Neurology Volume 72(3) January 20, 2009

This week in Neurology® Highlights of the January 20 issue Predictors of surgical outcome and pathologic considerations...

Author: American Academy of Neurology

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This week in Neurology® Highlights of the January 20 issue

Predictors of surgical outcome and pathologic considerations in focal cortical dysplasia

FBX07 mutations cause autosomal recessive, early-onset parkinsonian-pyramidal syndrome

Although surgical resection has been an important alternative treatment for patients with intractable epilepsy related to focal cortical dysplasia (FCD), the prognostic relevance of the degree of pathologic severity is controversial. This study assesses whether the pathologic subtypes of FCD affect surgical outcomes in patients with drug-resistant epilepsy. The authors also studied the prognostic roles of clinical factors and various diagnostic modalities in the surgical treatment.

The authors identify pathogenic FBXO7 gene mutations in two families with autosomal recessive, early-onset parkinsonian-pyramidal syndrome, delineating a novel genetically defined disease, designated as PARK15 (for monogenic parkinsonism-type 15). Unraveling the mechanisms of FBXO7-related disease may shed further light on the pathogenesis of parkinsonism and multiple-system brain degenerations.

See p. 211; Editorial, p. 206; see also p. 217

See p. 240

Incomplete resection of focal cortical dysplasia is the main predictor of poor postsurgical outcome Focal cortical dysplasia is the most important cause of focal intractable epilepsy in childhood. This study identifies predictors of favorable/unfavorable postoperative outcome that are crucial for appropriate selection of epilepsy surgery candidates. See p. 217; Editorial, p. 206; see also p. 211

Personality and lifestyle in relation to dementia incidence High neuroticism has been associated with a greater risk of dementia, and an active/socially integrated lifestyle with a lower risk of dementia. This study explores the separate and combined effects of neuroticism and extraversion on the risk of dementia and examines whether lifestyle factors may modify this association. See p. 253

Identification of a possible pathogenic link between congenital long QT syndrome and epilepsy Congenital long QT syndrome (LQTS), a potentially lethal cardiac channelopathy, often masquerades as epilepsy. A diagnostic consideration of epilepsy and treatment with antiepileptic drug medication were far more common in patients with type 2 LQTS (LQT2), caused by perturbations in the KCNH2-encoded potassium channel. See p. 224; Editorial, p. 208

VIEWS & REVIEWS

Genetics of epilepsy syndromes starting in the first year of life Molecular genetic analysis of causative genes for early-onset epilepsies is now possible but needs careful selection of tests and patients when offered as a diagnostic tool. This survey provides practical guidelines for optimal use. See p. 273

Incidence of acquired demyelination of the CNS in Canadian children This paper provides the first prospective national surveillance of acquired demyelination in the pediatric age group. It highlights the clinical features and emphasizes the importance of education in promoting awareness of demyelination and multiple sclerosis risk in children.

CLINICAL IMPLICATIONS OF NEUROSCIENCE RESEARCH

␥-Hydroxybutyric acid and its relevance in neurology ␥-Hydroxybutyric acid (GHB, oxybate) is a normal metabolite of ␥-aminobutyric acid (GABA) and has a widespread neuromodulatory role that is mediated both by GHB-specific and GABAB receptors. Exogenously administered GHB elicits intoxication, withdrawal, tolerance, and addiction; it also induces slow wave sleep and has been approved for treatment of narcolepsy. See p. 282

See p. 232

Podcasts can be accessed at www.neurology.org

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EDITORIAL

Epilepsy surgery patients with cortical dysplasia Present and future therapeutic challenges

Gary W. Mathern, MD

Address correspondence and reprint requests to Dr. Gary W. Mathern, Reed Neurological Research Center, 710 Westwood Plaza, Rm 2123, Los Angeles, CA 90095-1769 [email protected]

Neurology® 2009;72:206–207

Cortical dysplasia is the most common substrate in pediatric and the second or third most frequent etiology in adult epilepsy surgery patients.1 The histopathology was first described less than 30 years ago,2 and we are still learning about the clinical features of this disorder. Early surgical series involved few patients or focused on MRI positive cases,3,4 and reported inconsistent predictors of therapeutic success. Only recently have single centers published surgical cohorts of over 100 patients.5,6 This issue of Neurology® publishes two studies that together report on 315 epilepsy surgery patients with cortical dysplasia: Krsek et al., a mostly pediatric cohort from Miami Children’s Hospital, and Kim et al., a generally adult population from Seoul, Korea.7,8 Both centers have previously published on their cohorts.9,10 From these and other reports emerges a clearer image of the electro-clinico-pathologic characteristics of patients with cortical dysplasia. For example, most surgical patients have mild compared with severe forms of cortical dysplasia on histopathology. This is defined as mild malformations of cortical development (mMCD; 16%) or mild type I (59%) cortical dysplasia using the Palmini classification system (table).11 Only 25% of patients had severe Palmini type II cortical dysplasia. The incidence of type II cortical dysplasia was higher in the mostly pediatric cohort from Miami (35%) while type I dysplasia and mMCD was greater in the adult series from Seoul (83%). Likewise, MRI was reported as normal in 42% of the adult cohort compared with 34% of the pediatric population. Most of the normal MRI scans were in patients with mMCD and type I cortical dysplasia (figure). Extraoperative intracranial electrodes to localize ictal regions were used in 67% of children in the Miami series and in 86% of patients from Korea. Based on MRI and intracranial EEG findings, 31% of the cases from Miami and 33% of the patients from Seoul had incomplete resection of the seizure focus. Incomplete cortical resection was primarily due to overlap of ar-

eas of cortical dysplasia with functional motor, sensory, and language cortex. In both studies, the foremost predictor of postoperative seizure freedom was complete resection of the epileptogenic region. In the Miami series, 70% of patients were seizure-free with complete resections compared with 22% of those with incomplete resections. Similarly, the Seoul group reported that 82% of patients with complete resections were seizure-free compared with 47% with incomplete resections. These findings highlight present and future challenges in diagnosing and treating patients with medically refractory epilepsy with cortical dysplasia. Using current presurgical protocols, a substantial proportion of patients with cortical dysplasia will have negative MRI scans from mostly mMCD and mild type I disease. Hence, we need better neuroimaging and other diagnostic tools that accurately identify patients, especially those with mild cortical dysplasia. With better neuroimaging we can hopefully determine if areas of cortical dysplasia are not surgically removable without unacceptable neurologic deficits and avoid incomplete resections. Other future challenges include development of adjunctive therapies, which combined with resective surgery, can control seizures without disrupting normal cortical functions in areas of mild dysplasia. This may involve novel pharmacologic or other treatments incorporating findings of the neurobiology of cortical dysplasia suggesting that these regions have dysmature cellular and synaptic properties.12 For the practicing neurologist, these two studies have important clinical relevance. Specifically, negative MRI report in a patient with medically refractory epilepsy of undetermined etiology does not exclude that he or she harbors a subtle cortical dysplasia. It is from this kind of experience that experts recommend that patients be referred to a specialty center for a comprehensive epilepsy evaluation if seizures are uncontrolled after failure of two to three antiepilepsy drugs even if the MRI is negative.13 The

See pages 211 and 217 e-Pub ahead of print on November 12, 2008, at www.neurology.org. From the Department of Neurosurgery, The Brain Research Institute and The Mental Retardation Research Center, David Geffen School of Medicine, University of California, Los Angeles. Disclosure: The author reports no disclosures. 206

Copyright © 2009 by AAN Enterprises, Inc.

Table

Histopathologic grading of cortical dysplasia

Grade

Severity

Histopathologic description

mMCD

Very mild vs questionable dysplasia

Normal cortex with excess ectopic neurons in the molecular layer (Layer I) or subcortical white matter. Without cortical abnormalities, there is debate if this represents a “true” form of cortical dysplasia.

Type I

Mild

Cortical disorganization and dyslamination without dysmorphic neurons or balloon cells. My contain hypertrophic (large) or immature neurons.

Type II

Severe

Cortical disorganization and dyslamination with dysmorphic neurons and balloon cells. Sometimes referred to as Taylor type cortical dysplasia.

11

From Palmini et al. mMCD ⫽ mild malformations of cortical development.

other message is that surgical treatment of patients with cortical dysplasia is improving. With proper presurgical evaluation, including newer imaging technologies that reduce the need for invasive intracranial electrodes, more patients with cortical dysplasia can be identified with a 70% to 80% chance of becoming seizure-free after surgery.14,15 As exemplified by the limitations of the Miami and Seoul studies, our knowledge of the clinical characteristics of patients with cortical dysplasia will continue to evolve. Both studies were retrospective and took years to acquire their cohorts. Miami recruited patients from 1986 to 2006 (20 years) and the group from Seoul from 1995 to 2005 (10 years). In that span, the presurgical protocols and tools, like MRI, changed sometimes substantially. Future surgical studies would substantially benefit from multicenter prospective collaborations. This would recruit large cohorts quickly using uniform presurgical protocols, Figure

Patients with difficult-to-identify cortical dysplasia

(A) This 4-year-old patient had daily left body tonic and tonic-clonic seizures. Histopathology confirmed type IIA cortical dysplasia (severe dysplasia with cytomegalic but not balloon cells) in the right insula region (arrows). The MRI had originally been interpreted as normal. (B) This 15-year-old patient had two to three temporal lobe seizures per week. Histopathology showed type I (mild dysplasia without abnormal cells) involving the left anterior temporal lobe (arrow) without associated hippocampal damage. The presurgery MRI was reported as normal without cortical atrophy.

and more accurately accrue quantifiable clinical, EEG, neuroimaging, and other study parameters. Such a design would also allow studies to focus on patients with “purer” histopathologic forms of type I and type II cortical dysplasia. In that way we can learn about factors that characterize and predict treatment success in patients with epilepsy from cortical dysplasia besides incomplete resection.

REFERENCES 1. Harvey AS, Cross JH, Shinnar S, Mathern BW. Defining the spectrum of international practice in pediatric epilepsy surgery patients. Epilepsia 2008;49:146–155. 2. Taylor DC, Falconer MA, Bruton CJ, Corsellis JA. Focal dysplasia of the cerebral cortex in epilepsy. J Neurol Neurosurg Psychiatry 1971;34:369–387. 3. Cohen-Gadol AA, Ozduman K, Bronen RA, Kim JH, Spencer DD. Long-term outcome after epilepsy surgery for focal cortical dysplasia. J Neurosurg 2004;101:55–65. 4. Siegel AM, Cascino GD, Meyer FB, et al. Surgical outcome and predictive factors in adult patients with intractable epilepsy and focal cortical dysplasia. Acta Neurol Scand 2006;113:65–71. 5. Fauser S, Huppertz HJ, Bast T, et al. Clinical characteristics in focal cortical dysplasia: a retrospective evaluation in a series of 120 patients. Brain 2006;129:1907–1916. 6. Widdess-Walsh P, Kellinghaus C, Jeha L, et al. Electroclinical and imaging characteristics of focal cortical dysplasia: correlation with pathological subtypes. Epilepsy Res 2005;67:25–33. 7. Kim DW, Lee SK, Chu K, et al. Predictors of surgical outcome and pathologic considerations in focal cortical dysplasia. Neurology 2009;72:211–216. 8. Krsek P, Maton B, Jayakar P, et al. Incomplete resection of focal cortical dysplasia is the main predictor of poor postsurgical outcome. Neurology 2009;72:217–223. 9. Krsek P, Maton B, Korman B, et al. Different features of histopathological subtypes of pediatric focal cortical dysplasia. Ann Neurol 2008;63:758–769. 10. Chung CK, Lee SK, Kim KJ. Surgical outcome of epilepsy caused by cortical dysplasia. Epilepsia 2005;46 suppl 1:25–29. 11. Palmini A, Najm I, Avanzini G, et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62(6 suppl 3):S2–8. 12. Cepeda C, Andre VM, Levine MS, et al. Epileptogenesis in pediatric cortical dysplasia: the dysmature cerebral developmental hypothesis. Epilepsy Behav 2006;9:219–235. 13. Cross JH, Jayakar P, Nordli D, et al. Proposed criteria for referral and evaluation of children for epilepsy surgery: recommendations of the Subcommission for Pediatric Epilepsy Surgery. Epilepsia 2006;47:952–959. 14. Salamon N, Kung J, Shaw SJ, et al. FDG-PET/MRI coregistration improves detection of cortical dysplasia in epilepsy patients. Neurology 2008 (in press). 15. Wu JY, Sutherling WW, Koh S, et al. Magnetic source imaging localizes epileptogenic zone in children with tuberous sclerosis complex. Neurology 2006;66:1270–1272.

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EDITORIAL

Seizures and arrhythmias Differing phenotypes of a common channelopathy?

Jill V. Hunter, MBBS Arthur J. Moss, MD

Address correspondence and reprint requests to Dr. Jill V. Hunter, Baylor College of Medicine, 6621 Fannin Street, MC2-2521, Houston, TX 77030 [email protected]. org

Neurology® 2009;72:208–209

The human ether-a-go-go–related gene (HERG, now known as KCNH2) encodes an ion channel subunit IKr, which subserves a potassium current that repolarizes human heart ventricular cells. Mutations in KCNH2 cause type 2 long QT syndrome (LQT2) by disrupting IKr, increasing cardiac excitability, and even triggering the catastrophic arrhythmia torsades de pointes (TdP),1,2 ventricular tachyarrhythmias, and sudden death. In syncope related to ventricular tachyarrhythmias, loss of consciousness is abrupt in onset and offset. In contrast, epileptic seizures are abrupt in onset but have a slow offset associated with the postictal state. Cardiac syncope can progress into seizure activity if insufficient cerebral perfusion due to rapid ventricular tachyarrhythmia lasts for a minute or longer. In this issue of Neurology®, Johnson et al.3 examine a hypothesized commonality between the cardiac channelopathies associated with the long QT interval (especially LQT2) and CNS channelopathies that may cause seizure activity. They inferred this hypothesis from family histories utilizing the concept of “seizure phenotype,” defined as “the presence of either a personal or family history of seizures or epilepsy or a history of AED therapy.” Information was obtained from chart review of 343 consecutive unrelated patients clinically evaluated and genetically tested for long QT syndrome (LQTS) from 1998 to 2006. They found a personal history of seizure to be more frequent in LQT2 (39%), than either LQT1 or LQT3, with seizure phenotype in 47% of LQT2 patients. The authors speculate that “LQT2-causing perturbations in the KCNH2-encoded potassium channel may confer susceptibility for recurrent seizure activity.” This large prevalence with an apparent association to a specific mutation is intriguing, but must be interpreted with caution. The loosely defined family history may have resulted in some incorrect designation of a genetic relationship. Similarly, the criteria used to define seizure phenotype, including 24 hours of antiepileptic drug (AED) use, may have led to a spurious or

inaccurate diagnosis of epilepsy, especially in the absence of EEG documentation. Brain and heart share a similar ion channel milieu, and thus ion channel mutations resulting in a cardiac phenotype may also predispose to a CNS phenotype. The ion channels associated with LQTS were originally identified as myocardial specific, but recent reports make clear that the cardiac potassium channels, KCND2 (Kv4.2), KCNQ1 (Kv7.1), and KCNH2 (Kv11.1), are expressed in the CNS.4-6 This raises the question whether the seizures/syncope observed in some LQTS patients are cardiogenic in origin (TdP-type ventricular tachyarrhythmias) or epileptic seizures related to perturbations in CNS ion channel function. Patients with LQTS have been misdiagnosed with epilepsy and treated inappropriately with AEDs. However, some of these patients may have a neural channelopathy resulting in seizures and would benefit from therapy with AEDs. An important limitation of the present study, therefore, is the absence of information about the etiology (cardiac or neurogenic) of the personal seizure/ syncope phenotype associated with LQTS and especially LQT2. EEG testing is not standard of care for patients with LQTS and was therefore unavailable for this cohort. An additional diagnostic confound concerns the population of LQTS patients without (as yet) an identifiable ion channel mutation, i.e., those who are phenotype-positive (LQTS) and genotype-negative. A related issue is whether seizure disorders can be caused by single-gene mutations—as can happen with some forms of LQTS— or whether seizures result from mutations in susceptibility genes with other requirements for the expression of seizures. Seizure susceptibility genes could account for the finding that not all individuals with a mutation of the ion channel causing LQTS have a seizure history. Patients with LQTS 1 and 3 also seem to have an increased risk of seizure history above that of the general population (0.5–1% for epilepsy). Perhaps the LQT mutation is a necessary but not sufficient condition for the development of a seizure disorder.

See page 224 From Baylor College of Medicine (J.V.H.), Texas Children’s Hospital, Houston; and University of Rochester Medical Center (A.J.M.), Rochester, NY. Disclosure: The authors report no disclosures. 208

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Further studies are warranted, requiring a minimum of a three-generational family pedigree and EEG confirmation of seizure activity in addition to genetic testing. Should the findings of Johnson et al. be confirmed, the implications for therapy of a common channelopathy may be significant. EEG may become part of the workup for LQTS and ECG/Holter monitoring an integral part of the examination for a subset of patients with epilepsy. Pharmacogenetics profiling may one day apply to both the epilepsy and LQTS populations, focusing therapies on modulation of a specific ion channel that may treat both cardiac and CNS conditions.7 Such findings may lead to a better understanding and prevention of sudden unexpected death in epilepsy.8 REFERENCES 1. Fitzgerald PT, Ackerman MJ. Drug-induced torsades de pointes: the evolving role of pharmacogenetics. Heart Rhythm 2005;2:S30–S37.

2. 3.

4.

5.

6.

7.

8.

Moss AJ, Kass RS. Long QT syndrome: from channels to cardiac arrhythmias. J Clin Invest 2005;115:2018–2024. Johnson JN, Hofman N, Haglund CM, Cascino GD, Wilde AAM, Ackerman MJ. Identification of a possible pathogenic link between congenital long QT syndrome and epilepsy. Neurology 2009;72:224–231. Birnbaum SG, Varga AW, Yuan LL, et al. Structure and function of Kv4-family transient potassium channels. Physiol Rev 2004;84:803–33. Dalby-Brown W, Hansen HH, Korsgaard MP, et al. K(v) 7 channels: function, pharmacology and channel modulators. Curr Topics Med Chem 2006;6:999–1023. Haufe V, Chamberland C, Dumaine R. The promiscuous nature of the cardiac sodium current. J Mol Cell Cardiol 2007;42:469–477. Danielsson BR, Lansdell K, Patmore L, Tomson T. Phenytoin and phenobarbital inhibit human HERG potassium channels. Epilepsy Res 2003;55:147–157. Vatta M, Gertz SJ, Anderson AE. Epilepsy: Getting to the heart of the matter. European Paediatrics Review. Neurology Touch Briefings 2007;49–50.

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IN MEMORIAM

Sydney S. Schochet, Jr., MD (1937–2008)

Ludwig Gutmann, MD Jack E. Riggs, MD

Sydney S. Schochet, Jr., MD

Sydney S. Schochet, Jr., an icon in American neuropathology, died at his home in Gainesville, Florida, on September 18, 2008, surrounded by the family he loved so much. A superb clinician and teacher, he was revered by his colleagues, residents, and medical students. His 300 scientific publications spanned the spectrum of neuropathology. His weekly, well-attended teaching conferences were often conducted by residents whose authoritative presentations reflected the hours he spent with them in preparation—minus his soft Louisiana drawl. His effectiveness as a teacher resulted in numerous awards, including the McLachlan Award for outstanding basic science teaching on four occasions and the Outstanding Teacher Award in 1988 at the West Virginia University School of Medicine. Dr. Schochet was born February 7, 1937, in Chicago, the only son of Sydney S. Schochet, MD, and Mathilde Goldman Schochet. He received his undergraduate and medical degrees from Tulane University in 1959 and 1961. He trained in pathology and neuropathology at Tulane and the Armed Forces Institute of Pathology (AFIP), where he proudly served in the US Army as a major from 1967 to 1969. Dr.

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Schochet was a faculty member of the University of Iowa, University of Texas Medical Branch, and University of Oklahoma prior to spending 22 years at West Virginia University. He retired in 2003 and spent the last few years living in Gainesville, where the residents at the University of Florida honored him with the Didactic Teacher Award in 2006. Dr. Schochet had an enviable national and international reputation for his diagnostic, research, and teaching skills. He was a regular faculty member of the AFIP and AAN courses in neuropathology and served for many years on the AAN Residency Inservice Training Examination committee. His love of photography was reflected in the many high-quality slides that illustrated his conferences and lectures. Many of the neuropathology photos on the ABPN Part I examination and the AAN Resident In-service test were from Dr. Schochet’s vast collection. Sid, as all his friends and colleagues called him, was a quiet, unassuming man with a dry sense of humor. He liked to say that his name should be pronounced with a soft French inflection, although the more guttural pronunciation might be appropriate for a neuropathologist since it sounded like the word for the person certified to kill animals under Jewish law. Sydney S. Schochet was much more than an outstanding physician and educator. He was an exceptional human being who was always kind and caring and took immense satisfaction in the accomplishments of his coworkers. He was devoted to his wife, Pauline, two daughters, Katrina and Terri, and four grandchildren. He enjoyed working with his daughters on school science projects (his favorite was building a windup cockroach). More recently, he taught a granddaughter to wiggle her eyebrows as well as rub her belly and pat her head simultaneously. Pauline recalled that, years ago, her father predicted she would never find a husband who could meet her high expectations, “but he was wrong.”

ARTICLES

Predictors of surgical outcome and pathologic considerations in focal cortical dysplasia D.W. Kim, MD S.K. Lee, MD, PhD K. Chu, MD K.I. Park, MD S.Y. Lee, MD C.H. Lee, MD C.K. Chung, MD, PhD G. Choe, MD, PhD J.Y. Kim, MD, PhD

Address correspondence and reprint requests to Dr. Sang Kun Lee, Department of Neurology, Seoul National University Hospital, 28, Yongkeun dong, Chongno Ku, Seoul, 110-744, Korea [email protected]

ABSTRACT

Background: Although surgical resection has been an important alternative treatment for patients with intractable epilepsy related to focal cortical dysplasia (FCD), the prognostic relevance of the degree of pathologic severity is controversial and there has been only limited information regarding the prognostic factors involved in the surgical treatment of refractory epilepsy in patients with FCD.

Methods: We undertook the present study to assess whether the pathologic subtypes of FCD affect surgical outcomes in patients with drug-resistant epilepsy. We also studied the prognostic roles of clinical factors and various diagnostic modalities in the surgical treatment.

Results: A total of 166 consecutive patients were included. By univariate analysis, incomplete resection of epileptogenic area (p ⬍ 0.001), mild pathologic features (p ⫽ 0.01), and the presence of secondary tonic clonic seizures (2GTCS) (p ⫽ 0.05) were associated with poor surgical outcomes. There was a strong tendency for patients with severe pathologic features to have MRI abnormalities (p ⬍ 0.001). Incomplete resection of epileptogenic area (p ⬍ 0.001) and mild pathologic features (p ⫽ 0.02) were poor independent outcome predictors on multivariate analysis. The results of MRI, scalp EEG, fluorodeoxyglucose–PET, and ictal SPECT were not associated with surgical outcomes. Conclusions: Our study shows that there is a strong tendency for patients with severe pathologic features to have MRI abnormalities, and patients with incomplete resection, mild pathologic features, or the presence of secondary tonic clonic seizures have a high chance of a poorer surgical outcome. Neurology® 2009;72:211–216 GLOSSARY 2GTCS ⫽ secondary tonic clonic seizures; FC ⫽ febrile convulsion; FCD ⫽ focal cortical dysplasia; FDG ⫽ fluorodeoxyglucose; FDG-PET ⫽ fluorodeoxyglucose–positron emission tomography; FLE ⫽ frontal lobe epilepsy; IED ⫽ interictal epileptiform discharge; mMCD ⫽ mild malformations of cortical development; nTLE ⫽ neocortical temporal lobe epilepsy; OLE ⫽ occipital lobe epilepsy; PLE ⫽ parietal lobe epilepsy; SPECT ⫽ single-photon emission computed tomography.

Focal cortical dysplasia (FCD) is the most commonly encountered developmental malformation that causes refractory epilepsy. The pathologic features of FCD include mild disturbances, such as neuronal heterotopias or dyslamination, and severe abnormalities such as large, bizarre pyramidal neurons within the cortex and in the white matter, often associated with large balloon cells.1-3 With advances in neuroimaging techniques, in particular MRI, recent studies have revealed a higher prevalence of FCD than previously estimated and have improved the preoperative identification and classification of these abnormalities.4 Surgical resection has been an important alternative treatment for patients with intractable epilepsy related to FCD. Although it is very important to determine the prognostic factors in identifying ideal candidates for surgery and in predicting the prognoses of individual patients, there has been only limited information regarding the prognostic factors involved in the surgical treatment of refractory FCD. The prognostic relevance of the degree of pathologic severity Editorial, page 206 See also page 217 e-Pub ahead of print on November 12, 2008, at www.neurology.org. From the Department of Neurology (D.W.K.), Konkuk University Medical Center, Seoul; Comprehensive Epilepsy Center (S.K.L., K.C.), Department of Neurology, Seoul National University Hospital; Department of Neurology (K.I.P.), Seoul Paik Hospital, Inje University; Department of Neurology (S.Y.L.), Kangwon National University Hospital, Chuncheon; Departments of Internal Medicine (C.H.L.), Neurosurgery (C.K.C.), and Pathology (G.C.), College of Medicine, Seoul National University; and Department of Neurology (J.Y.K.), Hallym University Sacred Heart Hospital, Pyeongchon, Korea. Disclosure: The authors report no disclosures. Copyright © 2009 by AAN Enterprises, Inc.

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is controversial, because the presence of severe pathologic features has been associated with favorable surgical outcomes in some reports,5-7 whereas others have documented the opposite results.8,9 Moreover, few studies have analyzed the diagnostic and prognostic roles of modern presurgical evaluation techniques, such as MRI, fluorodeoxyglucose (FDG)-PET, and ictal single-photon emission computed tomography (SPECT), and their relationships to the severity of the pathology have not been adequately evaluated. In this study, we hypothesized that the degree of pathologic severity would be related to the sensitivity of diagnostic modalities and completeness of resection, and affect surgical outcomes of refractory epilepsy in patients with FCD. Therefore, we investigated the surgical outcomes in patients with FCD and evaluated the prognostic implications of the degree of pathologic severity, clinical factors, and the results of presurgical diagnostic modalities. METHODS Patients. Our study included 166 consecutive patients who had undergone surgical treatment for refractory epilepsy and had a pathologic diagnosis of FCD at the Seoul National University Hospital between November 1995 and June 2005. We excluded three patients with insufficient postoperative follow-up (less than 2 years), and 10 patients with other potentially epileptogenic lesions, such as tumors, vascular malformations, ischemic changes, and sequelae of trauma. We excluded 10 patients with malformations of cortical development other than FCD including periventricular heterotopia (2), polymicrogyria (7), and lissencephaly (1 patient). We also excluded 40 patients who had radiologic evidence of hippocampal sclerosis (HS) in addition to FCD, because it is possible that HS as well as FCD could act as epileptogenic areas in these patients. All patients had intractable epilepsy, despite taking the appropriate anticonvulsant drugs. We included only patients with focal resection and excluded patients with functional hemispherectomy or corpus callosotomy. Surgical outcome was divided into seizure-free and not seizure-free.

Magnetic resonance imaging. All patients underwent brain MRI. Standard MRI was performed on either a 1.0-T or a 1.5-T unit (Signa Advantages; General Electric Medical Systems, Milwaukee, WI), with conventional spin-echo T1-weighted sagittal and T2-weighted axial and coronal sequences in all patients. The section thickness and the conventional image gaps were 5 mm and 1 mm. T1-weighted three-dimensional magnetization-prepared rapid acquisition of gradient-echo sequences with 1.5 mm thick sections of the whole brain and T2-weighted fluid-attenuated inversion recovery images of 3 mm thick sections were also obtained in the oblique coronal plane of the temporal lobe. The angle of oblique coronal imaging was perpendicular to the long axis of the hippocampus. Spatial resolution was approximately 1.0 ⫻ 1.0 mm (matrix, 256 ⫻ 256 mm; field of view, 25 cm). Functional neuroimaging. PET was performed in 151 patients during the interictal period (no seizures for more than 24 212

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hours). Axial raw data were obtained using a PET scanner (ECAT EXACT 47; Siemens CTI, Knoxville, TN) 60 minutes after the IV injection of 18F-fluorodeoxyglucose (FDG, 370 MBq). Spatial resolution was 6.1 mm ⫻ 6.1 mm ⫻ 4.3 mm. FDG-PET images were assessed visually and by statistical parametric mapping analysis, as described previously.10 Ictal SPECT was performed in 121 patients during video-EEG monitoring. Technetium-99m (99MTc) was mixed with hexamethylpropyleneamine oxime (925 MBq) and injected as soon as a seizure started. Brain SPECT images were acquired within 2 hours of the injection. A triple-head rotating gamma camera (Prism 3000; Picker, Cleveland, OH) equipped with a high-resolution fan beam collimator was used. Interictal SPECT was also performed to identify perfusion changes. Side-by-side visual analysis of interictal and ictal images was performed and the subtraction method was used, as previously described.10 The results of FDGPET and SPECT were defined as localizing if the predominant hyperperfusion area or the predominant hypometabolic zone was confined to the resected lobe, and diffuse or multifocal abnormalities beyond the epileptogenic area were not considered as localizing even when they are within the epileptogenic hemisphere including the epileptogenic area.

Video-EEG monitoring. Interictal and ictal scalp EEGs were recorded in all patients using a video-EEG monitoring system, with electrodes placed according to the international 10 –20 system, with additional anterior temporal electrodes. In 143 patients for whom other methods gave inconclusive or discordant results, we used a combination of grids and strips for intracranial EEG. Grid and strip placements were determined by the results of presurgical evaluations. At least three habitual seizures were recorded during scalp and intracranial EEG monitoring. When necessary, preoperative and intraoperative functional mapping and intraoperative electrocorticography were also performed. A localizing pattern of ictal-onset rhythm/interictal spike was defined as a localized ictal rhythm/interictal spike in the electrodes of an epileptogenic lobe or two adjacent electrodes. In patients with multiple areas of ictal-onset rhythm/interictal spike, we defined as localizing only when the whole areas are finally considered epileptogenic, and did not include when the abnormalities were multilobar within the epileptogenic hemisphere including the epileptogenic area. Surgery and pathology. The surgical area was decided based on the clinical, neuroimaging, and electrophysiologic results. In our center, the resection margin for epilepsy of neocortical origin was defined by 1) the presence of either a discrete lesion on MRI with compatible ictal EEG or a massive and exclusive ictal-onset zone confirmed by intracranial EEG and 2) the absence of eloquent cortex. Complete resection was defined by resection of all of the MRI-visible lesions or areas of ictal onset, persistent pathologic delta slowing, ⬎1/second frequent spikes, and intermittent gamma wave by intracranial EEG. The diagnosis and classification of pathologic cortical dysplasia were according to the system of Palmini and colleagues: ectopically placed neurons only (mMCD), isolated architectural abnormalities (FCD 1A), additional immature or giant neurons (FCD 1B), presence of dysmorphic neurons (FCD 2A), and additional balloon cells (FCD 2B).1

Statistical tests. The association between the pathologic severity of FCD and the diagnostic sensitivity of various diagnostic modalities was tested with a linear association ␹2 test. We compared the clinical characteristics of the seizure-free group and the non–seizure-free group with Student t test or the Mann-

Whitney U test for continuous variables and with a ␹2 test for categorical variables. Variables with a p value of ⬍0.2 on univariate analysis were included in a multiple logistic regression model, to identify the predictors of surgical outcome. In multiple logistic regression, backward elimination was used to select variables to be maintained in the final model, using a p value of ⬍0.05 as the criterion for the significant associations. All analyses were conducted using SPSS version 12.0 (SPSS Inc., Chicago, IL) and STATA version 9.2 (STATA Corp., College Station, TX). A p value ⬍ 0.05 was considered significant.

16 of FCD 2B. There was a strong tendency toward better surgical outcomes at the last follow-up visit in patients with more severe abnormalities, because only 42.9% of patients with mMCD, 51.7% of those with FCD 1A, and 60.7% of those with FCD 1B became seizure-free, compared with 66.7% of patients with FCD 2A and 87.5% of those with FCD 2B who became seizure-free (p ⫽ 0.003).

RESULTS Patients and surgical outcomes. In our surgical cohort, 956 patients underwent surgical treatment for refractory epilepsy excluding vagus nerve stimulator implantation, and the pathologic diagnosis of FCD was made in 206 (21.5%) patients. Forty patients were excluded for the radiologic evidence of HS, and 166 (17.4%) patients were finally included for the present analysis. The subjects consisted of 102 men and 64 women, and the mean age of the patients at the time of surgery was 24.7 ⫾ 10.2 years (range, 3–51 years). The mean age at seizure onset was 11.5 ⫾ 7.0 years (range, 0.5–30 years) and the mean interval between seizure onset and surgery was 12.8 ⫾ 7.8 years (range, 0.5–37 years). A history of febrile convulsions was present in 18.1% (30/166) of patients, and 74.7% (124/166) of patients experienced secondary generalized tonic clonic seizures (2GTCS). The mean length of postoperative follow-up was 7.94 years. The proportion of seizure-free patients was 56.6% (94/166) during the follow-up for more than 2 years, whereas 43.4% (72/166) of patients did not become seizure-free. Changes of outcome during the follow-up period were observed in 17 of 166 patients. The running-down phenomenon was observed in 11 of 94 patients who became finally seizure-free, and reappearance of seizures after initially seizure-free state occurred in 6 of 72 patients who were not seizure-free.

genic area was performed in 111/166 (66.9%) patients, and the completeness of resection and pathologic severity were not associated, as shown in table 1. However, the completeness of resection was an important prognostic factor on univariate (table 2) and multivariate analysis (table 3). We could not perform complete resection in 55 patients because the epileptogenic zone included portions of eloquent area in 26 patients, and the epileptogenic area was often multifocal (10 patients) and widespread (19 patients).

Postoperative mortality and morbidity. No postoper-

ative mortality was found. Postoperative complications occurred in 15 (9.0%) patients. Postoperative infections occurred in seven patients including four meningitis, one skull osteomyelitis, and two scalp wound infections. Four incidents of postoperative hematoma were noted after intracranial electrode placement, which required emergency decompression. A postoperative neurologic deficit was noted in four patients, but all the neurologic deficits resolved spontaneously during the follow-up period. Surgical outcomes according to different pathologic subtypes of FCD. Pathologically, 21 patients showed

characteristics of mMCD, 89 patients of FCD 1A, 28 patients of FCD 1B, 12 patients of FCD 2A, and

Surgical outcomes according to completeness of resection. Complete resection of potently epilepto-

Diagnostic sensitivities of various modalities and their relationships to pathologic subtypes. MRI detected

focal abnormalities in the resected area in 42.2% of patients. Interictal scalp EEG showed unifocal epileptiform discharges to the resected lobe in 42.7% of patients, and ictal scalp EEG correctly localized them to the resected lobe in 76.5% of patients. FDG–PET showed concordant focal hypometabolism in 71.4% of patients and ictal SPECT showed concordant focal hyperperfusion in 60.1% of patients. The diagnostic sensitivities of various diagnostic modalities in relation to pathologic classification are summarized in table 1. There was a strong tendency to have identifiable lesions on MRI (p ⬍ 0.001) in patients with severe pathologic features. The diagnostic sensitivities of interictal/ictal EEG findings, FDG–PET, and ictal SPECT were not related to the severity of the pathologic features. Prognostic factors for seizure-free outcomes. On univariate analysis, the presence of 2GTCS (p ⫽ 0.05) was associated with poor surgical outcomes. The patients were divided into two subgroups: the mild pathologic group (mMCD, FCD 1A, and FCD 1B) and the severe pathologic group (FCD 2A and 2B). Patients in the severe pathologic group had a higher chance of becoming seizure-free than did patients in the mild pathologic group (p ⫽ 0.01; table 2). Because there was a strong tendency for patients with severe pathologic features to have focal lesions on MRI, the prognostic role of pathology may depend on the results of MRI. However, the severe pathologic features were related to a high rate of becoming seizure-free, even after adjustment for the effect of MRI (p ⫽ 0.04). Neurology 72

January 20, 2009

213

Table 1

Completeness of resection and diagnostic sensitivity according to pathologic subtypes mMCD

FCD 1A

FCD 1B

FCD 2A

FCD 2B

13/21 (61.9)

57/89 (64.0)

20/28 (71.4)

7/12 (58.3)

14/16 (87.5)

0.131

Diagnostic sensitivity

19.0 (4/21)

30.3 (27/89)

60.7 (17/28)

75.0 (9/12)

81.3 (13/16)

⬍0.001

False negative rate, %

81.0

69.7

39.3

25.0

18.7

Diagnostic sensitivity

61.9 (13/21)

40.4 (36/89)

50.0 (14/28)

16.7 (2/12)

37.5 (6/16)

False negative rate, %

38.1

59.6

50.0

83.3

62.5

Diagnostic sensitivity

76.2 (16/21)

79.8 (71/89)

75.0 (21/28)

66.7 (8/12)

68.8 (11/16)

False negative rate, %

23.8

20.2

25.0

33.3

31.2

Diagnostic sensitivity

66.7 (12/18)

66.3 (55/83)

78.3 (18/23)

41.7 (5/12)

93.3 (14/15)

False negative rate, %

33.3

33.7

21.7

58.3

Diagnostic sensitivity

80.0 (12/15)

56.3 (36/64)

40.9 (9/22)

71.4 (5/7)

76.9 (10/13)

False negative rate, %

20.0

43.7

59.1

28.6

23.1

Diagnostic sensitivity

68.4 (13/19)

85.9 (67/78)

86.4 (19/22)

80 (8/10)

92.9 (13/14)

False negative rate, %

31.6

14.1

13.6

20.0

Completeness of resection of potently epileptogenic zone

p Value

MRI

Interictal EEG 0.206

Ictal scalp EEG 0.352

FDG–PET (151) 0.269

6.7

Ictal SPECT (121)

Invasive monitoring (143) 0.189

7.1

Values are n (%) or %. mMCD ⫽ mild malformations of cortical development; FCD ⫽ focal cortical dysplasia; FDG-PET ⫽ fluorodeoxyglucose– positron emission tomography; SPECT ⫽ single-photon emission computed tomography.

Multivariate analysis identified incompleteness of resection (p ⬍ 0.001) and mild histopathologic features (p ⫽ 0.02) as independent significant prognostic factors for a poor surgical outcome (table 3). Consistent with our observation, complete resection of MRI-visible lesions or the epileptogenic zone has been associated with favorable surgical outcomes.5,9,11,12 Several studies documented the prognostic values of several clinical factors including seizures of short duration or early epilepsy surgery; we could not replicate these reports.12-14 Our study shows that patients with severe pathologic features have a higher chance of becoming seizure-free after surgery. We first assumed that the prognostic effect of pathologic severity in our study was attributable to the different sensitivity of MRI in detecting FCD, because patients with severe FCD pathology have a higher chance of presenting with abnormalities on MRI. However, the prognostic effect of pathologic severity was still strong after we statistically adjusted for the effect of MRI. The positive prognostic effect of severe pathologic features has also been documented in previous reports, which predominantly included patients with identifiable lesions on MRI.5-7,15 The prognostic role of pathologic severity may be related to the completeness of surgiDISCUSSION

214

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January 20, 2009

cal resection. Although the epileptogenic zone in FCD does not always correspond to the area with the most severe pathologic features, it is generally accepted that the epileptogenicity of FCD is usually associated with its pathologic severity. Because the grading of pathologic severity is usually based on the features of the most severely affected area, mild pathologic grades may reflect incomplete surgical resection of the epileptogenic zone in some patients. Alternatively, differences in prognoses may be associated with the different sensitivities of diagnostic modalities other than MRI. It can be assumed that severely dysplastic areas are more epileptogenic, regardless of the presence of an MRI abnormality, and that highly epileptogenic areas have a greater chance of accurate localization by other diagnostic modalities, such as scalp EEG and functional neuroimaging. The presence of 2GTCS was associated with poor surgical seizure-free outcomes in our study. The negative prognostic role of 2GTCS in epilepsy surgery for FCD has been observed in patients with mTLE,16,17 frontal lobe epilepsy,18 and FCD.11,12 The presence of 2GTCS may be a marker for more widespread cortical involvement and a widely distributed epileptogenic area,12 or continuing generalized epileptic activity may lead to widespread cortical struc-

Table 2

Clinical characteristics and outcomes after surgical resection in 166 patients with epilepsy with focal cortical dysplasia: Univariate analysis of prognostic factors for seizure-free outcomes for at least 2 years after surgery Seizure-free (n ⴝ 94, 56.6%)

Not seizure-free (n ⴝ 72, 43.4%)

Age at operation (y, mean ⴞ SD)

24.7 ⫾ 10.2

23.6 ⫾ 8.7

0.49

p Value

Age at onset (y, mean ⴞ SD)

11.4 ⫾ 7.5

11.5 ⫾ 6.4

0.96

Duration of epilepsy (y, mean ⴞ SD)

13.3 ⫾ 8.2

12.2 ⫾ 7.2

0.37

Men/women

56/38

46/26

0.57

Seizure frequency per month (mean ⴞ SD)

21.4 ⫾ 47.9

20.9 ⫾ 55.6

0.94

Presence of 2GTCS

64

60

0.05

Presence of FC

17

13

1.00

Complete resection of epileptogenic area

77

34

⬍0.001

MRI, concordant focal lesion

45

25

0.09

Interictal EEG, focal IED

41

30

0.80

Ictal EEG, localized ictal onset zone

68

59

0.15

PET, concordant focal hypometabolism (n ⴝ 151)

55/82

49/69

0.60

Ictal SPECT, concordant focal hyperperfusion (n ⴝ 121)

38/71

34/50

0.11

Invasive study

84

59

0.17

Mild pathologic characteristics

72

66

0.01

FLE

30

23

0.16

nTLE

24

28

PLE

8

8

OLE

9

3

23

10

Epilepsy syndrome

Multilobar

2GTCS ⫽ secondary generalized tonic clonic seizure; FC ⫽ febrile convulsion; IED ⫽ interictal epileptiform discharge; FLE ⫽ frontal lobe epilepsy; nTLE ⫽ neocortical temporal lobe epilepsy; PLE ⫽ parietal lobe epilepsy; OLE ⫽ occipital lobe epilepsy.

tural and functional changes, including secondary epileptogenesis in interconnected brain areas that are not removed by surgical resection.19 Whereas approximately 80% of patients became seizure-free after surgical treatment for mTLE related to HS or lesional epilepsy, such as primary brain tumor or vascular malformations,16,20 the efficacy of surgical treatment for FCD was consistently less favorable, with approximately 33–75% of individuals becoming seizure-free.5,6,12,21 Simple resection of the lesion, as performed in patients with mTLE or lesional epilepsy, may be a feasible surgical option as Table 3

Clinical characteristics and outcomes after surgical resection in 166 patients with epilepsy with focal cortical dysplasia: The final model of multivariate analysis of prognostic factors for seizure-free outcome for >2 years after surgery

Factors

OR

95% CI

Mild pathologic characteristics

0.305

0.108–0.857

Complete resection of epileptogenic area

4.939

2.405–10.140

p Value 0.02 ⬍0.001

long as the lesion is completely removed. However, the complete removal of FCD is often difficult because the demarcation of the lesion is frequently poor, and dysplastic tissues tend to be more extensive than is apparent on MRI.11,22 Furthermore, the seizures can originate in either the area of MRI abnormality or the area adjacent to the lesion, and MRI frequently does not show any abnormalities in patients with pathologically proven FCD.23 The frequent overlap of the epileptogenic zone with eloquent cortex may also be involved in the poor surgical outcomes.24 Evidence indicates that even patients with MRI abnormalities who have resective epilepsy surgery for FCD have worse surgical outcomes than those of patients who have surgery for other focal lesional epilepsy syndromes.25 The seizure-free rate in our study (56.6%) was no better than those of previous reports, but this result may be significant, considering that more than half the patients in our study failed to show definable abnormalities on MRI. Although a direct comparison of MRI and other diagnostic modalities may be difficult, it is clear that the localization of a lesion by MRI can both enhance our confidence and provide more information than do FDG–PET or ictal SPECT, especially when the localization is supported by an electrophysiologic study. MRI can also be helpful in planning appropriate intracranial electrode placement. Imaging findings may favor surgery in some patients who might otherwise be rejected for surgery.26 Although only a trend toward better surgical outcomes was observed in patients with definable lesions on MRI (p ⫽ 0.09), we believe that the recruitment of a large number of patients may confirm the prognostic value of MRI in epilepsy surgery for FCD. Received November 21, 2007. Accepted in final form July 14, 2008.

REFERENCES 1. Palmini A, Najm I, Avanzini G, et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62(suppl 3):2–8. 2. Sisodiya SM. Malformations of cortical development: burdens and insights from important causes of human epilepsy. Lancet Neurol 2004;3:29–38. 3. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia: a critical review of human tissue studies and animal models. Epilepsia 2007; 48(suppl 2):21–32. 4. Widdess-Walsh P, Diehl B, Najm I. Neuroimaging of focal cortical dysplasia. J Neuroimaging 2006;16:185–196. 5. Tassi L, Colombo N, Garbelli R, et al. Focal cortical dysplasia: neuropathological subtypes, EEG, neuroimaging and surgical outcome. Brain 2002;125:1719–1732. Neurology 72

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Chung CK, Lee SK, Kim KJ. Surgical outcome of epilepsy caused by cortical dysplasia. Epilepsia 2005;46(suppl 1): 25–29. Widdess-Walsh P, Kellinghaus C, Jeha L, et al. Electroclinical and imaging characteristics of focal cortical dysplasia: correlation with pathological subtypes. Epilepsy Res 2005;67:25–33. Palmini A, Gambardella A, Andermann F, et al. Operative strategies for patients with cortical dysplastic lesions and intractable epilepsy. Epilepsia 1994;35(suppl 6):57–71. Fauser S, Schulze-Bonhage A, Honegger J, et al. Focal cortical dysplasias: surgical outcome in 67 patients in relation to histological subtypes and dual pathology. Brain 2004; 127:2406–2418. Lee SK, Yun CH, Oh JB, et al. Intracranial ictal onset zone in nonlesional lateral temporal lobe epilepsy on scalp EEG. Neurology 2003;61:757–764. Cohen-Gadol AA, Ozduman K, Bronen RA, Kim JM, Spencer DD. Long-term outcome after epilepsy surgery for focal cortical dysplasia. J Neurosurg 2004;101:55–65. Fauser S, Bast T, Altenmuller DM, et al. Factors influencing surgical outcome in patients with focal cortical dysplasia. J Neurol Neurosurg Psychiatry 2008;79:103–105. Kral T, Clusmann H, Blu¨mcke I, et al. Outcome of epilepsy surgery in focal cortical dysplasia. J Neurol Neurosurg Psychiatry 2003;74:183–188. Siegel AM, Cascino GD, Meyer FB, Marsh WR, Scheithauer BW, Sharbrough FW. Surgical outcome and predictive factors in adult patients with intractable epilepsy and focal cortical dysplasia. Acta Neurol Scand 2006;113: 65–71. Hardiman O, Burke T, Phillips J, et al. Microdysgenesis in resected temporal neocortex: incidence and clinical significance in focal epilepsy. Neurology 1988;38:1041–1047. Jeong SW, Lee SK, Hong KS, Kim KK, Chung CK, Kim H. Prognostic factors for the surgery for mesial temporal

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lobe epilepsy: longitudinal analysis. Epilepsia 2005;46: 1273–1279. Hennessy MJ, Elwes RDC, Honavar M, Rabe-Hesketh S, Binnie CD, Polkey CE. Predictors of outcome and pathological considerations in the surgical treatment of intractable epilepsy associated with temporal lobe lesions. J Neurol Neurosurg Psychiatry 2001;70:450–458. Janszky J, Jokeit H, Schulz R, Hoppe M, Ebner A. EEG predicts surgical outcome in lesional frontal lobe epilepsy. Neurology 2000;54:1470–1476. Liu RSN, Lemieux L, Bell GS, et al. Progressive neocortical damage in epilepsy. Ann Neurol 2003;53:312–324. Britton JW, Cascino GD, Sharbrough FW, Kelly PJ. Lowgrade glial neoplasms and intractable partial epilepsy: efficacy of surgical treatment. Epilepsia 1994;35:1130–1135. Alexandre V Jr, Walz R, Bianchin MM, et al. Seizure outcome after surgery for epilepsy due to focal cortical dysplastic lesions. Seizure 2006;15:420–427. Sisodiya SM, Free SL, Stevens JM, Fish DR, Shorvon SD. Widespread cerebral structural changes in patients with cortical dysgenesis and epilepsy. Brain 1995;118:1039– 1050. Lee SK, Lee SY, Kim KK, Hong KS, Lee DS, Chung CK. Surgical outcome and prognostic factors of cryptogenic neocortical epilepsy. Ann Neurol 2005;58:525–532. Marusic P, Najm IM, Ying Z, et al. Focal cortical dysplasia in eloquent cortex: functional characteristics and correlation with MRI and histopathologic changes. Epilepsia 2002;43:27–32. Yun CH, Lee SK, Lee SY, Kim KK, Jeong SW, Chung CK. Prognostic factors in neocortical epilepsy surgery: multivariate analysis. Epilepsia 2006;47:574–579. Vattipally VR, Bronen RA. MR imaging of epilepsy: strategies for successful interpretation. Magn Reson Imaging Clin N Am 2004;14:347–372.

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Incomplete resection of focal cortical dysplasia is the main predictor of poor postsurgical outcome P. Krsek, MD, PhD B. Maton, MD P. Jayakar, MD, PhD P. Dean, ARNP B. Korman, MS G. Rey, PhD C. Dunoyer, MD E. Pacheco-Jacome, MD G. Morrison, MD J. Ragheb, MD H.V. Vinters, MD, PhD T. Resnick, MD M. Duchowny, MD

Address correspondence and reprint requests to Dr. Pavel Krsek, Department of Pediatric Neurology, Charles University, Second Medical School, Motol University Hospital, V Uvalu 84, CZ 15006 Prague 5, Czech Republic [email protected]

ABSTRACT

Background: Focal cortical dysplasia (FCD) is recognized as the major cause of focal intractable epilepsy in childhood. Various factors influencing postsurgical seizure outcome in pediatric patients with FCD have been reported.

Objective: To analyze different variables in relation to seizure outcome in order to identify prognostic factors for selection of pediatric patients with FCD for epilepsy surgery.

Methods: A cohort of 149 patients with histologically confirmed mild malformations of cortical development or FCD with at least 2 years of postoperative follow-up was retrospectively studied; 113 subjects had at least 5 years of postoperative follow-up. Twenty-eight clinical, EEG, MRI, neuropsychological, surgical, and histopathologic parameters were evaluated.

Results: The only significant predictor of surgical success was completeness of surgical resection, defined as complete removal of the structural MRI lesion (if present) and the cortical region exhibiting prominent ictal and interictal abnormalities on intracranial EEG. Unfavorable surgical outcomes are mostly caused by overlap of dysplastic and eloquent cortical regions. There were nonsignificant trends toward better outcomes in patients with normal intelligence, after hemispherectomy and with FCD type II. Other factors such as age at seizure onset, duration of epilepsy, seizure frequency, associated pathologies including hippocampal sclerosis, extent of EEG and MRI abnormalities, as well as extent and localization of resections did not influence outcome. Twenty-five percent of patients changed Engel’s class of seizure outcome after the second postoperative year.

Conclusions: The ability to define and fully excise the entire region of dysplastic cortex is the most powerful variable influencing outcome in pediatric patients with focal cortical dysplasia. Neurology® 2009;72:217–223 GLOSSARY FCD ⫽ focal cortical dysplasia; HS ⫽ hippocampal sclerosis; mMCD ⫽ mild malformations of cortical development; MST ⫽ multiple subpial transections; SGTCS ⫽ secondarily generalized tonic-clonic seizures; SS ⫽ standard scores.

Focal cortical dysplasia (FCD) and mild malformations of cortical development (mMCD) are recognized as the major cause of focal intractable epilepsy in childhood constituting up to 50% of cases in pediatric epilepsy surgery series.1,2 The currently used classification3,4 divides mMCD/FCD according to the severity of histologic findings. Favorable surgical outcomes have been reported in recent FCD series, with the postoperative rates of seizure-free patients varying between 50% and 75%.5–10 Appropriate selection of epilepsy surgery candidates based on identification of predictors of favorable/unfavorable postoperative outcome is crucial in these patients. Several Editorial, page 206 See also page 211 e-Pub ahead of print on November 12, 2008, at www.neurology.org. From the Department of Pediatric Neurology (P.K.), Charles University, Second Medical School, Motol University Hospital, Prague, Czech Republic; Department of Neurology (B.M., P.J., P.D., C.D., T.R., M.D.), Behavioral Medicine (B.K., G.R.), and Comprehensive Epilepsy Program (B.M., P.J., P.D., C.D., T.R., M.D.), Neuropsychology Section (B.K., G.R.), Department of Radiology (E.P.-J.), and Department of Neurological Surgery (G.M., J.R.), Brain Institute, Miami Children’s Hospital, FL; Pathology and Laboratory Medicine (Neuropathology) and Neurology (H.V.V.), Los Angeles, CA; and Department of Neurology (T.R., M.D.), University of Miami Miller School of Medicine, FL. Supported by grant IGA NR/8843-4 and VZ MZO 00064203. Disclosure: The authors report no disclosures. Copyright © 2009 by AAN Enterprises, Inc.

217

Table 1

Comparison of clinical data and neurologic and neuropsychological findings with seizure outcome

Characteristic

Engel I

Engel II

Engel III

Engel IV

Pre/perinatal risk factors Positive (n ⴝ 17) Negative (n ⴝ 132)

9 (53)

1 (6)

73 (55)

16 (12)

<2 (n ⴝ 75)

38 (51)

10 (13)

>2 (n ⴝ 74)

44 (59)

7 (9)

3 (18) 11 (8)

4 (24) 32 (24) p ⫽ 0.5886

Age at seizure onset, y 9 (12)

18 (24)

5 (7)

18 (24) p ⫽ 0.5510

Duration of epilepsy, y <5 (n ⴝ 57)

30 (53)

9 (16)

5 (9)

13 (23)

>5 (n ⴝ 92)

52 (56)

8 (9)

9 (10)

23 (25) p ⫽ 0.6243

Positive (n ⴝ 27)

13 (48)

5 (19)

1 (4)

Negative (n ⴝ 122)

69 (57)

12 (10)

13 (11)

7 (18)

5 (13)

Incidence of infantile spasms 8 (30) 28 (23) p ⫽ 0.3490

Incidence of status epilepticus Positive (n ⴝ 39)

18 (46)

Negative (n ⴝ 110)

64 (58)

10 (9)

9 (23)

9 (8)

27 (25) p ⫽ 0.3310

7 (15)

13 (27)

7 (7)

23 (23) p ⫽ 0.3285

29 (25)

Incidence of SGTCS Positive (n ⴝ 48)

22 (46)

6 (12)

Negative (n ⴝ 101)

60 (59)

11 (11)

Frequency of seizures Daily (n ⴝ 122)

67 (59)

9 (8)

9 (8)

Less frequent (n ⴝ 27)

15 (43)

8 (23)

5 (14)

7 (20) p ⫽ 0.1765

Neurologic findings Normal (n ⴝ 98)

54 (55)

10 (10)

Abnormal (n ⴝ 51)

28 (55)

7 (14)

12 (12) 2 (4)

22 (23) 14 (27) p ⫽ 0.3639

Neuropsychological findings (standard score categories) in 95 subjects SS <59 (n ⴝ 21)

9 (43)

2 (10)

3 (14)

7 (33)

SS 60–69 (n ⴝ 19)

9 (47)

1 (6)

3 (16)

6 (31)

SS 70–79 (n ⴝ 19) SS >80 (n ⴝ 36)

9 (47) 24 (67)

2 (11)

2 (11)

6 (31)

2 (5)

1 (30)

9 (25) p ⫽ 0.1613

Neuropsychological tests (standard score ranking) were available in a group of 95 patients. See text for explanation. The values in parentheses are percentages. SGTCS ⫽ secondarily generalized tonic-clonic seizures.

general epilepsy surgery variables are reported to be significant predictors of favorable postsurgical outcome: early indication for surgery, 8,11 regional ictal EEG patterns,12 detection of MRI lesion,8,13 and circumscribed resection.8,9 Temporal location of the surgery 9,12,14 and histopathologic types of dysplastic lesions (Krsek et al., submitted)9,13 were found to be important in FCD cases. Complete excision of the dysplastic cortex is often attributed to the contribution of intracranial EEG monitoring,5,15,16 MRI,6,17,18 both MRI and intracranial EEG,19 –21 or intraoperative histopathology.22 218

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January 20, 2009

However, the results remain highly controversial. Few studies report long-term seizure outcome after surgery for FCD.6,11,18,21,22 To address these issues we analyzed clinical, EEG, MRI, neuropsychological, surgical, and histopathologic data in relation to seizure outcome in a large cohort of children with histologically proven mMCD/FCD including 113 subjects with at least 5 years of postoperative follow-up. METHODS Patient selection. Patients who underwent resective epilepsy surgery at the Miami Children’s Hospital from March 1986 to June 2006 were retrospectively reviewed. Only subjects with a definite histologic diagnosis of mMCD/FCD were selected. Subjects with tuberous sclerosis complex, benign brain tumors, polymicrogyria, nodular heterotopia, SturgeWeber syndrome, and hemimeganencephaly were not included. We did not exclude patients with hippocampal sclerosis and a history of perinatal brain insults coexisting with mMCD/FCD. Two hundred subjects from a population of 567 patients operated on during this period met the above-mentioned criteria. Of these, 51 patients were excluded because they were either lost to follow-up (n ⫽ 21) or were postoperatively followed for less than 2 years (n ⫽ 30). A total of 149 patients were retained for analysis.

Presurgical evaluation. A complete history and a neurologic examination were obtained in every patient. All patients underwent preoperative scalp video EEG using the standard 10 –20 system of electrodes with additional electrodes applied in selected cases. Interictal epileptiform discharges and ictal EEG patterns were classified as either regional (appearing exclusively over a single lobe or in two contiguous regions such as centroparietal discharges) or non-regional (e.g., multilobar, hemispheric, or generalized). Patients with multiple ictal EEG patterns (such as regional and not-lateralized patterns) were always considered nonregional. MRI scans of patients imaged at 1.5-Tesla including FLAIR sequences (n ⫽ 108) were rigorously reevaluated. The remaining 41 children were either imaged at 0.3-Tesla or did not have available presurgical MRI data; their MRI results were excluded from the analysis. MRI data were re-evaluated independently by three experienced investigators (P.K., B.M., and E.P.J.). Presence of hippocampal sclerosis (HS) was evaluated based on combined histopathologic and MRI criteria. We confirmed the diagnosis of HS from the histopathologic findings or MRI evidence of hippocampal atrophy and signal intensity change if there was insufficient tissue for pathologic analysis. Presurgical neuropsychological data were reevaluated in 95 patients. For the purpose of the study, global functional ranking was determined because heterogeneous neuropsychological batteries had been utilized over the years in the assessment of the cases and also because some of the subjects could not be examined with common psychometric instruments due to pervasive intellectual or developmental impairments. The following categories of the ranking based on standard scores (SS) were distinguished: SS 1: moderate to severe impairment, IQ ⱕ59; SS 2: mild impairment, IQ 60 – 69; SS 3:

Table 2

Comparison of EEG, MRI findings, and hippocampal pathology with seizure outcome

Characteristic

Engel I

Engel II

Engel III

Engel IV

Normal (n ⴝ 83)

45 (54)

9 (11)

8 (10)

21 (25)

Slow (n ⴝ 66)

37 (56)

8 (12)

6 (9)

15 (23) p ⫽ 0.9925

Present (n ⴝ 106)

61 (58)

11 (10)

10 (9)

Absent (n ⴝ 43)

21 (49)

6 (14)

4 (9)

12 (28) p ⫽ 0.5594

3 (5)

15 (27)

Background activity

Focal slowing 24 (23)

Interictal epileptiform activity Regional (n ⴝ 55)

30 (55)

7 (13)

Nonregional (n ⴝ 94)

52 (55)

10 (11)

11 (12)

Regional (n ⴝ 54)

29 (54)

8 (15)

7 (13)

10 (18)

Nonregional (n ⴝ 95)

53 (56)

9 (10)

7 (7)

26 (27) p ⫽ 0.3689

Abnormal (n ⴝ 82)

45 (55)

10 (12)

6 (7)

21 (26)

Normal (n ⴝ 26)

14 (54)

2 (8)

1 (4)

Revealed (n ⴝ 71)

42 (59)

7 (10)

4 (6)

18 (25)

Not revealed (n ⴝ 37)

17 (46)

5 (14)

3 (8)

12 (32) p ⫽ 0.4542

21 (22) p ⫽ 0.5876

Ictal EEG patterns

MRI finding

9 (34) p ⫽ 0.3222

Detection of mMCD/FCD by MRI

MRI abnormalities other than mMCD/FCD Present (n ⴝ 18)

10 (56)

3 (16)

1 (6)

Absent (n ⴝ 90)

49 (54)

9 (10)

6 (7)

4 (22) 26 (29) p ⫽ 0.5782

Extent of MRI abnormality (changes typical for cortical malformations) Unilobar (n ⴝ 37)

21 (57)

5 (14)

2 (5)

Multilobar/hemispheric (n ⴝ 71)

38 (53)

7 (10)

5 (7)

9 (24) 21 (30) p ⫽ 0.6239

Histologically proven HS Present (n ⴝ 15) Not found (n ⴝ 134)

6 (40)

3 (20)

76 (57)

14 (10)

2 (13) 12 (9)

4 (27) 32 (24) p ⫽ 0.5555

HS proven by both MRI and histopathology Present (n ⴝ 22) Not found (n ⴝ 127)

9 (41)

3 (14)

73 (57)

14 (11)

4 (18) 10 (8)

6 (27) 30 (24) p ⫽ 0.3516

The values in parentheses are percentages. See text for diagnostic criteria of hippocampal sclerosis (HS). mMCD ⫽ mild malformations of cortical development; FCD ⫽ focal cortical dysplasia.

borderline intelligence, IQ 70 –79; SS 4: low average intelligence and above, IQ ⱖ80.

Surgical procedures and completeness of the resections. Forty-nine patients underwent one-stage excisional procedures. Pre-excision electrocorticography was used to define the epileptogenic zone and resection plane in these patients. Chronic invasive monitoring utilizing implanted subdural electrodes was performed in 100 patients including all cases with normal MRI scans. Completeness of the resection was determined by the epilepsy team composed of neurologists, neuroradiologists, and neurosurgeons at time of surgery and was subsequently confirmed by the authors using primary data. The resection was considered complete only if the region of the structural abnormality (if detected by MRI) and significant EEG abnormalities (defined below) were entirely removed. Resections were judged

incomplete if either the structural lesion or the electrographically abnormal region could not be completely excised because of proximity to the eloquent cortex. The criteria for evaluating intracranial EEG data have been published previously.23–25 In patients undergoing intraoperative electrocorticography, regions of active spiking with consistent focality, exhibiting rhythmic features such as trains of focal fast activity, or associated with focal attenuation of background were considered significant and resected. Infrequent spikes and spikes without consistent focality were ignored. In chronically implanted subjects, the seizure onset zone was the most critical factor in determining the epileptogenic region, especially in patients with normal MRI studies. It was defined as a region exhibiting focal rhythmic activity, bursts of highfrequency discharges, repetitive spiking, or electrodecremental patterns. Secondary foci that consistently activated during seizures were also included in the resection if they occurred in tissue adjacent or in regional proximity to the primary ictal focus. We defined secondary foci as cortical regions evidencing early spread of ictal activity and active independent spiking that were detected during intracranial recordings. Regions of frequent focal interictal spiking and background abnormalities with consistent focality were also considered significant. Slow waves occurring over widespread regions shortly after seizure onset were not regarded as critical for completeness of resection. Operative complications occurred in 27 subjects and included infections (n ⫽ 9), cerebrovascular accidents (n ⫽ 4), bleeding (n ⫽ 2), brain edema (n ⫽ 4), hydrocephalus (n ⫽ 1), CSF leak (n ⫽ 1), hemiparesis (n ⫽ 3), and cranial nerve paresis (n ⫽ 3). No subject died after the surgery.

Neuropathologic classification. All neuropathologic findings were reclassified by a neuropathologist (H.V.) and two experienced epileptologists (P.K. and M.D.) according to the current classification scheme.3,4 The following mMCD/FCD subcategories were recognized in our series. Twenty-nine patients had mild malformation of cortical development type II (mMCD) defined by the presence of abundant heterotopic white matter neurons and normal cortical architecture. The presence of a small number of ectopic neurons in the temporal lobe was not regarded as a significant pathologic finding. There were no patients with ectopically placed neurons only in or adjacent to layer 1 in our series (mMCD type I). Thirty-nine subjects were classified as FCD type Ia because of the presence of cortical dyslamination without cellular abnormalities. A microcolumnar arrangement of cortical neurons was frequently encountered in these patients, usually with an increase in the number of ectopic white matter neurons. Twenty-nine patients exhibited cortical dyslamination in conjunction with minor cellular abnormalities (giant and immature neurons) and were classified as FCD type Ib. The histopathologic findings of 28 subjects identified as FCD type IIa were characterized by more pronounced architectural and cytoarchitectural disturbances, especially dysmorphic neurons, but without balloon cells. Twenty-four patients with FCD type IIb exhibited the same features as the previous group but evidenced balloon cells that are pathognomic for this histopathologic subtype. Follow-up and outcome. Data regarding seizure outcome and antiepileptic drug treatment were obtained at 2, 5, and 10 years after the last surgery during outpatient visits and telephone contact. Surgical outcome was classified according to Engel’s classification scheme: Engel I, completely seizure-free, auras only or only atypical early postoperative seizures; Engel II, ⱖ90% Neurology 72

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Table 3

Comparison of surgical variables with seizure outcome

Characteristic

Engel I

Engel II

Engel III

Engel IV

11 (11)

25 (25)

Invasive monitoring (subdural electrodes) Yes (n ⴝ 100)

52 (52)

12 (12)

No (n ⴝ 49)

30 (61)

5 (10)

3 (6)

11 (23) p ⫽ 0.6770

Side of surgery Right (n ⴝ 73)

44 (60)

4 (6)

Left (n ⴝ 76)

38 (50)

13 (17)

Yes (n ⴝ 20)

10 (50)

3 (15)

No (n ⴝ 129)

72 (56)

14 (11)

9 (12)

16 (22)

5 (7)

20 (26) p ⫽ 0.0809

Reoperations 3 (15) 11 (8)

4 (20) 32 (25) p ⫽ 0.7279

Multiple subpial transections Yes (n ⴝ 8) No (n ⴝ 141)

3 (37.5) 79 (56)

0 (0) 17 (12)

1 (12.5) 13 (9)

4 (50)†

Seizure outcome in relation to clinical data. Influence of clinical variables and neurologic findings on seizure outcome is shown in table 1. There were no significant differences in seizure outcome between individual subgroups of patients, i.e., no clinical factor was a significant predictor of postsurgical seizure outcome.

32 (23)†

Seizure outcome in relation to EEG and MRI features.

Extent of surgery One lobe (n ⴝ 91)

46 (51)

12 (13)

Multilobar (n ⴝ 37)

19 (51)

Hemispherectomy (n ⴝ 21)

17 (81)

8 (9)

25 (27)

3 (8)

5 (14)

10 (27)

2 (9)

1 (5)

1 (5) p ⫽ 0.1942

Temporal vs extratemporal resections (in 91 cases with one lobe surgery) Temporal (n ⴝ 32)

17 (53)

4 (12.5)

4 (12.5)

Extratemporal (n ⴝ 59)

28 (47)

8 (14)

4 (7)

Complete (n ⴝ 103)

72 (70)

14 (13)

Incomplete (n ⴝ 46)

10 (22)

3 (6)

7 (22) 19 (32) p ⫽ 0.5792

Completeness of surgery 9 (9) 5 (11)

8 (8)* 28 (61)* p ⬍0.0001

The values in parentheses are percentages. *p ⬍ 0.0001. †Statistics not done because of small numbers of subjects with multiple subpial transactions.

seizure reduction or nocturnal seizures only; Engel III, ⱖ50% seizure reduction; and Engel IV, ⬍50% seizure reduction.

Statistical analysis. Overall relationships between seizure outcome (i.e., proportions of patients in individual Engel categories) and different variables of interest (e.g., clinical, EEG, and MRI features) were assessed primarily using ␹2 statistics within a contingency table format. Statistical analysis was performed twice: on the entire series and separately in a group of 127 subjects without hippocampal sclerosis. When overall significance tests indicated no relationship between rows and columns, follow-up procedures based on normal approximation were used. With ordered categorical data, the Cochran-Armitage test was used to determine trends undetectable through ␹2 or other methods. For binomial outcomes showing no relationships, post hoc statistical procedures were based on normal approximation, such as Fisher least significant difference. For multinomial outcomes, residual patterns were examined and categories were collapsed or omitted to determine the likely category within the contingency table responsible for the lack of independence. While none of the tables were extremely sparse (i.e., expected values ⬍1.0), some did contain expected values ⬍5. All p values were determined using exact statistical procedures (StatExact-5, Cytel Software Corporation). ␹2 results are presented as follows: ␹2 (␹2 statistic), p value. RESULTS There were 66 men and 83 women in our series, 144 children and adolescents (younger 220

than 20 years), and 5 young adults (aged 20 –25 years). At 2 years from (the last) surgery, numbers and proportions of patients in individual outcome categories were as follows: Engel I, 82 (55%); Engel II, 17 (12%); Engel III, 14 (9%); Engel IV, 36 (24%). A total of 28 clinical, EEG, MRI, neuropsychological, surgical, and histopathologic variables were analyzed.

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The influence of several scalp EEG parameters, MRI findings, and hippocampal pathology on seizure outcome is summarized in table 2. In general, EEG and MRI features were not predictive of postsurgical seizure outcome. We found no evidence of superior outcome for patients with highly localized interictal epileptiform activity and ictal EEG patterns compared to patients with non-regional epileptiform abnormalities. Importantly, there were no differences in seizure outcome between patients with normal and abnormal MRI scans. Seizure outcome in relation to hippocampal pathology. Fif-

teen patients had preoperatively detected HS, which was subsequently histopathologically confirmed. Seven more cases had insufficient tissue for pathologic analysis but demonstrated MRI evidence of hippocampal atrophy and signal intensity change. There was no difference in seizure outcome between subjects with and without HS. In order to exclude a possibility that the occurrence of HS modified our results, statistical analysis was done separately for the entire group of patients and after the exclusion of patients with HS. There were no differences between the groups. Seizure outcome in relation to neuropsychological ranking. Comparison of seizure outcome with neuro-

psychological findings is provided in table 1. A trend toward higher proportion of seizure-free patients in subjects with higher standard score ranking (above 80) was found (␹2 ⫽ 5.22, p ⫽ 0.1613). Seizure outcome in relation to surgical variables. The

relationship between different surgical variables and seizure outcome is presented in table 3. There was a highly significant relationship between completeness of resection and surgical outcome. Patients with complete resections had superior surgical outcomes compared to patients undergoing incomplete resec-

Figure

Proportions of mMCD/FCD subgroups of patients in individual seizure outcome categories given in percentages

tions who underwent MST. When reasons for the incomplete resections were analyzed, proximity of the dysplastic cortex to eloquent cortical regions was the most important factor. There were no differences in outcome if incomplete resections were determined by MRI or EEG criteria. Long-term seizure outcome. Data were available for

tions at the time of surgery (␹2 ⫽ 52.06, p ⬍ 0.0001). Eight children underwent multiple subpial transections (MST) because of an apparent overlap of the epileptogenic zone and eloquent cortex; their surgical outcomes were poorer in comparison with the entire group of patients. There was a trend toward better outcome in patients undergoing hemispherectomy compared to unilobar and multilobar resections (␹2 ⫽ 8.65, p ⫽ 0.1935). Other surgical variables did not influence outcome.

113 patients with at least 5 years of postsurgical follow-up, and 10-year follow-up was available in 55 patients. In 85 of 113 patients (75%), seizure outcome remained stable after the second year after the surgery. Three patients became seizure-free between the second and fifth postsurgical year, but none became seizure-free after the fifth postoperative year. Outcome improved (e.g., changed from Engel III to Engel II category) in six subjects after the second postsurgical year. Seizures recurred in 11 patients between the second and fifth postoperative year and in five after the fifth year. The condition worsened (e.g., changed from Engel II to Engel III category) in three patients after the second postoperative year. We found no differences in long-term outcome between subjects with complete and incomplete resections. Individual histopathologic subgroups of mMCD/FCD did not differ in proportions of patients who changed long-term outcomes. However, whereas almost all FCD type II patients who changed outcome after the second postsurgical year either recurred or worsened (10 of 11 patients), significant proportions of changed mMCD and FCD type I patients improved (3 of 8 and 5 of 10). Consequently, minor differences in seizure outcome that were apparent 2 years after the surgery (e.g., better outcome in FCD type II compared with FCD type I patients) disappeared at 5- and 10-year follow-up. Twenty-one patients (14%) were off medication at the end of the follow-up period.

Seizure outcome in relation to histopathology. A com-

DISCUSSION

FCD ⫽ focal cortical dysplasia; mMCD ⫽ mild malformations of cortical development.

parison of surgical outcome and neuropathologic classification of mMCD/FCD is presented in the figure. No differences between individual subgroups of mMCD/FCD were found (␹2 ⫽ 11.46, p ⫽ 0.4895). However, patients with FCD type IIa and IIb were over-represented in Engel I category while FCD type Ia and Ib subjects were more frequent in the Engel IV group. Analyses of patients groups in terms of the completeness of surgery. Since completeness of resections was

the single significant predictor of surgical outcome, individual variables were re-analyzed in subjects with complete and incomplete resections. There were no clinical, EEG, MRI, surgical, and histopathologic variables different between the groups, except for a higher proportion of subjects with incomplete resec-

Our study was conducted on the largest population of pediatric epilepsy surgery patients with histologically proven FCD to date and demonstrated that complete removal of dysplastic cortex is the single most important predictor of favorable seizure outcome. Seizure freedom was achieved in 70% of patients who had complete resections, but in only 22% with incomplete excision of the dysplastic region. Only 8% of subjects whose resections were judged complete showed no improvement in postoperative seizure frequency compared with 61% of patients after incomplete resections. Our results support the concept of intrinsic epileptogenicity of dysplastic lesions.5,16,26,27 It has been suggested that because FCD tissue typically colocalizes with the epileptogenic zone, structural lesion removal is both necessary and sufficient for the Neurology 72

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achievement of seizure freedom.5,19,26 Different practical approaches have been proposed to delineate dysplastic cortex. We utilized a definition of completeness of surgery based on careful evaluation of both anatomic (MRI) and physiologic (intracranial EEG) data.19,21 The reasons for incomplete resections are diverse but overlap of dysplastic and eloquent cortex was clearly the leading cause in our series. Colocalization of FCD with language or motor areas has been repeatedly demonstrated.20,28 MST have been proposed as an alternative to resection in this situation,29 but our experience with this procedure was discouraging. We suggest that a location of mMCD/FCD outside critical cortical areas facilitates their complete removal and remains the most important predictor of postsurgical outcome. Nevertheless, 22% of our patients who had incomplete surgeries are seizure-free. Similar proportions of “unexpectedly successful surgeries” were noted in other studies examining for completeness of FCD resections.6,9,13,16,17,21 It thus seems likely that the epileptogenic zone may occupy only a portion of dysplastic cortex in some FCD cases. This assumption is in agreement with a recent observation that histopathologic subtypes of FCD are not all equally epileptogenic and that no ictal onsets were found in balloon cells-containing regions.28 Seizure relief after an incomplete FCD removal remains probably the exception rather than the rule. Nevertheless, this possibility should be analyzed in detail since it might represent a background for “palliative” epilepsy surgery in cases with the overlapping epileptogenic and essential cortical areas. Unlike several previous reports, we did not find that other presurgical, surgical, and histopathologic parameters (such as early indication for surgery, circumscribed EEG and MRI abnormalities, presurgical intellectual capacities of patients, implantation of intracranial electrodes, extent and location of resections, and histopathologic subtypes of FCD) were predictors of postsurgical seizure outcome. Variations between studies might be due to differences in patient selection, use of diverse classification systems of cortical malformations, short follow-up, or an insufficient number of patients. Another possible explanation includes the diverse clinical, EEG, and imaging spectrum of FCD. We found that 75% of patients do not change seizure status after the second postoperative year. Late seizure recurrence or worsening of the condition occurred in 17% of subjects with long-term follow-up and were more frequent in FCD type II than in mMCD/FCD type I patients. On the other hand, 8% of patients improved or became seizure-free following the second 222

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postoperative year, particularly mMCD/FCD type I patients. Anticonvulsant medication was discontinued in 14% of all patients at the end of follow-up. Several studies analyzed long-term seizure outcome after surgery for FCD.6,11,18,21,22,30 Two6,21 reported the same proportion of seizure-free cases at 2 years postoperatively and at the end of long-term follow-up. Two others18,30 found that proportion of patients with favorable seizure outcomes decreased (from 84% to 70% and from 67% to 32% during a 10-year period, respectively). Only 14% of our patients were not taking medication at the end of follow-up. Other series reported that 47%18 and 32%30 of patients were off medication. Another study22 reported that after resection 59% of their FCD subjects were either on monotherapy or medication-free. These differences may reflect diverse approaches in postoperative management of patients among individual epilepsy surgery centers. At our center, AED therapy is tapered or discontinued only if there is seizure freedom for 2 or more years after the surgery, and no significant epileptiform features on the postoperative EEG. ACKNOWLEDGMENT Statistical analysis was conducted by David A. Ludwig, PhD, Department of Pediatrics, Department of Epidemiology & Public Health, Miller School of Medicine, University of Miami, FL; and Zbynek Hrncir, Department of Pediatric Neurology, Charles University, Second Medical School, Motol University Hospital, Prague, Czech Republic.

Received November 3, 2007. Accepted in final form August 5, 2008. REFERENCES 1. Wyllie E, Comair YG, Kotagal P, Bulacio J, Bingaman W, Ruggieri P. Seizure outcome after epilepsy surgery in children and adolescents. Ann Neurol 1998;44:740–748. 2. Harvey AS, Cross JH, Shinnar S, Mathern BW. ILAE Pediatric Epilepsy Surgery Survey Taskforce. Defining the spectrum of international practice in pediatric epilepsy surgery patients. Epilepsia 2008;49:146–155. 3. Palmini A, Lu¨ders HO. Classification issues in malformations caused by abnormalities of cortical development. Neurosurg Clin N Am 2002;13:1–16. 4. Palmini A, Najm I, Avanzini G, et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62(suppl 3):S2–8. 5. Chassoux F, Devaux B, Landre E, et al. Stereoelectroencephalography in focal cortical dysplasia: a 3D approach to delineating the dysplastic cortex. Brain 2000;123:1733– 1751. 6. Kloss S, Pieper T, Pannek H, Holthausen H, Tuxhorn I. Epilepsy surgery in children with focal cortical dysplasia (FCD): results of long-term seizure outcome. Neuropediatrics 2002;33:21–26. 7. Urbach H, Scheffler B, Heinrichsmeier T, et al. Focal cortical dysplasia of Taylor’s balloon cell type: a clinicopathological entity with characteristic neuroimaging and histopathological features, and favorable postsurgical outcome. Epilepsia 2002;43:33–40.

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Kral T, Clusmann H, Blumcke I, et al. Outcome of epilepsy surgery in focal cortical dysplasia. J Neurol Neurosurg Psychiatry 2003;74:183–188. Fauser S, Schulze-Bonhage A, Honegger J, et al. Focal cortical dysplasias: surgical outcome in 67 patients in relation to histological subtypes and dual pathology. Brain 2004; 127:2406–2418. Lawson JA, Birchansky S, Pacheco E, et al. Distinct clinicopathologic subtypes of cortical dysplasia of Taylor. Neurology 2005;64:55–61. Fountas KN, King DW, Meador KJ, Lee GP, Smith JR. Epilepsy in cortical dysplasia: factors affecting surgical outcome. Stereotact Funct Neurosurg 2004;82:26–30. Alexandre V Jr, Walz R, Bianchin MM, et al. Seizure outcome after surgery for epilepsy due to focal cortical dysplastic lesions. Seizure 2006;15:420–427. Hudgins RJ, Flamini JR, Palasis S, Cheng R, Burns TG, Gilreath CL. Surgical treatment of epilepsy in children caused by focal cortical dysplasia. Pediatr Neurosurg 2005; 41:70–76. Hirabayashi S, Binnie CD, Janota I, Polkey CE. Surgical treatment of epilepsy due to cortical dysplasia: clinical and EEG findings. J Neurol Neurosurg Psychiatry 1993;56: 765–770. Gambardella A, Palmini A, Andermann F, et al. Usefulness of focal rhythmic discharges on scalp EEG of patients with focal cortical dysplasia and intractable epilepsy. Electroencephalogr Clin Neurophysiol 1996;98:243–249. Francione S, Vigliano P, Tassi L, et al. Surgery for drug resistant partial epilepsy in children with focal cortical dysplasia: anatomical-clinical correlations and neurophysiological data in 10 patients. J Neurol Neurosurg Psychiatry 2003;74:1493–1501. Hong SC, Kang KS, Seo DW, et al. Surgical treatment of intractable epilepsy accompanying cortical dysplasia. J Neurosurg 2000;93:766–773. Kral T, von Lehe M, Podlogar M, et al. Focal cortical dysplasia: long-term seizure outcome after surgical treatment. J Neurol Neurosurg Psychiatry 2007;78:853–856. Paolicchi JM, Jayakar P, Dean P, et al. Predictors of outcome in pediatric epilepsy surgery. Neurology 2000;54: 642–647.

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Hader WJ, Mackay M, Otsubo H, et al. Cortical dysplastic lesions in children with intractable epilepsy: role of complete resection. J Neurosurg 2004;100:110–117. Hamiwka L, Jayakar P, Resnick T, et al. Surgery for epilepsy due to cortical malformations: ten-year follow-up. Epilepsia 2005;46:556–560. Cohen-Gadol AA, Ozduman K, Bronen RA, Kim JH, Spencer DD. Long-term outcome after epilepsy surgery for focal cortical dysplasia. J Neurosurg 2004;101: 55–65. Jayakar P, Duchowny M, Alvarez L, Resnick T. Intraictal activation in the neocortex: a marker of the epileptogenic region. Epilepsia 1994;35:489–494. Jayakar P, Duchowny M, Resnick TJ. Subdural monitoring in the evaluation of children for epilepsy surgery. J Child Neurol 1994;9(suppl 2):61–66. Turkdogan D, Duchowny M, Resnick T, Jayakar P. Subdural EEG patterns in children with Taylor-type cortical dysplasia: comparison with nondysplastic lesions. J Clin Neurophysiol 2005;22:37–42. Palmini A, Gambardella A, Andermann F, et al. Intrinsic epileptogenicity of human dysplastic cortex as suggested by corticography and surgical results. Ann Neurol 1995;37: 476–487. Otsubo H, Iida K, Oishi M, et al. Neurophysiologic findings of neuronal migration disorders: intrinsic epileptogenicity of focal cortical dysplasia on electroencephalography, electrocorticography, and magnetoencephalography. J Child Neurol 2005;20:357–363. Boonyapisit K, Najm I, et al. Epileptogenicity of focal malformations due to abnormal cortical development: direct electrocorticographic-histopathologic correlations. Epilepsia 2003;44:69–76. Hufnagel A, Zentner J, Fernandez G, Wolf HK, Schramm J, Elger CE. Multiple subpial transection for control of epileptic seizures: effectiveness and safety. Epilepsia 1997;38: 678–688. Mathern GW, Giza CC, Yudovin S, et al. Postoperative seizure control and antiepileptic drug use in pediatric epilepsy surgery patients: the UCLA experience, 1986 –1997. Epilepsia 1999;40:1740–1749.

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Identification of a possible pathogenic link between congenital long QT syndrome and epilepsy J.N. Johnson, MD* N. Hofman, MSc* C.M. Haglund G.D. Cascino, MD, FAAN A.A.M. Wilde, MD, PhD M.J. Ackerman, MD, PhD

Address correspondence and reprint requests to Dr. Michael J. Ackerman, Director, Long QT Syndrome Clinic and the Mayo Clinic Windland Smith Rice Sudden Death Genomics Laboratory, Mayo Clinic, Guggenheim 501, 200 First Street SW, Rochester, MN 55905 [email protected]

ABSTRACT

Background: Long QT syndrome (LQTS) typically presents with syncope, seizures, or sudden death. Patients with LQTS have been misdiagnosed with a seizure disorder or epilepsy and treated with antiepileptic drug (AED) medication. The gene, KCNH2, responsible for type 2 LQTS (LQT2), was cloned originally from the hippocampus and encodes a potassium channel active in hippocampal astrocytes. We sought to test the hypothesis that a “seizure phenotype” was ascribed more commonly to patients with LQT2.

Methods: Charts were reviewed for 343 consecutive, unrelated patients (232 females, average age at diagnosis 27 ⫾ 18 years, QTc 471 ⫾ 57 msec) clinically evaluated and genetically tested for LQTS from 1998 to 2006 at two large LQTS referral centers. A positive seizure phenotype was defined as the presence of either a personal or family history of seizures or history of AED therapy.

Results: A seizure phenotype was recorded in 98/343 (29%) probands. A seizure phenotype was more common in LQT2 (36/77, 47%) than LQT1 (16/72, 22%, p ⬍ 0.002) and LQT3 (7/28, 25%, p ⬍ 0.05, NS). LQT1 and LQT3 combined cohorts did not differ significantly from expected, background rates of a seizure phenotype. A personal history of seizures was more common in LQT2 (30/77, 39%) than all other subtypes of LQTS (11/106, 10%, p ⬍ 0.001). Conclusions: A diagnostic consideration of epilepsy and treatment with antiepileptic drug medications was more common in patients with LQT2. Like noncardiac organ phenotypes observed in other LQTS-susceptibility genes such as KCNQ1/deafness and SCN5A/gastrointestinal symptoms, this novel LQT2-epilepsy association raises the possibility that LQT2causing perturbations in the KCNH2-encoded potassium channel may confer susceptibility for recurrent seizure activity. Neurology® 2009;72:224–231 GLOSSARY AED ⫽ antiepileptic drug; LQT1 ⫽ type 1 LQTS; LQT2 ⫽ type 2 LQTS; LQTS ⫽ long QT syndrome; TdP ⫽ torsades de pointes.

Congenital long QT syndrome (LQTS) was first described as Jervell and Lange-Nielsen syndrome and Romano Ward syndrome in the late 1950s and early 1960s.1-3 LQTS is now understood as a collection of genetically distinct arrhythmogenic disorders resulting from genetic mutations in cardiac potassium and sodium ion channels, thus termed cardiac channelopathies.4 Recent investigations have shown that mutations in non-channel proteins can also cause LQTS.5,6 The trademark event for the patient with symptomatic LQTS is the potentially lethal ventricular dysrhythmia known as torsades de pointes (TdP).7 TdP can precipitate syncope, seizures, or sudden death, depending on whether the heart rhythm spontaneEditorial, page 208 e-Pub ahead of print on November 26, 2008, at www.neurology.org. *These two authors contributed equally to the research and writing of this article. From the Department of Pediatrics/Division of Pediatric Cardiology (J.N.J., C.M.H., M.J.A.), Department of Molecular Pharmacology and Experimental Therapeutics (C.M.H., M.J.A.), Department of Neurology/Division of Epilepsy (G.D.C.), and Department of Medicine/Division of Cardiovascular Diseases (M.J.A.), Mayo Clinic, Rochester, MN; and Departments of Clinical Genetics (N.H.) and Cardiology (A.A.M.W.), Academic Medical Center, Amsterdam, The Netherlands. Dr. Michael Ackerman’s research program was supported by the Mayo Clinic Windland Smith Rice Comprehensive Sudden Cardiac Death Program. M.J.A. is also an Established Investigator of the AHA and is supported by the NIH (HD42569). Dr. Arthur Wilde’s research program was supported by the Netherlands Heart Foundation (NHS 2000.059) and the Foundation Leducq (Grant 05 CVD, Alliance against Sudden Cardiac Death). Disclosure: Dr. Ackerman is a consultant for PGxHealth, Medtronic, and Pfizer. 224

Copyright © 2009 by AAN Enterprises, Inc.

ously reverts to normal rhythm or if the patient is defibrillated back to normal rhythm before death occurs.8,9 LQTS affects approximately 1 in 2,500 persons and patients with LQTS are often diagnosed with a seizure disorder, being fainters, or having “spells.”10 Numerous LQTS genotypespecific arrhythmogenic triggers have been identified.11,12 For example, swimming is relatively gene-specific for type 1 LQTS (LQT1)13,14 while events that occur in women during the postpartum period, as well as those triggered by auditory stimuli, often indicate the presence of type 2 LQTS (LQT2).12,15-18 The onset of TdP has been shown recently to be gene-specific as well, with LQT2 patients preferentially having pauses in cardiac rhythm prior to the onset of TdP.19 Besides the long appreciated sensorineural hearing loss observed in patients with bi-allelic KCNQ1 mutations (i.e., Jervell and LangeNielsen syndrome),5 the search for noncardiac phenotypic expression involving other organs continues. For example, patients with LQT3causing SCN5A mutations have been demonstrated to have an increased prevalence of gastrointestinal symptoms.20 In 1995, loss-of-function mutations in the 1,159 amino acid-containing alpha subunit of the rapidly activating delayed rectifying potassium channel, IKr, encoded by the human ether a go-go related gene, HERG, were discovered as the cause for LQT2.21 In addition, the Drosophila homolog of the HERG gene was found to be encoded by the aptly named seizure locus. Mutations in the seizure locus caused temperatureinduced hyperactivity followed by paralysis in Drosophila.22 Notably, the HERG gene, now annotated as KCNH2, was discovered originally in a hippocampal cDNA library in 1994, although murine ERG is expressed in other regions of the brain as well.23 Further studies reported that hippocampal expression of the ERG family of potassium channels is distributed preferentially to hippocampal astrocytes24 that may regulate neuronal excitability.24-27 Considering that the LQT2-associated HERG potassium channel is present and active in significant quantity in glial cells, and recognizing that blockage of potassium homeostasis in glial cells can be epileptogenic,24-26 it is tempt-

ing to speculate that patients with LQTS, particularly type 2 LQTS, may in fact exhibit neurally mediated seizures or epilepsy rather than simply a ventricular arrhythmia with subsequent collapse and seizure activity (i.e., torsadogenic seizures). Accordingly, we set out to test the hypothesis that either a prior personal or family history of seizure activity or treatment with antiepileptic drug (AED) medication were more common among patients with LQT2. METHODS Chart review. In this IRB-approved study, the medical records were reviewed for all unrelated patients (n ⫽ 343) clinically evaluated and genetically tested for LQTS between 1998 and 2006 at two LQTS referral centers: 1) Mayo Clinic’s Long QT Syndrome Clinic in Rochester, MN (n ⫽ 208) and 2) the Academic Medical Center, Amsterdam, The Netherlands (n ⫽ 135). The reviewers (J.N.J., N.H.) were blinded at all times to the patients’ genotypes. Classifications of patients were given based on genotype by a third author (C.M.H.) independent of the reviewers. Patients were classified as genotype positive LQTS if they had a clinically relevant genetic mutation in one of the known LQTS-susceptibility genes. Patients were classified as genotype negative LQTS if they had a clinical diagnosis of LQTS given by a physician specializing in LQTS (M.J.A., A.A.M.W.), but in whom genetic testing of the known LQTS-susceptibility genes was negative. A classification of normal was given if the patient had a negative genetic test as well as a negative clinical history for LQTS as determined by a physician specializing in LQTS (M.J.A., A.A.M.W.). A positive “seizure phenotype” was defined as the presence of either a personal or family history of seizures or epilepsy or a history of AED therapy. A family history of seizures was considered positive if the patient could name the affected family member, and if the family member shared genetic material with the proband. Family members with a history of seizures but who had married into the family were excluded. Vague histories of seizures in distant family members not clearly known by the proband and their immediate family were excluded. Only unrelated patients were reviewed to prevent the presence of a few large families from skewing the family history data. A patient was considered to have a positive treatment history if they had been placed on an AED medication for greater than 1 day, and if the medical treatment was specifically prescribed for a presumed seizure disorder. AED treatment for nonepileptic conditions (e.g., chronic pain) was not included. Given that the current standard of care for the evaluation of LQTS does not include an EEG, documented EEG evidence of epileptiform activity was not a requirement for inclusion in this study. We attempted to account for and exclude all acquired causes of seizures in the cohorts. Patients with a history of having a single seizure with documented fever were excluded. Also, patients with a history of seizures within months following a traumatic head injury (or patient’s family members with a similar history) were excluded. All other seizure histories were classified as a positive seizure phenotype.

Genetic testing. Patients in both the Mayo Clinic and Amsterdam cohorts had genetic testing performed using denaturing high performance liquid chromatography and direct DNA sequencing. For the Mayo cohort, comprehensive genetic testing Neurology 72

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Table 1

Demographic and genotype data

Cohort

Combined cohort

Mayo cohort

Amsterdam cohort

No. of patients

343

208

135

Age, y, mean ⴞ SD QTc, ms, mean ⴞ SD

27 ⫾ 18

24 ⫾ 15

32 ⫾ 22

471 ⫾ 57

467 ⫾ 50

476 ⫾ 65

p Value — 0.001* 0.15

% Female

68

68

67

% With positive “seizure phenotype”

29

24

36

0.023*

% With personal seizure history

20

13

30

⬍0.001*

% With seizure treatment history Total genotype positive patients

7 183 (53%)

4 105 (50%)

13 78 (58%)

0.94

0.004* 0.23

No. LQT1 (KCNQ1)

72

48

24

0.30

No. LQT2 (KCNH2)

77

39

38

0.06

No. LQT3 (SCN5A)

28

13

15

0.16

No. LQT5 (KCNE1)

4

4

0

0.16

No. LQT6 (KCNE2)

1

0

1

0.39

No. LQT7 (KCNJ2)

1

1

0

1

Total genotype negative patients

160

103

57

0.27

Genotype (ⴚ) phenotype (ⴙ)

70

31

39

0.002*

Genotype (ⴚ) phenotype (ⴚ)

90

72

18

⬍0.001*

Demographic data and genotype/phenotype classification of proband patients presenting to Mayo Clinic, Rochester, MN, and Academic Medical Centre, Amsterdam. p Values refer to the differences between the individual Mayo and Amsterdam cohorts. *p ⬍ 0.05.

for additional mutations in the five most common LQTSsusceptibility genes was performed routinely even after the first mutation was found, given that multiple mutations occur at a frequency of 5–10%. For the Amsterdam cohort, further genetic testing after first mutation identification was per protocol and was continued only if there was a severely prolonged QT interval or if the patient was symptomatic. The vast majority of the patients in the Amsterdam cohort of this study, however, were symptomatic, and thus had comprehensive assessment of at least KCNQ1 (LQT1), KCNH2 (LQT2), and SCN5A (LQT3), which accounts for approximately 75% of all LQTS and over 95% of the patients with genetically identifiable LQT1-12 genotypes.

Statistical analysis. Statistical analyses were performed with the assistance of the Mayo Clinic CTSA Service Center, Rochester, MN. All continuous variables were reported as the mean ⫾ SD. Proportions were analyzed and compared using a two-tailed Fisher exact test. Means were analyzed using the independent groups t test for means. QTc values were measured manually and calculated using the standard Bazett formula.28 A p value ⬍ 0.05 was considered to be significant.

Table 1 summarizes the demographics for the composite cohort and the individual Mayo Clinic and Amsterdam cohorts. Demographically, age was the sole significant difference between the two cohorts, with the Mayo cohort having a younger patient population (24 ⫾ 15 years) than the Amsterdam cohort (32 ⫾ 22 years, p ⫽ 0.001). Otherwise, there were no other significant differences found between the cohorts in terms of proportion of each ge-

RESULTS

226

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notype, QTc at diagnosis, or percent of patients with positive or negative genotypes. Overall, a positive seizure phenotype was recorded in 98/343 (29%) patients including 24% of the Mayo Clinic cohort and 36% of the Amsterdam cohort (table 1, p ⫽ 0.023). Twenty percent of the probands either had a personal history of seizures or were diagnosed clinically with epilepsy prior to the rendering of genotype proven LQTS. A personal seizure history was more common among the 135 unrelated cases in the Amsterdam cohort (30%) compared to the 208 unrelated cases in the Mayo Clinic cohort (13%, p ⬍ 0.001). In addition, 7% of the patients had received antiepileptic pharmacotherapy including 13% of the Amsterdam cohort compared to 4% of the Mayo Clinic cohort (p ⫽ 0.004). Among the 90 patients ultimately dismissed as normal (genotype negative/LQTS phenotype negative), 27% had a positive seizure phenotype, 17% had been diagnosed as having seizures, and 4% had been treated with AED medications (figure). In comparison, unrelated patients with genetically or clinically diagnosed LQTS had a similar prevalence of a positive seizure phenotype overall (31%, p ⫽ NS). However, subset analysis revealed that a positive seizure phenotype, a personal history of seizures, and a personal history of antiepileptic therapy was more common among patients with LQT2 (figure). In fact, a pos-

Figure

Prevalence of “seizure phenotype” in long QT syndrome subtypes

Percentage of patients from the combined cohort for each genotypic classification having a positive seizure phenotype (black bar), personal history of seizures (diagonal line), or history of treatment for seizures with antiepileptic drugs (AEDs, white bar). Not shown are patients with LQT5, LQT6, and LQT7 (n ⫽ 6), none of whom had a positive seizure phenotype. *Significance value of combined cohort of LQT2 patients compared to other long QT genotypes. **Significance value of combined cohort of LQT2 patients compared to the other LQTS genotypes (i.e. LQT1 and LQT3) and the genotype negative background rates. When compared to LQT3, LQT2 is more common (47% vs 25%) but did not achieve significance (p ⫽ 0.07).

itive seizure phenotype was recorded in nearly half (36/ 77) of the patients with LQT2 compared to 16/72 (22%, p ⬍ 0.002) with LQT1, 7/28 (25%, NS) with LQT3, and 15/70 (21%, p ⬍ 0.002) with genotype negative/phenotype positive LQTS. Analysis of the Mayo Clinic and Amsterdam cohorts separately indicate this increased prevalence of positive seizure phenotype in LQT2, 36% in the Mayo Clinic cohort and 58% in the Amsterdam cohort. Similarly, patients with LQT2 were far more likely to have a personal history of seizures or a preceding diagnosis of epilepsy (39%) compared to either the genotype negative/phenotype negative patients (17%) or the other subtypes of LQTS (11/106, 10%, p ⬍ 0.001, figure). This increased prevalence of personal seizures in LQT2 was evident in both cohorts independently: 26% vs 6% in the Mayo Clinic cohort (p ⬍ 0.008) and 53% vs 18% in the Amsterdam cohort (p ⬍ 0.001). A previous diagnosis of seizure disorder was more common among LQT2 patients seen in Amsterdam compared to the LQT2 patients evaluated at the Mayo Clinic (p ⬍ 0.02). Among the patients with LQT2 and a personal history of seizures (n ⫽ 30), 13 (17% of all LQT2 patients) had been treated with antiepileptic pharmacotherapy (9/20 from Amsterdam and 4/10 from Mayo Clinic) compared to only 12/266 (4.5%) non-LQT2

patients (p ⬍ 0.001) including 4% with LQT1, 0% with LQT3, 7% with genotype negative/phenotype positive LQTS, and 4% of patients that lacked both genetic and clinical evidence for a diagnosis of LQTS (figure). A summary of the mutations noted in patients who have both LQT2 and a positive seizure phenotype is shown in table 2. There were no associations seen between patients with a personal or family history of seizures and the location of the mutation in the potassium channel. DISCUSSION It is well known that the most common symptomatic triad stemming from TdP (the trademark dysrhythmia of LQTS) is syncope, seizures, or sudden death.7 Seizures have been viewed as the sequelae of prolonged cerebral hypoperfusion secondary to the cardiac dysrhythmia (i.e., torsadogenic seizures). Because these patients may be witnessed having an apparent generalized seizure, it is not uncommon for patients with LQTS to be misdiagnosed with epilepsy and treated with AED medication.7,29 Here, we demonstrate that among two independent cohorts of unrelated patients evaluated clinically and genotyped for LQTS, a seizure phenotype was far more commonly ascribed to patients with LQT2 Neurology 72

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Table 2

Mutations in patients with LQT2 and a positive “seizure phenotype”

Patient no.

Institution

History type

Exon

Nucleotide change

Coding effect

Location

1

Mayo

Personal

2

89 T⬎C

I30T

N-term

2

AMC

Personal

2

191 G⬎A

C64Y

N-term

3

Mayo

Personal

2

Del221-251

R73fs/31

N-term

4

AMC

Family

2

296 A⬎G

Y99S

N-term

5

AMC

Personal

2

296 A⬎C

Y99S

N-term

6

Mayo

Personal

4

526 C⬎T

R176W

N-term

7

AMC

Personal

4

Del578-582

A193fsX329

N-term

8

AMC

Personal

4

722 C⬎T

P241L

N-term

9

AMC

Personal

4

754delC

R252G

N-term

10

Mayo

Personal

4

916 G⬎T

G306W

N-term

11

Mayo

Personal

5

1096 C⬎T

R366X

N-term

12

Mayo

Family

5

1128 G⬎A

Q376sp

N-term

13

Mayo

Personal

6

1366 G⬎T

D456Y

S2

14

AMC

Personal

6

1501 G⬎C

D501H

S3

15

AMC

Personal

7

1600 C⬎T

R534C

S4

16

AMC

Personal

7

1674 C⬎G

A558P

S5

17

AMC

Personal

6/7

1129-2543_1678 Dup3682

I560fs/2

S5

18

AMC

Personal

7

1714 G⬎A

G572S

S5-Pore

19

AMC

Personal

7

1744 C⬎T

R582C

S5-Pore

20

AMC

Personal

7

1744 C⬎T

R582C

S5-Pore

21

Mayo

Personal

7

1838 C⬎T

T613M

Pore

22

AMC

Personal

7

1838 C⬎T

T613M

Pore

23

Mayo

Personal

7

1886 A⬎T

N629I

Pore

24

Mayo

Family

7

1898 A⬎G

N633S

Pore-S6

25

Mayo

Family

7

1918 T⬎G

F640V

S6

26

AMC

Personal

7

1920 C⬎A

F640L

S6

27

AMC

Personal

7

del1926-1934

del I642-V644

S6

28

AMC

Personal

7

1933 A⬎C

M645L

S6

29

AMC

Family

7

1945 T⬎C

S649P

S6

30

AMC

Personal

8

del2027-2028

Q676fsX721

S6-cNBD

31

AMC

Personal

8

2062 C⬎T

E698X

S6-cNBD

32

AMC

Personal

11

2616delC

P872fs877

C-term

33

Mayo

Personal

11

2626 G⬎T

E876X

C-term

34

Mayo

Family

12

2705delC

Q901fs/71

C-term

35

AMC

Personal

12

2775delG

G925fs/47

C-term

36

Mayo

Personal

13

3108insG

G1036fs/82X

C-term

Mutation data on the 36 LQT2 patients with a positive seizure phenotype presenting to either the Mayo Clinic, Rochester, MN, or the Academic Medical Centre (AMC), Amsterdam. Thirty of these patients had a personal history of seizures while six had a family history.

compared to either background rates or to patients with LQT1. A positive seizure phenotype was also more common in LQT2 compared to LQT3, but due to low total numbers of LQT3 patients, significance was not achieved. It is important to note that, while 39% of LQT2 patients had been labeled as having seizures, only a single patient with LQT3 had a personal history of seizures ( p ⬍ 0.001). 228

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Nearly half of the patients with LQT2 were classified as having either a personal or family history of seizures or history of AED therapy. In fact, patients with LQT2 were three to four times more likely to have been labeled with a personal history of seizures or formally given a diagnosis of epilepsy compared to patients without LQT2. In addition, nearly one out of five patients with LQT2 had been treated with

antiepileptic pharmacotherapy compared to less than 1 out of 20 patients with LQT1 and none of the 28 patients with LQT3. Just like previous phenotypegenotype associations that identified auditory triggers as a relatively LQT2-specific arrhythmogenic trigger18 and the postpartum period as a relatively LQT2-specific temporal period for increased risk among women,17,30 seizures/epilepsy can now be added as far more suggestive of LQT2 genotype status than any other genotype. KCNH2 was isolated originally from a hippocampal cDNA library.23 It is important to note that both the SCNA sodium channel family and KCNQ family of potassium channels have also been cloned in neuronal tissue.31-34 Indeed, it has been speculated that patients with SCN5A mutations could have an increased incidence of seizures.35 Our data only support an increased risk of seizure diagnosis in LQT2 compared to background. Neither LQT1 (KCNQ1) nor LQT3 (SCN5A) patients had a significantly greater seizure diagnosis frequency when compared to normal patients. Thus, even though there is similar neuronal expression of the two other main LQTSsusceptibility genes, LQT2 appears to be the only subtype commonly associated with a clinical expression of seizures. Intriguingly, KCNH2-encoded potassium channels are instrumental in potassium homeostasis in hippocampal glia.24 Pharmacologic studies have also shown a relationship between preventing voltagedependent potassium buffering in astrocytes and the development of epileptiform activity in the hippocampus.27,36 Voltage dependent buffering refers to the neurologic process whereby glia control the potassium concentration in the extraneuronal space in order to maintain normal neuronal conduction of action potentials.37 There has to be a constant extraneuronal potassium concentration in order for the action potential to effectively start and stop as needed. If this is perturbed, there is enhanced susceptibility for epileptic activity.36 Interestingly, blocking ERG potassium channels in astrocytes changes potassium concentrations extraneuronally,24 and such changes in potassium concentrations extraneuronally have been shown to be epileptogenic.36 Mechanistically, it is therefore conceivable that patients with LQT2 may have a decreased seizure threshold secondary to cerebral hypoperfusion stemming from a LQT2-precipitated cardiac dysrhythmia of TdP due in part to the perturbation in the neuronal KCNH2-encoded potassium channels that reside in the hippocampus. Alternatively, it is tempting to speculate that perhaps some of the “cardiac events” in patients with LQT2 are actually neurally mediated seizures secondary to the defect in the hippocampal

potassium channels encoded by KCNH2. If patients with LQT2 have a tendency for neurally mediated seizures, then KCNH2 may represent a novel candidate gene for certain types of epilepsy, particularly temporal lobe epilepsy. It should be noted that ERG potassium channelencoding genes have been shown to be expressed in numerous locations in mouse brain tissue, not limited to the hippocampus.38 Thus, epileptiform involvement of HERG mutations in humans may not only be limited to the hippocampus or temporal lobe. An alternate plausible explanation is that seizure activity could occur secondary to hypoxicischemic injury to the hippocampus from unrecognized arrhythmias. This theory would not however explain the specific predominance of seizure history in LQT2 patients compared to other LQTS patients. Unfortunately, erroneous treatment of LQTS patients with AEDs may in fact be harmful to the patient. Overall, 7% of patients in this study were treated with AED medications. Prior whole cellpatch clamp studies demonstrated that phenobarbital and phenytoin could block HERG-related currents potentially conferring susceptibility to druginduced TdP, particularly in predisposed patients.39 While this is intriguing, it is also important to note that several antiepileptic medications, including phenytoin, are in part sodium channel blockers and are potentially anti-arrhythmic. In fact, decades ago, phenytoin was commonly used pharmacotherapy for patients with LQTS. Clearly, the results from this LQT2-seizure genotype/phenotype analysis call for further cardiac and neurologic pharmacology interaction studies. Due to the retrospective study design, there exists the potential for reporting bias in the data collection process. However, the LQTS physicians (M.J.A., A.A.M.W.) solicited data regarding personal or family history of seizures or history of AED therapy uniformly and systematically in their clinical practices, and although not blinded to the patient’s genotype, the genotype was generally not established during the initial LQTS consultation. Regardless of the vantage point, a seizure phenotype was more common among patients with LQT2. However, statistical correction for multiple comparisons was not performed. In addition, we noted that the prevalence of a seizure phenotype in LQT2 was statistically greater among LQT2 patients evaluated in Amsterdam compared to those evaluated at the Mayo Clinic. It is possible that there could be an additional environmental/ethnic contribution. Considering that the patients in the Amsterdam cohort were significantly older on average than those in the Mayo cohort, it is also possible that the Amsterdam cohort simply had Neurology 72

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more life-years to have experienced a personal or family history of seizures. Regardless, the greater frequency of a personal history of seizures or prior diagnosis of epilepsy and prior treatment with AEDs among patients with LQT2 was completely concordant between both LQTS centers. Due to the acute and unpredictable onset of clinical events in patients with LQTS, EEGs during events were unavailable to the authors. A postictal EEG was not available either. This is not surprising, however, as an EEG is not viewed as standard of care in the clinical evaluation of patients with LQTS. Certainly, functional proof of neuronally mediated seizure activity in LQT2 patients while in normal sinus rhythm would lend credence to one of the postulated mechanisms. As stated previously, those labeled with a seizure phenotype in this study, in the absence of EEG evidence, may simply have expressed seizure activity following their cardiac-mediated syncopal episode. Regardless of the underlying mechanism, however, two independent cohorts have evidenced a proclivity for seizure labeling among patients with mutations in the KCNH2-encoded, cardiac/ neuronal-expressed potassium channel (LQT2). Just as it is debated whether all patients evaluated for generalized epilepsy should have a screening electrocardiogram, this new genotype–phenotype association will likely prompt a new dialogue as to whether patients with LQTS, particularly LQT2, should have a wake- and sleep-deprived EEG. ACKNOWLEDGMENT

8. 9.

10.

11.

12.

13.

14.

15.

16.

17.

18.

The investigators thank the patients who sought clinical evaluation at the respective LQTS clinics.

Received January 7, 2008. Accepted in final form July 16, 2008. 19. REFERENCES 1. Jervell A, Lange-Nielsen F. Congenital deaf-mutism, functional heart disease with prolongation of the Q-T interval and sudden death. Am Heart J 1957;54:59–68. 2. Romano C, Gemme G, Pongiglione R. Rare cardiac arrhythmias of the pediatric age: II: syncopal attacks due to paroxysmal ventricular fibrillation (presentation of 1st case in Italian pediatric literature) Clin Pediatr (Bologna) 1963;45:656–683. 3. Ward OC. A new familial cardiac syndrome in children. J Ir Med Assoc 1964;54:103–106. 4. Ackerman M.J. The long QT syndrome: ion channel diseases of the heart. Mayo Clin Proc 1998;73:250– 269. 5. Vatta M, Ackerman MJ, Ye B, et al. Mutant caveolin-3 induces persistent late sodium current and is associated with long-QT syndrome. Circulation 2006;114:2104–2112. 6. Mohler PJ, Schott JJ, Gramolini AO, et al. Ankyrin-B mutation causes type 4 long-QT cardiac arrhythmia and sudden cardiac death. Nature 2003;421:634–639. 7. Ackerman M.J: Cardiac channelopathies: it’s in the genes. Nat Med 2004;10:463–464. 230

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Incidence of acquired demyelination of the CNS in Canadian children

B. Banwell, MD J. Kennedy, MSc D. Sadovnick, PhD D.L. Arnold, MD S. Magalhaes, MSc K. Wambera, MD M.B. Connolly, MB, BCh J. Yager, MD J.K. Mah, MD N. Shah, MD G. Sebire, MD B. Meaney, MD M.-E. Dilenge, MD A. Lortie, MD S. Whiting, MD A. Doja, MD S. Levin, MD E.A. MacDonald, MD D. Meek, MD E. Wood, MD N. Lowry, MD D. Buckley, MD C. Yim, MD M. Awuku, MD C. Guimond, MSc P. Cooper, MD F. Grand’Maison, MD J.B. Baird, MD V. Bhan, MD A. Bar-Or, MD

Address correspondence and reprint requests to Dr. Brenda Banwell, Research Institute, The Hospital for Sick Children, 555 University Ave., Toronto, Ontario, M5G 1X8 Canada [email protected]

ABSTRACT

Background: The incidence of acquired demyelination of the CNS (acquired demyelinating syndromes [ADS]) in children is unknown. It is important that physicians recognize the features of ADS to facilitate care and to appreciate the future risk of multiple sclerosis (MS).

Objective: To determine the incidence, clinical features, familial autoimmune history, and acute management of Canadian children with ADS.

Methods: Incidence and case-specific data were obtained through the Canadian Pediatric Surveillance Program from April 1, 2004, to March 31, 2007. Before study initiation, a survey was sent to all pediatric health care providers to determine awareness of MS as a potential outcome of ADS in children. Results: Two hundred nineteen children with ADS (mean age 10.5 years, range 0.66 –18.0 years; female to male ratio 1.09:1) were reported. The most common presentations were optic neuritis (ON; n ⫽ 51, 23%), acute disseminated encephalomyelitis (ADEM; n ⫽ 49, 22%), and transverse myelitis (TM; n ⫽ 48, 22%). Children with ADEM were more likely to be younger than 10 years, whereas children with monolesional ADS (ON, TM, other) were more likely to be older than 10 years (p ⬍ 0.001). There were 73 incident cases per year, leading to an annual incidence of 0.9 per 100,000 Canadian children. A family history of MS was reported in 8%. Before study initiation, 65% of physicians indicated that they considered MS as a possible outcome of ADS in children. This increased to 74% in year 1, 81% in year 2, and 87% in year 3.

Conclusion: The incidence of pediatric acquired demyelinating syndromes (ADS) is 0.9 per 100,000 Canadian children. ADS presentations are influenced by age. Neurology® 2009;72:232–239 GLOSSARY ADEM ⫽ acute disseminated encephalomyelitis; ADS ⫽ acquired demyelinating syndromes; CI ⫽ confidence interval; CPSP ⫽ Canadian Pediatric Surveillance Program; mono-ADS ⫽ monolesional presentation; MS ⫽ multiple sclerosis; ON ⫽ optic neuritis; OR ⫽ odds ratio; poly-ADS ⫽ polylesional presentation; TM ⫽ transverse myelitis.

Multiple sclerosis (MS), characterized by relapses of neurologic dysfunction and MRI evidence of inflammatory lesions, is a common cause of neurologic disability among Canadian young adults. The onset and diagnosis of MS in children and adolescents is being increasingly recognized.1 MS diagnosis requires evidence of dissemination of inflammatory demyelination within the CNS and over time. A first attack of demyelination, or an acquired demyelinating syndrome (ADS), may occur as a transient illness or may represent the first attack of MS. ADS can be subdivided clinically into acute disseminated encephalomyelitis (ADEM), characterized by polyfocal deficits and encephalopathy; and clinically isolated syndromes, monofocal or polyfocal deficits without encephalopathy.2 The incidence of ADS in children is largely unknown, particularly because available studies tend to focus on only specific ADS presentations, such as ADEM,3,4 transverse myelitis (TM),5 or

Authors’ affiliations are listed at the end of the article. This study was funded by the Multiple Sclerosis Society of Canada and the Multiple Sclerosis Scientific Research Foundation and was performed in conjunction with the Canadian Paediatric Society Surveillance Program. Disclosure: The authors report no disclosures. 232

Copyright © 2009 by AAN Enterprises, Inc.

optic neuritis (ON).6,7 Prospective studies of children with ADS have shown that 16% to 25% are ultimately diagnosed with MS before age 18 years,8-10 emphasizing the importance of considering MS risk in this population. We defined the incidence and clinical and demographic features of ADS in Canadian children. Data were acquired through the Canadian Pediatric Surveillance Program (CPSP), an innovative national program designed to increase awareness of childhood disorders among Canadian pediatricians and pediatric subspecialists. Since its inception in 1996, the CPSP has provided surveillance of more than 34 pediatric disorders, including several neurologic conditions,11,12 and has facilitated knowledge translation through annual publications.13 METHODS Prestudy questionnaire and education materials. Before initiation of the 3-year surveillance study, a detailed educational overview of pediatric ADS, including definitions for each of the clinical presentations, and a prestudy questionnaire were mailed to 2,445 Canadian pediatric health care providers. The questionnaire consisted of the following three questions: 1. In the past 2 years, have you cared for any patient less than 18 years of age with acquired demyelination of the CNS? 2. Did you consider the possibility that the child might develop recurrent demyelination such as MS? 3. Were any of these children subsequently diagnosed with MS?

CPSP program. The CPSP uses a two-tiered reporting system. Each month, an initial reporting form is mailed to approximately 2,400 pediatric health care providers. A detailed reporting form is completed for each patient identified. Summaries of reporting practices and study highlights are published annually.14-16

Surveillance study. From April 1, 2004, to March 31, 2007, Canadian pediatricians and pediatric subspecialists (including child neurologists and pediatric ophthalmologists) received a monthly questionnaire asking them to reply with the age and sex of any child with ADS, or to indicate that they had not seen patients meeting the inclusion criteria during the month. A detailed reporting form was then mailed to all respondents asking the following: clinical features (a predesigned list was included on the reporting form), age at ADS, sex, country of birth, country of birth of the parents, history of immunization 1 month before ADS onset, family history of MS or other autoimmune disease in first and second generation relatives, presence or absence of white matter lesions on brain MRI, acute therapies, and duration of therapy. Physicians were asked whether they had considered the possibility of MS and whether they had discussed this possibility with the child and family. Inclusion and exclusion criteria. Patients had to be younger than 18 years, had to be Canadian residents, and had to have had no prior history of CNS demyelination. For inclusion, at least one of the following was required:

1. ON: defined by acute or subacute visual loss, typically associated with a relative afferent pupillary defect, restricted visual fields, pain with ocular movement, and with optic nerve swelling, abnormal signal or enhancement on CT or MRI of the orbits17. 2. TM: defined by weakness of the limbs, typically associated with a defined spinal sensory level, bladder or bowel dysfunction, and MRI evidence of spinal cord swelling, increased signal, or enhancement; spinal cord compression by extrinsic or intrinsic lesions was excluded.18 3. ADEM: defined by polylesional neurologic deficits, accompanied by encephalopathy (excessive irritability, somnolence, or coma).2 4. Monolesional demyelination (mono-ADS other): defined by neurologic deficits referable to a single CNS site distinct from the optic nerve or spinal cord. 5. Polylesional demyelination (poly-ADS): defined by multiple neurologic deficits implicating concurrent involvement of more than one site in the CNS in the absence of encephalopathy. The CPSP identified potential duplicate reports through careful screening of city and month of reporting, and age (in years) and sex of identified patients. Reporting physicians were contacted directly, duplicate cases were excluded, and only the primary physician involved in the care of the child completed the detailed reporting form. One author (B.B.) reviewed all detailed reporting forms to ensure that each patient met inclusion criteria, and to categorize the clinical presentation as ON, TM, ADEM, mono-ADS other, or poly-ADS. It was specifically noted whether children experienced fever or seizures during their ADS presentation. Children were excluded if they had culture-proven bacterial meningitis, viral encephalitis, demyelination of the peripheral nervous system (i.e., Guillain-Barre´ syndrome, chronic inflammatory demyelinating polyneuropathy), biochemical or radiologic suspicion of inherited or genetically defined leukodystrophy, metabolic, or mitochondrial disease, systemic and laboratory features suggestive of systemic lupus erythematosus or connective tissue disease, or radiation- or chemotherapy-associated white matter damage. Exclusion was determined by the reporting physician.

Statistical analyses. Descriptive statistics summarize clinical and demographic data. Comparisons across clinical presentations (categorized as ADEM, mono-ADS [which included TM, ON, and mono-ADS other], and poly-ADS) with respect to age (younger than 10 years and older than 10 years) were performed using ␹2 tests. This method also assessed the influence of sex and of season among the most common clinical presentations: ADEM, ON, and TM. Annual ADS incidence rates were calculated using Statistics Canada’s 2006 Census data19 and were compared using Poisson regression. Logistic regression, using a backward elimination selection procedure with a significance level for removal of 0.10, explored potential predictors (sex, age, clinical presentation, MRI evidence of white matter lesions) of reporting physicians’ decisions to prescribe treatment for the ADS event. RESULTS Prestudy questionnaire. Of the 2,445 pediatric health care providers, 611 (25%) completed and returned the questionnaire. Of these 611, 21% (130 physicians) reported having seen one or more children with CNS demyelination within the 2 years Neurology 72

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Table

returned (n ⫽ 10), or the child already met criteria for pediatric MS (n ⫽ 17). Clinical and demographic data were analyzed for 219 children with confirmed ADS (table).

Demographic characteristics

Age at demyelination, mean ⴞ SD (range), y

10.5 ⴞ 4.6 (0.66 –17.96)

Age as a function of clinical presentation, mean ⴞ SD (range), y ADEM (n ⴝ 49)

Incidence. The incidence of ADS in Canadian chil-

7.7 ⴞ 4.4 (0.9 –16.26)

Monolesional ADS TM (n ⴝ 49)

11.01 ⴞ 5.1 (0.66 –17.35)

ON (n ⴝ 51)

11.5 ⴞ 3.2 (3.19 –16.94)

Mono-ADS other (n ⴝ 24)

11.7 ⴞ 4.7 (1–17.91)

Polylesional ADS Poly-ADS (n ⴝ 35)

11.4 ⴞ 4.6 (2.17–17.96)

Both TM and ON (n ⴝ 8)

11.3 ⴞ 4.5 (4.81–17.5)

Sex, F:M

1.09:1.00

Child’s country of birth, n (%) Canada

193 (88)

Other

17 (8)

Not indicated

9 (4)

Parents’ country of birth, n (%) Canada Other Unknown Not indicated Ethnicity, n (%) European

Mother

Father

125 (57)

132 (62)

71 (32)

66 (30)

5 (2)

5 (2)

18 (8)

6 (7)

ADEM (n ⴝ 49)

ON (n ⴝ 51)

TM (n ⴝ 49)

33 (68)

29 (56)

30 (61)

Asian

6 (12)

5 (10)

5 (11)

African

1 (2)

0 (0)

0 (0)

Middle Eastern

1 (2)

1 (2)

1 (2)

Caribbean

1 (2)

1 (2)

1 (2)

South American

0 (0)

5 (10)

2 (4)

Aboriginal

0 (0)

2 (4)

3 (6)

Mixed

2 (4)

6 (12)

4 (8)

Not provided

5 (10)

2 (4)

3 (6)

Family history of autoimmune disease, n (%) Multiple sclerosis

17 (8)

Juvenile diabetes

9 (4)

Systemic lupus erythematosus

4 (2)

Thyroiditis

9 (4)

ADEM ⫽ acute disseminated encephalomyelitis; mono-ADS ⫽ monolesional presentation; TM ⫽ transverse myelitis; ON ⫽ optic neuritis; poly-ADS ⫽ polylesional presentation.

preceding the prestudy questionnaire, and 85 physicians stated that they had considered the diagnosis of MS and had discussed this possibility with the child and family. Surveillance study. Initial reporting forms were re-

ceived from 2,005 of 2,445 health care providers (82%) in year 1, 2,107 of 2,570 (82%) in year 2, and 2,033 of 2,606 (78%) in year 3. A total of 261 children were reported. Of these, 42 were excluded because they did not meet the inclusion criteria or their date of demyelination did not fall within the study period (n ⫽ 15), detailed reporting forms were not 234

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dren was calculated using national and provincial population data19 (figure 1). The annual average incidence was 0.9 per 100,000 Canadian children (95% confidence interval [CI] 0.8 –1.1/100,000) (year 1: 0.9/100,000 Canadian children [95% CI 0.7–1.1/ 100,000]; year 2: 1.0/100,000 [95% CI 0.9 –1.3/ 100,000]; year 3: 0.8/100,000 [95% CI 0.6 – 1.0/100,000]; p ⫽ 0.20). The lower rates in Quebec (0.6/100,000, 95% CI 0.3–1.1/100,000) and Saskatchewan (0.6/100,000, 95% CI 0.04 –2.4/ 100,000) and higher rate in Manitoba (1.6/100,000, 95% CI 0.5–3.7/100,000) were not significantly different. In Manitoba, one regional pediatric health center serves the majority of the children in the province. This center was actively involved in our national prospective study of demyelination, increasing the likelihood of case ascertainment. In addition to ascertainment issues, it is possible that unidentified clinical, demographic, or environmental factors may have influenced the higher rate in Manitoba and lower rates in Quebec and Saskatchewan; further studies are required to address this point. No children with ADS were identified in Prince Edward Island, Yukon, Northwest Territories, or Nunavut, likely because of the low pediatric population of these regions (34,214 resident children in PEI, 13,563 in the Yukon, 13,606 in the Northwest Territories, and 8,026 in Nunavut). The three most common clinical ADS presentations had similar incidence rates (ADEM: 0.2/100,000 Canadian children [95% CI 0.15– 0.3/100,000]; TM: 0.2/100,000 [95% CI 0.15– 0.3/100,000]; ON: 0.2/100,000 [95% CI 0.16 – 0.3/100,000]). Demographic characteristics. The female:male ratio was 1.09:1. The sex ratio remained consistent when evaluated as a function of age: 1.07:1 in children younger than 10 years, and 1.10:1 in children aged 10 years or older. Figure 2 displays the frequency of ADS as a function of age at presentation (median 11.1 years, range 0.5–18 years). Although all children were Canadian residents, 8% were born outside of Canada and 37% were firstgeneration Canadians (both parents born outside of Canada). The relative representation of ethnicities did not differ when evaluated as a function of the more common ADS presentations (ADEM, ON, or TM), and a family history of MS was reported in

Figure 1

Provincial incidence of acquired demyelinating syndromes in Canadian children (per 100,000; averaged over the 3 year study)

P.E.I. ⫽ Prince Edward Island; Sask. ⫽ Saskatchewan.

8%, juvenile-onset type 1 diabetes in 4%, thyroiditis in 4%, and systemic lupus in 2% (table). Clinical presentations. The table describes the mean

age for each of the ADS presentations. Children with ADEM were younger than children with other clinical presentations (p ⬍ 0.0001). Figure 3 delineates the relative frequency of ON, TM, ADEM, monoFigure 2

Distribution of acquired demyelinating syndromes (ADS) as a function of age

ADS other, and poly-ADS for the entire cohort and as a function of age at presentation (younger than 10 years, or aged 10 years and older). Children with ADEM were more likely to present when younger than 10 years, whereas children experiencing monolesionalADS (including ON, TM and other) were more likely to be older than 10 years (p ⬍ 0.001). The female:male ratio was highest for children with ON (1.43:1), whereas males were more likely to present with TM (0.85:1) and ADEM (0.81:1), although these relationships were not significant. The incidence of the three most common clinical presentations (ON, TM, ADEM) did not differ according to season (ON: p ⫽ 0.46; TM: p ⫽ 0.88; ADEM: p ⫽ 0.64) (figure 4). Nine children (4%) received vaccinations within 1 month of ADS onset: hepatitis B vaccine (n ⫽ 2), influenza vaccine (n ⫽ 1), measles, mumps, and rubella vaccine (n ⫽ 1), Gardasil (n ⫽ 1), pertussis vaccine (n ⫽ 1), Adacel (n ⫽ 1), unknown (n ⫽ 2). Fever was reported in 49 of 217 children (23%): 29 with ADEM, 3 with ON, 10 with TM, and 7 with polylesional ADS. Seizures occurred in 15 of 217 children (7%): 10 with ADEM, 1 with ON, 1 with TM, 2 with monolesional ADS other, and 1 with polylesional ADS. Detailed reporting forms for 2 patients did not indicate the presence or absence of fever or seizures. Neurology 72

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Figure 3

Relative proportion of children with each of the ADS presentations and representation of these ADS presentations as a function of age younger or older than 10 years

nisolone and 62% received oral prednisone (likely as an oral taper after IV therapy, although this was not specified), 17% received IV immunoglobulin, and 8% received other therapies. The presence of abnormal white matter on MRI was an important predictor of reporting physicians’ decisions to prescribe treatment for ADS (odds ratio [OR] ⫽ 2.23, p ⫽ 0.07). Clinical presentation (OR ⫽ 1.0, p ⫽ 0.98), age (OR ⫽ 0.8, p ⫽ 0.68), and sex (OR ⫽ 1.2, p ⫽ 0.62) did not emerge as predictors of the likelihood of treatment. Of the 33 children for whom corticosteroid therapy was not prescribed, 13 had ON, 4 had ADEM, 3 had TM, and 13 had mono-ADS (at sites extrinsic to the optic nerves or spinal cord) or poly-ADS. Data on clinical severity was not collected, likely an important additional predictor of physician decision to treat. Discussion about risk of recurrent demyelination. Cli-

nicians discussed recurrent demyelination and the potential for a future diagnosis of MS with 181 families (83% of all cases). When stratified by year of study, discussion of MS risk occurred with 74% of the families in study year 1, 85% in year 2, and 88% in year 3.

ADEM ⫽ acute disseminated encephalomyelitis; TM ⫽ transverse myelitis; ON ⫽ optic neuritis; ADS ⫽ acquired demyelinating syndromes; mono-ADS ⫽ monolesional presentation; poly-ADS ⫽ polylesional presentation.

MRI was performed in 208 children (95%) and was reported to be abnormal in 82% (n ⫽ 171). MRI evidence of white matter lesions occurred in more than 90% of children with demyelination referable to the brain but in only 40% of children with clinically isolated ON. MRI was reported as abnormal in 91% of children with TM, but the questionnaire did not specify whether this referred to MRI of the spine only. Treatment. Therapy was prescribed for more than 82% (180 children) at the time of their ADS presentation. Of these, 92% received IV methylpred236

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DISCUSSION Acquired inflammatory demyelination of the CNS affects 0.9 per 100,000 Canadian children per year, and there seems to be no seasonal influence. Incidence rates were similar for the most common clinical presentations, ADEM (0.2/ 100,000 per year), ON (0.2/100,000 per year), and TM (0.2/100,000 per year). A family history of MS in first- or second-degree relatives was reported by only 8% of the ADS patients. Although the frequency of familial MS in kindreds of adults with an initial demyelinating event is unknown, a family history of MS has been documented in 19.8% of 1,044 adult MS patients surveyed from a Canadian MS clinic.20 Familial MS rates in pediatric MS or ADS kindreds may be underestimated because family members are young and may still develop demyelination in the future. Families of children with ADS also reported other autoimmune disease (juvenile diabetes, 4%; thyroiditis, 4%; and systemic lupus erythematosus, 2%). A study of familial autoimmunity in Canadian adult MS patients reported a lower rate of juvenile diabetes (0.4%) but did not explore thyroiditis or lupus.21 A study in the United Kingdom did not reveal any increase in autoimmunity in MS families relative to families of healthy controls.22 We are unaware of any study exploring the frequency of autoimmune disease in relatives of healthy Canadian children. Of interest was the observation that 37% of children with ADS were first-generation Canadians, a

Figure 4

Seasonal distribution of acute disseminated encephalomyelitis, transverse myelitis, and optic neuritis

No specific seasonal effect is noted (ON: p ⫽ 0.46; TM: p ⫽ 0.88; ADEM: p ⫽ 0.64). ADS ⫽ acquired demyelinating syndromes; ADEM ⫽ acute disseminated encephalomyelitis; TM ⫽ transverse myelitis; ON ⫽ optic neuritis.

finding similar to our prior analysis of ethnicity in a pediatric population from Toronto, Canada.23 Unlike studies of pediatric MS where the sex ratio clearly demonstrates a female preponderance after puberty,1,24 the sex ratio for ADS in children did not change as a function of age. The absence of an agerelated female preponderance may relate to the variable MS risk associated with different ADS presentations. Male patients more commonly presented with ADEM and TM, clinical presentations considered less likely to be the harbinger of MS (reviewed in reference 1). Specific paraclinical features may be more predictive of MS risk than demographic variables. In our prior work, we reported that 68% of children with ON, accompanied by MRI evidence of at least one white matter lesion in the brain, will be diagnosed with MS within 2 years.25 Other studies of ON in children have reported subsequent MS diagnosis rates between 26% and 56%, although MRI features were variably reported.7,17 Of the 48 children with ON in the current study, 40% were reported to have an abnormal MRI and thus represent a subgroup considered to be at particularly high risk for a future diagnosis of MS. Another important potential influence on clinical presentation is age at ADS onset. Of the 219 children, 95 were younger than 10 years. As expected, an ADEM presentation was particularly prominent in this age group (37%). Whether the polylesional features, global impairment of alertness (encephalopathy), and widespread MRI lesion pattern

characteristic of ADEM delineate a unique pathobiology or an age-related propensity for heightened inflammation is an important area for future research. Our survey did not directly evaluate the physical or cognitive morbidity of ADS, and children with mild ADS symptoms may be underreported. However, corticosteroid therapies were prescribed for more than 80% of children, suggesting a relatively high degree of clinical acuity. The impact of ADS on the long-term health of Canadian children is the subject of ongoing research. To date, incidence studies of ADS in children have focused on ADEM or established MS.3,4 A study in San Diego County, California, used retrospective and prospective data from three regional centers, included persons younger than 20 years, and defined ADEM as any acute first demyelinating event associated with neuroimaging evidence of demyelination.4 The reported incidence of 0.4 per 100,000 may seem higher than the incidence of ADEM (0.2 per 100,000) in our study. However, applying their definition for ADEM to our population (n ⫽ 171 children with ADS and abnormal MRI) would actually yield a higher incidence of 0.7 per 100,000 (although we cannot comment on incidence in Canadians aged 18 –20 years). The incidence of ADEM reported in a national survey of German children (younger than 16 years) was 0.07 per 100,000.3 Although we included children up to age 18 years, we identified only two adolescents with ADEM between age 16 and 18 years. Thus, although our data suggest that ADEM may be more common in Canadian children, multinational collaborative studies with unified methodologies are required to validate this observation. Our study is strengthened by standardized clinical inclusion criteria and demographic collection forms. Nonetheless, we acknowledge the limitations of survey-based methodologies as response rate directly influences estimated disease incidence. Given that 78% to 82% of surveyed physicians completed the monthly forms, a response rate similar to other CPSP initiatives,26 we believe that our data represent at least an 80% estimate of the incidence of ADS in Canadian children. Important to our work was the education of pediatric health care providers on the potential risk of MS. The prestudy survey indicated that only 65% had considered the possibility of MS in their pediatric ADS patients. By study completion, more than 85% of reporting physicians stated that they had considered MS as a possible outcome of ADS in children. Although the increase in physician consideration of MS risk may reflect the fact that the initial survey queried physicians on prior care practices, and Neurology 72

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the subsequent survey was posed to respondents actively engaged in the care of children with ADS, we believe that the increase reflects the effectiveness of the educational materials provided to clinicians during the study. Our national, 23-site, prospective, Canadian study of ADS in children is now under way to define MS risk, as well as the clinical, demographic, immunologic, and neuroimaging features predictive of this outcome. AUTHORS’ AFFILIATIONS From The Hospital for Sick Children (B.B., J.K., S.M.), Toronto, Ontario; University of British Columbia (C.G., D.S.), Vancouver, British Columbia; Vancouver Coastal Health Authority (C.G., D.S.), UBC Hospital, British Columbia; Montreal Neurological Institute (D.L.A., A.B.O.), Quebec; McGill University (D.L.A., A.B.-O.), Montreal, Quebec; Victoria General Hospital (K.W.), British Columbia; Children’s Hospital of British Columbia (M.B.C.), Vancouver, British Columbia; Stollery Children’s Hospital (J.Y.), Edmonton, Alberta; Alberta Children’s Hospital (J.K.M.), Calgary, Alberta; The Children’s Hospital of Winnipeg (N.S.), Manitoba; Centre Hospitalier Universitaire de Sherbrooke (G.S.), Quebec; McMaster Children’s Hospital (B.M.), Hamilton, Ontario; The Montreal Children’s Hospital (M.-E.D.), Quebec; Centre Hospitalier Universitaire de Sainte-Justine (A.L.), Montreal, Quebec; Children’s Hospital of Eastern Ontario (S.W., A.D.), Ottawa, Ontario; Children’s Hospital of Western Ontario (S.L.), London, Ontario; Queen’s University (E.A.M.), Kingston, Ontario; Saint John Regional Hospital (D.M.), New Brunswick; IWK Health Centre (E.W.), Halifax, Nova Scotia; Royal University Hospital (N.L.), Saskatoon, Saskatchewan; Janeway Children’s Health and Rehabilitation Centre (D.B.), St. John’s, Newfoundland; Trillium Health Centre (C.Y.), Mississauga, Ontario; Windsor Regional Hospital (M.A.), Ontario; Rouge Valley–Centenary Hospital (P.C.), Scarborough, Ontario; Hoˆpital Charles LeMoyne (F.G.), Montreal, Quebec; Sudbury Regional Hospital (J.B.B.), Ontario; and Dalhousie MS Research Unit (V.B.), Halifax, Nova Scotia, Canada.

AUTHOR CONTRIBUTIONS

4.

5.

6.

7.

8.

9. 10.

11.

12.

13.

Statistical analyses were performed by S.M. and J.K.

ACKNOWLEDGMENT The authors thank all of the Canadian pediatric health care providers without whom this study could not have been performed. The authors also thank all of the 23 site coordinators: Courtney Fairbrother, Shelia Kent, Marjorie Berg, Catherine Riddell, Natarie Liu, Joan Kupchak, Christian Houde, Fabiola Breault, Stephanie Pellerin, Heather Davies, Heather Neuman, Laurie Wyllie, Chantal Horth, Mala Ramu, Edythe Smith, Alison Crowell, Doris Newmeyer, Vee McBride, Sharon J. Penney, Leanne Montgomery, Loris Aro, Julie Lafrenie`re, Louise Roberts, Laurie Robson, Trudy Campbell, Lucy Sagar, and Nancy Cacciotti; as well as Jennifer Hamilton and Melissa McGowan for their invaluable assistance.

Received May 12, 2008. Accepted in final form October 13, 2008. REFERENCES 1. Banwell B, Ghezzi A, Bar-Or A, Mikaeloff Y, Tardieu M. Multiple sclerosis in children: clinical diagnosis, therapeutic strategies, and future directions. Lancet Neurol 2007;6: 887–902. 2. Krupp L, Banwell B, Tenembaum S; for the International Pediatric MS Study Group. Consensus definitions proposed for pediatric multiple sclerosis. Neurology 2007;68: S7–S12. 3. Pohl D, Hennemuth I, von Kries R, Hanefeld F. Paediatric multiple sclerosis and acute disseminated encephalomy238

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Ramagopalan SV, Dyment DA, Valdar W, et al. Autoimmune disease in families with multiple sclerosis: a populationbased study. Lancet Neurol 2007;6:604–610. 22. Broadley SA, Deans J, Sawcer SJ, Clayton D, Compston DA. Autoimmune disease in first-degree relatives of patients with multiple sclerosis: a UK survey. Brain 2000; 123(pt 6):1102–1111. 23. Kennedy J, O’Connor P, Sadovnick AD, Perara M, Yee I, Banwell B. Age at onset of multiple sclerosis may be influenced by place of residence during childhood

rather than ancestry. Neuroepidemiology 2006;26:162– 167. 24. Ghezzi A, Deplano V, Faroni J, et al. Multiple sclerosis in childhood: clinical features of 149 cases. Mult Scler 1997;3:43–46. 25. Wilejto M, Shroff M, Buncic JR, Kennedy J, Goia C, Banwell B. The clinical features, MRI findings, and outcome of optic neuritis in children. Neurology 2006;67:258–262. 26. Health Canada. Evaluation of the Canadian Paediatric Surveillance Program. Canada Communicable Disease Report 30S2. Ottawa, Ontario; 2004.

Editor’s Note to Authors and Readers: Levels of Evidence coming to Neurology® Effective January 15, 2009, authors submitting Articles or Clinical/Scientific Notes to Neurology® that report on clinical therapeutic studies must state the study type, the primary research question(s), and the classification of level of evidence assigned to each question based on the classification scheme requirements shown below (left). While the authors will initially assign a level of evidence, the final level will be adjudicated by an independent team prior to publication. Ultimately, these levels can be translated into classes of recommendations for clinical care, as shown below (right). For more information, please access the articles and the editorial on the use of classification of levels of evidence published in Neurology.1-3 REFERENCES 1. French J, Gronseth G. Lost in a jungle of evidence: we need a compass. Neurology 2008;71:1634 –1638. 2. Gronseth G, French J. Practice parameters and technology assessments: what they are, what they are not, and why you should care. Neurology 2008;71:1639 –1643. 3. Gross RA, Johnston KC. Levels of evidence: taking Neurology® to the next level. Neurology 2008;72:8 –10.

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FBXO7 mutations cause autosomal recessive, early-onset parkinsonianpyramidal syndrome A. Di Fonzo, MD M.C.J. Dekker, MD P. Montagna, MD A. Baruzzi, MD E.H. Yonova, BSc L. Correia Guedes, MD A. Szczerbinska, BSc T. Zhao, BSc L.O.M. DubbelHulsman C.H. Wouters, PhD E. de Graaff, PhD W.J.G. Oyen, MD E.J. Simons G.J. Breedveld B.A. Oostra, PhD M.W. Horstink, MD V. Bonifati, MD

Address correspondence and reprint requests to Dr. Vincenzo Bonifati, Department of Clinical Genetics, Erasmus MC, P.O. Box 2040, 3000 CA Rotterdam, The Netherlands [email protected]

ABSTRACT

Background: The combination of early-onset, progressive parkinsonism with pyramidal tract signs has been known as pallido-pyramidal or parkinsonian-pyramidal syndrome since the first description by Davison in 1954. Very recently, a locus was mapped in a single family with an overlapping phenotype, and an FBXO7 gene mutation was nominated as the likely disease cause.

Methods: We performed clinical and genetic studies in two families with early-onset, progressive parkinsonism and pyramidal tract signs. Results: An FBXO7 homozygous truncating mutation (Arg498Stop) was found in an Italian family, while compound heterozygous mutations (a splice-site IVS7 ⫹ 1G/T mutation and a missense Thr22Met mutation) were present in a Dutch family. We also found evidence of expression of novel normal splice-variants of FBXO7. The phenotype associated with FBXO7 mutations consisted of early-onset, progressive parkinsonism and pyramidal tract signs, thereby matching clinically the pallido-pyramidal syndrome of Davison. The parkinsonism exhibits varying degrees of levodopa responsiveness in different patients.

Conclusions: We conclusively show that recessive FBXO7 mutations cause progressive neurodegeneration with extrapyramidal and pyramidal system involvement, delineating a novel genetically defined entity that we propose to designate as PARK15. Understanding how FBXO7 mutations cause disease will shed further light on the molecular mechanisms of neurodegeneration, with potential implications also for more common forms of parkinsonism, such as Parkinson disease and multiple system atrophy. Neurology® 2009;72:240–245 GLOSSARY DaTSCAN-SPECT ⫽ [123I]ioflupane single photon emission computed tomography; EMG/ENG ⫽ electromyography/electroneurography; IBZM-SPECT ⫽ [123I]iodobenzamide single photon emission computed tomography; ORF ⫽ open reading frame; PRR ⫽ proline-rich region; TMS ⫽ transcranial magnetic stimulation for the study of the motor pathways.

In 1954, Charles Davison described five patients with juvenile parkinsonism and pyramidal tract signs. In one case, necropsy disclosed lesions of the pallidum, ansa lenticularis, substantia nigra, and pyramidal tract.1 The term “pallido-pyramidal disease,” proposed by Davison for this entity, was adopted in the subsequent literature and textbooks2,3 [OMIM accession number: 260300]. Similar cases have been repeatedly reported,4-8 including several from India.9-12 The early onset, the type of familial aggregation, and frequent parental consanguinity suggest autosomal recessive inheritance. It became later evident that the parkinsonian component of this syndrome might exhibit good and sustained response to levodopa.4,6,8 Brain imaging was unremarkable,6-12 but PET showed marked decrease of striatal fluorodopa uptake.8 Recently, an Iranian kindred was reported with autosomal recessive, early-onset spastic paraplegia. Interestingly, 5 to 20 years after the appearance of pyramidal symptoms, 3 out of the 10 patients Supplemental data at www.neurology.org e-Pub ahead of print on November 26, 2008, at www.neurology.org. From the Department of Clinical Genetics (A.D., E.H.Y., L.C.G., A.S., T.Z., L.O.M.D.-H., C.H.W., E.d.G., E.J.S., G.J.B., B.A.O., V.B.), Erasmus MC, Rotterdam; Departments of Neurology (M.C.J.D., M.W.H.) and Nuclear Medicine (W.J.G.O.), Radboud University Medical Center, Nijmegen, The Netherlands; Department of Neurology (P.M., A.B.), University of Bologna; Dino Ferrari Center (A.D.), Department of Neurological Sciences, University of Milan; and Foundation I.R.C.C.S. Ospedale Maggiore Policlinico, Mangiagalli and Regina Elena, Milan, Italy. Supported by the “Internationaal Parkinson Fonds”–Netherlands, the Erasmus MC Rotterdam (Erasmus Fellowship), and the Netherlands Organization for Scientific Research (NWO, VIDI grant) to V.B. Disclosure: The authors report no disclosures. 240

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Figure 1

Genetic findings and neuroimaging in families with FBXO7 mutations

Black symbols in pedigrees denote affected individuals. To protect privacy, gender of unaffected siblings has been disguised and genotypes are not shown. (A) Pedigree of the Italian family and electropherograms of mutations. (B) Pedigree of the Dutch family and electropherograms of mutations. (C) Brain CIT-SPECT (DaTSCAN-SPECT) in the NIJ-002 patient, showing a severe presynaptic defect of the nigrostriatal dopaminergic systems. (D) Brain IBZMSPECT in the NIJ-002 patient, showing the integrity of postsynaptic striatal dopamine receptor-bearing compartment. (E, F) Normal brain MR images in the NIJ-002 (E) and BO-56 (F) patient.

developed akinetic-rigid parkinsonism without tremor (levodopa response was favorable in one patient, and not tested in the others).13 Linkage mapping in the Iranian family yielded a locus on chromosome 22, and a homozygous missense mutation in the gene encoding the F-box protein 7 (FBXO7) was proposed as the likely disease-causing variant.13 However, association of a single variant with disease does not prove causation, and the role of FBXO7 mutations remains to be demonstrated. Here, we report three novel pathogenic FBXO7

mutations in two families, showing unambiguously that recessive FBXO7 mutations cause a neurodegenerative disease with early-onset, parkinsonian-pyramidal phenotype. METHODS Two Caucasian families (one Dutch and one Italian), each containing two affected siblings, were studied. The disease segregation was compatible with autosomal recessive inheritance (figure 1). There was no history of parkinsonism in the previous generations, nor was there evidence of parental consanguinity. The clinical phenotype consisted of juvenile-onset, progressive parkinsonism with additional pyramidal signs (increased tendon reflexes, spasticity, and Babinski sign), and a prolonged course, thereby matching closely Davison syndrome. Levodopa Neurology 72

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Table

Clinical features in patients with FBXO7 mutations Patient code BO-53

BO-56

NIJ-002

NIJ-006

Gender

Female

Male

Female

Male

Onset age, y

10

13

18

19

Symptoms at onset

Arm tremor, writing difficulty, trunk stiffness, unsteadiness

Hand tremor, slowness of movements, unsteadiness

Tremor, nervousness

Slowness of movements, social withdrawal

Bradykinesia

⫹

⫹

⫹

⫹

Rigidity

⫹

⫹

⫹

⫹

Resting tremor

⫹

⫹

⫺

⫹

Action tremor

⫹

⫹

⫹

⫹

Postural instability

⫹

⫹

⫹

⫹

Dystonic features

⫹

⫹

⫺

⫺

Hyperactive tendon reflexes

⫹

⫹

⫹

⫹

Babinski sign

⫹

⫹

⫹

⫹

Other signs

Dysarthria, dysphagia, urinary incontinence, fecal incontinence

Slow saccades, reduced upgaze, dysarthria, dysphagia

Slow saccades, reduced upgaze

Dysphagia, reduced upgaze

Dementia

⫺

⫺

⫺

⫺

Levodopa response

⫹

⫹

⫹

⫹

Motor fluctuations

⫹

⫹

⫹

⫹

Dyskinesias

⫹

⫹

⫹

⫹

Behavioral disturbances

⫹

⫹

⫹

⫹

Brain MRI, EMG/ ENG, muscle biopsy

Brain MRI, IBZMSPECT, EMG/ENG, muscle biopsy

Brain MRI, IBZM-SPECT, EMG/ENG

Brain MRI

Signs at examination

Levodopa-induced side effects

Performed instrumental investigations Unremarkable

Abnormal

DaTSCANSPECT, TMS

⫹, present; ⫺, absent. EMG/ENG ⫽ electromyography/electroneurography; IBZM-SPECT ⫽ [123I]iodobenzamide single photon emission computed tomography; DaTSCAN-SPECT ⫽ [123I]ioflupane single photon emission computed tomography; TMS ⫽ transcranial magnetic stimulation for the study of the motor pathways.

therapy yielded variable extent of benefit on the parkinsonism in the different patients. The more important clinical features are summarized in the table, while the detailed case reports are provided in appendix e-1 on the Neurology® Web site at www. neurology.org. The relevant ethical authorities approved the study and written informed consent was obtained from all subjects. Genomic DNA was isolated from peripheral blood using standard protocols. The involvement of known genes causing early-onset autosomal recessive typical (parkin, PINK1, DJ-1) or atypical parkinsonism (ATP13A2), as well as those causing NiemannPick disease type C1 and C2 (NPC1, NPC2) and the neuronal ceroid lipofuscinosis-3 (CLN3), were excluded by haplotype analysis and direct gene testing (genomic sequencing of all exons for all genes; gene dosage for parkin, PINK1, DJ-1) (data not shown). All exons and exon–intron boundaries of the FBXO7 gene were PCR-amplified from genomic DNA. cDNA studies were also performed in samples from control subjects and mem242

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bers of the Dutch family. PCR primers, protocols, and sequencing methods are reported in appendix e-2. The novel sequence variants detected in patients were tested in ethnically matched healthy controls, using direct sequencing. Two Fbxo7 protein isoforms are known to be expressed from the usage of different open reading frame (ORF) start codons located on alternative 5=-exons (termed exon 1A and 1B) (figure 2). Here, FBXO7 variants are named according to the longest mRNA and protein isoforms (GenBank accession number NM_012179.3; NP_036311.3) (figure 2), and numbered at nucleotide level from the “A” of the ATGtranslation initiation codon.

We detected FBXO7 mutations in both families (figure 1). In the Italian family (BO), the two affected siblings carry a novel homozygous truncating mutation in exon 9 (c.C1492T, according to the longer FBXO7 transcript, predicted protein effect

RESULTS

Figure 2

The FBXO7 gene, transcripts, and protein isoforms

(A) Genomic structure of the FBXO7 gene (the genomic size is ⬃24 kb). (B) Schematic representation of the FBXO7 transcripts. (C) Fbxo7 protein isoform 1 with functional regions and position of mutations identified in this study. The Arg378Gly mutation identified previously by others in an Iranian family13 is also shown. Ubl ⫽ ubiquitin-like region; cdk ⫽ cdk6 interaction region; FP ⫽ region mediating homo- and heterodimerization of F-Box proteins, and interaction with the PI13 protein; PRR ⫽ proline rich region; R(ar)DP ⫽ highly conserved motif of still undetermined function (“ar” indicates any aromatic amino acids). (D) Alignment of Fbxo7 protein homologues in the regions targeted by the mutation identified in the Dutch patients. The closest homologues of the Fbxo7 protein were aligned using the program T-Coffee. GenBank accession numbers are as follows: NP_036311.3 [Homo sapiens]; XP_001153721.1 [Pan troglodytes]; NP_001033148.1 [Bos taurus]; NP_694875.2 [Mus musculus]; NP_001012222.1 [Rattus norvegicus]; NP_001012555.1 [Gallus gallus]; NP_001086668.1 [Xenopus laevis].

p.Arg498Stop). This mutation was present in heterozygous state in both the unaffected parents and in one of the unaffected siblings. The other unaffected sib did not inherit the mutation. In the Dutch family (NIJ), the two affected siblings are compound heterozygous for two novel FBXO7 mutations: a splice-

site mutation (IVS7 ⫹ 1G/T) and a single base substitution in exon 1A (c.C65T, predicted to lead to the missense protein change p.Thr22Met). This mutation affects only the FBXO7 isoforms containing exon 1A (figure 2). The unaffected mother only carried the heterozygous missense c.C65T mutation. Neurology 72

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Therefore, the remaining mutation (IVS7 ⫹ 1G/T) was most likely transmitted from the father, who was not available for genetic testing. In both families, direct sequencing revealed no additional diseaseassociated variants. The above-mentioned mutations were not found in ethnically matched controls: 364 chromosomes tested from the Italian general population (mutation c.C1492T); 300 chromosomes from the Dutch general population (mutation IVS7 ⫹ 1G/T); 348 chromosomes from the Dutch general population (mutation c.C65T). cDNA analysis from control blood cells and brain tissue confirmed the existence of the two known FBXO7 transcripts (termed isoform 1 and 2) and also revealed evidence for at least two novel in-frame isoforms (termed isoforms 3 and 4), expressed by alternative combinations of exons 1A or 1B with exons 2A or 2B (for details, see figure 2 and figure e-1). Direct sequencing of the full-length FBXO7 cDNA in one patient and the mother from the Dutch family confirmed the presence of the heterozygous c.C65T substitution, documenting therefore the expression of both alleles. The splice-site mutation (IVS7 ⫹ 1G/T) removes the invariable splice donor of intron 7, and is therefore expected to disrupt FBXO7 mRNA splicing. And indeed, multiple aberrant frame-shift splice variants were detected in the patient but not in the unaffected mother, resulting from the activation of premature cryptic splice sites in exon 7 (figure e-2). The missense mutation p.Thr22Met replaces a highly conserved amino acid in the N-terminal ubiquitin-like domain, one of the known functional domains of the Fbxo7 protein, which is only expressed in the two longer Fbxo7 isoforms (figure 2). This situation predicts that the two shorter Fbxo7 isoforms are unaltered, which is compatible with some small residual Fbxo7 functional activity in the Dutch patients. DISCUSSION The homozygosity and compound heterozygosity for pathogenic mutations detected in the patients from families BO and NIJ represent clearly disease-causing genotypes. Our data convey therefore different important messages. First, they show unambiguously that recessive FBXO7 mutations are a cause of neurodegeneration in humans, after the initial suggestive evidence provided by Shojaee and colleagues. Second, they show that the phenotypic spectrum associated with FBXO7 mutations is much broader and less stereotypic than suggested by Shojaee and colleagues, and it encompasses the combination of early-onset, progressive parkinsonism with associated pyramidal tract signs, matching 244

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therefore clinically the pallido-pyramidal syndrome of Davison. The disease progression seems slow, as patients were alive decades after symptoms onset. Response to levodopa of the parkinsonism is variable, sometimes marked and sustained, but often limited by severe motor and psychiatric side effects. Screening of the FBXO7 gene should be included in the diagnostic workup of patients with otherwise unexplained earlyonset parkinsonian-pyramidal syndromes. We agree with others6 that the term “parkinsonianpyramidal syndrome” should be preferred to “pallidopyramidal syndrome,” in the lack of pathology studies, and because the patients exhibit clinically a combination of parkinsonism and pyramidal disturbance. Moreover, on the basis of the clinical and imaging features, the disease caused by FBXO7 mutations should be listed among the monogenic parkinsonisms, and termed PARK15 (for monogenic parkinsonism type 15). The presynaptic nature of the parkinsonism is shown by the dramatic abnormality of DaTSCANSPECT, the beneficial effect of levodopa and the presence of levodopa-induced dyskinesias, and, in the patient NIJ-002, the low HVA and compensatory high biopterin levels in spinal fluid, normalized with levodopa therapy. A massive postsynaptic involvement is unlikely because of the normal MRI, normal IBZM-SPECT, and long-lasting beneficial levodopa effect, at least in some patients. The pyramidal signs together with abnormal TMS in patient NIJ-002 confirm the presence of pyramidal tract lesions. The normal MRI and the absence of dementia might help distinguish this disease from other forms of complicated early-onset parkinsonism (such as PARK9 or neurodegeneration with brain iron accumulation).14 The patients from the Italian family carrying a Fbxo7 homozygous truncating mutation exhibit a more severe phenotype than those in the Dutch family, who carry missense and splice mutations, and those in the Iranian family,13 with homozygous missense. This phenotypic variance might correlate with genotypes and with the possible residual Fbxo7 protein activity, but analysis of large number of patients with FBXO7 mutations is warranted. Little is known about the function of the Fbxo7 protein. When overexpressed, Fbxo7 shows mainly cytosolic, but also nuclear localization.15,16 Fbxo7 is a member of the F-box-containing protein (FBP) family, characterized by a ⬃40-amino acids domain (the F-box). FBPs serve as molecular scaffolds in the formation of protein complexes, and have been implicated in a range of processes, such as cell cycle, genome stability, development, synapse formation, and circadian rhythms.17 Through the interaction

between F-box and the Skp1 protein, FBPs become part of SCF (Skp1, Cullin1, F-box protein) ubiquitin ligase complexes, and play roles in ubiquitinmediated proteasomal degradation. However, FBPs might also be involved in ubiquitin-mediated, nonproteasomal pathways, and SCF-independent functions.17 Fbxo7 also contains an N-terminal ubiquitin-like domain, and a C-terminal proline-rich region (PRR), crucial for FBPs target specificity.15,17,18 Fbxo7 is known to interact with different proteins, including the hepatoma upregulated protein (HURP, a mitotic protein),18 the inhibitor of apoptosis protein 1 (cIAP1),16 and the proteasome inhibitor protein PI31.15 Finally, Fbxo7 has SCF-independent transforming activity by enhancing the interaction of cyclin-dependent-kinase CDK6 with its targets.19 Whether these or other, still unknown Fbxo7 interacting proteins are important for the neuronal function of Fbxo7 and for the mechanisms of neurodegeneration is unknown. Unraveling the Fbxo7 pathways in neurons and the mechanisms of Fbxo7linked disease will shed further light on the molecular mechanisms of multiple-system brain degeneration.

5.

6.

7.

8.

9. 10.

11.

12.

13.

14. ACKNOWLEDGMENT The authors thank all patients and relatives for their contributions, M.J.R. Janssen (Radboud University Nijmegen Medical Center) for help with neuroimages, and Tom de Vries-Lentsch, Erasmus MC, Rotterdam, for artwork.

15.

Received July 2, 2008. Accepted in final form September 16, 2008.

16.

REFERENCES 1. Davison C. Pallido-pyramidal disease. J Neuropathol Exp Neurol 1954;13:50–59. 2. Moller JC, Oertel WH. Other degenerative processes. In: Koller WC, Melamed E, eds. Parkinson’s Disease and Related Disorders, Part II: Handbook of Clinical Neurology, Vol. 84. Amsterdam: Elsevier; 2007:445–457. 3. Sutton JP. Other adult-onset movement disorders with a genetic basis. In: Pulst S-M, ed. Genetics of Movement Disorders. San Diego: Academic Press; 2003:511–540. 4. Horowitz G, Greenberg J. Pallido-pyramidal syndrome treated with levodopa. J Neurol Neurosurg Psychiatry 1975;38:238–240.

17.

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Tranchant C, Boulay C, Warter JM. Le syndrome pallidopyramidal: une entite meconnue. Rev Neurol 1991;147: 308–310. Nisipeanu P, Kuritzky A, Korczyn AD. Familial levodoparesponsive parkinsonian-pyramidal syndrome. Mov Disord 1994;9:673–675. Pradat PF, Dupel-Pottier C, Lacomblez L, et al. Case report of pallido-pyramidal disease with supplementary motor area involvement. Mov Disord 2001;16:762–764. Remy P, Hosseini H, Degos JD, et al. Striatal dopaminergic denervation in pallidopyramidal disease demonstrated by positron emission tomography. Ann Neurol 1995;38: 954–956. Kalita J, Misra UK, Das BK. Sporadic variety of pallidopyramidal syndrome. Neurol India 2003;51:383–384. Rajendran P, Aleem MA, Chandrasekaran R, Raveendran S, Ramasubramanian D. Familial Parkinsonian pyramidal syndrome. Neurol India 2000;48:297–298. Srivastava T, Goyal V, Singh S, Shukla G, Behari M. Pallido-pyramidal syndrome with blepharospasm and good response to levodopa. J Neurol 2005;252:1537– 1538. Panagariya A, Sharma B, Dev A. Pallido-pyramidal syndrome: a rare entity. Indian J Med Sci 2007;61:156– 157. Shojaee S, Sina F, Banihosseini SS, et al. Genome-wide linkage analysis of a Parkinsonian-pyramidal syndrome pedigree by 500 K SNP arrays. Am J Hum Genet 2008;82: 1375–1384. Bonifati V. Genetics of parkinsonism. Parkinsonism Relat Disord 2007;13(suppl 3):S233–S241. Kirk R, Laman H, Knowles PP, et al. Structure of a conserved dimerization domain within the F-box protein Fbxo7 and the PI31 proteasome inhibitor. J Biol Chem 2008;283:22325–22335. Chang YF, Cheng CM, Chang LK, Jong YJ, Yuo CY. The F-box protein Fbxo7 interacts with human inhibitor of apoptosis protein cIAP1 and promotes cIAP1 ubiquitination. Biochem Biophys Res Commun 2006;342:1022– 1026. Ho MS, Ou C, Chan YR, Chien CT, Pi H. The utility F-box for protein destruction. Cell Mol Life Sci 2008;65: 1977–2000. Hsu JM, Lee YC, Yu CT, Huang CY. Fbx7 functions in the SCF complex regulating Cdk1-cyclin B-phosphorylated hepatoma up-regulated protein (HURP) proteolysis by a proline-rich region. J Biol Chem 2004;279:32592– 32602. Laman H, Funes JM, Ye H, et al. Transforming activity of Fbxo7 is mediated specifically through regulation of cyclin D/cdk6. Embo J 2005;24:3104–3116.

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X-linked distal hereditary motor neuropathy maps to the DSMAX locus on chromosome Xq13.1-q21 M. Kennerson, PhD G. Nicholson, MD, PhD B. Kowalski, BSc(Hon) K. Krajewski, MSc D. El-Khechen, MSc S. Feely, MSc S. Chu, BMedSci M. Shy, MD, PhD J. Garbern, MD, PhD

Address correspondence and reprint requests to Dr. Marina Kennerson, Northcott Neuroscience Laboratory, ANZAC Research Institute, Concord, New South Wales, Australia [email protected]

ABSTRACT

Objective: To clinically characterize and map the gene locus in a three-generation family with an X-linked adult-onset distal hereditary motor neuropathy.

Methods: Microsatellite markers spanning the juvenile distal spinal muscular atrophy (DSMAX) locus were genotyped and analyzed using genetic linkage analysis. The promoter, untranslated and coding region of the gap junction ␤1 (GJB1) gene was sequenced. Nine positional candidate genes were screened for disease mutations using high-resolution melt (HRM) analysis.

Results: The family showed significant linkage to markers on chromosome Xq13.1-q21. Haplotype construction revealed a disease-associated haplotype between the markers DXS991 and DX5990. Sequence analysis excluded pathogenic changes in the coding and promoter regions of the GJB1 gene. Additional fine mapping in the family refined the DSMAX locus to a 1.44-cM interval between DXS8046 and DXS8114. HRM analysis did not identify disease-associated mutations in the coding region of nine candidate genes. Conclusion: We have identified a family with adult-onset distal hereditary motor neuropathy that refines the locus reported for juvenile distal spinal muscular atrophy (DSMAX) on chromosome Xq13.1-q21. Exclusion of mutations in the coding and regulatory region of the GJB1 gene eliminated the CMTX1 locus as a cause of disease in this family. Nine positional candidate genes in the refined interval underwent mutation analysis and were eliminated as the pathogenic cause of DSMAX in this family. The syndrome in this family may be allelic to the juvenile distal spinal muscular atrophy first reported at this locus. Neurology® 2009;72:246–252 GLOSSARY A ⫽ arm; ACRF ⫽ Australian Cancer Research Foundation; ALS ⫽ amyotrophic lateral sclerosis; CMT ⫽ Charcot–Marie– Tooth; CV ⫽ conduction velocity; dHMN ⫽ distal hereditary motor neuropathy; DSMAX ⫽ distal spinal muscular atrophy; F ⫽ female heterozygote; GJB1 ⫽ gap junction ␤1; HRM ⫽ high-resolution melt; kb ⫽ kilobase; L ⫽ leg; LPA ⫽ lysophosphatidic acid; M ⫽ affected male; MAP ⫽ motor action potential; Mb ⫽ megabase; MSR ⫽ muscle stretch reflex; NP ⫽ not performed; NR ⫽ no response; OMIM ⫽ Online Mendelian Inheritance in Man; SAP ⫽ sensory action potential; SNP ⫽ single nucleotide polymorphism; Temp sens ⫽ weakness markedly worsened in cold weather; UCSC ⫽ University of California Santa Cruz.

The hereditary disorders of the peripheral nerve form one of the most common groups of human genetic diseases. Charcot–Marie–Tooth (CMT) disease includes disorders of the peripheral nerve affecting both the motor and the sensory neurons. The disorder affects 1 in 2,500 people.1 Some neuropathies are predominantly motor disorders of the peripheral nervous system. These were previously called the “spinal” forms of CMT and distal “spinal muscular atrophy.” More recently, they have been called the distal hereditary motor neuropathies (dHMNs).2 dHMN is characterized by the wasting and weakness of the distal limb muscles, neurophysiologically by denervation on EMG, by normal or slightly reduced motor nerve conduction velocities, and by normal sensory nerve amplitudes and conduction velocities according to established clinical criteria.3 Supplemental data at www.neurology.org From Northcott Neuroscience Laboratory, ANZAC Research Institute (M.K., G.N., B.K., S.C.) and Molecular Medicine Laboratory (M.K., G.N.), Concord Hospital, Concord, New South Wales, Australia; and Department of Neurology (J.G., M.S., K.K., D.E.-K., S.F.), Wayne State University School of Medicine, Detroit, MI. Disclosure: The authors report no disclosures. 246

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dHMN is further classified based on the mode of inheritance, age at onset, and clinical patterns of weakness. Currently, 15 known loci that include autosomal dominant, autosomal recessive, and X-linked forms of the disease have been mapped, and seven genes have been identified.2 The biology of disease seen in dHMN is the selective death of motor neurons. Motor neurons represent a unique cell type with long axons (up to 1 meter) that require continuous maintenance from the cell body to the neuromuscular junctions. This organization requires that systems be in place to cope with the metabolic load to maintain efficient communication over these long distances. The genes identified to date highlight the involvement in two important pathways: axonal transport and RNA metabolism.4 These pathways include proteins involved in RNA processing,5 translation,6 stress response,7,8 apoptosis9 and retrograde transport.10 Families with dHMN have also shown overlap with clinical signs and pathology similar to amyotrophic lateral sclerosis (ALS) and CMT neuropathy. Clinical similarities between ALS4 and dHMN families later showed that both phenotypes had senataxin (SETX) mutations.5 Similarly, mutations in the dynactin gene (DCTN1) were found in both dHMN10 and ALS.11,12 Mutations in the HSP27 gene were identified in distal HMN2B and an axonal form of CMT (CMT2F).7 It is likely that identifying new genes involved in dHMN will be helpful in identifying additional pathways relevant to other motor neuron disorders as well as motor neuronal function. This study reports a family with X-linked recessive adult-onset distal hereditary motor neuropathy. We report genetic linkage studies that have mapped the disease locus in this family to the previously reported recessive distal spinal muscular atrophy (DSMAX) locus on chromosome Xq13.1-q21.13 METHODS Family ascertainment. We examined a threegeneration family with probable X-lined adult-onset hereditary motor neuron disease. Participants in the study gave informed consent in accordance with protocols approved by the Sydney Southwest Area Health Service Ethics Review Committee (Concord Hospital, New South Wales, Australia). Genomic DNA was extracted from whole blood using the PureGene Kit (Gentra

Systems) according to the manufacturer’s instructions. The known loci for X-linked CMT neuropathy (CMTX2, CMTX3, CMTX5, and Cowchock syndrome) were excluded. The coding region of the GJB1 gene (CMTX1) was excluded for pathogenic changes by sequence analysis of the proband (IV-5) (figure e-1 on the Neurology® Web site at www.neurology.org).

Clinical and electrophysiologic examination. The patients were evaluated by standard neurologic examination using the Medical Research Council scale. Nerve conduction studies were performed by standard techniques. Motor nerve conduction velocities and sensory nerve conduction velocities were performed at each evaluation. Sensory nerve conduction velocities were antidromic. Compound muscle action potential and sensory nerve action potential amplitudes were also recorded. Temperature was maintained at 34°C in the hands and feet for all visits. Surface electrodes were used in all studies.

Microsatellite analysis. Microsatellite markers DXS991, DXS1275, DXS8052, DXS559, DXS8046, DXS986, DXS6793, DXS995, DXS8076, DXS8114, DXS6803, DXS1196, DXS1217, and DXS990 were genotyped in the family. The forward primers were 5= labeled with the 6-FAM fluorochrome. Microsatellite markers were amplified using touchdown PCR protocols in 10-␮L reactions containing 25 ng DNA, 0.5 U Chromo AT Taq polymerase (TrendBio), 3 mM MgCl2, 200 ␮M dNTPs, 8 pmol primer, and 1X PCR enhancer (Invitrogen). The markers were sent to the Australian Cancer Research Foundation (ACRF) Facility, Garvan Institute of Medical Research (New South Wales, Australia) for size fractionation using the GeneScan LIZ 600 size standard. Genotypes were analyzed using GeneMarker version 5.1 software (SoftGenetics LLC). Linkage and haplotype analysis. Two-point linkage analysis was performed using the MLINK program from the Linkage package (version 5.1)14 in the Fastlink implementation (version 4.1p).15 Fully penetrant X-linked recessive inheritance was assumed with a disease allele frequency of 0.0001. Male and female recombination rates were considered equal. Marker alleles were set at 1/n, where n is the number of alleles observed. Extended haplotypes of family members were constructed according to the order of the Rutgers combined linkage–physical map of the human genome16 and based on the minimal number of intermarker recombinations. GJB1 gene promoter analysis. Primers were designed to amplify 1.285 kilobase (kb) of sequence upstream of the GJB1 gene methionine start codon. The region was amplified in four overlapping fragments that included the 5=UTR, the neural specific P1 promoter gene, and 1.2 kb upstream of the P1 promoter. Primer information is available on request. PCR reactions were performed in 30-␮L reactions containing 25 ng DNA, 0.5 U Chromo AT Taq polymerase, 3 mM MgCl2, 200 ␮M dNTPs, 8 pmol primer, and 1X PCR Enhancer. PCR amplification was performed on a MasterCycler (Eppendorf) with an initial denaturation of 95°C for 15 minutes followed by 35 cycles of 95°C for 30 seconds, appropriate annealing temperature for 30 seconds, and 68°C for 40 seconds, with a final extension of 68°C for 5 minutes. PCR products were purified (PureLink PCR purification kit, Invitrogen) and sequenced using standard BigDye Terminator Cycle Sequencing protocols at the ACRF Facility, Garvan Institute of Medical Research. Sequences were assembled using SeqMan II version 5.03 (DNASTAR Inc.). Candidate gene analysis. Candidate genes within the prioritized region of the DSMAX locus were identified through the Neurology 72

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Table 1

Clinical summary

Patient

Age, y

Age at onset, y

Weakness

Atrophy

Sensory

MSRs

Temp sens

IV-5

39

⬃13

Distal, L⬎A

Feet, lower legs, and hands

Mild distal toes and fingers

Absent Achilles

⫹

IV-6

44

10

Distal, L⬎A

Feet, lower legs, and hands

Mild distal lower legs

Absent Achilles, mildly brisk knees

⫹

IV-12

53

⬃30

Mild distal L ⬎A

Feet, lower legs, and hands

Normal

Normal

IV-21

57

⬃10

V-2

16

13

V-7

33

⬃25

Mild distal L ⬎A

Feet, lower legs, and hands

Mild distal reduction

Normal

Mild lower leg

—

Normal

Normal

Mild hand

High foot arches; mild hand atrophy

Normal

Normal

MSR ⫽ muscle stretch reflex; Temp sens ⫽ weakness markedly worsened in cold weather; L ⫽ leg; A ⫽ arm.

University of California Santa Cruz (UCSC) Genome Browser (Human March 2006 Assembly; NCBI Build 36.1). Mutation analysis of gene coding exons and flanking sequences was performed using high-resolution melt (HRM) protocols established in our laboratory.17 Samples were analyzed in duplicate and included affected males (n ⫽ 4), normal males (n ⫽ 4), carrier females (n ⫽ 2), and a noncarrier female (n ⫽ 1) from the family. To facilitate heteroduplex formation, male samples were mixed (1:1 vol/vol) with a known nonaffected male control from the family. Primers for HRM were designed using the LightScanner Primer Design Software (version 1.0.R.84, ID Technology). The size of amplicons designed ranged from 107 to 350 base pairs. Primer information is available on request. Samples were amplified and analyzed using previously described protocols.17 Amplicons in which affected and carrier melt profiles grouped together were sequenced using BigDye Terminator Cycle Sequencing protocols at the ACRF Facility, Garvan Institute of Medical Research. RESULTS Clinical characterization of family. The clinical syndrome in this family is of sex-linked distal weakness that begins in late childhood to early adulthood and is slowly progressive. The predominantly distal motor syndrome suggests a primary disorder of motor neurons. The presence of sensory symptoms, although a minor clinical component of the spectrum, suggests that the disorder may have more general effects on the peripheral nerve. A summary of the clinical (table 1) and electrophysiologic (table 2) findings in the family is presented. The proband (IV-5) is representative of the affected males and illustrates the clinical syndrome. His infancy and early developmental milestones were normal. As a child, he was able to keep up with his peers, and he learned to ride a bicycle and to ice skate. His earliest recalled symptoms were difficulty with ice skating, beginning around age 13–14 years. He developed slowly increasing difficulty with hand grip and with ankle strength thereafter, which were more noticeable initially during the winter. By age 16 years, he was unable to participate in sports and began to have increasing difficulty with walking and maintaining his balance because of ankle weakness, but he did not 248

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use ankle–foot orthotics until age 38 years. He continued to work as a handyman until age 40 years but currently remains active and independent with ambulation. On examination at age 40 years, his mental status and cranial nerve examinations were normal. He had normal proximal arm strength. Wrist, hand, and finger strength were moderately weak, and atrophy of hand muscles was present, although no fasciculations were observed. The hip flexors were mildly weak, and the knee flexors and extensors were moderately weak. The ankle and toe dorsi and plantar flexors were profoundly weak, and there was moderately severe atrophy of foot muscles. He had a steppage gait and was unable to tandem walk or to walk on his toes or heels. The sensory examination was minimally abnormal, with mild reduction in vibratory sense at toes. The biceps and triceps reflexes were preserved, but brachioradialis and ankle reflexes were absent. Plantar responses were neutral. The nerve conduction studies are summarized in table 2. The electromyogram showed increased insertional activity, with positive waves and fibrillations in selected distal arm and leg muscles. The clinical neurologic examinations of the proven heterozygous females were normal. Motor nerve conduction studies of IV-2 were normal, whereas her sensory conduction studies showed borderline abnormalities (table 2). Confirmation and refinement of the DSMAX locus.

Initial analysis of markers on Xq13.1-q21 included DXS8052 and DXS559. These markers flank the GJB1 gene. A maximum lod score of 3.46 at zero recombination was obtained for both markers (table 3). Although the coding and 5= untranslated region of the GJB1 gene had been excluded for pathogenic mutations, the lod scores suggested linkage to the CMTX1 locus. We subsequently examined the reported promoter of the GJB1 gene18; however, sequence analysis did not identify pathogenic mutations. The possibility of GJB1 gene dosage ef-

Table 2

Clinical motor and sensory electrophysiology Motor Median

Patient

Sex

Peroneal

Tibial

MAP, mV

CV, m/s

MAP, mV

CV, m/s

MAP, mV

CV, m/s

(⬎4)

(⬎48)

(⬎3)

(⬎41)

(⬎4)

(⬎41)

IV-5

M

2.62

61.5

NR

NR

2.80

31.9

IV-6

M

3.6

46.8

NR

NR

NR

NR

IV-12

M

12.0

57.5

1.0

NR

2.3

NR

IV-21

M

0.033

NR

NR

NR

NR

NR

V-2

M

6.3

64.7

2.6

NR

13.5

NR

V-7

M

8.0

56.8

NR

NR

NR

NR

IV-2

F

10*

56*

6

49

NP

NP

Sensory Median

Sural

SAP, ␮V

IV-5

M

CV, m/s

SAP, ␮V

CV, m/s

(⬎25)

(⬎49)

(⬎6)

(⬎43)

26.57

49.4

13.69

48.0

IV-6

M

17

50

NP

NP

IV-2

F

37

NP

5

NP

Abnormal values are in boldface type. *Values from ulnar nerve study; normal values are in parentheses. MAP ⫽ motor action potential; CV ⫽ conduction velocity; M ⫽ affected male; NR ⫽ no response; F ⫽ female heterozygote; NP ⫽ not performed; SAP ⫽ sensory action potential.

fects is highly unlikely in this family because carrier females heterozygous for the microsatellite markers flanking the GJB1 gene gave equal allele fluorescence intensity (data not shown). Additional markers were genotyped in the family and included DXS991,

Table 3

DXS1275, DXS8046, DXS986, DXS6793, DXS1196, DXS1217, and DXS990. Significant lod (ⱖ ⫹2) scores were obtained for five of the markers with the highest lod score being at DXS1217 (z ⫽ 3.80 at zero recombination) (table 3). A haplotype segregat-

Two-point lod scores between the disease and polymorphic markers on chromosome Xq13.1-q21

Marker

Position, cM/Mb

Recombination fraction, ⌰ 0.0

0.01

0.05

—∞

0.1

0.2

0.3

0.4

⫺0.31

⫺0.30

⫺0.48

0.54

0.44

0.26

DXS1275

58.3/68.4

3.46

3.41

3.18

2.89

2.27

1.59

0.84

DXS8052

58.67/69.7

3.46

3.41

3.18

2.89

2.27

1.59

0.84

DXS559

59.33/70.7

3.46

3.41

3.18

2.89

2.27

1.59

0.84

DXS8046

59.33/71.2

3.16

3.11

2.90

2.63

2.06

1.44

0.76

DXS986

59.96/79.2

0.80

0.79

0.74

0.67

0.52

0.37

0.19

DXS6793

59.96/80.6

0.30

0.30

0.28

0.25

0.20

0.15

0.08

DXS995

60.19/82.6

1.11

1.09

1.02

0.93

0.73

0.51

0.27

DXS8076

60.19/82.6

1.10

1.08

1.01

0.92

0.73

0.51

0.27

DXS8114

60.77/85.5

—∞

2.06

2.51

2.49

2.11

1.54

0.85

DXS991

56.06/55.5

DXS6803

60.84/86.3

—∞

⫺0.31

DXS1196

61.07/86.5

—∞

2.06

DXS1217

62.37/88.2

3.80

DXS990

64.12/92.8

—∞

0.3

0.48

0.54

0.44

0.26

2.51

2.49

2.11

1.54

0.85

3.73

3.49

3.16

2.48

1.74

0.92

⫺2.91

⫺1.54

⫺0.98

⫺0.48

⫺0.22

⫺0.08

Mb ⫽ megabase. Neurology 72

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ing with the disease in affected males and female carriers was identified between the markers DXS991 and DXS990. Two affected males showed a recombination. Individual V-10 showed a recombination between the markers DXS990 and DXS1217, whereas individual V-2 initially showed a more proximal recombination between the markers DXS6793 and DXS1196. To further refine the distal end of the DSMAX locus, additional markers between DXS6793 and DXS196 were genotyped (DXS995, DXS8076, DXS8114, and DXS6803). Individual V-2 showed an informative recombination between the markers DXS8114 and DXS8076. The DSMAX locus was subsequently refined to a 1.44-cM interval flanked by DXS8046 and a new distal flanking marker at DXS8114 (figure e-1). Transcript mapping and candidate gene analysis. The newly refined DSMAX locus flanked by the markers DXS8046 and DXS8114 corresponds to a 14.2megabase (Mb) interval and contains 56 full-length annotated genes (UCSC Genome Browser, Human March 2006 Assembly; NCBI Build 36.1). In addition, more than 70 partial transcripts were identified by querying the dbEST and UniGene databases. These expressed sequence tags are transcriptionally active and could represent either alternatively spliced messenger RNAs of the known annotated genes or novel transcripts. Nine candidate genes were selected from the refined region based on a role in neuron maintenance and database expression information from SOURCE and UniGene for relevant neural tissues (brain, spinal cord, and peripheral nerve). No aberrant melt curves (indicating a change in the amplicon DNA sequence) were observed in seven of the genes (PHKA1, NAP1L2, RNF12, SH3BGRL, GPR23, RPS4X, and CDX4). The subtractive–melt curves for affected/carrier and normal/noncarrier samples grouped together. In contrast, HRM analysis of exon 1 of the UPRT gene showed affected males and carrier females grouping separately to the normal males and noncarrier female (figure, A). Sequence analysis of male individuals from each group identified a reported intronic single nucleotide polymorphism (SNP; rs5981831). The affected males were hemizygous for the A allele, and nonaffected males were hemizygous for the G allele. Carrier females were heterozygous (A/G), and the noncarrier female was homozygous (G/G). Analysis of the rs5981831 SNP in the recombinant branch of the family did not further refine the DSMAX locus. HRM analysis of the PGK1 gene identified a reported intronic SNP (rs2007039) in the amplicon designed to analyze exon 6 of the gene. Sequence analysis of the two different groups showed the carrier female to be heterozygous (C/T) for the SNP, 250

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whereas samples from the other group were homozygous/hemizygous for the T allele (figure, B). DISCUSSION We have mapped an X-linked family with late-onset distal hereditary motor neuropathy to the DSMAX locus on chromosome Xq13.1-q21. The clinical findings in this family are of a primary motor disorder. Although several of the affected males did have mild sensory symptoms, they were not associated with electrophysiologic abnormalities, except in one case (IV-6), who was considered to have an acquired sensory neuropathy. This result suggests that the motor neuron is the major target of the disease. Mild sensory involvement has also been reported in ALS.19-21 The family’s clinical syndrome suggests that the disorder may be allelic to a juvenile form of X-linked distal spinal muscular atrophy reported in a Brazilian kindred.13 Clinically, the syndrome reported in the Brazilian family is more severe because onset is in childhood, but the syndromes are otherwise similar. It is likely that the two families will have different mutations in the same gene and could reflect the age-at-onset variability seen in disorders such as Werdnig–Hoffman and Kugelberg–Welander syndrome or Duchenne and Becker muscular dystrophy. Genetic analysis in this family suggested significant linkage to the CMTX1 locus. By thoroughly examining the GJB1 gene (both coding and regulatory sequences), we excluded this gene as a cause of the disease in the family. Examination of affected males did not identify recombination events that could further refine the proximal end of the DSMAX locus. A female individual (V-8) who carries a portion of the disease haplotype could be a carrier and potentially assist in refining the interval in this family. However, we have coded this individual as an unknown phenotype because it is difficult to clinically discriminate between carrier and noncarrier phenotypes. Based on the original mapping for the DSMAX locus and an informative recombination in an affected male in this family, the locus has been reduced to a 1.44-cM interval flanked by the markers DXS8046 and the newly defined distal marker DXS8114. The refined region corresponds to a 14.3-Mb interval and is gene rich. A number of candidate genes were selected for analysis based on neural cellular function (GPR23, NAP1L2, and RNF12) and expression in neural tissue (SH3BGRL, UPRT, RPS4X, and CDX4). Candidate genes chosen on functional relevance included ubiquitin ligase (RNF12), in which members of this family of proteins have been reported to be involved with axon outgrowth through the modulation of microtubule dynamics22;

Figure

Subtractive difference plots affected/carrier and normal/noncarrier samples for the amplicon containing exon 1 of the UPRT gene (A) and exon 6 of the PGK1 gene (B)

All samples were run in duplicate. For the UPRT gene, the samples grouped according to the genotype present in each individual. Sequence analysis of individuals from each group identified an intronic single nucleotide polymorphism (SNP) (rs5981831). The difference curve groups reflect the A allele segregating with affected and carrier individuals and the G allele segregating with the normal males and the noncarrier females. For the PGK1 gene, all samples were homozygous/hemizygous for the T allele, except for one carrier female who was heterozygous (C/T) for the SNP rs2007039.

a nucleosome assembly protein (NAP1L2) that interacts with the chromatin and alters transcription in neurons23; and a G protein– coupled receptor (GPR23) that binds lysophosphatidic acid (LPA). LPA is a potent lipid mediator and affects cell morphology, cell survival, and cell cycle progression in neuronal cells. Two genes screened in the current family are associated with diseases that have been mapped to the same region as DSMAX. PHKA1 encodes the ␣ subunit of muscle phosphorylase kinase, and defects in this gene cause X-linked muscle glycogenosis (Online Mendelian Inheritance in Man [OMIM] 300599).24 This disorder is characterized by slowly progressive, predominantly distal muscle weakness and atrophy. These clinical features were evident in the current family, and it was possible that different mutations to those identified for X-linked muscle glycogenosis could be the cause of DSMAX

in the family. This is not unprecedented, because mutations in different domains of the DNM2 gene cause both peripheral neuropathy25 and centronuclear myopathy.26 The enzyme phosphoglycerate kinase is encoded by PGK1, and mutations in this gene cause phosphoglycerate kinase 1 deficiency (OMIM 300653). Twenty-six families have been documented with clinically significant phosphoglycerate kinase deficiency, and 18 of these families have reported mutations in the PGK1 gene.27 This condition has a highly variable clinical phenotype that includes chronic hemolysis with or without mental retardation, myopathy, and neurologic involvement. Although no pathogenic mutations were identified in the coding region of this gene, a carrier female was identified with a reported SNP in the intronic region of the amplicon designed to scan exon 6 of this gene. HRM analysis was used for mutation analysis of the positional candidate genes. We have demonstrated that HRM analysis is a fast and efficient method for mutation scanning large multiexon genes such as PHKA1, which contained 32 coding exons, and PGK1, which had 11 exons. The rationale for using HRM analysis in this study was to detect changes in the DNA by identifying aberrant melt profiles that separate individuals into different groups based on their genotype. The results obtained for the PCR amplicon containing exon 1 of the UPRT gene showed that HRM analysis can successfully group the samples based on phenotype when a DNA change segregates exclusively with the disease. By using HRM analysis, we were able to efficiently exclude exons for a pathogenic role in the disease if affected/carrier individuals grouped with normal/ noncarrier individuals. This resulted in the sequencing burden being considerably reduced because only a few samples required sequencing. In instances where aberrant melt curves were identified, we observed 100% specificity using HRM analysis, because sequence analysis of aberrant melt curves corresponded to known reported DNA variations. This study provides confirmation of the DSMAX locus and has refined the candidate region to a size that is amenable to positional cloning strategies. Implementation of HRM analysis has provided a timeand cost-efficient mutation-scanning method for identifying the DSMAX gene. Identification of this gene will be important for understanding the selective demise of the lower motor neurons and may identify additional important common pathways relevant for other motor neuron disorders. AUTHOR CONTRIBUTIONS All two-point linkage analysis was conducted by M.K.

Received June 2, 2008. Accepted in final form October 3, 2008. Neurology 72

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REFERENCES 1. Skre H. Genetic and clinical aspects of Charcot-MarieTooth’s disease. Clin Genet 1974;6:98–118. 2. Irobi J, Dierick I, Jordanova A, Claeys KG, De Jonghe P, Timmerman V. Unraveling the genetics of distal hereditary motor neuronopathies. Neuromolecular Med 2006;8:131– 146. 3. De Jonghe P, Timmerman V, Van BC. 2nd Workshop of the European CMT Consortium: 53rd ENMC International Workshop on Classification and Diagnostic Guidelines for Charcot-Marie-Tooth Type 2 (CMT2-HMSN II) and Distal Hereditary Motor Neuropathy (Distal HMN-Spinal CMT) 26 –28 September 1997, Naarden, The Netherlands. Neuromuscul Disord 1998;8:426–431. 4. Van Den Bosch L, Timmerman V. Genetics of motor neuron disease. Curr Neurol Neurosci Rep 2006;6:423–431. 5. Chen YZ, Bennett CL, Huynh HM, et al. DNA/RNA helicase gene mutations in a form of juvenile amyotrophic lateral sclerosis (ALS4). Am J Hum Genet 2004;74:1128–1135. 6. Antonellis A, Ellsworth RE, Sambuughin N, et al. Glycyl tRNA synthetase mutations in Charcot-Marie-Tooth disease type 2D and distal spinal muscular atrophy type V. Am J Hum Genet 2003;72:1293–1299. 7. Evgrafov OV, Mersiyanova I, Irobi J, et al. Mutant small heat-shock protein 27 causes axonal Charcot-Marie-Tooth disease and distal hereditary motor neuropathy. Nat Genet 2004;36:602–606. 8. Irobi J, Van Impe K, Seeman P, et al. Hot-spot residue in small heat-shock protein 22 causes distal motor neuropathy. Nat Genet 2004;36:597–601. 9. Windpassinger C, Auer-Grumbach M, Irobi J, et al. Heterozygous missense mutations in BSCL2 are associated with distal hereditary motor neuropathy and Silver syndrome. Nat Genet 2004;36:271–276. 10. Puls I, Oh SJ, Sumner CJ, et al. Distal spinal and bulbar muscular atrophy caused by dynactin mutation. Ann Neurol 2005;57:687–694. 11. Puls I, Jonnakuty C, LaMonte BH, et al. Mutant dynactin in motor neuron disease. Nat Genet 2003;33:455–456. 12. Munch C, Sedlmeier R, Meyer T, et al. Point mutations of the p150 subunit of dynactin (DCTN1) gene in ALS. Neurology 2004;63:724–726. 13. Takata RI, Speck Martins CE, Passosbueno MR, et al. A new locus for recessive distal spinal muscular atrophy at Xq13.1-q21. J Med Genet 2004;41:224–229. 14. Lathrop GM, Lalouel JM, Julier C, Ott J. Strategies for multilocus linkage analysis in humans. Proc Natl Acad Sci USA 1984;81:3443–3446.

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15. Cottingham JrRW, Idury RM, Schaffer AA. Faster sequential genetic linkage computations. Am J Hum Genet 1993;53:252–263. 16. Kong X, Murphy K, Raj T, He C, White PS, Matise TC. A combined linkage-physical map of the human genome. Am J Hum Genet 2004;75:1143–1148. 17. Kennerson ML, Warburton T, Nelis E, et al. Mutation scanning the GJB1 gene with high-resolution melting analysis: implications for mutation scanning of genes for Charcot-Marie-Tooth disease. Clin Chem 2007;53:349– 352. 18. Neuhaus IM, Dahl G, Werner R. Use of alternate promoters for tissue-specific expression of the gene coding for connexin32. Gene 1995;158:257–262. 19. Hammad M, Silva A, Glass J, Sladky JT, Benatar M. Clinical, electrophysiologic, and pathologic evidence for sensory abnormalities in ALS. Neurology 2007; 69:2236–2242. 20. Isaacs JD, Dean AF, Shaw CE, Al-Chalabi A, Mills KR, Leigh PN. Amyotrophic lateral sclerosis with sensory neuropathy: part of a multisystem disorder? J Neurol Neurosurg Psychiatry 2007;78:750–753. 21. Pugdahl K, Fuglsang-Frederiksen A, de Carvalho M, et al. Generalised sensory system abnormalities in amyotrophic lateral sclerosis: a European multicentre study. J Neurol Neurosurg Psychiatry 2007;78:746–749. 22. Lewcock JW, Genoud N, Lettieri K, Pfaff SL. The ubiquitin ligase Phr1 regulates axon outgrowth through modulation of microtubule dynamics. Neuron 2007;56: 604–620. 23. Attia M, Rachez C, De Pauw A, Avner P, Rogner UC. Nap1l2 promotes histone acetylation activity during neuronal differentiation. Mol Cell Biol 2007;27:6093– 6102. 24. Wehner M, Clemens PR, Engel AG, Kilimann MW. Human muscle glycogenosis due to phosphorylase kinase deficiency associated with a nonsense mutation in the muscle isoform of the alpha subunit. Hum Mol Genet 1994;3: 1983–1987. 25. Zuchner S, Noureddine M, Kennerson M, et al. Mutations in the pleckstrin homology domain of dynamin 2 cause dominant intermediate Charcot-Marie-Tooth disease. Nat Genet 2005;37:289–294. 26. Bitoun M, Maugenre S, Jeannet PY, et al. Mutations in dynamin 2 cause dominant centronuclear myopathy. Nat Genet 2005;37:1207–1209. 27. Beutler E. PGK deficiency. Br J Haematol 2007;136: 3–11.

Personality and lifestyle in relation to dementia incidence

H.-X. Wang, PhD A. Karp, PhD A. Herlitz, PhD M. Crowe, PhD I. Ka˚reholt, PhD B. Winblad, MD, PhD L. Fratiglioni, MD, PhD

Address correspondence and reprint requests to Dr. Hui-Xin Wang, Aging Research Center, Dept. Neurobiology, Care Sciences and Society, Karolinska Institutet, Ga¨vlegatan 16, 113 30 Stockholm, Sweden [email protected]

ABSTRACT

Objective: High neuroticism has been associated with a greater risk of dementia, and an active/ socially integrated lifestyle with a lower risk of dementia. The aim of the current study was to explore the separate and combined effects of neuroticism and extraversion on the risk of dementia, and to examine whether lifestyle factors may modify this association.

Methods: A population-based cohort of 506 older people with no dementia from the Kungsholmen Project, Stockholm, Sweden, was followed up for an average of 6 years. Personality traits were assessed using the Eysenck Personality Inventory. Dementia was diagnosed by specialists according to DSM-III-R criteria.

Results: Neither high neuroticism nor low extraversion alone was related to significantly higher incidence of dementia. However, among people with an inactive or socially isolated lifestyle, low neuroticism was associated with a decreased dementia risk (hazard ratio [HR] ⫽ 0.51, 95% confidence interval [CI] ⫽ 0.27– 0.96). When compared to persons with high neuroticism and high extraversion, a decreased risk of dementia was detected in individuals with low neuroticism and high extraversion (HR ⫽ 0.51, 95% CI ⫽ 0.28 – 0.94), but not among persons with low neuroticism and low extraversion (HR ⫽ 0.95, 95% CI ⫽ 0.57–1.60), nor high neuroticism and low extraversion (HR ⫽ 0.97 95% CI ⫽ 0.57–1.65). Stratified analysis by lifestyle showed that the inverse association of low neuroticism and high extraversion in combination was present only among the inactive or socially isolated persons.

Conclusion: Low neuroticism in combination with high extraversion is the personality trait associated with the lowest dementia risk; however, among socially isolated individuals even low neuroticism alone seems to decrease dementia risk. Neurology® 2009;72:253–259 GLOSSARY AD ⫽ Alzheimer disease; CI ⫽ confidence interval; DSM ⫽ Diagnostic and Statistical Manual of Mental Disorders; EPI ⫽ Eysenck Personality Inventory; HPA ⫽ hypothalamus-adrenal cortex; HR ⫽ hazard ratio; MMSE ⫽ Mini-Mental State Examination.

Psychological stress has been related to neurodegenerative processes in the hippocampus in both human1,2 and animal studies.2 Stress is also associated with overactivation and dysregulation of the autonomic nervous system and hypothalamus-adrenal cortex (HPA) axis, which plays a role in coronary heart disease, infection, and accelerated aging.3 Certain personality characteristics are thought to reflect an individual’s level of stress over time and his or her capacity to cope with stressful events.4 The same environmental stressors are not likely to induce similar stress reactions in all people due to different personality traits.5 Persons with high neuroticism experience greater distress than those with low neuroticism in response to similar stressors,6,7 whereas studies on the relationship between extraversion and stress have led to conflicting results. At the biologic level, high neuroticism6,7 and high extraversion8 have been

From Aging Research Center (H.-X.W., A.K., A.H., I.K., B.W., L.F.), Dept. NVS, Karolinska Institutet, and Stockholm Gerontology Research Center, Stockholm, Sweden; and Department of Psychology (M.C.), University of Alabama at Birmingham. Supported by grants from the Swedish Council for Working Life and Social Research (No.2003-0386), the American Alzheimer Foundation, the Alzheimer Foundation Sweden, the Swedish Brain Power, Swedish Research Council, Gamla Tja¨narinnor Foundation, Fredrik and Ingrid Thurings Foundation, the Foundation for Geriatric Diseases and Loo and Hans Osterman Foundation for Geriatric Research at Karolinska Institutet, and the Centre for Health Care Science at Karolinska Institutet. Disclosure: The authors report no disclosures. Copyright © 2009 by AAN Enterprises, Inc.

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shown to be positively correlated with stress and high plasma levels of cortisol.9 Recently, high neuroticism has been shown to be a predictor for depression10 and Alzheimer disease (AD).11,12 A positive association between extraversion and psychological stress has been reported,8,13 but extraversion is also associated with the use of more effective coping strategies14 and more efficient utilization of social support.8 In turn, effective coping15 and social support16 have been shown to buffer against psychological stress. Having a rich social environment or an active lifestyle has also been found to be associated with reduced risk of dementia.17-20 In the present study, we tested the hypothesis that certain personality traits, such as high neuroticism, low extraversion, as well as their combinations, may have a risk or protective effect on dementia occurrence. In addition, since personality is associated with environmental factors such as social support,8 and these factors have been linked to dementia risk,18 we investigated whether an active and socially integrated lifestyle might act as a modifier in the association between personality and risk of dementia. Data were gathered from the Kungsholmen Project, a longitudinal population-based study on aging and dementia carried out in Stockholm, Sweden.21 METHODS Study population. The study population consisted of 506 older people who had no dementia at the second examination of the Kungsholmen Project (baseline of the current study) and who also completed the personality questionnaire. Among the 1,099 individuals who participated in the second examination, we excluded 238 subjects who met criteria for probable dementia and 140 who had physical or cognitive impairment (Mini-Mental State Examination [MMSE] score ⬍24) that hampered the personality assessment. These exclusion criteria left 721 eligible subjects who were asked to complete the personality inventory during the examination at our research center. Of the 544 participants who completed the personality questionnaire, 38 refused the second and third follow-up examinations, which were conducted an average of 3 and 6 years after the first follow-up assessment, leaving 506 (70.2%) subjects who were monitored for 6 years (figure). All enrolled participants consented to participate, and the Regional Research Committee at Karolinska Institute approved the study. In comparison with the 506 participants, the 177 nonparticipants who had missing information for personality were older and more likely to be women (p values ⱕ0.05), however, they had a similar distribution of education, cognitive functioning, vascular disease, and depressive symptoms or diagnoses of depression (p ⬎ 0.05). The 38 persons who dropped out of the follow-up had worse cognition, but were similar to the partici254

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Figure

Inception of the no-dementia cohort representing the study population (n ⴝ 506) and detection of the incident dementia cases (n ⴝ 144)

pants in terms of age, sex, educational level, vascular disease, and depressive symptoms or diagnoses of depression.

Incident dementia cases. The incident dementia cases were all subjects who developed dementia during the second and third follow-up periods. At each examination, all cohort subjects were clinically examined according to a standardized protocol including family and personal history collected by nurses, clinical examination performed by physicians, and psychological tests administered by trained personnel. When a subject was unable to answer questions, an informant, usually someone who was next of kin, was interviewed. A detailed description of the study design has been previously published.22 If the subject had moved, he or she was traced and asked to participate in the follow-up examinations. For those subjects who had died before the follow-up examinations, information regarding their health status during the follow-up was obtained from the Computerized Inpatient Register System, a registry of discharge diagnoses from all hospitals in Stockholm active since 1969. Dementia diagnosis was assigned according to DSM-III-R criteria,23 using a three-step diagnostic procedure22 both at baseline and follow-up examinations. Two physicians made independent preliminary diagnoses, and concordant diagnoses were accepted as final diagnoses, whereas a third opinion was requested in cases of disagreement. Diagnosis of dementia for subjects who died during the time interval between follow-up assessments was based on hospital records and death certificates.17,22 Measurement of personality traits: Neuroticism and extraversion. Personality was assessed using the Eysenck Personality Inventory (EPI). The EPI includes 57 items that measure neuroticism (24 questions) and extraversion (24 questions) in addition to a lie scale (9 questions).24 The subject responded

yes or no to each question, resulting in a range of scores from 0 to 24 for neuroticism and extraversion, and 0 –9 for the lie scale. The neuroticism scale assesses adjustment vs emotional instability and identifies individuals prone to psychological distress, unrealistic ideas, excessive cravings or urges, and maladaptive coping responses. Those with low scores are characterized as being calmer, more relaxed, unemotional, and self-satisfied.4 The extraversion measure assesses quantity and intensity of interpersonal interaction, activity level, need for stimulation, and capacity for joy. Those who score lower on extraversion are characterized as being more reserved, sober, task-oriented, and quiet.4 Lie scale serves as an indicator of test validity. Lying is suspected if rarely performed behaviors are endorsed by the respondent as being habitually performed, or if frequently performed nondesirable behaviors are denied.24 The internal consistency (Cronbach alpha) in the present study is 0.82 for neuroticism and 0.62 for extraversion.

Measurement of lifestyle factors. Information on lifestyle factors, including leisure activities as well as social network, was obtained from the subjects during a personal interview carried out by trained nurses. Subjects were asked about 1) regular engagement in any particular activities or organizations; 2) the types of activities or organizations; and 3) the frequency of participation. Grouping of social and leisure activities followed our previous study,19 which included mental, physical, social, and productive activities. A social network index, including both structural (availability of and contact with network resources) and qualitative (satisfaction with social contacts) aspects of social network, was proposed in our previous study.17 It includes four categories: rich, moderate, limited, and poor social network. The construct of our lifestyle variable is based on three of our previous studies, in which we reported that a rich or moderate social network and frequent participation in mental, social, and productive activities were related to a decreased risk of dementia.17,19,20 Taking into account both social network and leisure activities, a two-category lifestyle variable was created which constructed the most active group vs the less active ones, with an active and socially integrated lifestyle defined as frequent engagement in any of the four types of leisure activities, in addition to having either a rich or moderate social network. It was analyzed in relation to personality traits in the present study. Covariates. Information on age and sex was obtained from government records. Education was defined as the highest level achieved. Global cognitive functioning was assessed using the MMSE.25 Depressive symptoms were assessed by self-report of problems such as persistently feeling lonely or in a low mood. Diagnosis of depression followed DSM-IV criteria26 on the basis of information gathered by a structured interview using the Comprehensive Psychopathological Rating Scale. Information on vascular diseases was gathered from the Stockholm Computerized Inpatient Register System and diagnoses followed criteria from the International Classification of Disease, 8th Edition, which included heart disease (codes 410 to 414, and 427, 428), stroke (codes 430 to 438), and diabetes mellitus (code 250). Genomic DNA was prepared from peripheral blood samples at baseline, and APOE genotyping was conducted following standard procedures by two technicians who were blind to all other data for the study population.

Data analysis. Missing data on personality traits were replaced using the expectation maximization method27 for 30% of the sample who had 1–2 items missing; if more than 2 items were

missing, the scale was treated as missing. All multivariate models reported included the following covariates: age, sex, and education; presence or absence of vascular diseases; depressive symptoms or diagnoses of depression; and cognitive functioning. Multinomial logistic regression analysis was performed to examine potential differences between participants and nonparticipants. The hazard ratios (HR) and corresponding 95% confidence intervals (CI) for dementia associated with personality traits were estimated using Cox proportional hazard analyses. The time to event was assumed to be the midpoint between the first to second follow-up or between the second to third follow-up examinations or death. In the primary analyses, we began with crude analyses, followed by multivariate adjusted analyses. First we examined the separate associations for neuroticism and extraversion on risk of dementia using both continuous and dichotomous variables (median split). Second, we formed a four-category personality index based on the tabulation of the two dichotomized personality traits: 1) low neuroticism and high extraversion; 2) high neuroticism and high extraversion; 3) low neuroticism and high extraversion; and 4) low neuroticism and low extraversion. Third, we repeated all of the above analyses after stratifying by lifestyle (active vs inactive) defined as the combination of social network and participation in leisure activities. In the secondary analyses, in addition to the above covariates, we further adjusted for lie score, presence or absence of ApoE ␧4 alleles, and survival status. These factors were successively introduced into the basic multivariate models one at a time. In all the models, age and cognitive functioning were entered into the models as continuous variables. Sex (female vs male), education (8⫹ years vs ⬍8 years), depressive symptoms or diagnoses of depression (presence vs absence), vascular disease (presence vs absence), ApoE ␧4 alleles (presence vs absence), and survival status (survivors vs deceased) were entered as dichotomous variables. The personality index was introduced as an indicator variable with the high neuroticism and high extraversion category as reference group. To lend greater confidence in the results, we repeated the above analyses in subjects without any missing values for personality items. In addition, we explored potential moderators by performing stratified analyses by age (⬍85 years or 85⫹), sex, cognitive functioning (MMSE score ⬎26 vs ⬍27), and presence or absence of ApoE ␧4 alleles, vascular diseases, or depressive symptoms or diagnoses of depression. In order to address the possibility that preclinical cognitive dysfunction may have affected personality and other variables, we examined a sample including only incident dementia cases developed during the last 3-year follow-up. To eliminate potential diagnostic bias for the deceased subjects, additional analyses were conducted for survivors only.

During the 6 years of follow-up, 144 individuals developed dementia (table 1). As shown in table 2, neuroticism was not significantly related to dementia when entered into the model as a continuous or dichotomous variable. Similarly, extraversion was not associated with dementia risk. When combinations of the two personality traits were examined, in comparison to individuals who scored high on both neuroticism and extraversion, those with low neuroticism in combination with high extraversion were at the lowest risk for developing dementia,

RESULTS

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Table 1

Baseline characteristics of the no-dementia cohort and incident dementia cases occurring over 6 years

Baseline characteristics

No dementia (n ⴝ 362)

Incident dementia (n ⴝ 144)

p Value

Age, y, mean ⴞ SD

83.0 ⫾ 3.2

83.9 ⫾ 3.7

0.014

Female, n (%)

252 (69.6)

110 (76.4)

0.077

More than 7 years education,* n (%)

190 (52.8)

53 (37.1)

0.001

MMSE score, mean ⴞ SD

27.7 ⫾ 1.7

26.1 ⫾ 2.3

0.000

Having depressive symptoms or diagnosis of depression, n (%)

107 (29.6)

54 (37.5)

0.053

72 (19.9)

43 (29.9)

0.012

228 (63.0)

83 (57.6)

0.155

83 (25.9)

40 (30.8)

0.295 0.270

Having vascular disease, n (%) Having an active and socially integrated lifestyle, n (%) Any ApoE ␧4 alleles† Neuroticism, mean ⴞ SD High score above median, n (%)

8.0 ⫾ 3.8

8.4 ⫾ 3.5

186 (51.4)

72 (50.0)

9.7 ⫾ 2.6

9.7 ⫾ 2.9

189 (52.2)

67 (46.5)

Extraversion, mean ⴞ SD High score above median, n (%)

0.961

*Three and †55 persons with missing data.

whereas the other two groups had similar dementia risk (table 3). When personality traits were examined in relation to lifestyle, low neuroticism was associated with a decreased risk of dementia only among those who had an inactive or socially isolated lifestyle. No such phenomenon was observed for extraversion. A similar pattern was observed when the personality trait combinations were examined in relation to lifestyle. As compared with high neuroticism and high extraversion, the low neuroticism and high extraversion combination was significantly related to a decreased risk of dementia among those with an inactive or

Table 2

Hazard ratios (HR)* and 95% confidence interval (CI) for incident dementia in relation to the separate effect of neuroticism and extraversion HR (95% CI)

Neuroticism One unit decrease in neuroticism score

0.98 (0.93–1.03)

Low neuroticism vs high†

0.70 (0.48–1.03)

Extraversion One unit decrease in extraversion score

1.07 (0.95–1.09)

Low extraversion vs high†

1.25 (0.86–1.81)

*HR were derived from four different Cox proportional hazard models and adjusted for age, sex, education, ApoE ␧4 status, cognitive functioning, presence or absence of vascular diseases, and depressive symptoms. †High and low scores are divided according to median of the respective scores. 256

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socially isolated lifestyle, but not among those with an active and socially integrated lifestyle (table 4). Results were similar in the analyses excluding persons with missing values for any of the personality items. Controlling for lie score, ApoE ␧4 allele status, vascular disease, and survival status did not influence the above results. In addition, the results remained similar in the stratified analyses by age, sex, education, cognitive functioning, presence or absence of ApoE ␧4 alleles, depressive symptoms or diagnoses of depression, and presence of vascular disease. Furthermore, results were unchanged when analyses were restricted to survivors only or to new dementia cases that developed during the last 3-year follow-up. In a community-based cohort of 506 people who initially had no dementia, we found that neither high neuroticism nor low extraversion alone was significantly associated with the risk of developing dementia over a 6-year period. However, a lower risk of dementia was observed for persons with low neuroticism in combination with high extraversion as compared to individuals with high neuroticism and high extraversion. In addition, low neuroticism was associated with a significantly decreased dementia risk among persons with an inactive or socially isolated lifestyle. These findings provide further evidence that certain personality traits may play a role in dementia development, and that personality–lifestyle interactions may be especially important for determining dementia risk. Previous studies have shown that neuroticism captures substantial unique variance in discriminating healthy aging and very mild AD,28 and that older adults with high neuroticism (90th percentile) had double the risk of developing AD compared to those with low neuroticism (10th percentile)11,12 and cognitive decline.29 We did not find a significant association between high neuroticism and increased dementia risk, possibly due to the limited statistical power. Also, earlier studies have used different neuroticism scales.11,12,28,30 We found that neuroticism may interact with extraversion leading to different dementia risks. Prior studies have not examined the combined effect of neuroticism and extraversion in relation to dementia incidence. The decreased dementia risk among persons with low neuroticism and high extraversion in comparison to high neuroticism and high extraversion is not surprising. The high neuroticism and high extraversion combination has been linked to noveltyand excitement-seeking behavior in prior theories of personality31 and was found to be associated with HPA axis activity in response to laboratory-induced psychological stress.32 In contrast, people with low DISCUSSION

Table 3

Hazard ratios (HR)* and 95% confidence interval (CI) for incident dementia in relation to the combined effect of neuroticism and extraversion

Personality traits Neuroticism

Extraversion

Subjects, n

Cases, n

HR (95% CI)

High†

High†

163

46

1

Low

High

93

21

0.51 (0.28–0.94)

Low

Low

155

51

0.95 (0.57–1.60)

High

Low

95

26

0.97 (0.57–1.65)

*HR were derived from Cox proportional hazard model, and adjusted for age, sex, education, ApoE ␧4 status, cognitive functioning, presence or absence of vascular diseases, and depressive symptoms. †High and low neuroticism and extraversion are dichotomized according to median of the respective scores.

neuroticism have a more stable mood and better capacity to handle stressful situations without anxiety. Individuals who score high on extraversion usually have more optimistic outlooks on life in general4 and may be better equipped to cope with stressful events and therefore less prone to depression.33 For these reasons we expected that the combination of low neuroticism and high extraversion should lead to less stress, and, consequently, lower risk of dementia according to the stress-dementia hypotheses. Examination of personality in relation to lifestyle revealed that low neuroticism was associated with a decreased risk of dementia among those with an inactive or socially isolated lifestyle, suggesting that having an Table 4

Hazard ratios (HR)* and 95% confidence interval (CI) for incident dementia in relation to neuroticism and extraversion stratified by lifestyle Active and socially integrated lifestyle

Inactive or socially isolated lifestyle

Cases, n

HR (95% CI)

Cases, n

HR (95% CI)

One unit decrease in neuroticism score

83

1.02 (0.95–1.10)

61

0.93 (0.87–0.99)

Low score vs high†

42

0.81 (0.49–1.35)

30

0.51 (0.27–0.96)

One unit decrease in extraversion score

83

0.97 (0.88–1.06)

6

0.94 (0.84–1.05)

Low score vs high†

42

1.06 (0.62–1.80)

25

1.41 (0.81–2.46)

Neuroticism

Extraversion

Combined effect Neuroticism

Extraversion

High

High

29

1

Low

High

13

0.75 (0.32–1.74)

17

0.33 (0.12–0.90)

Low

Low

28

1.06 (0.54–2.07)

23

0.67 (0.28–1.58)

High

Low

13

1.04 (0.49–2.21)

13

0.88 (0.40–1.98)

8

1

*HR were adjusted for age, sex, education, ApoE ␧4 status, cognitive functioning, and presence or absence of vascular diseases, and depressive symptoms/diagnosis of depression. †High and low scores are divided according to median of the respective scores.

active and socially integrated lifestyle may buffer the negative effect of high neuroticism on dementia risk. These results are in line with a prior study showing that social support may attenuate the negative effect of depression on the progression of coronary heart disease.16 There is evidence that an active and integrated lifestyle may protect older persons against dementia,18 possibly through psychological mechanisms. More active and socially integrated subjects may be psychologically less stressed even if they score high on neuroticism or have distress-prone combinations of personality traits, such as the combination of high neuroticism and low extraversion or high neuroticism and high extraversion, since leisure activities and interactions with social network may help to reduce the stress. This is in line with a recent report on loneliness, where persons who perceived themselves as socially isolated or felt disconnected from others had an increased risk of dementia.34 The detected association may be interpreted as a possible consequence of biologic stress processes due to loneliness.35 Several strengths of the study increase confidence in the findings. First, the current study examined well-established personality traits assessed among community dwellers who were free of dementia both at baseline and at first follow-up. Second, the analyses were based on incident dementia cases detected during a 6-year-long follow-up. Third, we have taken into account a number of potential confounders in the analyses, including adjustment for baseline cognition. Fourth, all the findings were confirmed in several additional analyses. Some limitations of the study need to be addressed. First, personality was assessed only in 70% of the eligible subjects; missing personality data in 30% of the subjects may have affected the results. Second, the refusals and dropouts were older and had poorer cognitive performance than the participants. However, there is no reason to believe that potential misclassifications in dementia diagnoses were due to differences between participants and nonparticipants. Third, personality was assessed only at one occasion. Although individual differences in late adulthood have been observed,36 personality traits are generally stable throughout the life course.37 However, possible personality changes may precede clinical diagnosis of AD.38 Therefore, personality assessed at the first follow-up could still reflect personality changes due to preclinical dementia. However, our study cohort consisted of older people who were without dementia at both baseline and first follow-up examinations, and they were monitored for 6 years to detect new dementia cases. Fourth, we controlled for cognitive performance at first follow-up, which substantially lessens the likeNeurology 72

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lihood that neuropathologic changes were related to personality traits in our cohort. Furthermore, similar results were found in the subpopulation in which only new cases who developed dementia during the last 3 years of follow-up were included. In addition, it is likely that the occurrence of death is a noninformative event for dementia, because the likelihood that death was due to dementia in a dementia free cohort over 6 years is small. However, death might be a competing event for dementia leading to a biased estimation. The current study suggests that personality may influence the risk of dementia development, and that personality– environment interactions may be especially relevant. These findings have significant public health and clinical implications, as the negative effects of certain personality characteristics on dementia are likely to be stress-related, and could be buffered by an active and socially integrated lifestyle.

11.

12.

13.

14.

15.

16.

17. AUTHOR CONTRIBUTIONS H.-X. Wang conducted the statistical analysis.

18. ACKNOWLEDGMENT The authors thank all members of the Kungsholmen Project study group for their cooperation in data collection and management.

19.

Received March 1, 2008. Accepted in final form October 8, 2008. REFERENCES 1. Gallagher M, Landfield PW, McEwen B, et al. Hippocampal neurodegeneration in aging. Science 1996;274:484– 485. 2. Sapolsky RM. Depression, antidepressants, and the shrinking hippocampus. Proc Natl Acad Sci USA 2001; 98:12320–12322. 3. McEwen BS. Protective and damaging effects of stress mediators. N Engl J Med 1998;338:171–179. 4. Pervin LA, John OP. Personality: Theory and Research, 7th ed. Singapore: John Wiley & Sons., 1996. 5. Grant S, Langan-Fox J. Personality and the occupational stressor-strain relationship: the role of the Big Five. J Occup Health Psychol 2007;12:20–33. 6. Mroczek DK, Almeida DM. The effect of daily stress, personality, and age on daily negative affect. J Pers 2004;72: 355–378. 7. Millar K, Purushotham AD, McLatchie E, George WD, Murray GD. A 1-year prospective study of individual variation in distress, and illness perceptions, after treatment for breast cancer. J Psychosom Res 2005;58:335–342. 8. Swickert RJ, Rosentreter CJ, Hittner JB, Mushrush JE. Extraversion, social support processes, and stress. Pers Individ Dif 2002;32:877–891. 9. LeBlanc J, Ducharme MB. Influence of personality traits on plasma levels of cortisol and cholesterol. Physiol Behav 2005;84:677–680. 10. Kendler KS, Gatz M, Gardner CO, Pedersen NL. Personality and major depression: a Swedish longitudinal, population-based twin study. Arch Gen Psychiatry 2006; 63:1113–1120. 258

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Learn. Earn. Network. 2009 AAN Annual Meeting: An Excellent Value • Learn about the latest scientific advances in neurology • Earn valuable CME credit and fulfill Maintenance of Certification requirements • Network with your peers at exciting social events all week long • Enjoy the convenience and value of all this and more—in just one meeting Early registration and hotel deadline is March 20, 2009. Register today at www.am.com/AM2009.

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Autosomal dominant subcortical gliosis presenting as frontotemporal dementia

R.H. Swerdlow, MD B.B. Miller, MD, PhD M.B.S. Lopes, MD J.W. Mandell, MD, PhD G.F. Wooten, MD P. Damgaard, RN C. Manning, PhD M. Fowler, MD H.R. Brashear, MD

Address correspondence and reprint requests to Dr. Russell H. Swerdlow, University of Kansas School of Medicine, Landon Center on Aging, MS 2012, 3901 Rainbow Boulevard, Kansas City, KS 66160 [email protected]

ABSTRACT

Objective: To describe a multigenerational kindred with a frontotemporal dementia clinical syndrome (FTDS), extensive subcortical gliosis pathology, and autosomal dominant genetics.

Methods: Clinical, imaging, and pathologic evaluations of multiple family members. Results: Symptom onset commonly occurred in the fifth or sixth decade, although some kindred members did not develop obvious symptoms until their eighth decade. White matter changes were prominent on both MRI and CT imaging. Results from six brain autopsy evaluations showed consistent but varying degrees of pathology that, while unique, share some histologic similarities with leukodystrophies. These brains were notably devoid of both tau- and ubiquitin-containing inclusions.

Conclusions: Subcortical gliosis in this kindred arises from mutation of a novel gene or else represents a unique frontotemporal dementia clinical syndrome variant caused by mutation of an already known gene. Clinical relevance and research implications are discussed. Neurology® 2009; 72:260–267 GLOSSARY AD ⫽ Alzheimer disease; FTD ⫽ frontotemporal dementia; FTDS ⫽ frontotemporal dementia clinical syndrome; GFAP ⫽ glial fibrillary acidic protein; H-E ⫽ hematoxylin and eosin; HDLS ⫽ hereditary diffuse leukoencephalopathy with spheroids; MDRS ⫽ Mattis Dementia Rating Scale; MMSE ⫽ Mini-Mental State Examination; PD ⫽ Parkinson disease; PrP ⫽ prion protein; SPECT ⫽ single photon emission tomography; TDP43 ⫽ TAR DNA-binding protein 43; UVA ⫽ University of Virginia; VWM ⫽ vanishing white matter disease.

Progressive behavioral and cognitive decline are features of frontotemporal dementia (FTD).1-5 FTD syndromes (FTDS) are classified by whether they initially affect behavior, language expression, or semantic knowledge.6 Predominant unilateral or bilateral frontal or unilateral temporal presentations are described, and parietal lobe pathology is possible.7-10 Histopathologic findings variably consist of Pick bodies, tau inclusions, ubiquitin inclusions, TAR DNAbinding protein 43 (TDP43) inclusions, or no apparent protein aggregations.11,12 Approximately 40% of patients with FTD have a positive family history.13 Mutations in the tau and progranulin genes on chromosome 17 produce autosomal dominant FTDS, and are associated with tauopathy or ubiquitin inclusions.14-17 We now describe a multigenerational FTDS kindred with extensive subcortical gliosis and neither tau nor ubiquitin inclusions. METHODS Clinical descriptions. Six kindred members were evaluated at the Memory (five) and Movement (one) Disorder clinics of the University of Virginia (UVA). We report clinical and structural neuroimaging data, as well as results from relevant hematologic-serologic-urine, neuropsychological, and functional neuroimaging testing.

Pathology. Brain autopsies were available for six members of the kindred. We report essential gross and relevant microscopic findings. RESULTS A progressive disorder of behavior and cognition is suspected across three generations (figure 1). One instance of intermarriage occurred between distantly related cousins, but intermarriage for most branches is not suspected. Individuals with strong clinical or autopsy evidence of subcortical gliosis are

From the Department of Neurology (R.H.S.), University of Kansas School of Medicine, Kansas City; and the Departments of Neuropathology (B.B.M., M.B.S.L., J.W.M.) and Neurology (G.F.W., P.D., C.M., M.F., H.R.B.), University of Virginia School of Medicine, Charlottesville. Disclosure: The authors report no disclosures. 260

Copyright © 2009 by AAN Enterprises, Inc.

Figure 1

Frontotemporal dementia clinical syndrome (FTDS): Subcortical gliosis kindred

Squares represent males, circles represent females, and triangles represent individuals in which gender was not specified in order to enhance anonymity. Subjects for which a diagnosis of FTDS-subcortical gliosis is relatively certain are represented by filled squares/circles. Subjects with putative or possible FTDS are represented by hatched squares/circles.

discussed in detail. Brief descriptions of putatively or potentially affected subjects are provided. Subject IV-6. The UVA index case presented in 1995 at the age of 55 after 4 years of progressive behavioral change (including wandering), disorientation, poor memory, and decreased speaking. Examination revealed orientation only to person, markedly reduced spontaneous speech, and difficulty following multistep commands. There were nonfatiguing frontal release signs and appendicular paratonia. Brain CT showed severe, diffuse atrophy with frontal predominance and severe frontal lobe white matter attenuation. A differential diagnosis of Alzheimer disease (AD), Pick disease, and vascular dementia was considered. In subsequent years he developed mutism, bowel and bladder incontinence, hypertonia, myoclonic jerks, and gait abnormalities (small steps on a widened base). An FTD diagnosis was made before his death in 2001. At autopsy the brain weighed 1,120 g. There was atrophy of the bilateral frontal and tempo-

ral lobes with parietal and occipital sparing. Aside from the cerebellum where Purkinje cell loss was noted, neuron loss was minimal and very rare swollen neuronal cells were identified. No inflammation was seen. GFAP staining revealed hypertrophic astrogliosis within the superficial and deep white matter. There was loss of axonal processes with secondary myelin loss and prominent axonal spheroids (figure 2). Immunohistochemistry did not reveal tau, ubiquitin, or prion protein (PrP)-containing neuronal inclusions. Neurofilament staining occurred only in conjunction with axonal spheroids. Beta-amyloid staining revealed scant plaques and positive staining was present in a small proportion of arteries. Subject III-3. A 1976 non-UVA autopsy report said the patient manifested progressive movement (postural changes and bradykinesia) and cognitive decline during his sixth decade. Parkinson disease (PD) and AD were diagnosed. Pneumoencephalogram showed ventriculomegaly. Lumbar puncture was unremarkable. He died at the age of 59. The brain weighed Neurology 72

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Figure 2

not reveal tau, ubiquitin, TDP43, or PrP-containing neuronal inclusions. Neurofilament staining occurred only in conjunction with axonal spheroids. Neither Consortium to Establish a Registry for Alzheimer’s Disease (CERAD) nor dementia with Lewy body criteria were met. Based on the clinical history of dementia, extensive gliosis, and paucity of AD histology, a final diagnosis of frontotemporal dementia with subcortical gliosis was made.

Histologic findings from IV-6

Subject IV-2. At the age of 58, this subject developed

Representative histologic findings from the mid frontal lobe (A–D) and cerebellum (E) of IV-6 are provided. (A) Transition between cortex (bottom) and white matter (top). There is normal cortical morphology, myelinated U-fibers, and vacuolated demyelinated white matter (hematoxylin and eosin [H-E], x40 before 28.7% reduction). (B) Isometric gliosis involving white matter diffusely and cortex minimally (glial fibrillary acidic protein immunohistochemical stain, x40 before 28.7% reduction). (C) Diffuse white matter demyelination with relative sparing of U-fiber tracts (Luxol fast blue stain with H-E counterstain, x40 before 28.7% reduction). (D) Axonal dropout in white matter, with dystrophic changes including swelling and prominent axonal spheroids (Bielschowsky silver stain, x200 before 28.7% reduction). (E) The cerebellar cortex demonstrates a lack of significant gliosis in conjunction with Purkinje cell loss (H-E, x40 before 28.7% reduction).

1,150 g. Both frontal lobes were atrophic. No infarctions were seen. A marked increase in glia cells was noted. No Alzheimer changes or histologic evidence of PD were seen. Subject III-7. We autopsied but did not clinically ex-

amine this patient. She was institutionalized for dementia during her eighth decade and died at age 77. The brain weighed 1,150 g. The frontal gyri were mildly atrophic. There was marked ventricular dilation and an old cerebellar stroke. There was microvacuolation (spongiotic change) of the cerebral cortex within the outer laminae, reflecting prominent neuronal loss, which was accompanied by gliosis. Glial fibrillary acidic protein (GFAP) staining revealed subcortical hypertrophic astrogliosis. Axonal spheroids were observed. Immunohistochemistry did 262

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personality and behavioral changes. He neglected hygiene, lost interest in his hobbies, lost the ability to joke, began cursing excessively, developed stereotyped behaviors, and could not “finish what he started.” Otherwise, most activities of daily living were preserved. At age 61, outside neuropsychological testing revealed a dysexecutive state; MRI showed bifrontal atrophy and leukoariosis of the subcortical white matter capping the frontal horns (figure 3A). His vitamin B12 level was low. He was started on B12 and referred to the UVA MDC, where his Mini-Mental State Examination (MMSE) score was 25/30. He showed anosognosia, mildly impaired memory retention, and very impaired memory retrieval. Free hand and copy drawing were good. He named 11 animals over one minute and 1 F word over 1 minute. Set-shifting, motor sequencing, and ideomotor praxis were impaired. Aside from reduced odor discrimination and mild gegenhalten paratonia, his general neurologic examination was unremarkable. Behavioral and cognitive deficits progressed and at age 66 he died of pulmonary complications. The brain weighed 1,240 g. Severe frontal atrophy and diffuse white matter degenerative changes were evident (figure 4). There was extensive subpial gliosis with superficial laminar spongiotic change within the cortex (indicating neuronal loss), and extensive subcortical gliosis with moderate myelin loss. GFAP staining revealed hypertrophic astrogliosis (figure 4). White matter axonal dropout and dystrophic changes, including axonal spheroids, were seen. Immunohistochemistry did not reveal tau, ubiquitin, TDP43, or PrP-containing neuronal inclusions. Neurofilament staining occurred only in conjunction with axonal spheroids. Beta-amyloid immunochemistry revealed atypical cortical and white matter plaques with core but not mantle staining. There was amyloid deposition within multiple vessels. However, the hippocampus was unremarkable and other Alzheimer diagnostic features were lacking. Subject IV-15. At age 41, this man was referred to UVA for progressive walking, speaking, and memory decline. He had been fired from his longstanding job about 10 months earlier, and recently gained 15 pounds. MMSE score was 25/30. Dysarthria, re-

Figure 3

Neuroimaging findings

(A) Fluid-attenuated inversion recovery MRI axial images from IV-2 are shown. The superior images show frontal atrophy. Leukoariosis of the white matter capping the frontal horns is the most striking feature. (B) CT images from IV-15 show profound white matter hypodensities capping the frontal horns. (C) Fluid-attenuated inversion recovery MRI axial images from IV-19 show abnormally increased white matter signal of the centrum ovale and corona radiata.

duced repetition abilities, poor motor sequencing, and limb-kinetic apraxia were noted. Frontal release signs, extension plantar reflexes, and a parkinsonian gait were observed. Although an MRI obtained after a mild head injury 6 years earlier was reportedly normal, MRI at the age of 41 showed diffuse cortical atrophy and extensively increased white matter signal capping the frontal and occipital horns and rimming the rest of the lateral ventricles. Frontal leukoariosis was also observed on CT (figure 3B). Spinal fluid studies (including oligoclonal bands) were negative. Urine arylsulfatase A activity, serum arylsulfatase A activity, phytanic acid level, and plasma very long chain fatty acid levels were normal. Commercial spinocerebellar atrophy and triple repeat disorder genetic testing was negative. Progressive irritability, mutism, gait impairment, and bowel-bladder incontinence occurred. He died at age 43. The brain weighed 1,550 g and showed lateral ventricle enlargement with corpus callosum thinning. The hippocampus, midbrain, substantia nigra, pons, and cerebellum were normal. There was remarkable preservation of cortical neurons, which were nearly normal in density. Focal macrophage infiltrates were seen in all lobes and were present in white matter tracts and deep gray nuclei (figure 5).

GFAP staining showed hypertrophic astrogliosis. Degenerating white matter showed myelin loss, numerous dystrophic axons, and axonal swellings and spheroids. Immunohistochemistry did not reveal tau, ubiquitin, TDP43, or PrP-containing neuronal inclusions. Neurofilament staining occurred only in conjunction with axonal spheroids. Electron microscopy revealed no features specific for lysosomal storage disease. Subject IV-20. At the age of 46, this subject began

manifesting a progressive inability to express himself, apathy, social withdrawal, and sleep disturbance. He had a home repair business and lost the ability to use his tools. He presented to UVA approximately 1 year after symptom onset. His general neurologic examination was remarkable only for frontal release signs. MMSE and Mattis Dementia Rating Scale (MDRS) scores were 29/30 and 121/144. Speech was effortful and contained paraphasic errors. There were deficits on tests of visuospatial reasoning, cognitive processing speed, mental flexibility, and conceptualization. MRI showed ventriculomegaly, diffuse mild atrophy, and periventricular white matter disease. Single photon emission tomography (SPECT) showed reduced left subfrontal perfusion. Commercial genetic testing for notch3 gene mutation was negative. A skin biNeurology 72

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Figure 4

Gross brain and histologic findings from IV-2

opsy showed no evidence of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy. Over the next 3 years, he gradually lost the ability to walk and swallow. At death, the brain weighed 1,100 g. Hydrocephalus ex vacuo and prominent frontal lobe atrophy were present. The vasculature showed minimal atherosclerosis. Gross examination revealed diffuse loss of anterior white matter with cortical sparing. Microscopic sections of the cortex showed generalized atrophy, mild neuronal loss, and gliosis. Scant “balloon-like” neurons were seen in the frontal cortex. In addition to extensive gliosis in subpial and gray cortical locations, GFAP staining showed white matter hypertrophic astrogliosis. The white matter was notable for significant loss of myelin and large eosinophilic balls indicative of axonal degeneration. These spheroids were accompanied by a prominent reactive astrocytosis. The thalamus showed focal neuronal loss and gliosis and the midbrain was notable for increased reactive type II astrocytes. Immunohistochemistry did not reveal tau, ubiquitin, alpha-synuclein, or PrP-containing neuronal inclusions. Neurofilament staining occurred only in conjunction with axonal spheroids. A few diffuse neuritic plaques were seen on beta-amyloid immunostaining. Subject IV-19. This woman presented to UVA at the

Moderate atrophy and white matter degenerative change are evident in (A) frontotemporal and (B) parieto-occipital lobe coronal sections. Close-up views of (C) frontal and (D) parietal lobe coronal sections emphasize the severe white matter degeneration with relative cortical preservation. Representative histologic findings from the inferior parietal lobe (E–H) and hippocampus (I–K) of IV-2 are also provided. (E) Transition between cortex (bottom) and white matter (top). There is normal cortical morphology, myelinated U-fibers, and vacuolated, demyelinated white matter (hematoxylin and eosin [H-E], x40 before 28.7% reduction). (F) Isometric gliosis involving white matter diffusely and cortex minimally (glial fibrillary acidic protein immunohistochemical stain, x40 before 28.7% reduction). (G) Diffuse white matter demyelination with relative sparing of U-fiber tracts (Luxol fast blue stain with H-E counterstain, x40 before 28.7% reduction). (H) Axonal dropout in white matter, with dystrophic changes including swelling and prominent axonal spheroids (Bielschowsky silver stain, x200 before 28.7% reduction). (I) Normal hippocampal morphology (H-E, x40 before 28.7% reduction). There was no immunohistochemical evidence of beta-amyloid (J) or tau (K) accumulation (immunochemical stains for beta-amyloid [J] and tau [K], x40 before 28.7% reduction).

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age of 53 with 1 year of language and gait problems. Lower extremity tone was increased. Her gait was markedly apraxic and somewhat magnetic. The noncognitive examination was otherwise unremarkable. MMSE and MDRS scores were 27/30 and 132/144. Terseness of expression was noted. Neuropsychological testing showed executive dysfunction, constructional apraxia when attempting to copy complex but not simple figures (suggesting a deficit in visuomotor integration), and impaired fine motor dexterity. Her vitamin B12 level was borderline low, with an elevated homocysteine. MRI revealed moderate subcortical white matter disease without ventriculomegaly or extensive atrophy (figure 3C). SPECT showed decreased frontal cortical and subcortical perfusion. Vitamin B12 was repleted but gait and cognition continued to decline. She died at age 59. Autopsy was not performed. Putatively or possibly affected subjects. Only one pu-

tatively or possibly affected subject was evaluated at UVA. This was IV-8, who presented at age 51 with memory complaints. Aside from mild difficulty with motor sequencing, the bedside cognitive and general neurologic examinations were unremarkable. An MRI did not indicate white matter pathology. Neuropsychological testing showed an MDRS score of

Figure 5

Histologic findings from IV-15

Histologic micrographs from subject IV-15 (A, B: mid frontal lobe; C, D: inferior parietal lobe) are provided. Gliosis in this subject was multifocally floridly hypertrophic (A) with areas of ischemic infarction (B) and granulomatous inflammation (C), but no other evidence of infection (hematoxylin and eosin, x40 before 28.7% reduction). Axonal dystrophy with spheroids was quite prominent (D, Bielschowsky silver stain, x100 before 28.7% reduction).

130/144, impaired executive function, poor figure copy, and good memory. Repeat neuropsychological testing 3 years later (her last UVA evaluation) showed an MDRS of 123/144 and deficits otherwise similar in quality and quantity to those originally seen. By family report, 2 years later the subject began displaying increasingly poor social judgment. Subject II-1 died in a state psychiatric hospital at the age of 71 with a diagnosis of Pick disease. Subject II-7 died in a state psychiatric hospital at the age of 65 with dementia but no specific diagnosis. Subject III-1 died with a diagnosis of AD, and apparently did not develop dementia until late in her eighth decade. Subjects III-12, III-13, III-17, and III-26 died with diagnoses of AD at the respective ages of 67, 68, 62, and 52. Subject III-27 had clinical dementia at the time of death at the age of 48. Subject III-28 died in a state psychiatric hospital at the age of 45. Subject IV-3 died in a nursing home in her eighth decade. Subject IV-5 has been diagnosed with a neurologic disorder of the CNS and has abnormal white matter on neuroimaging. Subject IV-7 died in her fifth decade with a diagnosis of Pick disease. Subject IV-9 died in her fifth decade. Her clinical diagnoses included multiple sclerosis and Pick disease. Subject IV-21 died at age 50 with a diagnosis of Pick disease.

Paternal transmission indicates this is an autosomal dominant disorder. Penetrance is high but incomplete, and may result because senile symptom onset can occur. Absence of tau and ubiquitin pathology makes tau and progranulin gene mutation unlikely.17,18 Absence of ubiquitin and TDP43 staining also argues abnormal encoding or handling of TDP43 does not play a primary or etiologic role.19 With the exception of very limited congophilic angiopathy in two subjects, there was no evidence of beta-amyloid, alphasynuclein, or prion protein pathology. Hypertrophic astrogliosis without neuronal loss or only minimal neuronal loss in at least some subjects suggests this is a primary disorder of glia, not neurons. The constellation of predominant white matter involvement, roughly equal degrees of demyelination and axonal loss, and axonal spheroids are consistent with membranous lipodystrophy,20,21 dermatoleukodystrophy with neuroaxonal spheroids,22 the adult-onset variant of vanishing white matter disease (VWM),23 and hereditary diffuse leukoencephalopathy with spheroids (HDLS).24 The age at onset and lack of non-CNS manifestations are incompatible with the first two entities. Relatively late symptom onset in many affected members of our kindred and an absence of VWMcharacteristic oligodendroglial morphology argue DISCUSSION

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against VWM. Our kindred does share several clinical, radiologic, and neuropathologic features with HDLS.25,26 Since leukoencephalopathy and axonal spheroids are general markers of subcortical damage, the nosologic relationship between this kindred and HDLS-designated kindreds is unclear. HDLS is itself a relatively recently described entity, and it will be interesting to see whether future studies define HDLS as an etiologically distinct disease or as part of a clinicopathologic syndrome/spectrum. Elucidating the genetic basis of our kindred and the published HDLS kindreds could definitively resolve this issue. There were no ultrastructural features of lysosomal storage or long-chain fatty acid disease. Progressive familial leukodystrophy of late onset and Binswanger disease can manifest white matter pathology with subcortical gliosis,27,28 but our kindred mostly lacks extensive demyelination and does not show arteriolar pathology. An FTD subtype, dementia lacking distinctive histologic features, can also show subcortical gliosis.29-31 Subcortical gliosis as a pathologic entity was itself identified in 1949 and classified as a distinct disorder in 1967.32,33 Autosomal dominant subcortical gliosis kindreds are described.34 One was linked to chromosome 17q21-22 and shown to have an intronic tau gene mutation.35,36 Brains from affected kindred members had obvious tau pathology on immunohistochemical survey.36 In contrast, none of the brains from our kindred had any detectable evidence of tauopathy or abnormal tau immunohistochemistry. Executive dysfunction and frontal leukoariosis commonly occur in the absence of an amnestic state, and older individuals with this type of presentation may be diagnosed with the subcortical ischemic vascular dementia variant of vascular dementia.37-39 Determining the basis of this kindred’s autosomal dominant subcortical gliosis could potentially yield mechanistic insight into vascular-attributed cognitive impairment/dementia syndromes, as well as to what is commonly referred to as small vessel cerebrovascular disease. ACKNOWLEDGMENT The authors thank the members and relations of this kindred who encouraged them to prepare this article, and who also provided indispensable information used for the preparation of this manuscript. They also thank the National Prion Disease Pathology Surveillance Center for performing the PrP immunostaining analysis.

2.

3. 4.

5. 6.

7.

8.

9.

10.

11.

12. 13.

14.

15.

16.

17.

18. 19.

20. Received May 15, 2008. Accepted in final form October 14, 2008.

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Gans A. De ziekten van Pick en van Alzheimer. Nederlandsch Tijdschrift voor Geneeskunde, Amsterdam 1925; 2:1953. Frederick J. Pick disease: a brief overview. Arch Path Lab Med 2006;130:1063–1066. Kertesz A. Progress in clinical neurosciences: frontotemporal dementia-Pick’s disease. Can J Neurol Sci 2006;33: 141–148. Graff-Radford NR, Woodruff BK. Frontotemporal dementia. Semin Neurol 2007;27:48–57. Neary D, Snowden JS, Gustafson L, et al. Frontotemporal lobar degeneration: a consensus on clinical diagnostic criteria. Neurology 1998;51:1546–1554. Edwards-Lee T, Miller BL, Benson DF, et al. The temporal variant of frontotemporal dementia. Brain 1997;120: 1027–1040. Perry RJ, Hodges JR. Differentiating frontal and temporal variant frontotemporal dementia from Alzheimer’s disease. Neurology 2000;54:2277–2284. Swerdlow RH, Gripshover D, Brashear HR, Manning CA. Frontoparietal dementia: clinical and laboratory characterization of three patients. Soc Neurosci Abstr 2000;26: 2005. Cairns NJ, Brannstrom T, Khan MN, Rossor MN, Lantos PL. Neuronal loss in familial frontotemporal dementia with ubiquitin-positive, tau-negative inclusions. Exp Neurol 2003;181:319–326. Snowden J, Neary D, Mann D. Frontotemporal lobar degeneration: clinical and pathological relationships. Acta Neuropathol 2007;114:31–38. Bugiani O. The many ways to frontotemporal degeneration and beyond. Neurol Sci 2007;28:241–244. Chow TW, Miller BL, Hayashi VN, Geschwind DH. Inheritance of frontotemporal dementia. Arch Neurol 1999; 56:817–822. Cruts M, Gijselinck I, van der Zee J, et al. Null mutations in proganulin cause ubiquitin-positive frontotemporal dementia linked to chromosome 17q21. Nature 2006;442: 920–924. Baker M, Mackenzie IR, Pickering-Brown SM, et al. Mutations in progranulin cause tau-negative frontotemporal dementia linked to chromosome 17. Nature 2006;442: 916–919. Hutton M, Lendon CL, Rizzu P, et al. Association of missense and 5=-splice-site mutations in tau with the inherited dementia FTDP-17. Nature 1998;393:702–705. Davion S, Johnson N, Wientraub S, et al. Clinicopathologic correlation in PGRN mutations. Neurology 2007; 69:1113–1121. Tsuboi Y. Neuropathology of familial tauopathy. Neuropathology 2006;26:471–474. Leverenz JB, Yu CE, Montine TJ, et al. A novel progranulin mutation associated with variable clinical presentation and tau, TDP43 and alpha-synuclein pathology. Brain 2007;130:1360–1374. Kitajima I, Kuriyama M, Usuki F, et al. Nasu-Hakola disease (membranous lipodystrophy). Clinical, histopathological and biochemical studies of three cases. J Neurol Sci 1989;91:35–52. Minagawa M, Maeshiro H, Shioda K, Hirano A. Membranous lipodystrophy (Nasu disease): clinical and neuropathological study of a case. Clin Neuropathol 1985; 4:38–45.

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Plasma A␤, homocysteine, and cognition The Vitamin Intervention for Stroke Prevention (VISP) trial

A. Viswanathan, MD, PhD S. Raj S.M. Greenberg, MD, PhD M. Stampfer, MD, DrPH S. Campbell, MHS B.T. Hyman, MD, PhD M.C. Irizarry, MD

Address correspondence and reprint requests to Dr. Anand Viswanathan, Hemorrhagic Stroke Research Program, Massachusetts General Hospital Stroke Research Center, 175 Cambridge Street, Suite 300, Boston, MA 02114 [email protected]

ABSTRACT

Background: Amyloid-beta protein (A␤) plays a key role in Alzheimer disease (AD) and is also implicated in cerebral small vessel disease. Serum total homocysteine (tHcy) is a risk factor for small vessel disease and cognitive impairment and correlates with plasma A␤ levels. To determine whether this association results from a common pathophysiologic mechanism, we investigated whether vitamin supplementation–induced reduction of tHcy influences plasma A␤ levels in the Vitamin Intervention in Stroke Prevention (VISP) study.

Methods: Two groups of 150 patients treated with either the high-dose or low-dose formulation of pyridoxine, cobalamin, and folic acid in a randomized, double-blind fashion were selected among the participants in the VISP study without recurrent stroke during follow-up and in the highest 10% of the distribution for baseline tHcy levels. Concentrations of plasma A␤ with 40 (A␤40) and 42 (A␤42) amino acids were measured at baseline and at the 2-year visit. Results: tHcy levels significantly decreased with vitamin supplementation in both groups. tHcy were strongly correlated with A␤40 but not A␤42 concentrations. There was no difference in the change in A␤40, A␤42 (p ⫽ 0.40, p ⫽ 0.35), or the A␤42/A␤40 ratio over time (p ⫽ 0.86) between treatment groups. A␤ measures were not associated with cognitive change.

Conclusions: This double-blind randomized controlled trial of vitamin therapy demonstrates a strong correlation between serum tHcy and plasma A␤40 concentrations in subjects with ischemic stroke. Treatment with high dose vitamins does not, however, influence plasma levels of A␤, despite their effect on lowering tHcy. Our results suggest that although tHcy is associated with plasma A␤40, they may be regulated by independent mechanisms. Neurology® 2009;72:268–272 GLOSSARY A␤ ⫽ amyloid-beta; AD ⫽ Alzheimer disease; BMI ⫽ body mass index; DBP ⫽ diastolic blood pressure; MMSE ⫽ Mini-Mental State Examination; mRS ⫽ modified Rankin Scale; NIHSS ⫽ NIH Stroke Scale; SBP ⫽ systolic blood pressure; tHcy ⫽ total homocysteine; VISP ⫽ Vitamin Intervention in Stroke Prevention study.

Amyloid ␤-protein (A␤40, A␤42) deposition in the brain is a hallmark of Alzheimer disease (AD) and is thought to be the cause of cognitive impairment and dementia.1 Reduction of A␤ production is a candidate approach for treatment and prevention of cognitive impairment and dementia.2 Plasma total homocysteine (tHcy) levels are correlated with plasma A␤, although the biologic importance of this association is uncertain.3-5 Plasma tHcy has been implicated as a risk factor for small vessel cerebrovascular disease and the development of cognitive impairment and dementia.6-8 There are several potential implications of these associations in relation to AD, cognitive impairment, and microangiopathy. tHcy may increase the risk of AD by elevating A␤ levels. Alternatively, A␤ may increase the risk of microangiopathic changes through elevation of tHcy

From the Memory Disorders Unit (A.V., S.R., S.M.G., B.T.H., M.C.I.) and Stroke Service (A.V., S.M.G.), Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Boston; Harvard School of Public Health (M.S.), Boston; Department of Biostatistics (S.C.), Collaborative Studies Coordinating Center, University of North Carolina at Chapel Hill, Stroke Prevention and Atherosclerosis Research Center. M.C.I. is currently affiliated with WW Epidemiology, GlaxoSmithKline, Research Triangle Park, NC. Supported by NIH grants 5K23NS046327-04, P50AG05134, and 5R01AG026484-02 (Massachusetts General Hospital), the Harvard Center for Neurodegeneration and Repair, and NIH grant 5T32NS048005-05 (Harvard School of Public Health). Disclosure: Michael Irizarry is a stock and options-holding employee of GlaxoSmithKline. No pharmaceutical funding was used in this study. 268

Copyright © 2009 by AAN Enterprises, Inc.

levels. Neurotoxicity may be potentiated by the dual elevation of both tHcy and A␤. Finally, tHcy and A␤ may be markers of a pathogenic mechanism and independent of each other. Since plasma tHcy levels are readily modifiable by high-dose vitamin supplementation, we hypothesized that plasma A␤ levels may also be modifiable by vitamin supplementation. We thus aimed to test whether this association results from a common pathophysiologic mechanism between these biomarkers or if in fact they represent independent processes. The Vitamin Intervention in Stroke Prevention (VISP) was a randomized controlled trial designed to test the hypothesis that lowering tHcy levels with large doses of folic acid, pyridoxine, and vitamin B12 would reduce the incidence of recurrent stroke or myocardial infarction.9 Although the study did not show a benefit for the primary endpoint, tHcy was successfully lowered with vitamin therapy. We investigated plasma A␤ as an add-on component to the VISP study to test the hypotheses whether vitamin supplementation– induced reduction of tHcy over 2 years influences plasma A␤ levels. METHODS Subjects. Details of the VISP trial have been published previously.9 Briefly, the VISP trial enrolled a total of 3,680 adults who 1) were within 120 days of a mild or moderate ischemic stroke (modified Rankin Scale [mRS] score of ⱕ 3); 2) were 35 years or older; and 3) had a fasting tHcy level approximately greater than the 25th percentile for patients with stroke. Subjects were enrolled between September 1996 and December 2001 at 56 centers in the United States, Canada, and Scotland, and randomized to receive a high dose formulation (n ⫽ 1,827) containing 25 mg of pyridoxine, 0.4 mg of cobalamin (B12), and 2.5 mg of folic acid or the low dose formulation (n ⫽ 1,853) of 200 mcg of pyridoxine, 6 mcg of cobalamin, and 20 mcg of folic acid. Baseline VISP data included a standardized medical history, demographic variables, body mass index (BMI), stroke symptoms questionnaire, systolic and diastolic blood pressures, and a neurologic examination. Several scales which measure disability and cognition were administered to all patients (mRS, NIH Stroke Scale [NIHSS], Mini-Mental State Examination [MMSE]),9 and serum levels of folate, B12 levels, and a lipid profile were ascertained. tHcy levels were obtained while fasting and after methionine loading.9 The assembly of the cohort for this substudy is shown in the figure. We selected at random a group of 150 patients treated with high dose formulation and another group of 150 patients with low dose formulation (within sex and 10-year age strata) among participants who did not have a recurrent stroke during the trial and who were in the highest 10% of the distribution for baseline tHcy levels, in order to maximize potential observed

treatment effect. The sample numbers were selected based on power analysis and practical capacity for performing the outcome measures. In the current study, we had 85% power to detect an absolute difference of 7.5% in A␤ levels with high dose vitamin supplementation. Requirements for inclusion in the pool of potential subjects were availability of the following at both baseline and 2-year visits: blood samples, vitamin and tHcy levels, and MMSE. Individuals with less than 75% compliance by pill count were excluded. Plasma A␤ levels were measured in blood samples drawn at baseline and at the 2-year visit. This study was performed with approval by the ethics committees of all study institutions and administrative sites. Written informed consent was obtained from every potential participant. This substudy was performed with approval and in accord with the guidelines of our institutional review boards.

Blood collection. Blood was collected in polypropylene sterile plunger tubes containing potassium ethylenediamine tetraacetic acid. Samples were centrifuged at 1,380 g for 15 minutes, aliquoted with a protease inhibitor cocktail and frozen in dry ice, and stored at ⫺80°C.

Biochemical assays. Plasma tHcy was determined by highperformance liquid chromatography, as detailed in our previous studies.9-11 Plasma A␤40 and A␤42 concentrations were determined by sandwich ELISA using the BNT77 capture antibody and C-terminal specific detector antibodies BA27 and BC05 as previously described and validated.4,12 We have demonstrated this ELISA system to detect A␤40 or A␤42 at concentrations as low as 1 pmol/L and to detect both free and protein-bound A␤.4,12 All biochemical analyses were performed without knowledge of subjects’ clinical or radiographic information. Statistical analyses. For univariate analysis, ␹2 tests were used to compare two categorical variables and analyses of variance were performed to compare continuous variables distributions across groups. All p values were two-tailed and criterion for significance was p ⬍ 0.05. To determine whether vitamin treatment affected plasma A␤ levels, we adopted a linear mixed effects model for the data longitudinally measured over the 2-year period.13 This technique allows for analysis of time-independent and time-dependent variables to identify associations with these variables as well as their trajectories over time. The parameter estimates indicate how much change in plasma A␤ levels resulted from a one unit change in each risk factor. Analyses were also performed using the A␤42/A␤40 ratio, as it has been recently demonstrated that this ratio may be associated with an increased risk of dementia.14,15 For all models of the three outcome measures (A␤40, A␤42, and A␤42/A␤40 ratio), we investigated the effects of age, gender, clinical, and laboratory variables on change in plasma A␤ levels. Covariates that were associated with clinical scales in univariate analysis (p ⬍ 0.10) were considered in the final model.

The baseline clinical and demographic variables are presented in table 1. There were no significant differences in clinical or demographic variables between the two groups. There were no differences between A␤40 or A␤42 levels between treatment groups at baseline (p ⫽ 0.97 and 0.30, respectively). The median follow-up interval was 24.0 months. Levels of tHcy at 2-year follow-up declined in both treatment groups (change in tHcy at 2 years RESULTS

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269

Figure

Assembly of study sample from the Vitamin Intervention in Stroke Prevention (VISP) cohort

period (␤ ⫽ ⫺0.09, p ⫽ 0.44) and there was no significant difference in the change in A␤40 levels between treatment groups (␤ ⫽ 0.14, p ⫽ 0.40). Similarly, there was no significant change in either A␤42 levels or the A␤42-A␤40 ratio over time between treatment groups (p ⫽ 0.35 and p ⫽ 0.86) (table 2). There was no association between A␤40, A␤42 levels, or the A␤42/A␤40 ratio with MMSE at baseline (p ⫽ 0.10, p ⫽ 0.62, and p ⫽ 0.85, respectively) or follow-up (p ⫽ 0.28, p ⫽ 0.74, and p ⫽ 0.50, respectively). A␤ levels did not influence change in MMSE over the treatment period. In this study, we sought to define the relationship between plasma A␤ levels and homocysteine lowering in a cohort of subjects from the randomized controlled VISP trial.9 The current study demonstrates a strong association between tHcy and plasma A␤40 levels in subjects with ischemic stroke in this longitudinal analysis. These findings confirm and extend cross-sectional observational studies which have previously reported this association.3-5 However, despite the association of tHcy levels with A␤40, A␤40 levels were not influenced by vitamin treatment. The strength of this study stems from the fact that subjects had randomized assignment to treatment type and that these subjects were followed prospectively for recurrent stroke and other cardiovascular events over a 2-year period with complete follow-up of all patients. Elevated tHcy is a predictive factor for vascular disease, including ischemic heart disease and stroke.16 Several studies have suggested that elevated tHcy is also a risk factor for white matter disease,17 cognitive impairment,17-20 and AD.6-8 These associations may be explained by vascular21or direct neurotoxic22,23 effects of tHcy. Elevated plasma concentrations of A␤ are associated with microvascular disease in both populationbased epidemiologic studies and cohorts of subjects with cognitive impairment.4,24,25 In vitro studies have suggested direct physiologic or toxic effects of A␤ on the contractile/relaxation elements of the blood vessel wall.26-28 Data regarding plasma A␤ and the risk of cognitive decline are conflicting. Cohort studies have reported that elevated plasma A␤40 or A␤42 levels increase the risk of developing AD over 5– 8 years,14,15,29 although a fourth found that plasma A␤42 levels were not associated with cognitive decline over 30 months.30 Other studies have found low A␤40 or A␤42 levels associated with incident AD14,15 or more rapid cognitive decline in AD subjects.31 Finally, some have suggested that low conDISCUSSION

Recruitment for the VISP cohort is described in detail elsewhere.9 Briefly, patients with a presumptive diagnosis of acute ischemic stroke were screened. Patients with total homocysteine levels (tHcy) that exceeded defined thresholds were randomized for treatment with high- or low-dose vitamin therapy. A random sample of eligible subjects with the highest 10% of tHcy were included in the study.

4.73 ⫾ 8.98 in high treatment group and 1.66 ⫾ 7.79 in the low treatment group; p ⬍ 0.0001 and p ⫽ 0.009, respectively). Reduction of tHcy was significantly greater in the high treatment group [␤ (time ⫻ treatment group) ⫽ ⫺0.1289; p ⬍ 0.0001]. Baseline tHcy levels and tHcy levels at 2-year follow-up were significantly correlated with A␤40 levels (r ⫽ 0.25 and 0.29, respectively; p ⬍ 0.0001 for both comparisons). However, tHcy levels were not correlated with A␤42 levels at baseline (p ⫽ 0.50) or at 2-year follow-up (p ⫽ 0.20). Levels of A␤40, A␤42, and the A␤42-A␤40 ratio at baseline and follow-up are shown in table 2. A␤40 levels did not significantly change over the treatment 270

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Table 1

Baseline characteristics of subjects in cohort according to high or low vitamin treatment group Low-dose group (n ⴝ 157), n (%)

Characteristic

67.2 ⫾ 10.2

Age, y

High-dose group (n ⴝ 143), n (%) 66.7 ⫾ 11.2

Sex

p Value 0.68 0.72

Male

88 (51.4)

93 (48.6)

Female

55 (53.8)

64 (46.2)

Current smoker

25 (55.6)

20 (44.4)

0.64

Ever smoked

99 (51.0)

95 (49.0)

0.54

BMI* (kg/m2)

28.4 ⫾ 5.4

29.6 ⫾ 6.5

0.09

Homocysteine (␮mol/L)

14.6 ⫾ 5.7

15.7 ⫾ 7.9

0.16

Vitamin B12 level

356.8 ⫾ 213.8

369.3 ⫾ 502.8

0.78

Total cholesterol

202.9 ⫾ 42.6

203.3 ⫾ 52.2

0.95

27 ⫾ 3

27 ⫾ 3

0.99

MMSE mRS

1 (0, 2)

1 (0, 2)

0.36

NIHSS

0 (0, 1)

0 (0, 1)

0.77

SBP, mm Hg

141.5 ⫾ 18.4

DBP, mm Hg

78.2 ⫾ 10.7

141.9 ⫾ 20.4

0.85

78.6 ⫾ 9.9

0.71

98.55 ⫾ 6.67

98.61 ⫾ 7.43

0.94

A␤40 (pmol/L)

72.5 ⫾ 44.2

72.4 ⫾ 39.3

0.98

A␤42 (pmol/L)

18.3 ⫾ 17.8

24.4 ⫾ 69.9

0.33

Medication compliance,* %

Values are mean ⫾ SD, median (25th, 75th quartile), or n (%). *Measured at second follow-up visit. BMI ⫽ body mass index; MMSE ⫽ Mini-Mental State Examination; mRS ⫽ modified Rankin scale; NIHSS ⫽ NIH Stroke Scale; SBP ⫽ systolic blood pressure; DBP ⫽ diastolic blood pressure.

centrations of plasma A␤42 in combination with increased concentrations of plasma A␤40 are associated with an increased risk of cognitive impairment and dementia.14,15 This study did not find a significant treatment effect of high dose vitamins on plasma levels of A␤40 despite the effect of the high dose vitamins on lowering tHcy. This suggests that although tHcy is associated with plasma A␤40, they may have independent pathophysiologic mechanisms. This is in contrast to Flicker et al.,33 who detected an effect of tHcy lowering on plasma A␤40. These differences may be reflective of the patient population (stroke patients with high tHcy in the VISP study, community Table 2

Linear mixed model analysis of plasma A␤40, A␤42, and A␤42/A␤40 and plasma tHcy at baseline and 2-year follow-up Mean values at baseline

Mean values at 2-y follow-up

Treatment group ⴛ time p value

Plasma biomarkers

Low-dose

High-dose

Low-dose

High-dose

A␤40 (␮mol/L)

72.5 ⫾ 44.2

72.4 ⫾ 39.3

70.5 ⫾ 43.61

72.8 ⫾ 46.73

0.40

A␤42 (␮mol/L)

18.3 ⫾ 17.8

24.4 ⫾ 69.9

16.0 ⫾ 21.50

25.9 ⫾ 91.81

0.35

A␤42/A␤40

0.32 ⫾ 0.35

0.46 ⫾ 1.08

0.31 ⫾ 0.63

0.37 ⫾ 0.93

0.86

Homocysteine (␮mol/L)

14.6 ⫾ 5.7

15.7 ⫾ 7.9

12.9 ⫾ 5.3

11.0 ⫾ 4.3

⬍0.0001

dwelling older men in the study by Flicker et al.), confounding by dietary changes in folate consumption (VISP study), or differences in the form of A␤ measured by the assays (the assay used in this study measures protein bound A␤, and does not detect oligomeric forms). Additionally, given the definition of the subcohort as those VISP subjects in the highest quintile of tHcy at baseline, a component of the reduction of tHcy may represent regression to the mean rather than vitamin effects. The VISP study results may suggest that tHcy is merely a marker for vascular disease and risk of cognitive decline because tHcy lowering does not influence A␤40 levels.9 Although tHcy is associated with plasma A␤40, high dose vitamin treatment may differentially impact these two plasma markers.4 Finally, correlations between tHcy and other metabolites of the methylation cycle, such as S-adenosylhomocysteine, have been reported.34 How these metabolites respond to vitamin treatment remains to be elucidated. Further epidemiologic and therapeutic studies investigating the relationship between cerebrovascular disease and these potentially important plasma biomarkers (tHcy and A␤40, A␤42) are needed. Received July 3, 2008. Accepted in final form October 8, 2008.

REFERENCES 1. Selkoe DJ. Normal and abnormal biology of the betaamyloid precursor protein. Annu Rev Neurosci 1994;17: 489–517. 2. Greenberg SM, Bacskai BJ, Hyman BT. Alzheimer disease’s double-edged vaccine. Nat Med 2003;9:389–390. 3. Irizarry MC, Gurol ME, Raju S, et al. Association of homocysteine with plasma amyloid beta protein in aging and neurodegenerative disease. Neurology 2005;65:1402–1408. 4. Gurol ME, Irizarry MC, Smith EE, et al. Plasma betaamyloid and white matter lesions in AD, MCI, and cerebral amyloid angiopathy. Neurology 2006;66:23–29. 5. Flicker L, Martins RN, Thomas J, et al. Homocysteine, Alzheimer genes and proteins, and measures of cognition and depression in older men. J Alzheimer Dis 2004;6:329– 336. 6. Clarke R, Smith AD, Jobst KA, Refsum H, Sutton L, Ueland PM. Folate, vitamin B12, and serum total homocysteine levels in confirmed Alzheimer disease. Arch Neurol 1998;55:1449–1455. 7. McCaddon A, Hudson P, Davies G, Hughes A, Williams JH, Wilkinson C. Homocysteine and cognitive decline in healthy elderly. Dement Geriatr Cogn Disord 2001;12: 309–313. 8. Seshadri S, Beiser A, Selhub J, et al. Plasma homocysteine as a risk factor for dementia and Alzheimer’s disease. N Engl J Med 2002;346:476–483. 9. Toole JF, Malinow MR, Chambless LE, et al. Lowering homocysteine in patients with ischemic stroke to prevent recurrent stroke, myocardial infarction, and death: the Vitamin Intervention for Stroke Prevention (VISP) randomized controlled trial. JAMA 2004;291:565–575. Neurology 72

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Smolin LA, Schneider JA. Measurement of total plasma cysteamine using high-performance liquid chromatography with electrochemical detection. Anal Biochem 1988; 168:374–379. Malinow MR, Kang SS, Taylor LM, et al. Prevalence of hyperhomocyst(e)inemia in patients with peripheral arterial occlusive disease. Circulation 1989;79:1180–1188. Fukumoto H, Tennis M, Locascio JJ, Hyman BT, Growdon JH, Irizarry MC. Age but not diagnosis is the main predictor of plasma amyloid beta-protein levels. Arch Neurol 2003;60:958–964. Fitzmaurice G, Laird NM, Ware JH. Applied Longitudinal Analysis. New Jersey: John Wiley and Sons; 2004. Graff-Radford NR, Crook JE, Lucas J, et al. Association of low plasma Abeta42/Abeta40 ratios with increased imminent risk for mild cognitive impairment and Alzheimer disease. Arch Neurol 2007;64:354–362. van Oijen M, Hofman A, Soares HD, Koudstaal PJ, Breteler MM. Plasma Abeta(1-40) and Abeta(1-42) and the risk of dementia: a prospective case-cohort study. Lancet Neurol 2006;5:655–660. Homocysteine and risk of ischemic heart disease and stroke: a meta-analysis. JAMA 2002;288:2015–2022. Dufouil C, Alperovitch A, Ducros V, Tzourio C. Homocysteine, white matter hyperintensities, and cognition in healthy elderly people. Ann Neurol 2003;53:214–221. Lehmann M, Gottfries CG, Regland B. Identification of cognitive impairment in the elderly: homocysteine is an early marker. Dement Geriatr Cogn Disord 1999;10:12–20. Morris MS, Jacques PF, Rosenberg IH, Selhub J. Hyperhomocysteinemia associated with poor recall in the third National Health and Nutrition Examination Survey. Am J Clin Nutr 2001;73:927–933. Miller JW, Green R, Ramos MI, et al. Homocysteine and cognitive function in the Sacramento Area Latino Study on Aging. Am J Clin Nutr 2003;78:441–447. Faraci FM, Lentz SR. Hyperhomocysteinemia, oxidative stress, and cerebral vascular dysfunction. Stroke 2004;35: 345–347. Kruman Kumaravel TS II, Lohani A, Pedersen WA, et al. Folic acid deficiency and homocysteine impair DNA repair in hippocampal neurons and sensitize them to amyloid toxicity in experimental models of Alzheimer’s disease. J Neurosci 2002;22:1752–1762.

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VIEWS & REVIEWS

Genetics of epilepsy syndromes starting in the first year of life

Liesbet Deprez, PhD An Jansen, MD, PhD Peter De Jonghe, MD, PhD

Address correspondence and reprint requests to Prof. Dr. P. De Jonghe, VIB-Department of Molecular Genetics, Neurogenetics Research Group, University of Antwerp-CDE, Universiteitsplein 1, BE-2610 Antwerpen, Belgium [email protected]

ABSTRACT

Background: Incidence rates of epilepsy in children are highest during the first year of life. Most frequently, epilepsy results from a metabolic or structural defect in the brain. However, some infants have clearly delineated epilepsy syndromes for which no underlying etiology can be identified except for a genetic predisposition.

Methods: We reviewed the current knowledge on the genetics of epilepsy syndromes starting in the first year of life. We focus on those epilepsy syndromes without a clear structural or metabolic etiology.

Results: Recent molecular studies have led to the identification of the responsible gene defects for several of the monogenetic epilepsy syndromes with onset in the first year of life. Discussion: This knowledge has consequences for clinical practice as it opens new perspectives for genetic testing, improving early diagnosis, and facilitating genetic counseling. This overview of epilepsy syndromes and associated gene defects might serve as a basis for the selection of patients in whom genetic testing can be helpful. Neurology® 2009;72:273–281 GLOSSARY AD ⫽ autosomal dominant; AED ⫽ antiepileptic drug; AR ⫽ autosomal recessive; AS ⫽ absence seizures; BFIS ⫽ benign familial infantile seizures; BFNIS ⫽ benign familial neonatal-infantile seizures; BFNS ⫽ benign familial neonatal seizures; BMEI ⫽ benign myoclonic epilepsy of infancy; CAE ⫽ childhood absence epilepsy; DD ⫽ developmental delay; EFMR ⫽ epilepsy and mental retardation limited to females; EPI ⫽ epilepsy; FHM ⫽ familial hemiplegic migraine; FIME ⫽ familial infantile myoclonic epilepsy; FoS ⫽ focal seizures; FS ⫽ febrile seizures; GCS ⫽ generalized clonic seizures; GEFSⴙ ⫽ generalized epilepsy with febrile seizures plus; GTCS ⫽ generalized tonic-clonic seizures; ICCA ⫽ infantile convulsions and choreoathetosis; IS ⴝ infantile spasms; ISSX ⫽ X-linked infantile spasms; MR ⫽ mental retardation; MS ⫽ myoclonic seizures; PMR ⫽ psychomotor retardation; RTT ⫽ Rett syndrome; SB ⫽ suppression-burst; TLE ⫽ temporal lobe epilepsy; TS ⫽ tonic seizures; UCS ⫽ unilateral clonic seizures.

The age-specific incidence of epilepsy is high during the first year of life and declines throughout childhood. One reason for the elevated incidence is the large proportion of symptomatic epilepsies presenting at this age. Hypoxic-ischemic encephalopathy is the main cause of neonatal seizures. During infancy, other etiologies, including metabolic encephalopathies, chromosomal disorders, and brain malformations, play a major role.1 Several early-onset epilepsy syndromes have no underlying cause except for a genetic predisposition. Inheritance can be complex, involving several genetic and environmental factors, or monogenetic, caused by a single genetic defect. Recent identification of the causative genes for a number of early-onset epilepsies has created the possibility of genetic testing. The impact of genetic testing on clinical practice in general is relatively limited due to the small percentage of patients in whom a mutation is identified, the absence of specific treatment for patients harboring mutations, and complex implications for genetic counseling. However, genetic testing is useful for a subgroup of patients. Clinical differentiation between benign or malignant epilepsies might prove difficult at onset and may require follow-up time. Identification of the underlying cause, including genetic defects, provides information on prognosis, From the Neurogenetics Group (L.D., P.D.J.), Department of Molecular Genetics, VIB, Antwerpen; Laboratory of Neurogenetics (L.D., P.D.J.), Institute Born-Bunge, Antwerpen; University of Antwerp (L.D., P.D.J.), Antwerpen; Department of Pediatric Neurology (A.J.), Universitair Ziekenhuis Brussel; and Division of Neurology (P.D.J.), University Hospital of Antwerp, Antwerpen, Belgium. Supported by the Fund for Scientific Research Flanders (FWO), University of Antwerp and the Interuniversity Attraction Poles (IUAP) program P6/ 43 of the Belgian Science Policy Office (BELSPO). L.D. is a PhD fellow of the Institute for Science and Technology (IWT), Belgium. Disclosure: The authors report no disclosures. Copyright © 2009 by AAN Enterprises, Inc.

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avoiding unnecessary investigations and helping to select the appropriate treatment. When a genetic defect has been identified, information about the recurrence risk can be offered to the patient’s family. This survey reviews the epilepsy syndromes starting in the first year of life, which have a genetic origin in at least a subgroup of patients. IDIOPATHIC FOCAL EPILEPSIES Three familial focal epilepsy syndromes occur in the first year of life: benign familial neonatal seizures (BFNS), benign familial neonatal-infantile seizures (BFNIS), and benign familial infantile seizures (BFIS). All have similar clinical characteristics but differ in onset age and associated gene defect. These syndromes arise in previously normal neonates or infants. Afebrile focal and prominent motor seizures that secondarily generalize predominate, sometimes associated with apnea, cyanosis, and staring. Seizure frequency varies; some individuals experience a few attacks while others have several clusters per day. Interictal EEGs are normal and patients respond well to antiepileptic drugs (AEDs). Seizures remit after weeks or months. Developmental and intellectual outcome is usually normal. All three syndromes show autosomal dominant inheritance with high penetrance. The incidence of BFNS is 14.4 per 10,000 live births2; the incidences of BFNIS and BFIS are unknown. Early recognition of these benign familial focal epilepsy syndromes is important to prevent unnecessary investigations and may allow for early AED treatment cessation.

Benign familial neonatal seizures. BFNS starts on the second or third day of life, but onset may be delayed during the first month or even up to the third month. Seizures remit spontaneously after a few weeks or months. However, about 11% of patients experience infrequent seizures later in life, with good response to AEDs.2 Mutations in KCNQ2 and KCNQ3, encoding the voltage-gated potassium channel subunits Kv7.2 and Kv7.3, cause BFNS.3-5 Kv7.2 and Kv7.3 together form the heteromeric channel responsible for the M-current. This slowly activating and deactivating potassium current regulates subthreshold electrical excitability of neurons. Approximately 70 different KCNQ2 mutations have been identified including missense, frameshift, nonsense, splice-site mutations, and whole exon deletions. In KCNQ3, only missense mutations have been identified. Analysis in a cohort of unrelated BNFS patients yielded mutations frequencies of 56% (17/30) for KCNQ2 and 6.6% (2/30) for KCNQ3.6 De novo KCNQ2 mutations have also been identi274

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fied in patients with benign neonatal seizures without family history.7 Occasionally, KCNQ2 mutations may present with an aberrant phenotype of neonatal seizures evolving to drug-resistant epileptic encephalopathy and mental retardation. All but one of these mutations were inherited from a parent with BFNS, indicating the involvement of additional genetic or environmental factors.8 KCNQ2 mutations thus may increase the risk to develop a severe epileptic encephalopathy and a good prognosis is not guaranteed for all patients carrying a mutation, which has implications for counseling. Benign familial neonatal–infantile seizures. In BFNIS, seizures start between day 2 and 7 months of age and remit within the first year.9 BFNIS is caused by missense mutations in SCN2A, encoding the ␣2subunit of the voltage-gated sodium channel.10 This channel, consisting of a pore forming ␣-subunit and one or two regulatory ␤-subunits, is responsible for generation and propagation of action potentials in neurons. The prevalence of SCN2A mutations in a large cohort of BFNIS families has not yet been investigated. A preliminary study of SCN2A in unrelated patients with benign neonatal or infantile seizures without KCNQ2 mutations yielded a mutation frequency of 37.5% (3/8) (personal data). Benign familial infantile seizures. BFIS starts between

4 and 8 months and seizures spontaneously remit before the age of 3 years.11 BFIS is genetically heterogeneous with two loci identified (table), but the causative genes remain unknown. In several families, BFIS later in life cosegregates with paroxysmal choreoathetosis occurring spontaneously or induced by movements, exertion, startling, or anxiety. This syndrome, called infantile convulsions and choreoathetosis, shows linkage to one of the pure BFIS loci.12 BFIS associated with familial hemiplegic migraine has been reported in a single family which carried a mutation in ATP1A2, encoding the ␣2-subunit of the Na⫹, K⫹-ATPase.13 Benign infantile seizures also occur in isolated patients. This nonfamilial form has been subdivided in two types based on clinical characteristics: benign partial epilepsy in infancy with complex partial seizures and benign partial epilepsy in infancy with secondary generalized seizures.14 The International League Against Epilepsy classification combines the familial and nonfamilial forms into a single syndrome since there are no clear clinical differences.15 The genes involved in BFIS thus may be implicated in the nonfamilial form. The sporadic occurrence can be explained by de novo mutations or reduced penetrance.

Table

Genes and loci involved in the epilepsies starting in the first year of life without clear structural or metabolic etiology First report

Syndrome

Inheritance pattern

Locus

Locus name

Gene

Gene product

BFNS

Monogenetic, AD

20q13.3

EBN1

KCNQ2

Subunit voltage-gated K⫹ channel, Kv7.2

3, 4

8q24

EBN2

KCNQ3

Subunit voltage-gated K⫹ channel, Kv7.3

5

SCN2A

␣-subunit voltage-gated Na⫹ channel, Nav1.2

10

BFNIS

Monogenetic, AD

2q24

BFIS

Monogenetic, AD

19q12–q13.11

BFIS1

?

?

49

16p12–q12

BFIS2

?

?

50

ICCA

Monogenetic, AD

16p12–q12

?

?

12

BFISⴙFHM

Monogenetic, AD

1q21–q23

FHM2

ATP1A2

␣-subunit Na⫹, K⫹ATPase pump

13

FIME

Monogenetic, AR

16p13

EIM

?

?

31

FS

Monogenetic, AD

8q13–q21

FEB1

?

?

51

19p

FEB2

?

?

52

2q23–q24

FEB3

?

?

53

5q14–q15

FEB4

(MASS1)

Monogenic Audiogenic Seizure Susceptibility 1

16, 54

6q22–q24

FEB5

?

?

55

18p11.2

FEB6

(IMPA2)

Myo-inositol monophosphatase 2

17

(SEZ6)

Seizure-related 6

18

Complex

17q11.2 GEFSⴙ

Monogenetic, AD

19q13

GEFSP1

SCN1B

␤-subunit voltage-gated Na⫹ channel

21

2q24–q33

GEFSP2

SCN1A

␣-subunit voltage-gated Na⫹ channel, Nav1.1

22

5q34

GEFSP3/FEB8

GABRG2

␥-subunit GABAa receptor 23

(SCN2A)

␣-subunit voltage-gated Na⫹ channel, Nav1.2

28

2q24–q33

FSⴙCAE

FSⴙEPI FSⴙTLE

Ohtahara

Dravet

West

EFMR

2p24

GEFSP4

?

?

56

Complex

1p36

GEFSP5

(GABRD)

␦-subunit GABAa receptor

29

Monogenetic, AD

5q34

ECA2

GABRG2

␥-subunit GABAa receptor 26

Monogenetic, AD⫹ modifier

3p24–p23⫹18p

FEB9

?

?

57

Monogenetic, AD

21q22

FEB7

?

?

58

?

?

59

ETL2

?

?

60

Digenetic

1q25–q31⫹18qter

Monogenetic, AD

12q22–q23

Monogenetic, de novo

9q34.1

STXBP1

Syntaxin binding protein 1

33

Monogenetic, X-linked, recessive

Xp21

ARX

Aristaless-related homeobox protein

40

Monogenetic, de novo

2q24–q33

SCN1A

␣-subunit voltage-gated Na⫹ channel, Nav1.1

36

complex

5q34

(GABRG2)

␥-subunit GABAa receptor 27

Monogenetic, X-linked, recessive

Xp21

ARX

Aristaless-related homeobox protein

40

Monogenetic, X-linked, de novo

Xp22

CDKL5

Cyclin-dependent kinaselike 5

43

Monogenetic, X-linked, only females affected

Xq22

PCDH19

Protocadherin 19

48

The genes indicated between parentheses need additional evidence to prove their involvement. BFNS ⫽ benign familial neonatal seizures; AD ⫽ autosomal dominant; BFNIS ⫽ benign neonatal-infantile seizures; BFIS ⫽ benign familial infantile seizures; ICCA ⫽ infantile convulsions and choreoathetosis; FHM ⫽ familial hemiplegic migraine; FIME ⫽ familial infantile myoclonic epilepsy; AR ⫽ autosomal recessive; FS ⫽ febrile seizures; GEFS⫹ ⫽ generalized epilepsy with febrile seizures plus; CAE ⫽ childhood absence epilepsy; EPI ⫽ epilepsy; TLE ⫽ temporal lobe epilepsy; EFMR ⫽ epilepsy and mental retardation limited to females.

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EPILEPTIC SYNDROMES ASSOCIATED WITH FEBRILE SEIZURES Febrile seizures. Febrile sei-

zures (FS) occur between the age of 3 months to 5 years and affect 2–5% of children in Caucasian populations. Several inheritance patterns have been described including polygenic and autosomal dominant with reduced penetrance. Linkage analysis has identified six loci for pure FS (FEB1-6, table) and for two loci a causative gene has been proposed: MASS1 (Monogenic Audiogenic Seizure Susceptibility 1) in FEB416 and IMPA2 (myo-inositol monophosphatase 2) in FEB6.17 However, additional evidence is necessary to prove their pathogenic role. Mutation analysis of the functional candidate gene SEZ6 (Seizures related gene 6) identified heterozygous mutations in 35% of FS patients (21/60).18 However, more studies are required to confirm the contribution of SEZ6 in FS. Generalized epilepsy with febrile seizures plus. In

some families, FS constitute the main part of a broader phenotypic spectrum including febrile seizures plus (FS⫹) and afebrile seizures, i.e., generalized epilepsy with febrile seizures plus (GEFS⫹). FS⫹, the second most frequent phenotype, is characterized by FS extending beyond the age of 6 years or by afebrile generalized tonic-clonic seizures (GTCS) occurring in addition to FS. Less common phenotypes include FS⫹ associated with generalized seizures such as absences, myoclonic, or atonic seizures, but also focal seizures. Rarely, GEFS⫹ patients have isolated afebrile seizures in childhood. Most patients have benign spontaneously remitting childhood epilepsies and normal development. However, more severe phenotypes such as myoclonic-astatic epilepsy or Dravet syndrome have been described.19 Since GEFS⫹ is associated with both generalized and focal seizures, the use of the term autosomal dominant epilepsy with febrile seizures plus (ADEFS⫹) has been suggested.20 The clinical heterogeneity complicates the identification of the disorder in individual patients. In sporadic patients or small families, FS may be the only manifestation of GEFS⫹. GEFS⫹ has originally been described in large families showing autosomal dominant inheritance with 50 – 60% penetrance.19 Causal mutations in three genes—SCN1B,21 SCN1A,22 and GABRG2 23 — have been identified. SCN1B encodes the ␤1-subunit of the voltage-gated sodium channel and four different mutations have been identified. One study obtained an SCN1B mutation frequency of 4.1% (4/98) in patients with GEFS⫹ and FS.24 SCN1A, encoding the ␣1-subunit of the voltagegated sodium channel, is the most important GEFS⫹ gene, responsible for 5–10% of familial GEFS⫹ patients.25 All GEFS⫹-associated mutations 276

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are missense mutations. Mutations in GABRG2 have been identified in families with GEFS⫹ and FS associated with childhood absence epilepsy.23,26 GABRG2 encodes the ␥2-subunit of the GABAA receptor, a ligand-gated chloride channel responsible for fast synaptic inhibition in the mature brain. In a cohort of ⬎200 unrelated patients with GEFS⫹, FS, and IGE, only one patient carried a mutation, suggesting a GABRG2 mutation frequency below 1%.27 Mutations in the known GEFS⫹ genes account for less than 20% of families.19 In the majority of families and in almost all sporadic patients the cause remains unknown. Mutation analysis of these genes may be considered in patients with FS or FS associated with afebrile seizures and with a positive family history of FS or afebrile seizures. Genetic testing of isolated FS patients is less recommended since the mutation frequency in these patients is very low. In addition to the three GEFS⫹ genes, one SCN2A mutation has been reported in a small family in which the proband had FS and afebrile seizures.28 However, subsequent studies did not support a role for SCN2A in GEFS⫹. GEFS⫹ also frequently occurs in sporadic patients or in small families with a complex inheritance pattern involving more than one gene. GABRD, encoding the ␦-subunit of the GABAA receptor, has been proposed as susceptibility gene for GEFS⫹.29 Several loci have been identified for FS associated with different types of epilepsy (table), but the causative genes are not yet identified. Benign myoclonic epilepsy of infancy (BMEI) is characterized by brief generalized myoclonic seizures with onset between 4 months and 3 years. The etiology of BMEI is unknown but involvement of genetic factors has been suggested.30 Familial occurrence of myoclonic epilepsy of infancy has been described in a single family as familial infantile myoclonic epilepsy (FIME). Linkage analysis identified a recessive locus on chromosome 16p13 but the disease-causing gene is still unknown.31 OTHER IDIOPATHIC EPILEPSIES

MALIGNANT EPILEPSY SYNDROMES Some children with seizures beginning in the first year of life have a poor outcome with psychomotor retardation and refractory chronic epilepsy. These malignant epilepsies are often symptomatic and can have acquired or genetic causes. Genetic diseases frequently associated with early-onset epilepsy include chromosomal abnormalities (e.g., Wolf-Hirschhorn syndrome), inborn errors of metabolism (e.g., nonketotic hyperglycinemia), malformations of cortical development (e.g., lissencephaly), and diseases with a complex pathogenic mechanism (e.g., Rett syn-

drome). These disorders usually produce a complex clinical picture of which epilepsy is only a part. They can usually be diagnosed by detailed clinical investigation, neuroimaging, and biochemical analyses. An extensive description of the symptomatic epilepsies is beyond the scope of this review. Epileptic encephalopathies. In epileptic encephalopa-

thies, very frequent or severe seizures or subcontinuous paroxysmal interictal activity contribute to the progressive disturbance of cerebral function. The disorders selected for this review are characterized by predominance of early-onset seizures associated with developmental regression, absence of detectable metabolic or structural brain defects, and a proven genetic origin in at least part of the patients. Ohtahara syndrome. Ohtahara syndrome or early infantile epileptic encephalopathy syndrome represents only 0.2% of the epilepsies in children under the age of 15 years. It is characterized by early onset of intractable tonic spasms, suppression-burst pattern on interictal EEG, and poor prognosis. All patients have seizure onset in the first 3 months of life, with the majority in the first month. One third of patients have also focal seizures or asymmetric tonic seizures, but myoclonus is rare. The suppression-burst pattern on EEG consists of high-voltage activity alternating with nearly flat suppression phases. Ohtahara syndrome has a characteristic age-dependent evolution; it often evolves into West syndrome between the ages of 3 to 6 months and further into Lennox Gastaut between the ages of 1 and 3 years. Prognosis is poor, with severe psychomotor retardation and high mortality rate during infancy. Ohtahara syndrome is often symptomatic and structural brain abnormalities are the main cause.32 In some patients, however, no structural or metabolic etiology can be identified. Heterozygous mutations in STXBP1 (syntaxin binding protein 1 gene) have been identified in sporadic patients with Ohtahara syndrome. The mutations, including four missense mutations and one whole gene deletion, all arose de novo. The STXBP1 protein plays an essential role in synaptic vesicle release and secretion of neurotransmittors.33 The high mutation frequency of 35.7% (5/14) suggests the importance of STXBP1 mutations in this syndrome, making genetic testing useful in patients with Ohtahara syndrome without metabolic or structural brain defects. In addition, polyalanine expansions in ARX (Aristaless-related homeobox gene) have been found in two male patients with Ohtahara syndrome.34 ARX mutations were originally identified as the cause of X-linked recessive infantile spasms. Dravet syndrome. Dravet syndrome or severe myoclonic epilepsy of infancy is a rare epileptic syndrome

affecting approximately 1 in 40,000 children. Clinical cardinal features include febrile or afebrile generalized or hemiconvulsions starting in the first year of life, seizure evolution to a mixture of intractable generalized (myoclonic or atonic seizures, atypical absences) and focal seizures, normal early development, subsequent psychomotor retardation, and normal brain imaging at onset. Unlike suggested by the name severe myoclonic epilepsy of infancy, myoclonus is not present in all patients. Interictal EEGs are initially normal but progressively show generalized spike-and-waves, polyspike-and-waves, focal abnormalities, and photosensitivity. Seizures are difficult to treat, despite some sensitivity to well-chosen treatment regimens. Valproate, topiramate, and stiripentol may be helpful while lamotrigine and carbamazepine may exacerbate seizures. Since elevation of body temperature can trigger seizures, rigorous treatment of fever and avoiding hot baths are advocated.35 Dravet syndrome usually occurs in sporadic patients and heterozygous de novo SCN1A mutations have been found in 33–100% of patients.25,36 More than 250 different mutations have been identified, including missense, nonsense, frameshift, splice-site mutations, and chromosomal microdeletions encompassing SCN1A.37 Not all patients with Dravet syndrome are isolated; in some families, affected sibs carry the same SCN1A mutation due to germline mosaicism in one of the parents.38 The clinical features defining Dravet syndrome develop over time and the diagnosis is often not made until the second year of life. Identification of an SCN1A mutation may therefore facilitate early diagnosis and appropriate treatment. SCN1A mutations are not present in all patients diagnosed with Dravet syndrome, suggesting genetic heterogeneity. GABRG2 has been proposed as a susceptibility gene.27 Infantile spasms. West syndrome. West syndrome is characterized by a specific seizure type, i.e., epileptic spasms, a unique interictal EEG pattern termed hypsarrhythmia, and psychomotor retardation. Spasms start within the first year of life, mainly between 4 and 6 months of age. The hypsarrhythmia EEG pattern consists of random, high-voltage, nonsynchronous spikes and slow waves with variable duration and topography. Patients with epileptic spasms without hypsarrhythmia should not be diagnosed with West syndrome. Instead, infantile spasms (IS) can be used as a general term for age-related epilepsy syndromes manifesting epileptic spasms. Most patients with West syndrome have poor outcome, with chronic intractable epilepsy and mental retardation. Neurology 72

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Incidence of West syndrome ranges from 2–5 per 10,000 children.39 West syndrome is often symptomatic resulting from various causes including neonatal asphyxia, meningoencephalitis, tuberous sclerosis complex, and chromosomal abnormalities. However, the idiopathic form of West syndrome with a multifactorial genetic predisposition has also been described.39 X-linked infantile spasms. In some families, X-linked inheritance is present, and two forms have been described: recessive and dominant X-linked infantile spasms (ISSX). In recessive ISSX, male patients have IS associated with mental retardation and normal brain imaging. Female obligate carriers are asymptomatic. Recessive ISSX is caused by mutations in ARX 40 and in-frame expansions in the first or second polyalanine tract of the protein are the most common mutations. Expansion length seems to correlate with severity of the phenotype.34 In one family with an ARX missense mutation, the male patients had myoclonic seizures and GTCS as predominant seizures types.41 ISSX patients frequently develop severe generalized dystonia. Analysis of ARX in unrelated male patients with IS and developmental delay with unknown etiology identified mutations in 3.5% of patients (4/113).42 ARX mutation analysis should be considered in families containing multiple male patients manifesting IS, mental retardation, and dystonia. Dominant ISSX was first described in two female patients with early-onset severe IS and developmental arrest. Both patients carried a balanced X-autosome translocation disrupting CDKL5 (cyclin-dependent kinase like 5).43 To date, the phenotypic spectrum associated with CDKL5 mutations has broadened, including myoclonic encephalopathy, atypical Rett syndrome, and the early-onset seizure variant of Rett syndrome. Seizures start before the age of 3 months and are difficult to treat. Seizure types include generalized seizures such as tonic spasms, myoclonic seizures, and GTCS, besides focal seizures. All patients have mental retardation with normal brain imaging. Some patients manifest Rett-like features such as stereotypic hand movements and hand apraxia. All CDKL5 mutations arose de novo and most patients are isolated.44 Parental germline mosaicism has been reported.45 Mutations in CDKL5 are mainly found in female patients, suggesting that CDKL5 mutations are often gestational lethal in males. Frequency of CDKL5 mutations was 17% (7/42) in female patients with early-onset epilepsy associated with severe mental retardation of unknown etiology, and 10% (1/10) in female patients with IS.46 Mutation analysis of CDKL5 can be considered in female patients with severe in278

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tractable seizures starting in the first 3 months of life without a metabolic or structural defect. When IS and Rett-like features are also present, this genetic test is mandatory.44 Epilepsy and mental retardation limited to females. Epilepsy and mental retardation limited to females (EFMR) is an underrecognized disorder with X-linked inheritance but surprisingly only affecting females while sparing transmitting males. Seizure, cognitive, and psychiatric phenotypes show heterogeneity. Seizures start from the age of 6 to 36 months and may be precipitated by fever. Seizure types include GTCS, myoclonic and tonic seizures, absences, and focal seizures. Interictal EEGs are normal or show generalized and focal abnormalities. Most patients have normal early development followed by regression starting around seizure onset or later. Some patients, however, show normal development without regression while others have slow development from birth. Intellectual outcome varies from normal to severe disability. Psychiatric features form a characteristic part of EMFR and include autism spectrum disorders, obsessive features, and aggressive behavior.47 Seven different mutations of PCDH19 (protocadherin 19), including missense, nonsense, and frameshift mutations, have been reported as the cause of EFMR.48 EFMR is currently only recognized in rare large families as the unique inheritance pattern is the key to diagnosis. The frequency of this disorder in small families and isolated patients is unknown. Mutation analysis of PCDH19 should be considered in families containing only female patients with seizures starting in infancy, especially when associated with developmental delay. DISCUSSION Genetic factors play a role in several epilepsy syndromes starting in the first year of life. Molecular genetic studies have identified causative genes and loci for a number of these early-onset epilepsies (table), creating the possibility of genetic testing. Despite these breakthroughs, genetic testing has considerable limitations, as it is only possible for monogenetic epilepsies, whereas several epilepsy syndromes have a complex inheritance pattern. The figure contains guidelines for the selection of patients who can be considered for genetic testing. Some syndromes, including BFNS and Dravet syndrome, have an easily recognizable presentation and high mutation frequency in the known genes. For these disorders, genetic testing may confirm the diagnosis in early stages of the disease, avoiding unnecessary investigations and allowing optimal treatment. Molecular diagnosis in patients with an epileptic encephalopathy provides information about

Figure

Schematic overview of the phenotypic characteristics of patients in whom genetic testing may be useful

The characteristics indicated between parentheses are frequently present but are not mandatory. Frequencies indicated with (*) were obtained in patients with a positive family history. AED ⫽ antiepileptic drugs; AS ⫽ absence seizures; DD ⫽ developmental delay; FoS ⫽ focal seizures; FS ⫽ febrile seizures; GCS ⫽ generalized clonic seizures; GTCS ⫽ generalized tonic-clonic seizures; IS ⫽ infantile spasms; MR ⫽ mental retardation; MS ⫽ myoclonic seizures; PMR ⫽ psychomotor retardation; SB ⫽ suppression-burst; TS ⫽ tonic seizures; UCS ⫽ unilateral clonic seizures; RTT ⫽ Rett syndrome.

the recurrence risk and may allow preimplantation or prenatal testing. Asymptomatic female carriers of an ARX mutation have 50% chance to transmit the mutation to their sons, who will develop the severe epilepsy syndrome ISSX. In EFMR families, virtually all daughters (reduced penetrance has been described) of a man carrying the PCDH19 mutation will be affected. Dravet syndrome and dominant ISSX are usually seen in sporadic patients since causal mutations occur de novo. However, for both disorders rare familial cases due to parental germline mosaicism have been reported. These families have a higher recurrence risk than expected, making preimplantation or prenatal testing advisable. Unfortunately, the yield of several molecular analyses is low due to extensive genetic heterogeneity, as in FS and GEFS⫹, limiting their clinical application with the currently available costly technology. As a

consequence, absence mutation in a known gene does not exclude the disorder. There are further limitations; mutations in a particular gene may result in variable disease severity, making the prognosis of an individual mutation carrier difficult to predict. Apart from the exceptional situation of genetic counseling in the epileptic encephalopathies, predictive testing of asymptomatic relatives is not recommended, especially since development of epilepsy later in life cannot be prevented. Knowledge of underlying molecular mechanisms will allow the development of novel therapeutic strategies for specific epilepsy syndromes. One example is the use of retigabine for BFNS patients with a KCNQ2 or KCNQ3 mutation reducing the M-current. Retigabine, an AED which selectively enhances the current of M-type potassium channels, may therefore be used as a specific treatment.2 In clinical practice the use of retigabine may Neurology 72

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be limited due to the benign course of BFNS. The devastating epileptic encephalopathies are still intractable and this strategy will hopefully lead to the development of effective therapies, by specifically targeting the underlying pathomechanism.

16.

17.

18. ACKNOWLEDGMENT The authors thank Lieve Claes and Arvid Suls for reading the manuscript.

19.

Received May 20, 2008. Accepted in final form October 6, 2008.

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Scheffer IE, Turner SJ, Dibbens LM, et al. Epilepsy and mental retardation limited to females: an under-recognized disorder. Brain 2008;131:918–927. Dibbens LM, Tarpey PS, Hynes K, et al. X-linked protocadherin 19 mutations cause female-limited epilepsy and cognitive impairment. Nat Genet 2008;40:776–781. Guipponi M, Rivier F, Vigevano F, et al. Linkage mapping of benign familial infantile convulsions (BFIC) to chromosome 19q. Hum Mol Genet 1997;6:473–477. Caraballo R, Pavek S, Lemainque A, et al. Linkage of benign familial infantile convulsions to chromosome 16p12q12 suggests allelism to the infantile convulsions and choreoathetosis syndrome. Am J Hum Genet 2001;68: 788–794. Wallace RH, Berkovic SF, Howell RA, Sutherland GR, Mulley JC. Suggestion of a major gene for familial febrile convulsions mapping to 8q13-21. J Med Genet 1996;33: 308–312. Johnson EW, Dubovsky J, Rich SS, et al. Evidence for a novel gene for familial febrile convulsions, FEB2, linked to chromosome 19p in an extended family from the Midwest. Hum Mol Genet 1998;7:63–67. Peiffer A, Thompson J, Charlier C, et al. A locus for febrile seizures (FEB3) maps to chromosome 2q23-24. Ann Neurol 1999;46:671–678. Nakayama J, Hamano K, Iwasaki N, et al. Significant evidence for linkage of febrile seizures to chromosome 5q14q15. Hum Mol Genet 2000;9:87–91. Nabbout R, Prud’homme JF, Herman A, et al. A locus for simple pure febrile seizures maps to chromosome 6q22q24. Brain 2002;125:2668–2680. Audenaert D, Claes L, Claeys KG, et al. A novel susceptibility locus at 2p24 for generalised epilepsy with febrile seizures plus. J Med Genet 2005;42:947–952. Nabbout R, Baulac S, Desguerre I, et al. New locus for febrile seizures with absence epilepsy on 3p and a possible modifier gene on 18p. Neurology 2007;68:1374–1381. Hedera P, Ma S, Blair MA, et al. Identification of a novel locus for febrile seizures and epilepsy on chromosome 21q22. Epilepsia 2006;47:1622–1628. Baulac S, Picard F, Herman A, et al. Evidence for digenic inheritance in a family with both febrile convulsions and temporal lobe epilepsy implicating chromosomes 18qter and 1q25-q31. Ann Neurol 2001;49:786–792. Claes L, Audenaert D, Deprez L, et al. Novel locus on chromosome 12q22-q23.3 responsible for familial temporal lobe epilepsy associated with febrile seizures. J Med Genet 2004;41:710–714.

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CLINICAL IMPLICATIONS OF NEUROSCIENCE RESEARCH

␥-Hydroxybutyric acid and its relevance in neurology

Section Editor Eduardo E. Benarroch, MD

Eduardo E. Benarroch, MD

Address correspondence and reprint requests to Dr. Eduardo E. Benarroch, Mayo Clinic, 200 First Street SW, West 8A Mayo Bldg, Rochester, MN 55905 [email protected]

␥-Hydroxybutyric acid (GHB, oxybate) is formed from ␥-aminobutyric acid (GABA) in the normal human brain. GHB can also be administered exogenously and readily passes the blood– brain barrier. In the brain, GHB may have a widespread neuromodulatory role that is mediated both by GHB-specific and ␥-aminobutyric acid type B (GABAB) receptors. There has been widespread interest in GHB as a result of its emergence as a major recreational drug. Exogenously administered GHB elicits intoxication, withdrawal, tolerance, and addiction. GHB also induces slow wave sleep and has now been approved for treatment of narcolepsy. GHB may also have limited use as an anesthetic and in alcohol dependence and withdrawal. Accumulation of GHB may underlie some of the manifestations of succinic semialdehyde dehydrogenase deficiency (SSADH) in humans. The development of a murine model of this disease, together with that of specific GHB receptor ligands, has provided insights into the physiologic role of GHB in the brain and rationale for therapeutic intervention in neurologic disorders. These subjects have all been extensively reviewed.1-8 BIOCHEMISTRY AND PHARMACOLOGY OF

GHB is present in the normal brain and serves as both a precursor and a degradation product of GABA1-4 (figure). GABA is formed from glutamate by action of the glutamic acid decarboxylase and is degraded to succinic semialdehyde (SSA) by the GABA transaminase. GHB may be formed from SSA by action of the succinic semialdehyde reductase or can be converted back to SSA via the GHB dehydrogenase. Succinic semialdehyde is metabolized in the mitochondria by succinic semialdehyde dehydrogeGHB

nase (SSADH) to succinic acid, which enters the Krebs cycle or can be converted back to GABA. The distribution of GHB and its synthesizing enzyme in GABA-containing neurons suggests that GHB might colocalize with GABA in some inhibitory nerve terminals. GHB undergoes active vesicular uptake probably via the same vesicular inhibitory amino acid transporter that mediates the uptake of GABA and glycine. GHB is released by calcium (Ca2⫹)-dependent exocytosis and undergoes sodium-dependent presynaptic uptake.4 GHB binds to specific guanosine nucleotide binding (G) protein coupled receptors,9 which are distributed particularly in the hippocampus, neocortex, and thalamus.10 However, many of the pharmacologic and clinical effects of exogenously administered GHB are probably mediated via the GABAB receptor, where GHB might act both directly as a partial agonist and indirectly by serving as a precursor of GABA.4 There are several differences between GHB and the GABAB receptors (table 1). Studies in vitro indicate that GHB is a weak GABAB receptor agonist, with an affinity in the millimolar range, which is several orders of magnitude higher than the physiologic 1– 4 ␮M concentrations of GHB in the brain.11 The high GHB concentrations required to activate GABAB receptors may allow enough GHB to be converted to GABA. The conversion of GHB to GABA can be inhibited by ethosuximide and valproate.8 The functional GABAB receptor is a heterodimer consisting of the GABAB(1) and the GABAB(2) receptor and couples to various effector systems through Gi/o proteins (figure). Presynaptic GABAB receptors, as well as the GHB receptors, inhibit voltage-gated Ca2⫹ channels (VGCC) required for exocytosis, thus reducing neurotransmitter release. These receptors may act as inhibi-

GLOSSARY ACh ⫽ acetylcholine; AMPA ⫽ ␣-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid; DA ⫽ dopamine; EDS ⫽ excessive daytime sleepiness; GABA ⫽ ␥-aminobutyric acid; GABA-T ⫽ GABA transaminase; GAD ⫽ glutamic acid decarboxylase; GBL ⫽ ␥-butyrolactone; GH ⫽ growth hormone; GHB ⫽ ␥-hydroxybutyric acid; GHBDH ⫽ GHB dehydrogenase; GIRK ⫽ G-proteincoupled inward rectifying K⫹ channels; 5-HT ⫽ serotonin; NADPH ⫽ nicotinamide adenine dinucleotide phosphate; NMDA ⫽ N-methylD-aspartate; SSA ⫽ succinic semialdehyde; SSADH ⫽ succinic semialdehyde dehydrogenase deficiency; SSAR ⫽ succinic semialdehyde reductase; VGCC ⫽ voltage-gated Ca2⫹ channels; VIAAT ⫽ vesicular inhibitory amino acid transporter; VTA ⫽ ventral tegmental area. From the Department of Neurology, Mayo Clinic, Rochester, MN. Disclosure: The author reports no disclosures. 282

Copyright © 2009 by AAN Enterprises, Inc.

Figure

␥-Hydroxybutyric acid (GHB) and synaptic transmission

GHB serves as both a precursor and a degradation product of ␥-aminobutyric acid (GABA). GABA is formed from glutamate by glutamic acid decarboxylase (GAD) and is degraded to succinic semialdehyde (SSA) by GABA transaminase (GABA-T). GHB may be formed from SSA by action of the succinic semialdehyde reductase (SSAR) or converted back to SSA via the GHB dehydrogenase (GHBDH). SSA can be converted back to GABA or can be metabolized by succinic semialdehyde dehydrogenase (SSADH) to succinic acid. GHB may be colocalized with GABA in inhibitory nerve terminals and GHB undergoes active vesicular uptake probably via the vesicular inhibitory amino acid transporter (VIAAT) and undergoes presynaptic uptake. GHB binds to specific receptors (GHBR) and to GABAB receptors, which are both coupled to the G protein Gi/o. Presynaptic GABAB receptors, as well as the GHB receptors, inhibit voltage-gated Ca2⫹ channels (VGCC) required for exocytosis. Postsynaptically, GABAB receptors produce slow inhibitory postsynaptic potentials via G-protein-coupled inward rectifying K⫹ channels (GIRK). AMPA ⫽ ␣-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid; NMDA ⫽ N-methylD-aspartate; GBL ⫽ ␥-butyrolactone.

tory autoreceptors in GABA-containing neurons or as inhibitory heteroreceptors in neurons releasing glutamate, monoamines, or other neurotransmitters. Postsynaptically, GABAB receptor activation produces slow inhibitory postsynaptic potentials via G-proteincoupled inward rectifying K⫹ channels (GIRK), resulting in decreased neuronal excitability.3 PHYSIOLOGIC EFFECTS OF GHB Experimental evidence suggests that GHB might have a role as a neurotransmitter or neuromodulator, but its precise functions are unknown. The cloning and functional characterization of the GHB receptors9 and the development of selective GHB receptor antagonists12 have provided insight into the relative contribution of GHB and GABAB receptors to the effects of GHB in the brain. GHB affects several brain neurochemical systems4 (table 2). Electrophysiologic studies in rodent brain slices show dual effects of GHB depending on its

concentrations. At physiologic concentrations, GHB inhibits presynaptic VGCC and release of GABA or glutamate in the cortex, hippocampus, and thalamus; these effects are blocked by the GHB receptor antagonist NCS382. In contrast, the effects of millimolar concentrations of GHB, which are attained after its exogenous administration, are blocked by GABAB receptor antagonists. These effects include absence seizures in experimental animals and addiction, tolerance, intoxication, withdrawal, memory impairment, and increased slow wave sleep in humans. In humans, low oral doses induce short-term anterograde amnesia and potential for abuse; higher doses result in drowsiness and sleep; and still higher does lead to general anesthesia. Dosages higher than 50 mg per kilogram can result in coma, cardiorespiratory depression, seizures, and death. GHB-induced sleep, coma, and seizures. Important

targets of GHB are the thalamocortical circuits inNeurology 72

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Table 1

Differences between ␥-hydroxybutyric acid (GHB) and ␥-aminobutyric acid (GABA)B receptors

Receptor

GHB

GABAB

Affinity for GHB

High

Low

GABA binding

No

Yes

Affinity to baclofen

None

High

Coupling to Gi/o

Yes

Yes

Inhibition of adenylyl cyclase

Yes

Yes

Synaptic effects

VGCC inhibition

VGCC inhibition GIRK activation

Distribution in the cortex

Layer I–III

Layer IV–VI

Density in the hippocampus

High

Moderate

Density in the thalamus

Moderate

High

Density in the cerebellum

None

High

volved in the sleep-wake cycle and susceptibility to generalized seizure. In vitro studies indicate that GHB, acting via GABAB receptors, reduces neuronal activity in thalamic, hippocampal, and neocortical neurons. GHB administered to healthy volunteers13 or to patients with narcolepsy/cataplexy6,7 elicits an increase in slow-wave sleep (stage N3), which is characterized by the presence of delta waves in the EEG. Delta waves, like other sleep-related EEG patterns, critically depend on neuronal activity within the cortico-thalamo-cortical loop, reflecting both the intrinsic electrophysiologic properties of thalamic and cortical neurons and the balance between excitatory and inhibitory synaptic influences on these cells.14 GHB affects thalamocortical circuits via both presynaptic and postsynaptic GABAB receptors in a dose-dependent manner.3 Presynaptically, low GHB

concentrations primarily decrease glutamate release from corticothalamic terminals whereas intermediate concentrations inhibit GABA release from reticulothalamic terminals. High GHB concentrations directly hyperpolarize thalamocortical neurons via postsynaptic GABAB receptors. These combined presynaptic and postsynaptic effects of GHB lead to progressive hyperpolarization of thalamocortical neurons resulting in rhythmic burst activity and delta oscillations in thalamocortical circuits. At very high doses, GHB may elicit loss of excitability in thalamocortical networks, as may occur during overdose or coma.3 The effects of GHB in the thalamus may also predispose to development of generalized seizures. Absence seizures elicited by GHB administered either systemically or locally in the thalamus are the best pharmacologic model of human typical absence seizures.15 Since the sensitivity of presynaptic inhibitory GABAB heteroreceptors in glutamatergic corticothalamic terminals is higher than that of presynaptic autoreceptors in GABAergic reticular thalamic neurons, GHB elicits imbalance between excitatory and inhibitory drive to thalamocortical neurons. This, together with hyperpolarization of thalamocortical neurons induced via postsynaptic GABAB receptors, may result in increase in rhythmic burst firing of thalamocortical neurons and thalamocortical synchronization, which lead to development of absence seizures.3,15 GHB and drug addiction. GHB affects the mesolim-

bic dopaminergic circuit, which is critically involved in mechanisms of drug addiction.3 Elegant in vitro studies indicate that GHB, acting via GABAB receptors,16 may exert a bidirectional, dose-dependent ef-

Effects of ␥-hydroxybutyric acid on neurotransmitter systems

Table 2

Neurotransmitter system

Effect

Potential implications

GABA

Presynaptic inhibition of release

Absence seizures Drug addiction

Glutamate

Presynaptic inhibition of release

Increase slow wave sleep Absence seizures

Dopamine (DA)

284

Decreases activity of midbrain DA neurons and DA release in the striatum

Anticraving effect

Elicits upregulation of striatal D1 and D2 receptors

Addictive properties

Serotonin (5-HT)

Increases 5-HT turnover

Inhibition of REM sleep?

Acetylcholine (ACh)

Decreases ACh release in striatum and brainstem

Amnestic effects

Neurosteroids

Increases levels of progesterone and neuroactive steroids (such as allopregnanolone)

Potentiates GABAA receptor mediated sedative, hypnotic, anesthetic, and anticonvulsant effects

Growth hormone (GH)

Increases GH levels independently of GABA B receptors

Use to increase athletic performance

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fect on the overall output of dopaminergic neurons in the ventral tegmental area (VTA), a key brain reward area.17 These dual effects may reflect differences in the distribution of GIRK channels in local inhibitory GABAergic neurons and in output dopaminergic neurons in the VTA. At concentrations typically seen with recreational use, GHB preferentially inhibits local GABAergic neurons in the VTA, thereby increasing dopamine release in the nucleus accumbens, which is typical of the reinforcing effect of many drugs of abuse. At higher concentrations, GHB hyperpolarizes the dopaminergic VTA neurons, resulting in decrease in dopamine output, which may explain the anti-craving properties of GHB.17 Chronic GHB administration also induces a compensatory upregulation of dopaminergic D1 and D2 receptors and may also desensitize GHB and GABAB receptors in dopaminergic neurons; this could contribute to the addictive properties of GHB.4 GHB and neuroprotection. There is in vitro evidence that GHB may exert neuroprotective effects by several mechanisms.18 GHB may spare energy utilization, reduce oxidative stress, block excitotoxicity, and maintain vascular integrity. For example, under hypoxic or ischemic conditions, GHB was shown to spare ATP utilization and reduce the accumulation of lactate. The ability of GHB to shift the intermediary glucose metabolism to generate the reducing equivalents such as nicotinamide adenine dinucleotide phosphate (NADPH), which is required for the elimination of reactive oxygen species and lipid peroxides, may provide a basis for the effects of GHB against oxidative stress. CLINICAL CORRELATIONS Treatment of narcolepsy. Narcolepsy is characterized by excessive day-

time sleepiness (EDS), cataplexy, sleep paralysis, and hypnagogic hallucinations. Patients with narcolepsy enter REM sleep more rapidly than usual and their sleep is often fragmented, with reduced time spent in stage N3 sleep. Sodium oxybate, the sodium salt of GHB, has been approved by the Food and Drug Administration for treatment of the two major symptoms of narcolepsy, EDS and cataplexy, based on two randomized, double blind, placebo-controlled trials, as recently reviewed.6,7 The reduction of EDS may reflect improvement of sleep architecture, increasing slow-wave sleep duration and decreasing REM sleep duration and nocturnal awakenings. Succinic semialdehyde dehydrogenase deficiency. Suc-

cinic semialdehyde dehydrogenase deficiency (GHB aciduria) is an autosomal recessive disorder due to a mutation affecting the SSADH (ALDH5A1, alde-

hyde dehydrogenase 5 family, member A1) gene,8 resulting in impaired conversion of SSA to succinic acid and accumulation of GHB and GABA. SSADH has considerable phenotypic heterogeneity with predominance of neurologic and psychiatric manifestations. They include neurodevelopmental delay affecting predominantly expressive language, hypotonia, hyporeflexia, ataxia, inattention, hyperactivity, and sleep disturbances, including EDS. The clinical course is generally static, with improvement of gait ataxia and language over time, but about 10% of patients have progressive developmental regression, dystonia, choreoathetosis, or myoclonus. About half of these patients suffer generalized absence, myoclonic, or generalized tonic-clonic seizures. MRI shows cerebral and cerebellar (particularly vermian) atrophy and enhanced T2 signal in the globus pallidus, subcortical white matter, dentate nucleus, and substantia nigra. Proton MRS shows increase in brain GABA concentration. There is marked elevation of GHB and to a lesser extent GABA and reduced level of glutamine in the CSF. GHB aciduria may not always be detected during organic acid analysis; the gold standard for diagnostic confirmation is enzyme assay in white blood cells. The SSDH (Aldhh5a1) ⫺/⫺ knockout mice provide a model of human SSADH deficiency.5,8 These mice develop neonatal absence seizures and ataxia and tonic-clonic status epilepticus by the third to fourth week of age, leading eventually to death. In these mice, there is accumulation of GHB and GABA, as well as glutamine, arginine, dopamine, and guanidino species.19 The receptor antagonist NCS-382 or GABAB receptor antagonists increase survival in these animals. Therapeutic approaches to reduce GHB levels in SSADH deficiency include vigabatrin, which inhibits GABA transaminase, taurine, which may act as an antagonist of GABA B receptors and other drugs, as reviewed elsewhere.8 GHB intoxication, withdrawal, and dependence. GHB and its precursor ␥-butyrolactone (GBL) have become widespread drugs of abuse. GHB induces euphoria, anxiolysis, and disinhibition, and has also been used because of its anabolic growth hormonereleasing effect. GHB is currently a controlled substance classified by the National Institute on Drug Abuse as an illicit drug. However, GBL, which can be easily converted to GHB, is available in health food stores. The steep dose-response curve of GHB in the brain predisposes to overdosing. Acute GHB intoxication typically manifests with reduced level of alertness, ranging from sedation to coma, which may be associated with bradycardia, respiratory depression, hypothermia, vomiting, lack of gag reflex, and miosis or mydriasis. However, GHB intoxication Neurology 72

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may also produce paradoxical agitation, combativeness, or self-injury. The prolonged use of GHB can induce a dependence syndrome, which can be associated with a severe withdrawal phenomenon that resembles ethanol withdrawal and includes anxiety, insomnia, and tremor progressing to delirium, hallucinations, autonomic instability, and seizures. It has been proposed that the GABAB receptor agonist baclofen may be the drug of choice for future treatment trials for GHB dependency and withdrawal. GHB has emerged as a drug of abuse and appears to have an important role in the pathophysiology of SSADH deficiency. However, GHB is also an effective treatment for narcolepsy and may also have a role in the treatment of alcohol dependence and withdrawal,20 stimulus-induced paroxysmal drop attacks in Coffin-Lowry syndrome,21 tardive dyskinesia, and bipolar disorder.22 GHB may have neuroprotective effects in neurodegenerative disorders such as Alzheimer disease.18 The development of a murine model of SSADH, the cloning of a putative GHBR, and the development of novel GHB ligands may help determine the physiologic and pathophysiologic role of GHB in the CNS.

9.

10.

11.

PERSPECTIVE

REFERENCES 1. Maitre M. The gamma-hydroxybutyrate signaling system in brain: organization and functional implications. Prog Neurobiol 1997;51:337–361. 2. Wong CG, Gibson KM, Snead OC, 3rd. From the street to the brain: neurobiology of the recreational drug gamma-hydroxybutyric acid. Trends Pharmacol Sci 2004;25:29–34. 3. Crunelli V, Emri Z, Leresche N. Unravelling the brain targets of gamma-hydroxybutyric acid. Curr Opin Pharmacol 2006;6:44–52. 4. Drasbek KR, Christensen J, Jensen K. Gamma-hydroxybutyrate–a drug of abuse. Acta Neurol Scand 2006;114: 145–156. 5. Gupta M, Hogema BM, Grompe M, et al. Murine succinate semialdehyde dehydrogenase deficiency. Ann Neurol 2003;54 Suppl 6:S81–90. 6. Morgenthaler TI, Kapur VK, Brown T, et al. Practice parameters for the treatment of narcolepsy and other hypersomnias of central origin. Sleep 2007;30:1705–1711. 7. Robinson DM, Keating GM. Sodium oxybate: a review of its use in the management of narcolepsy. CNS Drugs 2007;21:337–354. 8. Knerr I, Pearl PL, Bottiglieri T, Snead OC, Jakobs C, Gibson KM. Therapeutic concepts in succinate semialdehyde dehydrogenase (SSADH; ALDH5a1) deficiency (gammahydroxybutyric aciduria). Hypotheses evolved from 25

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years of patient evaluation, studies in Aldh5a1-/- mice and characterization of gamma-hydroxybutyric acid pharmacology. J Inherit Metab Dis 2007;30:279–294. Andriamampandry C, Taleb O, Kemmel V, Humbert JP, Aunis D, Maitre M. Cloning and functional characterization of a gamma-hydroxybutyrate receptor identified in the human brain. Faseb J 2007;21:885–895. Castelli MP, Mocci I, Langlois X, et al. Quantitative autoradiographic distribution of gamma-hydroxybutyric acid binding sites in human and monkey brain. Brain Res Mol Brain Res 2000;78:91–99. Wu Y, Ali S, Ahmadian G, et al. Gamma-hydroxybutyric acid (GHB) and gamma-aminobutyric acidB receptor (GABABR) binding sites are distinctive from one another: molecular evidence. Neuropharmacology 2004;47:1146– 1156. Castelli MP, Pibiri F, Carboni G, Piras AP. A review of pharmacology of NCS-382, a putative antagonist of gamma-hydroxybutyric acid (GHB) receptor. CNS Drug Rev 2004;10:243–260. Van Cauter E, Plat L, Scharf MB, et al. Simultaneous stimulation of slow-wave sleep and growth hormone secretion by gamma-hydroxybutyrate in normal young men. J Clin Invest 1997;100:745–753. Llinas RR, Steriade M. Bursting of thalamic neurons and states of vigilance. J Neurophysiol 2006;95:3297–3308. Gervasi N, Monnier Z, Vincent P, et al. Pathway-specific action of gamma-hydroxybutyric acid in sensory thalamus and its relevance to absence seizures. J Neurosci 2003;23: 11469–11478. Pistis M, Muntoni AL, Pillolla G, et al. Gammahydroxybutyric acid (GHB) and the mesoaccumbens reward circuit: evidence for GABA(B) receptor-mediated effects. Neuroscience 2005;131:465–474. Cruz HG, Ivanova T, Lunn ML, Stoffel M, Slesinger PA, Luscher C. Bi-directional effects of GABA(B) receptor agonists on the mesolimbic dopamine system. Nat Neurosci 2004;7:153–159. Mamelak M. Alzheimer’ s disease, oxidative stress and gammahydroxybutyrate. Neurobiol Aging 2007;28:1340– 1360. Jansen EE, Verhoeven NM, Jakobs C, et al. Increased guanidino species in murine and human succinate semialdehyde dehydrogenase (SSADH) deficiency. Biochim Biophys Acta 2006;1762:494–498. Nava F, Premi S, Manzato E, Campagnola W, Lucchini A, Gessa GL. Gamma-hydroxybutyrate reduces both withdrawal syndrome and hypercortisolism in severe abstinent alcoholics: an open study vs. diazepam. Am J Drug Alcohol Abuse 2007;33:379–392. Havaligi N, Matadeen-Ali C, Khurana DS, Marks H, Kothare SV. Treatment of drop attacks in Coffin-Lowry syndrome with the use of sodium oxybate. Pediatr Neurol 2007;37:373–374. Berner JE. A case of sodium oxybate treatment of tardive dyskinesia and bipolar disorder. J Clin Psychiatry 2008;69: 862.

Clinical/Scientific Notes

M.A. van Es, MD F.P. Diekstra, MD J.H. Veldink, MD, PhD F. Baas, PhD P.R. Bourque, MD H.J. Schelhaas, MD, PhD E. Strengman E.A.M. Hennekam, PhD D. Lindhout, MD, PhD R.A. Ophoff, PhD L.H. van den Berg, MD, PhD

Supplemental data at www.neurology.org

A CASE OF ALS-FTD IN A LARGE FALS PEDIGREE WITH A K17I ANG MUTATION

Methods. A total of 39 unrelated FALS patients, negative for SOD1 mutations, were screened for ANG mutations. This study was approved by the local ethics committee and participants provided informed consent. DNA was isolated from venous blood and ANG mutation analysis was performed as described in appendix e-1. A total of 275 unrelated, healthy controls were taken from a prospective population-based study on ALS in The Netherlands and were also screened. 5 PMut (http://mmb2.pcb.ub.es:8080/PMut/) was used to predict the impact of an amino acid substitution on the structure and function of the protein.

Cases III-3, III-4, and IV-1 all presented with progressive upper and lower motor neuron loss of limbs. Case III-1 rapidly developed weakness in both arms with atrophy, fasciculations, and dyspnea, but no upper motor neuron signs. The patient died after 6 months from onset. Case III-2 initially presented with parkinsonism (bradykinesia, diminished postural reflexes, cogwheel rigidity [right arm], shuffling, short-stepped gait, and decreased spontaneous eye blink rate). There was no autonomic dysfunction and eye movements were intact. Dopaminergic treatment had little effect. After 5 years, the patient developed progressive weakness of the arms and legs with atrophy, fasciculations, and hyperreflexia. Interestingly, the patient also demonstrated symptoms characteristic of frontotemporal dementia (FTD), such as loss of interest in social contacts and family, short attention span, logopenia, verbal apraxia, perseveration, decreased personal hygiene, hyperorality, reckless behavior in traffic, sexual disinhibition, and apathy. Case I-2 and II-4 also appear to have been affected. However, no medical records were available. Patient I-2 developed limb weakness at age 70, leading to paralysis and death within 3 years. Patient II-4 developed speech impairment at age 60 and also died within 3 years. Patient II-2 (obligate carrier) died at age 50 from cardiovascular disease. Detailed clinical characteristics are provided in table e-2.

Results. We identified one mutation in one patient (122 A⬎T) (figure, A), leading to an amino acid substitution of lysine to isoleucine (K17I) (figure, B). PMut analysis predicted this mutation to be pathogenic. Sequence alignments of ANG in different species demonstrated high conservation (figure, C). Analysis of this pedigree revealed an autosomal dominant inheritance of the mutation (male to male transmission) (figure, D). DNA was available from 44 out of 62 family members (five affected individuals). All affected family members carried the K17I mutation. Ten carriers were identified, but all were under 50 years of age (except one who was 75 years old without symptoms or signs of ALS). The K17I mutation was not found in 275 control samples.

Discussion. Several ANG mutations in FALS have been reported, but clear segregation of mutations with the disease has not been shown. Here, we report the K17I mutation segregating with disease in a large pedigree. The fact that II-2 and a carrier (75 years of age) were without symptoms of ALS suggests incomplete penetrance of the mutation. This might explain why mutations in this codon have only been found in SALS. The K17I mutation was previously reported in three cases and K17E in two cases.3,6 This study provides a report of a patient with an ANG mutation and ALS, FTD, and parkinsonism. Five percent of patients with ALS also have FTD and up to 50% demonstrated mild cognitive impairment. Similarly, relatives of patients with ALS have an increased risk for developing PD. Therefore, genes involved in ALS are also considered candidate genes for

Approximately 90% of amyotrophic lateral sclerosis (ALS) cases are sporadic (SALS), but 10% are familial (FALS). Mutations in SOD1, Alsin, Dynactin, SETX, DJ-1, VAPB, and TDP-431 have been reported (table e-1 on the Neurology® Web site at www.neurology.org). After the identification of sequence variation VEGF in patients with ALS, mutations in another angiogenic gene (ANG) were identified in SALS and FALS.2,3 Studies in other populations have identified ANG mutations in patients with ALS, but also in healthy controls. This suggests that not all mutations are pathogenic.3,4

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Figure

Mutation analysis and partial pedigree

ANG is highly conserved between species, suggesting it has an important biologic function. Modeling of the K17I mutation using PMut predicted this to be pathogenic. Two functional studies demonstrated that the K17I mutation results in loss of function, possibly leading to insufficient ribosomes synthesis, decreased protein translation, and ultimately decreased motor neuron viability.6,7 We report segregation of the K17I mutation with FALS and a patient with FALS, FTD, and parkinsonism, which possibly implicates ANG in these diseases. From the Department of Neurology, Rudolf Magnus Institute of Neuroscience (M.A.v.E., F.P.D., J.H.V., L.H.v.d.B.), Department of Medical Genetics and Rudolf Magnus Institute of Neuroscience (E.S., R.A.O.), and Department of Medical Genetics (E.A.M.H., D.L.), University Medical Center Utrecht; Department of Neurogenetics (F.B.), Academic Medical Center, Amsterdam, The Netherlands; Division of Neurology (P.R.B.), University of Ottawa, Ontario, Canada; and Department of Neurology (H.J.S.), Radboud University Medical Center, Nijmegen, The Netherlands. Supported by Netherlands Organization for Scientific Research (NWO) and the Prinses Beatrix Foundation (PBF). Disclosure: The authors report no disclosures. Received June 12, 2008. Accepted in final form August 12, 2008. Address correspondence and reprint requests to Dr. Leonard H. van den Berg, Department of Neurology, Rudolf Magnus Institute of Neuroscience, University Medical Center Utrecht, Heidelbergaan 100, 3584 CX, Utrecht, The Netherlands; L.H.vandenBerg@ umcutrecht.nl Copyright © 2009 by AAN Enterprises, Inc.

ACKNOWLEDGMENT The authors thank the families for their participation.

1. 2.

3.

(A) Wild type sequence and the K17I mutation. (B) Three-dimensional structure of ANG modeling the K17I mutation in ball-and-stick representation. The figure was created using the program PyMOL (DeLano Scientific). (C) Sequence alignments of ANG in different species. Sequences were multiply aligned using Homologene (http://www.ncbi.nlm.nih.gov/sites/entrez/ query.fcgi?db ⫽ homologene). The numbering on top of the alignments correlates with the human amino acid sequence. Amino acid 17, which is the site of K17I, is indicated in red. (D) A simplified version of the pedigree is depicted to protect the privacy of the family. A partial four-generation pedigree is shown. All individuals marked in black have amyotrophic lateral sclerosis. All individuals carrying the122 A⬎T (K17I) mutation are marked in the pedigree. The obligate carrier (II-2) died at 50 years of age due to cardiovascular disease. No DNA was available from this individual. The spouse of II-2 tested negative for the mutation.

other neurodegenerative disorders. Indeed, an Italian study reported a SALS patient with a 132C¡T mutation and frontal lobe dysfunction.4

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4.

5.

6.

7.

Valdmanis PN, Rouleau GA. Genetics of familial amyotrophic lateral sclerosis. Neurology 2008;70:144–152. Greenway MJ, Alexander MD, Ennis S, et al. A novel candidate region for ALS on chromosome 14q11.2. Neurology 2004;63:1936–1938. Greenway MJ, Andersen PM, Russ C, et al. ANG mutations segregate with familial and ‘sporadic’ amyotrophic lateral sclerosis. Nat Genet 2006;38:411–413. Gellera C, Colombrita C, Ticozzi N, et al. Identification of new ANG gene mutations in a large cohort of Italian patients with amyotrophic lateral sclerosis. Neurogenetics 2008;9:33–40. van Es MA, Van Vught PW, Blauw HM, et al. ITPR2 as a susceptibility gene in sporadic amyotrophic lateral sclerosis: a genome-wide association study. Lancet Neurol 2007; 6:869–877. Wu D, Yu W, Kishikawa H, et al. Angiogenin loss-offunction mutations in amyotrophic lateral sclerosis. Ann Neurol 2007;62:609–617. Crabtree B, Thiyagarajan N, Prior SH, et al. Characterization of human angiogenin variants implicated in amyotrophic lateral sclerosis. Biochemistry 2007; 46:11810–11818.

A. Gonzalez-Duarte, MD S. Sullivan G.J. Sips T. Naidich, MD G. Kleinman, MD J. Murray S. Morgello, MD I. Germano, MD M. Mullen, MD D. Simpson, MD

Supplemental data at www.neurology.org

INFLAMMATORY PSEUDOTUMOR ASSOCIATED WITH HIV, JCV, AND IMMUNE RECONSTITUTION SYNDROME

A 37-year-old HIV-positive African woman developed severe chronic diarrhea. Her CD4⫹ T cell count was 25 cells/mm3. Within 1 month of initiation of highly active antiretroviral therapy (HAART) her plasma HIV viral load became undetectable and CD4⫹ T cell count rose to 96 cells/mm3, and continued to rise over the following months. Two months after the initiation of HAART she developed vertigo, loss of balance, incoordination, slurred speech, and tremor of the neck and limbs. Neurological examination revealed ocular abnormalities, dysarthria, and monotonic speech. She had bilateral limb dysmetria, past-pointing and endpoint tremor, impaired heel-knee-shin testing, head tremor, and a wide based, ataxic gait. Initial brain MRI revealed a confluent, nonenhancing area of signal abnormality predominantly involving the inferior right cerebellar hemisphere and extending to the posterior vermis, right cerebellar peduncle, and inferomedial aspect of the left cerebellar hemisphere. Six months later, MRI revealed progression of the cerebellar lesion, with nodular enhancement along the inferomedial aspect of the right cerebellar hemisphere. The patient remained clinically stable. MRI 8 months later revealed a large cystic ring-enhancing lesion in the location of the previously noted high signal intensity lesions of the cerebellum, with compression of the posterior fourth ventricle (figure, A). CSF revealed WBC 1, Prot 64, Gluc 49, and negative cytomegalovirus DNA PCR, Cysticercosis IgG Ab, Epstein-Barr virus DNA PCR, Venereal Disease Research Laboratory, and Cryptococcus Ag. Bacterial, viral, and fungal cultures were negative. JCV PCR was positive. Stereotactic biopsy of the cerebellar lesion, performed 17 months after the onset of neurologic symptoms, revealed giant cells with pleomorphic hyperchromatic nuclei, often multiple, surrounded by a dense infiltrate of lymphocytes and plasma cells (figure, B). The bizarre, pleomorphic cells were GFAP positive, demonstrated diffuse nuclear reactivity for p53 antigen, a high MIB-1 (Ki67) index, and focal, faint reactivity for polyoma virus T antigen (figure e-1 on the Neurology® Web site at www.neurology.org). Inflammatory infiltrates marked both for T and B cells (CD3, CD43, CD20, CD79a). The adjacent cerebellar folia were atrophic, with total loss of granular cell neurons, preservation of Purkinje cells, and infiltrates of lymphocytes and histiocytes. Four 10-␮m-thick sections of the mass were cut and utilized for DNA extraction; PCR was performed and demonstrated a 173 base pair band diagnostic of polyoma virus; its identity as JCV was

further confirmed with a BamHI digest which produces 2 DNA fragments of 120 and 53 base pairs (JCV, but not BKV or SV40 has this restriction site in the amplicon)1 (figure, C). A JCV-associated inflammatory pseudotumor was diagnosed. The patient has a stable pancerebellar syndrome 24 months after onset of neurologic symptoms. Discussion. The immune reconstitution inflammatory syndrome (IRIS) in HIV-infected patients receiving HAART is characterized by paradoxical clinical or radiologic deterioration despite an increasing CD4⫹ T cell count and decreasing HIV viral load.2 Foreign organisms become unmasked and trigger a disproportionate immune response.2 IRIS has been reported in association with JCV infection, the cause of progressive multifocal leukoencephalopathy (PML).3 The radiologic features of PML are generally characterized by focal high-signal lesions predominantly affecting white matter structures; in the cerebellum, it has been associated with atrophy of the folia, with or without white matter involvement. While PML generally shows minimal contrast enhancement, this is more frequent in IRIS-associated PML. The radiologic features in the current patient, demonstrating a cystic lesion with nodular enhancement were unusual, raised the possibility of a secondary neoplastic or infectious non–JCV-related lesion, and led to the eventual performance of a brain biopsy. PML is histologically characterized by the triad of oligodendroglial inclusions, demyelination, and bizarre, atypical astrocytes.4 In the cerebellum, selective loss of granular cell neurons, as seen in the present case, is common. PML may be associated with variable host inflammatory response. In the case of IRISassociated PML, there are appreciable inflammatory infiltrates, with a preponderance of T cells.4,5 Oligodendroglial nuclei with characteristic viral inclusions may be rare or absent, and bizarre pleomorphic or multinucleated cells may have astrocytic or histiocytic origins.6 Pathology in the current patient was unusual, as the combination of dense inflammation and bizarre glial cells resulted in a pseudotumor formation, heretofore unreported in JCV-associated IRIS. In our patient, oligodendroglial inclusions were not evident, although the abnormal morphology of giant cells was typical of JCV-transformed astrocytes, and the granular cell loss was characteristic of PML. The juxtaposition of the intense predominantly lymphoplasmacytic infiltrate surrounding these transformed cells is likely to represent an IRISinduced inflammatory response.7 This JCV-associated pseudotumor is an unusual manifestation of the spectrum of IRIS neuropathologies. Given its clinical and radiologic overlap with Neurology 72

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Figure

MRI and histologic examination of the cerebellum

other tumoral and infectious entities, clinicians must be alert to the differential diagnosis. From the Departments of Neurology, NeuroAIDS Program (A.G.-D., S.S., G.J.S., D.S.), Radiology (T.N.), Pathology (G.K., J.M., S.M.), Neurosurgery (I.G.), and Infectology (M.M.), Mount Sinai Medical Center, New York, NY. Supported in part by NIH grant R24MH59724. Disclosure: The authors report no disclosures. Received December 4, 2007. Accepted in final form August 14, 2008. Address correspondence and reprint requests to Dr. Alejandra Gonzalez-Duarte, Mount Sinai Medical Center, Annenberg 2nd Floor, Box 1052, New York, NY 10029; [email protected] Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

3.

4. 5.

6.

7.

(A) Gadolinium-enhanced T1 FLAIR MR image demonstrating a large cystic ring-enhancing lesion in the context of high signal intensity lesions of the cerebellum. (B) Excisional biopsy of the cerebellum demonstrating bizarre multinucleated giant cells surrounded by an inflammatory cell infiltrate (hematoxylin-eosin, original magnification 100⫻). (C) Ethidiumbromide stained gel demonstrating presence of JCV in the patient’s pseudotumor. Formalin-fixed, paraffin-embedded sections of the patient’s lesion (lanes 4 and 7) and autopsy-derived progressive multifocal leukoencephalopathy (PML) (lanes 5 and 8) and normal brain (lanes 3 and 6) were used to extract DNA for PCR. Lanes 2, 3, 4, and 5 show the results of PCR amplification of a 173 base pair segment of polyoma virus in a reaction run without template DNA (lane 2), with normal brain DNA (lane 3), DNA from the patient’s lesion (lane 4), and from an unrelated case of PML (lane 5). Lanes 6, 7, and 8 display BamHI digests of the PCR products run in lanes 3, 4, and 5, respectively. Both the patient’s lesion and the case of PML show specific, 120 and 53 base pair fragments that occur only with JCV, which has a BamHI restriction site in the amplicon (SV40 and BKV do not share this restriction site). Lane 1 contains a 100 bp DNA ladder.

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Arthur RR, Dagostin S, Shah K. Detection of BK virus and JC virus in urine and brain tissue by the polymerase chain reaction. J Clin Microbiol 1989;27:1174–1179. Murdoch DM, Venter WD, Van Rie A, Feldman C. Immune reconstitution inflammatory syndrome (IRIS): review of common infectious manifestations and treatment options. Aids Res Ther 2007;8:9. Roberts MT. AIDS-associated progressive multifocal leukoencephalopathy: current management strategies. CNS Drugs 2005;19:671–682. Brooks BR, Walker DL. Progressive multifocal leukoencephalopathy. Neurol Clin 1984;2:299–313. Safdar A, Rubocki RJ, Horvath JA, Narayan KK, Waldron RL. Fatal immune restoration disease in human immunodeficiency virus type 1-infected patients with progressive multifocal leukoencephalopathy: impact of antiretroviral therapy-associated immune reconstitution. Clin Infect Dis 2002;35:1250–1257. Gray F, Bazille C, Adle-Biassette H, Mikol J, Moulignier A, Scaravilli F. Central nervous system immune reconstitution disease in acquired immunodeficiency syndrome patients receiving highly active antiretroviral treatment. J Neurovirol 2005;11 (suppl 3):16–22. Martinez JV, Mazziotti JV, Efron ED, et al. Immune reconstitution inflammatory syndrome associated with PML in AIDS: a treatable disorder. Neurology 2006;67:1692– 1694.

Clinical/Scientific Notes

M.A. van Es, MD F.P. Diekstra, MD J.H. Veldink, MD, PhD F. Baas, PhD P.R. Bourque, MD H.J. Schelhaas, MD, PhD E. Strengman E.A.M. Hennekam, PhD D. Lindhout, MD, PhD R.A. Ophoff, PhD L.H. van den Berg, MD, PhD

Supplemental data at www.neurology.org

A CASE OF ALS-FTD IN A LARGE FALS PEDIGREE WITH A K17I ANG MUTATION

Methods. A total of 39 unrelated FALS patients, negative for SOD1 mutations, were screened for ANG mutations. This study was approved by the local ethics committee and participants provided informed consent. DNA was isolated from venous blood and ANG mutation analysis was performed as described in appendix e-1. A total of 275 unrelated, healthy controls were taken from a prospective population-based study on ALS in The Netherlands and were also screened. 5 PMut (http://mmb2.pcb.ub.es:8080/PMut/) was used to predict the impact of an amino acid substitution on the structure and function of the protein.

Cases III-3, III-4, and IV-1 all presented with progressive upper and lower motor neuron loss of limbs. Case III-1 rapidly developed weakness in both arms with atrophy, fasciculations, and dyspnea, but no upper motor neuron signs. The patient died after 6 months from onset. Case III-2 initially presented with parkinsonism (bradykinesia, diminished postural reflexes, cogwheel rigidity [right arm], shuffling, short-stepped gait, and decreased spontaneous eye blink rate). There was no autonomic dysfunction and eye movements were intact. Dopaminergic treatment had little effect. After 5 years, the patient developed progressive weakness of the arms and legs with atrophy, fasciculations, and hyperreflexia. Interestingly, the patient also demonstrated symptoms characteristic of frontotemporal dementia (FTD), such as loss of interest in social contacts and family, short attention span, logopenia, verbal apraxia, perseveration, decreased personal hygiene, hyperorality, reckless behavior in traffic, sexual disinhibition, and apathy. Case I-2 and II-4 also appear to have been affected. However, no medical records were available. Patient I-2 developed limb weakness at age 70, leading to paralysis and death within 3 years. Patient II-4 developed speech impairment at age 60 and also died within 3 years. Patient II-2 (obligate carrier) died at age 50 from cardiovascular disease. Detailed clinical characteristics are provided in table e-2.

Results. We identified one mutation in one patient (122 A⬎T) (figure, A), leading to an amino acid substitution of lysine to isoleucine (K17I) (figure, B). PMut analysis predicted this mutation to be pathogenic. Sequence alignments of ANG in different species demonstrated high conservation (figure, C). Analysis of this pedigree revealed an autosomal dominant inheritance of the mutation (male to male transmission) (figure, D). DNA was available from 44 out of 62 family members (five affected individuals). All affected family members carried the K17I mutation. Ten carriers were identified, but all were under 50 years of age (except one who was 75 years old without symptoms or signs of ALS). The K17I mutation was not found in 275 control samples.

Discussion. Several ANG mutations in FALS have been reported, but clear segregation of mutations with the disease has not been shown. Here, we report the K17I mutation segregating with disease in a large pedigree. The fact that II-2 and a carrier (75 years of age) were without symptoms of ALS suggests incomplete penetrance of the mutation. This might explain why mutations in this codon have only been found in SALS. The K17I mutation was previously reported in three cases and K17E in two cases.3,6 This study provides a report of a patient with an ANG mutation and ALS, FTD, and parkinsonism. Five percent of patients with ALS also have FTD and up to 50% demonstrated mild cognitive impairment. Similarly, relatives of patients with ALS have an increased risk for developing PD. Therefore, genes involved in ALS are also considered candidate genes for

Approximately 90% of amyotrophic lateral sclerosis (ALS) cases are sporadic (SALS), but 10% are familial (FALS). Mutations in SOD1, Alsin, Dynactin, SETX, DJ-1, VAPB, and TDP-431 have been reported (table e-1 on the Neurology® Web site at www.neurology.org). After the identification of sequence variation VEGF in patients with ALS, mutations in another angiogenic gene (ANG) were identified in SALS and FALS.2,3 Studies in other populations have identified ANG mutations in patients with ALS, but also in healthy controls. This suggests that not all mutations are pathogenic.3,4

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Figure

Mutation analysis and partial pedigree

ANG is highly conserved between species, suggesting it has an important biologic function. Modeling of the K17I mutation using PMut predicted this to be pathogenic. Two functional studies demonstrated that the K17I mutation results in loss of function, possibly leading to insufficient ribosomes synthesis, decreased protein translation, and ultimately decreased motor neuron viability.6,7 We report segregation of the K17I mutation with FALS and a patient with FALS, FTD, and parkinsonism, which possibly implicates ANG in these diseases. From the Department of Neurology, Rudolf Magnus Institute of Neuroscience (M.A.v.E., F.P.D., J.H.V., L.H.v.d.B.), Department of Medical Genetics and Rudolf Magnus Institute of Neuroscience (E.S., R.A.O.), and Department of Medical Genetics (E.A.M.H., D.L.), University Medical Center Utrecht; Department of Neurogenetics (F.B.), Academic Medical Center, Amsterdam, The Netherlands; Division of Neurology (P.R.B.), University of Ottawa, Ontario, Canada; and Department of Neurology (H.J.S.), Radboud University Medical Center, Nijmegen, The Netherlands. Supported by Netherlands Organization for Scientific Research (NWO) and the Prinses Beatrix Foundation (PBF). Disclosure: The authors report no disclosures. Received June 12, 2008. Accepted in final form August 12, 2008. Address correspondence and reprint requests to Dr. Leonard H. van den Berg, Department of Neurology, Rudolf Magnus Institute of Neuroscience, University Medical Center Utrecht, Heidelbergaan 100, 3584 CX, Utrecht, The Netherlands; L.H.vandenBerg@ umcutrecht.nl Copyright © 2009 by AAN Enterprises, Inc.

ACKNOWLEDGMENT The authors thank the families for their participation.

1. 2.

3.

(A) Wild type sequence and the K17I mutation. (B) Three-dimensional structure of ANG modeling the K17I mutation in ball-and-stick representation. The figure was created using the program PyMOL (DeLano Scientific). (C) Sequence alignments of ANG in different species. Sequences were multiply aligned using Homologene (http://www.ncbi.nlm.nih.gov/sites/entrez/ query.fcgi?db ⫽ homologene). The numbering on top of the alignments correlates with the human amino acid sequence. Amino acid 17, which is the site of K17I, is indicated in red. (D) A simplified version of the pedigree is depicted to protect the privacy of the family. A partial four-generation pedigree is shown. All individuals marked in black have amyotrophic lateral sclerosis. All individuals carrying the122 A⬎T (K17I) mutation are marked in the pedigree. The obligate carrier (II-2) died at 50 years of age due to cardiovascular disease. No DNA was available from this individual. The spouse of II-2 tested negative for the mutation.

other neurodegenerative disorders. Indeed, an Italian study reported a SALS patient with a 132C¡T mutation and frontal lobe dysfunction.4

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4.

5.

6.

7.

Valdmanis PN, Rouleau GA. Genetics of familial amyotrophic lateral sclerosis. Neurology 2008;70:144–152. Greenway MJ, Alexander MD, Ennis S, et al. A novel candidate region for ALS on chromosome 14q11.2. Neurology 2004;63:1936–1938. Greenway MJ, Andersen PM, Russ C, et al. ANG mutations segregate with familial and ‘sporadic’ amyotrophic lateral sclerosis. Nat Genet 2006;38:411–413. Gellera C, Colombrita C, Ticozzi N, et al. Identification of new ANG gene mutations in a large cohort of Italian patients with amyotrophic lateral sclerosis. Neurogenetics 2008;9:33–40. van Es MA, Van Vught PW, Blauw HM, et al. ITPR2 as a susceptibility gene in sporadic amyotrophic lateral sclerosis: a genome-wide association study. Lancet Neurol 2007; 6:869–877. Wu D, Yu W, Kishikawa H, et al. Angiogenin loss-offunction mutations in amyotrophic lateral sclerosis. Ann Neurol 2007;62:609–617. Crabtree B, Thiyagarajan N, Prior SH, et al. Characterization of human angiogenin variants implicated in amyotrophic lateral sclerosis. Biochemistry 2007; 46:11810–11818.

A. Gonzalez-Duarte, MD S. Sullivan G.J. Sips T. Naidich, MD G. Kleinman, MD J. Murray S. Morgello, MD I. Germano, MD M. Mullen, MD D. Simpson, MD

Supplemental data at www.neurology.org

INFLAMMATORY PSEUDOTUMOR ASSOCIATED WITH HIV, JCV, AND IMMUNE RECONSTITUTION SYNDROME

A 37-year-old HIV-positive African woman developed severe chronic diarrhea. Her CD4⫹ T cell count was 25 cells/mm3. Within 1 month of initiation of highly active antiretroviral therapy (HAART) her plasma HIV viral load became undetectable and CD4⫹ T cell count rose to 96 cells/mm3, and continued to rise over the following months. Two months after the initiation of HAART she developed vertigo, loss of balance, incoordination, slurred speech, and tremor of the neck and limbs. Neurological examination revealed ocular abnormalities, dysarthria, and monotonic speech. She had bilateral limb dysmetria, past-pointing and endpoint tremor, impaired heel-knee-shin testing, head tremor, and a wide based, ataxic gait. Initial brain MRI revealed a confluent, nonenhancing area of signal abnormality predominantly involving the inferior right cerebellar hemisphere and extending to the posterior vermis, right cerebellar peduncle, and inferomedial aspect of the left cerebellar hemisphere. Six months later, MRI revealed progression of the cerebellar lesion, with nodular enhancement along the inferomedial aspect of the right cerebellar hemisphere. The patient remained clinically stable. MRI 8 months later revealed a large cystic ring-enhancing lesion in the location of the previously noted high signal intensity lesions of the cerebellum, with compression of the posterior fourth ventricle (figure, A). CSF revealed WBC 1, Prot 64, Gluc 49, and negative cytomegalovirus DNA PCR, Cysticercosis IgG Ab, Epstein-Barr virus DNA PCR, Venereal Disease Research Laboratory, and Cryptococcus Ag. Bacterial, viral, and fungal cultures were negative. JCV PCR was positive. Stereotactic biopsy of the cerebellar lesion, performed 17 months after the onset of neurologic symptoms, revealed giant cells with pleomorphic hyperchromatic nuclei, often multiple, surrounded by a dense infiltrate of lymphocytes and plasma cells (figure, B). The bizarre, pleomorphic cells were GFAP positive, demonstrated diffuse nuclear reactivity for p53 antigen, a high MIB-1 (Ki67) index, and focal, faint reactivity for polyoma virus T antigen (figure e-1 on the Neurology® Web site at www.neurology.org). Inflammatory infiltrates marked both for T and B cells (CD3, CD43, CD20, CD79a). The adjacent cerebellar folia were atrophic, with total loss of granular cell neurons, preservation of Purkinje cells, and infiltrates of lymphocytes and histiocytes. Four 10-␮m-thick sections of the mass were cut and utilized for DNA extraction; PCR was performed and demonstrated a 173 base pair band diagnostic of polyoma virus; its identity as JCV was

further confirmed with a BamHI digest which produces 2 DNA fragments of 120 and 53 base pairs (JCV, but not BKV or SV40 has this restriction site in the amplicon)1 (figure, C). A JCV-associated inflammatory pseudotumor was diagnosed. The patient has a stable pancerebellar syndrome 24 months after onset of neurologic symptoms. Discussion. The immune reconstitution inflammatory syndrome (IRIS) in HIV-infected patients receiving HAART is characterized by paradoxical clinical or radiologic deterioration despite an increasing CD4⫹ T cell count and decreasing HIV viral load.2 Foreign organisms become unmasked and trigger a disproportionate immune response.2 IRIS has been reported in association with JCV infection, the cause of progressive multifocal leukoencephalopathy (PML).3 The radiologic features of PML are generally characterized by focal high-signal lesions predominantly affecting white matter structures; in the cerebellum, it has been associated with atrophy of the folia, with or without white matter involvement. While PML generally shows minimal contrast enhancement, this is more frequent in IRIS-associated PML. The radiologic features in the current patient, demonstrating a cystic lesion with nodular enhancement were unusual, raised the possibility of a secondary neoplastic or infectious non–JCV-related lesion, and led to the eventual performance of a brain biopsy. PML is histologically characterized by the triad of oligodendroglial inclusions, demyelination, and bizarre, atypical astrocytes.4 In the cerebellum, selective loss of granular cell neurons, as seen in the present case, is common. PML may be associated with variable host inflammatory response. In the case of IRISassociated PML, there are appreciable inflammatory infiltrates, with a preponderance of T cells.4,5 Oligodendroglial nuclei with characteristic viral inclusions may be rare or absent, and bizarre pleomorphic or multinucleated cells may have astrocytic or histiocytic origins.6 Pathology in the current patient was unusual, as the combination of dense inflammation and bizarre glial cells resulted in a pseudotumor formation, heretofore unreported in JCV-associated IRIS. In our patient, oligodendroglial inclusions were not evident, although the abnormal morphology of giant cells was typical of JCV-transformed astrocytes, and the granular cell loss was characteristic of PML. The juxtaposition of the intense predominantly lymphoplasmacytic infiltrate surrounding these transformed cells is likely to represent an IRISinduced inflammatory response.7 This JCV-associated pseudotumor is an unusual manifestation of the spectrum of IRIS neuropathologies. Given its clinical and radiologic overlap with Neurology 72

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289

Figure

MRI and histologic examination of the cerebellum

other tumoral and infectious entities, clinicians must be alert to the differential diagnosis. From the Departments of Neurology, NeuroAIDS Program (A.G.-D., S.S., G.J.S., D.S.), Radiology (T.N.), Pathology (G.K., J.M., S.M.), Neurosurgery (I.G.), and Infectology (M.M.), Mount Sinai Medical Center, New York, NY. Supported in part by NIH grant R24MH59724. Disclosure: The authors report no disclosures. Received December 4, 2007. Accepted in final form August 14, 2008. Address correspondence and reprint requests to Dr. Alejandra Gonzalez-Duarte, Mount Sinai Medical Center, Annenberg 2nd Floor, Box 1052, New York, NY 10029; [email protected] Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

3.

4. 5.

6.

7.

(A) Gadolinium-enhanced T1 FLAIR MR image demonstrating a large cystic ring-enhancing lesion in the context of high signal intensity lesions of the cerebellum. (B) Excisional biopsy of the cerebellum demonstrating bizarre multinucleated giant cells surrounded by an inflammatory cell infiltrate (hematoxylin-eosin, original magnification 100⫻). (C) Ethidiumbromide stained gel demonstrating presence of JCV in the patient’s pseudotumor. Formalin-fixed, paraffin-embedded sections of the patient’s lesion (lanes 4 and 7) and autopsy-derived progressive multifocal leukoencephalopathy (PML) (lanes 5 and 8) and normal brain (lanes 3 and 6) were used to extract DNA for PCR. Lanes 2, 3, 4, and 5 show the results of PCR amplification of a 173 base pair segment of polyoma virus in a reaction run without template DNA (lane 2), with normal brain DNA (lane 3), DNA from the patient’s lesion (lane 4), and from an unrelated case of PML (lane 5). Lanes 6, 7, and 8 display BamHI digests of the PCR products run in lanes 3, 4, and 5, respectively. Both the patient’s lesion and the case of PML show specific, 120 and 53 base pair fragments that occur only with JCV, which has a BamHI restriction site in the amplicon (SV40 and BKV do not share this restriction site). Lane 1 contains a 100 bp DNA ladder.

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Arthur RR, Dagostin S, Shah K. Detection of BK virus and JC virus in urine and brain tissue by the polymerase chain reaction. J Clin Microbiol 1989;27:1174–1179. Murdoch DM, Venter WD, Van Rie A, Feldman C. Immune reconstitution inflammatory syndrome (IRIS): review of common infectious manifestations and treatment options. Aids Res Ther 2007;8:9. Roberts MT. AIDS-associated progressive multifocal leukoencephalopathy: current management strategies. CNS Drugs 2005;19:671–682. Brooks BR, Walker DL. Progressive multifocal leukoencephalopathy. Neurol Clin 1984;2:299–313. Safdar A, Rubocki RJ, Horvath JA, Narayan KK, Waldron RL. Fatal immune restoration disease in human immunodeficiency virus type 1-infected patients with progressive multifocal leukoencephalopathy: impact of antiretroviral therapy-associated immune reconstitution. Clin Infect Dis 2002;35:1250–1257. Gray F, Bazille C, Adle-Biassette H, Mikol J, Moulignier A, Scaravilli F. Central nervous system immune reconstitution disease in acquired immunodeficiency syndrome patients receiving highly active antiretroviral treatment. J Neurovirol 2005;11 (suppl 3):16–22. Martinez JV, Mazziotti JV, Efron ED, et al. Immune reconstitution inflammatory syndrome associated with PML in AIDS: a treatable disorder. Neurology 2006;67:1692– 1694.

VIDEO NEUROIMAGES

Ocular flutter as the first manifestation of Lyme disease

A 33-year-old man with a history of tick bites presented with bursts of involuntary horizontal conjugate saccades, myoclonic head jerks, and truncal ataxia. A cerebral MRI was normal, and no antineuronal antibodies were found (anti-Hu, anti-Yo, anti-Ri). Despite negative serum antibodies for Borrelia burgdorferi, acute neuroborreliosis was suspected because of lymphocytic mild meningitis (19 white cells/mm3, protein 0.79 g/L) and apparent intrathecal synthesis of B burgdorferi IgM antibodies (ELISA titers 6.17, normal ⬍0.3), although false-positive IgM serologies can occur in this setting. Intravenous ceftriaxone treatment resulted in dramatic clinical improvement within a few weeks. In ocular flutter, saccadic intrusions are purely horizontal (see video), while in opsoclonus-myoclonus, a similar condition, they are multidirectional.1 Cerebral MRI studies are usually normal,2 lesions involving omnipause neurons in the pons, or the fastigial nucleus in the cerebellum being exceptional. Jesper Gyllenborg, MD, Dan Milea, MD, PhD, Copenhagen, Denmark Supplemental data at www.neurology.org

Disclosure: The authors report no disclosures. Address correspondence and reprint requests to Dr. Jesper Gyllenborg, Department of Neurology, Glostrup Hospital, University of Copenhagen, 57, Nordre Ringvej, DK-2600 Glostrup, Denmark; [email protected] 1. 2.

Wong AM, Musallam S, Tomlinson RD, Shannon P, Sharpe JA. Opsoclonus in three dimensions: oculographic, neuropathologic and modelling correlates. J Neurol Sci 2001;15:71–81. Peter L, Jung J, Tilikete C, Ryvlin P, Mauguiere F. Opsoclonus-myoclonus as a manifestation of Lyme disease. J Neurol Neurosurg Psychiatry 2006;77:1090–1091.

Copyright © 2009 by AAN Enterprises, Inc.

291

RESIDENT & FELLOW SECTION Section Editor Mitchell S.V. Elkind, MD, MS

Clinical Reasoning: A case of Wegener granulomatosis complicated by seizures and headaches Curiouser and curiouser

G. Gorman, MRCP M. Hutchinson, MD N. Tubridy, MD Address correspondence and reprint requests to Dr. Niall Tubridy, Department of Neurology, St. Vincent’s University Hospital, Elm Park, Dublin 4, Ireland [email protected]

SECTION 1

A 19-year-old man presented with a 5-day history of vomiting and passing dark urine. He gave a 6-month history of arthralgias and joint swelling, particularly affecting larger joints. He had recurrent epistaxis and recent episcleritis. He was noted to be thin and pale, with conjunctival suffusion and deformity of his nasal bridge. He was normotensive (115/60 mm Hg) and apyrexic (35.5°C). Blood parameters revealed normocytic normochromic anemia and acute renal failure. Renal biopsy showed extensive crescent formation with segmental necrotizing inflammation and fibrin deposition of the glomeruli. cANCA was strongly positive (titer 1280; ELISA PR3⫹) and he was diagnosed with Wegener granulomatosis. He commenced hemodialysis, steroids, and oral cyclophosphamide, and he was discharged home.

Figure

One month later, he presented with acute onset of throbbing headaches associated with nausea and photophobia, as well as recurrent generalized seizures necessitating sedation and intensive care unit admission. He was pyrexic (38°C) and hypertensive (165/95 mm Hg). Renal indices were abnormal (urea 29.1mmol/L; creatinine 755 ␮mol/L; potassium 5.4 mmol/L). Clinical examination revealed no focal neurologic deficits. Initial noncontrast brain CT was normal. Brain MRI with gadolinium showed small, ill-defined, nonenhancing high signal areas measuring 5–20 mm in several locations (figure, A). Questions for consideration: 1. What are the possible etiologies of the changes on brain MRI? 2. What additional diagnostic testing would you consider at this point?

Brain MRIs

(A) Axial fluid-attenuated inversion recovery (FLAIR) brain MRI showing small, ill-defined, nonenhancing high signal areas measuring 5–20 mm in several locations. (B) Axial FLAIR brain MRI showing progression in radiologic changes over a further 1-month interval. Bright confluent lesions are seen in both parieto-occipital regions. (C) Axial FLAIR brain MRI showing marked progression in radiologic changes over a further 2-month interval. Bright confluent areas are seen extensively involving the occipital and parietal lobes, and including the frontal lobes. (D) Axial FLAIR brain MRI showing full resolution of radiologic changes 6 months after commencement of IV cyclophosphamide.

GO TO SECTION 2

From the Department of Neurology, St Vincent’s University Hospital, Elm Park, Dublin, Ireland. Disclosure: The authors report no disclosures.

Copyright © 2009 by AAN Enterprises, Inc.

e11

SECTION 2

Full septic screen including midstream urine, chest x-ray, echocardiogram, and blood cultures were negative for active infection. MR angiography and formal four-vessel angiography were normal and revealed no beading. Postictal EEG was normal. A lumbar puncture was declined by the patient. He was commenced on oral phenytoin with titration of his immunosuppression and discharged home. He was readmitted 1 month later with further seizures and neutropenic sepsis. He was normotensive (120/60 mm Hg) and pyrexic (38.6°C). Neurologic examination was within normal limits. Focal areas of abnormal high signal were again noted on brain MRI and were identical to those previously reported (figure, A). ANCA levels were low (1:80). Cyclophosphamide was discontinued and mycophenolate was commenced. He was again admitted 1 month later with pyrexia (38.5°C) and diarrhea. He described brief episodes of visual blurring and central loss of vision. No evidence of central scotoma was detected clinically. Brain MRI now showed more extensive areas of high signal in both parieto-occipital regions (figure, B). CSF analysis showed normal protein and glucose parameters and was

acellular. Testing for JC Virus was negative. He commenced antimicrobial therapy and his immunosuppressive therapy was increased. He presented again 2 months later with recurrent partial seizures and severe throbbing headaches characteristic of migraine with aura. He described episodes of body image distortion (macrosomatognosia) with his hand “ballooning,” colored visual spectra phenomena characterized as a “kaleidoscope” effect or a light moving diagonally across his visual field from left to right, and an impaired sense of passage of time (a perceived increased speech velocity). These episodes heralded the onset of headache, seizures, or a combination of both. He had intractable hiccups and episodes of vertigo, ataxia, slurred speech, and nystagmus. He was normotensive (100/60 mm Hg), pyrexic (38.6°C), and neutropenic. Serum antiepileptic drug levels were within normal limits and cANCA was negative. Questions for consideration: 1. What is the term coined for the symptoms he is describing? 2. What is the postulated anatomic basis of his symptoms?

GO TO SECTION 3

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SECTION 3

The clinical picture at this stage was thought consistent with the Alice in Wonderland syndrome (AIW). The features of AIW were first reported by Coleman in 1932,1 but it was Todd2 who coined the term in 1955 to describe the phenomena of distorted space, time, and body image. Lewis Carroll made this phenomenon famous in “Alice’s Adventures in Wonderland” (1865) and it has been suggested that Alice’s transformations were based on Carroll’s own migrainous symptomatology; however, this remains controversial.3,4 AIW has been associated with migraine, epilepsy, drug intoxication, cerebral mass lesions (particularly involving the occipital and temporoparietal lobes), psychiatric disease, viral infections, pyrexia, and vasculitis.5,6 Perceptual errors of body schema and objects (kinesthetic illusions or metamorphopsia), of people appearing smaller (micropsia) or bigger (macropsia) than normal, visual hallucinations (Lilliputian hallucinations), and acceleration or deceleration of the passage of time have been attributed to AIW. It may also be associated with vertigo.7 The etiology and anatomic basis are

not fully understood. Body image distortion, vertigo, and metamorphopsia have been described from posterior parietal stimulation.8 Positive visual phenomena of flashing lights and colored visual spectra equated to a kaleidoscope and negative visual phenomena (scotoma) are both established occipital lobe phenomena. In addition, headache and vomiting may have an occipital origin.9 These symptoms prompted more brain imaging. Brain MRI showed diffuse white matter high signal abnormalities demonstrating dramatic interval progression involving the occipital, parietal, and temporal lobes in addition to bilateral cerebellar lesions with no enhancement with gadolinium (figure, C). The differential diagnosis of the underlying pathologic etiology was reversible leukoencephalopathy syndrome (RPLS) vs active cerebral vasculitis. Questions for consideration: 1. How would you manage a patient with Wegener granulomatosis, cerebral complications, and recurrent neutropenic sepsis? 2. How would you monitor response to treatment?

GO TO SECTION 4

Neurology 72

January 20, 2009

e13

SECTION 4

He was commenced on a modified dose of IV cyclophosphamide with close monitoring of bone marrow suppression. Sepsis and fluctuations in blood pressure were aggressively treated. Seizure control required quadruple antiepileptic drug therapy. Response to treatment was monitored by clinical status and serial C-reactive protein and c-ANCA measurements. Six months later he is well with full resolution of the radiologic changes (figure, D), seizures, and headaches. He remains dependent on dialysis. DISCUSSION We present a patient with cANCA-positive renal failure associated with progressive but reversible symmetric FLAIR-bright brain MRI lesions who developed features of AIW syndrome. The differential diagnosis initially included an infectious etiology, progressive multifocal leukoencephalopathy, Neuro-Behc¸et disease, acute disseminated encephalomyelitis, multiple sclerosis, cerebral vasculitis, and RPLS. CSF analysis showed no abnormalities suggesting inflammation, infection, or malignancy. MR and catheterization angiography revealed no evidence of vasculitis, although this would not completely exclude CNS vasculitis as Wegener disease typically affects small and medium vessels, which are not formally assessed by these methods. Serial brain MRI with gadolinium revealed no enhancement, however, also favoring a diagnosis of prolonged RPLS precipitated by renal failure, sepsis, and possible transient changes in blood pressure, not captured clinically. RPLS is caused by subcortical white matter edema predominantly involving the posterior regions of the brain, but gray matter and other regions including the brainstem and cerebellum may be involved. Although this is usually recognized as a single event,

e14

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recurrent episodes have been reported.10 The duration and recurrence of symptoms over a 4-month period despite changes in immunosuppression is atypical, and, despite instigation of these measures, there was clinical and radiologic progression. Aggressive immunosuppression was recommenced, albeit at modified doses, resulting in full resolution of symptoms and radiologic changes. We propose that the amended dosage of cyclophosphamide immunotherapy induced disease remission salvaging residual intrinsic renal function, reducing subclinical systemic blood pressure alterations, while reducing the risk of sepsis and direct chemotoxic effects of this therapy and thus improving cerebral autoregulation. REFERENCES 1. Coleman SM. Misidentification and non recognition. J Ment Sc 1933;79:42–51. 2. Todd J. The syndrome of Alice in Wonderland. Canad Med Assoc J 1955;73:701–704. 3. Blau JN. Somesthetic aura: the experience of “Alice in Wonderland.” Lancet 1998;352:582. 4. Podoll K, Robinson D. Lewis Carroll’s migraine experiences. Lancet 1999;353:1366. 5. Takaoka K, Takata T. “Alice in Wonderland” syndrome and Lilliputian hallucinations in a patient with a substance related disorder. Psychopathology 1999;32:47–49. 6. Hausler M, Raemaekers VT, Doenges M, et al. Neurological complications of acute and persistent Epstein-Barr virus infection in paediatric patients. J Med Virol 2002;68: 253–263. 7. Evans RW, Rolak LA. The Alice in Wonderland syndrome. Headache 2004;44:624–625. 8. Podoll K, Robinson D. Out-of-body experiences and related phenomena in migraine art. Cephalalgia 1999;19: 886–896. 9. Sveinbjornsdottir S, Duncan JS. Parietal and occipital lobe epilepsy: a review. Epilepsia 1993;34:493–521. 10. Lee V, Wijdicks E, Manno E, et al. Clinical spectrum of reversible posterior leukoencephalopathy syndrome. Arch Neurol 2008;65:205–210.

Correspondence

TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHY IN THE 21st CENTURY: NEUROSCIENCE FOR THE CLINICAL NEUROLOGIST

To the Editor: I read Brown’s article1 on transmissible spongiform encephalopathy (TSE) with interest. Other disease mechanisms apart from seeding by misfolded prion are under consideration in TSE. “Slow virus” infections might provide an explanation for the unusual characteristics of TSE agents, such as persistence for years and spread of the bovine spongiform encephalopathy agent to divergent species.2 Spiroplasma bacteria have been considered to be related to TSE, since they have been recovered from brain tissues of sheep with scrapie, cervids with chronic wasting disease, and patients with Creutzfeldt-Jakob disease by passage through embryonated eggs.3 Spiroplasma bacteria resist high temperatures, radiation, and fixatives.3 Prion protein may be induced on the cell surface by bacterial infection.3 Scrapie associated fibrils commonly found in TSE also appear identical to bacterial Spiroplasma fibrils.3 When Spiroplasma mirum was inoculated intracranially (IC) into deer, sheep, and goats, it caused spongiform encephalopathy. It had already been shown to cause spongiform encephalopathy in rodents. Spiroplasma spp was isolated from scrapie-affected sheep and from chronic wasting disease–affected deer by passage through embryonated eggs. When inoculated IC into sheep and goats, it caused spongiform encephalopathy in the animals.4 There may be a vector or reservoir in some instances in the spread of TSE. Hay mite preparations from five Icelandic farms with a history of scrapie caused spongiform characteristic of scrapie when injected into mice, indicating probably some vector was involved.5 An infectious agent, possibly a slow virus or Spiroplasma bacteria, may be responsible for some cases of TSE. Steven R. Brenner, St. Louis, MO

Interestingly, the NIH Laboratory where I spent my career bore the name “Slow, Latent, and Temperate Viruses of the CNS,” and for quite some time it looked as though a “slow virus” might be at the root of the problem (similar to the JC virus that causes progressive multifocal leukoencephalopathy). However, as new observations were made,6 the concept of a slow virus became less and less tenable, at least as the sole cause of disease, which certainly can no longer be seriously considered. Regarding the Spiroplasma hypothesis, the failure of a blinded study of rRNA species to detect any footprint of Spiroplasma in scrapie-infected hamster brain7 laid it to rest, unless one holds that the cause of scrapie is different from that of other prion diseases, an equally untenable proposition. Paul W. Brown, Bethesda, MD Disclosure: The author reports no disclosures. Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

3.

4.

5.

6.

7.

Brown P. Transmissible spongiform encephalopathy in the 21st century: neuroscience for the clinical neurologist. Neurology 2008;70:713–722. Manuelidis L. A 25 nm virion is the likely cause of transmissible spongiform encephalopathies. J Cell Biochem 2007;100:897–915. Bastian F. Spiroplasma as a candidate agent for transmissible spongiform encephalopathies. J Neuropathol Exp Neurol 2005;64:833– 838. Bastian F, Sanders D, Forbes W, et al. Spiroplasma spp. from transmissible spongiform encephalopathy brains or ticks induce spongiform encephalopathy in ruminants. J Med Microbiol 2007;56:1235–1242. Carp TI, Meeker, HC, Rubenstein R. Characteristics of scrapie isolates derived from hay mites. J Neurovirol 2000;6: 137–142. Brown P. The “brave new world” of transmissible spongiform encephalopathy (infectious cerebral amyloidosis). Molec Neurobiol 1994;8:79 – 87. Alexeeva I, Elliott EJ, Rollins S, et al. Absence of Spiroplasma or other bacterial 16S rRNA genes in brain tissue of hamsters with scrapie. J Clin Microbiol 2006;44:91–97.

Disclosure: The author reports no disclosures. INTRANASAL INSULIN IMPROVES COGNITION

Reply from the Author: Dr. Brenner considers the possibility that something other than the prion protein might be the cause of prion diseases. I sympathize with his caution, but not his choice of alternatives. 292

Neurology 72

January 20, 2009

AND MODULATES ␤-AMYLOID IN EARLY AD

To the Editor: At our department’s recent journal club conference, we discussed the report by Reger et al.1 on intranasal insulin in early Alzheimer disease (AD). Given findings of epidemiologic and transla-

tional research associating diabetes and dementia, these results are an intriguing extension of their earlier work but raised several questions.2 First, the sample was somewhat heterogeneous— comprising patients with AD but also mild cognitive impairment with amnestic features (aMCI). Some baseline differences were evident in cognition and serologic markers between treatment and placebo. For example, memory savings scores for baseline placebo and treatment, plus 21-day follow-up placebo arms, appeared to be different, but would be expected to be rather similar. Could these differences be related to incomplete randomization, possibly related to small sample size? The proportion of AD and aMCI patients by treatment and placebo groups would be useful to know. Several patients were unable to perform the Stroop Color-Word task or had missing bloodwork. From which groups were these data missing? Logical memory percent retention and Stroop performance speed were the neuropsychological tests presented. In the group’s previous work, additional tests included the Buschke Selective Reminding Test (SRT), the Self-Ordered Pointing Task, and a visual search task.2 That report focused on changes in total delayed logical memory and SRT relative to placebo, with no differences identified on tests of attention or processing speed. If other tests were performed but not reported, it would be helpful if the authors could provide a more comprehensive neuropsychological summary. That report also found a significant interaction with APOE genotype. Was APOE genotyping done for this study? Most analyses compared group means rather than differences in individual performance over test sequence yet the latter might be more informative. Concerning the correlation of Dementia Severity Rating Scale (DSRS) changes with the baseline Mattis Dementia Rating Scale (DRS) (figure 3), did these data satisfy an assumption of homoscedasticity prior to fitting the correlation line? Lower DRS in both groups appeared to be associated with higher variance in DSRS. Also, given a seemingly large fluctuation in DSRS over 21 days within one placebo outlier, was exclusion of this outlier considered? Different DSRS scales between the two groups further complicated interpretation. We appreciate the difficulty of undertaking such a study and satisfying all readers. Perhaps this forum will give the authors an opportunity to present other data. We look forward to the results of their ongoing 120-day trial.3 Mandip S. Dhamoon, MD, MPH, James M. Noble, MD, New York, NY Disclosure: The authors report no disclosures.

Reply from the Authors: We appreciate the careful consideration of our article by Drs. Dhamoon and

Noble. Our sample included both patients with aMCI and early AD, diagnosed by expert consensus. When diagnosed in this manner, the majority of patients with aMCI have prodromal AD.4 The sample was heterogeneous with respect to severity but likely not type of impairment. Most importantly, the two groups contained comparable numbers of aMCI and AD patients (aMCI comprised 50% of the placebo group and 62% of the insulin-treated group), and the groups had comparable dementia severity as indicated in table 1 by the DRS. Similarly, there were no significant baseline differences between groups on any cognitive measure, nor were there differences in missing data (two placebo- and one insulin-treated participant unable to complete the Stroop, with an identical pattern observed for amyloid). We agree that randomization is difficult for small pilot trials such as ours, and were fortunate that our groups were comparable on cognitive, demographic, and metabolic variables. Our cognitive battery was tightly focused for this pilot effort. We used the Hopkins Verbal Learning Test rather than the SRT because it is more easily administered. One additional test was included (SelfOrdered Pointing Test), but computer malfunction resulted in uninterpretable data for the majority of participants. We used a repeated measures analysis of covariance analytic approach, which is preferable with small samples, given the variability of difference scores for cognitive measures. Our analysis for the DSRS revealed an interaction with dementia severity as indexed by the Mattis DRS, which was then deconstructed by correlating Mattis scores and the change in Dementia Severity Rating over the 21-day period. We reported Pearson correlations, which as noted may be vulnerable to violation of homoscedasticity or outlier effects, particularly in small samples. However, the correlation pattern remains identical and statistically significant when Spearman rank order correlations are used which protect against the effects of such violations. Finally, the issue of APOE genotype raises an important point. In two previous studies of the acute effects of intranasal insulin, we observed an APOErelated pattern, such that participants without the APOE-e4 allele benefited from insulin, whereas e4 carriers showed no benefit.2,5 This APOE-related difference has been observed with other manipulations of insulin.6,7 The pattern may support a specialized role for insulin abnormalities in the pathophysiology of AD for non-e4 carriers, who comprise about half of patients with AD, but for whom no clear risk factor has been identified. In our pilot study, the insulin-treated group was evenly divided with respect to APOE-e4 carriage Neurology 72

January 20, 2009

293

(e4 – ⫽ 7, e4⫹ ⫽ 6). These cell sizes are too small to allow exploration of a differential APOE treatment response, but our ongoing larger clinical trial is powered to determine such an effect.3 We hope to soon have data to address this important question.

3.

4.

Suzanne Craft, PhD, Seattle, WA Disclosure on article to which this Correspondence refers: W.D. is a founding member of Kurve Technology, the maker and patent holder of the electronic atomizer used in this study. W.D. holds equity interest in excess of $10,000 in Kurve Technology. The remaining authors have nothing to disclose.

5.

6.

Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

Reger MA, Watson GS, Green PS, et al. Intranasal insulin improves cognition and modulates beta-amyloid in early AD. Neurology 2008;70:440 – 448. Reger MA, Watson GS, Frey WH 2nd, et al. Effects of intranasal insulin on cognition in memory-impaired older

7.

adults: modulation by APOE genotype. Neurobiol Aging 2006;27:451– 458. SNIFF 120: Study of Nasal Insulin to Fight Forgetfulness (120 Days). NCT00438568. Available at: http://www. clinicaltrials.gov. Accessed March 10, 2008. Gauthier S, Reisberg B, Zaudig M, et al. Mild cognitive impairment. Lancet 2006;67:1262–1270. Reger MA, Watson GS, Green PS, et al. Intranasal insulin administration dose-dependently modulates verbal memory and plasma amyloid-a in memory-impaired older adults. J Alzheimer Dis 2008 (in press). Craft S, Asthana S, Schellenberg GD, et al. Insulin metabolism in Alzheimer’s disease differs according to apolipoprotein E genotype. Neuroendocrinology 1999;295: 2750 –2757. Risner ME, Saunders AM, Altman JF, et al. Efficacy of rosiglitazone in a genetically defined population with mildto-moderate Alzheimer’s disease. Pharmacogenomics J 2006;4:246 –254.

VOLUNTARY PARTIAL RETRACTION Voluntary partial retraction of: Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes The authors state that they inadvertently published incorrect data in the analysis of one of the mutations in this manuscript. The data presented for the analysis of mutation AChR ⑀L78P (c.233C⬎T) was for the analysis of mutation AChR ⑀S278del (c.833_835delCTC). It is mutation ⑀S278del that causes a slow channel syndrome, not ⑀L78P. The study of the clinical features within the kinship harboring mutation ⑀S278del finds that the slow channel syndrome phenotype is revealed through the presence of a low expression mutation, ⑀R217L, on the second allele so the observational hypothesis in the original manuscript is valid. In the experiments described, the core AChR ⑀-subunit cDNA varies from the Genbank reference sequence (http://www.ncbi.nlm.nih.gov/entrz/viewer.fcgi?val⫽NM_000080) by two rare missense polymorphisms, ⑀V108A and ⑀T117S, that were present in the original clone isolated from a lambdagt10 human muscle cDNA library. Croxen R, Hatton C, Shelley C, Brydson M, Chauplannaz G, Oosterhuis H, Vincent A, Newsom-Davis J, Colquhoun D, Beeson D. Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes. Neurology 2002;59:162–168.

294

Neurology 72

January 20, 2009

Correspondence

TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHY IN THE 21st CENTURY: NEUROSCIENCE FOR THE CLINICAL NEUROLOGIST

To the Editor: I read Brown’s article1 on transmissible spongiform encephalopathy (TSE) with interest. Other disease mechanisms apart from seeding by misfolded prion are under consideration in TSE. “Slow virus” infections might provide an explanation for the unusual characteristics of TSE agents, such as persistence for years and spread of the bovine spongiform encephalopathy agent to divergent species.2 Spiroplasma bacteria have been considered to be related to TSE, since they have been recovered from brain tissues of sheep with scrapie, cervids with chronic wasting disease, and patients with Creutzfeldt-Jakob disease by passage through embryonated eggs.3 Spiroplasma bacteria resist high temperatures, radiation, and fixatives.3 Prion protein may be induced on the cell surface by bacterial infection.3 Scrapie associated fibrils commonly found in TSE also appear identical to bacterial Spiroplasma fibrils.3 When Spiroplasma mirum was inoculated intracranially (IC) into deer, sheep, and goats, it caused spongiform encephalopathy. It had already been shown to cause spongiform encephalopathy in rodents. Spiroplasma spp was isolated from scrapie-affected sheep and from chronic wasting disease–affected deer by passage through embryonated eggs. When inoculated IC into sheep and goats, it caused spongiform encephalopathy in the animals.4 There may be a vector or reservoir in some instances in the spread of TSE. Hay mite preparations from five Icelandic farms with a history of scrapie caused spongiform characteristic of scrapie when injected into mice, indicating probably some vector was involved.5 An infectious agent, possibly a slow virus or Spiroplasma bacteria, may be responsible for some cases of TSE. Steven R. Brenner, St. Louis, MO

Interestingly, the NIH Laboratory where I spent my career bore the name “Slow, Latent, and Temperate Viruses of the CNS,” and for quite some time it looked as though a “slow virus” might be at the root of the problem (similar to the JC virus that causes progressive multifocal leukoencephalopathy). However, as new observations were made,6 the concept of a slow virus became less and less tenable, at least as the sole cause of disease, which certainly can no longer be seriously considered. Regarding the Spiroplasma hypothesis, the failure of a blinded study of rRNA species to detect any footprint of Spiroplasma in scrapie-infected hamster brain7 laid it to rest, unless one holds that the cause of scrapie is different from that of other prion diseases, an equally untenable proposition. Paul W. Brown, Bethesda, MD Disclosure: The author reports no disclosures. Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

3.

4.

5.

6.

7.

Brown P. Transmissible spongiform encephalopathy in the 21st century: neuroscience for the clinical neurologist. Neurology 2008;70:713–722. Manuelidis L. A 25 nm virion is the likely cause of transmissible spongiform encephalopathies. J Cell Biochem 2007;100:897–915. Bastian F. Spiroplasma as a candidate agent for transmissible spongiform encephalopathies. J Neuropathol Exp Neurol 2005;64:833– 838. Bastian F, Sanders D, Forbes W, et al. Spiroplasma spp. from transmissible spongiform encephalopathy brains or ticks induce spongiform encephalopathy in ruminants. J Med Microbiol 2007;56:1235–1242. Carp TI, Meeker, HC, Rubenstein R. Characteristics of scrapie isolates derived from hay mites. J Neurovirol 2000;6: 137–142. Brown P. The “brave new world” of transmissible spongiform encephalopathy (infectious cerebral amyloidosis). Molec Neurobiol 1994;8:79 – 87. Alexeeva I, Elliott EJ, Rollins S, et al. Absence of Spiroplasma or other bacterial 16S rRNA genes in brain tissue of hamsters with scrapie. J Clin Microbiol 2006;44:91–97.

Disclosure: The author reports no disclosures. INTRANASAL INSULIN IMPROVES COGNITION

Reply from the Author: Dr. Brenner considers the possibility that something other than the prion protein might be the cause of prion diseases. I sympathize with his caution, but not his choice of alternatives. 292

Neurology 72

January 20, 2009

AND MODULATES ␤-AMYLOID IN EARLY AD

To the Editor: At our department’s recent journal club conference, we discussed the report by Reger et al.1 on intranasal insulin in early Alzheimer disease (AD). Given findings of epidemiologic and transla-

tional research associating diabetes and dementia, these results are an intriguing extension of their earlier work but raised several questions.2 First, the sample was somewhat heterogeneous— comprising patients with AD but also mild cognitive impairment with amnestic features (aMCI). Some baseline differences were evident in cognition and serologic markers between treatment and placebo. For example, memory savings scores for baseline placebo and treatment, plus 21-day follow-up placebo arms, appeared to be different, but would be expected to be rather similar. Could these differences be related to incomplete randomization, possibly related to small sample size? The proportion of AD and aMCI patients by treatment and placebo groups would be useful to know. Several patients were unable to perform the Stroop Color-Word task or had missing bloodwork. From which groups were these data missing? Logical memory percent retention and Stroop performance speed were the neuropsychological tests presented. In the group’s previous work, additional tests included the Buschke Selective Reminding Test (SRT), the Self-Ordered Pointing Task, and a visual search task.2 That report focused on changes in total delayed logical memory and SRT relative to placebo, with no differences identified on tests of attention or processing speed. If other tests were performed but not reported, it would be helpful if the authors could provide a more comprehensive neuropsychological summary. That report also found a significant interaction with APOE genotype. Was APOE genotyping done for this study? Most analyses compared group means rather than differences in individual performance over test sequence yet the latter might be more informative. Concerning the correlation of Dementia Severity Rating Scale (DSRS) changes with the baseline Mattis Dementia Rating Scale (DRS) (figure 3), did these data satisfy an assumption of homoscedasticity prior to fitting the correlation line? Lower DRS in both groups appeared to be associated with higher variance in DSRS. Also, given a seemingly large fluctuation in DSRS over 21 days within one placebo outlier, was exclusion of this outlier considered? Different DSRS scales between the two groups further complicated interpretation. We appreciate the difficulty of undertaking such a study and satisfying all readers. Perhaps this forum will give the authors an opportunity to present other data. We look forward to the results of their ongoing 120-day trial.3 Mandip S. Dhamoon, MD, MPH, James M. Noble, MD, New York, NY Disclosure: The authors report no disclosures.

Reply from the Authors: We appreciate the careful consideration of our article by Drs. Dhamoon and

Noble. Our sample included both patients with aMCI and early AD, diagnosed by expert consensus. When diagnosed in this manner, the majority of patients with aMCI have prodromal AD.4 The sample was heterogeneous with respect to severity but likely not type of impairment. Most importantly, the two groups contained comparable numbers of aMCI and AD patients (aMCI comprised 50% of the placebo group and 62% of the insulin-treated group), and the groups had comparable dementia severity as indicated in table 1 by the DRS. Similarly, there were no significant baseline differences between groups on any cognitive measure, nor were there differences in missing data (two placebo- and one insulin-treated participant unable to complete the Stroop, with an identical pattern observed for amyloid). We agree that randomization is difficult for small pilot trials such as ours, and were fortunate that our groups were comparable on cognitive, demographic, and metabolic variables. Our cognitive battery was tightly focused for this pilot effort. We used the Hopkins Verbal Learning Test rather than the SRT because it is more easily administered. One additional test was included (SelfOrdered Pointing Test), but computer malfunction resulted in uninterpretable data for the majority of participants. We used a repeated measures analysis of covariance analytic approach, which is preferable with small samples, given the variability of difference scores for cognitive measures. Our analysis for the DSRS revealed an interaction with dementia severity as indexed by the Mattis DRS, which was then deconstructed by correlating Mattis scores and the change in Dementia Severity Rating over the 21-day period. We reported Pearson correlations, which as noted may be vulnerable to violation of homoscedasticity or outlier effects, particularly in small samples. However, the correlation pattern remains identical and statistically significant when Spearman rank order correlations are used which protect against the effects of such violations. Finally, the issue of APOE genotype raises an important point. In two previous studies of the acute effects of intranasal insulin, we observed an APOErelated pattern, such that participants without the APOE-e4 allele benefited from insulin, whereas e4 carriers showed no benefit.2,5 This APOE-related difference has been observed with other manipulations of insulin.6,7 The pattern may support a specialized role for insulin abnormalities in the pathophysiology of AD for non-e4 carriers, who comprise about half of patients with AD, but for whom no clear risk factor has been identified. In our pilot study, the insulin-treated group was evenly divided with respect to APOE-e4 carriage Neurology 72

January 20, 2009

293

(e4 – ⫽ 7, e4⫹ ⫽ 6). These cell sizes are too small to allow exploration of a differential APOE treatment response, but our ongoing larger clinical trial is powered to determine such an effect.3 We hope to soon have data to address this important question.

3.

4.

Suzanne Craft, PhD, Seattle, WA Disclosure on article to which this Correspondence refers: W.D. is a founding member of Kurve Technology, the maker and patent holder of the electronic atomizer used in this study. W.D. holds equity interest in excess of $10,000 in Kurve Technology. The remaining authors have nothing to disclose.

5.

6.

Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

Reger MA, Watson GS, Green PS, et al. Intranasal insulin improves cognition and modulates beta-amyloid in early AD. Neurology 2008;70:440 – 448. Reger MA, Watson GS, Frey WH 2nd, et al. Effects of intranasal insulin on cognition in memory-impaired older

7.

adults: modulation by APOE genotype. Neurobiol Aging 2006;27:451– 458. SNIFF 120: Study of Nasal Insulin to Fight Forgetfulness (120 Days). NCT00438568. Available at: http://www. clinicaltrials.gov. Accessed March 10, 2008. Gauthier S, Reisberg B, Zaudig M, et al. Mild cognitive impairment. Lancet 2006;67:1262–1270. Reger MA, Watson GS, Green PS, et al. Intranasal insulin administration dose-dependently modulates verbal memory and plasma amyloid-a in memory-impaired older adults. J Alzheimer Dis 2008 (in press). Craft S, Asthana S, Schellenberg GD, et al. Insulin metabolism in Alzheimer’s disease differs according to apolipoprotein E genotype. Neuroendocrinology 1999;295: 2750 –2757. Risner ME, Saunders AM, Altman JF, et al. Efficacy of rosiglitazone in a genetically defined population with mildto-moderate Alzheimer’s disease. Pharmacogenomics J 2006;4:246 –254.

VOLUNTARY PARTIAL RETRACTION Voluntary partial retraction of: Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes The authors state that they inadvertently published incorrect data in the analysis of one of the mutations in this manuscript. The data presented for the analysis of mutation AChR ⑀L78P (c.233C⬎T) was for the analysis of mutation AChR ⑀S278del (c.833_835delCTC). It is mutation ⑀S278del that causes a slow channel syndrome, not ⑀L78P. The study of the clinical features within the kinship harboring mutation ⑀S278del finds that the slow channel syndrome phenotype is revealed through the presence of a low expression mutation, ⑀R217L, on the second allele so the observational hypothesis in the original manuscript is valid. In the experiments described, the core AChR ⑀-subunit cDNA varies from the Genbank reference sequence (http://www.ncbi.nlm.nih.gov/entrz/viewer.fcgi?val⫽NM_000080) by two rare missense polymorphisms, ⑀V108A and ⑀T117S, that were present in the original clone isolated from a lambdagt10 human muscle cDNA library. Croxen R, Hatton C, Shelley C, Brydson M, Chauplannaz G, Oosterhuis H, Vincent A, Newsom-Davis J, Colquhoun D, Beeson D. Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes. Neurology 2002;59:162–168.

294

Neurology 72

January 20, 2009

Correspondence

TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHY IN THE 21st CENTURY: NEUROSCIENCE FOR THE CLINICAL NEUROLOGIST

To the Editor: I read Brown’s article1 on transmissible spongiform encephalopathy (TSE) with interest. Other disease mechanisms apart from seeding by misfolded prion are under consideration in TSE. “Slow virus” infections might provide an explanation for the unusual characteristics of TSE agents, such as persistence for years and spread of the bovine spongiform encephalopathy agent to divergent species.2 Spiroplasma bacteria have been considered to be related to TSE, since they have been recovered from brain tissues of sheep with scrapie, cervids with chronic wasting disease, and patients with Creutzfeldt-Jakob disease by passage through embryonated eggs.3 Spiroplasma bacteria resist high temperatures, radiation, and fixatives.3 Prion protein may be induced on the cell surface by bacterial infection.3 Scrapie associated fibrils commonly found in TSE also appear identical to bacterial Spiroplasma fibrils.3 When Spiroplasma mirum was inoculated intracranially (IC) into deer, sheep, and goats, it caused spongiform encephalopathy. It had already been shown to cause spongiform encephalopathy in rodents. Spiroplasma spp was isolated from scrapie-affected sheep and from chronic wasting disease–affected deer by passage through embryonated eggs. When inoculated IC into sheep and goats, it caused spongiform encephalopathy in the animals.4 There may be a vector or reservoir in some instances in the spread of TSE. Hay mite preparations from five Icelandic farms with a history of scrapie caused spongiform characteristic of scrapie when injected into mice, indicating probably some vector was involved.5 An infectious agent, possibly a slow virus or Spiroplasma bacteria, may be responsible for some cases of TSE. Steven R. Brenner, St. Louis, MO

Interestingly, the NIH Laboratory where I spent my career bore the name “Slow, Latent, and Temperate Viruses of the CNS,” and for quite some time it looked as though a “slow virus” might be at the root of the problem (similar to the JC virus that causes progressive multifocal leukoencephalopathy). However, as new observations were made,6 the concept of a slow virus became less and less tenable, at least as the sole cause of disease, which certainly can no longer be seriously considered. Regarding the Spiroplasma hypothesis, the failure of a blinded study of rRNA species to detect any footprint of Spiroplasma in scrapie-infected hamster brain7 laid it to rest, unless one holds that the cause of scrapie is different from that of other prion diseases, an equally untenable proposition. Paul W. Brown, Bethesda, MD Disclosure: The author reports no disclosures. Copyright © 2009 by AAN Enterprises, Inc. 1.

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Brown P. Transmissible spongiform encephalopathy in the 21st century: neuroscience for the clinical neurologist. Neurology 2008;70:713–722. Manuelidis L. A 25 nm virion is the likely cause of transmissible spongiform encephalopathies. J Cell Biochem 2007;100:897–915. Bastian F. Spiroplasma as a candidate agent for transmissible spongiform encephalopathies. J Neuropathol Exp Neurol 2005;64:833– 838. Bastian F, Sanders D, Forbes W, et al. Spiroplasma spp. from transmissible spongiform encephalopathy brains or ticks induce spongiform encephalopathy in ruminants. J Med Microbiol 2007;56:1235–1242. Carp TI, Meeker, HC, Rubenstein R. Characteristics of scrapie isolates derived from hay mites. J Neurovirol 2000;6: 137–142. Brown P. The “brave new world” of transmissible spongiform encephalopathy (infectious cerebral amyloidosis). Molec Neurobiol 1994;8:79 – 87. Alexeeva I, Elliott EJ, Rollins S, et al. Absence of Spiroplasma or other bacterial 16S rRNA genes in brain tissue of hamsters with scrapie. J Clin Microbiol 2006;44:91–97.

Disclosure: The author reports no disclosures. INTRANASAL INSULIN IMPROVES COGNITION

Reply from the Author: Dr. Brenner considers the possibility that something other than the prion protein might be the cause of prion diseases. I sympathize with his caution, but not his choice of alternatives. 292

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AND MODULATES ␤-AMYLOID IN EARLY AD

To the Editor: At our department’s recent journal club conference, we discussed the report by Reger et al.1 on intranasal insulin in early Alzheimer disease (AD). Given findings of epidemiologic and transla-

tional research associating diabetes and dementia, these results are an intriguing extension of their earlier work but raised several questions.2 First, the sample was somewhat heterogeneous— comprising patients with AD but also mild cognitive impairment with amnestic features (aMCI). Some baseline differences were evident in cognition and serologic markers between treatment and placebo. For example, memory savings scores for baseline placebo and treatment, plus 21-day follow-up placebo arms, appeared to be different, but would be expected to be rather similar. Could these differences be related to incomplete randomization, possibly related to small sample size? The proportion of AD and aMCI patients by treatment and placebo groups would be useful to know. Several patients were unable to perform the Stroop Color-Word task or had missing bloodwork. From which groups were these data missing? Logical memory percent retention and Stroop performance speed were the neuropsychological tests presented. In the group’s previous work, additional tests included the Buschke Selective Reminding Test (SRT), the Self-Ordered Pointing Task, and a visual search task.2 That report focused on changes in total delayed logical memory and SRT relative to placebo, with no differences identified on tests of attention or processing speed. If other tests were performed but not reported, it would be helpful if the authors could provide a more comprehensive neuropsychological summary. That report also found a significant interaction with APOE genotype. Was APOE genotyping done for this study? Most analyses compared group means rather than differences in individual performance over test sequence yet the latter might be more informative. Concerning the correlation of Dementia Severity Rating Scale (DSRS) changes with the baseline Mattis Dementia Rating Scale (DRS) (figure 3), did these data satisfy an assumption of homoscedasticity prior to fitting the correlation line? Lower DRS in both groups appeared to be associated with higher variance in DSRS. Also, given a seemingly large fluctuation in DSRS over 21 days within one placebo outlier, was exclusion of this outlier considered? Different DSRS scales between the two groups further complicated interpretation. We appreciate the difficulty of undertaking such a study and satisfying all readers. Perhaps this forum will give the authors an opportunity to present other data. We look forward to the results of their ongoing 120-day trial.3 Mandip S. Dhamoon, MD, MPH, James M. Noble, MD, New York, NY Disclosure: The authors report no disclosures.

Reply from the Authors: We appreciate the careful consideration of our article by Drs. Dhamoon and

Noble. Our sample included both patients with aMCI and early AD, diagnosed by expert consensus. When diagnosed in this manner, the majority of patients with aMCI have prodromal AD.4 The sample was heterogeneous with respect to severity but likely not type of impairment. Most importantly, the two groups contained comparable numbers of aMCI and AD patients (aMCI comprised 50% of the placebo group and 62% of the insulin-treated group), and the groups had comparable dementia severity as indicated in table 1 by the DRS. Similarly, there were no significant baseline differences between groups on any cognitive measure, nor were there differences in missing data (two placebo- and one insulin-treated participant unable to complete the Stroop, with an identical pattern observed for amyloid). We agree that randomization is difficult for small pilot trials such as ours, and were fortunate that our groups were comparable on cognitive, demographic, and metabolic variables. Our cognitive battery was tightly focused for this pilot effort. We used the Hopkins Verbal Learning Test rather than the SRT because it is more easily administered. One additional test was included (SelfOrdered Pointing Test), but computer malfunction resulted in uninterpretable data for the majority of participants. We used a repeated measures analysis of covariance analytic approach, which is preferable with small samples, given the variability of difference scores for cognitive measures. Our analysis for the DSRS revealed an interaction with dementia severity as indexed by the Mattis DRS, which was then deconstructed by correlating Mattis scores and the change in Dementia Severity Rating over the 21-day period. We reported Pearson correlations, which as noted may be vulnerable to violation of homoscedasticity or outlier effects, particularly in small samples. However, the correlation pattern remains identical and statistically significant when Spearman rank order correlations are used which protect against the effects of such violations. Finally, the issue of APOE genotype raises an important point. In two previous studies of the acute effects of intranasal insulin, we observed an APOErelated pattern, such that participants without the APOE-e4 allele benefited from insulin, whereas e4 carriers showed no benefit.2,5 This APOE-related difference has been observed with other manipulations of insulin.6,7 The pattern may support a specialized role for insulin abnormalities in the pathophysiology of AD for non-e4 carriers, who comprise about half of patients with AD, but for whom no clear risk factor has been identified. In our pilot study, the insulin-treated group was evenly divided with respect to APOE-e4 carriage Neurology 72

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(e4 – ⫽ 7, e4⫹ ⫽ 6). These cell sizes are too small to allow exploration of a differential APOE treatment response, but our ongoing larger clinical trial is powered to determine such an effect.3 We hope to soon have data to address this important question.

3.

4.

Suzanne Craft, PhD, Seattle, WA Disclosure on article to which this Correspondence refers: W.D. is a founding member of Kurve Technology, the maker and patent holder of the electronic atomizer used in this study. W.D. holds equity interest in excess of $10,000 in Kurve Technology. The remaining authors have nothing to disclose.

5.

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Copyright © 2009 by AAN Enterprises, Inc. 1.

2.

Reger MA, Watson GS, Green PS, et al. Intranasal insulin improves cognition and modulates beta-amyloid in early AD. Neurology 2008;70:440 – 448. Reger MA, Watson GS, Frey WH 2nd, et al. Effects of intranasal insulin on cognition in memory-impaired older

7.

adults: modulation by APOE genotype. Neurobiol Aging 2006;27:451– 458. SNIFF 120: Study of Nasal Insulin to Fight Forgetfulness (120 Days). NCT00438568. Available at: http://www. clinicaltrials.gov. Accessed March 10, 2008. Gauthier S, Reisberg B, Zaudig M, et al. Mild cognitive impairment. Lancet 2006;67:1262–1270. Reger MA, Watson GS, Green PS, et al. Intranasal insulin administration dose-dependently modulates verbal memory and plasma amyloid-a in memory-impaired older adults. J Alzheimer Dis 2008 (in press). Craft S, Asthana S, Schellenberg GD, et al. Insulin metabolism in Alzheimer’s disease differs according to apolipoprotein E genotype. Neuroendocrinology 1999;295: 2750 –2757. Risner ME, Saunders AM, Altman JF, et al. Efficacy of rosiglitazone in a genetically defined population with mildto-moderate Alzheimer’s disease. Pharmacogenomics J 2006;4:246 –254.

VOLUNTARY PARTIAL RETRACTION Voluntary partial retraction of: Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes The authors state that they inadvertently published incorrect data in the analysis of one of the mutations in this manuscript. The data presented for the analysis of mutation AChR ⑀L78P (c.233C⬎T) was for the analysis of mutation AChR ⑀S278del (c.833_835delCTC). It is mutation ⑀S278del that causes a slow channel syndrome, not ⑀L78P. The study of the clinical features within the kinship harboring mutation ⑀S278del finds that the slow channel syndrome phenotype is revealed through the presence of a low expression mutation, ⑀R217L, on the second allele so the observational hypothesis in the original manuscript is valid. In the experiments described, the core AChR ⑀-subunit cDNA varies from the Genbank reference sequence (http://www.ncbi.nlm.nih.gov/entrz/viewer.fcgi?val⫽NM_000080) by two rare missense polymorphisms, ⑀V108A and ⑀T117S, that were present in the original clone isolated from a lambdagt10 human muscle cDNA library. Croxen R, Hatton C, Shelley C, Brydson M, Chauplannaz G, Oosterhuis H, Vincent A, Newsom-Davis J, Colquhoun D, Beeson D. Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes. Neurology 2002;59:162–168.

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Section Editors Christopher J. Boes, MD Kenneth J. Mack, MD, PhD

Book Review

PLUM AND POSNER’S DIAGNOSIS OF STUPOR AND COMA, 4TH EDITION

edited by Jerome B. Posner, Clifford B. Saper, Nicholas D. Schiff, and Fred Plum, 401 pp., Oxford University Press, 2007 $82.95 This text represents an extensive revision and update of a true classic in clinical neurology. Most of us grew up with the prior edition, now of distinguished vintage after some 25 years. In the preface, the authors clearly set forth the goal to set the clinical examination in the context of technological advances. The new edition contains nine chapters. The first reviews the basic pathophysiology of awareness and coma, again representing one of the best reviews of this material. The references have been updated and new data have been added to the classic functional anatomic discussion. The second chapter covers the examination of the comatose patient, again done better nowhere else. This chapter provides the clinician with the practical and conceptual information needed to evaluate the unresponsive patient. The third chapter deals with the structural causes of stu-

por and coma. This discussion has been updated to include the significance of tissue shifts as well as herniation. Yet the value of this chapter resides in the excellent clinical descriptions of the herniation syndromes and their anatomic correlates. Additional chapters cover specific structural causes of coma, as well as metabolic and psychogenic causes of unresponsiveness. These discussions have been updated, but retain the classic style of the prior edition. Brain death and prognosis are afforded individual chapters, and new data have been incorporated. The discussion of functional imaging in vegetative and minimally responsive states is timely and helpful. It is good to see a clinical classic updated, and this edition has been carefully edited to add relevant new data without altering the clinical feel of the prior work. The first three chapters alone provide adequate reason for this to be a “must reference” available to all neurology residents and clinicians. Reviewed by Daryl R. Gress, MD Disclosure: The author reports no disclosures. Copyright © 2009 by AAN Enterprises, Inc.

New Categories of Resident & Fellow Section Clinical Reasoning: Case presentations to aid in developing clinical reasoning skills. Right Brain: Neurology and the medical humanities — history, literature, and arts. Child Neurology: Patient case with detailed discussion about topic of focus. Pearls and Oy-sters: Clinical insights (pearls) and advice for avoiding mistakes (oysters). International: Educational exchanges, experiences in low and middle income countries. Emerging Subspecialities: History of fields such as Pain Medicine and Headache. Continue to submit articles about education research and educational topics, training videos, and teaching NeuroImages!

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Calendar

2009 Neurology® publishes short announcements of meetings and courses related to the field. Items must be received at least 6 weeks before the first day of the month in which the initial notice is to appear. Send Calendar submissions to Calendar, Editorial Office, Neurology®, Suite 214, 20 SW 2nd Ave., P.O. Box 178, Rochester, MN 55903 [email protected]

JAN. 16 –18 AAN Winter Conference will be held at Disney Contemporary Resort in Orlando, FL. American Academy of Neurology: tel (800) 879-1960; www.aan.com/winter. FEB. 9 –11 Case Studies in Epilepsy Surgery will be held at the Silver Tree and Snowmass Conference Center in Snowmass, CO. Contact Martha Tobin at (216) 445-3449 or (800) 2232273, ext 53449, or at [email protected] for seminar details. FEB. 9 –13 The 22nd Annual Practicing Physician’s Approach to the Difficult Headache Patient will be held at the Camelback Inn, Scottsdale, AZ. Approved for AMA PRA Category 1 credit. Diamond Headache Clinic Research & Educational Foundation: tel (877) 706-6363 or (733) 883-2062; [email protected]; www.dhc-fdn.org. FEB. 16 –17 Fifth Annual Update Symposium on Clinical Neurology and Neurophysiology will be held in Tel Aviv, Israel. Presented by Weill Cornell Medical College, Department of Neurology, and Tel Aviv University, Adams Brain Supercenter. www.neurophysiology-symposium.com. FEB. 20 –22 International Symposium on Stereotactic Body Radiation Therapy and Stereotactic Radiosurgery will be held at the Floridian Resort & Spa in Lake Buena Vista, FL. Contact Martha Tobin at (216) 445-3449 or (800) 223-2273, ext 53449, or at [email protected] for seminar details.

MAY 6 –10 International SFEMG Course and Xth Quantitative EMG conference will be held in Venice, Italy. tel 39041-951112; [email protected]; www.congressvenezia.it. MAY 8 The Office of Continuing Medical Education at the University of Michigan Medical School is sponsoring a CME conference entitled: Movement Disorders: A Practical Approach. It is located at The Inn at St. John’s in Plymouth, Michigan. tel (734) 763-1400; fax (734) 936-1641. MAY 11–12 Music and the Brain will be held at the InterContinental Hotel & Bank of America Conference Center in Cleveland, OH. Contact Martha Tobin at (216) 445-3449 or (800) 223-2273, ext 53449, or at [email protected] for seminar details. MAY 15–17 The Fifth International Conference on Alzheimer’s Disease and Related Disorders in the Middle East will be held in Limassol, Cyprus. www.worldeventsforum.com/alz. MAY 28 –30 6th International Headache Seminary. Focus on Headaches: New Frontier in Mechanisms and Management will be held at the Grand Hotel des Iles Borromees in Stresa (Italy); tel/fax 02 7063 8067; [email protected].

APR. 2– 4 The Innsbruck Colloquium on Status Epilepticus 2009 will be held at the Congress Innsbruck, Austria. [email protected]; www.innsbruck-SE2009.eu.

JUN. 8 –12 Leksell Gamma Knife® Perfexion™ Introductory Course will be held at the Gamma Knife Center in Cleveland, OH. Contact Martha Tobin at (216) 445-3449 or (800) 223-2273, ext 53449, or at [email protected] for seminar details.

APR. 3 5th Annual Contemporary Issues in Pituitary: Casebase Management Update will be held at the Cleveland Clinic Lerner Research Institute in Cleveland, OH. Contact Martha Tobin at (216) 445-3449 or (800) 223-2273, ext 53449, or at [email protected] for seminar details.

JUN. 12 Mellen Center Regional Symposium on Multiple Sclerosis will be held at the InterContinental Hotel & Bank of America Conference Center in Cleveland, OH. Contact Martha Tobin at (216) 445-3449 or (800) 223-2273, ext 53449, or at [email protected] for seminar details.

APR. 20 –22 Leksell Gamma Knife® Perfexion™ Upgrade Course will be held at the Gamma Knife Center in Cleveland, OH. Contact Martha Tobin at (216) 445-3449 or (800) 2232273, ext 53449, or at [email protected] for seminar details.

JUN. 19 –24 Epileptology Symposium will be held at the InterContinental Hotel & Bank of America Conference Center, in Cleveland, OH. Contact Martha Tobin at (216) 445-3449 or (800) 223-2273, ext 53449, or at [email protected] for seminar details.

APR. 25–MAY 2 AAN Annual Meeting will be held in Seattle, Washington State Convention & Trade Center, WA. American Academy of Neurology: tel (800) 879-1960; www.aan.com/am.

JUL. 7–10 SickKids Centre for Brain & Behaviour International Symposium. [email protected]; www.sickkids.ca/ learninginstitute.

APR. 26 The 21st Annual Symposium on the Treatment of Headaches and Facial Pain will be held at the New York, Marriott East Side Hotel in New York, NY. For further details please contact: Alexander Mauskop, MD; tel (212) 794-3550; fax (212) 794-0591; [email protected]. 296

MAY 3– 6 2nd International Epilepsy Colloquium, Pediatric Epilepsy Surgery Cite´ Internationale will be held in Lyon, France. http://epilepsycolloquium2009ams.fr.

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JUL. 16 –18 Mayo Clinic Neurology in Clinical Practice2009 will be held at the InterContinental Hotel, Chicago, IL. Mayo CME: tel: (800) 323-2688; [email protected]; http:// www.mayo.edu/cme/neurology-neurologic-surgery.html. SEP. 12–15 13th Congress of the European Federation of Neurological Societies will be held in Florence, Italy. For more informa-

tion: tel ⫹41 22 908 0488; http://www.kenes.com/efns2009/ index.asp; [email protected]. SEP. 25 Practical Pearls in Neuro-Ophthalmology–International Symposium in Honour of Dr. James Sharpe will be held on September 25, 2009 at the University of Toronto Conference Centre, Toronto, Ontario. For further information contact the Office of Continuing Education & Professional Development, Faculty of Medicine, University of Toronto: tel (416) 978-2719; (888) 5128173; fax (416) 946-7028; [email protected]; http:// events.cmetoronto.ca/website/index/OPT0907. OCT. 8 –11 The Third World Congress on Controversies in Neurology. Full information is available at: ComtecMed - Med-

ical Congresses, PO Box 68, Tel-Aviv, 61000 Israel; tel ⫹972– 3-5666166; fax ⫹972–3-5666177; cony@comtecmed. com; www.comtecmed.com/cony. OCT. 24 –30 19th World Congress of Neurology, WCN 2009, will be held in Bangkok, Thailand. www.wcn2009bangkok.com. NOV. 19 –22 The Sixth International Congress on Vascular Dementia will be held Barcelona, Spain. For further details, please contact: Kenes International 17 Rue du Cendrier, P.O. Box 1726, CH-1211, Geneva 1, Switzerland; tel ⫹41 22 908 0488; fax ⫹41 22 732 2850; [email protected]; http://www. kenes.com/vascular.

Save These Dates for AAN CME Opportunities! Mark these upcoming dates on your calendar for these exciting continuing education opportunities, where you can catch up on the latest neurology information. AAN Annual Meetings ● April 25—May 2, 2009, Seattle, Washington State Convention & Trade Center ● April 10 –17, 2010, Toronto, Ontario, Canada, Toronto Convention Centre

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In the next issue of Neurology® Volume 72, Number 4, January 27, 2009 www.neurology.org THE MOST WIDELY READ AND HIGHLY CITED PEER-REVIEWED NEUROLOGY JOURNAL

THIS WEEK IN Neurology®

299

Highlights of the January 27 issue

VIEWS & REVIEWS

368

EDITORIALS

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302

Idiopathic intracranial hypertension in men and the relationship to sleep apnea Michael Wall and Valerie Purvin

Vascular risk factors and dementia: How to move forward? A. Viswanathan, W.A. Rocca, and C. Tzourio

CLINICAL/SCIENTIFIC NOTES

375

Novel FHL1 mutations in fatal and benign reducing body myopathy S. Shalaby, Y.K. Hayashi, I. Nonaka, S. Noguchi, et al.

377

Elemental mercury neurotoxicity from self-injection H.H. Schaumburg, C. Gellido, S.W. Smith, et al. Linezolid inducing complex partial status epilepticus in a patient with epilepsy B.F. Shneker, P.D. Baylin, and M.E. Nakhla

ACGME, test thyself! Steven Feske

ARTICLES

304

Idiopathic intracranial hypertension in men B.B. Bruce, S. Kedar, G.P. Van Stavern, et al.

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Mutations in GBA are associated with familial Parkinson disease susceptibility and age at onset W.C. Nichols, et al., for the Parkinson Study GroupPROGENI Investigators

REFLECTIONS: NEUROLOGY AND THE HUMANITIES

Effect of aerobic training in patients with spinal and bulbar muscular atrophy (Kennedy disease) N. Preisler, G. Andersen, F. Thøgersen, et al.

NEUROIMAGES

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324

Comparing explorative saccade and flicker training in hemianopia: A randomized controlled study T. Roth, A.N. Sokolov, A. Messias, P. Roth, et al.

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TP53 codon 72 polymorphism is associated with age at onset of glioblastoma S. El Hallani, F. Ducray, A. Idbaih, Y. Marie, et al.

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Treatment of CNS sarcoidosis with infliximab and mycophenolate mofetil Michael Moravan and Benjamin M. Segal

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Treatment of intractable chronic cluster headache by occipital nerve stimulation in 14 patients B. Burns, L. Watkins, and P.J. Goadsby Epidemiology of ischemic stroke from atrial fibrillation in Dijon, France, from 1985 to 2006 Y. Be ´jot, D. Ben Salem, G.V. Osseby, et al. Factors associated with resistance to dementia despite high Alzheimer disease pathology D. Erten-Lyons, R.L. Woltjer, H. Dodge, et al. Biochemical indicators of vitamin B12 and folate insufficiency and cognitive decline C.C. Tangney, Y. Tang, D.A. Evans, and M.C. Morris

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Poems Arthur Ginsberg

Open-ring peripherally enhancing lesion of the cervical spine W. Pyle, K. Dastur, M. Rahman, and J. Tsay

RESIDENT & FELLOW SECTION

e15

Education research: Neurology residency training in the new millennium L.A. Schuh, J.C. Adair, O. Drogan, B.M. Kissela, et al.

PATIENT PAGE

e21

Alzheimer disease: What changes in the brain cause dementia? Kathleen A. Welsh-Bohmer and Charles L. White III

CORRESPONDENCE

382

Practice parameter: Assessing patients for falls

383

Lyme disease treatment

DEPARTMENTS

387 388

Book Review Calendar

FUTURE ISSUES

Abstracts In the Next Issue of Neurology®

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