in these patients.

References 1

2 3 4

5

We have also observed that the decline of 51Cr-EDTA GFR with aging begins earlier in life in SCA patients than in normal subjects (approximately 30 years vs. 40 years [14] of age, respectively). This decline occurs at a higher annual rate in SCA patients than in normal subjects (21.1 ml
min – 1 vs. 9.1 ml
min – 1 [14] per decade of life, respectively).

6

7 8

9

Different authors have demonstrated that young patients with SCA have supra-normal renal haemodynamics [11,12]. Vasodilation of the afferent glomerular arterioles, resulting from the increased renal production of prostaglandins by the kidney, is the most acceptable explanation for this finding [11,12].

10 11 12

13

Anaemia has been associated with a similar haemodynamic profile in an experimental model [18]. It cannot be the sole explanation for the changes seen in patients with SCA, however, since patients who receive multiple blood transfusions to achieve a normal haematocrit persist in maintaining supranormal GFR [19]. In fact, in the present study, we found no association between the haematocrit and the 51Cr-EDTA measurements of GFR.

14

15

16

17

Differently from other authors [20], we did not find association between urinary albumin and GFR, probably because we studied patients without renal dysfunction or in an initial phase of the renal disease. It may also be possible that albuminuria and GFR changes take place in an independent way in the early phase of the renal disease of SCA patients. Studies investigating the effects of angiotensin-converting enzyme inhibitors in patients with SCA and nephropathy found a significant reduction of proteinuria without a significant change in GFR [21] what could also suggest an independence of these parameters in SCA.

18

19

20 21

Platt OS, Brambilla DJ, Rosse WF, Milner PF, Castro O, Steinberg MH, et al. Mortality in sickle cell disease. Life expectancy and risk factors for early death. N Engl J Med 1994; 330:1639–1644. Ataga KI, Orringer EP. Renal abnormalities in sickle cell disease. Am J Hematol 2000; 63:205–211. Allon M. Renal abnormalities in sickle cell disease. Arch Intern Med 1990; 150:501–504. Aparicio AS, Mojiminiyi S, Kay JD, Shepstone BJ, De Ceulaer K, Serjeant GR. Measurement of glomerular filtration rate in homozygous sickle cell disease: a comparison of 51Cr-EDTA clearance, creatinine clearance, serum creatinine and beta 2 microglobulin. J Clin Pathol 1990; 43:370–372. Piepsz A, Pintelon H, Ham HR. Estimation of normal chromium-51 ethylene diamine tetra-acetic acid clearance in children. Eur J Nucl Med 1994; 21:12–16. Haycock GB, Schwartz GJ, Wisotsky DH. Geometric method for measuring body surface area: a height–weight formula validated in infants, children, and adults. J Pediatr 1978; 93:62–66. Brochner-Mortensen J. A simple method for the determination of glomerular filtration rate. Scand J Clin Lab Invest 1972; 30:271–274. Fleming JS, Zivanovic MA, Blake GM, Burniston M, Cosgriff PS. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Nucl Med Commun 2004; 25:759–769. Conover WJ. Practical nonparametric statistics. New York: John Wiley & Sons; 1971. Montgomery DC. Design and analysis of experiments 3rd edition. New York: John Wiley & Sons; 1991. De Jong PE, Statius Van Eps LW. Sickle cell nephropathy: new insights into its pathophysiology. Kidney Int 1985; 27:711–717. Allon M, Lawson L, Eckman JR, Delaney V, Bourke E. Effects of nonsteroidal antiinflammatory drugs on renal function in sickle cell anemia. Kidney Int 1988; 34:500–506. Hamilton D, Riley P, Miola U, Mousa D, Popovich W, al Khader A. Total plasma clearance of 51Cr-EDTA: variation with age and sex in normal adults. Nucl Med Commun 2000; 21:187–192. Grewal GS, Blake GM. Reference data for 51Cr-EDTA measurements of the glomerular filtration rate derived from live kidney donors. Nucl Med Commun 2005; 26:61–65. Blake GM, Gardiner N, Gnanasegaran G, Dizdarevic S. Reference ranges for 51Cr-EDTA measurements of glomerular filtration rate in children. Nucl Med Commun 2005; 26:983–987. Schmitt F, Martinez F, Brillet G, Giatras I, Choukroun G, Girot R, et al. Early glomerular dysfunction in patients with sickle cell anemia. Am J Kidney Dis 1998; 32:208–214. Granerus G, Aurell M. Reference values for 51Cr-EDTA clearance as a measure of glomerular filtration rate. Scand J Clin Lab Invest 1981; 41:611–616. Schrier RW, Earley LE. Effects of hematocrit on renal hemodynamics and sodium excretion in hydropenic and volume-expanded dogs. J Clin Invest 1970; 49:1656–1667. Statius van Eps LW, Schouten H, La Porte-Wijsman LW, Struyker Boudier AM. The influence of red blood cell transfusions on the hyposthenuria and renal hemodynamics of sickle cell anemia. Clin Chim Acta 1967; 17: 449–461. Guasch A, Cua M, Mitch WE. Early detection and the course of glomerular injury in patients with sickle cell anemia. Kidney Int 1996; 49:786–791. Falk RJ, Scheinman J, Philips G, Orringer E, Johnson A, Jennette C. Prevalence and pathologic features of sickle cell nephropathy and response to inhibitor of angiotensin-converting enzime. N Engl J Med 1992; 326: 910–915.

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Original article

An improved method for determining renal sufficiency using volume of distribution and weight from bolus 99mTc-DTPA, two blood sample, paediatric data Carl A. Wesolowskia, Paul S. Babynb and Richard C. Puetterc Objectives To find an improved method of determining renal sufficiency by exploring power functions for estimating normal value, E(arg), single compartment glomerular filtration rate (G1), rate constant (c) and renal sufficiency index, RSI = c/E(c) = G1/E(G1), using compartment volume (V ), patient mass ( W ), patient age (A), patient height (H), and sex (S). To present the best estimator of normal, E(G1) = f (V,W ).

glomerular filtration rate (GFR) and f (V,W ) was performed and yielded less than 5% precision error, 98% of the time. Normal RSI from f (V,W ) had the smallest standard deviation, 11.3%, no regression bias over a six-fold range on a Bland–Altman ratio plot, p = 0.4, and good agreement with clinical classification at 95% specificity (RSI > 0.8589, Cohen’s Kappa 0.70 ± 0.062 (mean ± bootstrap standard error).

Methods One hundred and thirty 99mTc-diethylenetriaminepentaacetic acid (99mTc-DTPA) combined imaging and G1 studies in 97 children were screened by findings and history to obtain 44 normal studies of patients 1.46– 18.5 years of age with blood samples at 109 (90–214) and 152 (120–246) min. Normal studies were used to generate predictive formulae.

Conclusions The best RSI from f (V,W ) is RSI = 90.927cV 0.35283W – 0.20185 and should detect mildly (14.1%) reduced renal sufficiency. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:963–970


c EðG1 Þ ¼ P xi i ¼

Results For power functions, f ðx1 ; x2 ; x3 ; . . . ; xn Þ, the statistically acceptable formulae in descending order of adjusted R2 were f (V, W ), f (V, A), f (V, H ), f (V ), f (W, A, H ), f ( W ), f (H ) and f (A). Relationships of the body surface area type, f (W, H, S) and f (W, H), were statistically unwarranted. Kleiber’s law, E(G1)p p W 3/4, 0.7618 with E(G1) = 6.9190W found here, allowed confirmation of GFRinulinE0.87G1. The best estimator is f (V,W ) = 10.998V 0.64717 W 0.20185, and may relate to a volumetric measure of body habitus. To verify methods, Monte Carlo simulation of the

Introduction The most important goal of this paper is to find the most sensitive method of detecting mild renal dysfunction. The ability to detect nephrotoxicity of chemotherapeutic agents, and the more general need for dosage adjustment for renal impairment and non-invasively detecting mild microscopic renal disease, are hampered by a lack of accurate estimation formulae of renal sufficiency. Renal sufficiency index, as defined here, is the ratio of supply of function, e.g., glomerular filtration rate (GFR), to demand for that function, e.g., some measure of body habitus. To find the most useful measure of body habitus, we examined multiple power functions for estimating single compartment glomerular filtration rate (G1), rate constant (g) and renal sufficiency index, RSI = g/E(g) = G1/E(G1), where E(g) and E(G1) are normal expectation values of g and G1. Useful formulae for E(g) and E(G1) were sought using all possible combinations of

Nuclear Medicine Communications 2006, 27:963–970 Keywords: kidney function tests, glomerular filtration rate, radioisotope renography methods, extracellular fluid volume–weight formula, renal sufficiency index

a Radiology, Memorial University of Newfoundland, Canada, bDiagnostic Imaging, The Hospital for Sick Children, Toronto, Canada and cCenter for Astrophysics & Space Sciences, University of California, San Diego, USA.

Correspondence to Dr Carl A. Wesolowski, Nuclear Medicine, The General Hospital, HSC, 300 Prince Philip Drive, St. John’s, NF, Canada, A1B 3V6. Tel: + 001 709 777 6132; fax: + 001 709 777 8267; e-mail: [email protected] Received 7 May 2006 Accepted 6 July 2006

compartment volume (V ), patient mass (W ), patient age (A), patient height (H ), and sex (S). Proposed explanations are discussed for the best estimator of normal G1, f (V, W ), and for Kleiber’s law (and other interesting estimators), as well as how to estimate GFRinulin from G1. Also discussed is the measurement precision, as well as problems associated with body surface area (BSA) formulae. (A list of the variables used in this paper, and the notation, is given in the appendix.) A constant infusion of inulin, with timed urinary collection and a mid-infusion or multiple blood samples, is usually taken as the ‘gold standard’ for measuring GFR, defined as inulin per minute of collection divided by the concentration of inulin per millilitre of plasma. However, due to its high molecular weight of approximately 5200 Da, inulin has the smallest volume of distribution of any known renal filterable marker. Furthermore, the

c 2006 Lippincott Williams & Wilkins 0143-3636


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964 Nuclear Medicine Communications 2006, Vol 27 No 12

volume of distribution of a constantly infused substance is only applicable to the test substance itself, and inulin volumes of distribution are only indirectly related to the volumes of distribution of other tracers. Current recommendations [1,2] for 51Cr-ethylenediaminetetraacetic acid (51Cr-EDTA) and 99mTc-diethylenetriaminetetraacetic acid (99mTc-DTPA) bolus methods suggest that G1, the one-compartment GFR, is numerically higher in value than GFRinulin, and propose a G1 correction by the method of Chantler and Barrett [3], i.e., GFRinulinE0.87G1. While this conversion factor was calculated for a different agent, 51Cr-EDTA, at different blood sampling times than are used here, nonetheless, using Kleiber’s law and constant infusion inulin data from Adolph [4], we demonstrate that this conversion factor applies to the results of this paper. While we apparently understand how to correct G1 for GFRinulin conditions as above, it is unclear that the onecompartment volume of distribution estimate for the 99m Tc-DTPA used here needs modification at all. This is because truth data for volumes of distribution is lacking [5] despite claims to the contrary [6]. Specifically, in 1981, Zierler [5] found many of the same problems with compartmental models that we find today. Among these are that for fitting a plasma concentration curve with a sum of exponentials, the fit quality and coefficients depend strongly on whether one fits the early or late time behaviour, that compartmental models are inherently non-testable, and that the adopted compartment models are often not realizable for a small number of compartments. Furthermore, while the recent experimental work of Finco [7] did not directly conclude that the noncompartmental, mean residence time calculations of Becker [8] diverge from inulin constant infusion calculations, they did suggest that this disparity existed for ‘commercial software’. Less ambiguously, for the onecompartment model, Finco [7] observed increasing inaccuracy for increasingly reduced function using very early (20 min) first sample times of two blood samples and for multiple early blood samples. To avoid this latter problem, the present study used first sampling times of greater than 90 min and a second sample after a further 45 min. As presented below, these sampling times seem to be adequate. The relative standard deviation (RSD) of our best fit function of RSI from f(V, W ) of 11.2% includes all inter-patient variability and experimental errors, and sets an absolute upper limit for all errors for discriminating between normal and abnormal results. If the sampling times were inadequate, systematic experimental errors would obviate finding an 11.2% RDS. However the contrary is found, and as presented below, RSI from f(V, W ) reduces the dispersion in RSI by a factor of 2 over that of BSA indexing of the same data, a result with very high statistical significance. Nonetheless, concerns

remain since both Zierler [5] and the experimental work of Finco [7] show that a single exponential and even a sum of exponential functions are problematic descriptors of the true plasma concentration curve. Indeed, one current recommendation made primarily to increase the precision of GFR measurements is to use a 2 h delay between blood samples [1]. Unfortunately, this recommendation is based on unspecified count rates from a 51Cr-EDTA data simulation [9] and the accuracy provided by the 2 h delay time between samples was not measured or discussed. So to ensure the precision of our results, we performed a Monte Carlo simulation using Poisson realizations of our data. These results allow us to determine error estimates for our fits that themselves can be shown to be accurate to 13% (i.e., the error in our error estimates are accurate to 13%). This allows for presentation of both best-fit parameter values along with their errors. Furthermore, this error analysis shows that our sample times are sufficiently late and have sufficient separation in time to reliably and accurately discriminate between proposed functional dependencies – see the results section below. The slope–intercept method used herein is a commonly used clinical method. Indeed, the data were obtained retrospectively from a clinical protocol. What is of much interest is that renal sufficiency can be accurately predicted using this data and that even if the single compartment model is imperfect, the fitting formulae derived using this method significantly reduce the variance in predicting renal sufficiency relative to previous methods. The focus of this work is not on calculating GFR, but on characterizing RSI. Of course, the method for calculating GFR in the present work relies upon calculating V of the injected agent. In this case, GFR from a bolus-injection one-compartment model (i.e., from the first order rate equation) is given by G1 ¼ gV ;

ð1Þ

where g is the rate constant for clearance of the exogenous marker. Equation 1 implies that two blood samples are needed to determine GFR. Logarithmic plots of renal data often show homogeneous variance, suggesting that power functions are the appropriate functional form for E(GFR) (Figs 1 and 2). Indeed, the most successful models of expectation of renal function to date, Kleiber’s law and the BSA formulae, are power function models. Consequently, the work presented here can be thought of as a generalization and extension of prior studies to investigate other power formulae with other variables. The current paper also includes joint probability testing of the suitability of hypothetical target formulae.

Methods This research was approved by the Research Ethics Board of the Hospital for Sick Children. One hundred and thirty

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An improved method for determining renal sufficiency Wesolowski et al. 965

Fig. 1

Fig. 2

V versus W r = 0.875

30

InV versus InW r = 0.923

3.5 3.0

25

2.5 20

V

InV

2.0 15

1.5 10

1.0 5

0.5 0

0 0

20

40

60

80

100

0

1

2

W

3

4

5

InW

There is good linear correlation, r = 0.875, between volume of distribution and weight for the entire population of 130 studies. This would suggest an G1 – E(G1) index where E(G1) = a1x1 + a2x2 + a3x3 + y + anxn. However, the correlation between ln V and ln W, r = 0.923, of Fig. 2 is better than for Fig. 1. This difference in fit quality is significant, p = 0.0422, with the (natural) log–log relationship of Fig. 2 removing the proportionate increase in variance with variable size seen in the linearlinear plot of Fig. 1.

In this paper, data that exhibit homogenous variance on log–log plotting are called proportional data. Proportional data implicate the use of linear–log regression, i.e., power functions, and ratios for modelling in preference to linear functions and differences. That is, the renal sufficiency index, RSI = G1/E(G1), where the estimation value of G1 is EðG1 Þ ¼ f ðx1 ; x2 ; x3    xn Þ ¼ K x1c1 x2c2 x3c3    xncn , a generalized power function, and G1 is the uncorrected single compartment, slope–intercept GFR, are preferred to the linear relationships illustrated in Fig. 1.

99m

Tc-DTPA combined imaging and G1 studies in 97 children were screened by findings and history to obtain 44 normal studies of patients 1.46–18.5 years of age from whom blood samples were drawn at 109 (90–214) and 152 (120–246) min. Normal studies were used to generate predictive formulae. Of the normal studies, 37 were performed during work-up for (non-renal) neoplasia and only seven for other conditions. There were 27/130 (21%) studies that could not be clearly classified and were grouped with the abnormal results. Among the 86 abnormal studies, 29 had neoplasia, 21 had renal transplants, 11 were investigated for other renal indications, and 25 for non-renal conditions. The data ranges are given in Table 1. Note that the normal period of renal maturation below approximately 18 months of age is not characterized here.

renal sufficiency index, RSI, such that any estimators, whether from BSA indexing or other E(GFR) or from E(g), follow the identities

Throughout this paper, the expectation value of GFR is modelled as

where K 0 = K/1000, and, V is in litres and not in millilitres as in Equations 1 and 3. Alternatively, E(g) could be fit directly to the data, but this would not allow for E(G1) equations without a V term, see the right hand expression in Equation 3. Equations 1 through 4 are used as follows. Suppose we find a function to estimate normal G1 to be f(V, W ), where V is in litres. This means that E(G1) = f(V, W ) = KV a W b, then RSI = G1/E(G1) = gV/(KV a W b) = gK – 1V 1 – aW – b and E(g) = KV a – 1 W b/1000.

c

c

c

EðG1 Þ ¼ K  x11 x22 x33    xncn n Y c ¼ xi i ¼ f ðx1 ; x2 ; x2 ; . . . ; xn Þ:

ð2Þ

i¼0

The fxi > 0g are selection(s) from the list of candidate predictor variables, i.e., of V, W, A, H, g, and S. We define a

RSI ¼

g G1 EðG1 Þ ¼ and EðgÞ ¼ EðgÞ EðG1 Þ V

ð3Þ

where substitution of the right hand expression for E(g) into the left hand equation results in the definition of G1 as per Equation 1. In other words, this definition is selfconsistent. RSI for each patient study is calculated from the left hand expression in Equation 3. An equation for E(g) is obtained from its corresponding E(G1), substituting Equation 2 into Equation 3 yields c

EðgÞ ¼

c

c

K x11 x22 x33    xcnn K 0 c1 c2 c3 ¼  x x x    xncn ; 1000 V V 1 2 3

ð4Þ

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966 Nuclear Medicine Communications 2006, Vol 27 No 12

Table 1

Minimum (Min) and maximum (Max) values for the range of the variables defined in the text

Range

V (l)

Normal, n = 44 Abnormal, n = 86 All, n = 130

Table 2

W (kg)

A (years)

H (cm)

Min

Max

Min

Max

Min

Max

Min

Max

F

M

2.53 1.34 1.34

24.2 18.1 24.2

10.1 4.35 4.35

75.8 88.8 88.8

1.46 0.285 0.285

18.5 18.0 18.5

82. 58.5 58.5

182. 181. 182.

12 32 44

32 54 86

z

f (V, W ) f (V, A) f (V, H ) f (V ) f (W, A, H) f(W) f (W, H ) f (W, H, S) f (H) f(A)

Min

G1(ml  min – 1) Max

0.00724 0.0138 0.000668 0.0127 0.000668 0.0138

Min

Max

31.5 5.69 5.69

213. 184. 213.

Statistics and probabilities of variables affecting G1 fit quality Probability of logarithm predictors having no effect on regression

E(ln(G1))

g (min – 1)

S

= ln f (V < 0.0001 < 0.0001 < 0.0001 < 0.0001

W

A

H

S)

0.0395 0.0807 0.0461

0.0756 0.7976 0.8032 < 0.0001

0.7372

< 0.0001

F statistic

R2 *

R2w

Accept Accept Weak Accept Weak Accept Reject Reject Accept Accept

374.48 360.77 349.76 661.56 109.18 306.41 149.83 97.77 187.62 184.60

0.9481 0.9462 0.9446 0.9403 0.8911 0.8795 0.8821 0.8800 0.8171 0.8146

0.9456 0.9436 0.9419 0.9389 0.8830 0.8766 0.8738 0.8710 0.8127 0.8102

Intercept

0.0173

< 0.0001 < 0.0001 < 0.0001 < 0.0001

Critical p = 0.1 < 0.0001 < 0.0001 0.0304 < 0.0001 0.0123 < 0.0001 0.1183 0.1309 < 0.0001 < 0.0001

All p < 0.05 relationships are in bold. Not shown are equations that include g as a predictor variable (No valid equations found). R is the coefficient of determination. w 2 R is an adjusted coefficient of determination index, R2=1 – (1 – R2)(n – 1)/(n – 1 – k), where k is the number of independent variables and n, the number of observations, is 44. z For f (V, W ), the variance inflation factor is a tolerable 7.921. * 2

The next expression is important for determining the relative likelihood of different models. To assess the joint probability of a given set of power function exponents fci g the full covariance of the variables is calculated. One way of doing this is to change coordinates to f~ci g by rotating into the coordinate system in which the eigenvectors of the covariance matrix are the basis vectors. In this coordinate system, the new variables are statistically independent and PðDc0 ; Dc1 ; Dc2 ; . . . ; Dcn Þ ¼ PðD~c0 ; D~c1 ; D~c2 ; . . . ; D~cn Þ n Y ð5Þ PðD~ci Þ ¼ i¼0

where the Dci are the departures of the ci from their optimized fit values. For Equation 5, K of Equation 2 was c incorporated into the notation using K ¼ eln K ¼ x00 , where x0 = e, and c0 = ln K. The uppercase P is used as shorthand for the two-tailed Student’s-t cumulative distribution function (CDF) probability. To estimate the errors in our results, a Monte Carlo simulation was performed. For each study, 20 different sets of Poisson realizations were made from eight measurements each from two sets of four counting vials. Each set of four vials contained two with plasma samples, one with blank background (tap water), and one with standard dilution fluid. On one occasion, a baseline

plasma sample at time zero was also counted (and modelled). Each set of Poisson realizations was run through the fitting procedure and the results tabulated for (1) GFR, (2) renal sufficiency index (RSI) from RSI = G1 / f(V, W ), (3) the one-compartment rate constant (g), and (4) the one-compartment volume of distribution (V ). RSDs were then estimated for the 20 trials (yielding roughly RSD ± 13%) for each patient study for each parameter. The population of RSDs was then analysed to yield expected errors of parameters.

Results For our data in general, power function modelling proved superior to linear modelling. An exception to this rule is the slight degrading of the partial correlation of V and H with G1 when power function modelling is compared to linear modelling (from r = 0.7629 to 0.7582, n = 130). Another result is that sex is a poor predictor variable (all p > 0.1). For these fits, values of S equal to 1 for female and 2 for male were assigned so that ln S exists and so that regressions using ln S would find appropriate scaling exponents. Multivariate linear regressions of lnG1 against all combinations of the logarithms of normal study data for V, W, A, H and S were performed (as per Equation 2 and Table 2) to produce the power function fits of Table 3.

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An improved method for determining renal sufficiency Wesolowski et al. 967

2

2 Table 3 All GFR fit equations, listed from best to worst R (R ). Equations from Table 2 and other sources. Formulae containing p < 0.05 predictor variables only are in bold

E(G1)

= Long form

Quality

CCIw

Notes

f(V,W )

10.988V 0.6472W 0.2019

Accept

33.95

f(V, A) f(V, H ) f(V ) f(W, A, H ) f(W ) f(W, H ) f(W, H, S) f(H ) f(A)

14.964V 0.7151A0.1109 4.3249V 0.7296H 0.2949 13.835V 0.8510 1056.2W 0.8404A 0.3563H – 1.2375 6.9190W 0.7618 10.029W 0.8053H – 0.1067 9.6009W 0.8053H – 0.1049S 0.0210 0.0193H1.7401 28.138A0.5785

Accept Weak Accept Weak Accept Reject H Reject S, H Accept Accept

18.94 100.6 7.835 332.5 11.12 166.5 187.0 38.42 5.215

Preferred formula, highest R Age limited High collinearity condition index Best single variable Very high collinearity condition index 73.3W 0.75, Kleiber’s law, kilocal./day BSA formula without sex BSA formula with sex High collinearity for single variable Low R 2

2

w

CCI, the collinearity condition index, measures collinearity for a fit model. CCI is the square root of the ratio of largest to smallest eigenvalues of the covariance matrix of the fit variables. Collinearity increases when a fit variable is more nearly a linear combination of other fit variables in a model, causing the constant associated with that variable to become unstable. For example, note how variable the exponent for H is in this table. The sex variable (where S = 1 for female and 2 for male) is equivalent to setting the female formula multiplier, S cS , to unity and multiplying for males by the factor 1.0140 for f (W,H,S ).

2

Table 2 shows the adjusted R2 indices (R ) for all of the power functions. The statistically acceptable power function formulae for predicting G1 in descending order of adjusted R2 were f(V, W ), f(V, A ), f(V, H ), f(V ), f(W, A, H ), f(W ), f(H ) and f(A). Relationships of the body surface area type, f(W, H, S ) and f(W, H ), were statistically unwarranted, degenerating to Kleiber’s law, f(W ). Kleiber’s law, with E(G1) = 6.9190W 0.7618 found for our data, can be explained by a fractal geometry of the body [10,11], with W 3/4 being the predicted scaling [4,10–15] and with the difference in exponents being insignificant ( P = 0.787, Equation 5). Adolph [4] obtains a similar (P = 0.851) exponent to ours from E(GFRinulin) = 5.921W 0.77 for humans, and the metabolically similar species, dogs, rabbits and rats [4]. Note that similarity of metabolism and proportionately linked renal function in this context requires similarity of comparative renal and total-body metabolic pathways and body temperatures, consideration of which we credit to Adolph. However, GFRinulin is significantly smaller than G1 for 99mTc-DTPA [6,16,17]. Indeed, using Adolph’s numbers we can independently recalculate a Chantler and Barratt [3] type estimator for the weight range of our population to be E(GFRinulin) = 0.879G1 (0.84–0.92, 95% confidence range, using the bootstrap method). This was obtained by regressing the normal data from this paper to the closest inulin exponent of W in the literature, i.e., Adolph’s result for the exponent of W. This appears to confirm Chantler and Barratt’s, 51Cr-EDTA 0.87 scale factor, is calculated for 99mTc-DTPA versus inulin, and is important because it answers the question ‘Under the conditions of our experiment, how does one estimate GFRinulin?’

The most likely candidate equations for predicting RSI are listed in Table 4. Using Equation 5, the best estimator of normal function, E(G1) = f(V, W ), did not differ in a statistically significant manner, P = 0.88, from V 2/3 W 3/4, where V 2/3 would represent an Euclidian area and W 3/4, a

Table 4 Renal sufficiency index (RSI) values from Equation 3 ranked from best to worst RSI from E(G1) rule:

Mean

SD

RSD

f (V, W ) f (V,A) f (V,H ) f (V ) f (W,A,H ) f ( W ), Kleiber’s law

1.0061 1.0063 1.0065 1.0070 1.0127 1.0142

0.11293 0.11549 0.11736 0.12049 0.16239 0.17404

0.11224 0.11476 0.11659 0.11966 0.16035 0.17159

Multiply by 100 for percentages. n = 44. SD is standard deviation. RSD is the relative standard deviation, also known as the coefficient of variation.

fractal length, applying arguments that parallel the fractal length scaling (W 1/4) and fractal volume scaling (W 3/4), explanations proposed [10,11] for Kleiber’s law [4,10,11,13–15]. The bootstrap 95% confidence limits for E(G1) = f(V, W ) = KV a W b are K (8.4884–14.249), a (0.47428–0.82005), b (0.03252–0.37119). No sub-selection of sampling times produced coefficients beyond these limits. RSI calculated from f(V, W ) had no significant regression bias over a six-fold range on Bland–Altman ratio plot, p = 0.39 (Figs 3 to 6). Table 4 shows that RSI from f(V, W ) has the best mean, standard deviation (SD) and relative standard deviation (RSD). Table 5 shows the RSI from f(V, W ) numerical predictive characteristics and good agreement from Cohen’s Kappa 0.70 ± 0.062, (mean ± bootstrap standard error). Also, there is agreement ( p = 0.26, Cochran’s Q) with expected prevalence of normal results in the clinical population (n = 59), when the 27/130 unclassifiable studies are reapportioned (using p = 0.5) as compares to the number predicted by f(V, W ) (n = 56) using the 95% sensitivity cut-off value of RSI = 0.859.

The simulation of error for the parameters is shown in frequency histograms of Fig. 7. It is of interest that over the range from 6.5 to 122.7% of expected renal function (as RSI from f(V, W )), and 5 to 185 ml  min – 1 GFR, 98%

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968 Nuclear Medicine Communications 2006, Vol 27 No 12

Fig. 3

Fig. 5

RSI f (W ) versus GFR

y = 0.000707x + 0.948

1.4

1.6 + 2SD

1.2 mean

1 0.8

− 2SD

0.6 0.4

1.4 RSI BSA (Haycock)

1.6

RSI f (W )

RSI BSA (Haycock) versus GFR

R 2 = 0.0322 p = 0.244

y = 0.00171x + 0.840 R 2 = 0.159 p = 0.00741 +2SD

1.2 mean

1 0.8

–2SD

0.6 0.4 0.2

0.2

0

0 0

25

50

75

0

100 125 150 175 200 225

25

50

75

100 125 150 175 200 225

Average G1 and f (W )

Average G1 and E(G1 BSA)

Figures 3 to 6 are Bland–Altman ratio plots, also known as Eksborg plots. The ratio plot type, e.g., RSI, is preferred to a difference plot for modelling proportional data. Figure 3 shows f ( W ), Kleiber’s law for GFR, arises in log space when the patient weight data is used to estimate G1. RSI = G1/ f ( W ) is an unbiased estimator ( p = 0.24 bidirectional) of averaged G1 and f ( W ).

RSI from BSA, is similar to Kleiber’s law but biased (p = 0.007). BSA is a power function, i.e., a proportional data model. No BSA formula arises from the data, so scaling of RSI BSA to the data was adjusted by assigning the mean value to be 1.

Fig. 4

Fig. 6

RSI f (V ) versus GFR 1.6 1.4

1.6 1.4

+ 2SD

1

mean

0.8

– 2SD

0.6 0.4

y = 0.000330x + 0.975 R 2 = 0.0176 p = 0.391 +2SD

1.2 RSI f(V,W )

1.2 RSI f (V )

RSI f(V,W ) versus GFR

y = 0.000388x + 0.971 R 2 = 0.0212 p = 0.346

1

mean

0.8

–2SD

0.6 0.4

0.2

0.2

0 0

25

50

75 100 125 150 175 200 225 Average G1 and f (V )

This figure shows that RSI from f (V ) ( = 0.07228gV 0.1490) is unbiased (p = 0.35) and has a lesser variance than RSI from f (W ).

0 0

25

50

75

100 125 150 175 200 225

Average G1 and f(V,W ) RSI from both weight and volume of distribution, f (V, W ), is the least biased estimator (p = 0.39) and has the least variance for estimating normal RSI of any variable combination, see Table 4.

of the values for GFR and RSI from f(V, W ) had less than 5% RSD with median errors of 0.9% and 1.8% RSD, respectively.

Discussion Multivariate linear regression and power function modelling provides a unifying theoretical framework for generating and evaluating new and existing predictive equations for GFR. Although not perfectly general, this method provides the ability to identify significant variables and relationships between these variables. An organized approach of this type is likely to minimize errors relative to other, less reliable methods of evaluating predictive relationships.

Kleiber showed that the exponent of weight for BSA formulae were too small and weight to the first power too large to predict metabolism between species [12]. Kleiber’s law is confirmed as statistically warranted by regression of ln G1 with the logarithms of the predictive variables of our data (Tables 2 and 3) and as a measure for which no significant bias is detected (Fig. 3). This confirms that GFR is a physiological marker of metabolism, and predicts that a change in metabolism will modify GFR (e.g., hypothyroidism reduces GFR).

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An improved method for determining renal sufficiency Wesolowski et al. 969

Specificity, agreements with clinical assessment (Kappa) and predicted proportion (Q) of normal cases for RSI from f (V, W )

Table 5 n = 130

RSI from f (V, W ) vs. clinic

RSI 95% specificity Predicted normal Cochran’s Qw Known normal Cohen’s Kappaz

*

0.8588975 59 vs. 56 1.29, p = 0.2568 59 vs. 44 0.70 ± 0.062, good

* Specificity is from 15-point interpolation of ROC curve data from RSI from f (V,W ). w Cochran’s Q statistic, probability of. z Cohen’s Kappa mean ± asymptotic standard error (bootstrap), grade. Cochran’s Q statistic and Cohen’s Kappa use normal and abnormal scores from clinical assessment for predicted prevalence of normal studies, n = 56, where certainly normal is n = 44, versus predicted normal studies from RSI from f(V,W ) is n = 59.

Fig. 7

GFR Coefficient of variation (RSD)

0.15

RSI f (V,W )




V

strong evidence suggesting that within a single species, i.e., in human children, the correlation of BSA to GFR is spurious, is caused by the use of weight to smaller-thanneeded powers in the BSA estimator equations, yielding a biased estimator of GFR (Fig. 5). Turner and Reilly suggest that BSA indexing of GFR should be abandoned [19]. However, this cannot be done today because of the large body of chemotherapy estimations that utilize BSA, and only rarely use Kleiber’s law. The current paper presents GFR predictors with significantly reduced RSD of RSI than past studies. For particular values of f(V, W ), 11.2% RSI RSD should provide the ability to diagnose hitherto unrecognized mild renal impairment non-invasively. Table 5 relates that a mere 14.1% reduction of RSI from f(V, W ) should be detectable. Figure 6 shows that the combination of V and W for predicting RSI has no significant bias ( p = 0.39). This lack of bias was slightly less than the also insignificant biases of the separate W and V power functions of Figs 3 and 4.

0.10

0.05

0.00 0 1020 30 40 0 1020 30 40 0 1020 30 40 0 10203040 Frequency

Results of Monte Carlo modelling. Twenty Poisson distributed samples (see text) were used to simulate measurements and then fit in the same manner as the original 130 patient studies. Shown are the resulting frequency histograms of the coefficients of variation (RSD or the standard deviations divided by the means) for each study for four parameters. The four frequency histograms from left to right are for (1) GFR, (2) renal sufficiency index (RSI) from RSI = G1 /f (V, W ), (3) the one-compartment rate constant (g), and (4) the one-compartment volume of distribution (V ). The RSD histograms of GFR and G1 are identical. Also, the RSD histograms of g and the half-time for plasma clearance T1/2 = ln 2/g are identical. Note that with the exception of 3/130 (2%) outliers each, both GFR and RSI from f (V, W ) are below 5% error.

According to Gehan and George [18], H is a necessary predictor of BSA with their estimators explaining 99% of the variation in BSA, the Du Bois’ formula explaining 98% and Boyd’s estimator explaining 97%. However, Table 2 shows that H is an unwarranted variable (P = 0.80) to use with W for predicting GFR. For uncorrected GFR, we find a significant difference between the Haycock or Du Bois’ formulae and f(W, H ), where PðD~cW ; D~cH Þ ¼ 0:012; 0:007, respectively from Equation 5. Using the currently recommended Haycock (or any other) BSA formula for children [1], yields an inferior predictor (RSD 18.1%) to Kleiber’s law (RSD 17.2%). In summary, the current paper provides

Any complete theory of body habitus should include geometric arguments. In addition to Kleiber’s law being consistent with f( W ), the fractal geometric arguments proposed [10,11] to explain Kleiber’s law also appear to be related to f(V, W ). The functional form of f(V, W ) is postulated to arise due to the importance of some fundamental volume, i.e., a geometric surface area (V 2/3) times optimized fractal length of metabolite delivery (W 1/4), critical to metabolism. As f(V, W ) has a superior 2 log-space R of 0.9436 and reduced RSD for RSI, f(V, W ) should likely be viewed as potentially replacing both f( W ), i.e., Kleiber’s law, and the BSA formulae when estimating renal sufficiency in children. To recapitulate, our best estimator, f(V, W ), yields the formulae: RSI = 90.927gV 0.35283W – 0.20185, expected normal rate constant E(g) = 0.010998V – 0.35283W 0.20185 and expected normal function E(G1) = 10.998V 0.64717W 0.20185 where GFRinulinE0.87G1. While the results of this work are very encouraging, they are preliminary. Further investigation would be needed to recommend implementation of these formulations for pharmacokinetic dosimetry, non-invasive detection of mild renal disease, correlation to other measures of renal sufficiency, and correlation to measures of metabolism or haemodynamic parameters.

Acknowledgements The authors are indebted to Dr David L. Gilday, for his advice and help with the procedure, data and preparation of the manuscript and to Maria Green, RTNM for her help with the procedure and the data.

References 1

Piepsz A, Colarinha P, Gordon I, Hahn K, Olivier P, Sixt R, et al. Guidelines for glomerular filtration rate determination in children. Eur J Nucl Med 2001; 28:BP31–BP36.

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970 Nuclear Medicine Communications 2006, Vol 27 No 12

2

3 4 5 6

7

8 9

10 11

12 13

14

Fleming JS, Zivanovic MA, Blake GM, Burniston M, Cosgriff PS. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Nucl Med Commun 2004; 25:759–769. Chantler C, Barratt TM. Estimation of glomerular filtration rate from plasma clearance of 51-chromium edetic acid. Arch Dis Child 1972; 47:613–617. Adolph E. Quantitative relations in the physiological constitutions of animals. Science 1949; 109:579–585. Zierler K. A critique of compartmental analysis. Annu Rev Biophys Bioeng 1981; 10:531–562. Peters AM, Henderson BL, Lui D, Blunkett M, Cosgriff PS, Myers MJ. Appropriate corrections to glomerular filtration rate and volume of distribution based on the bolus injection and single-compartment technique. Physiol Meas 1999; 20:313–327. Finco DR. Measurement of glomerular filtration rate via urinary clearance of inulin and plasma clearance of technetium Tc 99m pentetate and exogenous creatinine in dogs. Am J Vet Res 2005; 66:1046–1055. Becker R. A derivation of mean residence time. Eur J Drug Metab Pharmacokinet 1991; Spec No 3:15–16. De Sadeleer C, Van Laere K, Georges B, Piepsz A, Ham HR. Influence of time interval and number of blood samples on the error in renal clearance determination using a mono-exponential model: a Monte Carlo simulation. Nucl Med Commun 2000; 21:741–745. West GB, Brown JH, Enquist BJ. A general model for the origin of allometric scaling laws in biology. Science 1997; 276:122–126. West GB, Brown JH, Enquist BJ. The fourth dimension of life: fractal geometry and allometric scaling of organisms. Science 1999; 284:1677–1679. Kleiber M. Body size and metabolic rate. Physiological Reviews 1947; 27:511–541. West GB, Woodruff WH, Brown JH. Allometric scaling of metabolic rate from molecules and mitochondria to cells and mammals. Proc Natl Acad Sci USA 2002; 99(suppl 1):2473–2478. Maxwell LK, Jacobson ER. Allometric scaling of kidney function in green iguanas. Comp Biochem Physiol A Mol Integr Physiol 2004; 138:383–390.

15

Edwards NA. Scaling of renal functions in mammals. Comp Biochem Physiol A 1975; 52:63–66. 16 Moore AE, Park-Holohan SJ, Blake GM, Fogelman I. Conventional measurements of GFR using 51Cr-EDTA overestimate true renal clearance by 10 percent. Eur J Nucl Med Mol Imaging 2003; 30:4–8. 17 Holford NH. A size standard for pharmacokinetics. Clin Pharmacokinet 1996; 30:329–332. 18 Gehan EA, George SL. Estimation of human body surface area from height and weight. Cancer Chemother Rep 1970; 54:225–235. 19 Turner ST, Reilly SL. Fallacy of indexing renal and systemic hemodynamic measurements for body surface area. Am J Physiol 1995; 268:R978–R988.

Appendix: Variables and notation G1 g RSI BSA V W A H S E(arg) E(G1)

One-compartment GFR, ml  min – 1 Clearance rate constant, min – 1 Renal sufficiency index Body surface area, m2 Volume of distribution, ml or l Patient mass, kg Patient age, years Height, cm Sex; Female = 1, Male = 2 Estimation value of argument c c ¼ K  x1c1 x22 x33    xncn n Y c ¼ xi i ðproduct notationÞ i¼0

GFRinulin

¼ f ðx1 ; x2 ; x3 ; ; ; xn Þ (e.g., f (V, W ) GFR from constant infusion

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Original article 18

F-FDG PET imaging in assessing exudative pleural effusions

Bernard C. Duysinxa, Marie-Paule Larockb, Delphine Nguyena, Jean-Louis Corhaya, Thierry Burya, Roland Hustinxb and Renaud Louisa Background This study evaluates the accuracy of [18F]fluorodeoxyglucose positron emission tomography (18F-FDG PET) imaging with semi-quantitative analysis for differentiating benign from malignant pleural exudates and for guiding the search for the primary tumour of pleural metastases. Methods Whole-body 18F-FDG PET was performed in 79 patients with exudative pleurisy. Standard uptake values were normalized for body weight, body surface area, lean body mass (SUVbw, SUVbsa, SUVlbm) with and without correction for blood glucose levels. Thoracoscopy was systematically performed to reveal pathological diagnosis. Results All SUVs were significantly higher in all malignant pleural diseases (n = 51) than in benign (n = 28) (P < 0.001). Moreover SUVs were greater in the pleural metastases from pulmonary primaries (n = 25) and in mesotheliomas (n = 8) than in extrathoracic primaries (n = 18) (P < 0.01) with no significant difference between lung cancers and mesotheliomas. Receiver operating curve (ROC) analysis between benign and malignant lesions showed areas under the curves that ranged from 0.803 (SUVbsa g – 1)

Introduction Pleural effusions are a common disease medicine but their accurate diagnosis straightforward. In most of cases it is perform thoracocentesis. This procedure, useful, does not yield a precise diagnosis however [1,2].

in respiratory is often not necessary to although very in every case,

Pleural morphological imaging suffers from a lack of established specific criteria for malignancies [2–5]. Blind needle biopsies and, sometimes, eventually biopsies obtained during either pleuroscopy or open chest surgery, which yields the highest sensitivity for malignancies [6–8], may be needed. However, thoracoscopy is an invasive procedure and must be performed with caution in elderly patients and requires surgical facilities and trained staff. On the other hand, when a malignant pleural effusion is proved by histological biopsy, the search for the primary tumour requires the performance of several costly techniques.

to 0.863 (SUVbw). The cut-off value for SUVbw which gave the best accuracy (82.3%) was 2.2. When comparing thoracic with extrathoracic primaries the highest accuracy (80.4%) was found for a cut-off value of 2.6. Conclusion Semi-quantitative analysis of 18F-FDG PET imaging helps to differentiate malignant from benign pleural exudates and to distinguish between thoracic or extrathoracic primaries. Nucl Med Commun 27:971–976
c 2006 Lippincott Williams & Wilkins. Nuclear Medicine Communications 2006, 27:971–976 Keywords: positron emission tomography, fluorodeoxyglucose, pleural effusion, cancer Divisions of aPulmonary Medicine and bNuclear Medicine, University of Lie`ge, Belgium.

Correspondence to Dr Bernard Duysinx, Division of Pulmonary Medicine, CHU Sart Tilman B35, B-4000 Lie`ge, Belgium. Tel: + 32 436 67881; fax: + 32 436 68846; e-mail: [email protected]

Received 15 June 2006 Accepted 21 July 2006

accurately distinguish benign from malignant pulmonary tumours. Recent studies have shown the utility of 18FFDG PET in the evaluation of pleural diseases [10–20], particularly in diagnosis of mesothelioma. Most of these studies have investigated a low number of patients [10– 14,18,19]. Only a few studies have enrolled more than 30 patients [15–17,19]. In a previous study assessing 98 patients [17], we showed that 18F-FDG PET analysed by visual interpretation was helpful in the exploration for non-invasive pleural. To extend this study and to evaluate the usefulness of semiquantitative analysis we performed a new prospective study in 79 patients admitted to our hospital who required thoracoscopy for a diagnosis regarding pleural exudates. Before the invasive procedure, we performed whole-body 18F-FDG PET in each patient with semiquantitative analysis of pleural 18F-FDG uptake.

Material and methods Because of differences in glucose metabolism between normal and tumour cells, positron emission tomography using [18F]fluorodeoxyglucose (18F-FDG PET) [9] can

Patient selection

Seventy-nine consecutive patients (mean age of 62.8 ± 12.9 years; range 21–84 years; 47 males and 32

c 2006 Lippincott Williams & Wilkins 0143-3636


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972 Nuclear Medicine Communications 2006

2006, Vol 27 No 12

females) presenting with an exudative pleural effusion after thoracocentesis were investigated between 1999 and 2004. In chemical analysis, pleural effusion was considered as an exudate according to the criteria of Light et al. [21]; that is, if pleural effusion met at least one of the following criteria: Ratio of pleural fluid protein to serum protein greater than 0.5 K Ratio of pleural fluid lactic dehydrogenase to serum lactic dehydrogenase greater than 0.6 K Pleural fluid lactic dehydrogenase level greater than two thirds of upper limit of the normal value for serum lactic dehydrogenase

body mass SUVlbm ¼

Ctissue Dinj =lbm

where

 lbmf ¼ ð1:07bwÞ  148

 bw 2 bh

K

Neutrophilic pleurisy [22] and empyema were not included in this study. In all these patients the combination of chest X-ray, thoracic CT scanning (PQ 2000 4th generation, Picker, Cleveland, Ohio, USA) and thoracocentesis failed to give an aetiological diagnosis, thus justifying the realization of a thoracoscopy with pleural biopsies. 18

F-FDG PET study

An 18F-FDG PET study was performed on each subject before invasive procedures were carried out. 18F-FDG PET was initially performed with an UGM Penn PET 240H (UGM Medical Systems; Philadelphia, PA, USA) and later either with an ADAC UGM C-PET scanner (UGM Medical Systems, Pennsylvania, USA) or a Philips Allegro (Philips Medical Systems ALLEGRO, Brussels, Belgium). All patients fasted for 6 h prior to 18F-FDG PET and were tested for normal glucose levels before receiving the tracer. Emission scans from the base of the skull to the upper thighs were performed 73 ± 17 min, on average, after intravenous injection of 3.7 or 2.1 MBqkg – 1 of 18F-FDG, depending on the device. Images were constructed using an iterative algorithm and were fully corrected. The data were analysed by looking at the coronal, sagittal, and transverse slices alone and by cross-referencing. Regions of interest (ROIs) were placed on the most active pleural area of pathological regions which were first visually identified. The following standardized uptake values (SUVs) were calculated: SUVbw (normalized for body weight), SUVbsa (normalized for body surface area) and SUVlbm (normalized for lean body mass). SUVbw ¼

Ctissue Dinj =bw

where ctissue is the tissue concentration (in MBqml – 1) and Dinj is the injected dose (in MBq). The body weight (bw) is in grams. Corrections were then applied for lean body mass (lbm) [23] and body surface area (bsa) [24]. For lean

and

 2 bw lbmm ¼ ð1:1bwÞ  120 bh

where lbmf and lbmm are the lean body mass for female and male subjects, respectively; bw is the body weight (in grams), and bh is the body height (in centimetres). For body surface area SUVbsa ¼

Ctissue Dinj =bsa

where the body surface area (bsa) is in m2 and is given by bsa ¼ 0:007184bw0:425 bh0:725 These values were then corrected for glycaemia, assuming a normal blood glucose level of 5.55 mmoll – 1 (100 mgdl – 1) [25]: blood glucose : SUVg ¼ SUV 100 Normalization for blood glucose level gave SUVbw g – 1, SUVbsa g – 1 and SUVlbm g – 1. All SUVs are maximum pixel values (peak SUVs). Diagnosis of pleural exudate

The final diagnosis of the pleural effusion was obtained by invasive pleural biopsy during a thoracoscopy. When a diagnosis of benign disease was established, the patients were followed for at least 18 months to ensure absence of malignant pleural processes. The primary tumour was researched when pathology confirmed pleural metastases. Pleural malignancies were divided into three groups: pleural metastasis of extrathoracic cancer, pleural metastasis of primary lung cancer and mesothelioma. These two last sub-groups were considered as thoracic malignancies. Statistical analysis

The efficacy of 18F-FDG PET imaging in differentiating malignant from benign pleural lesions was evaluated using an unpaired t-test for each of six different SUVs. The four sub-groups were analysed by one-way analysis of variance (ANOVA) and by the Tukey–Kramer multiple comparisons tests. These analyses were obtained using the SAS version 9.1 software. ROCs and logistic regression analyses were calculated between benign and malignant pleural diseases, between extrathoracic tumour metastases and benign pleural lesions, between thoracic tumours (lung cancer and mesothelioma) and benign lesions, and finally between extrathoracic and thoracic tumours. Areas under

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SUVs in pleural malignancies Duysinx et al. 973

the curves (AUCs) were compared with a non-parametric approach [26]. Finally, clinical significance allowed the best estimation of the SUVbw cut-off for identifying pleural benign versus malignant diseases and extrathoracic versus thoracic pleural metastases.

Results Patient characteristics are presented in Table 1 and, apart from the gender ratio, are similar in each sub-group. There was no significant difference in blood glucose levels and delays after injection between sub-groups. Thoracoscopic biopsies showed benign lesions in 28 patients and malignant pleural effusions in 51. In the latter group, 18 were metastases of extrathoracic tumours (five breast cancers, three ovarian cancers, two kidney cancers, two gastric cancers, two pancreatic cancers, one colic tumour, one non-Hodgkin’s lymphoma, one sarcoma and one thymoma). Twenty-five pleural effusions were secondary to a pleural invasion of a lung cancer (five squamous non-small cell carcinomas, 10 adenocarcinomas, six large-cell carcinomas and four small-cell cancers). Mesothelioma was diagnosed in eight patients. Among the benign pleural diseases, the clinical context, bacteriological and histological data led to a final diagnosis of parapneumonic pleurisy (n = 5), tuberculosis (n = 3), chronic inflammatory changes (n = 12), asbestos pleurisy (n = 2), post-radic pleurisy (n = 5) and uraemic pleurisy (n = 1). The clinical course was that of a benign and selfTable 1

Patients’ characteristics

Characteristic

Benign pleurisies, n = 28

Malignant pleurisies, n = 51 Metastases Metastases Mesothelioma, of extrathor- of pulmonary n=8 acic tumour, tumour, n = 18 n = 25

Age (year) Gender Height (cm) Weight (kg) Blood glucose level (mmoll – 1) Delay after injection (min)

Table 2

64 ± 11 16M/12F 169 ± 9 66 ± 13 5.27 ± 1.5

72 ± 19

Quantification of

SUV SUVbw SUVbw g – 1 SUVlbm SUVlbm g – 1 SUVbsa SUVbsa g – 1

18

60 ± 18 8M/10F 167 ± 8 64 ± 11 5.49 ± 1.39

63 ± 12 17M/8F 170 ± 11 69 ± 16 5.94 ± 1.94

65 ± 9 6M/2F 169 ± 7 74 ± 27 4.94 ± 0.56

74 ± 13

73 ± 16

77 ± 20

limiting illness with complete resolution. So far, the follow-up for all patients with benign pleural disease has been at least 18 months with no disease recurrence or change to pleural malignancy diagnosis. Table 2 shows the average metabolic activity as measured using the various SUVs, according to the final diagnosis. Whatever the method used to calculate SUV, significant differences were observed between benign versus malignant group with higher values of SUVbw, SUVbsa and SUVlbm found in all malignant pleural exudates as compared to benign pleural lesions. The differences remained significant when the SUVs were normalized for blood glucose level. An example of SUVs in a benign versus a malignant pleural effusion is given in Fig. 1. ROC analysis calculated between these two groups demonstrated excellent sensitivity and specificity ratios whichever used SUV (Fig. 2). Normalizing for blood glucose level decreased significantly the diagnosis accuracy. SUVbw gave the highest AUC, although there was no statistical difference as compared to SUVbsa or SUVlbm (Table 3). From a clinical perspective a SUVbw of 2.2 gave the best accuracy (82.3%) for distinguishing between benign and malignant lesions (Table 4). Among malignancies, SUVs were higher in the pleural metastases from lung cancer and in mesotheliomas than in metastases from extrathoracic primaries (Table 2). In contrast to lung cancer, in which significant differences with extrathoracic tumours were observed for all SUVs (P < 0.01), a significant difference was found between metastasis of extrathoracic and mesothelioma only when the SUVs were normalized for blood glucose level (P < 0.05). There was no difference between lung cancers and mesotheliomas regarding any of the SUVs measured. Finally, when we compared the two sub-groups of pleural malignancy (extrathoracic mestastasis versus thoracic malignancy) (Fig. 3), a cut-off value of 2.6 for SUVbw gave the best accuracy, reaching 80.4% with 93.9%, 55.6%, 83.3% and 79.5%, respectively, for sensitivity, specificity, negative predictive value (NPV) and positive predictive value (PPV) (Table 5).

F-FDG uptake according to pathological results

Benign pleurisies n = 28

All malignant pleurisies n = 51

Metastases of extrathoracic tumour n = 18

Metastases of pulmonary tumour n = 25

Mesothelioma n = 8

1.9 ± 1.2 2.1 ± 1.6 1.4 ± 1.0 1.7 ± 1.3 0.04 ± 0.03 0.05 ± 0.04

4.7 ± 2.6*** 5.0 ± 3.2*** 3.6 ± 2.0*** 3.9 ± 2.5*** 0.11 ± 0.06*** 0.12 ± 0.08***

3.1 ± 1.8** 2.8 ± 1.5 2.4 ± 1.4** 2.2 ± 1.2 0.08 ± 0.05* 0.07 ± 0.04

5.5 ± 2.2*** 5.8 ± 2.8*** 4.3 ± 1.8*** 4.6 ± 2.3*** 0.13 ± 0.06*** 0.14 ± 0.08***

5.6 ± 3.8* 6.4 ± 4.6* 4.3 ± 2.8* 4.9 ± 3.4* 0.13 ± 0.09* 0.15 ± 0.11*

*

P < 0.05; P < 0.01; P < 0.001. Each sub-group is compared with benign pleurisies. SUV: standardized uptake value; bw, body weight; lbm: lean body mass; bsa: body surface area. **

***

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974 Nuclear Medicine Communications 2006

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Areas under the curves obtained from receiver operating characteristic curves between benign pleural lesions versus all malignant pleural exudates and between pleural metastases of extra-thoracic primaries versus thoracic malignancies (mestastases of lung cancer and mesotheliomas)

Table 3

Fig. 1

SUV

Area under the curve Benign Benign pleuri- Benign pleuri- Metastases of pleurisies vs. sies vs. metas- sies vs. metas- extrathoracic malignant tases of tases of tumour vs. mepleurisies extrathoracic thoracic tumour tastases of tumour thoracic tumour

SUVbw SUVbw g – 1 SUVlbm SUVlbm g – 1 SUVbsa SUVbsa g – 1

0.863 0.816*** 0.855 0.819*** 0.834 0.803***

0.751 0.648 0.737 0.647 0.702 0.632

0.924 0.891*** 0.919 0.894* 0.906 0.877*

0.779 0.812 0.773 0.818 0.763 0.800

*

P < 0.05; P < 0.01; *** P < 0.001. All comparisons were made with SUVbw. Abbreviations are as in the footnote to Table 2. **

Sensitivity, specificity and predictive values for different cut-off values for SUVbw obtained with receiver operating characteristics analysis between benign and malignant pleural effusions

Table 4

SUVbw

Cut-off value

Sensitivity (%) Specificity (%) NPV (%) PPV (%) Accuracy (%)

Example of patient with benign pleural disease (SUVbw,max = 1.1) (a), and pleural metastases from cancer in the right lung (SUVbw, max = 3.5) (b).

2.2

2.4

2.6

2.8

90.2

86.3

82.3

76.5

68.6

64.3

75

78.6

85.7

89.3

78.2 82.1 81

75 86.3 82.3

71 87.5 81

66.7 90.7 79.7

60.1 92.1 75.9

NPV: negative predictive value; PPV: positive predictive value. Other abbreviations are as in the footnote to Table 2.

Fig. 3

1.0

1.0

0.8

0.8

0.6 SUV bw SUV bw/g SUV1bm SUV1bm/g SUV bsa SUV bsa/g

0.4 0.2 0.0

Sensitivity

Sensitivity

Fig. 2

2

0.6 0.4

SUV bw SUVbw/g SUV1bm SUV1bm/g SUVbsa SUVbsa/g

0.2 0.0

0.0

0.2

0.4

0.6

0.8

1.0

1-Specificity Receiver operating characteristic curves of benign versus malignant pleural disease.

0.0

0.2

0.4

0.6

0.8

1.0

1-Specificity Receiver operating characteristic curves between pleural metastases of extrathoracic primaries and thoracic malignancies.

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SUVs in pleural malignancies Duysinx et al. 975

Sensitivity, specificity and predictive values for different cut-off of SUVbw obtained with receiver operating characteristics analysis between metastases of extrathoracic primaries and thoracic malignancies

Table 5

SUVbw

Sensitivity (%) Specificity (%) NPV (%) PPV (%) Accuracy (%)

Cut-off 2.4

2.6

2.9

3.5

4.7

93.9 38.9 77.8 73.8 74.5

93.9 55.6 83.3 79.5 80.4

84.8 61.1 68.7 80 76.5

72.7 61.1 55 77.4 68.6

51.5 83.3 48.4 85 62.7

Abbreviations are as in the footnotes to Tables 2 and 4.

Discussion The diagnosis of malignant pleural effusion is sometimes a challenging medical problem. The differentiation of malignant from benign pleural effusion is often difficult with the currently available non-invasive imaging techniques. A computed tomography (CT) scan alone cannot usually differentiate benign from malignant pleural disease [2–5], while thoracocentesis and blind or CTguided pleural biopsy lack sensitivity [1,6–8]. On the other hand, pathological proof of malignant pleural metastasis makes it necessary to seek the primary tumour, often using a large number of procedures, which may be a problem in patients in poor general condition. In this study, we have assessed for the first time the value of semi-quantitative 18F-FDG PET imaging analysis in the investigation of pleural effusions. Our results confirm the data available [10–20] including our previous series [17] regarding the application of 18F-FDG PET imaging. Our previous results, obtained using visual interpretation of the images, indicated that 18F-FDG PET was useful and far superior to pleural fluid cytology (62%) and blind pleural biopsy (44%) [6–8], and was virtually similar to pleuroscopy alone (95%) [7] for differentiating benign from malignant pleural lesions. The sensitivity of PET imaging usually ranges from 88.8 to 100% and the negative predictive value from 67 to 100% in diagnosing malignancy. In the present study, we show that a semi-quantitative analysis using the SUVs provides results of similar sensitivity for assessing pleural malignancy. From a methodological perspective, the SUV normalized for body weight, and not corrected for blood glucose level, is the best parameter for identifying malignant involvement, as demonstrated by the AUC obtained by ROC analysis (Fig. 1 and Table 3). There was no statistical difference between any of the SUV methods (normalized for body weight, body surface area or lean body mass) except when correcting for blood glucose level, which decreased the AUC in all cases. The quantification of 18F-FDG uptake in pleural effusion has, however, the advantage to yield cut-off values for distinguishing between benign and malignant effusions as well as between extrathoracic versus intrathoracic malignancies. Moreover, in opposition to visual analysis, the SUV quantification of metabolic

imaging gives an objective interpretation independent of the physician’s expertise. The most significant contribution of 18F-FDG PET to the management of pleural effusion is perhaps its ability to exclude malignant lesions. 18F-FDG PET should allow the clinician to avoid performing invasive diagnostic escalation in cases where a benign lesion is suspected by SUVbw less than 2.2 and may help to prevent complications associated with an invasive procedure, as well as unnecessary hospitalization of these patients. Considering the high sensitivity and negative predictive value of 18 F-FDG PET, it may provide valuable information, noninvasively, to evaluate pleural disease when pleural fluid analysis is impossible, the results are questionable or when it is difficult to obtain enough pleural fluid. Each sub-group of pleural malignancies displayed higher metabolic activity than benign lesions. The average SUV from extrathoracic malignancies was also lower than that measured in the group of thoracic tumours. This, in fact, is not surprising and may be explained by the pathological type of the extrathoracic tumours, some of which showed very low SUV: two breast cancers (SUVbw: 0.6 and 1.3), one kidney cancer (SUVbw: 1.4), one pancreatic adenocarcinoma (SUVbw: 1.4) and one gastric adenocarcinoma (SUVbw: 2.1). As the treatment for pleural malignancy depends on the histological nature of the tumour, increased 18F-FDG uptake should prompt invasive exploration to guide the treatment but ROC analysis between sub-groups of pleural metastases suggests that a cut-off value of 2.6 for the SUVbw may separate the metastases of extrathoracic cancer from thoracic malignancies. Thereby, a value for SUVbw provided by PET imaging may help to orientate the search for the primary tumour. As many as 15% of patients diagnosed with non-small cell lung cancer (NSCLC) will present with pleural effusion [27]. Identifying malignant pleural involvement associated with lung cancer is of paramount importance as it contributes prognostic information and is useful in the selection of patients amenable to surgery. T4 lung stage tumour includes malignant pleural effusion and defines locally extensive tumour, excluding surgical resection. Patients with primary lung cancer and no or mild 18FFDG uptake in the pleura (i.e. less than a value of 2.2 for SUVbw) should be considered as potentially resectable. It is of note that all post-radic pleurisies of patient with lung cancer treated by radiotherapy showed a very low SUVbw ( < 2.2) and there were no false positive results. We thus believe that 18F-FDG PET imaging can help in pleural staging of NSCLC and facilitate decision-making as to when to start invasive procedures. Patient with asbestos-related pleural disease require attention to identify malignant mesothelioma. In this

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976 Nuclear Medicine Communications 2006

2006, Vol 27 No 12

indication, some studies are interesting with regard to metabolic imaging with 18F-FDG PET using visual interpretation [11,17,19] as well as quantification with SUV [12]. Benard et al. [12,13] have already postulated that the uptake of 18F-FDG was significantly higher in malignant mesothelioma than in benign lesion and that a SUV cut-off of 2.0 provided a sensitivity of 91% and a specificity of 100%. These results are confirmed in our limited series of mesotheliomas, as one out of eight showed a SUVbw lower than 2.0 (SUVbw = 0.9). On the other hand, we did not find any significant difference in the SUV values between mesothelioma and pleural metastasis of the lung. It is thus unlikely that 18F-FDG PET imaging could be helpful in distinguishing these two types of thoracic tumour, for which the radiological presentation can be very similar. As we found that a cut-off value of 2.2 for SUVbw distinguished benign pleurisy from malignant pleural disease, including mesothelioma, we believe that monitoring SUVbw from 18F-FDG PET imaging might be particularly helpful in the follow-up of patients with asbestos-related pleural disease to pick up any malignant transformation.

Conclusions In this series of 79 patients, we confirm the high diagnostic accuracy of 18F-FDG PET imaging for identifying pleural malignancy. Furthermore, SUVs are helpful in giving objective metabolic information and guiding the search for the primary tumour. SUVbw values of 2.2 and 2.6 seem to be the best cut-offs for distinguishing benign from malignant pleural effusions and extrathoracic pleural metastasis from thoracic primary malignacies, respectively. The prognostic value of SUVs needs to be assessed in a prospective study.

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Original article

SPECT/CT imaging using a spiral CT scanner for anatomical localization: Impact on diagnostic accuracy and reporter confidence in clinical practice Paul J. Roach, Geoffrey P. Schembri, Ivan A. Ho Shon, Elizabeth A. Bailey and Dale L. Bailey Purpose To evaluate the incremental benefit in routine clinical practice of computed tomography (CT) scans acquired for anatomical localization on an integrated SPECT/CT which incorporates a spiral CT scanner, in comparison with conventional planar and SPECT scanning. Methods The first 50 studies acquired on the integrated system were evaluated by two experienced nuclear medicine physicians who were aware of the patient’s clinical history. These included bone scans, gallium scans, octreotide scans, sestamibi parathyroid scans and MIBG scans. For each patient study, abnormalities were assessed on planar and SPECT images for location and provisional diagnosis and a quantitative scale was used to assess reporter confidence. The fused SPECT/CT images were then reviewed and the location and provisional diagnosis noted and reporter confidence was assessed using the same quantitative scale.

10% and a significant change in 9% over planar/SPECT imaging. The confidence of diagnosis was improved moderately in 10% and improved significantly in a further 10% of cases. For the final scan interpretation, there would have been no change in 44% patients, a minor change in 30% and a significant change in 26% with the use of SPECT/CT. Conclusion Use of integrated SPECT/CT with a high spatial resolution, spiral CT used for anatomical localization improves accuracy and reporter confidence in clinical practice. As a result, final reports were different in 56% of the cases, including being significantly different in 26% patients compared to reporting with planar/SPECT c 2006 Lippincott alone. Nucl Med Commun 27:977–987 Williams & Wilkins. Nuclear Medicine Communications 2006, 27:977–987 Keywords: hybrid imaging, image fusion, SPECT/CT

Results There were 129 abnormalities detected in 50 patient studies. For localization of abnormalities, the inclusion of the CT resulted in a minor change in 16% of cases and a significant change in 11% over planar/SPECT imaging alone. The confidence of localization was improved moderately in 19% and improved significantly in 6%. For diagnosis, SPECT/CT resulted in a minor change in

Department of Nuclear Medicine, Royal North Shore Hospital, Sydney, Australia.

Introduction

emission tomography (PET) imaging has been dramatic. The medical literature has increasingly demonstrated the clinical benefits of hybrid scanners over either PET or CT alone in cancer imaging and nearly all PET scanners sold today are now hybrid machines [4,5].

Functional imaging techniques, such as nuclear medicine, have been used for many years to provide important clinical information about various disease processes. These techniques provide unique information about the function of the body and often demonstrate disease prior to abnormalities being detected on anatomical imaging modalities, such as plain X-ray, computed tomography (CT) or magnetic resonance imaging (MRI) [1,2]. However, functional imaging is hampered by a lack of anatomical detail and the complementary nature of functional and anatomical imaging modalities is being increasingly recognized [3]. In 2001, the first hybrid PET/CT camera was commercially released and the impact of this advance on positron

Correspondence to: Dr Paul Roach, Department of Nuclear Medicine, Royal North Shore Hospital, Pacific Highway, St Leonards, Sydney, NSW, Australia. Tel: + 61 2 9926 8375; fax: + 61 2 9906 1124; e-mail: [email protected] Received 23 May 2006 Accepted 22 July 2006

While the impact of hybrid imaging on PET scanning is undisputed, there is less literature relating to its impact on conventional nuclear medicine and single photon emission computed tomography (SPECT) imaging. As many SPECT radiopharmaceuticals are relatively targetspecific, it has been suggested that the impact of hybrid imaging in SPECT imaging will be even greater [2,6]. In 2000, GE Medical Systems commercially released a hybrid SPECT scanner that integrates a dual-head variable angle gamma camera with a low dose X-ray tube

c 2006 Lippincott Williams & Wilkins 0143-3636

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978 Nuclear Medicine Communications 2006, Vol 27 No 12

[7]. Numerous series and case studies have reported the improved diagnostic accuracy of this hybrid scanner over SPECT imaging alone in conditions such as lymphoma [8], infection [9], bone disease [10], neuroendocrine tumours [11,12], parathyroid adenomas [13,14], thyroid cancer [15,16], adrenal tumours [17], cavernous haemangioma [18,19] as well as with lymphoscintigraphy [20,21], hepatic artery perfusion scintigraphy [22] and a combination of common clinical scanning indications [23]. As this camera uses a relatively low performance Xray scanning device (compared with a spiral diagnosticquality CT), image acquisitions are slow (in the order of 10 min) and the images produced are not of high diagnostic quality [1,7–9,13,15,16,22]. The lengthy acquisition time is less than ideal as it may increase the chances of patient movement which can result in misregistration between the emission and transmission data, thus reducing the overall accuracy of the fusion process [15] and restricts patient throughput. In 2005, we developed a hybrid SPECT/CT scanner which integrates a Philips Skylight variable angle, dualhead gamma camera with a single-slice Picker PQ5000 spiral CT [24]. Later the same year, commercial hybrid SPECT/CT systems which integrate current generation gamma cameras with multi-slice CT scans, with configurations varying from single slice up to 64 slice machines, became available. Compared with the GE ‘Hawkeye’ system, the use of spiral CT scanning results in much faster image acquisitions ( < 1 min) and images produced are of much higher quality, and should therefore produce higher diagnostic accuracy. However, the impact on routine clinical practice of systems which incorporate true spiral CT technology has not been previously reported and is unknown. The aim of this study was to evaluate the incremental benefit in routine clinical practice of anatomical localization provided by an integrated SPECT/CT scanner which incorporates a spiral CT scanner compared with routine planar/SPECT scanning.

Materials and methods Patients and study types

This was a retrospective study that analysed the first 50 studies performed on the hybrid scanner. Patients who underwent spiral SPECT/CT were those who would have previously undergone SPECT imaging of at least one region of the body, either to evaluate an abnormality seen on planar scanning, or as a routine study based on the likely benefit in various clinical conditions, e.g., lymphoma. In our department, SPECT imaging of at least one region is routinely performed in all patients having 67Ga imaging for lymphoma or infection as well as patients who have 111In-octreotide or meta-iodobenzylguanidine

(MIBG) imaging for neuroendocrine tumours. For other scan types, e.g., 99mTc bone scans, SPECT is performed in selected cases based on the clinical question and the body region being assessed. As there have been several publications demonstrating the clinical benefit of hybrid imaging (using a low-dose X-ray tube system) in each of the conditions studied, ethics committee approval was not required for this study. Imaging protocols Planar imaging

All patients in this series had planar whole-body scans or spot views performed on either the Philips Skylight variable angle dual-head gamma camera (16 mm thick NaI(Tl) crystal) (Philips Medical Systems, Milpitas, California) or a Picker 2000XP (Picker, Cleveland, Ohio) fixed dual-head gamma camera. SPECT and SPECT/CT imaging

The SPECT/CT system that we have devised requires that the gamma camera bed is removed from its usual position and replaced by the CT bed for a SPECT/CT study [24]. The CT bed has been modified so that routine CT and SPECT scanning can be performed without moving the patient. The CT bed moves with ease along a track, which can be locked in either the CT position or SPECT position. This reduces misregistration artefacts as no change in patient positioning is required between studies. As the CT and SPECT scanners are positioned in a fixed, reproducible spatial relationship it is possible to measure the y-axis and z-axis translations to achieve accurate co-registration. There are no x-axis translations required as the isocentre of CT scanner is aligned with the axis-of-rotation of the SPECT scanner. The software fusion of the CT and SPECT data has been previously verified by scanning a variety of line and point sources and complex objects to confirm the accuracy of alignment [24]. For all patients studied in this series, the CT scan was performed first and then the bed positioned so that the SPECT study could be done immediately after the CT acquisition. Low-dose CT acquisition parameters were used: 30–50 mA, 130 kVp, 512  512 matrix, pitch of 1.5, table speed of 1 cm
s – 1 and field of view (FOV) of 400– 450 mm. Typically, the acquisition time for the CT to cover this FOV was 40–50 s per study. Patients were instructed to breathe normally and the intention was to acquire the CT during tidal breathing. Routine CT studies were reconstructed using a slice thickness of 4 mm, which is reduced to 1.5–2 mm for imaging small structures such as parathyroid and sentinel node imaging. On completion of the CT scan, the bed translates to the SPECT position, the bed height was set to match the default SPECT bed height, and the study acquired using default SPECT parameters with the detector heads in a 1801 opposed configuration and a 128  128 matrix, with

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Imaging using a spiral CT scanner Roach et al. 979

31 steps, step-and-shoot mode, a non-circular orbit and a time per projection of 15–25 s depending on the type of studying being acquired. High resolution collimators (Philips ‘VXGP’) were used for all 99mTc studies, medium energy, general purpose collimators used for 111In and 67 Ga studies, and high energy general purpose collimators used for 131I studies. CT and SPECT studies were acquired with the patient’s arms above the head in 48 cases and with the arms at the patient’s side in two cases. The SPECT studies were acquired on a Philips Jetstream acquisition station and were processed and displayed using a HERMES workstation (Nuclear Diagnostics AB, Stockholm). During SPECT acquisition, the CT study was reconstructed and automatically transferred to the workstation. Each SPECT study was reconstructed using ordered subset expectation maximization (OSEM) algorithm, with eight subsets, four iterations and 128 slices. Interactive three-dimensional post-filtering was applied to the transverse sections. Registration of SPECT data to CT data was performed using the Multimodality program on the workstation which uses a mutual information cost function to register the data sets. Auto-registration using only two degrees of freedom (y-axis and z-axis translation) is required to register these data sets. In six studies (12%), manual adjustments were used to improve accuracy of registration, but this was only required in studies with either low counting statistics (67Ga, n = 4) or limited anatomical detail on SPECT (111In-octreotide, n = 2). It should be noted that while image fusion using our system is based on software registration (as opposed to PET/CT and the new commercial SPECT/CT scanners which use spatial bed position), even with a fully integrated PET/CT (and therefore presumably SPECT/CT), it is widely recognized that there is often the need for a final manual adjustment to achieve optimum registration accuracy [25]. The reconstruction and registration process took typically less than 10 min per study, dependent on the number of slices in the CT data set. The volumes were then saved as a co-registered pair and viewed using a fusion display program. Image interpretation

Each study was reviewed by two of three experienced nuclear medicine physicians who were aware of the clinical history but blinded to the results of other investigations. If any of the reporting physicians had reported the study previously during routine clinical practice, that reporter was excluded and the other two physicians were used to report the scan for the purposes of this study. The reporters were not specialist radiologists but all had undergone formal training in crosssectional CT anatomy and informal supervision in CT reporting. Scans were reviewed as per our routine clinical practice, i.e., planar images were reviewed on film on a viewing box and SPECT studies reviewed interactively on the workstation.

For each abnormality, or possible abnormality, seen on SPECT or planar imaging, the location of the lesion and the provisional or differential diagnosis was recorded as would be done clinically in our department. Also confidence for both location and diagnosis was rated using the following five-point scale: (1 = little confidence, 2 = some confidence, 3 = moderate confidence, 4 = good confidence, 5 = high confidence). The two reporters reviewed each case concurrently and consensus was used in the event of disagreement. The SPECT/CT fused images were then reviewed on the workstation and the same assessment of lesion location and the provisional diagnosis was made and our confidence for both location and diagnosis was recorded using the same five-point scale. When determining whether there was significantly more information provided by SPECT/CT, a change of at least two points was required using the five-point scale. For example, if confidence changed from 2 to 3, this was considered to be only a minor change, but if confidence increased from 2 to 4, this was considered to be a significant change. Finally, we assessed whether the SPECT/CT had resulted in a change in final diagnosis and scan interpretation. This was assessed semi-quantitatively using a three-point scale with 0 indicating no change, 1 indicating a minor change and 2 indicating a significant change. This was somewhat arbitrary but localizing a lesion from one lymph node to another was considered to be a minor change whereas localizing an abnormality, for example, in bone instead of lung, was considered to be a significant change. This scoring was based on the reporter’s assessment of their scan interpretation and was not based on whether patient management would be altered. The same scaling was used to assess any impact on our final diagnosis for each abnormality.

Results The 50 patient studies included 99mTc-diphosphonate bone scans (n = 13), 67Ga citrate scans (infection n = 18, lymphoma n = 10), 111In-octreotide scans (n = 5), 99mTcsestamibi parathyroid scans (n = 2) and 123I-MIBG or 131 I-MIBG scans (n = 2). The clinical indications for each are shown in Table 1. The series comprised 29 males and 21 females. In these 50 scans, SPECT and SPECT/CT studies of the chest were performed in 24 cases (including the neck and skull in eight cases), the abdomen/pelvis in 25 cases and the lower limbs in one patient. Site analysis

A total of 129 abnormalities (sites), or possible abnormalities, were identified in these 50 patient studies. For location of lesions, the addition of SPECT/CT data resulted in a minor change in 16% of sites and a significant change in 11% of sites (see Table 2). There

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980 Nuclear Medicine Communications 2006, Vol 27 No 12

Table 1

Number of patients, scan type and clinical indication

Study type

Clinical indication

Sub-total

Total

Bone scan

Infection Staging carcinoma Low back pain Foot pain Neck pain Infection–PUO Infection–otitis externa Infection–low back pain Lymphoma Carcinoid Cushing’s disease Hyperparathyroidism Carcinoid Phaeochromocytoma therapy

4 4 3 1 1 14 1 3 10 4 1 2 1 1

13

Gallium scan

Gallium scan Octreotide scan Parathyroid MIBI MIBG Total

Table 2

Bone scan Gallium scan–infection Gallium scan–lymphoma Octreotide scan Parathyroid MIBG Total

Table 3

Patient number

Study type

2 3

Bone scan Bone scan

4 16

Bone scan Gallium–infection

19 31

Gallium–infection Gallium–infection

2 2

32

Gallium–infection

50

34

Gallium–lymphoma

35

Gallium–lymphoma

38

Gallium–lymphoma

39

Gallium–lymphoma

43

Gallium–lymphoma

18

10 5

Additional information provided by SPECT/CT by site

Study type

Number of sites 41 48 29 5 3 3 129

No change 34 32 19 4 3 2 94

(82%) (67%) (66%) (80%) (100%) (67%) (73%)

Minor change 4 11 5 1

(10%) (23%) (17%) (20%) 0 0 21 (16%)

Significant change 3 (8%) 5 (10%) 5 (17%) 0 0 1 (33%) 14 (11%)

Additional information provided by SPECT/CT by patient

Study type Bone scan Gallium scan–infection Gallium scan–lymphoma Octreotide scan Parathyroid MIBG Total

Number of patients 13 18 10 5 2 2 50

No change 4 7 5 4 2

Minor change

(31%) 6 (39%) 7 (50%) (80%) 1 (100%) 0 1 22 (44%) 15

(46%) (39%) 0 (20%) 0 (50%) (30%)

Additional information provided by SPECT/CT compared with SPECT in 13 patients with final reports considered to be significantly different due to use of SPECT/CT

Table 4

Significant change 3 (23%) 4 (22%) 5 (50%) 0 0 1 (50%) 13 (26%)

was a minor change in confidence of lesion localization in 19% and a significant change in 6% of cases. For final diagnosis, adding the CT data resulted in a minor change in 10% of sites and a significant change in 9% of sites. There was a minor change in confidence of final diagnosis in 10% and a significant change in 10% of cases. Patient analysis

Table 3 shows the impact of SPECT/CT on overall final reports in the 50 patients. SPECT/CT resulted in no change to the final overall scan interpretation in 44% of patients, a minor change in 30% and a significant change in 26% of the 50 patients in the series. The details of how the use of SPECT/CT resulted in a significant change in these 13 patients are listed in Table 4.

43

131

I-MIBG therapy

SPECT findings

SPECT/CT findings

Facet joint arthritis L2 Osteomyelitis L2 R petrous temporal Mastoid osteomyelitis osteomyelitis T4 osteomyelitis T4/5 discitis Psoas abscess Psoas abscess + L2 osteomyelitis T10 infection/arthritis T8/9 discitis Focal infection RUQ Physiological bowel activity Arthritis R shoulder L humeral head osteomyelitis Prevertebral lympho- Vertebral lymphoma ma of lymph nodes Left lower abdominal Physiological bowel lymphoma activity Paravertebral disease Lymphoma T11 pedicle and facet joint Physiological bowel Iliacus lymphoma activity L3 lymphoma Degenerative change/ compression R adrenal bed L1 metastasis

Results by study type 99m Tc-diphosphonate bone scans

Bone scans were performed in 13 patients with the clinical indications listed in Table 1. The use of SPECT/ CT resulted in a change in site localization in 10% (minor change) and 8% (significant change). Final scan interpretation was significantly altered based on the information provided by SPECT/CT in 23% of patients (see Tables 3 and 4). The improved localization which was provided by SPECT/CT appeared to be most helpful in the spine where it led to better differentiation of vertebral abnormalities and better definition of the exact vertebra, or part of the vertebra, involved. Accurate localization of vertebral levels can be particularly problematic in the cervical spine and we found SPECT/CT to be particularly helpful in this area, with the affected vertebral level changed as a result of its use in one case. In the axial skeleton, SPECT/CT provided more accurate localization which allowed for conditions such as facet joint arthritis, vertebral osteomyelitis and disciitis to be better differentiated (two cases). There was less impact on the appendicular skeleton with no cases providing additional information over SPECT alone. There was a greater impact for patients being investigated for infection (with three patients having a significant change in final scan interpretation and one having a minor change from a total of four patients) than for evaluation of bone metastases (with an impact only seen in one case, resulting in the level of vertebral involvement being altered in the cervical spine). Hybrid imaging had an impact on all three patients being investigated for low back pain, but this was regarded as minor in each case. In this group, SPECT/CT differentiated an osteophyte from

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Imaging using a spiral CT scanner Roach et al. 981

what was interpreted to be physiological activity in the right ureter, differentiated osteophyte formation from costovertebral arthritis and demonstrated partial compression in a degenerative vertebral body that was considered to be predominantly degenerative disease on planar/SPECT imaging alone. 67

Ga citrate scans

Gallium scintigraphy was performed in 18 cases with suspected infection. Of these, 14 cases were for pyrexia of unknown origin (PUO), three were for low back pain and one was for otitis externa. A total of 48 abnormalities, or possible abnormalities, were detected in these patients and SPECT/CT resulted in a minor change in the localization of 23% and a significant change in 10%. Final scan interpretations were significantly altered in 22% of patients. Of the patients being assessed for PUO, SPECT/CT resulted in a major change in the scan interpretations of four patients (see Table 4) and a minor change in seven others. Minor changes included differentiation of bone marrow hyperplasia from anterior osteophytes (one case) and more accurate differentiation of bone and soft tissue involvement in the thoracolumbar spine (three cases). In the three cases of low back pain, only one case was altered by the use of SPECT/CT. In this patient, hybrid imaging demonstrated uptake to involve the anterior vertebrae superficially whereas uptake was thought to be localized to prevertebral soft tissue based on SPECT alone. The one case of otitis externa studied had a minor change to the final scan interpretation as SPECT/CT was better able to demonstrate the extent of soft tissue involvement. An example of a case of suspected infection where SPECT/CT significantly impacted the final diagnosis is shown in Fig. 1. Lymphoma was the clinical indication for 10 patients studied in this series. Of these, five patients had their final scan interpretation significantly altered by the use of SPECT/CT (see Table 4) with no change being noted in the other five patients. Of 29 sites of abnormality, there was a minor change noted in the localization of 17% abnormalities and a significant change noted in another 17% of sites. The value of SPECT/CT is highlighted in the case of an 81-year-old man with disseminated nonHodgkin’s lymphoma described in Fig. 2. 111

In-octreotide scans

Octreotide scanning was performed in five patients in this series. In only one case did SPECT/CT impact on final diagnosis by differentiating pathological uptake from physiological gallbladder uptake. It should be noted that three cases revealed no abnormal uptake of 111 In-octreotide so the true impact of hybrid imaging with this radiopharmaceutical may be underestimated by this small series.

123

I-MIBG and

131

I-MIBG scans

MIBG scintigraphy was performed on two patients in this series. In one case, diagnostic scintigraphy was performed with 123I-MIBG to evaluate liver metastases and assess other sites of uptake in a patient with carcinoid syndrome. The ability of SPECT/CT to confirm uptake of MIBG in the liver lesions was considered to be of a useful advantage of hybrid imaging. The other patient was scanned 8 days after receiving an 8 GBq ablation for disseminated phaeochromocytoma and the benefit of SPECT/CT is outlined in Fig. 3. In his case, SPECT/CT localized a lesion to the L1 vertebral body, instead of the right adrenal bed region as was suspected on SPECT only. The appearances were deceptive on planar imaging and SPECT due to the patient’s marked scoliosis but SPECT/CT clearly revealed the exact anatomical location of the lesion. This finding was considered to be a significant change over planar/ SPECT alone. 99m

Tc-sestamibi parathyroid scans

Parathyroid scintigraphy using 99mTc-sestamibi was performed on two patients in this series. Both patients had parathyroid adenomas localized to the thyroid region. SPECT/CT was considered to provide no additional information over SPECT alone. Neither patient had an abnormality in the mediastinum.

Discussion The last decade has seen major developments in the successful image fusion of the unique functional information provided by nuclear medicine techniques with the exquisite anatomical information provided by radiological modalities, such as CT and MRI [2]. While comparing different scans side by side (so-called ‘visual fusion’) is still commonly practised in many institutions, there have been significant advances in both software (i.e., scans acquired on different devices during separate scanning procedures) and hardware (on a combined, hybrid scanner in a single imaging session) image fusion in recent years [2]. While software fusion is becoming more reliable, its accuracy continues to be limited by the flexible and deformable nature of the body which can often make realignment of organs and structures across different imaging examinations difficult [2]. In addition, the optimal protocol and positioning for a radiological procedure can be very different to a SPECT procedure, an example being the use of a breath-hold and its impact on the conformity of the thorax. Hardware fusion has the advantage of eliminating potential causes of misregistration such as those related to different bed shapes, patient positioning and changes to gastrointestinal or urinary tract contents [1]. However, misregistration due to respiratory motion can still occur in the chest and upper abdomen [1]. Despite this potential limitation, the success of integrated PET/CT has heralded a new era in medical imaging in which hardware fusion is being increasingly

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Nuclear Medicine Communications 2006, Vol 27 No 12

Fig. 1

(a)

Posterior

Anterior (b)

CT

SPECT

SPECT/CT

Transaxial

Coronal (a) Gallium infection imaging. Anterior and posterior planar images of an 84-year-old male with Staphylococcus aureus septicaemia. Planar images show right paravertebral uptake which was considered to be likely to represent a psoas abscess (arrowed). Right abdominal uptake was considered likely to represent physiological bowel activity. CT and MRI (done 10 days earlier) revealed thoracolumbar degenerative changes and right psoas enlargement. No abscess was seen. (b) Transaxial (top row) and coronal (bottom row) images of SPECT (left), CT (middle) and fused SPECT/CT (right) of the mid abdomen show increased gallium accumulation in the upper right psoas region. It was considered on SPECT imaging only that uptake was located in muscle and probably in an adjacent soft tissue collection. SPECT/CT revealed the uptake was actually located in a large anterolateral osteophyte with extension into the adjacent L2 vertebral body which indicated infection in bone in addition to the psoas muscle. SPECT/CT confirms abdominal uptake elsewhere is located in bowel, indicative of physiological bowel activity. The enlarged right psoas was aspirated in its mid portion (arrowed) under CT control and as only 2 ml blood was removed, it was assumed that this was a haematoma. However, cultures did grow Staphylococcus aureus and as SPECT/CT showed much greater gallium accumulation more superiorly, aspiration more superiorly would presumably have indicated more active infection. Hybrid imaging can help guide biopsy sites in such cases. Note that the CT quality was degraded due to the patient’s body habitus (weight 105 kg) and the positioning of the patient’s arms at his side (due to inability to keep them above his head).

accepted as the most reliable and accurate method to ensure precise image co-registration in medical imaging today. With its poorer spatial resolution, we have previously suggested that SPECT has even more to gain from accurate image fusion that PET [2,6]. In addition, many radiopharmaceuticals used in SPECT imaging are highly specific and therefore scans may demonstrate few, if any, anatomical landmarks making accurate image interpreta-

tion difficult. This applies to some commonly used radiopharmaceuticals, such as 131I, 111In-octreotide and radiolabelled leucocytes, as well as newer agents such as radiolabelled monoclonal antibodies, receptor agents and peptides [2,15]. As these agents are likely to be used increasingly in clinical practice in the future, there is a clear need to ensure that software techniques and hardware are developed which will facilitate accurate image fusion for current and future SPECT radiopharmaceuticals.

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Imaging using a spiral CT scanner Roach et al. 983

Fig. 2

(a)

Anterior

(b)

Posterior

SPECT

CT

SPECT/CT

Conronal

Transaxial (a) Gallium lymphoma imaging. This patient was an 80-year-old male with recurrent B-cell non-Hodgkin’s lymphoma. Anterior and posterior planar images show multiple abnormalities above the diaphragm. It was considered that most of the abdominal uptake represented physiological bowel activity. (b) Abdominal coronal images (top row) of SPECT (left), CT (middle) and fused SPECT/CT (right). It was considered on SPECT that abdominal uptake represented bowel activity, although SPECT/CT was able to differentiate bowel accumulation from uptake in the left iliacus region. CT (middle) confirms lymphomatous involvement (arrowed). Lower thoracic images (bottom row) at the T10 level show focal accumulation in the left paravertebral region. It was uncertain on SPECT alone whether this was in soft tissue, bone or pleura. SPECT/CT confirmed bone involvement with destruction of the pedicle evident on CT.

Since the release of the GE Hawkeye SPECT system in 2000, there have been an increasing number of series and case reports that have documented the improvement in diagnostic accuracy in various settings where it has been used. This is a hybrid system that uses a low performance

X-ray tube and it has been reported to provide adequate information for the precise assessment of SPECT findings in most cases [1]. In a study with a similar design to ours, Schillaci et al. [23] assessed the impact of hybrid imaging using this scanner in 81 patients with

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984 Nuclear Medicine Communications 2006, Vol 27 No 12

Fig. 3

(a)

Posterior

Anterior (b)

SPECT

CT

SPECT/CT

(a) MIBG therapy. The patient was a 60-year-old male with a past history of left phaeochromocytoma resected 8 years earlier. Diagnostic 123I-MIBG scintigraphy revealed metastatic disease involving the right upper abdomen and lungs. Anterior and posterior planar imaging (left) performed eight days after an 8 GBq ablation showed multiple abnormalities, the most intense being in the right adrenal area. Note no abnormality in the midline. On coronal SPECT/CT imaging (right), uptake is seen in the L1 vertebral body, an unexpected and unknown finding. The patient’s marked scoliosis (seen on CT) explains why the vertebral uptake is located to the right of the midline. Uptake in a pulmonary metastasis is also noted. (b) Transaxial SPECT (left), CT (middle) and SPECT/CT (right) at the L1 level shows uptake in the right adrenal bed with extension posteromedially into the vertebral body. The bony metastasis can be seen on the CT.

various clinical situations using different radiopharmaceuticals and found that hybrid imaging improved accuracy in B40% of cases compared with SPECT alone. They reported that hybrid imaging provided correct localization of SPECT findings in 28% patients, allowed exclusion of disease at sites of physiological radiopharmaceutical uptake in 10% of cases and defined the functional significance of CT lesions in 3% of patients. More recently, several series have been published assessing the impact of this camera in specific conditions, such as lymphoma, thyroid cancer, infection imaging and the evaluation of liver lesions. Palumbo et al. [8] reported in a small series of 24 patients with lymphoma that hybrid imaging provided additional data in 54% patients and resulted in a change in therapeutic approach in a third of all patients. In a series of 82 patients with suspected infection studied with either gallium or radiolabelled leukocytes, Bar-Shalom et al. [9] reported that hybrid

imaging provided additional information for infection diagnosis and localization in 48% of patients. In the evaluation of hepatic haemangiomas, Schillaci et al. [18] reported a significant impact of hybrid imaging in four of 12 patients studied. In a large series of patients with thyroid cancer, Tharp et al. [16] reported that hybrid imaging provided incremental diagnostic information over planar imaging in 57% of 71 patients. As a result, there was a change in management in 41% of patients by avoiding unnecessary therapy and allowing appropriate surgical planning in cases where surgery was warranted. The evaluation of neuroendocrine tumours is one area where it is considered that the impact of hybrid imaging may be particularly important [23]. In a series of 72 patients with various neuroendocrine tumours, Krausz et al. [26] reported that hybrid imaging improved localization in 23 of 44 positive studies. As a result, 14% of patients had their management altered by modifying

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Imaging using a spiral CT scanner Roach et al. 985

therapy, optimizing surgery or avoiding futile surgical procedures. A significant impact has also been reported in MIBG imaging where the false positive rate can be reduced by differentiating abnormal sites of uptake from sites of physiological uptake, such as the kidneys, spleen and heart [17]. A valuable impact in bone scintigraphy has also been noted with an improvement demonstrated in both the localization and characterization of lesions [10]. In this series, 85% of lesions were correctly localized by hybrid imaging and 81% of equivocal SPECT lesions were accurately characterized. While there is consistent literature supporting the benefit of hybrid imaging with the Hawkeye in an increasing number of clinical conditions, this system has the limitation that the anatomical information is provided by a low performance X-ray tube, rather than a current generation spiral CT scanner [1]. In addition to relatively lengthy image acquisition times, the images provided are low resolution and not of the quality seen with current generation diagnostic quality CT scanners [1]. To our knowledge, this is the first report on the impact of hybrid SPECT scanning using a spiral CT scanner for anatomical localization in a clinical setting where a variety of different radiopharmaceuticals and clinical indications have been assessed. Our findings of a change in location in 28% of sites, change of diagnosis in 19% of sites and change in final reports noted in 56% of cases (with 26% being considered significant) confirm the advantages of hybrid imaging in clinical practice. The impact of hybrid imaging in our series is greater than the 41% quoted in the series using a low-dose X-ray scanner by Schillaci et al. [23], but it should be noted that it is difficult to directly compare the two series given the different types of clinical indications and radiopharmaceuticals used. Compared to published literature, the clinical impact of hybrid scanning using a spiral scanner appears to be of a similar, or greater, magnitude to that noted in many series which have used the Hawkeye system, although this is probably due to our use of the CT data for anatomical localization only. In our series, there was a higher impact of SPECT/CT on a per patient basis compared with per site. This is explained by the fact that sites where hybrid imaging significantly affected final interpretations were distributed across a large number of the patients in our series, rather than being concentrated in just a small number of patients. This supports the notion that while not all sites of disease (or sites of physiological distribution) will benefit from hybrid imaging, there will be some gain in a majority of patients when multiple areas are being assessed. It should also be noted that while there were reasonable patient numbers in this series to assess the impact of hybrid imaging on bone and gallium scintigraphy, it is harder to draw conclusions on its impact on parathyroid scintigraphy or 111In-octreotide imaging

due to small patient numbers. As few of our patients had disease demonstrated on 111In-octreotide, the impact of hybrid imaging will undoubtedly be underestimated. Similarly, the most likely benefit of hybrid scanning in parathyroid imaging will be in the evaluation of the ectopic lesions (especially in the mediastinum) although both of our cases in this series had adenomas in the neck, an area where pinhole and SPECT techniques have well documented high accuracy [27,28]. While spiral and multidetector CT scanning will produce superior image quality compared to a low-performance Xray tube and improved accuracy is therefore to be expected, the overall incremental impact in clinical practice over the low performance X-ray scanner may only be limited given the inherent relatively poor resolution of SPECT imaging. Much of the benefit of hybrid imaging comes from the ability to localize a SPECT radiopharmaceutical to a particular organ, or region of that organ, rather than requiring exact precision with millimetre accuracy. It may be more likely that the real benefit of spiral and multidetector CT scanning over low performance X-ray systems will be in the acquisition times of each study, or when imaging small structures such as lymph nodes. Compared with the 10 min acquisition required for the fixed slice thickness (5 or 10 mm) Hawkeye system, spiral scanners will acquire their data with finer axial sampling in less than 1 min. This results in faster overall acquisition times, less impact on patients (especially those that are sick and need to have imaging performed as rapidly as possible), and will facilitate the use of hybrid imaging, particularly in busy departments where camera time may be limited and long X-ray acquisition times may be a disincentive for the use of hybrid imaging. With the advent of multislice CT hybrid systems, there is the potential for high diagnostic quality CT scanning to be performed concurrently with, or sequentially to, SPECT imaging. This is likely to be of particular benefit in oncology imaging where essential, but complementary, information is provided by anatomical and functional imaging. High resolution CT scanning should allow for better characterization of lesions compared with low performance hybrid scanners, and may reduce the need for subsequent diagnostic quality studies to evaluate abnormalities seen on SPECT. In our series, CT data were used only for anatomical localization, and studies were reported by nuclear medicine physicians with limited training in cross-sectional anatomy and CT reporting. For this reason, the overall impact of hybrid imaging that we have demonstrated is almost certainly underestimated as a greater benefit would be expected if full diagnostic studies were performed (i.e. with intravenous and oral contrast) and if interpretation was done by radiologists fully trained in CT reporting.

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986 Nuclear Medicine Communications 2006, Vol 27 No 12

Nevertheless, as new commercial SPECT/CT are installed in imaging departments around the world, many centres will utilize these scanners in the same way that we have done and the benefit that we have shown is important to note, especially for centres that will use CT data only for anatomical localization and where studies will be read by nuclear medicine physicians. It is also likely that newer applications, such as coronary artery imaging and visualization of cerebral blood flow will also be facilitated by multislice hybrid technology as will more accurate attenuation correction for various radionuclides [1]. The potential advantage of spiral CT over hybrid imaging using a low-performance X-ray tube has been noted in the evaluation of the abdomen with gallium scintigraphy where misleading information has been demonstrated with the low-performance system [9]. However, the use of spiral CT may increase radiation exposure to patients, although it is possible to minimize this increase. In our series, we routinely set acquisitions to 30 mA, although this is increased to 50 mA in larger patients ( > 90 kg) to avoid suboptimal image quality (see Fig. 1). With the exception of the patient shown in Fig. 1, all CT images appeared to be of acceptable quality for our purposes. Based on published data, the additional radiation exposure from a 30–50 mA CT scan of the thorax or abdomen is in the order of 1.0–2.0 mSv [29]. This is higher than the quoted figure of 0.5 mSv for the GE Hawkeye which acquires images at 2.5 mA [30]. This additional radiation exposure is small compared with the effective dose received from several commonly used radiopharmaceuticals, such as 67Ga (48 mSv per 400 MBq injection), 99mTc-diphosphonate (4.6 mSv per 800 MBq injection), 99mTc-sestamibi for parathyroid imaging (6.8 mSv per 800 MBq injection) and 111In-octreotide (11 mSv per 200 MBq injection) [31,32]. It is also worth noting that we often perform ‘localization’ scans when imaging neuroendocrine tumours as a complement to MIBG imaging to locate the kidneys using, for example, 99m Tc-mercaptoacetyltriglycine (MAG3), which imparts a radiation dose of r 4 mSv for 300 MBq injected activity. Use of a hybrid scanner obviates the need for the use of this additional radiopharmaceutical. For radionuclide therapy, the additional radiation exposure from a low dose CT is small compared to that received from the radiopharmaceutical. However, it should be recognized that if the use of hybrid spiral CT scanning may facilitate the accurate characterization of abnormalities seen on SPECT (and even low-dose SPECT/CT scans), there will be less need for subsequent diagnostic CT scans in many cases and, as a result, the overall radiation exposure to such patients will actually be reduced. Nevertheless, care will be needed with the use of multidetector CT scanners to ensure that any additional radiation exposure is appropriate in each

individual clinical setting. Multidetector CT scans may be associated with high radiation exposures in the order of 9–28 mSv for chest examinations and up to 30 mSv for the abdomen and pelvis [33,34]. This may be even more significant for newer generation scanners which can be associated with higher radiation doses. For instance, effective dose equivalents of up to 15 mSv have been quoted with use of a new 64-slice machine for coronary artery imaging [35]. There are several limitations of our study which should be noted. This is a retrospective series and the impact on patient management and clinical follow-up were not assessed. As with other reports that have addressed the impact of new imaging technology in a variety of clinical settings and indications, the mix of patients at our institution may not be considered indicative of other centres, hence it could be argued that the findings may be considered to be relatively institution-specific. Our series did, however, combine patients undergoing a variety of commonly performed nuclear medicine procedures (e.g., bone scans and gallium scintigraphy) with clinical questions frequently assessed in most major referral hospital settings. The use of SPECT imaging with radiopharmaceuticals such as MIBG and 111 In-octreotide would now appear to be relatively routine, hence inclusion of these study types would appear to be well suited to assess the true impact of hybrid imaging in overall clinical practice today. Gallium scintigraphy for lymphoma is being increasingly replaced by PET/CT [36], but our findings will be of relevance to many centres where access to PET is limited and gallium is still routinely used for staging and follow-up of patients with lymphoma. As hybrid machines with diagnostic quality spiral CT scanners are installed over the next few years, more data will emerge on the use of these hybrid scanners in specific clinical indications and practitioners will then be better able to assess the real impact of the technology in their own individual practice. It should also be noted that the benefit of hybrid imaging that we have demonstrated should be compared with SPECT imaging only. Our series did not assess the impact on all patients having nuclear medicine scans, but rather compares SPECT/CT only with those patients who would have had SPECT as part of their routine imaging. This does not include myocardial perfusion scanning, which is routinely performed with SPECT at our institution, and which was not assessed in this study. In conclusion, this is a preliminary series reporting the impact of hybrid SPECT/CT imaging incorporating spiral CT technology in clinical practice. We found hybrid imaging improved the final diagnosis and reporter confidence in a significant number of patients studied. In those patients who would have previously undergone SPECT-only imaging, hybrid SPECT/CT resulted in a

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Imaging using a spiral CT scanner Roach et al. 987

change in lesion localization in 28% of sites and final reports were different in 56% cases, with a significant difference noted in 26% cases. Prospective studies assessing specific clinical indications and effects on patient management will be required to assess the true impact across various diagnostic settings. A comparison of the clinical utility of hybrid scanners using low performance X-ray scanners with newer multislice machines in the same patients would also be a valuable study to assess the additional benefits of these ‘next generation’ scanners in clinical practice. These future studies should help further define the appropriate place for this new technology in various diagnostic imaging settings.

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Schillaci O. Hybrid SPECT/CT: a new era for SPECT imaging? Eur J Nucl Med Mol Imaging 2005; 32:521–524. Roach PJ, Bailey DL. Combining anatomy and function: the future of medical imaging. Int Med J 2005; 35:577–579. Coleman R, Delbeke D, Guiberteau M, Conti PS, Royal HD, Weinreg JC, et al. Concurrent PET/CT with an integrated imaging system: intersociety dialogue from the Joint Working Group of the American College of Radiology, the Society of Nuclear Medicine, and the Society of Computed Body Tomography and Magnetic Resonance. J Nucl Med 2005; 46: 1225–1239. Czernin J, Schelbert H. PET/CT imaging: facts, opinions, hopes, and questions. J Nucl Med 2004; 45(suppl):1S–3S. Osman MM, Cohade C, Fishman EK, Wahl RL. Clinically significant incidental findings on the unenhanced CT portion of PET/CT studies: Frequency in 250 Patients. J Nucl Med 2005; 46:1352–1355. Bailey DL. Is PET the future of nuclear medicine? Eur J Nucl Med Mol Imaging 2003; 30:1047–1049. Bocher M, Balan A, Krausz Y, Shrem Y, Lonn A, Wilk M, Chisin R. Gamma camera-mounted anatomical X-ray tomography: technology, system characteristics and first images. Eur J Nucl Med 2000; 27:619–627. Palumbo B, Sivolella S, Palumbo I, Liberati AM, Palumbo R. 67Ga-SPECT/ CT with a hybrid system in clinical management of lymphoma. Eur J Nucl Med Mol Imaging 2005; 32:1011–1017. Bar-Shalom R, Yefremov N, Guralnik L, Keider Z, Engel A, Nitecki S, Israel O. SPECT/CT using 67Ga and 111In labelled leukocyte scintigraphy for diagnosis of infection. J Nucl Med 2005; 47:587–594. Horger M, Eschmann SM, Pfannenberg C, Venthein R, Besenfelder H, Claussen CD, et al. Evaluation of combined transmission and emission tomography for classification of skeletal lesions. Am J Roentgenol 2004; 183:655–661. Amthauer H, Denecke T, Rohlfing T, Ruf J, Bohmig M, Gutberlet M, et al. Value of image fusion using single photon emission computed tomography with integrated low dose computed tomography in comparison with a retrospective voxel-based method in neuroendocrine tumours. Eur Radiol 2005; 15:1456–1462. Even-Sapir E, Keidar Z, Sachs J, Engel A, Bettman L, Gaitini D, et al. The new technology of combined transmission and emission tomography in evaluation of endocrine neoplasms. J Nucl Med 2001; 42:998–1004. Gayed IW, Kim EE, Broussard WF, Evans D, Lee J, Broemeling LD, et al. The value of 99mTc-sestamibi SPECT/CT over conventional SPECT in the evaluation of parathyroid adenomas or hyperplasia. J Nucl Med 2005; 46:248–252. Kaczerik K, Prager G, Kienast O, Dobrozensky G, Dudczak R, Niederle B, et al. Combined transmission and Tc-99m sestamibi emission tomography for localisation of mediastinal parathyroid glands. Nuklearmedizin 2003; 42:220–223.

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Original article

Relationship between cancer cell proliferation, tumour angiogenesis and 201Tl uptake in non-small cell lung cancer Seigo Fujitaa, Shigeki Nagamachia, Ryuichi Nishiia, Hideyuki Wakamatsua, Shigemi Futamia, Shozo Tamuraa, Yasunori Matsuzakib, Toshio Onizukab, Kinta Hatakeyamac and Yujiro Asadac Purpose To investigate whether 201Tl uptake is associated with cell proliferation and angiogenesis in non-small-cell lung carcinoma (NSCLC). Methods Eighty-four patients with scheduled NSCLC underwent 201Tl single photon emission computed tomography (SPECT) imaging: 15 min (early scan) and 240 min (delayed scan) after intravenous injection of 111 MBq of 201Tl chloride. 201Tl indices were calculated on early images (early ratio: ER) and delayed images (delayed ratio: DR). The retention index (RI) was also calculated from these two parameters. Using surgically resected cancer specimens (54 adenocarcinoma, 24 squamous cell carcinoma (SCC), six large-cell carcinoma), immunohistochemical stains for both Ki-67 (MIB-1 index) and CD34 were performed to examine the proliferative activity and the micro-vessel density (MVD), respectively. Results The mean value of 201Tl index was 1.69 ± 0.77 (ER) and 2.31 ± 1.08 (DR). The average RI was 42.6 ± 42.9%, respectively. Both DR and RI positively correlated with MIB-1 index (r = 0.68, P < 0.05 and r = 0.52, P < 0.05). When we analyse adenocarcinoma and SCC separately, there was a significant positive correlation (r = 0.62, P < 0.05) between RI and MIB-1 index in adenocarcinoma

Introduction Non-small cell lung cancer (NSCLC) account for 75–80% of all lung cancer, and surgical cure is achieved in only 15–20% of the patients [1]. Although the incidence has been declining modestly in men, it has shown a progressive increase in women, NSCLC is one of the most difficult cancers to cure. A large number of biological prognostic factors for lung cancers have been described [2,3]. Altered regulation of the cell cycle, which is related to tumour proliferative activity, is hallmark of human cancers [4,5]. The expression of Ki-67 antigen can be used as a simple histopathological marker for cell proliferation. A monoclonal antibody against Ki-67 antigen detects a human nuclear antigen that is present only in proliferating cells, but is absent in quiescent cells [6–8]. The relationship between Ki-67 antigen expression and the disease-free survival after surgery of NSCLC has been reported [9].

but not in SCC (r = 0.20, P = NS). The value of ER positively correlated with MVD (r = 0.75, P < 0.05). It demonstrated strong positive correlation with both histological types (adenocarcinoma: r = 0.80, P < 0.05, SCC: r = 0.66, P < 0.05). Conclusion 201Tl SPECT imaging is effective non-invasive method for assessing both the proliferation and the angiogenesis in NSCLC. Both DR and RI are useful indicators for assessing cancer cell proliferation in lung adenocarcinoma. ER is a useful marker for assessing the tumour angiogenesis in NSCLC. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:989–997
Nuclear Medicine Communications 2006, 27:989–997 Keywords: non-small cell lung cancer, proliferation, MVD, angiogenesis

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Tl SPECT, MIB-1 index, cell

a Department of Radiology, bDepartment of Second Surgery and cDepartment of First Pathology, Miyazaki Medical College, Japan.

Correspondence to Dr Seigo Fujita, Department of Radiology, Miyazaki Medical College, 5200 Kihara, Kiyotake, Miyazaki Prefecture 889-1692 Japan. Tel: + 0081 985 85 1510; fax: + 0081 985 85 3392; e-mail: [email protected] Received 15 May 2006 Accepted 29 August 2006

As well as this, tumour angiogenesis is also an important prognostic factor in various cancers including NSCLC [10–16]. Increase of micro-vessel density (MVD) is a predictor of poor survival [10]. The MVD has been measured with an anti-CD34 monoclonal antibody that is specific for endothelial cells and has been used to measure angiogenesis in tumour samples [10]. Therefore it is important to be aware of biological processes, such as proliferation and angiogenesis, when predicting a prognosis or planning therapy. Usually, identification of lung cancer is accomplished on chest radiograph or computed tomography (CT), but these modalities do not reflect biological behaviour. In contrast, radionuclide imaging such as [18F]fluorodeoxyglucose (18F-FDG) positron emission tomography (PET) [17] or 30 -deoxy-30 -[18F]fluorothymidine (18F-FLT) PET [18,19] has been used recently for non-invasive functional

c 2006 Lippincott Williams & Wilkins 0143-3636


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990 Nuclear Medicine Communications 2006, Vol 27 No 12

imaging. It is useful not only for differential diagnosis but also for estimating biological behaviour, including proliferative activity. However, it needs special equipment such as a cyclotron and a PET camera, and the system is still expensive. In a clinical setting the fasting conditions should be set before the examination. The image quality of 18F-FDG PET might deteriorate in a diabetic patient. Moreover, physiological accumulation of the mediastinum might influence the diagnosis. Sometimes, 18F-FDG PET gives false positive results because of its inherent inability to discriminate inflammatory processes from tumour infiltration since high level metabolic changes occur in both instances. Although 18 F-FLT is an appropriate tracer that visualizes cell proliferation, some drawbacks, such as lower sensitivities in tumour detection, have been reported [19]. Conventionally, 201Tl SPECT has also been advocated to distinguish benign from malignant lesions [20–25] and to identify lymph node metastasis [26]. Several studies suggested a correlation between tumour uptake and membrane potential [27–29]. Takekawa et al. [26] suggested a relationship between 201Tl uptake and tumour viability. We previously reported that the 201Tl index was a useful indicator for predicting mediastinal lymph nodes metastasis in NSCLC [30]. In the current study, we investigated whether 201Tl uptake could reflect tumour cell proliferation and tumour angiogenesis, both important indicators of the biological behaviour of NSCLC.

Materials and methods Patients

Eighty-four patients referred for evaluation of a lung mass or nodule from April 2003 through August 2005, who had a diagnosis of lung cancer confirmed by histopathology were enrolled in this study. Two physicians blinded to each patient’s clinical status and to results of other imaging studies interpreted each imaging study. Staging diagnosis was based on the TNM classification. There were 59 men and 25 women, ranging in age from 40 to 83 years (mean: 65.8 ± 9.7 years). Histopathological diagnoses of the 84 primary lesions included adenocarcinoma (n = 54), squamous cell carcinoma (SCC, n = 24) and large cell carcinoma (n = 6). The post-operative TNM staging of patients was as follows: stage I (A 20, B 20), II (A 6, B 9), III (A 16, B 9) and IV 4. All patients gave their informed consent prior to participation. Approval of Miyazaki Medical College ethics committee was obtained before the current study. Imaging protocol

Chest CT scans were performed with X-Vigor (Toshiba, Tokyo, Japan), Aquilion (Toshiba, Tokyo, Japan) and LightSPEED (General Electric Medical Systems, USA)

systems. Images were obtained with 10 mm collimation, at 10 mm intervals, in the supine position, from the lung apices to the adrenal glands. The scanning parameters (120 kVp, 170–200 mA, a 1 s scanning time, 30–35 cm field of view with a 512  512 matrix) varied slightly, depending on the patient’s size. Images were printed with settings for mediastinum and lung display. An intravenous bolus injection of 100 ml of the non-ionic contrast medium Iopamidol (300 mg I  ml – 1; Bracco Industria Chemica, Milan, Italy) was used. 201

Tl SPECT images were acquired at both 15 min and 240 min after radiotracer (111 MBq) injection. Images were acquired with a large field of view, dual-detector camera and computer system (E-CAM; Siemens, USA, and Prism 2000; Picker, USA), equipped with a lowenergy, high-resolution, parallel-hole collimator. The dual-detector camera was rotated over 1801 in a circular orbit. Images were acquired in 64 steps over 3601 of rotation at the rate of 30 s/view. Data were collected digitally (64  64 matrix for 45 s per view in a step-andshoot mode) in a dedicated computer system (E-soft; Siemens, USA, and Odyssey; Picker, USA). A 20% window centred on the 68 keV mercury X-ray of 201Tl provided energy discrimination. Continuous transaxial tomograms were reconstructed after filtered back-projection with a Butterworth filter to reduce statistical noise (cut-off frequency: 0.22 cycle/pixel, order 8). Coronal and sagittal tomograms of 1 pixel thickness were reconstructed from the trans-axial images. 201Tl SPECT study was done within 1 week after chest CT imaging in all patients. The reconstructed SPECT images were displayed under the condition with lower cut-off level at 30% of the maximal count [24,25,30]. Each 201Tl SPECT image was analysed visually with respect to chest CT or fusion image with chest CT by computer system (E-soft; Siemens, USA). Along the contour of a tumour maximal diameter (T) and on the contralateral side over the normal lung area (N), regions of interest (ROIs) were drawn in the transverse image. Using the mean per pixel obtained from these ROIs, early (ER) and delayed (DR) T/N ratios and retention indices [RI = (DR – ER)  100/ER] were calculated.

Immunohistochemical study

Specimens surgically excised were fixed in 10% formalin and embedded in paraffin. Sections (4 mm thick) were stained with haematoxylin–eosin (H&E) for morphological studies, and serial sections were examined by immunohistochemistry using the avidin–biotin complex immunoperoxidase method as described previously [31]. Briefly, the sections were deparaffinized, and incubated in 3% H2O2 in methanol for 30 min to block endogenous peroxidase activity. For retrieval of Ki-67 antigen, the sections were heated for 10 min in 10 mmol  l – 1 citrate buffer solution (pH 6.0) in a microwave oven. The sections were then incubated with primary monoclonal antibodies against Ki-67 (clone MIB-1, diluted in

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Cell proliferation, tumour angiogenesis and

1:50; DakoCytomation, Kyoto, Japan) and CD-34 (clone QBEnd 10, diluted in 1:50; DakoCytomation) at 41C overnight. Intervening washes with phosphate-buffered saline were followed by incubation with biotinylated secondary antibodies for 30 min. After further washes, peroxidase-conjugated streptavidin (Strept ABComplex, DakoCytomation) was applied. Visualization was carried out for 2 min with 0.05% 3,30 -diaminobenzidine containing hydrogen peroxide. Finally, the nuclei were counterstained with haematoxylin. As a negative control, normal mouse IgG was used instead of the primary antibodies. Evaluation of the specimens

All slides were evaluated without any knowledge of clinico-pathologic data. The percentage of Ki-67-positive cells was calculated by counting more than 1000 tumour cells in random high-power fields (10  40), and recorded as the MIB-1 index. Evaluation of MVD was also assessed by counting positively stained tissues. Briefly, any CD34positive blood vessels with lumina as well as cell clusters without a lumen and single cells that were clearly separate from adjacent tumour cells and other connective tissue elements were considered to be countable microvessels. Six areas containing high vascular components were chosen, and counts of micro-vessels were observed with a high-power field (10  40). MVD was defined as the total number of micro-vessels in six high-power fields per specimen. The histology of each specimen was classified according to the WHO criteria [32] and the pathological stage of each tumour was recorded using the TNM staging system [33]. Statistical analysis

All variables are expressed as the mean ± standard deviation (SD). The Student t-test and one-way ANOVA (with Bonferonni correction when appropriate) were applied to evaluate differences for continuous parameters. Correlations among 201Tl indices (ER, DR, RI) and immunohistochemical data (MIB-1 index, MVD) were analysed using the Pearson correlation test. Data analyses were performed using StatView ver.5.0 for windows statistical application program. In all assessments, P < 0.05 was considered significant.

Results 201

Tl SPECT images demonstrated uptake in the primary lesions in all patients. On the chest CT scan, these primary lesions were enhanced with iodinated contrast medium. The average of 201Tl index of all tumours’ ER, DR and RI was 1.69 ± 0.77, 2.31 ± 1.08, 42.6 ± 42.9%, respectively. In patients with adenocarcinoma, the mean 201 Tl index was 1.58 ± 0.74 (ER), 2.13 ± 1.01 (DR) and 42.5 ± 46.2% (RI). In patients with SCC, the mean 201Tl index was 1.85 ± 0.74 (ER), 2.48 ± 1.12 (DR) and 36.6 ± 39.4% (RI), respectively. Every 201Tl index showed no significant difference between adenocarcinoma

201

Tl uptake in NSCLC Fujita et al. 991

and SCC (Table 1). There was a positive correlation between DR (r = 0.68, P < 0.05), RI (r = 0.52, P < 0.05) and MIB-1 index. In contrast, there was no significant correlation between ER and MIB-1 index. (Fig. 1). When we analyse each histopathology type separately, there was a significant positive correlation (r = 0.62, P < 0.05) between RI and MIB-1 index in adenocarcinoma but not in SCC (r = 0.20, P = NS) (Fig. 2). Within adenocarcinoma, the MIB-1 index of well differentiated adenocarcinoma was relatively lower than those of moderately group or poor differentiated adenocarcinoma, but there was no statistical significance. Both DR and RI were significantly higher in poor differentiated adenocarcinoma than those of well differentiated adenocarcinoma (Table 2). Within each 201Tl index, only RI demonstrated positive strong correlation with MIB-1 index in every differentiated-type adenocarcinoma. Although DR correlated positively with MIB-1 index in both poorly differentiated adenocarcinoma and moderately differentiated adenocarcinoma, there was a weak correlation in well differentiated adenocarcinoma (Table 3). Regarding MVD, the value of poorly differentiated adenocarcinoma was relatively higher than those of moderately differentiated group or well differentiated type, but there was no statistical significance. There was a strong positive correlation (r = 0.75, P < 0.05) between ER and MVD. However, neither DR nor RI correlated with MVD (Fig. 3). When we analyse it according to the histological type separately, there was a strong correlation (r = 0.80, P < 0.05) between ER and MVD in adenocarcinoma (Fig. 4a). Although the correlation coefficient was smaller than that of adenocarcinoma, it also showed strong correlation (r = 0.66, P < 0.05) between ER and MVD in SCC (Fig. 4b). When we analysed it according to histological differentiation degree separately within adenocarcinoma, the correlation coefficient between 201 Tl index and MVD in the poorly differentiated type was higher than that of the moderately differentiated type or the well differentiated group (Table 4). Figure 5 shows a representative case of a pulmonary nodule with intense 201Tl uptake: the ER was 1.64, DR was 3.44, and RI was 110% (Fig. 5a,b). Histopathological evaluation of a surgical specimen confirmed a poorly differentiated adenocarcinoma. The MIB-1 index for this patient was 58.9% (Fig. 5c) and the MVD was 179 (Fig. 5d). 201 Table 1 Tl indices of patients with non-small cell lung carcinoma (NSCLC)

Tumour NSCLC (n = 84) Adenocarcinoma (n = 54) Squamous cell carcinoma (n = 24)

Early ratio

Delayed ratio

Retention index (%)

1.69 ± 0.77 1.58 ± 0.74 1.85 ± 0.74

2.31 ± 1.08 2.13 ± 1.01 2.48 ± 1.12

42.6 ± 42.9 42.5 ± 46.2 36.6 ± 39.4

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Nuclear Medicine Communications 2006, Vol 27 No 12

70

70

60

60

60

50 40 30

MIB-1 index (%)

70

MIB-1 index (%)

MIB-1 index (%)

Fig. 1

50 40 30

20

20

10

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y =7.8x + 23 r= 0.68, P< 0.05

0 0

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2 ER

3

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40 30

y = 0.14x + 38 r= 0.52, P < 0.05

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r = 0.14, P=NS 0

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0

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3 DR

4

10 0

5

–50

0

50 100 150 RI (%)

Correlation between 201Tl indexes and MIB-1 index. Both the delayed ratio (DR) and retention index (RI) significantly correlate with MIB-1 index. There is no significant correlation between the early ratio (ER) and MIB-1 index.

70

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50

MIB-1 index (%)

MIB-1 index (%)

Fig. 2

40 30 20

40 30 20

y = 0.15x + 32

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r= 0.20, P= NS

r = 0.62, P< 0.05 0

0 – 50

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150

0

50 100 RI (%)

150

There is a significant correlation (r = 0.62, P < 0.05) between retention index (RI) and MIB-1 index in adenocarcinoma (left). There is not a significant correlation (r = 0.20, P = NS) between RI and MIB-1 index in SCC (right).

Table 2

Comparison of MIB-1 index and the MIB-1 index (%)

Well n = 18 Mod n = 25 Poor n = 9

32.3 ± 18.3 44.8 ± 14.0 45.0 ± 21.4

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Tl indices for adenocarcinoma 201

Micro-vessel density

108 ± 45.8 125 ± 99.8 144 ± 81.7

Tl index (%)

Early ratio

Delayed ratio

Retention index

1.42 ± 0.47 1.63 ± 0.72 1.80 ± 1.17

1.85 ± 0.56 2.13 ± 0.95 2.72 ± 1.61

35.7 ± 33.9 40.7 ± 53.8 61.4 ± 44.0

Well: well-differentiated carcinoma; mod: moderately differentiated carcinoma; poor: poorly differentiated carcinoma.

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Cell proliferation, tumour angiogenesis and

Discussion 201

Following the reports that Tl accumulates in lung cancer [22,23], various studies have dealt with the differentiation of malignant tumour from benign lesions [24,25], the usefulness for the staging [34] and for determining the therapeutic effect [35], and predicting lymph node metastasis [30]. The current study demonstrated that 201Tl uptake of NSCLC is associated with the cell proliferation and tumour angiogenesis. There are various factors influencing 201Tl uptake in malignant tumours: Na–K-ATPase activity, tumour perfusion, tumour cell viability, permeability of cell membrane, the co-transport system, and the calcium ion channel system [20–23]. These mechanisms are supported by an experiment showing that Na–K-ATPase is associated with active transport of 201Tl into a tumour cell [20–23]. Kier [36] reported that Na–K-ATPase activities enhanced in plasma membranes of high metabolic cells such as cancer cells. They demonstrated that increased Na–K-ATPase activity in metastasis cells was associated with cell-surface fluidity and metastatic potential. In addition, Na–K-ATPase activity is known to correlate with the growth rate of normal cells or mutated cells [37]. The

Table 3 Comparison of correlation coefficient between indices and MIB-1 index for adenocarcinoma

Well n = 18 Mod n = 25 Poor n = 9

201

Tl

Early ratio

Delayed ratio

Retention index

0.282 0.152 0.277

0.359 0.553 0.561

0.579 0.586 0.581

Abbreviations as in the footnote to Table 2.

201

Tl uptake in NSCLC Fujita et al. 993

association between 201Tl uptake and tumour cell proliferation has also been reported [38]. Previous studies have reported that cell proliferation is an important prognostic factor in different types of cancer, including NSCLC [10–16]. We selected the MIB-1 antibody to investigate relationship between Ki-67 expression and prognosis or postoperative recurrence in NSCLC [9,39–43]. Immunocytochemical investigation by means of flow cytometry has demonstrated that the antigen recognized by the MIB-1 antibody is closely associated with the cell nuclear Ki-67 antigen [44]. Ki-67 antigen expression starts in the S phase, and progressively increases through the S and G2 phases, reaching its maximum at mitosis. After division, the cells return to the G1 phase with a stock of Ki-67 antigen. The Ki-67 level decreases progressively during this phase and finally disappears in cells with a long G2 phase [45–47]. With this background, the MIB-1 index has been reported to be an effective marker of proliferative activity in various malignant tumours, including NSCLC [9,39–43,48]. The current study demonstrated that both DR and RI correlated positively with the MIB-1 index, which indicated that 201Tl uptake reflects proliferative activity in part. On the other hand, Ishibashi et al. have reported that 201Tl imaging is useful for estimating cancer cell proliferation in SCLC [38]. However, they did not find significant correlation between the cell proliferation and 201 Tl uptake in NSCLC. Although the exact reason is unknown, they did not involve histological type, which may influence the discrepancy. In the current study, the correlation between MIB-1 and RI was good in adenocarcinoma in comparison with SCC, which often contains

400

400

300

300

300

200

100

MVD

400

MVD

MVD

Fig. 3

200

200

100

100

y = 89x − 9 r = 0.75, P < 0.05 0 0

1

2 ER

3

4

0

1

2

3 DR

4

5

-50

0

50 100 150 RI (%)

There is a strong correlation (r = 0.75, P < 0.05) between ER and micro-vessel disease (MVD). There is no correlation between DR, RI and MVD. (Abbreviations as in the legend to Fig. 1).

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994 Nuclear Medicine Communications 2006, Vol 27 No 12

central necrosis that causes cavity formation. Such a phenomenon leads to heterogeneous uptake of the tracer, which results in poor correlation between 201Tl uptake and MIB-1 index of SCC. As other markers that denote SCC activity, such as chromosomal aberrations [49], are known, future studies should involve those indices.

Fig. 4

(a) 400

Although we investigated the relationship between the degree of cell histological differentiation and MIB-1 index, we did not find a close association. Cancer proliferation activity in NSCLC, therefore, cannot be presumed by the histological differentiation. As RI correlated positively with MIB-1 of adenocarcinoma in every differentiated type, it is likely that this is the most effective index for assessing cell proliferation activity. Angiogenesis, namely, neovascularization from pre-existing vasculature, is also an important prognostic indicator in different types of cancer, including NSCLC [10–16,50,51].

MVD

300

200

100 y = 96x − 13 r = 0.80, P < 0.05 0

1

2

4

3 ER

(b) 300

MVD

200

100

y = 62x + 37 r = 0.66, P < 0.05 0

1

2 ER

3

4

There is a strong positive correlation (r = 0.80, P < 0.05) between ER and MVD in adenocarcinoma (a). There is also a positive correlation (r = 0.66, P < 0.05) between ER and MVD in SCC (b). (Abbreviations as in the legends to Figs 1 and 3).

Table 4 Comparison of correlation coefficient between indices and micro-vessel disease for adenocarcinoma

Well n = 18 Mod n = 25 Poor n = 9

201

Tl

Early ratio

Delayed ratio

Retention index

0.829 0.785 0.961

0.533 0.254 0.797

0.281 0.394 0.443

Abbreviations as in the footnote to Table 2.

Angiogenesis consists of sprouting and non-sprouting (the enlargement, splitting and fusion of pre-existing vessels) processes and both can occur concurrently [50]. The growth of NSCLC is usually dependent on angiogenesis, which is regulated by complex mechanisms in the presence of various angiogenesis-related molecules [50]. Conventionally, the MVD has been measured with an anti-CD34 monoclonal antibody that is specific for endothelial cells and has been used to measure angiogenesis in tumour samples. Increase of MVD is a useful predictor of poor survival [15]. The current study demonstrated strong correlation between ER and MVD. Traditionally, 201Tl uptake during the early phase is said to reflect tumour cell perfusion closely related with vascularity. However, accounting for the imaging time after tracer injection and the clearance time of 201Tl from the blood, ER did not seem to reflect merely cell perfusion. The current study confirmed that ER expressed the angiogenesis that will be required for tumour growth and proliferation. Indeed, poorly differentiated adenocarcinoma showed prominent strong correlation between MVD with ER, both well differentiated group and moderately differentiated group also demonstrated showed positive correlation between MVD with ER. The value of ER is, therefore, effective marker for estimating cancer cell angiogenesis in adenocarcinoma. Non-invasive assessment of cell proliferation and angiogenesis in cancer treatment is very important in both for predicting prognosis and for determining the therapy protocol [52]. In planning radiotherapy, the delineation of tumour contour and planning target volume (PTV) are usually performed on transverse CT. Recently the PTV is usually created in the three-dimensional manner which is irregular and often makes it more difficult to achieve a highly conformal three-dimensional plan such as intensity-modulated radiotherapy (IMRT) [53]. Therefore, the effective planning of IMRT is an important issue

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Cell proliferation, tumour angiogenesis and

201

Tl uptake in NSCLC Fujita et al. 995

Fig. 5

A 71-year-old female patient with poorly differentiated lung adenocarcinoma. Co-registration with CT and 201Tl SPECT image showed intense pulmonary nodule uptake in left lower lung field (a: early image, b: delayed image). 201Tl indexes are ER = 1.64, DR = 3.44 and RI = 110%. Immunohistochemical staining with Ki-67 showed strong positive staining. The MIB-1 index was 58.9% (c). Immunohistochemical staining with CD34 also showed strong positive staining in vessels. The value of MVD was 179 (d). (Abbreviations as in the legends to Figs 1 and 3).

in management of NSCLC [54]. Grosu et al. reported that new functional imaging methods such as imaging of hypoxia, cell proliferation, angiogenesis, apoptosis and gene expression lead to the identification of different areas of a biologically heterogeneous tumour mass that can be targeted individually using IMRT [55]. In addition, a biological dose distribution can be generated, the so-called dose painting [50]. As 201Tl SPECT can provide useful information of biological characteristics, in part, we can select cancer therapy appropriately taking into account the degree of 201Tl uptake.

Diagnosis by SPECT alone has limitations in evaluating tumours adjacent to the structures with intense physio-

logical uptake such as the heart or diaphragm. Co-registration of 201Tl SPECT images with CT scans using software is one of the alternative methods for resolving the problem [56,57]. They can be performed by a computer algorithm to combine the images in one display using anatomical landmarks. However, in clinical practice, it is time consuming and somewhat difficult to accomplish due to respiratory motion. Recently, integrated SPECT/CT with simultaneous, co-registered anatomical and functional imaging has been introduced and used for clinical purposes [58]. It should be more accurate than SPECT alone in the anatomical localization of lung cancer [59]. Furthermore, it will improve quantification by utilizing the absorption correction. Such a new device may be very useful in future studies.

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996 Nuclear Medicine Communications 2006, Vol 27 No 12

In conclusion, the 201Tl index, both DR and RI, are useful indicators for assessing cancer cell proliferation in lung adenocarcinoma. Similarly, ER is a useful marker for assessing tumour angiogenesis in NSCLC. Because both cell proliferation and angiogenesis are important biological indicators of NSCLC both for determining therapies [30] and assessing the prognosis, 201Tl indices will be useful for planning and monitoring therapy.

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Original article

Preparation of 99mTc-Nanocoll for use in sentinel node localization: Validation of a protocol for supplying in unit-dose syringes Lesley M. O’Briena, Rodger Duffinb and Alistair M. Millara Objectives To determine if 99mTc-Nanocoll is affected by storage in a syringe, passage through an R-Lock or mixing with Patent Blue V dye. Methods A Nanocoll kit was reconstituted at 280 MBq/ 5 ml. Samples of 0.5 ml were drawn into 3 ml Plastipak syringes. After 1 h and 6 h, adsorption of 99mTc on a syringe was measured and the following tests of quality were performed on samples from the kit and a syringe: 99m Tc-pertechnetate impurity by thin-layer chromatography, the percentage of 99mTc on particles > 100 nm by Nuclepore filtration and particle size by photon correlation spectroscopy. These tests were also applied to samples that had been passed through an R-Lock or mixed with Patent Blue V. Each experiment was performed five times.

difference (P > 0.05, n = 5) between 99mTc-Nanocoll from the original kit and a syringe at either 1 h or 6 h. Similar results were obtained with samples that had been passed through an R-Lock or mixed with Patent Blue V. Conclusions A capped 3 ml Plastipak syringe is a suitable container in which to supply 99mTc-Nanocoll. Neither passage through an R-Lock nor mixing with Patent Blue V affects the quality of 99mTc-Nanocoll. Nucl Med Commun c 2006 Lippincott Williams & Wilkins. 27:999–1003 Nuclear Medicine Communications 2006, 27:999–1003 Keywords:

99m

Tc-Nanocoll, stability, sentinel node

a Radiopharmacy, The Royal Infirmary of Edinburgh and bELEGI Colt Laboratory, The Queen’s Medical Research Institute, University of Edinburgh, UK.

Results In all samples, adsorption of 99mTc on syringes was < 1%, 99mTc-pertechnetate impurity was < 2%, > 95% of 99mTc was labelled to particles < 100 nm in diameter, the mean particle diameter was 6.6 nm and the particles had a diameter of < 12 nm. All tests showed no significant

Correspondence to: Dr A.M. Millar, Department of Pharmacy, The Royal Infirmary of Edinburgh, Little France Crescent, Edinburgh, EH16 4SA, UK. Tel: + 0044 (0) 131 242 2929; fax: + 0044 (0) 131 242 2931; e-mail: [email protected]

Introduction

appropriate than our standard practice of supplying radiopharmaceuticals in unit dose vials. Supplying 99m Tc-Nanocoll in a syringe should minimize (1) the handling of radioactive material in the theatre, (2) the exposure of theatre staff to radiation, (3) the potential for radioactive contamination, and (4) the disposal of radioactive waste. There are many reports of the quality of 99m Tc radiopharmaceuticals being compromised by their containers [1–9]. One of the aims of this work was therefore to determine if a capped syringe is a satisfactory container in which to supply 99mTc-Nanocoll.

Technetium-99m Nanocoll Injection (99mTc-Nanocoll) is a radiopharmaceutical that is used in sentinel node localization. This is an increasingly popular nuclear medicine technique with its value in the management of melanoma and breast cancer being widely reported. 99m Tc-Nanocoll is a radiolabelled colloid of human albumin with a mean particle size that is measured in the nanometre size range. It is prepared using a commercially available freeze-dried kit. We were asked to supply 99mTc-Nanocoll directly to an operating theatre in which breast surgery is performed. This theatre is in a hospital several miles from the radiopharmacy. The proposal is to administer 99m Tc-Nanocoll (20 MBq/0.5 ml) by subareolar injection immediately before the start of surgery and perform intraoperative sentinel node localization with a gamma probe. Operating theatre personnel have less experience in the handling of radioactive material than the typical client of the radiopharmacy. For this reason, we decided that supplying 99mTc-Nanocoll in a syringe would be more

Received 14 March 2006 Accepted 31 August 2006

The protocol for administration is to remove the cap from the syringe containing the 99mTc-Nanocoll, fit a nonreturn valve (R-Lock) and hypodermic needle, inject the contents of the syringe sub-areolarly, remove the syringe from the R-Lock leaving the needle in situ, fit a syringe containing blue dye (Patent Blue V) and inject the dye through the R-Lock. The other aims of this work were therefore to determine if the quality of 99mTc-Nanocoll is affected by passage through an R-Lock or mixing with Patent Blue V.

c 2006 Lippincott Williams & Wilkins 0143-3636

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1000 Nuclear Medicine Communications 2006, Vol 27 No 12

Materials and methods Preparation of

99m

Tc-Nanocoll

Sodium Chloride Injection (0.9%) BP (Braun, Melsungen, Germany) then Sodium Pertechnetate [99mTc] Injection BP from a Drytec generator (GE Healthcare, Amersham, UK) were injected into a Nanocoll kit (GE Healthcare) to give a radioactive concentration of 280 MBq/5 ml. The vial was inverted a few times then incubated at room temperature for 10 min. The vial was shaken gently, a vent needle was inserted through the cap and a 0.5 ml aliquot of the solution was drawn into a 3 ml luer-lock Plastipak syringe (Becton Dickinson, Franklin Lakes, NJ, USA). The needle was replaced with a sterile cap (Angiokard Medizintechnik, Friedeburg, Germany) and the syringe was stored at room temperature. Measurement of

99m

Tc adsorption on the syringe

99m

The degree of Tc adsorption was determined using a modification of a method used previously to measure the adsorption of 99mTc on glass vials [2]. An empty 3 ml syringe and cap were weighed. A needle was fitted and a 0.5 ml sample of 99mTc-Nanocoll was drawn into the syringe. The needle was discarded and the cap was fitted. After storage for the test period, the capped syringe was weighed and the activity of 99mTc was measured in a radionuclide calibrator. The contents were expelled from the syringe. The emptied capped syringe was weighed and its activity was measured. The percentage of 99mTc adsorbed on the syringe was calculated from the difference between the megabecquerels per gram of the solution in the syringe and the megabecquerels per gram of the expelled solution. Measurement of unbound

99m

Tc

99m

The level of unbound Tc was determined by thin-layer chromatography over a distance of 10 cm on a 2.5  20 cm ITLC-SG plate (Pall, Ann Arbor, MI, USA) with a mobile phase of methanol and water (85:15) [10]. The distribution of 99mTc on the plate was measured with a scintillation detector on a Mini-Scan radiochromatogram scanner (Bioscan, Washington, USA). Chromatograms were recorded and analysed with LauraLite 3 radiochromatography software (LabLogic, Sheffield, UK). The percentage of unbound 99mTc-pertechnetate was calculated by expressing the activity at the solvent front as a percentage of the total activity on the plate. Measurement of

99m

Tc on particles > 100 nm

This was performed by the method of Davis et al. [11]. A 100 nm pore size Nuclepore track-etched polycarbonate membrane (Whatman, Brentford, UK) was assembled in a 13 mm diameter Swinnex filter holder (Millipore, Billerica, MA, USA). A 0.1 ml sample of 99mTc-Nanocoll was introduced into the filter holder. A syringe containing 10 ml sodium chloride injection was fitted to the holder and the count rate from the sample was measured by placing it 30 cm from a sodium iodide scintillation

detector. The solution in the syringe was passed through the filter. Once all the liquid had passed through and the bubble point was reached, radioactive liquid was present in the void between the underside of the membrane and the outlet of the holder. To ensure that the detected radioactivity was solely from 99mTc trapped on the filter, the membrane was removed to a fresh holder and the count rate from it was measured. The 99mTc detected on the filter represented 99mTc labelled to particles > 100 nm in diameter. This was quantified by expressing the count rate after filtration as a percentage of the count rate before filtration. Measurement of particle size

The mean particle diameter and the percentage of particles less than 12 nm in diameter were measured by photon correlation spectroscopy using a PS90 Particle Size Analyzer (Brookhaven Instrument Corporation, New York, USA). This instrument provides a measuring size range of 2 nm to 3 mm. Samples were prepared by reconstituting Nanocoll kits as described above but using 0.5 ml of a decayed 99mTc generator eluate. This was necessary as the analyser is located in a laboratory that is not licensed for the handling of radioactive material. The above tests were performed on samples that had been stored in the original kit vial and 3 ml syringes for 1 and 6 h. To determine if the quality of 99mTc-Nanocoll is affected by passage through the non-return valve used during injection into the patient, a sample was prepared as described above and passed though an R-Lock (Codan, Lensahn, Germany). Adsorption in the R-Lock was measured as described above. Unbound 99mTc, 99mTc on particles > 100 nm, mean particle diameter and the percentage of particles less than 12 nm in diameter were measured on the 99mTc-Nanocoll that passed through the R-Lock. To detect any interaction between 99mTc-Nanocoll and the dye that is injected after it, a 500 ml sample was mixed with 50 ml Patent Blue V Sodium 2.5% (Guerbet, Roissy, France). Unbound 99mTc and 99mTc on particles > 100 nm were measured in the mixture. Particle sizing by photon correlation spectroscopy was not possible due to attenuation of the laser light by the blue dye. Each experiment was performed five times. Each set of measurements was made on a different lot of Nanocoll. Where appropriate, statistical analysis was performed using the paired t-test. A P-value > 0.05 was taken to demonstrate no significant difference.

Results The results of the measurements comparing 99mTcNanocoll in the original kit vial and syringes are shown

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Stability of

in Table 1. Adsorption of 99mTc on the syringes was less than 1% and statistical analysis showed no significant difference between the 1 h and 6 h samples. The amount of unbound 99mTc, as determined by thin-layer chromatography, was well within the typical pharmacopoeial limit of 5% specified for other 99mTc radiopharmaceuticals. The percentage of 99mTc that passed through a 100 nm filter was consistently greater than 95% indicating that most of the 99mTc was labelled to particles less than 100 nm in diameter. The results from the photon correlation spectroscopy measurements showed no change in particle size distribution in either original kit vials or syringes over the 6 h period of the study. From the 25 measurements of particle size, the mean particle diameter was 6.6 ± 1.4 nm. The typical size distribution is shown in Fig. 1. For each quality measure that was performed to compare samples stored in vials and syringes, statistical analysis showed no significant difference between the results after storage for both 1 h and 6 h. The results of the measurements performed to detect any deleterious effects caused by passing 99mTc-Nanocoll through an R-Lock or mixing it with Patent Blue V are shown in Table 2. Neither procedure was found to affect the quality of 99mTc-Nanocoll.

Kit reconstitution was performed at the maximum volume of 5 ml to provide the highest possible specific activity while remaining within the manufacturer’s recommendations. High specific activity was recommended by Gommans et al. [13] who showed that nodal uptake is increased as the number of injected particles is decreased. The value of this precaution is uncertain as Veronesi et al. [14] have shown the opposite effect. A possible reason for this discrepancy is the site of injection. Gommans et al. used a combination of intradermal and subdermal/parenchymal areolar injections while Veronesi used an intratumoural injection. As our injection technique was to be similar to that of Gommans et al., the high specific activity preparation seemed the more appropriate. In addition to the possible benefit of administering the minimum number of particles, reconstitution at the maximum volume offers a financial advantage in that it provides up to ten patient doses from one Nanocoll kit.

Fig. 1

60

Results of quality measurements performed on

Quality measure

Tc adsorbed on syringe (%) Unbound 99mTc (%) 99m Tc on particles > 100 nm (%) Mean particle diameter (nm) Particles < 12 nm in diameter (%)

99m

Percentage of total number of particles

The effect of containers on the quality of 99mTc radiopharmaceuticals has been the subject of several reports. Adsorption onto vials and syringes [1–7] and reductions in radiochemical purity caused by containers [8,9] are recognized problems. Agitation is also known to affect the particle size of a particular radiocolloid [12]. Against this background of potential problems, we considered it prudent to investigate the stability of 99m Tc-Nanocoll in syringes before starting to supply it in this way. To achieve this, various measures of quality were used to compare 99mTc-Nanocoll samples taken from a reconstituted kit against samples that had been withdrawn from the kit and stored in syringes. The kit was reconstituted and stored according to the manufacturer’s instructions. The results from the samples taken from the kit were therefore assumed to reflect the highest quality that can be achieved. GE Healthcare specifies that 99m Tc-Nanocoll should be administered within 6 h of

99m

Tc-Nanocoll in unit-dose syringes O’Brien et al. 1001

preparation. Samples were therefore tested at 1 h and 6 h after reconstitution of the Nanocoll kit.

Discussion

Table 1

99m

40

20

0 0

15

10

5

Particle diameter (nm) Particle size distribution of Nanocoll.

Tc-Nanocoll stored in original kit vials and syringes

1 h after preparation

6 h after preparation

Kit

Syringe

Kit

Syringe

– 1.7 ± 0.3 1.9 ± 0.5 6.0 ± 1.8 100

0.6 ± 0.6 1.7 ± 0.2 1.8 ± 0.9 6.5 ± 1.3 100

– 0.2 ± 0.1 2.0 ± 1.0 7.4 ± 1.1 100

0.5 ± 0.2 0.3 ± 0.1 2.0 ± 1.4 6.0 ± 1.3 100

Each value in the mean ± standard deviation of five results.

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1002 Nuclear Medicine Communications 2006, Vol 27 No 12

Table 2 Results of measurements to determine the effect of an R-lock and Patent Blue V on the quality of 99mTc-Nanocoll Quality measure 99m

Tc adsorbed on syringe and R-lock (%) Unbound 99mTc (%) 99m Tc on particles > 100 nm (%) Mean particle diameter (nm) Particles < 12 nm in diameter (%)

After passage through R-lock

Mixed with Patent Blue V

0.6 ± 0.5

–

0.6 ± 0.3 2.3 ± 0.5 7.1 ± 1.4 99 ± 1

0.4 ± 0.2 2.0 ± 0.8 – –

Each value is the mean ± standard deviation of five results.

No adsorption on the Plastipak syringe was detected up to 6 h after preparation. This might not be true for all brands of syringe. Unbound 99mTc-pertechnetate was measured using the TLC system specified by GE Healthcare [10]. This showed no difference between the kit and syringe samples. It is interesting to note that labelling improved with time. When measuring the size of a particulate radiopharmaceutical, the important measure is the size of the particles to which the radionuclide is labelled. A purely physical measurement of size may be misleading as a small number of large particles might carry a disproportionate amount of the radionuclide. Measuring radionuclide retention on a filter membrane with pores of uniform diameter is therefore the method of choice. Unfortunately, filters with a sufficiently small pore diameter to assess colloids in the nanometre size range are not available. A pore diameter of 100 nm is the smallest that is readily available and through which samples can be passed using pressure from a syringe. This is reflected by the manufacturer’s data on 99mTc-Nanocoll which specifies that ‘at least 95% of the human albumin colloidal particles has a diameter of < 80 nm’. It is likely that this specification is related to what can be measured rather than the true size of the colloid. Our filtration measurements show that the samples comply with the manufacturer’s specification. If the particle size is in the range 0 to 20 nm, our filtration measurements do not exclude the presence of aggregates that are less than 100 nm in diameter.

change in particle size throughout the course of our experiments. Administration through an intravenous cannula is known to affect the quality of certain 99mTc radiopharmaceuticals [15–17]. The technique of using a non-return R-Lock to simplify the injection of dye after the 99m Tc-Nanocoll therefore had the potential to affect the quality of the radiopharmaceutical by either adsorption of 99mTc onto some component of the R-Lock or destabilization of the colloid. Fortunately, the R-Lock was not found to affect 99mTc-Nanocoll. Also, using Patent Blue V injection to expel the residual 99mTcNanocoll from the R-Lock results in mixing of the two solutions and the consequent potential for interaction. This was therefore investigated and no interaction was detected. Particle sizing by photon correlation spectroscopy was not feasible due to the attenuation of the laser light by the blue dye. The possibility of a change in particle size cannot therefore be excluded. However, the filtration measurements demonstrate that the product complies with the manufacturer’s specification. On the thin-layer plate, the blue dye migrates at the solvent front while the 99mTc remains at the origin. This demonstrates that Tc is not complexed by Patent Blue V. In conclusion, a capped 3 ml Plastipak syringe is a suitable container in which to supply 99mTc-Nanocoll and neither passage through an R-Lock nor mixing with Patent Blue V affects the quality of 99mTc-Nanocoll.

References 1 2 3 4

5 6

7

The measurements using photon correlation spectroscopy show that there is no change in particle size over the 6 h of the study. Although this does not provide any information about the size of the particles to which the 99m Tc is labelled, taken with the filtration data, it suggests that particle aggregation does not occur. Our value of approximately 7 nm for the mean particle diameter does not agree with the 12 nm reported by Gommans et al. [13]. The only explanation that we can offer for this discrepancy is that the measurements were made using different instruments. In the context of our work, the important finding is that we did not detect any

8 9

10 11 12 13

Porter WC, Dworkin HJ, Gutkowski RF. Vial retention of technetium-99m sulphur colloid. Am J Hosp Pharm 1975; 32:1141–1143. Millar AM. The adsorption of 99Tcm-dimercaptosuccinic acid onto injection vials. Nucl Med Commun 1984; 5:195–199. Millar AM, Stewart E. The adsorption of 99Tcm radiopharmaceuticals onto injection vials – letter to the editor. Nucl Med Commun 1985; 6:115–116. Elliott AT, Murray T, Hilditch TE, Whateley TL. Investigation of factors affecting adhesion of 99Tcm labelled colloids to glass vials. Nucl Med Commun 1990; 11:375–381. Graham D, Millar A. Factors affecting the sorption of 99mTc-tetrofosmin onto glass vials. Nucl Med Commun 1997; 18:335. Jansson BA, Goransson MB, Agren BN. Adsorption of some technetium99m radiopharmaceuticals onto disposable plastic syringes. J Nucl Med Technol 1998; 26:196–199. Gunasekera RD, Notghi A, Mostafa AB, Harding LK. Adsorption of radiopharmaceuticals to syringes leads to lower administered activity than intended. Nucl Med Commun 2001; 22:493–497. Slater DM, Anderson M, Garvie NW. Syringe extractables: effects on radiopharmaceuticals. Lancet 1983; ii:1431–1432. Waight CC, Beattie LA, O’Brien LM, Millar AM. Preparation of 99mTcMAG3:Radiochemical purity is affected by the residence time of sodium chloride solution in a three-part syringe. Nucl Med Commun 2006; 27:197–200. GE Healthcare, Amersham, UK. Nanocoll package insert, June 2003. Davis MA, Jones AG, Trindade H. A rapid and accurate method for sizing radiocolloids. J Nucl Med 1974; 15:923–928. Hilditch TE, Elliott AT, Murray T, Whateley TL. Formation of large particles in a 99 m Tc -tin colloid preparation. Nucl Med Commun 1986; 7:845–850. Gommans GMM, van Dongen A, van der Schors TG, Gommans E, Visser JFM, Clarijs WWJ, et al. Further optimisation of 99mTc-Nanocoll sentinel node localisation in carcinoma of the breast by improved labelling. Eur J Nucl Med 2001; 28:1450–1455.

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Stability of

14

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Veronesi U, Paganelli G, Viale G, Luini A, Zurrida S, Galimberti V, et al. A randomised comparison of sentinel-node biopsy with routine auxillary dissection in breast cancer. N Engl J Med 2003; 349:546–553. Hegge FN, Hamilton GW, Larson SM, Ritchie JL, Richards P. Cardiac chamber imaging: a comparison of red blood cells labeled with Tc-99m in vitro and in vivo. J Nucl Med 1978; 19:129–134.

16

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Tc-Nanocoll in unit-dose syringes O’Brien et al. 1003

Millar AM, Wathen CG, Muir AL. Failure in labelling of red blood cells with 99m Tc: interaction between intravenous cannulae and stannous pyrophosphate. Eur J Nucl Med 1983; 8:502–504. Hambye AS, Vandermeiren R, Vervaet A, Vandevivere J. Failure to label red blood cells adequately in daily practice using an in vivo method: methodological and clinical considerations. Eur J Nucl Med 1995; 22:61–67.

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BNMS report

Training of staff for the delivery of PET/CT services in the UK Alan C. Perkins, Isky Gordon, James Read, Beverly Ellis and On behalf of the Professional Standards and Education Committee of the British Nuclear Medicine Society. (Committee members: R. Allen S.E.M. Clarke C. Garner, A.J. Hilson J.W. Frank, D. McCool, A. Nicol, M.C. Prescott, P.J. Ryan, R.A. Shields and W.B. Tindale) Evidence for the cost effectiveness of PET/CT imaging is now driving the widespread introduction of PET/CT services throughout the UK. The provision of PET/CT facilities will require a workforce of medical, scientific, technical and engineering staff who are adequately trained and fit for purpose. Suitably trained staff in this speciality are scarce. The development and accreditation of training courses and other educational resources for training programmes in all disciplines will therefore be required at a national and regional level. The implementation of PET/CT training can be achieved more cost-effectively by developing multi-professional learning resources whenever

possible. It is intended that the recommendations would be implemented by close co-operation of both public and private healthcare providers together with educational c 2006 establishments. Nucl Med Commun 27:1005–1010 Lippincott Williams & Wilkins.

1. Introduction

maintaining competency need to be set and a mechanism needs to be in place for competency assessment. Where appropriate, web-based learning should be explored in order to minimize the impact of such training on the provision of other NHS nuclear medicine services. The BNMS multi-professional education initiative could provide a suitable resource but development of PET/CT modules would require seed funding.

The strategy and standards for the provision of PET services in the UK have been published [1,2]. This document on ‘The training of staff for PET/CT services’ is directed at all health care professionals who will either be involved in PET/CTunits or those who require a PET/ CT service for their patients. This information should also be valuable for those in the independent sector involved in providing any part of a PET/CT service. This is not the curriculum for full training of health care professionals, but rather a document that will inform the appropriate committees and bodies who will develop the full curriculum for each health care professional group.

2. Summary of training and educational issues The provision of PET/CT services will require a workforce of medical, scientific, technical and engineering staff. Staff trained in this speciality are scarce. Training programmes will be required at a national and regional level. In addition, mutual recognition of overseas qualifications would make it easier for staff to move between countries in order to develop services. The establishment of PET/CT training resources could be achieved more cost-effectively by developing multi-professional learning resources whenever possible. Such multi-professional learning programmes could be phased in as the demand increases but they must be designed to meet the needs of all the relevant professional groups. Standards for achieving and

Nuclear Medicine Communications 2006, 27:1005–1010 Keywords: PET/CT, Staff training. Correspondence to Professor A.C. Perkins, Department of Medical Physics, University Hospital, Nottingham, NG7 2UH, UK. Tel: + 0044 0115 9709192; fax: + 0044 0115 9422745; e-mail: [email protected]

With appropriate funding it may be possible to release PET/CT trainers for 1 to 2 days per week (and backfill their positions) so that structured, mentored training can take place in their host institutions. This applies to all health care professionals involved even though there may be difficulty with backfill in certain professional groups due to few numbers nationally. This could be coordinated centrally so that trainees/consultants/senior professionals have the opportunity of visiting at least two different centres and can be exposed to a wider variety of different cases. Improvement in electronic communication within the NHS could allow some mentored teaching and reporting to be undertaken remotely from the trainee/ consultant’s home institution. With coordinated planning and appropriate funding, current UK PET expertise can be expanded to meet the skills shortage in all PET disciplines that otherwise threatens the development of PET services in the UK, both in the short and long terms. The following are required to achieve this expansion:

c 2006 Lippincott Williams & Wilkins 0143-3636

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1006 Nuclear Medicine Communications 2006, Vol 27 No 12

Development and accreditation of training courses and other educational resources for all disciplines. Funding to set up new training courses and other educational resources (e.g., e-learning) K Funding for training at centres that have relevant expertise K Salary support to allow backfilling of posts to release trainers K Training of staff prior to commencement of a new PET service and/or replacement of experienced staff moving from training centres to other NHS sites or the private sector K A multi-professional approach (where possible) to maximize more cost-effectiveness of training programmes K Involvement of the NHS, academic institutions and independent sector providers of PET services in PET training, in view of the shortage of training opportunities K Clear identification of the costs of providing PET/CT education so that these overheads can be taken into account when comparing the costs of PET service delivery between training and non-training centres.

training in radionuclide radiology during their first 3 years training. The training requirements for the radiology SpR will be the same as that for the nuclear medicine SpR.

K

3. Medical training for PET/CT Medical personnel are likely to be predominantly radionuclide radiologists and nuclear medicine physicians. Training requirements are expected to differ for specialist registrars (SpRs) and currently practising consultants. The former will receive full formal training in PET/CT whilst the latter may need more individualized training depending on existing skills and previous training and exposure to either PET or CT. An important aspect of delivery of CT training is the need for an emphasis on training in PET/CT as a standalone modality, as separate training in PET and CT is unlikely to be adequate given some of the pitfalls and artefacts etc, that are peculiar to the (PET/CTa PET + CT). 3.1 Future trainees (SpRs) (a) Nuclear medicine physician trainees

3.2 Existing consultants

Practising consultants should obtain full training in all aspects of PET/CT. However, where this ideal cannot be met, dual reporting between a nuclear medicine physician with PET experience and a radiologist with CT experience may be an acceptable compromise. (a) Nuclear medicine physicians

Supplementary CT training will be required if solo reporting is anticipated. The learning outcomes and level of competence should be the same as those proposed for future trainee nuclear medicine physicians. (b) Radiologists

It is envisaged that PET/CT will be normally undertaken by accredited radionuclide radiologists. PET/CT is not currently part of the training programme and this group would have to obtain separate training to the radionuclide radiology training. Individual radiologists without full radionuclide radiology training but with suitable PET training and experience, including the basic science knowledge for PET, tracer theory and knowledge of the limitations of the technique, may also undertake PET/ CT reporting at the discretion of the ARSAC certificate holder and approval of their employer. Individual training requirements will need to be decided by a consultant with ARSAC certification in PET. These will depend on previous experience and skills but a minimum of 300 mentored PET/CT studies are required before solo reporting. Additional training in a PET/CT training centre, accredited by SAC of JCHMT, will be required if a consultant radionuclide radiologist wishes to hold an ARSAC certificate. 3.3 Categories of competence

Two categories of involvement are envisaged with each requiring different knowledge bases and acquired competencies: Category A: PET ARSAC certificate holder Category B: PET/CT reporter (operator under IRMER and with ARSAC certificate holder’s agreement).

Both PET/CT and SPECT/CT will be fully covered during their 4-year training programme. This is under the auspices of the Specialist Advisory Committee (SAC) for Nuclear Medicine of the JCHMT.

K

(b) Radiologist trainees

An ARSAC certificate holder, in conjunction with other appropriate professionals would be able to accept and manage referrals, protocol design and implementation and undertake IRMER practitioner duties, in addition to the abilities of a PET/CT reporter. For example, a PET/ CT reporter would be able to report independently

Currently, training in PET falls within the year 6 curriculum and thus full PET training is not contained within the year 5 curriculum for radionuclide radiology. Placing PET/CT fully into the year 5 curriculum can only be considered when the radiology SpRs have adequate

K

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Training staff for UK PET/CT services Perkins et al. 1007

PET/CT examinations and be involved in PET/CT research, teaching and audit. 3.4 Participation at multi-disciplinary meetings

It is essential that the results of PET/CT examinations are effectively integrated into patients’ management plans. Ideally there should be sufficient consultants with category A or category B competence to attend all relevant multi-disciplinary meetings (MDMs). If a PET/CT examination requires re-interpretation following discussion at an MDM, then the study should be referred back to a consultant with category A or B competence for review and re-presented at the MDM at a later date.

4. Nuclear medicine technologists and radiographers 4.1 Current status

Nuclear medicine technologists include those from both radiography and clinical technology backgrounds. There is a national shortage of nuclear medicine technologists. Radiographers currently receive minimal theoretical training in PET within BSc courses and no practical PET experience is included in present training courses. It is recommended that radiographer first-degree syllabus includes an increase in the nuclear medicine component. For clinical technologists, basic training includes minimal theoretical knowledge and no practical experience in PET. Formal teaching of PET is included in certificate and diploma/MSc courses in nuclear medicine, i.e., 2 h formal teaching in PET equipment, 1.5 h in CT basics and 4 h PET & PET/CT applications & techniques. Visits to PET centres are also included but no practical handson experience is currently required for portfolios of experience because access to appropriate sites is limited. The EANM currently offer short courses for PET technologists. Some distance learning is available from the University of Sydney. Practical experience may be gained by technologists and radiographers participating in a structured training rota to a PET centre with appropriately trained staff. 4.2 Future needs

There is a worldwide shortage of nuclear medicine technologists and one can not expect to recruit into PET from this source. Yet, recruitment of new nuclear medicine technologists is essential especially as many technologists currently in nuclear medicine departments may go into PET thus leaving a bigger problem in the gamma camera section of nuclear medicine. Only by providing a readily accessible, high quality, training programme that also allows for an expanded role of the nuclear medicine technologist will recruitment improve. The expansion of PET services in the UK will require development of additional training resources in order to address the current shortage of nuclear medicine technologists trained in PET. There is a need to establish in the UK specific PET/CT certificates of competence,

diploma and/or MSc modules that are accredited by appropriate bodies (e.g., Society & College of Radiographers, IPEM, BNMS, CANME). Future training programmes for clinical technologists are currently under development by collaboration between the Institute of Physics and Engineering in Medicine and the higher educational institutions. A combined multi-professional course attended by both radiographers and clinical technologists working in nuclear medicine (and possibly others, e.g., physicists) is likely to be more cost-effective, even though each craft group is likely to begin the course with some of the necessary knowledge and skills. Practical experience will need to be offered by all current and all future PET/CT service providers. Close and co-operative working between the NHS and independent sector is essential for all training programmes. Once technologists are trained it will be expected that each will maintain a record of continuing professional development.

5. Medical physicists There is an overall shortage of medical physicists coming into the field of nuclear medicine. As with all nuclear medicine investigations to satisfy legal requirements medical physicists are required to act as IR(ME)R 2000 ‘medical physics experts’ in departments undertaking PET/CT examinations. This necessitates a detailed knowledge of radiation physics, radiation dosimetry, equipment operation and equipment quality assurance. In-depth education, training and experience will be required before individuals can take on this role. This professional group may also contribute to the production of procedure protocols, image analysis and in some instances the reporting of patient scans. 5.1 Trainees

Currently, training in medical physics following an honours degree is via the NHS clinical scientist training scheme. This scheme currently involves the IPEM diploma/MSc plus a further 2 years advanced training leading to state registration as a healthcare clinical scientist with the Health Professions Council. In future, new training schemes may be led by the higher education sector. Placements in PET/CT or PET and CT should be encouraged as part of advanced placement programmes in nuclear medicine once comprehensive nuclear medicine experience has been gained (such training would also be required for SPECT/CT procedures in nuclear medicine departments). At the present time the number of suitable training sites and availability of staff to perform the training is limited, but it is expected that this will grow as more centres are established. Taught MSc courses or even professional doctorates could be readily modified to include theoretical aspects of PET & PET/CT. A higher level of training would be required

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1008 Nuclear Medicine Communications 2006, Vol 27 No 12

to support research that requires the use of non-routine or complex PET methodology. 5.2 Existing staff

Nuclear medicine physicists already in post could gain the additional experience necessary to obtain IRMER medical physics expert status with respect to PET/CT by: (1) a short placement in PET/CT or PET + CT or (2) short teaching courses. An appropriate course could be organized through the professional bodies such as BNMS, IPEM, EANM or through higher educational teaching establishments. It is likely that medical physics support will come from nuclear medicine specialists, at least initially. Training departments outside the UK could also play an important role in training UK professionals. Nuclear medicine physicists may also contribute to reporting of patient scans. Training and experience in PET/CT will require placements at centres undertaking these scans. Resources will need to be made available to fund all the developments above. In particular, as the existing PET expertise is sparse and stretched, it can not be automatically assumed that existing centres will be able to take on substantial additional training commitments unless appropriate and substantial resources are put in place to allow them to do this.

6. Radiopharmaceutical scientists Radiopharmaceutical scientists are mainly radiopharmacists (Royal Pharmaceutical Society of Great Britain (RPSGB) registered and those who are internationally recruited but are not eligible for RPSGB registration) or radiochemists by background. However, some of these scientists may have a physical or life science background and have developed specialist expertise in the science and practice of radiopharmacy. There is a critical shortage of senior level radiopharmaceutical scientists (recognized radiopharmacy experts) who can direct and manage a PET radiopharmacy service that complies with the standards required by the Medicines and Healthcare Products Regulatory Agency (MHRA) for the preparation of PET radiopharmaceuticals suitable for use in patients. The expansion of PET will place a significant burden on the current small pool of senior level radiopharmacy experts. There are serious issues regarding the state registration of radiopharmaceutical scientists. Radiopharmacists who have undertaken an approved pharmacy training can be registered with the Royal Pharmaceutical Society of Great Britain (RPSGB). However, internationally recruited experienced radiopharmacists (including those recruited

at the head of radiopharmacy service level) may not be eligible for registration by the RPSGB. Radiochemists and radiopharmaceutical scientists from other scientific backgrounds are not eligible for registration by the RPSGB. Although prior to July 2005, radiopharmaceutical scientists (including radiopharmacists and radiochemists) were able to register as a clinical scientist via the Healthcare Professions Council (HPC), there is now no mechanism for state registering radiopharmaceutical scientists via the HPC route. There is a need to provide a register for radiopharmaceutical scientists under the clinical scientist grouping. There is an urgent need for a recruitment programme for this group of scientists and clearly defined training and career pathways as this lack of career pathway and the current uncertainty regarding registration with the HPC will seriously hinder recruitment. This shortage of radiopharmaceutical scientists is not only in the UK but worldwide so even if recruitment from overseas is possible it is likely to be limited. There will still be an acute need to train this group of healthcare professionals in the UK to provide conventional radiopharmacy services and additional PET radiopharmacy provision. The current group of senior level radiopharmaceutical scientists are willing and able to provide training to produce a viable and competent workforce to maintain radiopharmacy services in the future. Currently, there is no structure that would enable this to happen. As the issue is likely to be wider than trust level it is essential that there is some strategic guidance as trusts, in the current financial climate, are unlikely to set up and fund training grades from which they may not benefit. The training would consist of theoretical training, e.g., MSc in radiopharmaceutics and PET radiochemistry and practical training involving placements in conventional radiopharmacy and PET radiopharmacy under the direction and supervision of a recognized radiopharmacy expert. It is essential that training prepares those being trained both for conventional radiopharmacy and PET radiopharmacy in all areas of which a severe shortage of radiopharmaceutical scientists exists. The small group of recognized experts in radiopharmacy and PET radiochemistry could assist in the development of PET radiopharmacy services using a hub-and-spoke type model similar to that of the training proposed in the document Nuclear medicine and radionuclide imaging. A strategy for provision in the UK. A report of the Intercollegiate Standing Committee on Nuclear Medicine. This would require further recruitment to support the expansion in the long term. A higher level of training than that needed to maintain a clinical service would be required to support research.

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Training staff for UK PET/CT services Perkins et al. 1009

For those departments undertaking clinical research, an Investigational Medicinal Products (IMPs) manufacturer’s authorization (MA) for the preparation of new radiopharmaceuticals or PET radiotracers may be required. A ‘qualified person’ defined under the EC Directive 2001/83/EC who has the appropriate knowledge and training for the release of radiomedicinal products is required to be named on the IMP MA. There is an urgent need for the training requirements of how to achieve this in practice to be addressed in collaboration by the Nuclear medicine and radiopharmacy community, Department of Health, MHRA and QP assessors from the Royal Pharmaceutical Society of Great Britain, Royal Society of Chemistry and Institute of Biology. If this issue is not addressed then clinical trials in nuclear medicine may cease and this will affect patient care in the future.

7. Radiopharmaceutical technologists Radiopharmaceutical technologists may be from pharmacy technician, nuclear medicine technologist, radiography or chemistry backgrounds. There is a shortage of trained radiopharmaceutical technologists in conventional radiopharmacy services and the expansion of PET will exacerbate the existing situation. There will still be a need to recruit and train radiopharmaceutical technologists for conventional and PET radiopharmacy services. As with radiopharmaceutical scientists there is a lack of a defined career pathway for this group of staff and this situation will seriously hinder recruitment and retention if not addressed alongside the nuclear medicine technologist and radiography groupings. Radiopharmaceutical technologists from a conventional radiopharmacy training would need to undergo additional training to provide PET radiopharmacy services and those recruited from a chemistry or laboratory background would require additional training in the manufacture of radiomedicinal products and good manufacturing practice (GMP). Currently, there is no structured training provision for this group of staff specific to PET or conventional radiopharmacy. Structured training programmes need to be developed for radiopharmaceutical technologists. In a similar manner to radiopharmaceutical scientists there will need to be a syllabus for the core of knowledge and courses to provide this training locally or nationally. Training should also include supervised practice in radiopharmacy both conventional and PET. Currently, these staff are registering on the voluntary HPC clinical technologists register or RPSGB pharmacy technician register. The RPSGB pharmacy technician register only recognizes pharmacy technicians who have undertaken an approved pharmacy technician route and does not specifically recognize radiopharmaceutical technologists. Radiopharmaceutical technologists who have science degree backgrounds are not eligible for registra-

tion via this route. Currently, the voluntary register for clinical technologists is registering radiopharmaceutical technologists from all backgrounds who have the required experience alongside medical physics and nuclear medicine technologists. It would be appropriate for all radiopharmaceutical technologists to be on one state register such as the clinical technologists register as nuclear medicine technologists are also on this register and may undertake radiopharmacy as part of their nuclear medicine activities. Although in some situations staff may have dual registration for both the clinical technologists register and the RPSGB pharmacy technician register depending on the activities they undertake. There needs to be a mechanism for registering radiopharmaceutical technologists on the clinical technologists register when the voluntary register closes and becomes an HPC clinical technologists register. This registration anomaly needs to be addressed urgently by the appropriate authorities.

8. Cyclotron engineers There are very few ready trained personnel available within the UK. There is also a shortage of suitably skilled engineers who could be trained to take on this work and there are no formal higher education based training courses currently available. Cyclotron operators/engineers tend to be drawn from a range of backgrounds in the engineering and radiation sectors. Training is usually acquired on the job, supplemented by visits to overseas manufacturing facilities and user group meetings. Training courses for such engineers are required and a career structure needs to be established. Secondment of UK staff with appropriate backgrounds to overseas centres may be necessary. It is anticipated that the industry would be in a position to assist with this form of specialized training. Since there is little available in the way of training matter the following text has been included to scope the breadth of training material. Cylotron engineers must be trained as radiation workers. They must have a high awareness of safety requirements and be familiar with industrial safety requirements including radiation safety, high voltage safety, hydraulics, vacuum systems, gas systems and bottle farms and helium and water cooling systems. They are expected to have a basic knowledge of the following subjects related to basic accelerator and cyclotron theory: Physics of the atom DC electronics, op-amps, timers, logic circuits K Basic electricity RF systems and magnetism K Basic heat transfer and fluid flow for liquid-phase systems K RF amplifiers, transmission, and antenna K Physics background to negative ion formation K K

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1010 Nuclear Medicine Communications 2006, Vol 27 No 12

K K

Microsoft Windows operating system Microsoft Excel, Windows Explorer and Internet Explorer

In practice the engineer must be competent to service, maintain and calibrate complex equipment being aware of common failures and problems and ensuring continued operation and minimal downtime of equipment. An important component of training would therefore be in troubleshooting, response and recovery in the following areas AC and DC power distributions, power failure, RF tuning, cooling water failure, vacuum failure mechanisms, target system arrangement and operation.

Current educational resources K K K K

Acknowledgements The BNMS would like to thank The Royal College of Radiologists and The Institute of Physics and Engineering in Medicine for their support with this document. Special thanks also go to Dr Paul Marsden at St Thomas’ Hospital, London.

References 1

2

PET/CT in the UK. A strategy for development and integration of a leading edge technology within routine clinical practice. London: Royal College of Radiologists, August 2005. ISBN 1 905034 05 9. Standards for delivering a PET service within the UK. Report of the Intercollegiate Standing Committee on Nuclear Medicine. http:// www.rcr.ac.uk/docs/radiology/worddocs/standardsPETcentre2.doc

Nuclear medicine MSc EANM or SNM PET/CT courses Brighton interactive CT anatomy training modules Short courses and symposia, for example:  Guys & St Thomas’ Clinical PET Course  BIR PET/CT symposia  RMH PET/CT symposium  PETential PET/CT symposium  SNM course CT for the nuclear medicine physician  BIR course CT for the nuclear medicine physician  Alliance Medical PET/CT training course  Conferences by the BNMS, EANM and SNM

Proposed education resources

Proposed electronic learning, for example:  CT content of RCR e-learning modules for medics  BNMS multi-professional e-learning project K ULCH is setting up a training programme for physicists in PET/CT K A new MSc programme in ‘Radiopharmaceutics and PET Radiochemistry’ has recently been set up by King’s College London. K

Appendix: Training resources Whilst most of the current UK PET/CT units already have many visitors obtaining PET/CT exposure, adequate resources to provide structured mentored training to the level required for ARSAC/IRMER is lacking. It is anticipated that training will be acquired from a range of sources, the most important being structured mentored training in a PET/CT unit. Resource locations

Current PET/CT units are at: Guys & St Thomas Clinical PET Centre The Institute of Nuclear Medicine, UCH K The Royal Marsden Hospital K Belfast Royal Victoria Hospital K Mt Vernon Hospital K Overseas training sites K K

The King’s College programme will help to address the international shortage of experienced personnel in this discipline. The course will comprise full-time or parttime postgraduate education and training in radiochemical aspects of PET and radiopharmaceutical science for suitably qualified science or pharmacy graduates wishing to pursue a career in these areas in NHS, industry or academia. Radiopharmacy and cyclotron workplace training and placement will be included. Industry and professional organizations have recognized this is a priority area and are providing some financial support for the course but more financial support is required. This course started in October 2005. Potential UK sites for practical training include St Thomas’ Hospital, PETNet, GE Healthcare, GSK. A robust career structure needs to be established to encourage pharmacists and chemists into the field.

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NEWS AND VIEWS DECEMBER 2006 News and Views is the newsletter of the British Nuclear Medicine Society. It comprises articles and up to date, relevant information for those working within the nuclear medicine community both nationally and internationally. Readers are invited to submit material, meeting announcements and training opportunities to the Editors: Mr Mike Avison, Medical Physics Department, Bradford Royal Infirmary, Duckworth Lane, Bradford, West Yorkshire, BD9 6RJ, UK. Tel: ( + ) 44 (0)1274 364980, E-mail: [email protected] or Mrs Maria Burniston, Medical Physics Department, St James’s University Hospital, Beckett Street, Leeds, LS9 7TF, UK. Tel: ( + )44 (0)113 206 6930, E-mail: [email protected] Nuclear Medicine Communications, 2006, 27:1011

Autumn meeting of the BNMS

This year the autumn meeting of the BNMS was held in Homerton College, Cambridge. This proved an excellent venue with superb lecture facilities in leafy, peaceful surroundings. The invited speakers covered a well-chosen and wide range of subjects from molecular imaging (Dr Alexander McEwan) to the national strategy for diagnostic imaging (Dr Erika Denton). The latter talk was a bravura performance, incorporating almost every government nitiative, project and target relevant to nuclear medicine. The excellent invited lecture on parathyroids was delivered by Mr Gordon Wishart and provided a view from the referrer’s perspective; although this was a referrer who is clearly engaged with the imaging department. His very clear intraoperative photographs described graphically what is really involved in the move to minimally invasive surgery and its attendant challenges. A thought-provoking talk by the local team described some of the common lymph drainage patterns for the breast and arm, demonstrating the probable explanation why even with sentinel lymph node techniques a small proportion of patients will still develop lymphoedema in the arm. As has become standard in recent years, a significant body of lectures addressed the area of PET. Dr Isla

McKenzie presented compelling evidence that a subgroup of phaeochrocytoma are much more easily detected with DOPA PET than with pentetreotide imaging while Dr Doug Abrams described the challenges of production and delivery of FDG in Canada. Overall, the standard of presentations was excellent and the organization flawless. Professor Peter Ell honoured

In September, the World Nuclear Association, honoured Professor Peter Ell by awarding him the World Nuclear Association award. The award is given for distinguished contributions to the peaceful use of nuclear technology. Professor Ell is a long-standing member of BNMS, a co-founder of the European Association of Nuclear Medicine and served as editor-in-chief of the European Journal of Nuclear Medicine from 1990 to 2003. He has made major contributions to nuclear medicine over many years, including editing one of the most comprehensive text books, Nuclear Medicine in Clinical Diagnosis and Treatment. Meeting Announcements British Nuclear Cardiology Society: Annual Conference

Date: Monday 4 December 2006 Venue: National Heart and Lung Institute, London Website: www.bncs.org.uk

British Nuclear Medicine Society: Annual Conference

Dates: 19–21 March 2007 Venue: Manchester, UK Website: www.bnms.org.uk ICNC 2007: 8th International Conference of Nuclear Cardiology

Dates: 29 April to 2 May 2007 Venue: Prague, Czech Republic Website: http://www.icnc8.org Education and Training EANM Learning Courses

Dates: Weekend courses throughout 2007 Venue: EANM PET Learning Facility, Vienna, Austria Contact: EANM executive Secretariat on + 43 1 2128030, fax + 43 1 21280309 Website: www.eanm.org/education/ esnm/esnm_intro.php Email: [email protected] EANM distance learning in nuclear cardiology

This course is designed for physicians who actively participate in the performance and/or interpretation of nuclear cardiology studies. The course is intended to provide a detailed review of the critical elements needed to carry out the technical aspects of nuclear cardiology studies as well as the most common clinical indications. Website: http://www.eanm.org/edu Online/edu_start.php?navId = 332

c 2006 Lippincott Williams & Wilkins 0143-3636

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Abstracts

Abstracts for the Autumn Meeting of the British Nuclear Medicine Society Cambridge, UK, 4–5 September 2006

Nuclear Medicine Communications 2006, 27:1013–1031

PET SESSION Invited talk: Molecular imaging in the era of molecular medicine: how do we get from here to there?

Alexander McEwan Cross Cancer Institute, Alberta, Canada A1 Audit of cancer-related PET–CT in a regional cancer centre

A. Anstee and W.L. Wong Paul Strickland Scanner Centre, Mount Vernon Hospital, Northwood, UK. Background The provision of PET–CT in the United Kingdom lags behind many countries in western Europe. The Department of Health recently published guidelines for the expansion of this service [1]. Purpose To audit cancer-related PET–CTs referred from a regional cancer centre during one financial year and correlate this data with the Department of Health guidelines. Methods Patients referred from the Mount Vernon Cancer Centre (serving a population of 2 million), during the financial year April 2005–2006 were identified. The original request forms were reviewed; clinical indication and cancer type were recorded. Results One thousand, one hundred and eightyseven cancer-related PET–CTs were performed, representing 74.2% of the predicted 800 scans per million population. Lung cancer-related scans most accurately reflected the published guidelines (85.2%). Cancer groups under-represented were lymphoma (18.3%) and cancers with level B evidence (43.5%). Targets were exceeded for oesophageal cancer (207%) and cancers with level C evidence (188.3%). The lead clinicians for the different cancer types were subsequently interviewed regarding the referral patterns. Discussion Wide variation is seen in the number of cancer-related PET–CTs performed. The role of education, particularly multi-disciplinary team meetings, increases the number of scans requested. Financial constraints are a deterrent to requesting PET–CT in some cancer groups.

Reference 1. A Framework for the Development of Positron Emission Tomography (PET) Services in England. Gateway Number 5265. London: Department of Health, October 2005. A2 Implementation of a mobile PET CT service: NHS experience

M.B. Hanney, E. Lorenz, V. Prakash, C. Powell-Wiffen and W.B. Tindale Sheffield Teaching Hospitals NHS Foundation Trust, UK. The framework for future provision of PET imaging in the UK will require many NHS trusts to develop imaging services based around mobile scanners. Although such services will be developed in conjunction with private service providers, their implementation and integration into the trust’s imaging service requires significant input from the host site. We present our recent experiences of implementing, managing and developing a PET–CT service provided by a mobile unit. The required infrastructure and preparatory work is discussed, together with role of clinical, scientific and other staff groups in the day to day running of the service. The inherently limited frequency and capacity of the mobile service has required the development of referral criteria to allow access for those patient groups most likely to benefit whilst ensuring that demand is restricted to a manageable level and acceptable waiting times are maintained. Current referral criteria allow the unit to be used at full capacity whilst maintaining an average turnaround time of 10 working days from receipt of request to issuing a report. The value of the service has been demonstrated by an audit of 171 patients showing management changes in 55% of cases. A3 Is the whole-body PET/CT scan a luxury procedure for investigating pulmonary lesions compared to a thoracic and upper abdominal study?

H. Cheow, A. Winship, S. Rankin, D. Landau and M. O’Doherty King’s College London School of Medicine at Guy’s, King’s College and St. Thomas’ Hospitals, UK.

c 2006 Lippincott Williams & Wilkins 0143-3636


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1014 Nuclear Medicine Communications 2006, Vol 27 No 12

Materials and methods A retrospective review of PET/CT studies (skull base to upper thighs) was performed for either pulmonary nodules or lung cancer staging between January and June 2005. Abnormalities on PET or CT were identified and categorized into location- thorax, upper abdomen, head & neck and other sites. Result Two hundred and forty-nine patients were included in this study. Two hundred and twenty-nine (77.9%) patients showed no lesion outside thoracic region on either CT or PET. In addition to the thoracic lesion, 23 (7.8%), 18 (6.1%) and 12 (4.1%) patients had lesions found in one other region in either the upper abdomen, head and neck, or others respectively. Twelve (4.1%) patients had lesions found in three regions. Comparing with the standard CT scan for lung malignancy covering the area of lung apices to the liver, 18 (6.1%) patients had metastatic disease outside this area. Of these, six (2.0%) cases were upstaged. Conclusion If PET/CT were performed from lung apices to the lower border of the liver, 93.9% of the lesions relevant to the diagnosis were included. 2.0% of the cases will be understaged. References 1. Aquino SL, Fischman AJ. Does whole-body 2-[18F]fluoro-2-deoxy-D-glucose positron emission tomography have an advantage over thoracic positron emission tomography for staging patients with lung cancer? Chest 2004; 126:755–760. 2. Stroobants SG, D’Hoore I, Dooms C, et al. Additional value of whole-body fluorodeoxyglucose positron emission tomography in the detection of distant metastases of nonsmall-cell lung cancer. Clin Lung Cancer 2003; 4:242–247. A4 18F-FDG PET–CT in the investigation of the cancer of unknown primary

S. Hana and I.R. McDougallb a Nuclear Medicine Department, Glasgow Royal Infirmary, Glasgow, UK, and bNuclear Medicine Department, Stanford University Medical Centre, Stanford, California, USA. Introduction Detecting primary tumour in patients, who present with metastatic cancer of unknown primary remains a diagnostic challenge. 18F-FDG PET–CT has been shown to have potential to demonstrate the primary cancer, which was not identified by conventional diagnostic workup. Patients and methods The case records of 20 patients (11 male, 9 female, age 15–84 years, mean 58 years) who had 18 F-FDG PET–CT for the evaluation of metastatic cancer of unknown primary in 2003–2005 were retrospectively reviewed. There were 11 patients with cervical nodal metastases and 9 patients with extra-cervical metastases: liver (2), bone (2), retroperitoneal (3), mediastinal (1) and abdominal wall (1). Results PET–CT revealed already known metastatic sites as well as further metastatic foci. PET–CT correctly

identified the primary cancer in 3 out of 11 patients (27%) with metastatic masses in the neck and allowed more accurate staging for appropriate therapy. They were nasopharyngeal carcinoma (2) and tongue base cancer (1). However, PET–CT failed to detect the primary tumour in any of the 9 patients who presented with extra-cervical metastases. Conclusions This study demonstrates that 18F-FDG PET–CT can play a role in identifying primary tumour in metastatic cancer of unknown primary in head and neck region only. A5 Unsuspected soft tissue and skeletal metastases detected by PET/CT when compared to conventional imaging

O. McNallya, S. Hughesb, J. Clarkeb and T. Lyncha a NI Cancer Centre, Belfast City Hospital and bRoyal Victoria Hospital, Belfast, UK. A central tenant of modern oncological practice is the need for an accurate staging system that allows determination of prognosis and the formulation of an appropriate management plan. For such a system to be effective it must be supported by a reliable, reproducible imaging pathway. Conventional imaging protocols use ultrasound, CT and MRI in an attempt to stage the spread of disease. The introduction of combined PET/CT imaging has impacted significantly on the detection of distant disease and often leads to both staging and management change. PET/CT has made us rethink the assumptions that have previously been made about the patterns of disease dissemination. We illustrate the exquisite use of PET/CT imaging in the identification of unsuspected skeletal and soft tissue metastases in patients with a range of primary malignancies. We demonstrate metastatic deposits that were not found using conventional imaging, many of which have no associated abnormality on either CT or MRI, even with retrospective review. Isolated gluteal deposits from non-small cell lung cancer, pancreatic tail secondary disease from a colonic adenocarcinoma and multiple examples of deposits in morphologically normal muscle, soft tissue and viscera are included in this collection. GENERAL SESSION Invited talk: National strategy for diagnostic imaging

Erika Denton Norfolk and Norwich Hospital, UK. A6 How do service improvement strategies relate to nuclear medicine?

A. Swift, C.A. Segasby, C. Taylor, C.M. Brown, A. Pickles, W. Inman and W.B. Tindale Medical Imaging and Medical Physics Directorate, Sheffield Teaching Hospitals Foundation Trust, Sheffield, UK. The NHS faces its biggest challenge: to improve service quality, meet targets but reduce costs. Trusts face difficult decisions and are scrutinizing all services. Work has recently been under way to assess our directorate’s

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Abstract of the BNMS meeting, September 2006

performance: patient pathways have been process mapped and capacity and demand data acquired. Initial conclusions led to key changes such as reorganization/ relocation of radiological services with rationalization of equipment/rooms. However, it became clear that many models are not easily applied to nuclear medicine due to the multistage nature of patient pathways en route to a gamma camera. This work considers the limitations of service improvement models when applied to nuclear medicine. Particular issues addressed include the inability of gamma camera occupancy to truly reflect capacity and nuclear medicine inefficiencies originating from external sources. Staff involvement at all levels is critical for success and the needs of the patient must be placed centrally in developing future workflow models. Initial outcomes include identification of bottlenecks (e.g., poor referral–receipt times; poor patient compliance with pretest instructions) and ‘black holes’ for the patient (e.g., the delay from referral to appointment confirmation). By following an appropriately focused approach, improvements in quality and service can be achieved in parallel with reductions in expenditure. A7 Improving the service: IT, IHE, PACS and nuclear medicine

N.J.G. Brown BIR IHE-UK, London, UK. Integrating information technology (IT) services within the work practices of a nuclear medicine service can be a challenge, especially when the system has been designed for use in radiology. The Integrating the Healthcare Enterprise (IHE) organization exists to enable users and suppliers to work together obtain the smooth working of IT systems in healthcare. It achieves this by carefully defining the tasks that need to be performed and defining IHE integration profiles which specify the way to use standards that are already implemented in supplier systems to enable smooth IT support for the tasks. Suppliers can submit their software for testing at annual test sessions where they show that their system can operate correctly with three other supplier systems. The work of IHE in the UK is managed by a steering committee whose members include representatives of BIR, RCR, CoR, IPEM and, recently joined, BNMS. The technical committee includes 14 company subscriber members. Some key details of the IHE nuclear medicine integration profile are described including: the use of HL7 order and placer numbers; DICOM accession numbers and performed procedure steps; and the use of IHE displayable reports for nuclear cardiology. IHE webbased document access is also described. A8 Valvular regurgitation and diastolic function

I.P. Clements, D.O. Hodge and C.G. Scott Mayo Clinic, Rochester, Minnesota, USA.

1015

Background To determine whether valvular regurgitation affects diastolic function? Methods Radionuclide angiocardiography (ERNA) was performed on 760 patients (aged > 44 years) referred for cardiac evaluation; seventy-three had valvular regurgitation. Two groups were identified the first (395) with LVEF > 0.49, and the second (365) with LVEF < 0.50. Rapid LV filling was assessed from the peak-filling rate (PFR, mls – 1m – 2). Three patterns of PFR were defined in 37 same-age subjects with no cardiac pathology; PFR < 10th, between the 10th and the 90th, and > 90th percentile value, respectively, defined delayed relaxation, normal filling or restrictive filling. Results When the LVEF was > 0.49, 30 patients had regurgitation; in regurgitation PFR was significantly greater (348 ± 266 vs. 229 ± 108, P < 0.0001) and the incidences of delayed relaxation, normal and restrictive filling were significantly different (23%, 47% and 30% vs. 44%, 48% and 8%, respectively; P = 0.0002). When LVEF < 0.50, 43 patients had valvular regurgitation; in regurgitation PFR was significantly greater (339 ± 182 vs 220 ± 102, P < 0.0001) and the incidences of delayed relaxation, normal and restrictive filling were significantly different (21%, 46% and 32% vs 49%, 44% and 7% respectively, P = 0.0001). Conclusion In independent determinations of LVEF, regurgitation was associated with increased PFR and a 50% reduction in delayed relaxation and a 4-fold increase in restrictive filling. A9 Investigation of normal ranges for LVEF MUGA studies using three different processing workstations

S.C. Hiscocka, D. Polleya, M.J. Evansa, J. FitzGeralda, D. Hallb and F. Zananirib a Royal United Hospital, Bath, and bUnited Bristol Healthcare Trust, Bristol, UK. A study has been undertaken to establish normal ranges for LVEF MUGA studies using the processing techniques used at the Royal United Hospital (Bath), Bristol Royal Infirmary and Southmead Hospital (Bristol). A retrospective study of patients who had LVEF MUGA studies performed at the RUH between August 2003 and July 2006 was undertaken in order to identify normal studies. Patients were included who had a MUGA LVEF result with normal wall motion and who also had left ventricular function qualitatively identified as normal on echocardiogram. These studies were transferred in interfile format and processed on each of the other workstations by two operators. All studies were reprocessed to give intra- and inter-operator variability. The results have been used to establish normal ranges for each of the processing methods. These normal ranges and distributions have been compared and the possibility of assigning a national single cut-off value, as recommended by the NICE guidelines for Herceptin, will be discussed.

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1016 Nuclear Medicine Communications 2006, Vol 27 No 12

PHYSICS SESSION Invited talk: Progress with combined PET/MRI

T. Adrian Carpenter Wolfson Brain Imaging Centre, Cambridge, UK. A10 Systematic measurement of PET and CT scan misalignment for a GE Discovery ST PET–CT scanner

E.J. Somer, P.J. Schleyer and P.K. Marsden Guy’s, King’s and St Thomas’ School of Medicine at King’s College London, UK. The scanning couch of the GE Discovery ST (DST) consists of a support, taking either the CT or PET position, and a sliding table-top that carries the patient through the gantry during image acquisition. The misalignment of PET and CT images of a registration phantom scanned with the table top in retracted, intermediate and extended positions relative to the supporting pillar was measured with couch loadings to represent a 70 kg patient orientated either head-in or feet-in to the scanner bore. The tests were performed on two identical DST scanners. PET–CT alignment along the y-axis (vertical) for both scanners was dependent upon the position of the tabletop relative to the supporting pillar. Errors ranging between 1.3 mm (retracted) and – 1.8 mm (extended) were minimized when the couch was in an intermediate position, independent of patient orientation. Errors along the z-axis (axial) were reduced as the tabletop moved from the retracted (2.4 mm) to the extended (0.6 mm) position; this effect was also present on the second scanner (1.7 mm to 0 mm). Errors on the x-axis did not follow a particular trend for either scanner. The largest rotational error was 0.71 and the maximum RMS misalignment was 3.1 mm. Applications assuming accurate image alignment should account for these errors. A11 A review of acquisition modes on a newly acquired PET/CT imager

R. Peare, F.I. McKiddie, E.J. Davidson, L. Schweiger and M.E. Brooks John Mallard Scottish PET Centre, Aberdeen, UK. A review was taken of the protocols used for whole-body FDG PET imaging on our newly acquired GE Discovery STE PET/CT scanner. This scanner has both 2-D and 3D acquisition modes. The protocol currently used in our centre is that recommended by the applications team. The default acquisition is 3-D imaging with a bed time of 2.5 min. Other centres with the same or similar scanners are using different protocol, with some centres acquiring in 2-D mode. There was particular concern about the level of noise present in the liver on the patient images. Phantom and patient studies were performed in both 3-D and 2-D with bedtimes of 5 min each. The 3-D data was acquired in list mode so that other bed times could be examined. The default reconstruction parameters were found to be suboptimum and were altered before further

analysis. Contrast-to-noise ratio and observer experiments were used to determine the optimum parameters. Initial studies show that the current bed time of 2.5 min is too short. An acquisition time of at least 4 min per bed is required in 3-D mode. Further patient studies are ongoing to compare 2-D and 3-D modes in clinical cases. A12 Checking CT–SPECT alignment using a cylindrical flood phantom

I. Murray, J. Anton and N. Burtally Barts and the London NHS Trust, UK. Modern hybrid SPECT–CT systems rely upon an assumption that both the reconstructed CT and SPECT images are inherently registered to each other without the need to use registration algorithms. In order to test this assumption it is possible to use a phantom arrangement consisting of radio-opaque sources filled with a gamma emitting radionuclide. The purpose of this study was to test an alternative method using a cylindrical flood phantom used for routine SPECTuniformity quality control. A standard uniformity acquisition was made on a GE VG gamma camera including a Hawkeye acquisition. The data was iteratively reconstructed 9 times. For each reconstruction the attenuation map was offset in the y direction between – 4 and + 4 voxels. The central slices of the reconstructed slices were summed together yielding 9 uniformity images relating to the different attenuation map offsets. The standard deviation over the ‘uniform’ region was calculated and plotted against the offset of the attenuation map. The position of the minimum value of these plots was then taken to represent the mis-alignment in the y-axis direction. This was repeated using offsets in the x-axis direction. This method provides a simple means of checking the alignment between CT and SPECT images without the need for an additional phantom or camera time. A13 Acceptance testing of a brand new, fully diagnostic hybrid SPECT–CT camera: Physicists, factories and funny phantom activity

R.C. Fernandez, K. Adamson and S. Allen Guy’s and St Thomas’ NHS Foundation Trust, London, UK. Independent, scientifically rigorous acceptance testing is always essential and is especially important for hybrid systems which can perform fully diagnostic CT and are therefore capable of delivering large radiation doses to both patient and staff. The Philips Precedence, installed at Guy’s in January 2006, is such a hybrid system and is the first of its kind to be installed in Europe. This paper details acceptance testing of this system, during which numerous complex problems were encountered, which required extensive dialogue between physicist, vendor and factory to resolve. Examples include availability and transparency of factory specifications, incorrect COR calibrations at installation, and ‘phantom’ radioactive contamination attributed to software glitches. Resolving

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Abstract of the BNMS meeting, September 2006

these problems prolonged the duration of acceptance testing, which had implications on staffing resources and clinical throughput. After acceptance testing each modality independently, the fused system requires appropriate testing. SPECT–CT alignment was verified and is monitored regularly. We propose easily available, in-house test phantoms which can be used specifically for this purpose, and make recommendations for type/frequency of quality control (QC) checks. Our experience should benefit other centres to plan and complete an effective acceptance testing schedule as well as set up appropriate procedures for routine QC of new hybrid imaging systems. A14 Evaluation of collimator response modelling in 3-D OSEM iterative SPECT reconstruction

A.D. Scotta,b and L. Livieratosa a Guy’s and St Thomas’ Hospital, London, and bKing’s College Hospital, London, UK. Modelling of collimator response (CRM) as part of iterative reconstruction has been proposed as a resolution recovery technique in SPECT. Recently, such schemes have become commercially available from a number of manufacturers as an option within routine reconstruction software. The aim of this study was to evaluate the use of CRM (Astonish, Philips Medical Systems) on SPECT data from the Philips Precedence camera. Ceramic point sources (diameter, B2 mm) concealing 99mTcO4 were inserted at various depths in an anthropomorphic torso phantom (Data Spectrum) for measurements of spatial resolution with filtered back-projection, MLEM, OSEM and Astonish with and without attenuation correction (AC). The effect on noise levels was assessed by the coefficient of variation within a ROI defined on a 20 cm cylinder of uniform activity at varying acquisition times. Attenuation maps were derived from CT images in both cases. Results show that CRM provides improved FWHM and FWTM over the other reconstruction schemes and significantly reduces the depth dependency of spatial resolution without affecting conservation of counts. AC further improves quantitative accuracy of activity ratios as assessed by well-counter measurements of the sources. CRM demonstrated lower noise levels with less dependency on acquisition time. A15 The effect of reconstruction techniques on quantification in DaTSCAN imaging

E. Kalogiannia, D.J. Toweya, P.G. Bainb, J.W. Franka and K.S. Nijrana a Hammersmith Hospitals NHS Trust and bUniversity Department of Neuroscience, Imperial College London, UK.

tions were used to give series of transverse slices, which were then analysed using BRASS (NUD, Stockholm). The images were visually assessed as essential tremor (ET, 15), Parkinson’s disease (PD, 7), early PD (8). The BRASS results were then compared with this assessment. Using just the first two groups, both FBP and OSEM reconstructions showed good agreement with visual assessment. OSEM was found to be slightly better than FBP in delineating disease. Putamen uptake was found to be a better indicator than caudate or striatum uptake. But when visual assessment indicates early disease the ability of quantification to identify PD from ET is reduced. The results show significant differences between ET and PD patients. However, when early stage disease is imaged quantification fails to separate them from the normal range. Reasons for this failure include changes in the nonspecific binding and the use of gross regions, both of which will mask smaller changes in striatal uptake. It was found that neither reconstruction technique allowed quantification to identify early disease. ENDOCRINE/SKELETAL/INFECTION SESSION Invited talk: Pre-operative localization for primary hyperparathyroidism

Gordon C. Wishart Addenbrooke’s Hospital, Cambridge, UK. A16 Should multimodality imaging be used in localizing suspected parathyroid adenomas?

F. Sundram, S.A. Mostafa and A. Notghi Department of Nuclear Medicine, City Hospital, Birmingham, UK. Background Preoperative localization using functional imaging (FI) or ultrasound (US) can identify suspected parathyroid adenomas (PA) .We investigated the role of both techniques. Methods Seventeen patients (14 F, 3 M) with biochemical hyperparathyroidism had FI (dual-isotope subtraction with 99mTc-MIBI and 123I) and also US neck performed by an experienced operator. Histopathology of resected specimens was noted. Results These are given in Table 1. Of 4 positive isotope studies, 4 had PA (sensitivity 100%). Two US were positive (sensitivity 50%). Of 13 negative isotope studies, 9 were negative on US. One symptomatic patient had surgery, revealing a PA (false negative on both studies). The remaining 8 patients had no surgery. In 4 Table 1 Isotope

Positive (n = 4)

To assess the effect of reconstruction techniques on the quantification of DaTSCANs, data for a group of 30 patient scans were retrospectively analysed. Filtered back-projection (FBP) and iterative (OSEM) reconstruc-

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Negative (n = 13)

Ultrasound Positive

Negative

2 2 PA

2 2 PA

4 2 PA ( < 1 cm) 2 osteomalacia

9 8 no surgery 1 PA

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1018 Nuclear Medicine Communications 2006, Vol 27 No 12

US positive but isotope negative patients, 2 PA ( < 1 cm) were noted (US sensitivity 100%, isotope sensitivity 50%). Two remaining patients had osteomalacia due to parathyroid hyperplasia. Conclusion In localizing suspected PA, isotope studies demonstrate high sensitivity. US has a moderate false positive rate. Patients with negative isotope studies (usually smaller PA) should undergo US by an experienced operator. A17 The impact of radionuclide imaging in the investigation of painful knee prostheses

A. Cai, H. Mohan and S.E.M. Clarke Department of Nuclear Medicine, Guy’s and St Thomas’ NHS Foundation Trust, UK. Background Infection of knee prosthesis is a serious complication of total knee arthroplasty associated with high patient morbidity. Confirming the diagnosis may be difficult clinically. Aim To assess the impact of 99mTc-diphosphonate (MDP) bone scintigraphy and 111In white cell (WBC) scans in the management of patients with painful knee prosthesis. Method Scan results and clinical outcome in 28 patients (14 male, 14 female, mean age 40 years) with 34 total knee arthroplasties who had both a 2-phase 99mTc-MDP and 111In WBC scan performed at Guy’s and St Thomas’ Hospital for suspected infection between 2002 and 2005 were studied. Biochemical markers, radiographic reports, intervention and subsequent outcome were all noted from case files. Results There is a close relationship (97.5% concordance) between the pattern of bone scintigraphy on blood pool phase and 111In WBC scans. Our results show that certain scan patterns are associated with high specificity and sensitivity for detecting infection. 87.5% (n = 14) of patients with positive radionuclide scans subsequently underwent further surgery. Further surgery in patients with a negative scan accounted for only 7.7%. Conclusion Bone scintigraphy patterns including blood pool correlate well with white cell scan results, and the results significantly influence subsequent patient management. A18 Optimization of sacro-iliac index

H. Richardson and M. Al-Janabi King’s College Hospital and Queen Elizabeth Hospital, London, UK. Background Quantification of the ratio of sacro-iliac uptake to other bone uptake (SI index) in 99mTc-MDP scans may reveal underlying inflammatory disease in the SI joint (‘sacroiliitis’). However, no definitive method has been published. This uncertainty has prevented acceptance of SI index quantification, and it is still viewed as a

controversial indicator [1]. Different techniques shall be investigated to assess the efficacy of SI index. Method A group of 48 normal bone scans (26 female, 22 male) were selected from our archive on the basis of a completely normal consultant’s report. These were compared to 46 patients (29 female, 19 male) who had been specifically referred for SI index. The best analysis technique was investigated by comparing the distribution of the ‘normal’ group SI index with the ‘referred’ group SI index. Results The traditional profile method was found to have no diagnostic power. An ROI method (using ROI over SI and the ‘trough’ between SI and sacrum) was found to separate the normal from the referred group. By excluding non-bilateral results an optimal sensitivity:specificity of approximately 90%:80% can be achieved. Conclusions Comparing SI ROI against a trough ROI is the best method for detecting abnormal SI index, and gives a good indication of sacroiliitis. Reference 1. Lin, W-Y et al. Influence of age and gender on quantitative sacroiliac joint scintigraphy. J Nucl Med 1998; 39. A19 111In-labelled WBC imaging in the detection of fever of unknown origin: a 2-year audit

N. Seshadri, P. Burli and K. Balan Department of Nuclear Medicine, Addenbrooke’s Hospital, Cambridge, UK. Background 111In-labelled white blood cell (111In-WBC) imaging is often used for diagnosing fever of unknown origin (FUO). Its diagnostic performance, however, is variable, with a broad range of sensitivities and specificities being reported. Aim To evaluate the diagnostic performance of 111InWBC scintigraphy in the detection of FUO in the light of improved diagnostic performance of other imaging modalities in our institution. Materials and methods A retrospective review of patients undergoing whole-body 111In-WBC scanning for FUO over a 2-year period was performed. Of the 61 patients investigated, follow-up was complete in 54 cases. Results The studies were true positive in 11 patients, true negative in 23 patients, false positive in 11 patients and false negative in 9 patients. The figures for sensitivity, specificity, positive predictive value and negative predictive value were 55%, 68%, 50% and 72%, respectively. There was no significant change in the sensitivity in post-operative cases (55%). The specificity, however, was much higher (80%) with a PPV of 83% and a NPV of 50%. CRP and leukocyte counts did not differ significantly between patients with true positive and true negative studies. Conclusion The results confirm that the accuracy of 111 In-WBC imaging in our institution is similar to what is

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Abstract of the BNMS meeting, September 2006

reported in the literature. Further, although the overall sensitivity is not good it does fare well in the investigation of post-operative patients. A20 Investigation of infection in replacement joints

C.L. Paterson, A. Bolster and J.B. Neilly Glasgow Royal Infirmary, UK. A prospective audit was performed at Glasgow Royal Infirmary of patients referred for investigation of suspected osteitis or infected hip or knee joint prosthesis over the period August 2004 to December 2005. The imaging algorithm included a 3-phase bone scan followed if necessary by an indium-labelled white cell scan and bone marrow scan combination. Patients with normal peri-prosthesic uptake on phase 3 of the bone scan had no further imaging. The reports of 37 patients who had undergone all three scans were reviewed. The bone scan of 10 of the 37 patients studied showed negative dynamic flow and blood pool images. Following the 111In white cell scan and 99mTc bone marrow scan it was determined that, of these 10 patients, none had infection of their replacement joint. From these preliminary results we conclude that if the first two phases of a 3-phase bone scan are negative it may not be necessary to perform further nuclear medicine examinations to exclude infection in patients with a replacement joint. The advantages of this include reduced patient and staff doses and decreased departmental costs. This work is part of an ongoing prospective audit of outcomes in patients referred for infection imaging. RENAL SESSION A21 Glomerular filtration rates measured with EDTA and iohexol

51

Cr-

N.J. Birda, A.R. Michellb and A.M. Petersa a Department of Nuclear Medicine, Addenbrooke’s Hospital, Cambridge, and bWilliam Harvey Research Institute, Department of Biochemical Pharmacology, St Bartholomew’s Medical School, London, UK. Aims To compare reproducibilities of glomerular filtration rates (GFR) independently measured with 51CrEDTA and iohexol and to compare different methods of calculating GFR. Methods Twenty normal volunteers were studied on 2 or 3 occasions and 60 patients once. Independent administrations of 51Cr-EDTA and iohexol were followed by independent blood samples at 20, 40, 60, 120, 180 and 240 min. Plasma iohexol concentration was measured by X-ray fluorescence (XRF) in all 110 studies and additionally by HPLC in 14. GFR was calculated by 3 methods: from (A) the area under the full clearance curve, (B) the area under the curve using the last 3 samples and (C) the slope using the last 3 samples (GFR per unit of extracellular fluid volume).

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Results For both tracers the best reproducibility was for GFR calculated with method B. Reproducibility with iohexol measured with HPLC was significantly better than for iohexol measured with XRF or for 51Cr-EDTA. Considering all the studies there was significantly better agreement between the tracers with method B than with method A or C (P < 0.001). Conclusions GFR based on HPLC of iohexol may be superior to 51Cr-EDTA. Method C is more susceptible to experimental error than method B. A22 Calculation of split renal function from contrast enhanced multidetector CT: comparison with renal scintigraphy

S.M. McDonalda, F. Sebastianb, N.J. Hayesa, Z. Abubackera and J.C. Fowlera a Department of Radiology, Luton and Dunstable Hospital NHS Trust, and bDepartment of Radiology, University College London Hospitals NHS Foundation Trust, UK. Introduction Recent studies have demonstrated that contrast enhanced multidetector CT (CEMDCT) can be used to assess split renal function. We aimed to validate this technique using our recently acquired GE LightSpeed MDCT and to test the reproducibility of the method by measuring inter-observer variability. Method All patients who were investigated with both renal scintigraphy and CEMDCT were identified. Those who underwent intervention between the two studies were excluded. Seven patients (4 F:3 M) with a spectrum of renal disease were included in the final cohort. Split renal function was calculated from 3-D reconstructed CEMDCT images by a single observer blinded to the scintigraphy results using a standardized method. The CT examinations were independently evaluated by four additional observers to assess the inter-observer variability. Results CT derived measurements of split renal function demonstrated excellent correlation with scintigraphy (r = 0.96, P = 0.0004) and inter-observer agreement was good to very good (k = 0.79–0.90). Conclusion CEMDCT provides a valid and reproducible alternative to nuclear medicine techniques for the assessment of differential renal function. The method correlates well with scintigraphy and appears to be transferable between CT manufacturer platforms. It has the additional advantages of low inter-observer variability, anatomical definition, and the potential for reduced radiation burden to the patient. A23 DMSA scintigraphy: Does the quantitative assessment of normal relative function agree with the clinician’s interpretation?

J.M. Anton, A.H. Elmegadmi and N.W. Garvie St Bartholomew’s and Royal London Hospitals, UK. Background The normal range for relative function on DMSA scintigraphy is defined scientifically as 50% ± 5%,

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using the geometric mean (GM) technique, However, DMSA scans are also assessed visually by the clinician, and necessarily categorized as ‘normal’ or ‘abnormal’. Aim To determine the extent to which the GM (scientific) and clinical (subjective) interpretations correlate. Methodology Eighty-one adult patients undergoing DMSA scintigraphy were reviewed. Forty-five scans were graded ‘normal’ clinically (group I) by two observers blinded to the GM result, and 36 were deemed to have visible evidence of unilateral disease (group II). All patients with visual evidence of bilateral disease were pre-excluded. Results In group I, 37/45 (82%) had GM relative values within the normal range. If this range were extended to 50% ± 8% all group I patients would have been quantitatively normal. In group II, 31/36 patients (86%) with a visible unilateral abnormality had relative GM values outside the normal range. The overall accuracy of clinical assessment of normality/abnormality was 84% (sensitivity 79%, specificity 88%). Conclusion There is reasonable correlation between the clinical (subjective) and scientific (objective) assessment of DMSA scans. Where a discrepancy exists, it may be worthwhile performing SPECT, to exclude defects not visible on planar scintigraphy. A24 131I therapy in a thyrotoxic haemodialysis patient: Ascertaining the facts after the prior risk assessment

A. Mackie and E. McCauley Regional Medical Physics Department, University Hospital of North Durham, UK. Aim To quantify staff radiation risks and ensure safe/ legal disposal of radioactive waste. Method A thyrotoxic patient undergoing therapy (361 MBq 131I) underwent serial whole-body imaging and dose-rate measurement (0.3, 0.5, 1 and 2 m), pre- and post-haemodialysis, for 28 days. Solid-waste activity was measured with a gamma camera. A gamma counter measured blood and dialysate-waste activity. Staff wore an electronic personal dosimeter in a breast pocket. Environmental monitors were deployed. Results A mono-exponential fit to whole-body geometric mean counts demonstrated a half-life of 7.45 days. Mean dose rate at 1 m, pre- and post-dialysis, gave a half-life of 7.07 days. Approximately 2–7% of retained activity was lost per dialysis session; most discharged to drains. Activity concentration of blood and dialysate in waste was 0.2 and 0.006 Bqml – 1MBq – 1 131I retained respectively. Staff received 65 mSv. Dose at the adjacent treatment bay was < 0.2 mSv (at 1.5 m). Solid-waste activity was 35 kBq 48 h post-therapy; half-life of 8.8 days. The dialyser contains 60% of total-waste activity. Conclusion Results for whole-body retention of 131I are in keeping with previously reported data; but absolute

and distribution of activity in the waste differs and is probably dialyser specific. TECHNOLOGISTS/NURSES SESSION A25 Comparison between 4 DM SPECT and QGS for the assessment of left ventricular function

J. John, K. Massey and V.R. Lele Department of Nuclear Medicine, Jaslok Hospital and Research Centre, Mumbai, India. Aim To compare two different software package (QGS and 4 DM SPECT) for gated myocardial perfusion SPECT, for assessing left ventricular end diastolic and end systolic volumes (EDV and ESV) of left ventricular ejection fraction (EF). Methods Fifty patients with known or suspected coronary artery disease underwent same-day stress (296 MBq (8 mCi)) and rest (888 MBq (24 mCi)) myocardial perfusion scan using 99mTc-MIBI. Images were acquired with triple head gamma camera (PRISM 3000 XP) using LEHR collimators. Acquisitions over 3601 with gating of rest images using 8 frames/cycle was done. Data were analysed using the QGS software supplied on the Odyssey VP workstation. They were then transferred to an Esoft workstation via DICOM and analysed using 4 DM SPECT software. Images were reconstructed on the Esoft workstation using filtered back-projection with a Butterworth filter, order 5 and cut-off 0.40, and then processed with 4 DM SPECT. On the Odyssey workstation reconstruction was done using a low-pass filter, order 5 and cut-off 21, and processed using QGS. These values were found to be optimum for the 2 types of software. EDV, ESV and EF were calculated by using this software. Results Correlation between the results of QGS and 4 DM SPECT showed: (1) similar EDV, ESV and EF in 35 patients (70%); (2) similar EF values but varying EDV and ESV in 8 patients (15%); and (3) similar EDV and ESV but varying EF values in 14% of patients. Conclusion For any one group of patients, there is a 16% variation in the EDV and ESV obtained by the 2 software packages (QGS and 4 DM SPECT) in 15% of those patients. The EF also varied by 14%. It is advisable for departments to adhere to one particular package for consistency of results. A26 Oesophageal transit in patients with gastro-oesophageal reflux disease: Which is the best labelled food to swallow?

N.B. Smitha, L.K. Hardinga, M. Hawashb and B. Spychalb Departments of aPhysics and Nuclear Medicine, and bSurgery, Sandwell and West Birmingham NSH Trust, City Hospital, UK. We have examined oesophageal transit in 40 patients with dysphagia prior to surgery and at 6 weeks and 6 months later. The meals were liquid (water), semi-solid (egg), and solid (toast) each labelled with 30 MBq 99mTc-DTPA. Patients fasted for 12 h prior to the test and sat in front of

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Abstract of the BNMS meeting, September 2006

the camera (GP collimator), positioned with the cricoid cartilage at the top of the field of view. A practice swallow was performed to familiarize the patient with the procedure, and for each meal 240  0.5-s frames acquired in a 128  128 matrix, with dry swallows performed every 30 s. The procedure was repeated for each of the 3 meal types. Condensed images were created enabling data to be visualized without ROIs and to demonstrate slow transit, hold-up or retrograde motion. The average transits of the 3 swallows from mouth to stomach were calculated for each of the meal types. Liquid and semi-solid swallows were consistent but not as discriminating as toast, in terms of hold-up. Toast showed impaired transit, which was variable between swallows, a problem identified in several of the patients. We recommend toast and 3 swallows as the best procedure to demonstrate dysphagia. A27 Development of a safer needle recapping device through design collaboration with radiopharmacy staff

E.J. Meittunena and J.R. Ballingerb a SafetyEdge.Net, Inc., Preston, Lancashire, and bGuy’s and St Thomas’ NHS Trust, London, UK. Purpose Needle-stick injuries are reported frequently in national injury databases [1,2]. A new concept needle recapping device was developed through the use of a user design development process that promotes collaboration with radiopharmacy staff. Methods Radiopharmacy staff were observed for ergonomics and safety criteria involved in the processes that were associated with the injuries. No currently available devices provided adequate solutions. Macro-design concept requirements were surveyed from staff and generations of prototypes were developed and evaluated based on user needs criteria which included ease of use, ergonomics, suitability for sterilization, and ease of decontamination. Concept design evaluations were accomplished for device generations on four occasions in the USA and once in the UK. Results The current prototype constructed in aluminium and plastic can be sterilized for use in a class A environment. Firmly positioned by use of a suction cup, it allows one-handed operation with fixed or swivel needles and can be easily decontaminated. Conclusion User design collaboration processes are time consuming, but this study shows that such an implementation of a new device concept can meet the attributes required of staff in a typical radiopharmacy and thus be more readily implemented. References 1. Jagger J, et al. N Engl J Med 1988; 319:284–288. 2. Hung JC, et al. J Nucl Med Technol 1999; 27:290–293. A28 Benefit of monitoring GFR injection sites

H.M. Scicluna and L. Pike

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Addenbrooke’s Hospital, Cambridge University Hospitals NHS Foundation Trust, UK. Measurement of GFR is often requested as part of the patient care pathway. Optimal outcome for quality care requires accurate measurement of renal function using intravenous injection of 51Cr-EDTA. Extravasation of the isotope at the time of injection would invalidate the result [1] and could be detrimental to the patient if undetected. An audit was performed to assess whether monitoring of the injection site post-injection is an effective method for detecting extravasation that was not apparent during administration. A protocol for monitoring of the injection site post-administration of 51Cr-EDTA was formulated to support the procedure. Following the injection and withdrawal of intravenous access, the background activity is ascertained with a mini 900 scintillation monitor. Then the site of injection and contra-lateral site are monitored and recorded on the GFR datasheet and any difference is noted. In a period of 1 month 82 patients were monitored for the study, extravasation was detected in 3 cases. Two proceeded with the GFR study as levels of extravasation were estimated to be < 1% whilst 1 was abandoned. Clinically, this indicates a benefit of monitoring GFR injection sites. Monitoring also provides reassurance for the injector in some cases. Reference 1. Fleming J, Zivanovic M, Blake G, Burniston M, Cosgriff P. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Nucl Med Commun 2004; 25:759–769. ONCOLOGY SESSION Invited talk: DOPA–PET imaging in the diagnosis and assessment of phaeochromocytoma

Isla McKenzie Addenbrooke’s Hospital, Cambridge, UK. Invited talk: Perils and pitfalls of PET radiopharmaceutical production

Doug Abrams Cross Cancer Institute, Alberta, Canada. A29 Lymphatic drainage pathways of the breast and upper limb

C.K. Solankia, T.M. Bennett Brittonb, A. Wonga, N.G. Hartmana, S.E. Pinderc, E. Provenzanoc, D. Eatond, A.D. Purushothame and A.M. Petersf Departments of aNuclear Medicine, cHistopathology, and dMedical Physics, and bCambridge Breast Unit, Addenbrooke’s Hospital, Cambridge, UK, eDepartment of Academic Oncology, King’s College, London, UK, and fDepartment of Applied Physiology, Brighton and Sussex Medical School, UK. Purpose A proportion of patients undergoing sentinel lymph node biopsy (SLNB) for breast cancer develop lymphoedema of the upper limb. We hypothesize that

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1022 Nuclear Medicine Communications 2006, Vol 27 No 12

this may be due to shared lymphatic drainage between the breast and upper limb in this group of patients, and the aim of this study was to investigate the extent of the similarity between the axillary lymphatic drainage basins of the breast and upper limb. Methods Patients with breast cancer due to undergo axillary lymph node dissection were injected pre-operatively with 40 MBq of 99mTc-human immunoglobulin (HIG) intradermally into the peri-areolar region of the ipsilateral breast and 3 MBq of 111In-HIG intradermally into the dorsum of the ipsilateral hand, or vice versa. Axillary lymph nodes were removed, separated, and wellcounted for the presence of 99mTc and 111In. Results In all, 15 patients were entered into the study. In 13/15 patients the ‘hottest’ lymph node for 99mTc was different from the ‘hottest’ lymph node for 111In. In 2/15 patients the ‘hottest’ lymph node for 99mTc was also the ‘hottest’ lymph node for 111In suggesting a common drainage pathway for the ipsilateral breast and upper limb. Conclusion Whilst the majority of patients have different pathways of lymphatic drainage for the ipsilateral breast and upper limb, in some patients the drainage pathway is through a common sentinel lymph node. These patients may be at increased risk of developing lymphoedema when undergoing SLNB. A30 Correlation of tumour histology with tumour to background ratio in 99mTc-depreotide SPECT imaging of the lung

G. Ainslie, A. Bolster and J.B. Neilly Glasgow Royal Infirmary, UK. Aim To determine the effectiveness of a measure of tumour to background ratio (T:B) as a quantification tool in 99mTc-depreotide SPECT imaging. Method The raw data were reconstructed using iterative reconstruction followed by a post reconstruction low pass filter. Values of T:B were obtained from the transverse slices by defining a region of interest around the site of the tumour and around a region of normal lung uptake (background region: same slice and lung). In cases where the tumour site was too large, background regions were defined in the contra-lateral lung. Results Case notes of 24 patients undergoing NeoSPECT scanning were reviewed. To date, histology was available in 12 patients, 9 of whom (group 1) had nonsmall-cell lung cancer (NSCLC); and 3 (group 2) had benign histology (TB, organizing pneumonia, bronchiectasis). The mean T:B for group 1 was 2.14 while the mean T:B for group 2 was 2.09. Due to small numbers statistical comparison was not possible. Conclusion The results demonstrate that T:B ratios for NeoSPECT uptake in the primary NSCLC lesions is of the magnitude of approximately 2:1. These results are similar in magnitude to those lesions with benign histology.

A31 Longitudinal follow-up of patients with medullary thyroid cancer

R. Karugaba and S.E.M. Clarke Department of Nuclear Medicine, Guy’s and St Thomas’ NHS Foundation Trust, London, UK. Background Medullary thyroid cancer (MTC) is rare. Its behaviour is usually indolent and calcitonin levels may be elevated for a number of years before imaging identifies the site(s) of recurrence Aim To determine the value of combined planar and tomographic radionuclide imaging with 99mTc(V)-dimercaptosuccinic acid (99mTc(V)-DMSA) and 111In-octreotide in the longitudinal follow-up of patients with MTC. Method Scan results were reviewed in 19 patients (17 female, 2 male, mean age 68 years). Seventy combined studies were performed over a mean period of 7.5 years (range 1–18 years). Studies were repeated at 6-month intervals if calcitonin levels were rising rapidly and at 1year intervals if calcitonin levels were elevated but plateaued or rising slowly. Results Twenty-eight (37.3%) combined studies were abnormal and, similarly, 28 (37.3%) were negative. In 15 (20%) combined studies the 99mTc(V)-DMSA alone was positive and in 4 ( 5.3%) the 111In-octreotide alone was positive. In 11 99mTc(V)-DMSA and 7 111In-octreotide studies the tomographic study alone was positive. In 13 patients, the combined studies became positive during the course of follow-up. Conclusion Combined imaging with planar and tomographic 99mTc(V)-DMSA/ 111In-octreotide identified sites of recurrent disease in 62.7% of follow-up studies performed. Radionuclide imaging provides an effective method of monitoring MTC patients with biochemical evidence of relapse. A32 Experience with 90y-ibritumomab (zevalin) in the management of refractory non-hodgkin’s lymphoma

S. Hana, A.H. Iagarub, H.J. Zhub and M.L. Gorisb a Nuclear Medicine Department, Glasgow Royal Infirmary, UK, and bNuclear Medicine Department, Stanford University Medical Centre, California, USA. Introduction Radioimmunotherapy has been shown to be an effective treatment for refractory non-Hodgkin’s lymphoma (NHL). We present our experience with Zevalin, a radiolabelled monoclonal antibody targeting CD20, in the management of refractory NHL. Patients and methods The case records of 25 patients (19 male, 6 female; mean age 57 years, range 36–85 years), who had outpatient Zevalin therapy in 2000–2005 for refractory NHL were reviewed retrospectively. Types of NHL were follicular/mixed follicular (12), mantle cell (9), marginal zone/MALT (2), diffuse large cell (1) and immunoblastic (1). Patients were preloaded with Rituximab (250 mgm – 2) on days 0 and 7, followed by diagnostic 111In 185 MBq (5 mCi) on day 0 and ther-

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Abstract of the BNMS meeting, September 2006

apeutic 15 MBqkg – 1 (0.4 mCikg – 1) 90Y dose of labeled Zevalin on day 7 respectively. Results Overall response rate was 16/25 (64%). Nine of 25 patients (36%) achieved complete response and 7/25 patients (28%) had partial response. Four of 25 patients (16%) showed mixed response and 2/25 patients (8%) revealed progressive disease. In 3/25 patients (12%) there was no change in disease status. Grade 3 or 4 haematological toxicity developed in 11/25 (44%) of cases but it was transient and reversible. Conclusion Zevalin radioimmunotherapy is a reliable adjunctive treatment for NHL refractory to conventional treatment. SCIENTIFIC POSTERS ABSTRACTS P1 Tracer uptake in the sinuses on routine whole-body bone scans

T.A. Szyszko, J. Williams, J. Murrell and J. Frank The Hammersmith Hospitals NHS Trust, London, UK. Background and purpose To study the incidence of tracer uptake within the sinuses on whole-body bone scans. This uptake is seen incidentally on many bone scans and this study was undertaken to see whether this correlates with known sinusitis, allergies or hayfever. Methods Fifty bone scans carried out at Charing Cross and Hammersmith Hospitals were reviewed. The patients were asked to complete a questionnaire enquiring about coryza or sinusitis within the week preceding the scan and any history of allergy or hayfever. Patients with known head and neck disease were excluded from the study. Uptake within the sinuses was then assessed as either present or absent compared with uptake within skull vault. Results Overall, 11 of the 18 patients (61%) with infection, allergies or known sinusitis had uptake within their sinuses. Specifically, 75% of patients who had a coryza and 75% of the patients who reported sinusitis had uptake in the sinuses; but only 55%of patients with allergies or hayfever had uptake in the sinuses. Interestingly, of the 33 patients who denied any nasal or sinus symptomatology, 14 (42%) still had uptake within the sinuses on the bone scan. Conclusion There is no correlation between known sinusitis, allergies, hayfever or infection with uptake in the sinuses on bone scan. P2 Evaluation of dual-head compared with single-head lymphoscintigraphy quantification

M. Azimi and N. Gulliver St George’s Hospital, London, UK. Background Conventional lymphoscintigraphy is currently acquired anteriorly only. This means that for static views, counts acquired will be depth dependent. The depth of the injection site (space between toes) is superficial, but the depth of the pelvic inguinal and iliac nodes can be up to 10 cm.

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Aim To determine the difference between anterior only and geometrical mean of anterior/posterior views, and to assess the variation of their difference with body mass index (BMI). Method Eleven patients underwent static scans with 99m Tc-nanocolloid on a dual-headed camera. ROI analysis yielded total counts in both feet (at injection and 2 h later) and in both groins (after 2 h). Percentage uptake in groins and percentage retention in feet were calculated. Transmission images were obtained to measure node depth. Results Percentage differences between attenuation corrected anterior only and geometrical mean were plotted against the patient’s BMI. Statistical analysis of the two methods showed that the mean difference between attenuation corrected anterior only and geometrical mean was 14% in right and 12% in left. Conclusion There is a small systematic difference between anterior and anterior/posterior views due to depth of nodes. P3 Typographical accuracy of reports issued by the nuclear medicine department

M.J. Kaduthodil, E. Lorenz and V. Prakash Sheffield Teaching Hospitals NHS Trust, UK. Purpose An imaging report is permanent part of the patient’s medical record and, therefore, they are legally important documents. A typographical error can alter the accuracy of the report which may have detrimental impact on the patient’s management. Reporting staff have a duty to issue reports that are accurate. We looked at our typographical errors and compared it with accepted national standard [1]. Methods One hundred completed nuclear medicine scans were analysed retrospectively. This included 25 bone, 25 lung, 25 octreotide and 25 MIBI parathyroid scans. A copy of the authorized report for each investigation was checked against the handwritten original. Each error identified was graded 1–3 for severity and 0–3 for significance. Results 85% of reports were completely error free. 13% contained grade 1 errors; all grade 0 for significance. 2% contained higher grade errors which were deemed to be significant. This falls short of the locally agreed standard of 97% of reports should be issued error free [1]. CHANGES: The reports will be checked in a quiet environment free from interruptions. Re-audit is planned for 6 months. Reference 1. Clinical governance and revalidation: a practical guide for radiologists. London: The Royal College of Radiologists, 2000. P4 Improving lachrymal scintigraphy at the MRI

G. Ellula, M. Worrallb and R. Lawsona

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1024 Nuclear Medicine Communications 2006, Vol 27 No 12

a

Manchester Royal Infirmary, and bChristie Hospital NHS Trust, Manchester, UK.

Lachrymal scintigraphy is an established non-invasive imaging technique of the lachrymal drainage system with a very low radiation burden. Increased demand for lachrymal scintigraphy at the Manchester Royal Infirmary has prompted us to investigate ways of improving the imaging methodology. A lachrymal apparatus phantom was created using a Perspex facial mask (originally used for radiotherapy purposes) and small bore tubing. 99mTccolloid (1.5 MBq), comparable to that currently used for the scanning of patients, was administered to the phantom and 5-min images of both eyes were obtained using a pinhole collimator with 3, 4 and 6 mm inserts, LEGP and LEHR collimators. The 3 mm pinhole collimated image was the best resolved. The low sensitivity was deemed not to affect its clinical usefulness but administering a higher activity of 99mTc is feasible since the DRL is 4 MBq. If a lachrymal scan suggests a ‘pre-lachrymal sac’ blockage a single eye view close to the pinhole aperture could be useful in attempting to resolve the canalicular apparatus. In order to assess if this was possible, we performed further single eye views of the phantom using the 3 mm pinhole collimator. The results of this were encouraging, suggesting that this was achievable. P5 Validation of a platelet marker for use in noninvasive imaging studies in the rabbit

H. Saihkaya, C. Pagea, J. Ballingerb and K. Rickardsa a King’s College University of London, bGuy’s and St Thomas’ Hospital, London, UK. 123

I-labelled meta-iodobenzylguanidine (123I-MIBG) might be a useful marker for studying the role of platelets in rabbit models of inflammatory diseases. We have investigated the effect of serotonin and oxidative stress on MIBG uptake by rabbit platelets and determined whether MIBG uptake affects platelet chemotaxis. Platelet-rich plasma (PRP) separated from citrated rabbit blood was either co-incubated with 123I-MIBG and serotonin (1  10 – 8 to 1  10 – 4 M) for 4 h or incubated with MIBG for 4 h before co-incubation with serotonin for 2 h or incubated with 100 mM menadione, washed with cold PBS and then incubated with 123I-MIBG for 4 h. Following centrifugation, radioactivity was measured in the platelet pellets. N-formyl-methionyl-leucyl-phenylalanine (fMLP: 1  10 – 8 to 1  10 – 4 M) was loaded in the bottom wells of a chemotaxis plate. Following 4 h incubation with 123I-MIBG, PRP was placed on top of a 3 mm filter and, after 70 min incubation, migrated cells in the lower chamber were counted. Only high concentrations of serotonin (1  10 – 5 and 1  10 – 4 M) reduced MIBG uptake and retention whilst menadione showed no significant effect on platelet MIBG uptake suggesting that alterations in plasma serotonin concentration and

oxidative stress are unlikely to affect MIBG uptake in vivo. MIBG had no significant effect on fMLP-induced platelet chemotaxis so should not affect platelet recruitment to sites of inflammation. P6 Limitations of emission based methods for attenuation correction in myocardial perfusion imaging

J. Maraisa, D.N. Taylora, A.R. Denmana, C.A. Phillipsb and P.D. Pictonb a Department of Medical Physics, Northampton General Hospital NHS Trust, and bUniversity of Northampton, UK. A source of degradation in SPECT imaging is photon attenuation. The determination of an accurate, patientspecific attenuation map, which represents the spatial distribution of linear attenuation coefficients, is fundamental to performing attenuation compensation. Methods for generating the attenuation map either use a transmission source or the emission data itself. Emission-based methods avoid the major obstacles in transmission based methods. Such methods include calculated methods, consistency conditions criteria and computer simulations. Limitations of these emissionbased methods are the spillover of activity into the myocardium and the complexity of the correction algorithms. To overcome this, Stodilka et al. [1] suggested an attenuation correction for brain SPECT using attenuation distributions inferred from a head atlas. The same principles could be applied to create a cardiac atlas. We have proposed the application of the same principles to create a cardiac atlas. The problem in the thorax is more complex than in the head. In this poster we examine the problems inherent in such an approach and produce results to illustrate potential solutions. It is concluded that registration of thorax cross section is possible prior to iterative reconstruction but further work is required to produce a universally applicable clinically useful correction. Reference 1. Stodilka RZ, et al. J Nucl Med 2000; 41:1569–1578. P7 MUGA scanning: Do you know how your camera works?

S. Redgate and A. Harris Sheffield Teaching Hospitals NHS Foundation Trust, UK. Purpose To assess gating on our cameras prior to the introduction of radionuclide ventriculography (MUGA) scanning for herceptin monitoring. Method Using an ECG simulator, gated 3 min images were acquired on each camera. The heart rate was changed from 65 bpm to 43 bpm, or 43 bpm to 65 bpm at 1 min, in both time and phase mode. Gated images of a point source were also acquired. Results These are listed in Table 1.

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Abstract of the BNMS meeting, September 2006

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Table 1 Camera

Operation in time mode

Operation in phase mode

1

As expected

2

As expected

The R–R interval at the centre of the Updated the as an average of the beat acceptance window was used previous n accepted beats. to calculate the frame duration, not the instantaneous R–R interval. As expected Not updated

3

As expected

As expected

4 5

Frame durations were 9–19% higher As expected than expected and consequently As expected low counts or zero counts were registered in the end frames.

Conclusion The cameras all worked differently and not always as described in the literature and manufacturer’s documentation, highlighting the importance of testing camera gating. P8 Audit of administered doses and assessment of residual activities remaining in syringes in myocardial perfusion imaging with 99mTc-tetrofosmin

R. Meades, J. Nadarajah, Z. Win and A. Al-Nahhas Department of Nuclear Medicine and Imaging, Hammersmith Hospital, London, UK. Background Administration of the correct activity in myocardial perfusion imaging (MPI) is extremely important in achieving good quality images with the optimum dose being based on the patient weight adjusted ARSAC DRL. However, it is recognized that Myoview (99mTc-tetrofosmin) may stick to the tubing of cannulae and extension tubing used in its administration leading to administered doses being less than optimal [1]. A previous audit resulted in a change to our protocol, including the drawing up an extra 100 MBq for each patient dose to compensate for the residual activity. This audit was performed to complete the audit cycle and examine the effect of the change in protocol. Method Residual activities were measured for 56 patients and percentage differences of the administered doses from the desired doses were calculated. Residual doses were found to be 102 ± 27 MBq (mean ± standard deviation) which is comparable to those in the previous audit. The percentage difference between administered dose and desired dose was found to have reduced from – 6% ( ± 9%) to 1% ( ± 10%) (means ± standard deviations). Conclusion This audit has shown that drawing up an extra 100 MBq for each patient dose has resulted in an increased accuracy of administered doses in MPI. Reference 1. Gunasekera RD, Notghi A, Mostafa AB, Harding LK. Adsorption of radiopharmaceuticals to syringes leads to lower administered activity than intended. Nucl Med Commun 2001; 22:493–497.

Beats acceptance window

Normalization of end frames No

No

Updated as a weighted average of the Yes previous n beats, accepted or not. Not updated Occasionally Not updated Occasionally

P9 Potential pitfalls in PET–CT imaging

C. Turner, K. Tung, J. Smart, V. Batty, P. Kemp and S. Harden Southampton General Hospital, UK. Positron emission tomography (PET)–CT imaging is an increasingly important technique for the diagnosis and staging of a wide range of neoplasms. Although still a relatively new medical imaging tool, it is rapidly becoming an integral component in the management of oncological diseases. PET – CT imaging is based on the principle of detecting coincident annihilation photons released during the decay of the glucose analogue [18F]fluorodeoxyglucose (FDG). FDG accumulates in metabolically active tumour cells more than in normal tissue, but it is not a tumour-specific substance. Increased FDG uptake may be physiological or occur in a range of benign pathological causes including healing bone, regional response to infection, degenerative change, post-operative, active granulomatous disease, aseptic inflammatory response and response to chemoradiotherapy. This poster demonstrates a variety of such potential diagnostic pitfalls with emphasis on the PET– CT imaging appearances. Recognition of these benign entities and correlation with clinical findings allows improved diagnostic accuracy to enable optimal patient management. P10 The effect of time interval between thyroidectomy and radioiodine ablation therapy on extra-thyroidal involvement in differentiated thyroid carcinoma

K.L. Dixon and L. Jansson Poole Hospital NHS Trust, UK. In patients with differentiated thyroid carcinoma, extrathyroidal recurrence in the time between thyroidectomy and radioiodine ablation therapy (RAT) has been observed [1]. This study sets out to determine the effect on patient prognosis. Retrospective analysis was performed on 26 patients referred to Poole Hospital for RAT following thyroidectomy between 2002 and 2006, who underwent both an immediately pre- or post-ablation whole-body scan (WBS) and a follow-up WBS at 4–6 months. The immediate WBS was used to indicate extra-

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1026 Nuclear Medicine Communications 2006, Vol 27 No 12

thyroidal involvement, and the follow-up WBS to suggest short-term prognosis. The average time between thyroidectomy and RAT was 80 ± 5 days. Extra-thyroidal involvement was indicated in 8 patients who had an average time between thyroidectomy and RAT of 89 ± 5 days. Of these patients, 7 (88%) were disease clear on their follow-up WBS. The 18 patients who showed only thyroidal involvement had an average time between thyroidectomy and RAT of 76 ± 4 days, and of these patients 11 (61%) were disease clear on their follow-up WBS. This study indicates that the incidence of extrathyroidal involvement increases with time between thyroidectomy and RAT, but also that RAT may be more effective on these patients. Reference 1. Kumar A, Bal CS. Indian J Pediatr 2003; 70:707–713. P11 A dynamic phantom for validation of renogram analysis software in nuclear medicine

H.J. Cowarda and L. Livieratosb a King’s College Hospital, London and bGuy’s and St Thomas’ Hospital, London, UK. Validation of clinical analysis software is a traditionally difficult area, as clinical data with known values are not possible to obtain and data from physical phantoms are often over-simplistic. The use of the Zubal phantom, a segmented CT data set, was investigated to create activity distributions in the human body to mimic the uptake of the radiopharmaceutical 99mTc-MAG3 after intravenous administration. Activity distributions were assigned to the human model using custom IDL routines, guided by bio-kinetics of a 3-exponential model of total body retention, and a kidney transit time of 4 min, as accepted by ICRP [2]. The effects of photon attenuation, noise and gamma camera resolution were simulated. Time–activity curves and renogram analysis software were used to successfully validate the phantom. Results showed that dynamic renal studies could be successfully simulated and used for validation purposes of analysis software. One major advantage to this phantom, compared to the ICRP dosimetric phantom, is that all tissue compartments are of realistic shape and size. Hence results using this phantom may be more accurate both for image validation purposes and dosimetric applications. Future use of this dynamic phantom may include the validation of new software, or comparison of alternative software. References 1. Zubal IG. Med Phys 1994; 21:299. 2. Stabin M. J Nucl Med 1992; 33:33–40. P12 An intercomparison of three techniques for determining relative function on DMSA scintigraphy: Do the differences matter?

J.M. Anton, A.H. Elmegadmi and N.W. Garvie

St Bartholomew’s and The Royal London Hospitals, UK. Introduction In DMSA scintigraphy, relative function can be assessed by geometric mean (GM), arithmetic mean (AM) or posterior count (PC) ratios. Does it matter which technique is used? What is the correlation between the individual techniques, and how frequently would any variance render a normal result (50% ± 5%) abnormal, or vice versa? Methodology One hundred and three adult DMSA studies were analysed, using GM, AM and PC ratios. Renal outlines were defined manually, and background subtraction was performed. Results The values were plotted to obtain the lines of regression. The best correlation was obtained when GM was plotted against AM (R2 = 0.989). GM against PC, and AM against PC, yielded correlation coefficients of 0.919 and 0.947, respectively. If the GM result is viewed as the ‘standard’, use of the AM or PC techniques would have rendered a normal result abnormal, in 2% and 8% of cases. Conversely, an abnormal result would have been deemed normal, in 4% and 6% of cases, respectively. Conclusion There is good correlation between the GM and AM techniques, and, only rarely did any minor variance result in abnormal normal reassignment. The PC technique does not appear as reliable. P13 An audit of glomerular filtration rate calculation methodology across three hospital sites and its impact on chemotherapy dose prescription in oncology

D. Phillipsa, M.A. Richardsonb, R.G. Ardleya, S.T. Chandlerc, D. Wrightc and S. Gokulb Regional Medical Physics Department, aThe University Hospital of Hartlepool, bThe James Cook University Hospital, Middlesbrough, and cDarlington Memorial Hospital, UK. On Teesside, three nuclear medicine departments provide Glomerular filtration rate (GFR) measurements for chemotherapy dosage calculation (via the Calvert formula) for patients referred to the Middlesbrough Cancer Centre and its satellite clinics. The current GFR calculation software, NIRAS (RMPD), uses the Chantler method of slope–intercept correction and the Du Bois and Du Bois (1916) formula for body surface area. In response to the publication of BNMS GFR guidelines [1] and the need to replace dated software the Excel Spreadsheet for calculation of GFR (‘The Cosgriff Spreadsheet’) [2] was purchased by all three centres. This software utilizes Brochner-Mortensen (1972) first exponential correction and the Haycock (1978) body surface area correction in accordance with the BNMS guidelines. For a 3-month period all GFR referrals for chemotherapy prescribing were calculated using the existing NIRAS software and Cosgriff ’s Spreadsheet (n = 134). This allowed a direct comparison of the two methods on measured GFR and their subsequent impact on prescribed chemotherapy dose for this patient

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Abstract of the BNMS meeting, September 2006

population. The mean percentage difference between the NIRAS and Cosgriff methods was 0.32 ± 0.45 (mean ± SE) for absolute GFR but with a range of over 33% ( – 23.45% to 9.68%). 90% of the audited patient group would receive a ± 5% difference in chemotherapy dose, depending on their GFR. This is not regarded as clinically significant. References 1. Fleming JS, Zivanovic MA, Blake GM, Burniston M, Cosgriff PS. Guidelines for the measurement of glomerular filtration rate using plasma sampling. Available at www.bnms.org, 2004. 2. Cosgriff PS. www.nuclearmedicine.org.uk (version 4.6). P14 Investigation into the effect of haemolysis on blood samples taken for glomerular filtration rate assessments

J.M. Price, E. Gawthorpe, M.B. Barnfield and M.T. Burniston Leeds Teaching Hospitals NHS Trust, Leeds, UK. Background and aim BNMS guidelines [1] pertaining to blood samples for GFR assessments state, ‘A small degree of haemolysis occurs occasionally ythis should not unduly affect the GFR result, but marked haemolysis will invalidate the measurement.’ The project aim was to investigate to what extent haemolysis would affect the GFR result and give an indication of when a blood sample should be rejected. Method 99mTc-DTPA was added to a volume of whole blood to give a tracer concentration similar to that found in a patient undergoing GFR assessment. This was separated into batches, with each batch placed in an ultrasound bath and exposed to differing durations of ultrasound to induce various haemolysis levels. These samples and a control were then assayed in a multi-well counter and the effect of haemolysed samples on subsequent GFR measurements simulated. Results and conclusion Increased ultrasound exposure resulted in haemolysis leading to a reduced plasma count. The average count from each batch was calculated and a 4% range found between non-haemolysed and maximally haemolysed samples. Even significant degrees of haemolysis, greater than normally seen in clinical practice were shown to introduce only a small inaccuracy in the plasma count and GFR. Sample haemolysis should not necessitate repeat assessments. Reference 1. Fleming JS, et al. Nucl Med Commun 2004; 25:759–769. P15 V/Q scanning: How, when and why? A consultant survey

S. Ping-Lim and P.D. Strouhal The Royal Wolverhampton Hospitals NHS Trust, UK.

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Purpose Despite BTS guidelines for imaging of suspected acute PE, local confusion seemed to exist regarding referral criteria, how positive V/Q scans compare to positive CTPA, what some PIOPED reporting terminology means and what action to take following inconclusive results. Method One hundred and forty-five consultants were emailed questionnaires, with a 6-week deadline for replies. Two bottles of wine were offered as an incentive to a randomly selected respondent if over half replied. Questions included frequency of V/Q requesting; if preimaging chest X-rays and D-dimer assays were requested; if formal pre-test probabilities were assessed. Consultants’ views were asked about pre-test probability assessment, how V/Q and CTPA scans compare and their management plans for given combinations of pre-test probability and V/Q scan probability of PE; and also their understanding of ‘indeterminate’ results. Results Only 25 completed replies were received. 84% infrequently request V/Q scans, but nearly all obtain chest X-rays and D-dimers first. Only 8% formally assess pre-test probability of PE; 96% know to proceed to CTPA for ‘indeterminate’ V/Q scans but 44% believe this result differs to an ‘intermediate’ scan. 96% respondents thought a negative CTPA much more reliable than negative V/Q scan. Conclusion PIOPED terminology is poorly understood, V/Q scanning remains underrated and our consultants do not like wine. Reference 1. The British Thoracic Society Standards of Care Committee, Pulmonary Embolism Guideline Development Group. BTS guidelines for the management of suspected acute pulmonary embolism. Thorax 2003; 48:470–484. P16 Healthy passive cigarette smokers have increased pulmonary alveolar permeability

C. Beadsmoore, H. Cheow, K. Szczepura, P. Ruparelia and A.M. Peters Addenbrooke’s Hospital, Cambridge, UK. Background and aim To determine whether passive smoking has a measurable effect on lung function in otherwise healthy subjects? There is current interest concerning passive smoking but no objective evidence showing that it has any impact on lung function. Methods The pulmonary clearance rate of 99mTc-DTPA was measured in 21 healthy volunteers after inhalation as a radio-aerosol and compared between healthy cigarette smokers, passive smokers and non-smokers. All volunteers had normal lung function. Results Clearance half-times in healthy passive smokers (n = 5) were longer than in healthy smokers (n = 6) but clearly shorter compared with healthy non-smokers

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1028 Nuclear Medicine Communications 2006, Vol 27 No 12

(n = 10) with respective mean values of 45.2 (SD 8.3), 24.3 (8.6) and 80.3 (20) min. Conclusion Passive smoking has a functional impact on the lung blood/gas barrier. P17 The impact of British Thoracic Society guidelines on referral patterns for V/Q scintigraphy in patients with suspected pulmonary embolism

T.D. Barwick and N.W. Garvie Nuclear Medicine Department, Royal London Hospital, UK. Background BTS guidelines (2003) recommend CTPA as the primary imaging strategy for PE, although V/Q scintigraphy can be used initially, provided certain criteria, including normal chest radiography, are fulfilled. Implementation of these guidelines should therefore diminish the demand for V/Q scintigraphy, and specifically reduce the proportion of indeterminate probability (IP) scans, with a concomitant rise in CTPA studies (particularly following IP scans). Are these effects observed in clinical practice? Methods V/Q scans were reviewed over two 12 month periods, before and after publication of the BTS guidelines. The V/Q scans were totalled, and subdivided according to probability (modified PIOPED criteria). CTPA rates following IP scans were documented. Results Total number of V/Q scans reduced by 19.5% from 503 (2002) to 405 (2005), with a corresponding increase in CTPA examinations. The proportions of high probability, IP, and normal scans were 12.5%, 6.6% and 80.9% in 2002, and 12.1%, 6.9% and 81% in 2005, respectively. The percentage of IP patients receiving subsequent CTPA was 51.5% in 2002, and 60.6% in 2005. Conclusions Since 2003, there has been a significant but non-selective reduction in all probability types of V/Q scans. There has been a rise in the number of patients with IP results being referred for CTPA. P18 Perfusion:ventilation count rates in gas lung V/Q scanning

99m

Tc-techne-

C. Findlay, S. Allan, J. Morrison and A. Nicol Nuclear Medicine Department, Southern General Hospital, Glasgow, UK. Background Lung V/Q scans may be performed using Tc-technegas then 99mTc-MAA administrations. As 99m Tc is used for both imaging procedures, a perfusion count rate considerably greater than the ventilation count rate is required. Local DRLs for V/Q are 20 MBq 99mTctechnegas and 100 MBq 99mTc-MAA. However, the activity administered for ventilation will vary due to the nature of the administration procedure. Aims To investigate the perfusion:ventilation count rates obtained in clinical practice and to determine the influence of BMI, perfusion count rate (cpsQ) and ventilation count rate (cpsV) on this ratio (cpsQ/cpsV). Methods A retrospective analysis was performed on 50 consecutive patients. BMI, cpsQ, cpsV and cpsQ/cpsV were 99m

recorded. Correlation analysis was performed to investigate the relationships between cpsQ/cpsV and BMI, cpsQ, cpsV. Results Average cpsQ/cpsV was 4.9 (SD = 1.3) with a minimum of 2.3. 14% of patients had a ratio < 4.0. cpsQ/ cpsV was found to be correlated with BMI, cpsQ, cpsV (Pearson coefficient, P < 0.001). Conclusions Variation in cpsQ/cpsV was observed in clinical practice. This ratio was dependent on BMI, cpsQ and cpsV. Local practice includes an assessment of cpsQ/ cpsV for individual patients. P19 Defining a low-dose CT protocol for SPECT/CT localization scans on the Philips Precedence SPECT/CT camera

K.L. Adamsona, B.R. Walmsleyb and D.J. Gallagherb Department of aNuclear Medicine and bRadiation Safety, Guy’s and St Thomas’ NHS Foundation Trust, London UK. Aim To determine a low-dose CT protocol that could be used for SPECT/CT image localization scans on the Philips Precedence SPECT/CT camera. Methods A Rando body phantom was scanned using the default Abdo/Pelvis helical CT protocol (100 mAs, 120 kV, collimation 16  1.5, slice width 3 mm, rotation time 0.5 s). A further six scans were performed varying the current–time product (25–200 mAs), slice width (1.5– 3 mm), collimation (16  1.5 to 16  0.75) and rotation time (0.5–0.75 s). The pitch was kept constant. For each scan, the same three transaxial slices (lung, spine and pelvis) were visually inspected and the CT number and relative noise measured in two representative regions. The dose for each scan was recorded. Results Visually, there was no loss of information from any scan. The relative noise varied as 1=pcffi (where c is the current–time product, in mAs), as expected. Two lowdose clinical protocols were proposed: a standard localization protocol (120 kV, 50 mAs, slice width 3 mm, 0.5 s) and a second protocol for multi-planar reconstruction reviewing, which incorporated a thinner reconstructed slice width (1.5 mm). Conclusion The Rando phantom lacks clinical detail but has been beneficial in defining low-dose localization CT protocols. These protocols are now in clinical use and may be further optimized following review by radiologists and nuclear medicine consultants. P20 Introduction of SPECT/CT fusion imaging in a district general hospital

I. Hagana, A. Talbota, F. Koeberga, M. Evansb and D. Dunlopa Departments of aNuclear Medicine and bMedical Physics, Royal United Hospital, Bath, UK. The object of this presentation is to describe our initial experience of SPECT/CT fusion imaging in a busy district general hospital with an emphasis on some of the multidisciplinary challenges faced and opportunities

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Abstract of the BNMS meeting, September 2006

created by the introduction of this technique. Installation of a hybrid SPECT/CT scanner (GE Infinia Hawkeye) in our department was completed in March 2006. Although purchased largely to allow improved attenuation correction for myocardial studies, its arrival has allowed us to begin performing SPECT/CT fusion imaging in selected cases. Encouraged by the work of others [1,2] we have found it to be particularly useful in the assessment of endocrine tumours such as phaeochromocytomas and as a problem-solving tool in some skeletal scintigraphic studies, and we present some illustrative cases. In order to take advantage of the undoubted benefits of this technology, our physicists, technicians and radiologists have all had to adapt to the multidisciplinary implications of its introduction. These include radiation protection issues, quality assurance, CT training for nuclear medicine technicians, the need for flexibility in scheduling work lists, the creation of new imaging protocols, and the need for radiologist supervision of studies. References 1. Fielding PA, et al. Nucl Med Commun 2006; 27:299. 2. Reid RH, et al. Nucl Med Commun 2006; 27:287. P21 Evaluation of NED and AEGIS personal extremity dosimeters

B.L. Gawthorpe and A. Burns Leeds Teaching Hospitals NHS Trust, UK. Background An advanced extremity gamma system (AEGIS ED2) and a nuclear educational device (NED) were lent to the trust for evaluation. The two were compared and evaluated for use within the nuclear medicine department. Methods Tests were performed to assess the energy response, angular sensitivity, linearity and accuracy of each device. The devices were compared in terms of their ease of use and their suitability for a range of planned applications including finger monitoring during QC, treatment and dispensing, and environmental monitoring in patient and staff areas. Both devices were found to be useful for real-time monitoring of extremity dose. Results The AEGIS proved to be a more versatile, reliable and user-friendly device but suffered from very poor response at some angles where sensitivity was reduced to 19% of the maximum sensitivity. The NED showed an exaggerated response to lower energy photons. When compared to other dosimeters the relative response from the NED was three times higher for 99mTc (140 keV) than for 131I (predominantly 360 keV). This effect was further increased when scattering material, such as lead shields, was present. The NED was also difficult to set up correctly and gave anomalous readings due to frequent user errors. P22 Assessment of staff extremity doses

L. Pike

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Addenbrooke’s Hospital, Cambridge, UK. Background In order to comply with the ALARP principle it is important to monitor and review work practices involving radiation [1]. One such practice is the administration of therapeutic doses of 131I. At Addenbrooke’s Hospital outpatient therapies for thyrotoxicosis involve the dispensing and administration of 400, 600 or 800 MBq of 131I in liquid form. In order to ensure procedures are effective and doses are being kept ALARP, the extremity doses for the three physicists who carry out the therapies were measured over the course of the day. Method An AEGIS ED2 dosimeter with a silicon detector was used to log the doses. The detector was placed on the tip of the index finger of the dominant hand and was worn throughout vial preparation, dispensing, administration and waste disposal. Times of activities were recorded to differentiate doses received for certain procedures. Results The average extremity dose for each physicist was 0.07, 0.11 and 0.07 mSvMBq – 1 dispensed. Preparation of 131I stock vials was found to contribute greatest to the total dose and typically increased it by a factor of 2. However, even in the worst-case scenario the annual extremity dose would not exceed 1/10th of the annual dose limit. Reference 1. Ionising Radiation Regulations 1999 (SI 1999 No. 3232). London: HMSO. P23 Are you measuring your true skin dose when handling FDG?

R.J. Mortona, D. Murraya, A. Zonoozia, M. Pryorb, G. Planta, L. Bellamya and J.R.W. Halla a Guildford Diagnostic Imaging, Lodestone Patient Care, and b Department of Radiation Protection, Royal Surrey County Hospital, UK. Background The Ionising Radiation Regulations 1999 state that the annual dose limit to the skin is 500 mSv, averaged over 1 cm2 (classification: 150 mSvy – 1). Hence it is essential to measure the maximum skin dose when handling 18F-FDG. Aim To map the dose received by the hands when handling 18F-FDG. Method Thirty thermoluminescence dosimeters were used to map the dose to the front and back of the hands while the operator was manipulating and injecting (with appropriate tungsten shielding) FDG under routine conditions. Results (1) Dominant hand. The dose received at the base of the index and ring fingers was 1=2 and 1=6, respectively, of that at the tip of the index finger. The dose received to the back of the index finger was half that of the palm side while both sides of the ring finger received equal dose. (2) Non-dominant hand. All fingers received up to 3 times

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1030 Nuclear Medicine Communications 2006, Vol 27 No 12

the doses of the corresponding dominant hand except for the index finger which received 1=3 of the dose of the dominant hand. Conclusions Measuring the dose at the base of the finger could grossly under estimate the annual skin dose. This could potentially change the classification of staff working with FDG. TECHNOLOGIST POSTER ABSTRACTS P24 Timing and timely scanning for GI bleeds

S. Doyle, N. Ridley, L. Jackson and J. Cook The Great Western Hospital, Swindon, UK. Background and purpose All patients referred with GI bleed of unknown origin have immediate imaging (for 1 h post-injection) and subsequent imaging performed up to 24 h post-injection, unless the immediate scan is reported as positive. The standard protocol is: (1) all patients are scanned on the day of request; (2) scanned at 60 min post injection; and (3) delayed imaging is performed to detect subsequent bleeding, unless the initial scan is reported as positive. The purpose of the study was to review a change in GI bleed scanning protocol. Materials and methods A 2-year retrospective audit of nuclear medicine GI bleed scans was performed. Results were obtained from 12 patients imaged using in-vivo 99m Tc-labelled red blood cells (400 MBq). Results (1) 100% of scans were performed on the day of request. (2) 100% of scans were performed immediately post-injection. (3) Three of the 12 (25%) of immediate scans were positive. (a) Nine of the 12 (75%) of immediate scans were negative. (b) All 9 patients (100%) with negative immediate scans were scanned up to 24 h later. (c) Three of the 9 (33%) were negative after delayed imaging. (d) Six of the 9 (66%) were reported as positive for GI bleed on the delayed scan. Conclusion The retrospective audit showed the standard protocol for GI bleed scans was adhered to in 100% of patients scanned. The results demonstrate the benefits of delayed imaging in detection of GI bleeding. P25 Usefulness of late imaging of sentinel lymph nodes in breast cancer

A. Fullbrook, L. Wright, J. Ward, R.J. Morton and J.R.W. Hall Department of Nuclear Medicine, Frimley Park Hospital, UK. Background At Frimley Park Hospital sentinel lymph node (SLN) images are acquired the day before surgery. 99m Tc-nanocolloid (40 MBq) is injected periareolar and anterior, lateral and oblique images acquired 3 h postinjection. Since July 2005, 185 SLN studies have been performed of which 9 ( < 5%) have shown no nodes at 3 h. Aim To look at late imaging of patients who show no tracer uptake at 3 h. Method Patients who show no tracer uptake at 3 h were imaged 6 h post-injection. A 10 min oblique view of the

relevant breast was acquired using a LEHR collimator and 128  128 matrix. Results Of the 9 negative node images at 3 h, 4 (44%) patients showed uptake in the sentinel node at 6 h post injection. Two patients had low activity nodes found at surgery using a gamma probe. One patient had 2 nodes seen with blue dye only. Histology showed tumour infiltration. One patient had no detectable nodes (including blue dye). One patient with a negative scan had ductal carcinoma in situ and therefore no nodes were removed at surgery. Conclusion Late imaging can improve the SLN localization if the 3-h image is negative. P26 Paediatric DMSA imaging: Optimal collimator choice

J. Davison, S.J. McGranaghan, E. McCauley and A. Mackie Regional Medical Physics Department, University Hospital of North Durham, UK. The aim of the study was to determine which gamma camera collimator, low energy all purpose (LEAP) or lowenergy, ultra-high resolution (LEUHR), produce DMSA images maximizing image quality and reporter confidence, and to confirm which of the recommendations of the European guidelines for paediatric DMSA imaging [1] is most important; choice of collimators or total counts acquired. Thirty-five patients were imaged with both LEAP and LEUHR collimators and reported by two blinded observers. Images were scored on a scale of 1 to 5, from normal through equivocal to abnormal, and also assessed for quality, including the ability to see the kidney’s internal architecture. Results were analysed using proportional statistics. The LEUHR images showed significantly improved clarity of renal outline, compared to those acquired using the LEAP collimators, but with significantly poorer count statistics. The clinical outcome was identical for 31 out of 35 (89%) patients initially. On review, a consensus score was reached on a further 3 patients. One patient demonstrated an abnormality only on the LEAP image, which was not visualized on the LEUHR image. Choice of collimator will most likely be selected by the reporter’s own preference, either for renal clarity or for image counts. Reference 1. Paediatric Committee of The European Association of Nuclear Medicine. Guidelines of 99mTc-DMSA scintigraphy in children. Available at www.eanm.org. P27 Does an increase in administered activity produce a significant improvement in the image quality in relation to the myocardial SPECT scan?

H. Spence Diana, Princess of Wales Hospital, Grimsby, UK.

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Abstract of the BNMS meeting, September 2006

Purpose To investigate the relationship between image quality and administered activity at varying tissue equivalent thicknesses. Methods Images were acquired at 5 differing administered activities and using 5 varying tissue equivalent thicknesses around the phantom. Images were qualitatively interpreted as a myocardial perfusion imaging patient according to a 4-point scale by 3 experienced observers. Weighted kappa statistics were used for intraand inter-observer agreement. Contrast was used as a quantitative measure of image quality to investigate the relationship between image quality, administered activity and the size of the phantom using tissue equivalent material. Results Using weighted kappa statistics intra-observer agreement was found to be moderate (k = 0.49), whilst inter-observer agreement ranged from poor to moderate (k = 0.29–0.50). A positive relationship was established between contrast and administered activity (r = 0.99, P < 0.001). A negative relationship was found between

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contrast and an increase in size of the phantom (r = 0.97, P < 0.001). However, only a moderate relationship was found in the patient data when comparing thoracic circumference to image contrast (r = 0.62, P < 0.001). Conclusion A relationship exists between image quality, phantom size and administered activity. Further study is required for the more complex patient model. CASE REPORT POSTERS P28 Serial cerebral perfusion SPECT images using 99m Tc-HMPAO in post-operative paediatric moyamoya disease

B.F. Lee and N.T. Chiu National Cheng-Kung University, Tainan, Taiwan. P29 Disseminated cryptococcal infection: detection by 18 F-FDG-PET

N. Seshadri and K. Balan Department of Nuclear Medicine, Addenbrooke’s Hospital, Cambridge, UK.

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Nuclear Medicine Communications 2006, Vol 27

i

Index to authors Volume 27 2006 Abe, K, 893 Akın, H, 387 Akman, S, 715 Akoglu, E, 191 Alavi, A, 11, 231 Al-Housni, MB, 113 Ali, N, 165 Al-Nahhas, A, 477, 547, 685, 709 Andreadis, C, 11 Anjos, DA, 395 Antke, C, 267 Antsaklis, A, 911 Aoki, Y, 143 Aras, G, 261, 377, 873 Arıcı, M, 61 Arumugam, P, 843 Asada, T, 151 Asada, Y, 987 Asakura, H, 887 Ayala, A, 31 Aydın, F, 715 Aygit, CA, 91 Aytemir, K, 61 Babyn, PS, 961 Bach-Gansmo, T, 185 Baden, M, 247 Baete, K, 765 Bailey, DL, 975 Bailey, EA, 975 Ballinger, JR, 543 Banerjee, S, 271, 661 Barai, S, 425 Barbaro, D, 627 Bardie`s, M, 559 Barros, FB, 957 Basu, S, 413 Bauchet, L, 137 Bautz, W, 521 Bax, JJ, 317 Bayrakdar, R, 179 Baysal, EY, 715 Beattie, LA, 197 Beiki, D, 567 Benlier, E, 91 Bergh, J, 347 Berkarda, S, 447 Bernard, X, 815 Berthold, F, 171 Besser, GM, 165 Biggi, A, 213 Bizais, Y, 431, 559 Blackwood, KJ, 807 Bloch, W, 171 Bockisch, A, 669 Bodet-Milin, C, 431 Bol, A, 815 Boni, G, 627 Bourguet, P, 363 Brandau, W, 669 Brenner, W, 739 Brink, I, 17 Britton, KE, 165

Brockhuis, B, 943 Brogsitter, C, 853, 951 Brown, SM, 611 Brusa, L, 381 Buchert, R, 739 Bui, F, 583 Burchert, W, 853, 951 Burri, H, 105 Bury, T, 969 Buscombe, JR, 589 Butler, T, 937 Buursma, RA, 25 Caglar, M, 61 Camargo, EE, 395, 957 Canizales, AL, 165 Capriotti, G, 757 Caraccio, N, 439 Carlier, T, 431, 559 Carrasquillo, JA, 31 Carrerea, D, 507 Carrio, I, 551 Cascini, LG, 213 Cavarec, M-B, 559 Cawley, M, 937 Cecchin, D, 583 C¸elikel, C¸, 387 C¸ermik, TF, 447 Chakraborty, S, 661 Chandra, P, 231 Chang, C-J, 779 Chatterton, BE, 695 Chawla, S, 795 Cheetham, AM, 113 Chen, CC, 31 Chen, G, 603 Chen, J, 551 Chen, S, 603 Chen, YH, 859 Cheng, CY, 859 Cheow, HK, 101 Cherng, S-C, 859 Chevalier, J, 137 Chew, SL, 165 Chianelli, M, 757 Chiang, S, 11 Choe, YS, 919 Choi, JY, 919 Choi, Y, 919 Chou, Y-H, 779 Chung, HW, 919 Chung, J-K, 773 Chuttani, K, 619 Clausen, M, 739 Cobbe, SM, 927 Comte, F, 137 Conant, E, 231 Concia, E, 645 Consoli, V, 645 Conti, PS, 795 Cooper, M, 455 Cooper, MS, 543 Coppens, A, 815

Corhay, J-L, 969 Corstens, FHM, 515 Costa, FF, 957 Coubes, P, 137 Couturier, O, 431, 559 Csiki, Z, 501 Culell, P, 785 Currie, GM, 57 Czerniecki, B, 231 Damia, S, 231 Danieli, R, 381 Dardano, A, 439 Das, T, 661 de Klerk, JMH, 223 de Murphy, AC, 371 De Palma, D, 213 De Sadeleer, C, 255 De Sutter, J, 529 de Vries, EFJ, 25 de Vries, J, 791 De Winter, O, 529 Debets, JMH, 677 Debnath, MC, 271 Dede, F, 191, 387 DeJongste, MJL, 791 Del Corral, P, 31 Demakopoulos, N, 119 Dempsey, MF, 611 Denizot, B, 363 Derejko, M, 943 Dessogne, P, 5 Deulofeu, P, 785 Di Martino, F, 439 Dietlein, M, 171, 353 Dou, K-F, 333 Drake, WM, 165 Dresel, S, 733 Dubaniewicz, M, 943 Duffin, R, 997 Duman, GD, 387 Dupont, P, 765 Dustan, K, 455 Duysinx, BC, 969 Ebdon-Jackson, S, 203 Edenbrandt, L, 127, 417 Eftekhari, M, 567 Eilles, C, 865 Eising, EG, 669 El-Ali, HH, 127 Elgazzar, A, 321, 831 Ellis, B, 1003 Ellul, G, 843 Erba, P, 633 Erba ¸s, B, 45, 877 Erdil, YT, 191 Erdogan, G, 359 Erdogan, MF, 359 Ergu¨n, EL, 877 Ergu¨n, LE, 45 Eriksson, E, 347 Etchebehere, ECSC, 395, 957

Fabbro, M, 137 Fallahi, B, 567 Fanti, S, 1, 685 Fard-Esfahani, A, 567 Farsakoglu, Z, 191 Farshid, G, 695 Ferran, N, 507 Ferro-Flores, G, 371 Fertig, V, 865 Fezoylidis, I, 119 Filippi, L, 381 Firat, F, 447 Fischer, T, 171, 353 Fleming, JS, 701 Foley, RR, 165 Folks, RD, 551 Fox, PT, 573 Fraile, M, 785 Franz, M, 267 Freudenberg, L, 669 Fuchs, E, 865 Fujita, S, 987 Fujitaka, K, 247 Fusco, FM, 205 Fuste´, F, 785 Fuster, D, 11 Futami, S, 987 Gabruk-Szostak, B, 353 Gaeta, GB, 205 Galajda, Z, 501 Galt, JR, 551 Galuska, L, 501 Ga´mez, C, 507 Ganguly, S, 271 Garai, I, 501 Garcia, EV, 551 Garin, E, 363 Gemmel, F, 529 Georgoulias, P, 119 Ghiotto, F, 113 Ghisellini, F, 645 Gholamrezanezhad, A, 567 Ghoorun, S, 765 Gildehaus, FJ, 733 Giurato, L, 745 Goethals, I, 529 Golan, H, 689 Gordon, I, 1003 Go¨rges, R, 669 Grassetto, G, 583 Green, M, 695 Gregianin, M, 1 Groenewald, W, 765 Grosset, DG, 931 Grosset, KA, 931 Grossman, AB, 165 Grosso, M, 439 Gru¨ning, T, 853, 951 Guan, L, 11 Gubern, JM, 785 Guirao, S, 507 Gu¨ltekin, SS, 261

c 2005 Lippincott Williams & Wilkins 0143-3636

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ii Nuclear Medicine Communications 2006, Vol 27

Gu¨nay, EC, 877 Gu¨ngo¨r, F, 715 Gursoy, A, 359 Gutman, F, 5 Gu¨ven, AG, 715 Gyo¨rke, T, 17

Jessurun, GAJ, 791 Jiang, L, 603 Jiang, X, 603 Johnsrud, K, 185 Jones, IW, 853, 951 Julia´n, FJ, 785

Haba, R, 887 Hacker, M, 733 Hadley, DM, 611, 931 Hahn, K, 733 Hainsworth, JES, 461 Ham, HR, 255 Hameed, A, 495 Hamilton, D, 937 Hanada, K, 535 Hansen, CL, 803 Harada, N, 143 Harrison, P, 461 Hasegawa, M, 81 Hatakeyama, K, 987 Hauet, RJ, 137 He, Z-X, 333 Herry, J-Y, 363 Heuts, EM, 677 Hirao, K, 151 Hirata, Y, 151 Hitomi, K, 535 Hitomi, Y, 535 Hitzel, A, 5 Ho Shon, IA, 975 Hofman, M, 837 Hogg, S, 937 Hol, KP, 185 Holte, Ø, 185 Honda, H, 893 Hoofwijk, AGM, 677 Hosono, M, 535 Hospers, GAP, 25 Huang, W-S, 779, 859 Huh, WS, 919 Huisman, H, 515 Hulsewe´, KWE, 677 Hussain, R, 589 Hustinx, R, 969

Kaanders, JHAM, 515 Kabakcı, G, 61 Kadanali, A, 179 Kadanali, S, 179 Kahraman, S, 61 Kamel, N, 359 Kaneko, K, 893 Kang, HJ, 773 Kanno, T, 481 Kano, T, 143 Kantarci, M, 489 Karabulut, E, 61 Karlsen, J, 185 Kawakami, Y, 71 Kawamoto, T, 81 Kazdal, HZ, 715 Kelion, AD, 113 Kengen, RAM, 677 Khalil, MM, 321, 831 Khalil, W, 321, 831 Khan, A, 495 Khan, AU, 495 Khonsari, M, 853, 951 Kim, B-T, 919 Kim, IK, 773 Kiratli, PO¨, 877 Kirime, E, 535 Kiyak, M, 91 Kobe, C, 353 Koc¸ak, U, 45 Koch, W, 733 Koga, H, 893 Koivisto, T, 157 Kollias, J, 695 Komeya, Y, 535 Kondo, K, 143 Kong, H, 807 Kontos, A, 119 Koutsikos, J, 911 Koyun, M, 715 Krause, J, 733 Krause, K-H, 733 Kubota, T, 37 Ku¨c¸u¨k, NO¨, 261, 377 Ku¨c¸u¨k, ON, 873 Kudo, T, 535 Kuikka, JT, 157 Kulak, AH, 873 Kulak, H, 377 Kumar, R, 231 Kumar Mishra, A, 619 Kurien, S, 425 Kuwert, T, 521

Iagaru, A, 795 Ibi ¸s, E, 261, 377, 873 Iida, H, 823 Ikoma, Y, 723 Ilias, I, 31 Inanir, S, 191 Ito, H, 723 Iwamoto, T, 151 Iwasaka, T, 247 Jacobsson, H, 347 Jager, PL, 317, 791 Jakobsson, D, 417 Jamart, J, 815 Janer, J, 785 Jenkins, PJ, 165 Jensen, M, 171 Jentzen, W, 669 Jeong, MJ, 773 Jerabek, PA, 573

la Fouge`re, C, 733 Ladonne, J-M, 5 Lambert, B, 223 Lange, A, 17 Lapi, P, 627

Larisch, R, 267 Larock, M-P, 969 Larsson, SA, 347 Lass, P, 943 Lawson, RS, 843 Laymon, CM, 901 Lazaris, D, 911 Lazzeri, E, 633, 645 Lazzeri, M, 439 Lee, C-H, 779 Lee, CM, 773 Lee, EJ, 919 Lee, JY, 773 Lee, K, 573 Lee, K-H, 919 Lee, MS, 859 Lee, SD, 773 Lejeune, F, 363 Lejeune, J-J, 363 Leon, FA, 507 Li, P, 603 Lima, CSP, 957 Lima, MCL, 395, 957 List, A, 937 Liu, X-J, 333 Ljungberg, M, 127 Llatjo´s, M, 785 Lopez, M, 507 Louis, R, 969 Lumachi, F, 583 Ma, K-H, 779 Maeda, J, 723 Maeda, M, 481 Maenner, P, 865 Mahmoudian, B, 61 Mainta, E, 911 Mako´, E, 17 Malhotra, A, 425 Mamede, M, 31 Manni, C, 381 Mansi, L, 213 Maria Bo¨rner, S, 171 Mariani, G, 1, 439, 627, 709 Mariano-Goulart, D, 137 Mariscal, A, 785 Marshall, VL, 931 Martin, B, 739 Martin, W, 927 Martin-Comin, J, 507 Martinez-Ballarin, I, 507 Marx, K, 353 Marzola, CM, 583 Massari, L, 645 Mather, SJ, 461 Matsuda, H, 151 Matsunaga, N, 71 Matsuzaki, Y, 987 Matteucci, F, 213 Mazo-Ruiz, MFC, 957 McGhie, J, 113 McKay, J, 837 McKillop, JH, 203 Melin, J, 815 Melissinou, M, 911 Menendez, L, 795

Menichetti, F, 645 Mesbah, H, 363 Mester, J, 739 Meucci, G, 627 Mihara, F, 893 Mikhaeel, NG, 101 Millar, AM, 197, 997 Mirpour, S, 567 Mishra, P, 619 Miura, M, 81 Miyazaki, C, 143 Moka, D, 353 Mondal, A, 619 Monson, JP, 165 Monzani, F, 439 Morioka, T, 893 Moser, E, 17 Motherwell, DW, 927 Mulder, NH, 25 Mu¨ller, H-W, 267 Mu¨ller, SP, 669 Mumcuog˘lu, EU¨, 45 Murata, Y, 81 Murray, I, 165 Mustonen, T, 157 Nadig, MR, 425 Nagamachi, S, 987 Nair, N, 413 Nakamura, F, 481 Nakamura, S, 247 Nakao, R, 723 Nandurkar, D, 837 Nanni, C, 685 Nar, F, 45 Narang, R, 425 Nardiello, S, 205 Naylor, V, 113 Nemoto, K, 151 Nemoto, M, 143 Nevin, SM, 951 Nguyen, D, 969 Nguyen, J-M, 431 Nicol, A, 611 Nikolaus, S, 267 Nishii, R, 987 Nishimura, T, 37 Nishimura, Y, 535 Nishiyama, Y, 887 Noiret, N, 363 No¨mayr, A, 521 Nunan, TO, 203 Nuyts, J, 765 O’Brien, LM, 197, 997 O’Doherty, MJ, 101, 833 Ogusu, T, 481 Oh, YH, 919 Ohishi, H, 143 Ohkawa, M, 887 Ohlsson, M, 417 Ohnishi, T, 151 Ohya, N, 893 Okada, H, 481 Okuyama, C, 37 Olofsson, F, 417

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Index to authors iii

Ones, T, 191 Onizuka, T, 987 Ormsby, PL, 853 Orsini, P, 627 Ota, M, 723 Otte, A, 595, 753 Otto, C, 171 Ouchi, Y, 481 Owens, J, 611 Oyen, WJG, 515 Ozener, C, 191 Pacak, K, 31 Palmer, J, 127 Pandit, M, 843 Panwar, P, 619 Papanastassiou, V, 611 Papantoniou, V, 911 Parisella, GM, 633 Pasquini, C, 627 Patel, CD, 425 Patonay, L, 501 Patterson, J, 611, 931 Paula, RB, 395 Pedraza-Lo´pez, M, 371 Pelizzo, MR, 709 Pelizzo, RM, 1 Perkins, AC, 1003 Pe´terffy, A´, 501 Petrie, MC, 927 Piepsz, A, 255 Pierantozzi, M, 381 Pimlott, S, 611 Piquenot, J-M, 5 Plowman, PN, 165 Prandini, N, 633, 645 Prato, FS, 807 Prescott, MC, 843 Puetter, RC, 961 Puig, P, 785 Querellou, S, 431 Rakotonirina, H, 363 Rambaldi, FP, 213 Ramı´ rez-Iglesias, TM, 371 Ramos, CD, 395, 957 Rampin, L, 1 Rampling, R, 611 Ratib, O, 105 Raza, H, 495 Read, J, 1003 Reza Ay, M, 339 Ricart, Y, 507 Rigau, V, 137 Righetti, A, 105 Roach, PJ, 975 Roca, M, 507 Rodrı´ guez-Corte´s, J, 371 Roelants, V, 815 Ro¨mer, W, 521

Rossi, B, 633 Rossi, M, 137 Rotureau, L, 455 Roux, J, 363 Rubello, D, 1, 685, 709, 833 Rull, M, 785 Rutgers, V, 25 Saad, STO, 957 Sadik, M, 417 Saghari, M, 567 Salancı, VB, 45 Salaun, P-Y, 431, 559 Salvatori, M, 1 Sanson, A, 5 Santos, AO, 395, 957 Sarikaya, A, 91, 447 Sarkar, BR, 271 Sarma, DH, 661 Sasaki, M, 893 Satoh, K, 887 Sawamoto, H, 893 Scha¨fer, O, 17 Schembri, GP, 975 Schicha, H, 171, 353 Schillaci, O, 381 Schinagl, DAX, 515 Schmidt, M, 171 Schnall, M, 231 Schneider, E, 669 Schoenberger, J, 865 Schoma¨cker, K, 171, 353 Schuster, S, 11 Sebastian, C, 165 Seiki, Y, 143 Seki, C, 723 S¸en, F, 191, 387 Serra, C, 785 Seven, B, 489 Sharma, R, 619 Shibuya, H, 81 Shields, RA, 843 Siegel, Y, 689 Signore, A, 213, 633, 757 Simi, U, 627 Singh, A, 477 Sitte, W, 267 Skretting, A, 185 Slart, RHJA, 317, 791 Szawek, J, 943 Sohlberg, A, 823 Sola`, M, 785 Somer, EJ, 601 Somsen, GA, 105 Sotiropoulou, M, 911 Spitz, R, 171 Staal, MJ, 791 Stanzione, P, 381 Stodilka, RZ, 807 Strobel, D, 521 Suga, K, 71

Sugiura, T, 247 Sugo, N, 143 Suhara, T, 723 Sundaram, S, 803 Sutbeyaz, Y, 489 Suurkula, M, 417 Suzuki, K, 723 Szabados, L, 501 Szikszai, A, 865

Varga, J, 501 Varoglu, E, 179, 489 Venkatesh, M, 661 Ve´ra, P, 5 Verberne, HJ, 105 Verkeyn, JMA, 677 Vogel, WV, 515 von Borczyskowski, D, 739 von Schoultz, E, 347

Takano, A, 723 Takavar, A, 567 Takekuma, M, 481 Tamura, S, 987 Tang, L, 603 Tang, Z, 603 Tarı, P, 377 Tatsch, K, 733 Tawadros, S, 171 Thein, R, 689 Thompson, SL, 753 Tokmak, E, 377 Tonge, CM, 843 Tønnesen, HH, 185 Top, H, 91 Torizuka, T, 481 To¨zu¨n, N, 387 Traino, AC, 439 Tregnaghi, A, 583 Tress, W, 267 Tsiouris, S, 911 Tsopelas, C, 695 Tsujii, N, 535 Tsuka, Y, 247 Tuglular, S, 191 Turco, A, 627 Turkington, TG, 901 Turog˘lu, HT, 387 Turoglu, T, 191 Turzo, A, 431, 559

Waight, CC, 197 Wakamatsu, H, 987 Walrand, S, 815 Watabe, H, 823 Watanabe, H, 81 Webb, M, 547 Weber, SA, 171 Weinstein, S, 231 Wesolowski, CA, 961 Whatley, M, 31 Wheat, JM, 57 Wieczorek, D, 943 Wilczek, B, 347 Wilke, F, 739 Win, Z, 477 Wyper, D, 611

Uccioli, L, 745 Ug˘ur, O¨, 45 Umemura, S, 247 Ushijima, Y, 37 Uslu, C, 489 Uslu, H, 179, 489 Vaalburg, W, 25 Valette, F, 559 Vallejos, V, 785 Valsamaki, P, 911 van Dalen, JA, 515 Van de Veire, N, 529 van der Ent, FWC, 677 van der Pol, HAG, 677 van Veldhuisen, DJ, 317 Vandenberghe, S, 237 Vanninen, E, 157 Vanninen, R, 157 Vanoverschelde, J-L, 815

Xaplanteris, P, 119 Xydis, K, 119 Yagyu, Y, 535 Yamamoto, Y, 887 Yamashita, F, 151 Yamashita, T, 71 Yang, M-F, 333 Yang, SP, 859 Yang, Y-J, 333 Yardımcı, Y, 45 Yasuno, F, 723 Yee, S-H, 573 Yeh, C-B, 779 Yildirim, M, 179, 489 Yildirim, U, 489 Yin, D, 603 Yokoe, K, 887 Yokota, K, 143 Yoon, J-K, 919 Yoshida, S, 247 Yoshikawa, E, 481 Yoshiura, T, 893 Yu, J, 603 Zaidi, H, 339 Zajic, T, 17 Zaki, M, 71 Zanca, M, 137 Zeniya, T, 823 Zerva, C, 911 Zhuang, H, 11, 231 Zucchetta, P, 583

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iv Nuclear Medicine Communications 2006, Vol 27

Index to subjects Volume 27 2006

Page numbers with ‘‘A’’, ‘‘P’’ or ‘‘T’’ after them represent posters, abstracts or technical notes. Page numbers preceded by ‘‘A’’ represent conference proceedings, which were paginated separately from the journal. abdominal activity 959 absorbed dose 603 acceptance testing 1022A acquisition modes 1022A adenosine stress 281A age 447 albuminuria 965 alginate 185 alkaline phosphatase A10 alkoxysilane 293A Alzheimer’s disease 37, 151, 535, 739 angiogenesis 995 angiographic now-reflow phenomenon 247 animal models A13, 185, 271, 363, 573, 695, 1030P ankle pain 299A anti-CD33 antibodies 299P anti-thyroglobulin autoantibodies 873 antibiotics 645 antitubercular drugs 645 aortic graft infection 303P apocrine metaplasia 911 apoptosis 353 radiation-induced 81 arrhythmias 291A ARSAC 203 arthroplasty 645, 1024A arthroscopy 689 artificial neural networks 937 asthma 281A atrial fibrillation 303P attention-deficit/hyperactivity disorder 267, 733 attenuation correction 339, 765, 843, 853, 1030P audit A9, 283A, 1031P, 1032P autoantibodies 873 axillary dissection 231 bombesin 371 bone infections 307P, 633 bone inflammation 290A bone marrow biopsy 11 bone metastases A10, 294A, 417, 619 bone mineral density 307P bone pain 291A bone scans 417, 865, 877 abnormalities 307P bone scintigraphy 17, 307P, 308P, 521, 919 bone SPECT 291A bowel overlap 281A brain abscess 213 brain imaging 302P, 339

brain perfusion 151 brain SPECT 765 brain tumors 143 breast attenuation artifact 804 breast cancer 5, 287A, 310T, 677, 773, 785, 1036P bone metastases A10 chemotherapy response prediction 297A, 347 detection 911 metastasis 231 see also scentinel lymph nodes breast, lymphatic drainage 1027A 11

C-choline 296A calcitonin 359 cancer breast See breast cancer cell proliferation 995 colon 302P esophageal 296A lung A12, A14, 298, 507, 977, 995 pancreatic 286A prostate A10 thyroid 1, A3, A4, 165, 261, 353, 359, 495, 567, 627, 669 of unknown primary 1020A uterus 481 vulval A3 see also tumors cardiac blood pool studies 292A cardiac function 529 cardiac glycosides 271 cardiac imaging 807 cardiac index A3, A8 cardiac pacemakers 281A cardiac request forms A8 cardiac SPECT 551, 859 cardiac syndrome X 791 cardiac volume 301P cardiology 807 cell signaling A3 central nervous system stimulants 733 cerebral blood flow 535, 943 cerebral oxygen metabolism 573 cerebral perfusion 157 Charcot joint 745 chemotherapy response 306P chest pain 282A chest radiography 297A children bone mineral density 307P congenital hyperinsulinism 290A DMSA scanning 305P, 1036P glomerular filtration rate 255, 305P indirect radionuclide cystography A7 pyelonephritis 715 wrist injury 291A

c 2005 Lippincott Williams & Wilkins 0143-3636

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Index to subjects v

chronic obstructive pulmonary disease 71, 285A cis-enhancer reporter system 773 clinical technologists A5 cognitive impairment 37, 151, 943 collimator response modeling 1023A colloid scintigraphy 387 colon cancer 302P compliance 298A computed tomography 281A, 287A, 288A, 843 ankle/foot pain 299A colon cancer 302P congenital hyperinsulinism 290A contrast enhanced multidetector 1025A image size 515 lung cancer 298A lymphoma 299A neuroendocrine tumors 287A non-small-cell lung cancer 297A pheochromocytoma 583 scan misalignment 1022A computer assisted diagnosis 417 computer simulation 285A congenital hyperinsulinism 290A copper complexes 309P coronary arteriogram 301P coronary artery disease 61, 333, 529 coronary artery mycotic aneurysm 301P coronary bypass surgery 501 correction model 127 cost-benefit ratio 753 Counter Intelligence A7 51 Cr-EDTA 255, 306P, 965 glomerular filtration rate 1025A CT See computed tomography cystic fibrosis A6 DaTSCAN 294A, 295A, 296A, 306P, 937, 1023A dementia 37, 151, 295A, 306P, 535, 739, 943 depression 535 diabetes mellitus, myocardial perfusion 301P diabetic foot 745, 757 diagnosis 11, 481, 529, 715, 931 pelvic inflammatory disease 179 diagnostic accuracy 289A diagnostic work-up 205 diastolic function 1021A DMSA scintigraphy 1025A, 1032P pediatric 1036P l-[[beta]-11C]dopa 723 dopamine synthesis 723 dopamine transporter 296A, 733, 779, 931 dose optimization A5 dosimeters 1035A dosimetry 439, 669 driver exposure 301P drug delivery A1 drug-induced parkinsonism 931 dual isotope lung perfusion/ventilation 303P dual X-ray absorptiometry 307P dual-isotope scintigraphy 865

ductal breast hyperplasia 911 ductal carcinoma in situ 785 see also breast cancer echocardiography 425 ejection fraction A3, 57, 292A, 293A, 300P see also left ventricular ejection fraction elderly patients 529, 535 electrocardiography A8 emphysema 887 end systolic volume 292A endobronchial lesions 296A endocarditis 213 epilepsy 45 temporal lobe 893 ERIC1 antibody 171 esophageal cancer 296A esophageal transit 1026A essential tremor 931, 937 estrogen receptor 773, 911 Evans blue dye 695 Ewing sarcoma 17 exercise testing 333, 791 expert systems 739 external beam radiotherapy 302P extra-cardiac radioactivity 859 extracellular fluid volume-weight formula 969 18

F 293A F-FDG 213, 302P, 405 Alzheimer’s disease 739 breast cancer 231, 297A cancer of unknown primary 1020A endobronchial lesions 296A esophageal cancer 296A Ewing sarcoma 17 lung lesions 507 lymphoma 299A malignant lymphoma 11 non-small-cell lung cancer 297A pheochromocytoma 31 pleural effusions 977 pulmonary metastases 795 Takayasu’s arteritis 547 uterine cancer 481 vertebral osteomyelitis 303P 18 F-FDOPA congenital hyperinsulinism 290A pheochromocytoma 287A 18 F-FEAU, herpes simplex virus thymidine kinase 25 femoral head avascular osteonecrosis 919 fever of unknown origin 205, 213, 1024A filtered backprojection 765 finger dose A13 fluorodeoxyglucose see FDG fluorodopamine 31 foot pain 299A Fourier analysis 105 Fourier transformation 304P fractal analysis 157, 296A free muscle flap transfers 91 18

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vi Nuclear Medicine Communications 2006, Vol 27

68

Ga-DOTA-octreotate 287A 68Ga-DOTATATE 287A, 288A 68Ga-DOTATOC 284A, 288A gallbladder ejection fraction A14 gallium scan 213 gamma camera A2, 284A, 285A computer simulation 285A lung cancer A14 gastric emptying scintigraphy 431 gastrin-releasing peptide receptor 371 gastroesophageal reflux disease 309P, 1026A gastrointestinal bleeding 1036P gated blood-pool radionuclide ventriculography 105 gated cardiac acquisition A8 gated myocardial perfusion SPECT 57, 291A, 321 gated SPECT 292A cardiac index A8 gender 447 geometric mean imaging 701 glioma 137, 611 glomerular filtration rate 407, 965, 969, 1025A calculation 1032P children 255, 305P hemolysed samples 1033P monitoring injection site 1027A Graves’ disease 439, 559 guidelines 203, 297A, 298A, 305P

hallucinations 295A head misalignment 284A heart failure A3, 105 heart rate 281A heart-to-liver ratio 859 hemodialysis 61, 1026A hemolysis 1033P hepatobiliary excretion 859 hepatocellular carcinoma 363 herpes simplex thymidine kinase 25 viral gene expression 611 hibernating myocardium A1 histiocytes 695 Hodgkin’s lymphoma 286A Hu¨rthle cell carcinoma 377 hybrid imaging 521, 983 hydroxyapatite 661 hyperparathyroidism 282A, 709 hyperthyroidism A4, 299P hypoxia imaging 309P 123

I cardiac SPECT 551 thyroid cancer 1 thyroid uptake A4 123 I-FIAU, glioma 611 123 I-FP-CIT 381, 931 123 I-IMP 893 123 I-iomazenil 893 123 I-MIBG

neuroendocrine tumors A12 pheochromocytoma 583 124 I 237 125 I, thyroid cancer 165 125 I-annexin V 81 131 I 284A, 293A, 1026A Graves’ disease 439, 559 monoclonal antibodies 171, 286A salivary gland uptake 669 thyroid cancer A4, 261, 353, 495, 865 131 I-lipiodol 363 image artifact 737, 843, 959 image fusion 515, 521, 843, 983 image quality 737 image reconstruction 807, 823, 901, 1023A immunoreactivity 293A 111 In, white cell imaging 1024A indirect radionuclide cystography A7 inflammatory arthropathy 308P information systems 289A injectables 185 interobserver variability A11 intestinal uptake 877 iodogen 293A iohexol 1025A ionizing radiation, response to A3 ischemia 281A, 282A ischemic heart failure 317 juvenile nasopharyngeal angiofibroma 489 Ki-67 911 kidney gamma camera imaging 285A see also renal knee prosthesis 1024A knee SPECT 689 81 mKr, dual isotope lung perfusion/ventilation 303P Kupffer cells 387 lachrymal scintigraphy 1029P large vessel vasculitis 547 lead shielding 310P left ventricular ejection fraction A9, 321, 425 normal range 1021A left ventricular function 1026A left ventricular volume 292A lemon juice 859 lesion volume determination 283A leucocyte scintigraphy 179, 213 levodopa 931 linear discriminant analysis 535 literature review 205 lithium 299A 177 Lu-labeled hydroxyapatite 661 lung cancer A12, A14, 507, 977, 995 staging 298A lung metastases 261 lung scintigraphy A11

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Index to subjects vii

lung ventilation A7, 71 lung ventilation scintigraphy A6 lung/heart ratio 119 lymphatic drainage 1027A lymphatic mapping A3 lymphedema A14 lymphoma 286A, 299A, 753, 1028A PET 11 lymphoscintigraphy 289A, 308P, 677, 695, 1029P magnetic resonance imaging epilepsy 45 pheochromocytoma 583 mammoscintigraphy 347 maximum likelihood 765 meta-analysis 589, 757 meta-123I iodobenzylguanidine A3 metastatic disease 291A methylphenidate 733 metoclopramide 959 MIB-1 index 995 MIBG 283A microPET 573 MIRD-based models 559 molecular imaging 611, 807, 1019A monoclonal antibodies 171, 286A, 561 Monte Carlo simulation 127 motion correction 292A MUGA scan processing A6, 1021A, 1030P multimodality imaging 1023A musculoskeletal sarcoma 795 myeloid leukemia 299P myocardial acquisition 300P myocardial defects 127 myocardial infarction 247 myocardial perfusion imaging A2, 57, 119, 127, 247, 281A, 292A, 300A, 300P, 791, 837, 959, 1030P, 1031P arrhythmias 291A artifact 959 attenuation correction 853, 1030P audit A9 diabetes mellitus 301P effect of food and water on A9 elderly patients 529 exercise SPECT 119 quantitative accuracy A9 restoration of 247 scintigraphy 113 myocardial stem cell therapy 807 myocardial stress 282A myocardial tomography 333 myocardial viability A2, 915 myometrial infiltration 481 13

N-ammonia 794 Na+/K+-ATPase 271 neck, ultrasound 709 necrosis 353 needle capping 1027A neural networks 417

neuroblastoma 171 neuroectodermal tumors 288A neuroendocrine tumors A12, 283A, 287A, 288A, 300P neutrophile trafficking 285A NICE A9, 289A non-alcoholic steatohepatitis 387 non-Hodgkin’s lymphoma See lymphoma non-research use 286A non-small-cell lung cancer 297A nuclear cardiology service 289A nuclear medicine 601 departments A13, 1029A imaging 283A report writing 1029A service provision 298A, 1020A, 1021A 15

O-labeled oxygen 573 obese patients A8, 915 occult infection 213 octreoscan 282A octreotide A12, 283A scintigraphy A13 optical imaging 317 osteomyelitis 633, 645, 745, 757 ouabagenin conjugates 271 Paget’s disease of bone 307P pancreatic cancer 286A pancreatic imaging 477 parathyroid adenoma 1023A parathyroid imaging 473 parathyroid lesions 282A parathyroid scintigraphy 543 parkinsonian syndrome 937 Parkinson’s disease 294A, 295A, 381, 931, 943 passive smoking 1033P pelvic inflammatory disease 179 penile carcinoma 289A, 298A peptide radiopharmaceuticals 294A percutaneous coronary intervention 247 PET 101, 284A, 288A, 302P, 601 Alzheimer’s disease 739 aortic graft infection 303P breast cancer 231 cardiac syndrome X 791 [methyl-14C]choline A13 colon cancer 302P congenital hyperinsulinism 290A dopamine synthesis 723 endobronchial lesions 296A epilepsy 893 Ewing sarcoma 17 herpes simplex virus 25 image size 515 lesion volume determination 283A lung cancer 298A lymphoma 299A malignant lymphoma 11 myocardial viability 915 neuroendocrine tumors 287A, 288A

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viii Nuclear Medicine Communications 2006, Vol 27

neurological 339 non-small-cell lung cancer 297A oncology 685 pheochromocytoma 31, 287A pleural effusions 977 respiratory motion correction 285A scan misalignment 1022A Takayasu’s arteritis 547 three-dimensional 237 uterine cancer 481 PET/CT mobile service 1019A potential pitfalls 1031P pulmonary metastases 795 soft tissue/skeletal muscle metastases 1020A staff training 1011 whole-body scan 1019A phantoms 1022A, 1032P pheochromocytoma 31, 287A, 583 photoreduction 561 Pichia pastoris 299P pinhole SPECT 823 platelet markers 1030P pleural effusions 977 positron emission tomography see PET preparation 197, 455, 1005 primitive neuroectodermal tumor 17 procalcitonin 715 prognosis 137, 529 progressive supranuclear palsy 381 prostate cancer A10 prosthesis-related infections 633, 645 prosthetic joint infections 307P pulmonary alveolar permeability 1033P pulmonary angiography A11 pulmonary embolism A11, 297A, 1034P pulmonary emphysema 71 pulmonary metastases 795 pulmonary vein stenosis 303P pyelonephritis 715 quality control 255, 304P quantitative analysis 701 radial harvesting 501 radiation protection A6, 284A radiation synovectomy 603, 661 radiation-induced apoptosis 81 radio-guided surgery 1, 709 radioactivity 603 radiochemical purity 197, 455, 543 radiographer reporting 288A, 290A radioimmunoscintigraphy 171 radioimmunotherapy 286A, 299P, 753 pancreatic cancer 286A radioiodine 165, 299P, 495 Hu¨rthle cell carcinoma 377 thyroid cancer A3, 359, 567 see also various iodine isotopes

radioiodine ablation 627 radiometabolic treatment 627 radionuclide bone scan 297A radionuclide imaging 417 radionuclide therapy 223, 561 radionuclide ventriculography 293A radiopharmaceuticals 223, 294A, 757, 1005 radiotherapy 81 localized 185 188 Re 223 188 Re-DMSA 294A 188 Re-rhenium sulfide 603 188 Re-SSS lipiodol 363 region of interest template 37 regional cerebral blood flow 151 registration 521 correction 843 renal clearance 191 renal function evaluation 305P renal function impairment 191 renal function tests 969 renal reflux A7 ambulatory monitoring A5 renal scintigraphy 304P, 1025A renal sufficiency 969 renal transplantation 919 renogram analysis 1032P renography 305P, 701, 969 replacement joints, infection 1025A reporter gene imaging 807 reporting 288A, 289A reproducibility 191 resolution recovery 823 respiratory effort 301P respiratory motion correction 285A resynchronization therapy 105 right ventricular ejection fraction 293A rituximab 293A sacro-iliac index 1024A safety 1027A salivary glands 395 radioiodine uptake 669 scintigraphy 405, 447 scan misalignment 1022A scatter correction 295A, 310P SCID mice 171 scintigraphy 501 bone 17, 307P, 308P, 521, 919 DMSA 1025A, 1032P, 1036P free muscle flap transfers 91 gastric emptying 431 lachrymal 1029P leucocyte 179, 213 lung A11 myocardial perfusion 113, 300P parathyroid lesions 282A pediatric wrist injury 291A renal function evaluation 305P salivary glands 405, 447

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Index to subjects ix

sentinel lymph nodes 5 wrist registration 290A scintimammography 308P, 589, 911 SEHCAT procedure A5 sentinel lymph nodes A1, A4, A5, 231, 289A, 298A, 310P, 695, 785, 833, 1005, 1036P breast cancer 310T, 677 scintigraphy 5 septal penetration 551, 901 service provision 298A, 1020A service standards 1011 severe combined immunodeficiency see SCID sickle cell anemia 965 silent ischemia 61 silent myocardial ischemia 61 single photon emission computed tomography see SPECT SISCOM 45 skeletal muscle metastases 1020A Smartvent A7 sodium iodide symporter 287A, 773 soft copy display screens 304P soft tissue metastases 1020A solid tumors 413 SPECT 37, 287A ADHD 267 ankle/foot pain 299A artifacts 804 bone 291A brain 306P brain perfusion 151 brain tumors 143 breast cancer 347 cardiac 551, 807, 859 chemotherapy response 306P epilepsy 893 gamma camera 284A gated 57 glioma 137 hybrid 521 ictal/interictal 45 juvenile nasopharyngeal angiofibroma 489 knees 689 left ventricular function 1026A myocardial perfusion 113, 119, 292A, 1036P Parkinson’s disease 381, 943 pinhole 823 pulmonary emphysema 71 quality control 304P septal penetration 551, 901 silent myocardial ischemia 61 subarachnoid hemorrhage 157 SPECT/CT 521, 983 acceptance testing 1022A alignment 1022A introduction of 1034P low-dose CT protocol 1034P spinal surgery 291A splenic shift 387 split renal function 1025A spondylodiskitis 633, 645

ST depression 281A statistical mapping 739 stem cells 807 streptavidin 561 stress-rest protocol 113 stroke index A3 subarachnoid hemorrhage 157 survival 137 syringe shields A13, 284A Takayasu’s arteritis 547 99m Tc 308P juvenile nasopharyngeal angiofibroma 489 ouabagenin conjugates 271 99m Tc-depreotide 290A, 507 lung 1027A lung cancer A12, A14 99m Tc-DTPA leakage 310T renal sufficiency determination 969 99m Tc-EDDA/HYNIC-[Lys3]-bombesin 371 99m Tc-HMPAO, leucocyte scintigraphy 179, 301P 99m Tc-HSA, hand perfusion 501 99m Tc-HYNIC 294A 99m Tc-labelled red cells 489 99m Tc-MAA, dual isotope lung perfusion/ventilation 303P 99m Tc-MAG3 clearance 191 preparation of 197 renal scintigraphy 304P 99m Tc-methylene diphosphonate 689, 865 bone scans 877 99m Tc-MIBI 61, 282A myocardial viability 915 scintimammography 589 thyroid cancer 261, 873 99m Tc-Nanocoll 1005 99m Tc-pertechnetate 287A, 395 salivary gland scintigraphy 447 99m Tc-phenylhydrazine 294A 99m Tc-sestamibi breast cancer 347 free muscle flap transfers 91 low protocol 709 myocardial perfusion 127, 837 myocardial tomography 333 parathyroid imaging 473 parathyroid lesions 282A radiochemical purity 455, 543 scintimammography 911 thyroid cancer 1 99m Tc-sulesomab 307P 99m Tc-tetrofosmin 113, 853 cardiac SPECT 859 myocardial perfusion 119, 247, 300A, 959, 1031P 99m Tc-trans-1,2-cyclohexyldinitrilotetramethylene phosphonic acid 619 99m Tc-TRODAT-1 SPECT 733 Tourette’s syndrome 779 99m Tc-VDMSA 911

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x Nuclear Medicine Communications 2006, Vol 27

Technegas A7, A11, 887 thallium 137 thymidine kinase 25 thyroglobulin 567 thyroid cancer 1, A3, A4, 165, 261, 353, 359, 495, 567, 627, 669, 833, 865, 873 extra-thyroidal involvement 1031P follow-up 1027A thyroid clinic 309P thyroid stimulating hormone, recombinant 627 thyroid volume 439 thyrotoxicosis 1026A 201 Tl 271, 804 brain tumors 143 gated SPECT myocardial perfusion scintigraphy A2 non-small cell lung cancer 995 perfusion SPECT 425 TNM staging 101 Tourette’s syndrome 779 tracer uptake 1029P training A7, 1011 trastuzumab 561 treadmill exercise 281A tremulous disorders 296A triple energy window scatter 284A tube voltage 339 tumors 81 brain 143 localization A14 neuroectodermal 288A neuroendocrine 283A, 287A, 288A, 300P solid 413 see also cancer ultrasound neck 709 parathyroid lesions 282A

ultrasound-guided fine needle aspiration 298A uterine cancer 481 V/Q lung scintigrams A11, 1033P, 1034P valvular regurgitation 1021A vascular parkinsonism 931 ventilatory impairment 887 ventricular mechanical dyssynchrony 105 ventriculography 105, 293A verapamil 247 vertebral osteomyelitis 303P virtual learning A6 visual interpretation 295A vulval malignant melanoma A3 white cell imaging A3, 1024A white cell labeling A3 whole-body scanning 285A, 567, 1019A, 1029P wrist injury 291A wrist registration scintigraphy 290A 133

X dynamic SPECT 887 gas washout 71 X-rays 853

86

Y 237 Y internal radiotherapy 185 syringe shields 284A 90 Y-DOTA-lantreotide 300P 90 Y-ibritumomab 595, 1028A 90 Y-zevalin A4 90

zevalin 286A see also 90Y-ibritumomab

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