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APPENDIX
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EBMT Officers
EBMT Coordination Offices
President Dietger Niederwieser
EBMT Secretariat & JACIE Accreditation Office
Division of Haematology & Oncology University Hospital Leipzig Johannisallee 32 A 04103 Leipzig GERMANY Tel: +49 341 971 3050 Fax: +49 341 971 3059
[email protected]
Hospital Clínic Villarroel 170, 08036 Barcelona SPAIN Tel: +34 93 454 9543 Fax: +34 93 453 1263
[email protected] [email protected]
Secretary Per Ljungman
12th Floor Tower Wing Guy's Hospital Great Maze Pond London SE1 9RT UNITED KINGDOM
Department of Haematology Karolinska University Hosp/Huddinge 141 86 Stockholm SWEDEN Tel.: +46 8 585 80000 Fax: +46 8 774 8725
[email protected]
Treasurer (2002 – 2008) Harry Schouten University Hospital Maastricht Dept. Hematology P.O. Box 58000 6202 AZ Maastricht The Netherlands Tel.: +31 43 3 877025 Fax: +31 43 3 875006
[email protected]
EBMT Clinical Trials & Registry Office
Registry Tel: +44 207 188 8408 Fax: +44 207 188 8411
[email protected]
Clinical Trials Tel: +44 207 188 8402 Fax: +44 207 188 8406
[email protected]
EBMT Data & Study Office Faculté de Médecine Saint-Antoine 27, rue Chaligny, 75571 Paris Cedex 12 FRANCE Tel: +33 1 40 46 95 07 Fax: +33 1 40 46 96 07
[email protected]
EBMT Clinical Trials & Study Office Department of Medical Statistics & Bioinformatics, Postzone S-05-P LUMC, PO Box 9600 2300 RC Leiden THE NETHERLANDS
[email protected] HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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EBMT Committees Cord Blood Committee Chair: Eliane Gluckman
Outreach Committee Co-Chairs: Vladimir Koza
[email protected]
[email protected]
Eliane Gluckman Developmental Committee Co-Chairs: Katarina Leblanc Willem Fibbe
Prospective Clinical Trials Chair: Gérard Socié
[email protected]
[email protected]
Education Committee Chair: Tamás Masszi
Quality Assessment of Autografts Chair: Francesco Lanza
[email protected]
[email protected]
JACIE Executive Committee President: Ineke Slaper-Cortenbach
Registry Committee Chair: Carmen Ruiz de Elvira
[email protected]
[email protected]
[email protected]
Vice-President: Jane Apperley
[email protected] Nuclear Accident Committee Chair: Ray Powles
[email protected]
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Statistical Committee Chair: Myriam Labopin
[email protected]
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EBMT Working Parties
Please refer to the following website address:
http://www.ebmt.org/5WorkingParties/wparties1.html
EBMT Nurses Group Board President Erik Aerts
Secretary Michelle Davies
University Hospital Zürich Dept. of Medicine 8091 Zürich SWITZERLAND Tel.: +41 44 255 2633 Fax: +41 44 255 9781
[email protected]
Adult Leukaemia Unit Christie Hospital Wilmslow Road Manchester M20 4BX UNITED KINGDOM
[email protected]
President Elect Arno Mank Academic Medical Centre Dept. Hematology Meibergreef 9, NL-1105 AZ Amsterdam NETHERLANDS Tel: +31 20 566 7905 Fax: +31 20 566 9030
[email protected]
Treasurer Joachim Blankart AK St. Georg Dept. of Hematology G3, STE Lohmuehlenstrasse, 5 20099 Hamburg GERMANY
[email protected]
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Affiliated Organisations/Websites International Organisations: ASBMT ASCO ASH BMDW EBMT EHA EORTC ESH EUROCORD IBMTR ISCT Europe ISEH NETCORD WMDA
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www.asbmt.org www.asco.org www.hematology.org www.bmdw.org www.ebmt.org www.ehaweb.org www.eortc.be www.esh.org www.eurocord.org www.ibmtr.org www.celltherapysociety.org/Related_Organizations/ ISCT_Europe/ www.iseh.org www.netcord.org www.worldmarrow.org
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Published by
Via Martin Piaggio, 17/6 16122 Genoa, Italy tel +39 10 83794229 - fax +39 10 83794260
[email protected] - www.accmed.org
© 2008 EBMT and ESH Printed by Litoprint (Genoa, Italy)
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CHAPTER 1
Stem cell transplant organisations
J. Apperley, A. Keating
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CHAPTER 1 • Transplant societies
Haematopoietic stem cell transplantation (HSCT) is a professional medical activity and like other medical specialities, lends itself to the development of groups of individuals with similar and/or overlapping interests. It is therefore not surprising that there are a number of national and international HSCT societies that carry out activities with common aims, i.e. the continuous development of their speciality and improvement in outcome for patients. Such activities usually revolve around one or more annual meetings at which new information can be presented, discussed and disseminated. The organisations may have accompanying infrastructures for education, professional standards, production of guidelines and lobbying. However there is one additional characteristic of the HSCT societies that sets them apart from many other organisations and that is the collection, analysis and reporting of outcome of all transplants performed by their member centres. Worldwide there are three international HSCT societies - European Group for Blood and Marrow Transplantation (EBMT) - Center for International Blood and Marrow Transplantation Research (CIBMTR) - Asia Pacific Blood and Marrow Transplantation (APBMT). There are several national HSCT societies in Europe (Table 1) and there are welldeveloped national groups elsewhere including those in Australia, Canada, Japan, New Zealand and North America. The largest of these is the ASBMT in the USA and their official journal is Biology of Blood and Marrow Transplantation.
Table 1: National HSCT Societies and Registries in Europe Country
E-mail address
Austrian SCT Registry
[email protected]
Czech SCT Registry
[email protected]
Societé Francaise de Greffe de Moelle et de Thérapie Cellulaire
[email protected]
Deutsches Register für Stammzelltransplantationen (DRST)
[email protected]
Italian National BMT Registry (GITMO)
[email protected]
Dutch National Registry (TYPHON)
[email protected]
Spanish Haematopoietic Transplantation Group & Cellular Therapy (GETH)
[email protected]
Swiss National Stem Cell Transplant Registry (STABMT)
[email protected]
Turkish Transplant Registry
[email protected]
British Society of Blood and Marrow Transplant
[email protected]
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Laboratory processing of haematopoietic stem cells typically involves a group of professionals with skills that are complementary to those of the average transplant physician and they are represented by the International Society of Cellular Therapy. There is clearly a considerable overlap with the interests of the physician orientated HSCT societies and indeed these professional groups work closely together, most notably in the development of an accreditation system that has established and inspects against, a series of standards for the clinical, collection and laboratory facilities, i.e. FACT-JACIE. The principles of this accreditation system will be discussed in Chapter 4 and will not be discussed in detail in this Chapter. All these societies (EBMT, CIBMTR, APBMT, ISCT) have a similar structure in which a Board of Directors (usually elected by the professional membership) provide and manage the infrastructure, which in turn permits the work of the society. Although the terminology may differ a number of committees or working groups oversee the scientific work and parallel groups of individuals focus on policy issues such as education, meeting organisation, legal and regulatory matters etc. In addition the provision of donors (bone marrow, peripheral blood derived stem cells, cord blood, cellular sub-populations) has also necessitated the development of local and national donor organisations, whose focus is rather different from the professional societies. They have established mechanisms for the recruitment, assessment, provision and follow-up of donors, and are all members of an umbrella organisation, the World Marrow Donor Association (WMDA), that oversees the development of standards and guidelines for the management of these volunteer individuals. NETCORD was developed to share best practice in cord blood procurement. In order to facilitate rapid identification of suitable HLA-matched unrelated donors for individual patients, all the donor registries submit their typing data to an organisation known as Bone Marrow Donors Worldwide (BMDW).
1. European Group for Blood and Marrow Transplantation (EBMT) 1.1. History Over the past three decades the EBMT has grown from a small group of enthusiasts to a society representing more than 500 teams from 57 countries. In 1975 teams from Paris, Leiden, London and Basel met for the first time to discuss general problems and individual patients, and to draw up protocols with a view to improving results. They continued to meet regularly in the Swiss or French Alps and the group, then called the European Cooperative Group for Bone Marrow Transplantation, attracted an ever-increasing number of participants. In 1979, the European Foundation for Bone Marrow Transplantation (EBMT) was formally established with legal status in 20
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the Netherlands. Four Working Parties were created, each with its own registry, i.e. Aplastic Anaemia, Acute Leukaemia, Inborn Errors and Transplantation Immunology. In 1989, a new constitution created the “European Group for Bone Marrow Transplantation”, the abbreviation EBMT being retained. Subsequently, the working parties were restructured and new ones added, i.e. Acute Leukaemia, Aplastic Anaemia, Autoimmune Disease, Chronic Leukaemia, Immunobiology, Inborn Errors, Infectious Diseases, Late Effects, Lymphoma, Nursing, Paediatrics and Solid Tumours. In 1995 the name of the group was changed once more to “European Group for Blood and Marrow Transplantation”, though the acronym “EBMT” again remained. 1.2. Aims The overall objective of the EBMT is to improve the outcome of HSCT by a number of connected activities. The most long-standing of these activities is the collection, validation and analysis of outcome data of all transplants performed by member teams. However the work of the group has expanded to include accreditation, education and outreach, prospective clinical trials and regulatory affairs. 1.3. Organisation EBMT members may be full members, associate members, individual members or corporate patrons. Full and associate memberships are granted to teams rather than individuals. EBMT members have the right to propose and elect office holders, stand for election, attend annual meetings and participate in the Working Parties and EBMT studies. Full membership implies a duty to report individual transplant data to the EBMT registries. The EBMT Board is composed of elected individuals including the President, Secretary, Treasurer and the Working Party Chairpersons. The Board co-ordinates EBMT activities and is responsible for the budget. The organisational structure is shown in Figure 1. The Working Parties are the heart of EBMT, performing retrospective and prospective studies relating to their particular disease or other interest. However the Board of the EBMT have long recognised the need for activities that cross Working Party activities and this has led to the development of EBMT committees. Unlike the Working Party Chairs, who are elected by the member centres and who have permanent positions on the EBMT Board, the Chairs of the committees are appointed by the Board. Committees are designed to be responsive to the needs of the organisation and can be dissolved which there is no longer any requirement for the particular activity. The journal Bone Marrow Transplantation is the official journal of the EBMT.
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Figure 1: Organisational structure of EBMT EBMT Board Working Parties
Committees
Registries
Administration
Acute Leukaemia Aplastic Anaemia Autoimmune Chronic Leukaemias Immunobiology Infectious Diseases Inborn Errors Late Effects Lymphoma Nurses group Paediatrics Solid tumours
Accreditation Clinical Trials Education Developmental Graft Engineering Nuclear Accidents Outreach Statistical
Data Managers Study Coordinators Statisticians
EBMT Secretary EBMT Treasurer Executive Secretary JACIE Office
1.4. Committees The current committees are shown in Table 2. The work of some of these committees will be discussed in more detail below. 1.4.1. Education and Outreach The Education committee was established in 1995 with the intention to provide a more structural approach to training in HSCT. Together with the European School for Haematology (ESH) they have successfully organised the annual residential training course in which senior transplant physicians and scientists interact in formal and informal settings with less experienced colleagues. This committee have also issued this ESH-EBMT transplant manual, now in its fifth edition. Most recently this committee has developed an Outreach project to assist transplant units in economically disadvantaged countries. The EBMT are not alone in this concept and are actively integrating their ideas with the ESH, and are aware of similar programmes within the American Society of Hematology (ASH), CIBMTR, the European Hematology Association (EHA) and the World Health Organisation (WHO). This project supports educational initiatives including training courses in clinical and laboratory techniques, exchange programmes, fellowships, clinical trial participation and twinning programmes.
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Table 2: EBMT Committees Sub-committee
Responsibilities
Accreditation
JACIE, audit and activity survey
Developmental
Emerging stem cell therapies for nonhaematological disorders
Education
Training courses, handbook (jointly with ESH), outreach initiatives
Nuclear accident
Establishment of network of units with haematological expertise to respond to the need for care of neutropenic patients in the event of nuclear accidents
Outreach
Dissemination of excellence in HSCT
Prospective clinical trials
Infrastructure for prospective clinical trials
Quality assessment of autografts
Standardisation of assessment of haematopoietic cell product quality
Statistics
Ensure statistical accuracy within EBMT studies. Develop new statistical tools, production of guidelines
1.4.2. Prospective Clinical Trials This committee was established in 2003 to assist the EBMT in their progress from a retrospective study organisation to a prospective clinical trials group. The introduction into European Legislation of the EU Directive on Clinical Trials (2001/20/EC) has highlighted the necessity of creating an appropriate infrastructure for the conduct of these trials. Most recently the EBMT together with the CIBMTR and the University of Central Lancashire (UCLAN) have been successful in obtaining funding from the European Commission to develop international prospective clinical trials within the transplant community, a project that has the acronym CLINT. 1.4.3. Accreditation The Accreditation committee has a number of diverse functions, which have developed over many years. The committee was originally responsible for a simple form of accreditation for allogeneic sibling transplant in which centres requested accreditation on the basis of performance of a minimum number of transplants annually. Each year, one-third of centres is chosen at random and alerted to the possibility of an audit of data and centre validation. Later, a proportion of these “at risk” centres are visited by senior transplant physicians and data are checked at source.
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One of the most valuable activities of this committee and indeed of EBMT is also one of the simplest. In 1990, Professor Alois Gratwohl began to collect information relating to the annual transplant activity of all transplant centres in Europe (including those centres that are not EBMT members). As a result the EBMT now hold important data relating to activity, geographical variation, changing trends and the development of new technologies. Activity is published annually and has proved invaluable to the funding agencies (public and private) in identifying transplant needs. More recently the bulk of the work if the accreditation committee has been the development and introduction of the FACT-JACIE accreditation programme for all transplant units. This is discussed in more detail in Chapter 4. 1.5. Data collection and data-flow Since its inception the essence of EBMT has been to evaluate and optimise the outcome of patients treated by HSCT. In order to do this it was always deemed necessary to collect patient data, and so the EBMT registry was formed. As of today the registry holds information on almost 300,000 transplants performed by the member teams, and this has proved an invaluable resource in the analysis of factors affecting transplant outcome, in identifying complications and their optimal treatment, in determining trends in management and in evaluating new technologies. Member teams are required to report all their transplant activity. “Minimal Essential Data” (MED) are collected on a number of different forms. MED-A data form the basis of the registry. Relatively simple and limited data collected in this way allow the analyses of overall survival, disease free survival, transplant related mortality, and relapse risk. These data are identical to those collected by the CIBMTR on their Transplant Essential Data (TED) form, and the facility exists to allow cross reporting to both registries. More detailed information is collected on MED-B forms which relate to the specific disease and to the procedure (allograft or autograft). The submission of these data by members is voluntary as opposed to MED A data which are obligatory. Information relating to specific studies is collected via MED-C forms. Until recently the term “form” literally referred to hard copy in virtually all cases. Paper forms were sent to the EBMT and transcribed onto a central electronic database, but the complexity of the programme and the relative lack of familiarity with computers at the time precluded widespread use of this approach. The EBMT has now developed an on-line Internet based reporting system, Project Manager Internet Server or ProMISe. Data reported in this way are received simultaneously by the EBMT registries in Genoa, London, Leiden and Paris but can also be downloaded for local use. This system has revolutionised data reporting with increasing numbers of centres adopting ProMISe. In 2006, 458 users from 364 centres accessed the database to enter data for more than 25,000 transplants. 24
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CHAPTER 1 • Transplant societies
The EBMT has welcomed the development of national transplant societies and has understood the desire of some of these groups to have their own national databases. In the past, for member centres in countries with national registries, data flowed from the centre to the national registry and then to EBMT. Centres in countries without a national registry reported directly to EBMT. Now if the centre is utilising ProMISe, the data are deposited simultaneously in the national and EBMT registries.
2. Center for International Blood and Marrow Transplantation Research (CIBMTR) 2.1. History The CIBMTR is a relatively new organisation having developed in 2004 from an affiliation of the International Blood and Marrow Transplant Registry (IBMTR), a sister organisation of EBMT based at the Medical College of Wisconsin, and the research interests of the US National Marrow Donor Program (NMDP). The IBMTR was established in 1972 to link transplant centres internationally and to collect data derived from allogeneic transplants for the purposes of analysis and reporting. As autografting became more extensively used these transplants were added to the IBMTR databases but collection was confined to procedures performed in North and South America. The NMDP was established in 1987 to facilitate the search, identification, and procurement of volunteer unrelated donor haematopoietic stem cells. At that time the NMDP also had the remit to collect and analyse data emanating from these alternative donor transplants, a task which has now been subsumed into the activities of CIBMTR. The NMDP, like several of the unrelated donor registries, also maintains a tissue biobank of donor and recipient samples. The CIBMTR now has 459 members in 52 countries. Based on data collected in the Centers for Disease Control Hospital Surveys and the US Government Accounting Office and worldwide surveys of transplant activity, approximately 40% of allogeneic transplants worldwide and about 50% of autografts performed in North and South America are registered with the CIBMTR. 2.2. Aims The CIBMTR is a clinical research programme whose major mission is to provide a resource of data and statistical expertise to the HSCT community. They aim to define key areas for future research, secure funding for research, design and implement clinical studies and make available their resources including the clinical database and the tissue bank.
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2.3. Organisation The organisational structure of the CIBMTR is shown in Figure 2. The Affiliation Board and Executive Director have managerial oversight. The Chief Scientific Director has primary responsibility for administrative and scientific operations. CIBMTR Scientific Working, Executive and Advisory Committees provide policy and scientific oversight for this work. The activities of the CIBMTR are funded primarily by grants and contracts from the US government. The CIBMTR Assembly is the voting membership, comprised of a single representative from each CIBMTR Research Center. The CIBMTR Advisory Committee meets biannually to review scientific and other activities of the CIBMTR. The CIBMTR Executive Committee is a subcommittee of the Advisory Committee that provides ongoing advice and counsel to the CIBMTR Statistical Center between meetings of the Advisory Committee.
Figure 2: Organisational structure of CIBMTR
NMDP - Research
Affiliation Committee
Assembly
Executive Director
Advisory Committee
Executive Committee
MCW - IBMTR
Senior Research Advisor
Chief Scientific Director Working/Steering Committees Statistical Methodology
Clinical Trials Support
BMT CTN*
Immunobiology
RCI BMT
Observational Research
Clinical Outcomes
Health Policy
* The BMT CTN Data and Coordinating Center is a Collaboration of CIBMTR, NMDP and the EMMES Corporation; NMDP: National Marrow Donor Program; IBMTR: International Bone Marrow Transplant Registry; MCW: Medical College of Wisconsin; BMT CTN: Blood and Marrow Transplant Clinical Trials Network; RCI BMT: Resource of Clinical Investigations in Blood and Marrow Transplantation 26
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2.4. Committees The Scientific Working Committees are analogous to the EBMT Working Parties and set priorities for observational studies. There are 17 Working Committees: Acute Leukaemia; Chronic Leukaemia; Lymphoma; Plasma Cell Disorders; Solid Tumours; Paediatric Cancer; Non-malignant Marrow Disorders; Immune Deficiencies/Inborn Errors of Metabolism; Autoimmune Diseases; Graft Sources and Manipulation; Graft versus Host Disease; Late Effects and Quality of Life; Immunobiology; Infection and Immune Reconstitution; Regimen-related Toxicity and Supportive Care; Health Services and Psychosocial Issues; Donor Health and Safety. Membership on CIBMTR Working Committees is open to anyone willing to take an active role in studies using CIBMTR data and/or resources. The CIBMTR also functions as the Data and Coordinating Centre of the US Blood and Marrow Transplant Clinical Trials Network (BMT CTN). The BMT CTN was established by the National Heart, Lung, and Blood Institute (NHLBI) and the National Cancer Institute (NCI) in order to design, develop, and execute prospective clinical trials to improve the safety, applicability, and efficacy of HSCT. Sixteen core centres were awarded cooperative agreements with the NHLBI/NCI and a broader network of over 60 transplant centres have participated in eight prospective trials enrolling over 1,500 patients in the first four years of activity. 2.5. Data collection The CIBMTR collects data at two levels. Registration data (Transplant Essential Data–TED) are identical to the Med-A data of the EBMT and the facility exists to allow cross reporting to both registries. All CIBMTR teams contribute registration data. Research data are collected on subsets of registered patients and includes comprehensive pre- and post-transplant clinical information.
3. The Asia Pacific Blood and Marrow Transplantation (APBMT) The Asia Pacific Blood and Marrow Transplantation (APBMT) group is an organisation, akin to EBMT and CIBMTR, which allows HSCT physicians in Asian countries to share their experience and develop co-operative studies. The APBMT group was established in 1990 and initial meetings were held in China and Japan. Since 1994 annual meetings have been an integral part of the organisation. The APBMT aims to promote all aspects associated with HSCT including basic and clinical research. A registry to collect the results of HSCT performed in member centres began in 2006.
4. Worldwide Network for Blood and Marrow Transplantation (WBMT) In 2007 the concept developed of an organisation that would unite the three
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international HSCT societies (EBMT, CIBMTR and APBMT) together with the WMDA with the aim of unifying data collection and reporting systems, developing harmonised guidelines, providing an effective professional lobby to the various national and international regulatory agencies and strengthening the research database. The name Worldwide Network for Blood and Marrow Transplantation was adopted and the constitution is currently in development.
5. International Society for Cellular Therapy (ISCT) 5.1. History ISCT was established in 2002 as a professional organisation for those working or interested in cell-based research, processing, manipulation, and clinical translation. It evolved from the US and European branches of the International Society for Hematotherapy and Graft Engineering (ISHAGE) when the change in name and focus was considered essential to encompass the use of cellular therapy for a wider range of diseases and applications. ISHAGE had itself been established in 1991 when it had become increasingly clear that an organisation was required to represent the interests and needs of those involved in graft manipulation. 5.2. Aims The mission of ISCT is to provide a global forum for the development and validation of standardised technology and for representation of the membership to other professional organisations and regulatory and governmental bodies. They also have a strong commitment to education and training. 5.3. Organisation In contrast to the other HSCT organisations, the membership of the ISCT is largely composed of individuals although there are arrangements for laboratory and corporate membership. In 2007, the Society had over 1200 members from 6 continents. Leadership of the ISCT is provided by an Executive Committee which manages the affairs of the Society and oversees the work of the committees and working groups. An advisory committee, composed largely of previous elected officers, provides input from the membership to the Executive Committee and also advise on the long-term strategy of the Society. ISCT is composed of several standing committees, representing all facets of ongoing activities of the Society, and short-term working groups, designed to address immediate concerns within cell therapy, drafting position statements and guidance for broad dissemination or submission to the FDA. The current committees and groups are shown in Table 3. 28
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Table 3: Working Parties and Standing Committees of ISCT Working Parties
Standing Committees
- Microbial testing - Storage & transportation - Devices - Facility sanitisation - Homologous use - Release testing
- Cell therapy commercialisation - Cell and tissue evaluation - Education and publication - Gene therapy - Haematopoietic stem cells - Immunotherapy & dendritic cells - Laboratory practices - Regulatory affairs - Europe - Regulatory affairs - America - Regulatory affairs - Asia - Mesenchymal and tissue stem cells
ISCT organises an annual meeting of diverse groups of investigators, practitioners, and technologists with a focus on research and education. Cytotherapy is the official journal of ISCT.
6. World Marrow Donor Association (WMDA) 6.1. History The WMDA was created in 1994 to address obstacles faced when transplantation involved donors and recipients in different countries. It is a voluntary organisation of representatives of HSCT donor registries, cord blood banks, and other institutions and individuals involved in the use of unrelated donors. Currently the WMDA represents 11.8 million donors in 67 stem cell registries and 292,000 units in 47 cord blood banks from 48 countries. The individual donor registries are too numerous to mention here but include the National Marrow Donor Program (NMDP) in the United States, the Anthony Nolan Trust in the UK and the German. 6.2. Aims To improve and simplify stem cell donation for donors and patients. 6.3. Organisation (Figure 3) The membership of the WMDA is comprised of both donor (blood, marrow and cord blood) organisations and individuals. Much of the work of the WMDA is done through its working groups (Figure 3), which are focussed on developing best practice in the medical, ethical, technical, quality and financial activities of international HCT as they relate to volunteer donors. Importantly, the WMDA established an
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Figure 3: Organisational structure of WMDA President Vice Presidents Past President Chief President Elect President Emeritus North & South America Operating Asia, Australia, Officer Secretary General Pacific Islands, Europe Treasurer
Working Groups Clinical Registries Information Technology Ethics Quality Assurance
Accreditation Committee
Secretaries North & South America Asia, Australia, Pacific Islands, Europe
WMDA Administrative Office Leiden, NL
accreditation program for donor registries in 2003, to certify adherence to WMDA standards for registry operation. These standards cover the general organisation of the registry, donor recruitment and consent procedures, donor characterisation and evaluation, information technology, facilitation of search requests, second and subsequent donations, graft procurement and transport, assessment of donor and transplant outcomes and financial and legal liabilities. The standards are exacting and to date 12 registries have received accreditation. The aim is clearly to extend the process to all their registry members.
7. Alliance for harmonisation of cellular therapy accreditation (AHCTA) The process of HSCT has recently come under extensive scrutiny by national and international regulatory agencies and this has resulted in increasing stringency in the legislation regarding the procurement and use of stem cells. In 2006 it became clear that there was a real risk that regulations could be introduced nationally that would conflict with processes elsewhere in the world. This is particularly relevant to the requirements for import and export of cells. As it is common practice in alternative donor transplantation to use stem cells collected in one country for a patient treated in another country there was some urgency 30
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to present our concerns in a unified manner. Furthermore there was clearly a need for the relevant parties to agree common policies, procedures and protocols. AHCTA was developed to harmonise the standards of all the involved organisations (Figure 4) with the objective of creating a single set of quality, safety and professional requirements for cellular therapy. These standards will comprehensively cover all aspects of the process from assessment of donor eligibility to transplantation and clinical outcome. AHCTA regards regulatory authorities as partners in the application of these global standards, essential to their successful adoption and will endeavour to inform and support these authorities in the area of cellular therapy regulation.
Figure 4: Membership of AHCTA European Federation for Immunogenetics
European Group for Blood & Marrow Transplantation
Foundation for the Accreditation of Cellular Therapy
Joint Accreditation Committee ISCT-EBMT
International NETCORD Foundation
World Marrow Donor Association
International Society for Cellular Therapy (Europe)
8. Bone Marrow Donors Worldwide (BMDW) 8.1. History BMDW began as an initiative of the Immunobiology Working Party of the EBMT in 1988. It represents a voluntary collaborative effort of the stem cell donor registries and cord blood banks to provide centralised information on the HLA phenotypes and other relevant data of unrelated stem cell donors and cord blood units and to make this information easily accessible to the physicians of patients in need of HSCT. The first edition of a publication containing the donor files of eight registries with a total of 155,000 volunteer stem cell donors was
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issued in 1988. The current number of donors and cord blood units in the BMDW database is almost 12 million! 8.2. Aims The original goal was to collect the HLA phenotypes of volunteer stem cell donors and cord blood units, and to maximise the chance of finding a stem cell donor or cord blood unit by providing access to all stem cell donors and cord blood units available in the world. 8.3. Organisation The BMDW Editorial Board consists of one representative of each stem cell donor registry or cord blood bank participating in BMDW, and meets twice a year to discuss achievements and necessary improvements. The office of BMDW remains in Leiden in the Netherlands and the service is provided and managed by Europdonor Foundation. The HLA types and matching programmes are available online. More recently programmes for mismatched donors and cord blood donors have been added together with a haplotype frequency analysis on phenotype data submitted by the participating registries. Only HSCT professionals who have received authorisation and participating registries can access these services. A username and password are required, and a secure protocol has been implemented to encrypt all transmitted data.
9. NETCORD 9.1. History NETCORD is a global non-profit organisation, which was established in 1998 to connect the existing cord blood banks in order to share best practice in this rapidly evolving field. 9.2. Aims The goal of the organisation is to maintain compliance with quality standards so as to ensure uniformity and quality assurance of the cord blood units supplied by their members. In addition they provide access for registered transplant units to a centralised web-based search process, support research to constantly improve the processes of procurement and processing and develop educational programmes to increase the donor pool and the subsequent use of the products. 9.3. Organisation Member cord blood banks are expected to have inventories of at least 1000 cord 32
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CHAPTER 1 • Transplant societies
blood units of documented high quality. Thus far, NETCORD has 23 members and as of late 2007 has an inventory of 153,830 units, an increase of nearly 30,000 units or 23% in the past 12 months. NETCORD has facilitated the use of over 3,000 cord blood units worldwide both for children and adults. In order to ensure high and uniform quality of all cord blood (CB) units, NETCORD established quality standards in collaboration with FACT for the collection, cryopreservation, storage and release of units. All NETCORD banks are expected to obtain accreditation by NETCORD-FACT and to comply with their quality standards. To facilitate direct searches for CB units by transplant centres and to avoid allocation conflicts for patients, NETCORD developed the Virtual Office (VO), which contains a real-time search programme for compatible and available units. Details include high resolution HLA typing, volume, cell number and infectious disease markers. By submitting a search request to the VO, transplant centres no longer have to search multiple databases of the different cord blood banks, but receive a single unified search report on all acceptable units that are contained in the inventory. The collection of clinical outcome data is coordinated by CIBMTR or Eurocord. The Eurocord registry has operated on behalf of EBMT since 1995. Thanks to the NETCORD-Eurocord collaboration, 3286 cord blood transplantations have been reported to the Eurocord registry from 177 EBMT (64% of cases) and 187 non-EBMT (36% of cases) centers from 43 European and nonEuropean countries. In summary, the intense activity within all these societies reflects the on-going enthusiasm for HSCT and cellular therapy. Further information can be obtained from the specific websites (Table 4).
Table 4: Web addresses of transplant related organisations EBMT
www.ebmt.org
ESH
www.esh.org
CIBMTR
www.cibmtr.org
APBMT
www.apbmt.org
ISCT
www.celltherapysociety.org
WMDA
www.worldmarrow.org
AHCTA
www.ahcta.org
BMDW
www.bmdw.org
NETCORD
www.netcord.org
Eurocord
www.eurocord.org
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*
CHAPTER 2
Biological properties of haematopoietic stem cells
A. Wodnar-Filipowicz
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1. Introduction Until a few years ago, an interest in stem cells, mostly of haematopoietic origin, was limited to a relatively small representation of scientists and clinicians seeking to understand the role of these rare cells in tissue homeostasis and to utilise their remarkable potential to regenerate an adult tissue following transplantation. Over the last few years, stem cells have captured the imagination of scientists, clinicians and the lay public alike with the promise of representing a future remedy for the major degenerative diseases of our civilisation and even dysfunctions associated with normal ageing. The progress in technologies to detect and enumerate stem cells in vivo led to discovery of stem cells residing in most mammalian tissues, contributing to their generation, homeostasis and probably repair. A major breakthrough has been achieved in the development of methods to propagate human embryonic stem (ES) cells in culture and to drive their in vitro differentiation into specialised human tissues. The concept of cancer stem cells has emerged, as cells responsible for generation and persistence of tumours. Here we discuss the stem cell field by summarising the current knowledge of the phenotype of these cells, their interactions with the microenvironment in which they reside, and the mechanisms regulating their functions. The central place will be taken by haematopoietic stem cells (HSCs), representing the first-discovered, the best-understood and, at present, the only clinically-applicable population of stem cells. We also summarise the current state of knowledge on the functional plasticity of somatic tissue stem cells and on human ES cells. The therapeutic relevance gained from the basic research findings will be emphasised.
2. Stem cell definition Stem cells are defined as a population of undifferentiated cells with the capacity to divide for indefinite periods, to self-renew and to generate a functional progeny of highly specialised cells. This common definition includes cells present in different physical locations and having fundamentally different proliferative properties and functions (Table 1). A fertilised egg (zygote) represents a totipotent stem cell, a cell with unrestricted differentiation potential and the only cell with the capacity to give rise to all cells necessary for the development of foetal and adult organs. ES cells forming a cluster of cells inside the blastocyst are pluripotent stem cells, capable of generating a variety of specialised cell types, but limited in their differentiation potential by the inability to support the development of a foetus. Further specialisation results in generation of multipotent stem cells residing in adult somatic tissues. Their physiological functions are to replenish mature cell populations of the given tissue
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Table 1: Definition of stem cell types Stem cell
Developmental properties
Fertilised egg
- Totipotent - Unrestricted differentiation potential
ES cells
- Pluripotent - Give rise to a variety of specialised cell types
Adult somatic stem cells
- Multipotent - Limited to specific tissues
or organ, and to respond to stress by repairing the damage. HSCs represent the prototype of multipotent adult tissue stem cells (1). In humans, HSCs can be found in cord blood as a result of stem cell migratory properties during foetal development, whereas post-natally, the only organ harbouring HSCs and pursuing active multilineage haematopoiesis is the bone marrow.
3. Characteristics of stem cells in the bone marrow More than 40 years of research on bone marrow-derived stem cells, initiated in the 1960s by Till and McCulloch, marked an ongoing improvement in methods to quantitate and isolate these cells. Assays for clonogenic precursors of the myeloerythroid lineages in vitro, defined as long term culture-initiating cells (LTCICs) and committed colony forming units (CFUs) were followed by the development of a model of immunocompromised non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, which allows the study of the repopulating ability of human haematopoietic cells in vivo (2). These functional assays are paralleled by progress in the phenotypic characterisation of haematopoietic cells by flowcytometry, owing to monoclonal antibodies specifically recognizing cell surface molecules (Figure 1). The phenotypic properties of murine HSCs have been precisely defined as cells devoid of lineage markers and expressing the stem cell antigen (sca1) and the receptor c-kit. The Lin-sca1+c-kit+ (LSK) cell population has self-renewing and long-term repopulating activity in vivo. Other cell surface markers defining the HSC compartment in mice include the tie2 and flt3 receptors and CD150. Characteristically, c-kit, flt3 and tie2 function as receptors for early-acting haematopoietic growth factors: stem cell factor, flt3 ligand and angiopoietin, which act as key positive regulators of haematopoiesis (3). The most primitive human HSCs express CD34 and lack CD38 cell surface antigens and have the capacity to reconstitute a sublethally irradiated NOD/SCID host. The CD34+CD38- HSC compartment, which constitutes ±0.1% of bone marrow cells, is 36
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Figure 1: The immunophenotypic characteristics of human and mouse long-term repopulating haematopoietic stem cells (LT-HSC)
Human HSC
Mouse HSC
LinCD34+ CD38c-kit+ CD133+ SP
Linc-kit+ sca1+ Tie2+ CD150+ CD244CD48SP
LT-HSC
Lin: lineage; SP: side population cells
heterogeneous and contains also c-kit-, flt3-, and CD133-expressing cells. Both mouse and human HSCs are present among the side population (SP) cells which express the drug transporter protein Abcg2 and therefore have the ability to actively efflux the DNA-intercalating dye Hoechst. Human HSCs remain at present less well defined than murine HSCs. Despite the availability of methods which greatly facilitate and enhance the precision of studies with well defined cell populations, it is most likely that pure human HSCs have not yet reached the hands of the scientists. Even less defined remain the properties of stem cells from tissues other than bone marrow, primarily because the isolation of stem cells of skin, muscle, brain or liver remains difficult. Nevertheless, the flow cytometry-based characterisation of somatic tissue stem cells indicates that several cell surface markers are shared, including CD34, c-kit, sca1 and CD133, underlying common features of these rare and not easilyaccessible stem cell populations. Haematopoiesis occurs in close physical contact with stroma lining the bone marrow niches. Recently, much attention has focused on the differentiation properties of stroma cells themselves. A marrow stromal cell population, termed mesenchymal stem cells, has been shown to give rise to numerous tissue types, including cartilage, bone, fat, and muscle (4). In addition, a minor population of adherent multipotent adult progenitor cells (MAPCs) was found to be capable of differentiation into functional hepatocytes, endothelial cells, skeletal myeloblasts, osteoblasts, chondrocytes and, importantly, haematopoietic cells (5). This suggests the capacity of the bone marrow for multilineage tissue regeneration is present in
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both haematopoietic and non-haematopoietic stem cell populations. The therapeutic implications of non-haematopoietic bone marrow cells have already been clinically tested in the treatment of osteogenesis imperfecta in transplanted children (6) (see also Chapter 34). Improvement of clinical parameters suggests that bone marrow derived mesenchymal progenitor cells carry a potential for repair of bone and cartilage tissue.
4. Stem cell niches The concept of the stem cell niche defines a microenvironment, where important interactions between adhesion molecules and their ligands, and between cytokines, chemokines and their corresponding receptors control the fate of stem cells and their progeny (7). In the adult bone marrow, HSCs are located in the trabecular endosteum, where osteoblastic cells are critical components sustaining the quiescence or selfrenewal of HSCs, the properties essential for long-term haematopoiesis (8). Both intrinsic and extrinsic mechanisms define the state of either quiescence or cycling and differentiation of HSCs (Figure 2). The intrinsic mechanisms include transcription Figure 2: The mechanisms regulating the HSC niche
Extrinsic mechanisms PTH
nerves Osteoblast
Cadherins VCAM - VLA ICAM - LFA SDF-1 - CXCR4
cell-cell adhesion migration
quiescence self-renewal expansion
SCF - c-kit Flt3-L - Flt3 Ang-1 - tie2 Jagged/Delta - Notch Wnt - Frizzled
Signal transduction Gene expression HSC
Intrinsic mechanisms
A graphical representation of extrinsic and intrinsic mechanisms in the niche based on the functional interaction of the parathyroid hormone (PTH) and nervous system with osteoblasts, the homing of HSCs in response to chemokines (SDF-1), the physical interactions involving the cadherins and integrins (VLA), and regulation of HSC quiescence, self-renewal and expansion by cytokines: stem cell factor (SCF), Flt3 ligand (Flt3-L), angiopoietin-1 (ang-1), notch ligands (jagged and delta) and wnt ligands, followed by signaling downstream from the cognate receptors and initiating the gene expression 38
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factors and epigenetic regulators acting through chromatin remodelling. The extrinsic mechanisms are dictated by the environment of stromal and osteoblastic cells. Chemokines are responsible for HSC homing into the niche. The direct physical interaction between HSCs and the niche cells are mediated by adhesion molecules, such as integrins and cadherins. The membrane-bound and locally secreted cytokines define the HSC fate by initiating specific signalling pathways within the cell. The most prominent examples are stem cell factor, flt3 ligand, angiopoietin, Notch ligands and wnt ligands, which act synergistically. Also ex vivo, these cytokines allow an expansion of HSC numbers. According to the most recent findings, the extrinsic environmental cues in the HSC niche include hormonal regulators, such as parathyroid hormone, and the influence of a sympathetic nervous system, both signalling through the osteoblastic niche component, as well as the regulation by oxidative conditions in the niche. The information on the structure and cellular composition of the niche is only beginning to be revealed, and particularly in the human system the knowledge is only just emerging.
5. Leukaemic stem cells The available data suggest that leukaemia is a stem cell disease, in which the stem cell self-renewal mechanisms are preserved but the tight growth control is lost due to malignant transformation. As an additional mechanism, the oncogenic events might be imposing the self-renewal capacity at the level of committed progenitor cells (9). Consequently, leukaemic stem cells (LSCs) share many molecular mechanisms that regulate the function of normal HSCs. At present, phenotypic features specific for LSCs are not defined. LSCs are thought to reside within the CD34+CD38- cell population, which contains transplantable cells giving rise to human leukaemia in NOD/SCID mice. HSC-characteristic adhesion molecules, such as integrins, are involved in LSC interaction with stroma. Hence, dissimilarities between normal and leukaemic cells most likely have their origin in differences in intracellular signal transduction pathways (10). As an example, oncogenic lesions in the cytokine receptors, c-kit and flt3, are responsible for constitutive activation of downstream signalling in cells at the earliest stages of haematopoietic differentiation, resulting in HSC to LSC transition. A better understanding of the properties of LSCs is of major therapeutic relevance for the design of LSC-targeted therapies (Figure 3). Conventional chemotherapy-based treatment of leukaemia, and cancer in general, is primarily directed against the bulk of malignant cells, and thus does not eliminate the abnormal stem cells. These cells are the origin of cancer recurrence and are responsible for relapse. Current efforts are focussed on the development of methodology to isolate these cells to better
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Figure 3: The therapeutic relevance of LSCs
Conventional cancer therapy
Specific cancer stem cell therapy
Leukaemia
Leukaemia relapse
Leukaemia regression
Leukaemic blasts are depicted in grey, and LSCs in blue
homogeneity, and to dissect the differences in molecular mechanisms used by normal HSC and LSC for their self-renewal and interaction with the microenvironment in the bone marrow (11).
6. Embryonic stem cells Human ES cells can be isolated from the blastocyst 4–5 days after fertilisation, and cultured in vitro to give rise to immortalised cell lines. Depending on the culture conditions, differentiation into cells bearing characteristics of various somatic tissue types including haematopoietic, neural, muscle and other tissues, can be achieved. This work, initiated in 1998 by Thompson et al. (12) who defined methods to isolate and propagate human ES cells from the fertilised oocyte at the 30 cell stage, is of significant value for studies on human developmental biology. Importantly, in vitro cultured human ES cells represent valuable tools for drug screening. The most publicised and controversial aspect of ES-related research is associated with the origin of these cells, being a donated surplus human blastocyst from in vitro fertilisation procedures. Destruction of human embryos in order to obtain the ES cells has been of serious ethical concern. Novel findings describe derivation of human ES cell lines from single blastomeres at the 8-cell stage, without embryo destruction. ES cells are the subject of intense research which aims at understanding the molecular basis of “stemness”. In parallel, cellular biology techniques have been
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developed which allow manipulations such as nuclear transfer from a somatic cell to the enucleated oocyte, and the further generation of ES cell lines bearing genetic information defined by the donated nucleus. The multilineage developmental potential of human ES cells opens new therapeutic avenues for the restoration of damaged or diseased tissue. The possibility to provide these cells with genetic information from the patient by nuclear transfer, is an approach which – in the future – might yield transplantable tissue of a full immunological compatibility. Ultimately, the choice of an approach will require the consideration of advantages and disadvantages associated with the cell source for transplantation (Table 2).
Table 2: Potential therapeutic value of ES cells and adult somatic stem cells Stem cell source
Advantages
Disadvantages
ES cells
- perfect plasticity - easy to programme - stable in culture - no risk of infection transmission
- ethical concern - limited source - rejection danger - carcinogenicity?
Adult somatic stem cells
- accessible source (HSCs) - no rejection (autologous) - no ethical concern - no carcinogenicity
- less/no plasticity?
7. Stem cell plasticity The term “adult stem cell plasticity” defines the ability of tissue-specific stem cells to acquire, under certain microenviromental conditions, the fate of cell types different from the tissue of origin and belonging to all three germ layers, i.e. similar to the differentiation ability of ES cells. Traditionally, the development of adult stem cells has been depicted along a well-defined path of a linear and irreversible progression concluding in terminally differentiated cell types. Furthermore, the differentiation and regenerative potential of adult stem cells has been regarded as restricted to tissues in which they reside. These traditional concepts have been challenged in the recent years by numerous studies performed by transplantation of stem cells derived from bone marrow and other organs, and which demonstrated the presence of cells of interest in tissues other than those in which they normally reside. The findings from murine studies, which suggested that stem cells may be recruited out of a circulation and engaged in regeneration of diverse tissues at distal sites, have initiated a search for “unusual” locations of donor-derived stem cells
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in patients receiving organ transplants. In some cases transitions have been documented, and are likely to reflect a healing response by cells summoned to the site of injury and instructed by the local environment of the damaged tissue. However, donor-host cell fusions rather than functional stem cell plasticity may represent the underlying mechanism (13). This concept of plasticity of somatic tissue stem cells has a potential clinical impact and may revolutionise tissue transplantation therapies and regenerative medicine. According to this novel view, at least a subset of stem cells may alter their fate in a manner that is more plastic and dynamic then previously thought, causing a fascination with these cells that has spread to nearly all clinical disciplines. There are more questions raised then answers. Following a phase of excitement and rapid accumulation of results in favour of stem cell plasticity, research in this field is now going through a less spectacular phase of verification of the existing data with refined techniques. It is too soon to discard the basic paradigm of developmental biology of the mesodermal, endodermal and ectodermal germ layer origin of mammalian organs, but a need for possible revisions to the unidirectional view of cell fate in post-embryonic development may arise.
8. Conclusions and future perspectives The ultimate goal for regenerative medicine is to channel the multipotent and/or pluripotent stem cells with high proliferative capacity into specified differentiation programs within the body for a multitude of therapeutic uses. These envisaged uses may include the generation of neurons for treatment of Alzheimer’s disease, Parkinson’s disease or spinal cord injuries, the generation of insulin-secreting pancreatic cells for the treatment of diabetes, or the generation of heart muscle cells for treatment of congenital disorders or heart attacks. Recent major technical advancements in the isolation, expansion and controlled differentiation of human ES cells and adult stem cells from at least some tissues, and additionally, the establishment of nuclear transfer techniques from the somatic cell to an enucleated donor oocyte opened a number of potential new therapeutic approaches for the restoration of damaged or diseased tissue. The new challenge in stem cell biology is related to understanding the molecular and the functional programmes of leukaemic versus normal stem cells. The principle of self-renewal and lineage-making decisions of stem cells of different tissue-origin requires understanding in molecular terms. Gene expression profiles bring evidence of the overlapping genetic programs of ES cells, haematopoietic and other adult tissue stem cells, both normal and transformed (14). Determining how epigenetic features relate to the transcriptional signatures of ES and various types 42
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of adult stem cells, is a new key challenge for the future (15). Understanding of the stem cell niche is essential for advancing the approaches to control developmental pathways by both cell-autonomous and microenvironmental cues. All this information will be used as guidance for specifically targeting the fate of normal and malignant stem cells in clinical settings.
Acknowledgments This work has been supported by the Swiss National Science Foundation grant 3100-110511.
References 1. Bryder D, Rossi DJ, Weissman IL. Hematopoietic stem cells: The paradigmatic tissue-specific stem cell. Am J Pathol 2006; 169: 338-346. 2. Bhatia M, et al. A newly discovered class of human hematopoietic cells with SCIDrepopulating activity. Nat Med 1998; 4: 1038-1045. 3. Arai F, SudaT. Maintenance of quiescent hematopoietic stem cells in the osteoblastic niche. Ann NY Acad Sci 2007; 1106: 41-53. 4. Pittenger MF, et al. Multilineage potential of adult human mesenchymal stem cells. Science 1999; 284: 143-147. 5. Jiang Y, et al. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 2002; 418: 41-49. 6. Horwitz EM, et al. Clinical responses to bone marrow transplantation in children with severe osteogenesis imperfecta. Blood 2001; 97: 1227-1231. 7. Scadden DT. The stem-cell niche as an entity of action. Nature 2006; 441: 1075-1079. 8. Wilson A, Trumpp A. Bone-marrow haematopoietic-stem-cell niches. Nat Rev Imunol 2006; 6: 93-106. 9. Huntly BJ, Gilliland DG. Leukaemia stem cells and the evolution of cancer-stem-cell research. Nat Rev Cancer 2005; 5: 311-321. 10.Rizo A, et al. Signaling pathways in self-renewing hematopoietic and leukemic stem cells: Do all stem cells need a niche? Hum Mol Genet 2006; 15 Spec No 2: R210-9. 11.Clarke MF, et al. Cancer Stem Cells – Perspectives on Current Status and Future Directions: AACR Workshop on Cancer Stem Cells. Cancer Res 2006; 66: 9339-9344. 12.Thomson JA, et al. Embryonic stem cell lines derived from human blastocysts. Science 1998; 282: 1145-1147. 13.Serafini M, Verfaillie CM. Pluripotency in adult stem cells: State of the art. Semin Reprod Med 2006; 24: 379-388. 14.Forsberg EC, Bhattacharya D, Weissman IL. Hematopoietic stem cells: Expression profiling and beyond. Stem Cell Rev 2006; 2: 23-30. 15.Spivakov M, Fisher AG. Epigenetic signatures of stem-cell identity. Nat Rev Genet 2007; 8: 263-271.
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Haematopoiesis takes place in the following adult human organs: a) Bone marrow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Peripheral blood . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Spleen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Liver . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. The stem cell compartment in the bone marrow consists of: a) Clonogenic CFU and LTC-IC progenitors only . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Haematopoietic, mesenchymal and endothelial cell progenitors . . . . . . . . . . c) NOD/SCID repopulating cells only . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Haematopoietic, liver and neural stem cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Cell surface antigen CD34 is expressed on: a) Long-term repopulating haematopoietic stem cells . . . . . . . . . . . . . . . . . . . . . . . . b) Short-term repopulating haematopoietic stem cells . . . . . . . . . . . . . . . . . . . . . . . . c) Haematopoietic and non-haematopoietic stem and progenitor cells . . . . . . d) Lineage-committed progenitors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Embryonic stem cells are characterised by: a) Lineage-restricted differentiation potential . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Potential to become a variety of specialised cell types . . . . . . . . . . . . . . . . . . . . c) Ability to generate placenta . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Ability to form a blastocyt . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Osteoblast components of the stem cell niche are involved in: a) Supporting stem cell self-renewal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Supporting the stem cell differentiation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Inhibiting the bone marrow stroma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Promoting the stem cell exit to the peripheral blood . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 3
Immunogenetics of allogeneic HSCT
*
3.1
The role of HLA in HSCT J.M. Tiercy
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CHAPTER 3.1 • HLA and donor matching
1. Introduction Tissue compatibility is determined by genes of the major histocompatibility complex (MHC), known as the HLA system in man, that are clustered on the short arm of chromosome 6. The HLA region is a multigenic system that encodes structurally homologous cell surface glycoproteins characterised by a high degree of allelic polymorphism in human populations. Immune responses against incompatible HLA antigens represent a major barrier to haematopoietic stem cell transplantation (HSCT). The accuracy of histocompatibility testing and matching criteria will therefore have important consequences on transplant outcome. This is particularly true in the case of transplantation with HSC from unrelated donors, where serologically hidden incompatibilities may account for the increased rate of post-transplant complications.
2. HLA antigens The homologous HLA Class I (HLA-A, -B, -C) and Class II (HLA-DR, -DQ, -DP) antigens are codominantly expressed and differ in their structure (Figure 1), tissue distribution and characteristics in peptide presentation to T-cells (1). The biological function of HLA molecules is to present peptide antigens to T-cells, thereby playing a central role in T-cell-mediated adaptive immunity. HLA Class I molecules, which are expressed on most nucleated cells, are composed of an a-chain (encoded in the MHC) non covalently associated with b2-microglobulin (encoded on chromosome 15) (Figure 1). The two outermost a1 and a2 domains of the heavy chain form the peptide binding site. Peptides (usually of 8–10 amino acids) presented by Class I
Figure 1: Schematic representation of HLA Class I and Class II molecules
The two polymorphic domains a1/a2 of HLA Class I are encoded by exons 2 and 3, respectively, and exon 3 codes for the most proximal a3-domain that interacts with b2-microglobulin (b2m) and CD8. The HLA Class I b-chain is non covalently associated with b2m. The Class II molecules are heterodimers composed of an a and a b-chain. The most distal domains of each of the two chains form the peptidebinding site
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molecules are derived from proteolytic degradation of cytoplasmic proteins by the proteasome. These are transported across the endoplasmic reticulum where they bind to Class I antigens. Pathogen-derived peptides presented to Class I antigens are usually recognised by CD8+ cytotoxic T-lymphocytes (CTL) (1). HLA Class II antigens are expressed on a subset of cells of the immune system comprising dendritic cells, B cells, activated T-cells, macrophages, collectively referred to as antigen presenting cells (APC). They are heterodimers composed of the two membrane-bound a- and b-chains that are encoded by two genes that colocalise in the MHC. The peptide-binding pocket is formed by the most distal domains of the two chains. Extracellular antigens internalised by endocytosis/phagocytosis are degraded in an endocytic compartment into peptides of 10–30 amino acids that bind Class II molecules. HLA Class II-peptide complexes expressed on the membrane are usually recognised by CD4+ T-helper cells (1). Peptide-HLA complexes are the ligands of clonally distributed T-cell receptors (TCRs). TCRs are also able to recognise allogeneic HLA molecules at a high frequency, so that 1–10% of the peripheral blood lymphocytes of a donor can respond to a given allo-MHC antigen (2). Immune responses against incompatible HLA antigens may be extreme, such as in the case of graft-versus-host disease (GvHD) mediated by alloreactive cytotoxic T-lymphocytes (CTL), and thus represent a major barrier to HSCT.
3. Genomic organisation of the HLA system The MHC comprises 12 classical HLA genes located on a 3.6 Mb segment of the short arm of chromosome 6. Three HLA Class I genes (A, B, C) (Figure 2) encode for the heavy chains of HLA-A, B and -C antigens. Polymorphic residues are essentially located in the a1- and a2-domains encoded by exons 2 and 3, respectively, which form the peptide binding site. HLA Class II antigens (DR, DQ, DP) are heterodimers encoded by an a-chain and a b-chain gene (e.g. DRA/DRB1 or DQA1/DQB1) that co-localise at the centromeric part of the MHC (Figure 2) (1, 3). Essentially all of the polymorphism is located on exon 2 of b-chain genes, whereas the DRA gene is non polymorphic, and DQA1 and DPA1 loci exhibit a lower level of polymorphism (Figure 1). The HLA-DR sub-region presents an additional level of complexity since a second polymorphic DRB gene may be present, i.e. DRB3 in DR11/DR12/DR13/DR14/DR17/DR18 haplotypes, DRB4 in DR4/DR7/DR9 haplotypes, and DRB5 in DR15/DR16 haplotypes (Figure 2). Because of the codominant expression of HLA genes, a heterozygous individual may therefore express up to 12 different HLA antigens.
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Figure 2: HLA as a multigenic and polymorphic system
Schematic representation of the 12 HLA Class I and II loci in the MHC that comprises >200 genes on the short arm of chromosome 6. The corresponding number of antigens (as defined by serology) and alleles (as defined by nucleotide sequence) are indicated for each locus (3). About 10% of the alleles assigned by the HLA Nomenclature Committee are characterised by silent substitutions, and 2% are null alleles (3). Lower part of the figure: the HLA-DR subregion presents an additional level of complexity, with the presence of a second DRB gene in most haplotypes: DRB3, DRB4 or DRB5, which encode respectively the DR52, DR53, and DR51 antigens. For the nomenclature: see Appendix 1
4. Allelic polymorphism HLA genes are the most polymorphic loci of the human genome accounting for a total of 2584 alleles currently identified in worldwide populations (Figure 2) (3). Polymorphism of the HLA system was initially detected by serology, i.e. by using typing reagents derived from sera of multiparous women, or individuals who have been immunised by multiple transfusions. In the early 1980’s molecular cloning of the first HLA genes opened the way for a complete understanding of the molecular basis of HLA diversity and for the development of a variety of DNA-based typing techniques. Most of the serologically defined specificities are now subdivided into numerous alleles, and this number is still continuously growing (3). A regular update of the new alleles at the various HLA loci is available on the web (www.ebi.ac/imgt/hla) (Figure 2). Appendix 1 lists the HLA-A/B/C/DR/DQ serotypes and the corresponding groups of alleles detected by high resolution typing. The variability within a given serotype was recognised by cellular assays long before the DNA sequencing era. For example CTLs were shown to discriminate between two HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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HLA-A2 serologically identical individuals. Figure 3 shows a schematic alignment of the a1/a2 domain sequences with unique residues shared by all A2 alleles (G62, T142-H145). These residues are recognised by the monospecific and/or polyspecific allo-antisera used as typing reagents. Each of the 103 different A2 alleles expressed at the cell surface differs by a very limited number of residues, usually 1–4, and such allele mismatches can be efficiently recognised by CTLs (4–7). Although many of the Class I and II antigens comprise a large number of alleles worldwide, a limited number of alleles are found in any given population at a gene frequency >0.1% (8, 9). The combination of HLA alleles inherited on the same segment of chromosome 6 is referred to as a haplotype, for example the A1-B8-DR3 or the DRB1*1501DQB1*0602 haplotypes. Because of linkage disequilibrium HLA disparities at a given locus will frequently be associated with incompatibilities at an adjacent locus, as observed in many instances for B-Cw and DRB1-DQB1 bi-locus groups.
Figure 3: Schematic representation of HLA-A2 alleles showing common residues on the a1/a2-domains that are recognised by alloantisera
Open boxes mark residues that differ between the 0201-0207, 9201, and 9215 alleles. For example 0201 and 0206 alleles differ at one single position (Phe9 vs. Tyr9). Such a serologically silent mismatch is recognised by CTLs (see Figure 4) 50
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5. HLA typing: Methods and resolution levels Serology (microlymphocytotoxicity) is still a method of choice for low resolution typing in most laboratories, at least for HLA-AB, due to its simplicity and low cost. However the lack of monospecific alloantisera, the low-resolution power of the method and the requirement for viable lymphocytes, all contributed to the development of genomic DNA typing techniques, initially for Class II, later on for Class I typing. DNA typing techniques are based on the nucleotide sequence information of the polymorphic DNA segments, using PCR technology. A number of HLA typing methods based on DNA sequence variations have been developed, mainly using PCR-SSP (sequence-specific primers) amplification, or reverse PCR-SSOP (sequence-specific oligonucleotide probes) hybridisation on solid support (e.g. microbead arrays), or direct sequencing (8, 10). Three different levels of resolution for HLA DNA typing are usually recognised (Table 1). Low-resolution, also referred to as generic typing or 2-digit typing, corresponds to the identification of broad families of alleles that cluster into serotypes (e.g. A*02), and is thus the equivalent of serological typing (A2). High resolution, or 4-digit
Table 1: HLA nomenclature and levels of resolution HLA (a)
Definition
Resolution
A2
Refers to the A2 antigen defined by monospecific/ polyspecific antisera Any of the A*02 alleles (A*0201-0299 and A*9201-9215) Either one of these 4 alleles, other DRB1*11 alleles are excluded Allele defined Allele with a substitution in the coding sequence that leads to a stop codon (null allele), this allele is not expressed 2 DRB1*0301 alleles that differ by a silent substitution in exon 2, this difference is functionally silent, but may influence DNA typing A*2402 allele with low expression due to a mutation in intron 2 sequence, this allele is expressed at a level that is undetectable by serological typing
low
A*02 DRB1*1102/1103/ 1111/1114 A*0201 A*0232N DRB1*030101 and DRB1*030102 A*24020102L
low intermediate high high high
high
(a) DNA nomenclature: the first 2 digits refer to the serotype (A*02), the 3rd and 4th digits define substitution(s) in the coding sequence (A*0201), the 5th and 6th digits describe synonymous substitution(s), and the 7th and 8th 2 digits refer to substitution(s) in intron or 5’ or 3’ sequences. N and L mark alleles with respectively no or low surface expression. The suffix S means an allele encoding a protein that is expressed as a secreted molecule only and the suffix Q means an allele with a mutation that has been shown previously to have a significant effect on cell surface expression but without confirmation, and therefore with a questionable expression status
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typing, allows the discrimination of the individual alleles within each serotype (e.g. A*0201). Intermediate resolution HLA typing of a potential donor for this patient might give as a result A*0205/08/22: that means that the donor can be either A*0205, or A*0208, or A*0222, but definitively not A*0201 or any of the other A2 alleles. This level of typing results from the fact that these 3 alleles share common sequence determinants and are therefore identified in the same hybridisation group pattern. It is obviously very practical for rapid donor selection. In the example given above, such a donor typed as A*0205/08/22 would be disregarded for further analysis, whereas a parallel donor with the HLA type A*0201/01L/09/43N/66/75 would be selected and tested further in order to disclose compatibility at the allele level with the A*0201-positive patient. The accreditation program for the histocompatibility laboratories set up by the European Federation for Immunogenetics (EFI) has defined minimal criteria for HLA typing in related and unrelated HSCT (www.efiweb.org/standards.html).
6. HLA matching in related HSCT The best donor is a HLA genotypically matched sibling as identified by family typing. The family study allows not only the identification of a potential related donor, but also the confirmation of the patient’s genotype, which is important if an unrelated donor search is initiated. HLA-A, B, DR low resolution typing (serology or 2-digit DNA typing) is able in most cases to determine the paternal and maternal haplotypes present in the patient and a potential sibling donor. Thus ABDR typing can confirm genotypic identity for the whole set of HLA genes, i.e. a 12/12 match (Figure 4). Because of weak linkage disequilibrium between DP and the DR/DQ loci, a low level of DP-mismatched sibling donors (1–2%) are identified due to recombination. An HLA-A/B or B/DRB1 recombination event is detected by routine HLA-A/B/DR typing in about 2% of families. For a given patient the probability of having a genotypically identical sibling donor is 25% for each sibling, whereas the probability of having a haploidentical donor (i.e. with one shared haplotype) is 50%. When the patient has a very frequent ABDR haplotype it may be worth searching for a phenotypically identical donor in the blood-related members of the extended family, particularly when information on consanguinity is provided, for example when there is sound information that there has been intermarrying between aunts/uncles on paternal and aunts/uncles on maternal side, so that cousins may be HLA genotypically identical. In some families, apparent homozygosity of one of the parents may prevent formal identification of genotypic identity. As illustrated in the example shown in Table 2, both siblings 1 and 2 are phenotypically identical to the patient, based on ABDR 52
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Figure 4: Matching criteria in related and unrelated HSCT
By definition an HLA-genotypically identical sibling donor is compatible at the allele level at all loci on both chromosomes (12/12 match). In unrelated HSCT, matching for A/B/C/DRB1/DQB1 loci is usually searched for (10/10 match). In addition to DRB1 compatibility, some centres also consider DRB3/DRB5 polymorphism. DRB3 mismatches occur frequently in DR13 haplotypes. Because of strong linkage disequilibrium with DRB1 (at least in Caucasoids), the DRB5 locus is usually not tested. In DR15/16 haplotypes, DRB5 mismatches usually co-occur with DRB1 disparities. Searching for a 12/12 match implies DPB1 typing. Donors with an 8/8 match (not shown) or a 6/6 match apply, respectively, when HLAC/DP, or HLA-C/DQ/DP are not tested
low resolution typing. However typing for HLA-C and HLA-DRB1 at the allele level shows that sibling 1 is actually compatible with the patient, whereas sibling 2 has inherited a different maternal haplotype with 2 mismatches. In case of mismatched related HSCT, the risk of GvHD (recipient alleles absent in the donor) and graft failure (donor alleles absent in the recipient) increase with the number of HLA disparities on the non-shared haplotype (reviewed in ref. 11). A differential effect of Class I and Class II mismatches has been described, in which aGvHD risk was associated with Class II disparities (12).
7. HLA matching in unrelated HSC transplantation When no HLA-genotypically identical sibling donor is available, transplantation with HSC from HLA-ABCDRB1/DQB1-allele matched unrelated donors can result in comparable disease-free survival rates, notably for good-risk patients (11). Donor
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Table 2: Low versus high resolution HLA typing in homozygous haplotypes Haplotype
Low resolution typing
High resolution typing
father
a b
A1-B8-DR3 A31-B18-DR15
A*0101-B*0801-Cw*0701-DRB1*0301 A*3101-B*1801-Cw*1203-DRB1*1501
mother
c d
A2-B51-DR11 A2-B51-DR11
A*0201-B*5101-Cw*0501-DRB1*1101 A*0201-B*5101-Cw*1203-DRB1*1104
patient
a c
A1-B8-DR3 A2-B51-DR11
A*0101-B*0801-Cw*0701-DRB1*0301 A*0201-B*5101-Cw*0501-DRB1*1101
sibling 1
a c
A1-B8-DR3 A2-B51-DR11
A*0101-B*0801-Cw*0701-DRB1*0301 A*0201-B*5101-Cw*0501-DRB1*1101
sibling 2
a d
A1-B8-DR3 A2-B51-DR11
A*0101-B*0801-Cw*0701-DRB1*0301 A*0201-B*5101-Cw*1203-DRB1*1104
High resolution typing may disclose hidden incompatibilities in families where the 4 parental genotypes cannot be unambiguously determined. In this example the mother is ABDR homozygous by serology, but in fact has 2 different haplotypes on the basis of high resolution typing
identification has been largely facilitated by the Bone Marrow Donor Worldwide (BMDW) Registry (www.bmdw.org) which provides access to >11 million HLA-typed donors and cord blood units available in the national registries. Compared to HSCT from HLA genotypically identical sibling donors, unrelated HSCT is associated with an increased frequency of post-transplant complications, which are mainly due to undisclosed HLA incompatibilities not detected by serology (8, 13). Allele mismatches usually involve difference(s) in the peptide-binding site (Figure 5) that influence
Figure 5: Examples of common serologically hidden HLA Class I incompatibilities that involve single amino acid mismatches recognised by CTLs
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T-cell recognition either by direct contact with TCR or indirectly by modulating the repertoire of peptides bound by the HLA molecule. 7.1. Matching criteria for unrelated donors Whereas genotypically identical siblings share by definition the same alleles at all loci, the degree of allele matching in two unrelated individuals strictly depends on how many loci are considered for high resolution typing. Figure 4 illustrates the three most common situations: the gold standard matching comprises the analysis of HLAA, -B, -C, -DRB1, and -DQB1 loci. When a patient and a donor share the same 5 alleles on both haplotypes the situation is referred to as a 10/10 match. If HLA-C or -DQB1 typing is omitted, this will be an 8/8 match, and if only the A/B/DRB1 loci are considered, it becomes a 6/6 match. Obviously because of the multiple possible BC and DRB1-DQB1 associations, a 6/6 match may in fact correspond to a 6/10 match, i.e. to an incompatible combination (Table 3). In the case of non-malignant diseases the graft versus leukaemia effect that may be mediated by DP incompatibilities is not necessary and DPB1 matching might be considered when several HLA-A/B/C/DR/DQ-compatible donors are available (12/12 match).
Table 3: Hidden incompatibilities in HLA-ABDRB1 allele-matched donors Matching
6/6
8/10
A*
B*
Cw*
DRB1*
DQB1*
patient
0201/1101
0801/3503
NT
0301/1101
NT
donor
0201/1101
0801/3503
NT
0301/1101
NT
patient
0201/1101
0801/3503
0701/0401
0301/1101
0201/0301
donor
0201/1101
0801/3503
0701/1203
0301/1101
0201/0501
7.2. Allele matching: Relative importance of individual loci In the early 1990’s the role of HLA matching was hampered by the poor resolution achieved by HLA typing, particularly for HLA Class I alleles. Based on high resolution typing methods recent studies (14–18) have reached the almost general consensus that allele-level matching does improve transplant outcome after both myeloablative and reduced-intensity conditioning regimen (reviewed in ref. 11). The HLA effect on transplant outcome is modulated by the disease risk with an effect of single disparities reported to be significant for low risk disease patients (16). However the relative importance of individual loci still remains an open issue. Whereas
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the effect of A/B/C/DRB1 mismatches have been well documented by most retrospective large scale studies, the role of DQ and DP incompatibilities appear more controversial (reviewed in ref. 11). The National Marrow Donor program (NMDP) Histocompatibility Committee has recommended allele-level typing for HLAA/B/C/DRB1 (8/8 match) (13). There is however some evidence suggesting a trend of an additive HLA-DQ effect in HSCT from donors with multiple mismatches (16). Recent data on T-cell-depleted unrelated HSCT have documented that HLA-DPB1 matching was associated with increased risk of relapse, irrespective of the compatibility status for the other loci (19). A comparison of serological versus allele class I mismatches in CML patients suggested that qualitative differences may influence the risk of graft failure, with a higher risk in serotype-mismatched patients (15, 20). Contrasting outcomes have been reported in studies with patients from different ethnic backgrounds: in the Japanese Marrow Donor Program (JMDP) study (14), HLAA/B/C/DRB1 mismatches were found to be significant risk factors for grades III–IV acute GvHD, whereas the U.S. National Marrow Donor program (NMDP) data revealed a DRB1 effect with no contribution of HLA-DQ/DP or HLA-B/C (15) mismatches. HLAA/B/C/DRB1, but not DQ/DP mismatches decreased survival in the NMDP study (15), whereas in the JMDP study only A/B/DRB1 disparities were associated with mortality (14). Differences between studies may involve selection criteria of each transplant centre, patients age or other pre-transplant risk factors, experience in treating GvHD, as well as the relevance of the GvL effect in CML patients. Also when comparing studies from varying population groups, major differences between mismatched allele combinations may contribute to contrasting clinical outcomes. Selection of cord blood units is generally based on HLA-A/B-low and -DRB1-high resolution matching, taking into account that cell dose is the most important parameter affecting outcome. Data on the effect of HLA matching in cord blood transplantation are difficult to interpret, since most reports lack HLA-C/DQ and high resolution HLA-AB typing results. Nevertheless cord blood appears less HLA restricted than adult HSCT, in other words HLA incompatibilities may be less detrimental (21).
8. Probability estimates of finding a matched unrelated donor Identification of an unrelated HLA allele-matched HSC donor is a costly and time consuming procedure. To improve search efficiency, an estimate of the probability of identifying a 10/10 HLA matched donor allows an early decision to transplant with HSC from an alternative donor (haploidentical, cord blood, autologous), or to accept varying degrees of allele-mismatches for searches with a low probability of success, depending on the clinical context and local guidelines. Based on the current 56
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knowledge of allele and haplotype frequencies in different populations the probability of identifying a 10/10 matched donor at the start of the search is highly predictable (22). The number of ABDR-matched (low resolution) donors available in the Registry for a given patient often reflects the chance of finding a donor compatible at the allele level. However, the following parameters have a negative impact on this probability: • presence of a rare allele in the patient (<5% within a given serotype, e.g. B*4405) • HLA-ABDR haplotype that does not belong to the 10 most frequent haplotypes in Caucasoids • unusual B-DR association (e.g. B65-DRB1*0101) • unusual DR-DQ association, e.g. DRB1*1501-DQB1*0603 • presence of an antigen that is split into >2 alleles occurring at a frequency >10% of a given serotype in the population, e.g. B35, B44, DR4, DR11, DR13 • presence of B*2705, B18, B*4402, B*4403, B51 (higher risk of HLA-C mismatch) • <3 donors available in the BMDW registry. Because of different distribution of alleles and HLA haplotypic associations, allele matching will also be more difficult when patients and potential donors belong to different ethnic groups. For European Caucasoid patients a 40–50% probability of identifying a 10/10 matched donor has been reported (22, 23).
9. Choice of donor based on HLA typing Vigorous T-cell responses are generated against allogeneic tissues when the graft and the host express different MHC molecules. Single amino acid differences in the antigen recognition site of a HLA molecule can initiate graft rejection or GvHD. The choice of the donor will therefore depend on the degree of HLA identity between donor and recipient. Looking for a related donor 1. HLA-A/B/DR typing (serology or low resolution DNA typing) of patient, the sibling(s) and both parents to determine the haplotypes. 2. If a sibling is geno-identical the transplant can be performed after a confirmatory typing on a second blood sample from patient and donor but without further HLA investigations. 3. If one sibling is a monozygotic twin this is a syngeneic transplant, this donor can be chosen but some of the GvL effect might be lost. 4. If both parents share a common haplotype or if one parent is HLA-ABDR homozygous, this is considered to be a pheno-identical transplant. High
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resolution Class I and II typing is recommended to define the exact degree of compatibility. 5. If a family donor with only one HLA difference (5/6 match) is found, the transplant can be performed despite the possibility of an increased risk of GvHD. This is a related one antigen mismatched donor transplant. High resolution Class I and II typing is recommended to define the exact degree of compatibility. 6. Parents and/or siblings sharing only one haplotype with the recipient are considered as a haploidentical-related donor transplant. Looking for an unrelated donor 1. One should obtain a complete Class I and Class II high resolution typing for the patient (HLA-A, -B, -C, -DRB1 and -DQB1). Several levels of compatibility are defined: 12 out of 12 (A/B/C/DRB1/DQB1/DPB1) 10 out of 10 (A/B/C/DRB1/DQB1) or 8 out of 8 (A/B/C/DRB1) (see section 7). The level of HLA incompatibility (low or high resolution typing) accepted can vary from centre to centre but the priority is to look for the highest degree of compatibility at the allele level. DPB1 typing should be included in case several 10/10 matched donors are available. 2. Look at the BMDW (Bone Marrow Donor Worldwide) database to evaluate the probability of finding a donor, and determine which registries have suitable donors. 3. Send a search request to the bone marrow donor registries which have suitable donors in order to perform a confirmatory typing (high resolution) to find the best HLA matched unrelated donor. 4. Search simultaneously for a cord blood unit through BMDW and Netcord in order to find an unrelated cord blood donor. If several HLA identical bone marrow donors are found, choose the male, ABO identical, and/or CMV-negative donor. If there is no well matched (10/10, 9/10, or possibly 8/10) unrelated donor or if the time frame is too short, choose a cord blood unit, as long as the number of nucleated cells is >2 x 107/kg and there are no more than 2 HLA mismatches (4/6).
10. Conclusions The HLA system, with 100 serologically defined specificities and over 2500 alleles, represents a major barrier to HSC transplantation. There is now a broad consensus that selection of unrelated HSC donor by high resolution molecular typing technology contributes to a better clinical outcome. However the relative importance of individual loci still remain to be better defined, and multicentre studies should contribute to resolving this issue. The difficulty in reaching clear consensus among 58
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the clinical studies, particularly with respect to the role of single locus mismatches, possibly results from subtle differences in patients groups with respect to patient and donor selection criteria, pre-transplant risk factors or GvHD prophylaxis. Additional immunogenetic factors such as minor histocompatibility antigens, cytokine and chemokine genes, or activating/inhibitory killer immunoglobulin-like receptor (KIR) genes, may play a role in determining transplant outcome. Donor HLA matching criteria should take into account parameters such as the time frame allowed by the patient’s disease and the probability of identifying a well matched donor based on the patient’s HLA phenotype.
References 1. Klein J, Sato A. The HLA system. First of two parts. New Engl J Med 2000; 343: 702-709. 2. Afzali B, Lechler RI, Hernandez-Fuentes MP. Allorecognition and the alloresponse: Clinical implications. Tissue Antigens 2007; 69: 545-556. 3. Marsh SGE, Albert ED, Bodmer WF, et al. Nomenclature for factors of the HLA system, 2002. Tissue Antigens 2002; 60: 407-464. 4. Fleischhauer K, Kernan NA, O'Reilly RJ, et al. Bone marrow-allograft rejection by T lymphocytes recognizing a single amino acid difference in HLA-B44. N Engl J Med 1990; 323: 1818-1822. 5. Rufer N, Tiercy J-M, Breur-Vriesendorp B, et al. Histoincompatibilities in ABDR-matched unrelated donor recipient combinations. Bone Marrow Transplantation 1995; 16: 641-646. 6. Oudshoorn M, Doxiadis II, van den Berg-Loonen PM, et al. Functional versus structural matching: Can the CTLp test be replaced by HLA allele typing? Hum Immunol 2002; 63: 176-184. 7. Scott I, O’Shea J, Bunce M, et al. Molecular typing reveals a high level of HLA class I incompatibility in serologically well matched donor/recipient pairs - Implications for unrelated bone marrow donor selection. Blood 1988; 92: 4864-4871. 8. Tiercy JM, Villard J, Roosnek E. Selection of unrelated bone marrow donors by serology, molecular typing and cellular assays. Transpl Immunol 2002; 10: 215-221. 9. Hurley CK, Fernandez-Vina M, Hildebrand WH, et al. A high degree of HLA disparity arises from limited allelic diversity: Analysis of 1775 unrelated bone marrow transplant donorrecipient pairs. Human Immunol 2007; 68: 30-40. 10.Little AM, Marsh SG, Madrigal JA. Current methodologies of human leukocyte antigen typing utilized for bone marrow donor selection. Curr Opin Hematol 1998; 5: 419-428. 11.Petersdorf EW. Risk assessment in hematopoietic stem cell transplantation. Best Pract Res Clin Haematol 2007; 20: 155-170. 12.Ottinger HD, Ferencik S, Beelen DW, et al. Hematopoietic stem cell transplantation: Contrasting the outcome of transplantations from HLA-identical siblings, partially HLAmismatched related donors, and HLA-matched unrelated donors. Blood 2003; 102: 11311137. 13.Hurley CK, Wagner JE, Setterholm MI, Confer DL. Advances in HLA: Practical implications for selecting adult donors and cord blood units. Biol Blood Marrow Transplant 2006; 12
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(1 Suppl 1): 28-33. 14.Morishima Y, Sasazuki T, Inoko H, et al. The clinical significance of human leukocyte antigen (HLA) allele compatibility in patients receiving a marrow transplant from serologically HLA-A, HLA-B, and HLA-DR matched unrelated donors. Blood 2002; 99: 4200-4206. 15.Flomenberg N, Baxter-Lowe LA, Confer D, et al. Impact of HLA class I and class II high resolution matching on outcomes of unrelated donor bone marrow transplantation. Blood 2004; 104: 1923-1930. 16.Petersdorf EW, Anasetti C, Martin PJ, et al. Limits of HLA mismatching in unrelated hematopoietic cell transplantation. Blood 2004; 104: 2976-2980. 17.Chalandon Y, Tiercy J-M, Schanz U, et al. Impact of high resolution matching in allogeneic unrelated donor stem cell transplantation in Switzerland. Bone Marrow Transplant 2006; 37: 906-916. 18.Carreras E, Jiminez M, Gomez-Garcia V, et al. Donor age and degree of HLA matching have a major impact on the outcome of unrelated donor hematopoietic cell transplantation for chronic myeloid leukemia. Bone Marrow Transplant 2006; 37: 33-40. 19.Shaw BE, Marsh SG, Mayor NP, et al. HLA-DPB1 matching status has significant implications for recipients of unrelated donor stem cell transplants. Blood 2006; 107: 1220-1226. 20.Petersdorf EW, Hansen JA, Martin PJ, et al. Major-histocompatibility-complex class I alleles and antigens in hematopoietic-cell transplantation. New Engl J Med 2001; 345: 17941800. 21.Gluckman E, Rocha V. Donor selection for unrelated cord blood transplants. Curr Op Immunol 2006; 18: 565-570. 22.Tiercy J-M, Nicoloso de Faveri G, Passweg J, et al. The probability to identify a 10/10 HLA allele-matched unrelated donor is highly predictable. Bone Marrow Transplantation, 2007; 40: 515-522. 23.Tiercy J-M, Bujan-Lose M, Chapuis B, et al. Bone marrow transplantation with unrelated donors: What is the probability of identifying an HLA-A/B/Cw/DRB1/B3/B5/DQB1matched donor? Bone Marrow Transpl 2000; 26: 437-441.
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Appendix 1. List of HLA-A, -B, -Cw, -DR and -DQ serotypes with their corresponding groups of alleles (June 2007) serotype A1 A2 A203 A210 A3 A11 A23 (9) A24 (9) A2403 A25 (10) A26 (10) A29 (19)
alleles A* 0101-0125 0201-0299, 9201-9215 0203 0210 0301-0329 1101-1130 2301-2315 2402-2476 2403, 2410 2501-2506 2601-2634 2901-2916
B7 B703 B8 B13 B14 B15 B18 B27 B2708 B35 B37 B38 (16) B39 (16) B3901 B3902 B40 B4005 B41 B42 B44 B45
alleles B* 0702-0754 0703 0801-0833 1301-1317 1401-1407N 1501-1599, 9501-9529 1801-1826 2701-2737 2708 3501-3575 3701-3712 3801-3816 3901-3941 3901 3902 4001-4074 4005 4101-4108 4201-4209 4402-4453 4501-4507, 5002
serotype A30 (19) A31 (19) A32 (19) A33 (19) A34 (10) A36 A43 (10) A66 (10) A68 (28) A69 (28) A74 (19) A80
B53 B54 (22) B55 (22) B56 (22) B57 (17) B58 (17) B60 (40) B61 (40) B62 (15)
B63 (15) B64 (14) B65 (14) B67 B70 (15) B71 (70)
alleles A* 3001-3021 3101-3117 3201-3215 3301-3310 3401-3408 3601-3604 4301 6601-6606 6801-6838 6901 7401-7412N 8001 alleles B* 5301-5312 5401-5411 5501-5526 5601-5620 5701-5712 5801-5815 4001,4007,4010,4014, 4031,4034,4048,4054 4002-4004,4006,4009, 4016,4027,4029 1501,1504-07,1515, 1520,1524,1525,1527, 1528,1530,1532,1535, 1539,1545,1548,1570, 1571,1573,1582,1584, 1516-1517 1401 1402 6701-6702 1509,1537,1551 1510,1518,1580,1593 HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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B46 B47 B48 B49 (21) B50 (21) B51 (5) B5102 B5102 B52 (5)
4601-4610 4701-4705 4801-4816 4901-4905 5001-5002,5004 5101-5148 5102 5102 5201-5211
B72 (70) B73 B75 (15)
serotype Cw1 Cw2 Cw3 Cw4 Cw5 Cw6 Cw7 Cw8
alleles Cw* 0101-0118 0202-0218 0302-0340 0401-0427 0501-0516 0602-0616N 0701-0748 0801-0814
serotype Cw9 Cw10 Cwx Cwx Cwx Cwx
DR1 DR1 DR15 (2) DR16 (2) DR3 DR4 DR7 DR8
alleles DRB1* 0101-0110 0101-0116 1501-1522 1601-1611 0302-0335 0401-0464 0701, 0703-0712 0801-0832
DR52
DR53 blank
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DRB3*0101-01 DRB3*0201-0218 DRB3*0301-0303 DRB4*0101-0106 DRB4*01030102N, 0201N,0301N
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B76 (15) B77 (15) B78 B81 B82 B83
1503,1546 7301 1502,1508,1511,1521, 1531 1512,1514 1513 7801-7805 8101-8102 8201-8202 8301
Cwx Cwx
alleles Cw* Cw*0303 Cw*0302,0304 1202-1221 1402-1408 1502-1520 16011602,1604, 1606-1609 1701-1704 1801-1803
DR9 DR10 DR11 (5) DR12 (5) DR13 (6) DR14 (6)
alleles DRB1* 0901-0906 1001 1101-1162 1201-1215 1301-1379 1401-1466
DR51
DRB5*0101-0112 DRB5*0201-0205
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CHAPTER 3.1 • HLA and donor matching
DQ2 DQ3 DQ4 DQ5 (1)
alleles DQB1* 0201-0205 0301-0320 0401-0402 0501-0505
DQ6 (1) DQ7 (3) DQ8 (3) DQ9 (3)
alleles DQB1* 0601-0630 0301, 0304 0302, 0305, 0310 0303
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The biological function of HLA molecules is to present peptide antigens to T-cells. The peptide binding site of HLA Class I molecules is composed of: a) An a-chain and a b-chain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The proximal part of the a-chain. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The distal part of the a-chain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The a-chain and the b2-microglobulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. A heterozygous individual can express at the cell surface a maximum of: a) 10 different HLA Class I and Class II antigens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 12 different HLA Class I and Class II antigens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 13 different HLA Class I and Class II antigens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 14 different HLA Class I and Class II antigens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. A HLA-ABDR haplotype is: a) A combination of alleles encoded on the same chromosome . . . . . . . . . . . . . . b) A combination of alleles shared by two unrelated individuals . . . . . . . . . . . . . c) A combination of alleles that differ between a patient and a partially incompatible sibling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) A combination of alleles that are frequent in a given population . . . . . . . . . 4. In which patient/donor combination would you expect a higher risk of HLA-C mismatches? a) Two HLA-ABDR phenotypically identical siblings. . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Two HLA-ABDR phenotypically identical unrelated individuals . . . . . . . . . . . . c) Two HLA-ABDR phenotypically identical cousins . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Two HLA-ABDR genotypically identical siblings . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 3.1 • HLA and donor matching
5. The HLA-ABDR typing of a patient’s family leads to the identification of a potential sibling donor who differs from the patient by one single DR antigen. The father is homozygous for HLA-AB antigens and heterozygous for HLA-DR. The possible reason for the DR-incompatibility is: a) A recombination event between HLA-A and -B in one paternal haplotype. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A recombination event between HLA-B and -DR in one paternal haplotye . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Inheritance of 2 different paternal HLA-ABDR haplotypes . . . . . . . . . . . . . . . . . d) Either of the 3 possibilities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 3
Immunogenetics of allogeneic HSCT
*
3.2
KIR: Beneficial effects of natural killer cell alloreactivity in haploidentical HSCT A. Velardi, L. Ruggeri, A. Mancusi, F. Aversa, M. F. Martelli
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CHAPTER 3.2 • NK cells in haploidentical transplantation
1. Introduction Recent studies have shown natural killer (NK) cells influence the outcome of haploidentical haematopoietic transplantation in a remarkable way, with a favourable outcome where NK cells are alloreactive in the donor-recipient direction (1-4). NK cell activation is regulated by a balance between inhibitory and activating receptors, called killer-cell Ig-like receptors (KIRs). In humans, currently 16 inhibitory KIR genes and pseudo-genes are known, which codify for inhibitory and activating KIRs. Inhibitory KIRs recognise amino acids in the COOH-terminal portion of the MHC class I a1 helix (reviewed in refs. 3, 5, 6). They possess two (KIR2D) or three (KIR3D) extra-cellular C2-type Ig-like domains and a long cytoplasmic tail (L) containing immunoreceptor tyrosine-based inhibition motifs (ITIM) which recruit and activate SHP-1 and SHP-2 phosphatases for inhibitory signal transduction. KIR2DL1 recognises HLA-C alleles characterised by a Lys80 residue (HLACw4 and related, “Group 2” alleles). KIR2DL2 and KIR2DL3 (which are allele variants) recognise HLA-C with an Asn80 residue (HLA-Cw3 and related, “Group 1” alleles). KIR3DL1 is the receptor for HLA-B alleles sharing the Bw4 supertypic specificity (Table 1). Another type of human NK cell inhibitory receptor involved in HLA recognition is CD94-NKG2A. It binds to the non-conventional class I molecule HLA-E. Several HLA class I alleles provide signal sequence peptides that bind HLA-E and allow its expression at the cell surface. Consequently, it is expressed in every individual. Inhibitory KIRs, CD94/NKG2 and HLA-class I genes determine individual NK cell repertoires during development. As they are located on different chromosomes, receptors and ligands segregate independently in human pedigrees. The HLA class I genotype selects a self-tolerant repertoire by dictating which KIR and/or NKG2A
Table 1: HLA-class I allele specificity of the main KIRs expressed by human NK cells KIR gene
Encoded protein
HLA specificity
KIR2DL1
P58.1 receptor
HLA-C group 2 (e.g., -Cw2, -Cw4, -Cw5, -Cw6) (Lys80)
KIR2DL2/3
P58.2 receptor
HLA-C group 1 (e.g., -Cw1, -Cw3, -Cw7, -Cw8) (Asn80)
KIR3DL1
P70/NKB1 receptor
Bw4 alleles (e.g., HLA-B27)
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receptor combinations are to be used as inhibitory receptors for self HLA class I (7). Consequently, every functional NK cell in the mature repertoire expresses at least one inhibitory receptor for self HLA: co-expression of two or more receptors is less frequent.
2. Inhibitory KIRs and alloreactivity Since inhibitory KIRs recognise specific groups of HLA class I molecules, i.e., HLAC group 1, HLA-C group 2, HLA-Bw4 alleles, NK cells with the potential to exert alloreactions use KIRs as inhibitory receptors for self (1-6). NK cells which express, as their only inhibitory receptor for self, a KIR for the HLA class I group which is absent on allogeneic targets, sense the missing expression of the self class I KIR ligand and mediate alloreactions (“missing self” recognition). Importantly, most individuals possess a full complement of inhibitory KIR genes and can exert NK cell alloreactions (3, 4, 6). In particular, 100% of individuals possess the KIR2DL2 and/or KIR2DL3 receptors for HLA-C group 1 alleles. If they have HLA-C group 1 allele(s) in their HLA type, these individuals possess HLA-C1-specific NK cells which are alloreactive against cells from individuals who do not express HLA-C group 1 alleles. Ninety-seven percent of individuals possess the KIR2DL1 receptor for HLAC group 2. If they possess HLA-C group 2 allele(s) in their HLA type, these individuals have HLA-C2-specific NK cells which mediate alloreactions against cells from individuals who do not express HLA-C group 2 alleles. Finally, ~90% of individuals possess the KIR3DL1 receptor for HLA-Bw4 alleles. When they have HLABw4 allele(s) in their HLA type, these individuals may have HLA-Bw4-specific NK cells that are alloreactive against Bw4-negative cells. These KIR ligand mismatches often occur in haploidentical donor recipient transplant pairs.
3. Clinical effects of NK cell alloreactivity When exerted in the donor-versus-recipient direction, NK cell alloreactivity emerged as a crucial factor in improving outcomes of haploidentical transplantation (1-4). It reduced the risk of leukaemia relapse, did not cause graft versus host disease (GvHD) and markedly improved event-free survival (EFS) in a series of haploidentical transplants (57 acute myeloid leukaemia (AML) patients, 20 of whom were transplanted from NK alloreactive donors) (2). In an updated analysis (4), 112 highrisk AML patients received haploidentical transplants from NK alloreactive (n=51) or non-NK alloreactive donors (n=61). Transplantation from NK-alloreactive donors was associated with: - a significantly lower relapse rate in patients transplanted in any CR (3 vs. 47%) (p<0.003); 68
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- better EFS whether patients transplanted in relapse (34 vs. 6%, p=0.04) or in remission (67 vs. 18%, p=0.02); - overall reduced risk of relapse or death (relative risk vs. non-NK-alloreactive donor: 0.48 [95%CI: 0.29-0.78], p<0.001). The 67% probability of surviving event-free for acute myeloid leukaemia patients transplanted in any remission from NK alloreactive donors is in the range of best survival rates after transplantation from unrelated donors and cord blood units. The 34% EFS of patients transplanted in chemo-resistant relapse from NK alloreactive donors is also satisfactory. Transplantation from non-NK alloreactive haploidentical donors appears justified only for AML patients in remission and even in these patients it is associated with only an 18% EFS. Lack of an NK alloreactive donors is a contra-indication to transplant for patients in chemo-resistant relapse as very few survive. Several observations suggest alloreactive NK cells are responsible for favourable transplantation outcomes. Transfer of human alloreactive NK cells to NOD-SCID mice eradicated previously transplanted human AML cells (2). KIR ligand mismatches correlated with the ability of donor NK cell clones to kill cryopreserved haematopoietic recipient cells, including leukaemia cells (1, 2, 4). Most importantly, as depicted in Figure 1, upon transplantation from NK alloreactive donors the engrafted stem cells give rise to an NK cell repertoire which includes donor-vs-recipient alloreactive NK clones which kill cryopreserved haematopoietic recipient cells, including leukaemia cells (1). Donor versus recipient alloreactive NK clones are detectable in vivo in recipients for up to 1 year after transplant (4).
4. Selection of an NK alloreactive donor In the clinical studies above, NK alloreactive donors were found for ~50% of patients, which approaches the maximum because, in fact, 1/3 of the population expresses class I alleles belonging to all three class I groups recognised by KIRs and block NK cells from every donor. How is an NK alloreactive donor selected? Recipients who express alleles belonging to 1 or 2 of the 3 class I allele groups recognised by KIRs may find NK alloreactive donors. Donors who are HLA-C group mismatched with their recipients possess highfrequency NK clones which are alloreactive against recipients’ target cells (1-4). Thus, high-resolution HLA-C typing is a good predictor of NK cell alloreactivity. Since 3% of individuals do not possess the KIR2DL1 gene, the combination of a KIR2DL1negative donor and a recipient without HLA-C group 2 alleles could result in a 1.5% incidence of false positivity. KIR2DL1 gene typing of the donor may be necessary to assess the NK alloreactive potential of this combination.
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Figure 1 NK repertoire Host targets CD16 CD8 CD4
cells/cmm
600 400 200 0
High-intensity conditioning
0
50
100 days
150
200
KIR2DL1
HLA-C group 2 Cw2/Cw4
KIR2DL2/3
Missing group 1
KIR3DL1
HLA-B51
T cell
Stem Stem Stem Stem Stem Stem Stem Stem Stem Stem Stem
Mega-dose stem cells from “allo NK” donor No post-transplant immune suppression
Haematopoietic stem cell transplantation from a KIR ligand-mismatched donor who possesses donor versus recipient alloreactive NK clones in his/her repertoire gives rise to transient post-transplant generation of a repertoire which is identical to the donor’s, including the alloreactive clones. In fact, alloreactive clones can be isolated from the circulation of successfully engrafted recipients and can kill cryopreserved recipient targets, including leukaemic cells (1, 4)
In HLA-Bw4 mismatches, even when the KIR3DL1 gene is present in the donor (~90% of individuals), NK repertoire studies show alloreactive NK clones are non-detectable ~1/3 of individuals (4). In some allelic variants in the HLA-Bw4 inhibitory NK receptor gene KIR3DL1 may not allow full receptor expression at the cell membrane and affect NK cell inhibition by HLA-Bw4 ligand (8); others apparently express alloreactive NK clones in very low frequencies. Thus, for HLA-Bw4 mismatches, functional assessment of the donor NK repertoire appears necessary. As approximately half of unrelated donor transplants are mismatched for one or more HLA class I alleles, donor vs. recipient NK cell alloreactivity may also occur in this setting. However, some retrospective studies show no advantage in transplantation from KIR ligand-mismatched donors (9, 10). Unrelated donor transplant protocols are heterogeneous in conditioning regimens, patient populations and underlying diseases. They use T-cell-deplete bone marrow harvests (or, less frequently, peripheral blood progenitors) which contain ~4 log more T-cells and up to 1 log fewer stem cells than haploidentical grafts. Relatively few transplanted stem cells, combined with the high T-cell graft content and post-transplant immune suppression have been associated with poor reconstitution of potentially alloreactive, KIR-bearing NK cells (11). Nonetheless, other studies have observed an increased GvL effect (12–15). 70
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CHAPTER 3.2 • NK cells in haploidentical transplantation
A marked survival advantage was reported in patients who received anti-thymocyte globulins pre-transplant (which provided in vivo T-cell depletion) and a graft containing 2-3-fold more nucleated cells than usual in unrelated-donor transplants (12). Prospective studies are needed to determine whether strategies (high doses of stem cells, T-cell depletion, no post-transplant immune suppression), which in haploidentical transplantation harness donor-vs-recipient NK cell alloreactivity, can be implemented to improve outcome in unrelated donor transplants.
5. The “missing ligand” model Since the original report that NK cell alloreactivity in haploidentical transplantation rests upon KIR ligand mismatching and donor NK cell recognition of “missing self” on recipient targets (2), the “missing ligand” model has been proposed as a powerful algorithm for predicting favourable transplant outcomes not only in haploidentical transplants (16) but also in matched sibling (17) and in unrelated donor transplants (18). Under the perturbed conditions that exist after a haematopoietic stem-cell transplant, it hypothesises that NK alloreactions occur when KIR ligand-matched donors possess an “extra” KIR for which neither donor nor recipient have an HLA ligand. These donors may carry KIR-bearing NK cells in an anergic/regulated state which, upon transfer into the recipient, are hypothesised to become activated and exert a GvL effect. However, even though self-tolerant NK cells which do not express inhibitory receptors for self-MHC have been described (19) no studies have as yet determined whether tolerant NK cells acquire/resume cytotoxic effector function after transplant. When an adult series of AML patients who were transplanted from haploidentical donors was analysed according to the “missing ligand” algorithm, the “missing ligand” transplant recipients disappointingly had a worse prognosis than patients transplanted from NKalloreactive (KIR ligand mismatched) donors (4). While differences in diseases, age of patients and transplantation protocols, such as ATG vs. no ATG in the conditioning or peripheral blood CD34+ cells vs. bone marrow as a source of haematopoietic cells, may account for these conflicting results, the analysis shows that donor NK cell recognition of “missing self” on recipient targets is essential for triggering powerful NK cell alloreactions that impact beneficially on transplantation outcomes.
6. Effects of donor activating KIR genetics on donor vs. recipient NK cell alloreactivity Activating KIRs, which regulate NK and T-cell functions, are molecular homologues of the inhibitory KIRs with shorter cytoplasmic tails (S) (reviewed in refs. 3, 5, 6) and a charged residue in their transmembrane domain that allows association with
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ITAM containing signalling polypeptides. Knowledge of their ligand specificity is limited. Studies have reported a weak interaction between KIR2DS1 and Lys80 HLAC molecules, despite its homology to KIR2DL1, and an even weaker interaction between KIR2DS2 and Asn80 HLA-C molecules, despite its homology to KIR2DL2 and KIR2DL3. Unlike inhibitory KIRs, activating KIRs exhibit extensive variation in gene number and content, which leads to heterogeneity within the general population and diverse ethnic groups (reviewed in ref. 6). Indeed, activating KIRs may not even be present in approximately 25% of Caucasians who are homozygous for the so-called group A KIR gene haplotypes which contain inhibitory KIR genes and the KIR2DS4 activating KIR gene (encoding for a non-functional protein in 2/3 of individuals). On the other hand, 75% of Caucasians are either heterozygous or homozygous for B haplotypes which carry not only inhibitory KIR genes but also various combinations of activating KIR genes (KIR2DS1-2-3-5 and KIR3DS1).
7. Activating KIRs and results of haploidentical HSCT We, therefore, evaluated the role of donor activating KIR genetics in haploidentical haematopoietic transplantation (Mancusi et al., manuscript submitted for publication). In a series of 84 haploidentical transplants for AML, the impact of donor KIR genetics (group A vs. group B KIR gene haplotypes) was assessed separately in NK alloreactive and non-NK alloreactive transplants. Forty-seven recipients were transplanted from NK alloreactive donors (12 with group A KIR gene haplotypes vs. 35 with B haplotypes) and 37 recipients from non-NK alloreactive donors (8 with group A KIR gene haplotypes vs. 29 with B haplotypes). KIR gene haplotypes had no impact in non-NK alloreactive transplants. In transplants from NK alloreactive donors, presence of group B haplotype KIR genes in the donors was associated with reduced incidence of TRM (largely infection-related) (B vs. A haplotypes: 20 vs. 67% TRM, p<0.005). In multivariate analyses against disease status at transplant, age, patient and donor sex, conditioning regimens, and the number of CD34+ and CD3+ cells in the graft, it was the only significant variable predicting protection from TRM (RR: 0.24; 95%CI: 0.14-0.42; p<0.01) and resulted in a trend towards better EFS (60 vs. 33%, p<0.1). When the number of activating KIR genes in the donor was taken into account, donors carrying ≥ 3 activating KIR genes provided significant protection from TRM and significantly better EFS compared with A haplotype donors (TRM: 12 vs. 67%, p<0.003) (EFS: 71 vs. 33%, p=0.02). In multivariate analysis, transplantation from alloreactive donors carrying ≥3 group B haplotype activating KIR genes was the only variable predicting protection from TRM (RR: 0.40; 95%CI: 0.27-0.58; p<0.02) and significantly improved EFS (RR: 0.56; 95%CI: 0.320.98; p<0.05). Thus, while NK-alloreactive donors protect against leukaemia relapse, 72
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CHAPTER 3.2 • NK cells in haploidentical transplantation
those who also carry activating KIRs protect against infectious mortality and help improve survival. The chance of finding an NK alloreactive donor is ~ 50% of haploidentical transplants and the odds of finding an NK alloreactive donor carrying activating KIRs are ~ 30% of haploidentical transplants.
8. Activating KIRs and protection against infection Protection against infection may be mediated directly by NK cells or indirectly through other mechanisms. Activating KIRs could enhance NK cell cytokine secretion and cytotoxicity against pathogen infected cells in the context of missing self. A notable example of direct recognition of a pathogen by activating NK receptor is provided by murine CMV protein m157 and the murine Ly49H NK receptor (20). In humans, progression to AIDS is slower in patients who have both KIR3DS1 and the HLA-Bw4 allotype, the putative ligand of KIR3DS1 (21). More NK cells expressing NKG2C are present in CMV-exposed individuals, suggesting this activating NK cell receptor plays a role in the immune response to this infection (22). Therefore associations between activating NK receptors and enhanced immunity against infections have been documented. Activating KIRs could also help control infections indirectly through the interaction between NK cells and dendritic cells (DCs) (reviewed in ref. 23). NK cells regulate DC homeostasis and maturation. Mature DCs can, in turn, activate NK cells. In vivo NK/DC interactions in lymphoid organs or non-lymphoid tissues can lead to Th1 polarisation. NK cells in lymph nodes provide the early IFN-g production, which is essential for Th1 polarisation. Consequently, the interaction between NK cells and DCs influences the quality and the strength of adaptive immune response. The clinical data suggest either or both these mechanisms could operate in haploidentical transplants from NK-alloreactive donors who possess activating KIRs. Whatever the mechanisms, these studies have improved criteria for donor selection.
9. Conclusions Transplantation from a full HLA haplotype mismatched family member is nowadays a viable option for patients with acute leukaemia at high risk of relapse who urgently need a transplant and who do not have a matched donor (24). The mismatched transplant relies for its success on the combined action of: - high-intensity conditioning regimens to ensure the lowest possible residual leukaemia burden and optimal immunosuppression; - high doses of haematopoietic cells to ensure engraftment across the HLA barrier; - extensive T-cell depletion of the graft to prevent GvHD (with no post-transplant immunosuppression);
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- donor versus recipient NK cell alloreactivity to enhance anti-leukaemia effects; - NK-alloreactive donors who also carry activating KIRs to protect against infectious mortality.
References 1. Ruggeri L, Capanni M, Casucci M, et al. Role of natural killer cell alloreactivity in HLAmismatched hematopoietic stem cell transplantation. Blood 1999; 94: 333-339. 2. Ruggeri L, Capanni M, Urbani E, et al. Effectiveness of donor natural killer cell alloreactivity in mismatched hematopoietic transplants. Science 2002; 295: 2097-2100. 3. Ruggeri L, Aversa F, Martelli MF, Velardi A. Haploidentical transplantation and natural killer cell recognition of missing self. Immunol Rev 2006; 214: 202-218. 4. Ruggeri L, Mancusi A, Capanni M, et al. Donor natural killer cell allorecognition of missing self in haploidentical hematopoietic transplantation for acute myeloid leukemia: Challenging its predictive value. Blood 2007; 110: 433-440. 5. Moretta L, Moretta A. Killer immunoglobulin-like receptors. Curr Opin Immunol 2004; 16: 626-633. 6. Parham P. MHC class I molecules and KIRs in human history, health and survival. Nat Rev Immunol 2005; 5: 201-214. 7. Yawata M, Yawata N, Draghi M, et al. Roles for HLA and KIR polymorphisms in natural killer cell repertoire selection and modulation of effector function. J Exp Med 2006; 203: 633-645. 8. Pando MJ, Gardiner CM, Gleimer M, et al. The protein made from a common allele of KIR3DL1 (3DL*004) is poorly expressed at cell surfaces due to substitution at position 86 in Ig domain 0 and 182 in Ig domain 1. J Immunol 2003; 171: 6640-6647. 9. Davies SM, Ruggieri L, DeFor T, et al. Evaluation of KIR ligand incompatibility in mismatched unrelated donor hematopoietic transplants. Blood 2002; 100: 3825-3827. 10.Farag SS, Bacigalupo A, Eapen M, et al. The effect of KIR ligand incompatibility on the outcome of unrelated donor transplantation: A report from the center for international blood and marrow transplant research, the European blood and marrow transplant registry, and the Dutch registry. Biol Blood Marrow Transplant 2006; 12: 876-884. 11.Cooley S, McCullar V, Wangen R, et al. KIR reconstitution is altered by T cells in the graft and correlates with clinical outcomes after unrelated donor transplantation. Blood 2005; 106: 4370-4376. 12. Giebel S, Locatelli F, Lamparelli T, et al. Survival advantage with KIR ligand incompatibility in hematopoietic stem cell transplantation from unrelated donors. Blood 2003; 102: 814-819. 13.Elmaagacli AH, Ottinger H, Koldehoff M, et al. Reduced risk for molecular disease in patients with chronic myeloid leukemia after transplantation from a KIR-mismatched donor. Transplantation 2005; 79: 1741-1747. 14.Beelen DW, Hottinger HD, Ferencic S, et al. Genotypic Inhibitory Killer Immunoglobulinlike Receptor Ligand Incompatibility Enhances the Long-term Antileukemic Effect of Unmodified Allogeneic Hematopoietic Stem Cell Transplantation in Patients with Myeloid Leukemias. Blood 2005; 105: 2594-2600. 74
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CHAPTER 3.2 • NK cells in haploidentical transplantation
15.Dawson MA, Spencer A. Successful use of haploidentical stem-cell transplantation with KIR mismatch as initial therapy for poor-risk myelodysplastic syndrome. J Clin Oncol 2005; 23: 4473-4474. 16.Leung W, Iyengar R, Turner V, et al. Determinants of antileukemia effects of allogeneic NK cells. J Immunol 2004; 172: 644-650. 17.Hsu KC, Keever-Taylor CA, Wilton A, et al. Improved outcome in HLA-identical sibling hematopoietic stem-cell transplantation for acute myelogenous leukemia predicted by KIR and HLA genotypes. Blood 2005; 105: 4878-4884. 18.Hsu KC, Gooley T, Malkki M, et al. KIR ligands and prediction of relapse after unrelated donor hematopoietic cell transplantation for hematologic malignancy. Biol Blood Marrow Transpl 2006; 12: 828-836. 19.Fernandez NC, Treiner E, Vance RE, et al. A subset of natural killer cells achieves selftolerance without expressing inhibitory receptors specific for self-MHC molecules. Blood 2005; 105: 4416-4423. 20.Smith HRC, Heusel JW, Mehta IK, et al. Recognition of a virus-encoded ligand by a natural killer cell activation receptor. Proc Natl Acad Sci USA 2002; 99: 8826-8831. 21.Martin MP, et al. Epistatic interaction between KIR3DS1 and HLA-B delays the progression to AIDS. Nature Genetics 2002; 31: 429–434. 22.Guma M, et al. Imprint of human cytomegalovirus infection on the NK cell receptor repertoire. Blood 2004; 104: 3664–3671. 23.Degli Esposti MA, Smyth MJ. Close encounters of different kinds: Dendritic cells and NK cells take centre stage. Nature Reviews Immunology 2005; 5: 112-124. 24.Aversa F, Tabilio A, Velardi A, et al. Treatment of high risk acute leukemia with T-celldepleted stem cells from related donors with one fully mismatched HLA haplotype. N Engl J Med 1998; 339: 1186-1193.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which molecules are recognised by inhibitory KIRs? a) HLA class II molecules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) HLA-E . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) HLA class I molecules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All of the above . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which of the following statements is correct in pre-clinical models and in the clinical practice of haploidentical haematopoietic transplantation? a) Donor vs. recipient alloreactive NK cells cause GvHD. . . . . . . . . . . . . . . . . . . . . . .
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b) Donor vs. recipient alloreactive NK cells increase the incidence of leukaemia relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Donor vs. recipient alloreactive NK cells mediate a GvL effect . . . . . . . . . . . . d) Donor vs. recipient alloreactive NK cells cause rejection . . . . . . . . . . . . . . . . . . . 3. From the data in ref. 4 of this Chapter, the event-free survival of acute myeloid leukaemia patients transplanted in any remission from haploidentical NK alloreactive donors is: a) 30-40% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 40-50% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 50-60% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 60% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. From the data in ref. 4 of this Chapter, the event-free survival for acute myeloid leukaemia patients transplanted in chemo-resistant relapse from NK alloreactive donors is: a) 10% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 10-20% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 20-30% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 30% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. From the study by Mancusi et al. reported in this Chapter, transplantation from NK alloreactive donors who also possess activating KIR genes is associated with: a) Decreased incidence of leukaemia relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Increased incidence of GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Decreased incidence of infectious mortality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Increased incidence of infectious mortality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 3
Immunogenetics of allogeneic HSCT
*
3.3
Non-HLA immunogenetics and role in transplant outcome A.M. Dickinson
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CHAPTER 3.3 • Non-HLA immunogenetics
1. Introduction While HLA typing remains the central means of selecting donors and the major factor determining haematopoietic stem cell transplant (HSCT) outcome, the sequencing of the human genome has brought to light myriads of single nucleotide polymorphisms whose significance in determining an individual’s immunological phenotype and their possible role in influencing the outcome of an allogeneic stem cell transplant is only just beginning to be explored. This review discusses the field of non-HLA immunogenetics and the patient and/or donor genotypes which may influence transplant outcome, including occurrence and severity graft versus host disease (GvHD), infectious episodes and overall survival (OS). Results associated with polymorphisms of cytokine and immune-related genes, genes of uncertain function, and minor histocompatibility antigens will be summarised.
2. Cytokine gene polymorphisms (CGPs) Cytokine gene polymorphisms occurring within the 5’ or 3’ regulatory sequences of genes may alter the structure of the transcription factor binding sites within gene promoters and therefore alter the amount of cytokine produced, for example, upon allogeneic stimulation or infection. Many of the reported cytokine gene polymorphisms occur within apparent regulatory regions of the gene. Within normal populations high or low producers of cytokines naturally exist due to the inherited gene polymorphisms. Initial studies within the solid organ transplant setting demonstrated that patients with high-producer tumour necrosis factor (TNF) and low-producer interleukin 10 (IL-10) genotypes were more likely to reject their solid organ graft. These studies were extended to the HLA matched sibling and matched unrelated donor (MUD) HSCT. A number of cytokine gene polymorphisms have now been associated with GvHD and/or transplant outcome. The majority of the studies have been carried out in single centre series of HLA matched sibling transplants and some MUD transplant patients. 2.1. Influence of CGPs on GvHD and post-HSCT complications Acute graft versus host disease (aGvHD) incidence following allo-HSCT between HLA identical siblings is 30–80% and can be fatal in up to 50% of cases. GvHD is induced by release of pro-inflammatory cytokines IL-1, IL-6, IL-8, and TNF-a during the "cytokine storm" following both radiotherapy and chemotherapy conditioning regimens. This is amplified by the activation of transplanted donor T-cells via reaction to upregulated HLA and adhesion molecules on recipient target cells. Tissue damage then ensues from activated T-cells and NK cells and release of predominantly Th1 type cytokines (IL-2, IFN-g, TNF-a).
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Although T-cells are central to the initiation of GvHD, pro-inflammatory mediators such as IL-1 and TNF-a are capable alone of inducing the pathological changes of GvHD. Growing evidence suggests that inflammatory cytokines are also involved in transplant outcomes other than GvHD such as interstitial pneumonia and VOD. The impact of gene polymorphisms on the susceptibility to infection and drug metabolism or transport (pharmacogenomics) post-transplant is now being identified. Taken together it is now evident that the genetic make up of the recipient and the donor can strongly influence the success or failure of HSCT. The CGPs studied for association with outcome in allo-HSCT, to date, are summarised in Table 1.
Table 1: Non-HLA gene polymorphisms and their associations with GvHD and HSCT outcome CGP
Proposed function of polymorphism
Effect of recipient genotype (RG) or donor genotype(DG) on HSCT outcome
TNFd3 (a)
- Pro-inflammatory - Upregulates TNF-a production
TNF receptor TNFRII 196R (b)
- Unknown (TNFRII receptor stimulates T-cell proliferation and allo-immune responses)
IL-10-1064(12-15) (c) IL-10 haplotype
- Anti-inflammatory - Haplotype associated with decreased production of IL-10
IL-6-174 (d)
- Pro-inflammatory - G allele associated with increased IL-6 production - Pro-inflammatory - Lower in vitro IFN-g production - Pro-inflammatory
RG: Increases aGvHD in HLA matched sibling BMT. No association in CBT DG: More severe GvHD in MUD transplant RG acute/DG chronic Incidence of acute and chronic GvHD in HLA-matched siblings RG: Increase in GvHD in BMT RG & DG associated with aGvHD No association in CBT RG: Increases acute and chronic GvHD in HLA-matched sibling cohorts RG: Increased aGvHD in HLAmatched sibling cohorts
IFN-g (e) Intron 1 allele IL-1 gene family IL-1a 889; intron 6 VNTR (f) IL-1 Ra VNTR VNTR intron 2 (g)
- Anti-inflammatory - Down-regulates proinflammatory effects of IL-1
DG: associates with cGvHD in HLA-matched siblings Improved OS decreases TRM; DG or RG in MUD transplants DG decreases aGvHD and RG increases cGvHD in HLAmatched siblings continue
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CGP
Proposed function of polymorphism
Effect of recipient genotype (RG) or donor genotype(DG) on HSCT outcome
TGF-b TGF-509 (h)
- C allele at –509 associated with increased TGF-b production
RG: No association with TGF-b –509 with incidence of GvHD or outcome in HLA-matched sibling cohort. Also association with GvHD incidence and high TGF-b producers in paediatric HLA-matched sibling cohort
TGF-b1 codon 10 leucine/proline
Alteration in protein structure
Development of aGvHD in MUD paediatric transplants
NOD2/CARD15 SNP 8, 12, 13 (i)
Altered NFK-b function
Two or more variants associate with severe GvHD and TRM, HLA mismatched siblings and MUD
(a) (5, 6, 10, 25); (b) (7, 8); (c) (5, 6, 9, 10, 25, 26); (d) (6, 26); (e) (27); (f) (28, 29); (g) (30, 31); (h) (27, 32–34); (i) (17–19)
Both the patient and donor genotype are analysed with respect to transplant outcome, and the main findings are reviewed below and in recent reviews (1–4). 2.1.1. TNF genes The TNF-a gene is located within the class III region of the major histocompatibility complex (MHC). Therefore, in HLA-identical sibling HSCT recipient and donor genotype will be identical and may equally or additively affect TNF production and transplant outcome. Initial HSCT studies associated recipient genotype (d3/d3) of the TNF-d microsatellite with increased severity of aGvHD (grade III-IV) in CsA-alone treated HLA-matched sibling transplants (5). A larger cohort of transplants that received cyclosporine plus methotrexate prophylaxis demonstrated an association of recipient TNFd/d3 genotype with increased mortality (6). A Japanese MUD transplant study described an association of the TNF-863 and 857 polymorphisms in donors and/or recipients with a higher incidence of GvHD grade III-IV and a lower rate of relapse (7), however in HLA-A, -B and -DRb1 matched pairs only GvHD outcome remained significant. Studies have also investigated the role of the TNF-a (-308) and TNF-b (+1069) in HSCT and GvHD, but results have been inconsistent. A role of TNF-a-308 in HLA-matched
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sibling transplants was initially based on TNF-a secretion studies, not confirmed in larger cohorts. These polymorphisms, TNF-a and/or TNF-b, have been associated with toxic complications in both HLA matched sibling and MUD cohorts. TNF-a*2 (associated with higher TNF-a production) is associated with the TNF-b*1 allele contributing to more severe toxic complications than TNF-b* or TNF-a*1 homozygotes. Mullighan et al. (3) also analysed the TNF-488A allele in two independent cohorts of HLA matched sibling transplants, and this allele was associated with more severe aGvHD and cGvHD and early death. TNF-a polymorphisms are in linkage disequilibrium with HLA Class I and Class II genotypes within the MHC Class III region. Since HLA genotypes are known to influence GvHD development, associations with TNF-a genotype may interact or have additive effects with the HLA associations (3). A polymorphism (196 R/M) in the TNF receptor II gene in MUD HSCT found that recipients of TNFRII-196R-positive donors had a higher incidence of severe GvHD and a lower rate of relapse than from TNFRII 196M homozygous donors (7). Recipient TNFRII 196R allele was associated with aGvHD incidence but the TNFRII 196RR genotype in the donor was associated with (increased) incidence of extensive cGvHD in an HLA-matched sibling cohort (8). 2.1.2. IL-10 gene The SNPs and microsatellites of the IL-10 gene resolve into several conserved haplotypes. Three common haplotypes of the promoter region between -1082 and -592 represent high, intermediate, and low production of IL-10. The low producer (ACC) haplotype in the recipient associated with severe aGvHD grade III-IV in CsAalone (6) and CsA + MTx (methotrexate) (7) treated HLA-matched sibling cohorts. IL-10 low producer haplotype associations have recently been confirmed as playing a role in predicting outcome and GvHD (grades III-IV) HLA matched sibling transplants in two separate large (>400) cohorts (9). A recent study by Bettens et al. associated the IL-10-1064CA microsatellite with greater than 12 recipients and linked with the low producer IL-10-1082*A allele with worse survival (10). 2.1.3. IL-6 gene Possession of the G allele (IL-6-174 polymorphism) in the recipient (HLA-matched sibling transplant) associated with both acute and chronic GvHD, this allele has also been correlated with high serum IL-6 levels in normal individuals. 2.1.4. IFN-g gene Allele 2 of the CA repeat microsatellite in intron 1 has been associated with high 82
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in vitro IFN-g production, whereas allele 3 has been linked to lower IFN-g production. The possession of allele 2 within the 874A allele coincides with the NFK-b binding site which may give rise to changes in the IFN-g gene transcription. Lack of IFN-g can lead to accelerated acute GvHD in IFN-g knockout mouse models. Possession of allele 3 in the recipient genotype in HLA-matched sibling transplants was associated with development of aGvHD and increased risk of EBV infections. Other studies have shown association with lack of the 2/2 genotype with GvHD (11, 12). 2.1.5. IL-1 gene family IL-1 receptor antagonist (IL-1Ra) binds IL-1 and can lead to induction of other cytokines. HLA-matched siblings transplant studies have demonstrated an association of the IL-Ra VNTR (allele 2) (which downregulates IL-1 production) in the donor genotype with less severe aGvHD, and in the recipient genotype with cGvHD. Carriage of allele 2 (donor genotype) in either the VNTR or -889 polymorphisms of IL-1a gene associated with cGvHD. A study of paediatric MUD transplant group found that IL-1a-889 in either donor or recipient was associated with improved survival and decreased TRM, but not with GvHD. IL-1 polymorphisms and association with HSCT outcome has recently been reviewed by Cullup and Stark (13). 2.1.6. TGF-b gene Several studies have reported association of TGF-b genotype with GvHD. A polymorphism (-509 C/T) in the promoter region is associated with variation in plasma concentration of TGF-b (C allele with higher production), and amino acid substitutions at codons 10 (leuÆpro) and 25 (argÆpro), which alters protein structure. A small study on HLA-identical sibling BMT showed no association of the TGF-b-509 polymorphism with either GvHD or outcome. However a study in 67 paediatric patients showed an association with the TGF-b1 codon 10 polymorphism in the donor genotype and development of aGvHD. TGF-b1 receptor II polymorphism (1167 C/T) genotype in the recipients in the same study was also associated with the development of GvHD.
3. Minor histocompatibility antigens and transplant outcome GvHD occurring after HLA matched sibling transplants is initiated by T-cells recognising minor histocompatibility antigens (mHag) (peptides derived from intracellular proteins of restricted polymorphisms coded by autosomal or Y chromosome genes, presented by HLA molecules) and are polymorphic proteins with differences between patient and donor (reviewed in (14)). T-cell clones recognising mHags were initially isolated after BMT in patients developing GvHD or graft
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rejection. The male specific mHags encoded by the Y chromosome have been shown to be involved in HLA-matched sex mismatched HSCT. The tissue expression of some mHag (e.g., HA-1 and HA-2) is limited to the haematopoietic system whereas other mHag (e.g., H-Y, HA-3) are ubiquitously expressed on normal tissues. Mismatches between patient and donor for HA-1, HA-2, 4, and 5 are associated with increased GvHD incidence. The precise characterisation of the peptide sequence of haematopoietic tissue-restricted or cancer cell-restricted mHa make them ideal targets for immunotherapy mHag specific DLI has led to complete remission of relapsed leukaemia in 3 patients, mHag HA-1 is also expressed on epithelial cancer cells and not on normal epithelial cells giving rise to the concept of the use of mHag in more diverse cancer therapies.
4. Other non-HLA-encoded genes implicated in transplant related complications Other non-HLA encoded genes have also been implicated not only in GvHD but also in TRM, infectious episodes and outcome in HLA-matched sibling transplants. The steroid hormone receptor supergene family includes the estrogen receptor and the vitamin D receptor (VDR), both have marked effects on the development of the immune system and variations in VDR are associated with autoimmune and immune dysfunctional diseases. Vitamin D analogues can prolong graft survival and prevent GvHD in animal studies. Polymorphism in the VDR intron 8 region (15), and in intron 1 of the estrogen receptor alpha (ERa) genes have been associated with both occurrence of GvHD, post-transplant infection and likelihood of survival following allo-BMT. Donor and recipient genotype in genes that regulate the host response to microorganisms (myeloperoxidase (MPO), mannose binding lectin (MBL) and Fcg receptors (FcgRIIa, IIIa, IIIb)) have been associated with infections after BMT. First infection post-transplant has been associated with the R-131 allele in the patient FcgRIIa genotype and occurrence of severe bacterial infections increased when MPO donor genotype was AG or AA. TRM was influenced by FcgRIIIb genotype and donor MPO genotype (see Table 2). These studies further define and improve understanding of the mechanisms involved in host defence against infection during BMT. 4.1. Innate immunity Investigations into other non-cytokine genes, associated with innate immunity, have been studied including pattern recognition receptors (PRRs) which activate the innate and adaptive arms of the immune response against pathogens. PRRs include the transmembrane receptors, Toll-like receptors (TLRs), which have the ability to activate antigen presenting cells (APCs) (16, 17). Polymorphisms in the TLR4 gene, 84
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Table 2: Non-HLA genes associated with risk of infections and TRM Gene
Alleles
Effect
R or D genotype
MPO (a)
AG/AA
D
FcgRIIa (a)
R-131
FcgRIIIb (a)
HNA1a/HNA1a
Increased bacterial infection Increased risk of non-leukaemic death Increased first overall infections (infections occurring before d 80) Delayed neutrophil recovery Severe infection Increased risk of non-leukaemic death Increased TRM rates Quick neutrophil recovery Decreased infection Increased risk of major infection
MBL (b)
HNA1b/HNA1b HNA1a/HNA1b HYA haplotype MBL2
R
D
D D/R D/R
(a) (31); (b) (35). MPO: myeloperoxidase, an enzyme found primarily in the lysosomes of neutrophils. A single base substitution (G > A) in the promoter region of the MPO gene (–463) decreases expression and alters binding site for the transcription factor; FcgRIIa: Fcg receptor, polymorphisms of the receptor affect receptor affinity and specificity; FcgRIIIb: Fcg receptor HNA1a and HNA1b, isoforms affect different ligand binding of antibodies; MBL: mannose-binding lectin
for example, have been studied and associated with hypo-responsiveness to lipopolysaccharides with a reduced risk of aGvHD, but increased risk of Gramnegative bacteraemia. Polymorphisms in the TLR1 and TLR6 genes have been associated with invasive aspergillosis following HSCT; patient genotype being associated with increased risk of infection. Intracellular PRRs associated with inflammatory bowel disease, such as the intracellular group of NOD-like receptors (nucleotide-binding oligomerisation domain containing receptors) have also been studied in cohorts of HSCT patients. There are three SNPs in NOD2/CARD15 (SNP8, SNP12, SNP13) gene, involved in defective NF-kB responses and possession of two or more NOD2/CARD15 variants has been associated with more severe GvHD and increased transplant-related mortality (TRM) (18). Further studies showed that in HLA-matched siblings the genetic risk could be modulated depending on the type of gastrointestinal decontamination pre-transplantation (17). In T-cell-depleted transplants, possession of NOD2/CARD15 SNPs associated with lower disease-free survival (19) and the results illustrate that altered immune responses associated with NOD gene variants may be dependent on type of immunomodulation, e.g. bacterial decontamination and by the type of immunosuppression (e.g. T-cell depletion).
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5. Pharmacogenomics in HSCT Knowledge of an individual’s drug response has the potential to improve HSCT outcome, and several polymorphisms of genes involved in the metabolism of drugs employed in conditioning regimens and prophylaxis strategies have been associated with GvHD, survival, toxicities and platelet recovery following HSCT (3). A SNP (C677T) associated with lower methylenetetrahydrofolate reductase (MTHFR) activity was linked to increased mucositis and delayed platelet engraftment in HSCT recipients receiving methotrexate, but results have not been consistent. These findings were confirmed by Robien et al. (20), but, Kalayoglu-Besisik et al. (21) found no associations with HSCT outcome. Other studies have involved HLA matched siblings and MUD transplant cohorts with differing results.
6. Gene associations with transplant outcome – Problems of interpretation The cytokine gene polymorphisms examined in our initial studies were selected on the basis of their association with altered cytokine production in vivo or their demonstrated association with transplant-related pathology, and or their association with immune dysfunction/autoimmune diseases. Typically, alleles linked with increased expression of pro-inflammatory cytokines such as TNF-a, IL-6, and IFNg show associations with complications such as GvHD, whereas alleles associated with increased expression of anti-inflammatory cytokines (IL-10 and IL-1Ra) show protection effects (see Table 1). However, as more studies are reported this model appears too simple to account for all the findings. Several problems surround the interpretation of the role of cytokine variations in transplant outcome. HLA association: In HLA matched sibling transplants HLA-A3 is associated with a higher and DR1 with a lower risk of GvHD. These associations may represent HLA types best able to present specific antigens relevant to disease processes. Alternatively, the effect on GvHD may reflect Class III region TNF polymorphisms forming an extended haplotype with the HLA Class I and Class II region. Determining what precisely occurs at this locus is therefore complex. In the MUD transplant setting the role of HLA will be of a paramount importance in interpretation of non-HLA immunogenetic data. High resolution tissue typing and standardisation of results is necessary across BMT centres for comparative studies. GvHD increases proportionally with the degree of HLA Class I and II disparity between patient and donor. Mismatching for HLA-C alleles increases the risk of graft failure, GvHD and mortality. Disparities between HLA sequence polymorphisms detected by serology are termed antigen mismatches; those identified only by DNA-based typing are termed allele mismatches. The risk of mortality after MUD transplants is increased by mismatches at a single allele at HLA A B C or DRB1. The significance of these mismatches must 86
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be taken into account prior to further analysis of the data for the role of non-HLA immunogenetics in the MUD transplant setting. 6.1. Population differences in gene frequencies In vivo disease association may not necessarily correlate with in vitro cytokine expression or production. This could be due to genetic variation in allele frequencies between different ethnic populations resulting in different associations being found. Therefore, comparisons between populations are difficult unless local population allele frequency is simultaneously assessed and are taken into account. For example, IL-10 GCC haplotype in Japan is of a very low frequency compared to Europe. One study compared the incidence of aGvHD and TRM in HLA identical sibling transplants within the Japanese, white and African Americans, Scandinavians and the Irish (22), and showed differences between the American groups and the Irish compared to the Japanese or Scandinavians. The results indicate that genetic differences other than HLA may affect transplant outcome, and several studies on allele frequencies in transplant populations worldwide are currently being undertaken. 6.2. Different transplant procedures Analysis of the complex data emerging on the role of non-HLA immunogenetics on stem cell transplant outcome is compounded by a number of factors including the heterogeneity of the diagnoses and clinical state of the recipients, GvHD prophylaxis regimes, and conditioning strategies. Most of the research has been carried out within the HLA-matched sibling setting. Larger studies are now needed together with multivariate analysis in both HLA-matched siblings and matched unrelated donor transplants, especially those undergoing homogeneous clinical trials.
7. Future developments Current clinical studies are aimed at the development of a clinical and genetic risk score which will extend the predictive power of genetic polymorphisms. To this aim large cohorts of HLA-matched and MUD transplants need to be complied to fully investigate the impact of non-HLA immunogenetics on transplant related complications, and this is underway both at the European and International level including involvement of the International Bone Marrow Transplant Registry. Combined analysis of CGPs suggest that a risk index for GvHD could be developed alongside clinical risk factors such as gender, age, CMV status, and minor histocompatibility mismatches. Combining the EBMT risk score for CML with genetic factors has led to a scoring system which can further subdivide the patients for good, intermediate and poor prognosis (23). Further research into the possible
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role of pharmacogenomics (24) in determining outcome may allow risk prediction to tailor prophylaxis and therapy for GvHD on a disease specific or individual patient basis.
Acknowledgments Funding for current research on non-HLA gene polymorphisms is from European Commission grants TRANS-NET (MRCN-CT-2004-512253) and StemDiagnostics (LSHB-CT-2007-037703), the Leukaemia Research Fund and the Tyneside Leukaemia Research Association.
References 1. Dickinson AM. Risk assessment in haematopoietic stem cell transplantation: pre-transplant patient and donor factors: non-HLA genetics. Best Pract Res Clin Haematol 2007; 20: 189207. 2. Dickinson AM, Charron D. Non-HLA immunogenetics in hematopoietic stem cell transplantation. Curr Opin Immunol 2005; 17: 517-525. 3. Mullighan CG, Bardy PG. Advances in the genomics of allogeneic haemopoietic stem cell transplantation. Drug Development Research 2004; 62: 273-292. 4. Dickinson AM, Harrold J, Cullup H. Haematopoietic stem cell transplantation: Can our genes predict clinical outcome? Expert Rev Mol Med 2007; 9: 1–19. 5. Middleton PG, Taylor PRA, Jackson G, et al. Cytokine Gene polymorphisms associating with severe acute graft-versus-host disease in HLA-identical sibling transplants. Blood 1998; 92: 3943-3948. 6. Cavet J, Middleton PG, Segall M, et al. Recipient tumour necrosis factor-alpha and interleukin-10 gene polymorphisms associate with early mortality and acute graft-versushost disease severity in HLA matched sibling bone marrow transplants. Blood 1999; 94: 3941-3946. 7. Ishikawa Y, Kashiwase K, Akaza T, et al. Polymorphisms in TNFA and TNFR2 affect outcome of unrelated bone marrow transplantation. Bone Marrow Transplant 2002; 29: 569-575. 8. Stark GL, Dickinson AM, Jackson GH, et al. Tumour necrosis factor receptor type II 196M/R genotype correlates with circulating soluble receptor levels in normal subjects and with graft-versus-host disease after sibling allogeneic bone marrow transplantation. Transplantation 2003; 76: 1742-1749. 9. Lin MT, Storer B, Martin PJ, et al. Relation of an interleukin-10 promoter polymorphism to graft-versus-host disease and survival after hematopoietic-cell transplantation. N Engl J Med 2003; 349: 2201-2210. 10.Bettens F, Passweg J, Gratwohl A, et al. Association of TNFd and IL-10 polymorphisms with mortality in unrelated hematopoietic stem cell transplantation. Transplantation 2006; 81: 1261-1267. 88
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11.Bogunia-Kubik K, Mlynarczewska A, Wysoczanska B, et al. Recipient interferon-gamma 3/3 genotype contributes to the development of chronic graft-versus-host disease after allogeneic hematopoietic stem cell transplantation. Haematologica 2005; 90: 425-426. 12.Mlynarczewska A, Wysoczanska B, Karabon L, et al. Lack of IFN-gamma 2/2 homozygous genotype independently of recipient age and intensity of conditioning regimen influences the risk of aGVHD manifestation after HLA-matched sibling haematopoietic stem cell transplantation. Bone Marrow Transplant 2004; 34: 339-344. 13.Cullup H, Stark G. Interleukin-1 polymorphisms and Graft versus Host Disease. Leuk Lymphoma 2005; 46: 517-523. 14.Goulmy E. Human minor histocompatibility antigens. Curr Opin Immunol 1996; 8: 75-81. 15.Middleton PG, Cullup H, Dickinson AM, et al. Vitamin D receptor gene polymorphism associates with graft-versus-host disease and survival in HLA-matched sibling allogeneic bone marrow transplantation. Bone Marrow Transplant 2002; 30: 223-228. 16.Napolitani G, Rinaldi A, Bertoni F, et al. Selected Toll-like receptor agonist combinations synergistically trigger a T helper type 1-polarizing program in dendritic cells. Nature Immunol 2005; 6: 769-776. 17.Holler E, Rogler G, Brenmoehl J, et al. Prognostic significance of NOD2/CARD15 variants in HLA-identical sibling hematopoietic stem cell transplantation: Effect on long term outcome is confirmed in 2 independent cohorts and may be modulated by the type of gastrointestinal decontamination. Blood 2006; 107: 4189-4193. 18.Holler E, Rogler G, Herfarth H, et al. Both donor and recipient NOD2/CARD15 mutations associate with transplant-related mortality and GvHD following allogeneic stem cell transplantation. Blood 2004; 104: 889-894. 19.Granell M, Urbano-Ispizua A, Arostegui JI, et al. Detrimental effect of NOD2/CARD15 polymorphisms in T-cell depleted allogeneic stem cell transplantation. Bone Marrow Transplant 2006; 37: S79; P431. 20.Robien K, Ulrich CM, Bigler J, et al. Methylenetetrahydrofolate reductase genotype affects risk of relapse after hematopoietic cell transplantation for chronic myelogenous leukemia. Clinical Cancer Res 2004; 10: 7592-7598. 21.Kalayoglu-Besisik S, Caliskan Y, Sargin D, et al. Methylenetetrahydrofolate reductase C677T polymorphism and toxicity in allogeneic hematopoietic cell transplantation. Transplantation 2003; 76: 1775-1777. 22.Oh H, Loberiza FR, Jr., Zhang MJ, et al. Comparison of graft-versus-host-disease and survival after HLA-identical sibling bone marrow transplantation in ethnic populations. Blood 2005; 105: 1408-1416. 23.Dickinson AM. A Distinct Pattern of Non-HLA Polymorphisms Predicts an Increased Risk for GvHD without Benefit of GvL in HLA Matched Sibling Transplants for Chronic Myeloid Leukemia (CML). BMT Tandem meeting, Keystone, Colorado 2007 (Abstract). 24.Leather HL. Drug interactions in the hematopoietic stem cell transplant (HSCT) recipient: what every transplanter needs to know. Bone Marrow Transplant 2004; 33: 137-152. 25.Kogler G, Middleton PG, Wilke M, et al. Recipient cytokine genotypes for TNF-alpha and
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IL-10 and the minor histocompatibility antigens HY and CD31 codon 125 are not associated with occurrence or severity of acute GVHD in unrelated cord blood transplantation: a retrospective analysis. Transplantation 2002; 74: 1167-1175. 26.Socié G, Loiseau P, Tamouza R, et al. Both genetic and clinical factors predict the development of graft-versus-host disease after allogeneic hematopoietic stem cell transplantation. Transplantation 2001; 72: 699-706. 27.Cavet J, Dickinson AM, Norden J, et al. Interferon-gamma and interleukin-6 gene polymorphisms associate with graft-versus-host disease in HLA-matched sibling bone marrow transplantation. Blood 2001; 98: 1594-1600. 28.Cullup H, Dickinson AM, Cavet J, et al. Polymorphisms of interleukin-1alpha constitute independent risk factors for chronic graft-versus-host disease after allogeneic bone marrow transplantation. Br J Haematol 2003; 122: 778-787. 29.MacMillan ML, Radloff GA, DeFor TE, et al. Interleukin-1 genotype and outcome of unrelated donor bone marrow transplantation. Br J Haematol 2003; 121: 597-604. 30.Cullup H, Dickinson AM, Jackson GH, et al. Donor interleukin 1 receptor antagonist genotype associated with acute graft-versus-host disease in human leucocyte antigen-matched sibling allogeneic transplants. Br J Haematol 2001; 113: 807-813. 31.Rocha V, Franco RF, Porcher R, et al. Host defense and inflammatory gene polymorphisms are associated with outcomes after HLA-identical sibling bone marrow transplantation. Blood 2002; 100: 3908-3918. 32.Grainger DJ, Heathcote K, Chiano M, et al. Genetic control of the circulating concentration of transforming growth factor type beta 1. Human Mol Gen 1999; 8: 93-97. 33.Tambur AR, Yaniv I, Stein J, et al. Cytokine gene polymorphism in patients with graftversus-host disease. Transplant Proc 2001; 33: 502-503. 34.Hattori H, Matsuzaki A, Suminoe A, et al. Polymorphisms of transforming growth factorbeta1 and transforming growth factor-beta1 type II receptor genes are associated with acute graft-versus-host disease in children with HLA-matched sibling bone marrow transplantation. Bone Marrow Transplant 2002; 30: 665-671. 35.Mullighan CG, Heatley S, Doherty K, et al. Mannose-binding lectin gene polymorphisms are associated with major infection following allogeneic hemopoietic stem cell transplantation. Blood 2002; 99: 3524-3529.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which cytokine gene polymorphism has most consistently been associated with GvHD? a) TNF . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) IL-10 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90
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c) IL-1 gene family . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) TGF-b . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2. Identify genes of the innate immune system a) HA-1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) TGF-b . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) NOD2/CARD15 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) HLA-A3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. What outcome of the transplant procedure may be influenced by the non-HLA genetics of the patient and donor? a) Transplant related mortality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Incidence and severity of GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Survival . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All three . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. What are the potential future developments of studying non-HLA genetics? a) Improve donor selection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Aid in design of new therapeutic regimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Development of a prognostic risk score for individual patients . . . . . . . . . . . . d) All of a), b) & c) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5. In MUD and sibling transplants what are the considerations to be taken into account in studying non-HLA immunogenetics? a) HLA matching and linkage disequilibrium between HLA and cytokine genes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Clinical risk factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Population genetics and allele frequency . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All three . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
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JACIE accreditation of transplant centres
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1. Introduction JACIE (The Joint Accreditation Committee of ISCT-Europe and EBMT) is a non-profit body established for the purposes of assessment and accreditation in the field of haematopoietic stem cell transplantation (HSCT). The Committee was founded in 1998 by the European Group for Blood and Marrow Transplantation (EBMT) and the International Society for Cellular Therapy (ISCT), the two leading scientific organisations involved with HSCT in Europe (1). JACIE modelled itself on the USbased Foundation for the Accreditation of Cellular Therapy (FACT), established in 1996 by the ISCT and the American Society for Blood and Marrow Transplantation (ASBMT). JACIE actively collaborates with FACT in establishing standards for the provision of quality medical and laboratory practice in HSCT, and the two organisations now issue joint FACT-JACIE standards, which are applicable internationally (2, 3). The primary aim of JACIE is to improve the quality of HSCT in Europe by providing a means whereby transplant centres, HSC collection facilities and processing facilities can demonstrate high quality practice. This is achieved through external inspection of facilities to ensure compliance with the FACT-JACIE standards. An additional and wider aim is to ensure harmonisation between FACT-JACIE standards and other national/international standards, including the EU Tissues & Cells Directive (Directive 2004/23/EC) and the related implementing Commission Directives 2006/17/EC and 2006/86/EC (4–6). JACIE accreditation is voluntary, but provides a means whereby transplant facilities can demonstrate that they are working within a quality system covering all aspects of the transplantation process and thus show compliance with the requirements of insurance companies or national and/or international regulatory authorities. The current organisation of JACIE is shown in Figure 1. The structure ensures wide consultation with over 20 European countries now represented on the Board in addition to nursing, paediatrics and cord-blood representatives. In 2007 an Accreditation Committee was established which reviews all inspection reports and advises the JACIE Board. Accreditation of HSC transplant facilities is through on-line submission of documentation and an on-site visit by a team of trained inspectors. Centres may apply for accreditation as complete programmes comprising clinical programme, collection facility and a processing laboratory or, for example, as a single collection or processing facility which may serve a number of clinical programmes.
2. The FACT-JACIE standards The FACT-JACIE standards cover all aspects of clinical transplant programmes, collection facilities (BM collection and PBPC collection) and processing of HSC, as
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Figure 1: Structure of JACIE JACIE Board Executive Committee
JACIE Office
EBMT - ISCT
Accreditation Committee
JACIE BOARD Officers (JACIE Executive Committee) President; Vice-President; Medical Director; Accreditation Committee Chairman; Executive Officer Sector Representatives NETCORD; Nursing; Paediatric National Representatives Currently: Austria; Belgium; Czech Republic; Denmark; Estonia; Finland; France; Germany; Greece; Hungary; Italy; The Netherlands; Norway; Poland; Slovakia; Slovenia; Spain; Sweden; Switzerland; Turkey; United Kingdom (Elected by National Transplant Societies) JACIE ACCREDITATION COMMITTEE Accreditation Committee Chairman; Medical Director; 1-2 Clinical Representatives; 1-2 Collection Representatives; 1-2 Processing Representatives. (Appointed by the Executive Committee from a pool of experienced inspectors) JACIE OFFICE Executive Officer; Administrative Officer
shown in Table 1. The standards also apply to the use of therapeutic cells (TC) derived from blood or marrow, including donor lymphocytes and MSCs. The standards cover the clinical use of Cord Blood HSC by clinical programmes but not the collection of banking of cord blood, which are covered by the related Netcord-FACT standards and inspected/accredited by FACT-Netcord (7, 8). Within each subsection of the standards are detailed lists of specific requirements; for example the standard on donor evaluation and selection contains over 35 specific items relating to clinical evaluation, laboratory testing, informed consent etc. The current edition of the standards is the 3rd edition, dated 2007. The 4th edition is currently in preparation. 94
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Table 1: Contents of the FACT-JACIE standards (3rd edition 2007) Clinical programme
Collection (BM and PBPC)
Processing
General - including programme organisation and transplant activity
General
General
Facilities - inpatient and outpatient
Facilities – theatre/apheresis facilities
Facilities – laboratory size and organisation
Staffing – qualifications and training – medical, nursing and other staff
Staffing – qualifications and training
Staffing – qualifications and training
Quality Management Programme – including adverse event reporting, audit and outcome review
Quality Management Programme – including adverse event reporting, audit, validation of procedures and equipment, and outcome review
Quality Management Programme – including adverse event reporting, audit, validation of procedures and equipment, and outcome review
Policies and Procedures (SOPs) – requirements for SOPs including format, document control, lists of specific areas to be covered
Policies and Procedures – as left
Policies and Procedures – as left
Donor evaluation, selection and Donor evaluation/care at time testing – allogeneic and of collection autologous
Process Controls – processing techniques, testing, assays, validation
Administration of high dose therapy and infusion of HSC
Labelling-specific requirements Labelling-specific requirements for labelling and documentation for labelling and documentation during and after collection during and after processing and at distribution
Clinical research
Collection procedure (BM or PBPC) – including paediatrics donors
Distribution and Release – review of products prior to release, documentation to be sent with product
Data management – including Storage – conditions for accuracy of recording transplant storage data
Storage – conditions for storage including temperature control, alarm systems
Records – departmental and medical
Transport – within facility and/or to external facility
Receipt and transportation – acceptance, quarantine, transport of fresh/frozen products
Records – including electronic records
Disposal – including consent, documentation
Direct Distribution to Clinical Programme – requirements for record keeping
Records – including electronic records
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The standards are accompanied by a manual which contains the standards together with detailed guidance on the interpretation and measures required to demonstrate compliance. Each standard is followed by specific questions relating to that standard and these questions form the basis of the inspection checklist, which must be completed prior to inspection by the applicant centre and verified by the inspector during the inspection (Figure 2). The complete standards and the accompanying guidance manual are available on the JACIE website, together with other useful information relating to inspection and accreditation (3).
Figure 2: Example from checklist INSPECTION CHECKLIST
APPLICANT
B6.000 D O N O R E V A L U A T I O N , SELECTION AND MANAGEMENT
Y
N
INSPECTOR
N/A
Y
N
N/A
B6.100 Are there donor evaluation procedures in place to protect the safety of the haematopoietic progenitor cell donor D* and recipient? Do these evaluation procedures assess the potential for disease transmission D from the donor to the recipient? Do these evaluation procedures assess the risks to the donor from the D collection procedure? Are donor evaluation and selection test results documented?
D
B6.110 Are there written criteria for donor evaluation and selection?
D
*D: deficiency in compliance if answer is N
2.1. General aspects of the standards Although the standards are very specific in certain areas, e.g. labelling requirements, some essential principles apply throughout. The first of these is a requirement for documentation of policies, procedures, actions, requests and so on. This does not just apply to the need to have written policies and procedures, but extends to all 96
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aspects of transplant activity. For example, the initial diagnosis of a patient must be documented by source material or reports. A request from the clinical unit to the laboratory for issue of cells must be made in writing. A potential donor must not only be properly evaluated for eligibility, but the programme must have clear written criteria for what constitutes an eligible donor and must clearly document whether the donor meets these criteria. The second is a requirement that personnel must not only be appropriately qualified, they must be trained in the procedures they regularly perform and their competency to perform the task after training must be assessed and documented. Similarly there is a requirement for validation of all equipment and procedures. Validation is a term used to describe the activity required to prove that any procedure, process, equipment, material, activity or system actually leads to the expected results. For example a new apheresis machine must be shown to produce the expected results in terms of cell yields. Also important is the requirement for close cooperation and interaction between the different parts of the programme, especially important where a clinical programme may use a distant collection and/or processing facility. Mechanisms for transfer of information e.g. regarding donor assessment and consent, results of collection, request for product issue, adverse events etc must be clearly established and often require transfer of written information. The Quality Management Programme is essential to ensure that all these aspects of the programme run smoothly. 2.2. Quality management (QM) An active quality management programme (QMP) is essential to the FACT-JACIE standards. A QMP is a mechanism to ensure that procedures are being carried out by all staff members in line with agreed standards. In a transplant programme, this ensures that the clinical, collection and laboratory units are all working together to achieve good communication, effective common work practices and increased guarantees for patients. It is a means of rapidly identifying errors or accidents and resolving them so that the possibility of repetition is minimised. It assists in training and clearly identifies the roles and responsibilities of all staff. Once the required level of quality has been achieved, the remaining challenge is to maintain this standard of practice. With a working quality management system in place and adequate resources, the fundamental elements necessary to sustain the programme are continued staff commitment and vigilance. The culture and systems for quality management are well-established in laboratories but are relatively new in clinical units and many programmes have experienced difficulty setting up a QMP to cover the clinical programme and collection facility.
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It is recommended that a programme have a dedicated quality manager, ideally with specific training in QM, not only to develop the QMP to meet the initial requirements for accreditation but to ensure continued adherence to standards. To assist centres in developing a QM programme JACIE has recently published “A practical guide to implementing quality management in a stem cell transplantation (SCT) programme”, a copy of which will be distributed to all EBMT centres, and which will be available via the JACIE website (3). This contains detailed explanation of different aspects of QM together with example scenarios and documents. The QMP must address at a minimum: - Organisational structure - Third party agreements - Process developments and review - Personnel requirements, training and competency - Outcome analysis - Audits - Management of products with positive anti-microbial culture - Detection and reporting or errors, accidents and adverse events - Record review and document control - Validation/qualification of equipment, reagents and procedures - Inventory control - Product tracking - Process controls Within a programme there can be one QM system covering all areas, i.e. clinical, collection and processing, or the laboratory may have a separate system, and other combinations are possible. If there is more than one system then there should be clear interaction between them. 2.3. Policies and procedures There are specific requirements for how policies and procedures should be written, to ensure that all essential aspects are covered. They must be available to all relevant staff at all times. Electronic access is perfectly acceptable but there must be at least one hard copy available at all times and staff must know where to access this. For each section – clinical, collection and processing – there is a list of specific policies and procedures that must be covered. These are shown in Table 2. All SOPs must be reviewed annually. 2.4. Clinical programme, collection facility and processing facility standards The following paragraphs will comments on some particular aspects of the standards, but for full details the reader should refer to the standards and manual. 98
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Table 2: List of required policies and procedures All sections
-
Donor and patient confidentiality Emergency and safety procedures Infection control, biological, chemical and radiological safety Quality Management and improvement Personnel training Competency assessment Errors, accidents and adverse events Outcome analysis Audits Corrective actions Biological product deviations Product tracking Facility maintenance and monitoring Disposal of medical and biohazard waste Disaster response
Clinical programme
-
Donor and patient evaluation, selection and treatment Donor and patient consent Infection prevention and control (clinical unit) Administration of the preparative regimen Administration of HSC Blood product transfusion
Collection facility
-
Donor treatment, screening and consent Management of paediatric donors, if applicable Product collection (BM and/or PBSC) Labelling (including forms and samples) Expiration dates Storage Release and exceptional release Transportation Reagent and supply management Equipment maintenance and monitoring
Processing facility
-
Product receipt Processing and process control Prevention of cross-contamination RBC compatibility testing Processing of ABO incompatible products Cryopreservation and thawing Labelling (including forms and samples) Expiration dates Storage Release and exceptional release Product recall Transportation Reagent and supply management Equipment maintenance and monitoring Environmental control Hygiene and use of personal protective attire
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2.4.1. Definition of a programme The standards define a Clinical Transplantation Programme as an integrated medical team housed in geographically contiguous or proximate space with a single Programme Director and common staff training programs, protocols, and quality management systems. Programmes that include non-contiguous institutions in the same area must share common protocols, staff training procedures, quality management systems, and review of clinical results and evidence of regular interaction. This means that two separate units should not combine together for the purposes of JACIE accreditation unless they are truly working together in everyday practice. Where two separate units are working together, they must be within one hours travelling time to ensure close and regular interaction. 2.4.2. Clinical programme activity To ensure continuing proficiency in a transplant programme, a minimum volume of patients must be treated per year. The current minimum activity requirements for each 12-month period are as follows: - For allogeneic transplantation: at least 10 new patients - For autologous transplantation only: at least 5 new patients - For allogeneic and autologous transplantation there is no minimum requirement for autograft numbers - For combined adult and paediatric programmes, a minimum of 5 new adult patients and 5 new paediatric patients - For programs utilising more than one clinical site for transplantation, a minimum of 5 new patients must be transplanted per site. 2.4.3. Donor evaluation This section covers information, consent, and requirements for evaluation including medical history and testing for infectious disease markers. The medical history for allogeneic donors must include a number of specific items relating to possible transmission of infections and non-infections disease. The use of a standard donor history questionnaire is recommended. Testing for infection disease markers must include: HIV-II, hepatitis B virus, hepatitis C virus and Treponema pallidum. HTLV1 and 2 testing only required as per governmental regulations, which in the EU requires that donors are tested if they are in a risk-group for infection. These tests are required for all donors, allogeneic and autologous. CMV testing is also required for allogeneic donors. The tests must be performed within 30 days prior to collection of cells. If more than one collection is performed the tests will need to be repeated if they are out of this time period. 100
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Other required tests include at least: ABO group and Rh type and pregnancy assessment for all female donors of childbearing potential. For allogeneic donors HLA-A, B, DR typing must be performed by an EFI-accredited laboratory. 2.4.4. Data collection The Programme must collect all the data contained in the EBMT MED-A forms. Prior to inspection the team must submit records of this data for 10 consecutive patients and on the day of inspection the inspectors verify the data against the source material. It is important to have records of the tests used to confirm the original diagnosis as well as of all the peri-transplant data. 2.4.5. Bone marrow collection If a programme that regularly uses bone marrow (BM), this must be collected in a facility that meets the FACT-JACIE standards. This means that the BM collection facility must also be inspected and accredited. The minimum activity required to apply for accreditation is one collection per year, with 3 collections during the 3year period of each accreditation cycle. Even where only one harvest is performed per year, all requirements of the standards must be met, e.g. relevant SOPs must be in place and regularly reviewed, with evidence of continued training of staff and review of procedures. 2.4.6. Apheresis collection The minimum activity requirement for an apheresis collection facility (CF) is 10 HSC collection procedures per year, with 30 during the 3-year period of each accreditation cycle. A CF must have a Director and a Medical Director. These can be the same person, but the Director does not have to be medically qualified. 2.4.7. Processing facility standards A CF must have a Director and a Medical Director. These can be the same person, but the Director does not have to be medically qualified. There must be appropriate and validated assays and test procedures for the evaluation of cellular therapy products. Procedures must be demonstrated to result in acceptable target cell viability and recovery. Where processing includes exposure to the environment this must take place in an environment with specified air quality and cleanliness. This means that the Director should establish through validation and testing what air quality is appropriate. The FACT-JACIE standards do not specify any defined level of air quality. However in EU member states the laboratory must comply with the requirements of the EU Directive on air quality.
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3. The inspection and accreditation process 3.1. Initial application The centre implements measures as described in the JACIE accreditation manual, and then applies for inspection by submitting basic information about the programme/facility and a number of supporting documents including a selfassessment checklist (Figure 2). The application information and checklist must be submitted in English but all other documentation, including Standard Operating Procedures (SOPs) is accepted in the language of the centre. 3.2. Inspection An on-site visit is carried out by a team of trained inspectors, usually one per facility (clinical/collection/processing). Inspectors are medical, scientific or other professional persons working in HSCT, with specific qualifications and experience for inspecting clinical, collection and/or processing facilities (9). Inspectors must attend a JACIEsponsored training course and pass an examination. Where a clinical programme performs adult and paediatric transplants, an adult and a paediatric inspector will attend. Inspectors may also be from another country but should be either native or fluent speakers of the relevant language. In countries where it is not possible to assign an inspector who speaks the language, a local expert is requested to assist with translation of interviews and documents as necessary. An inspection visit generally lasts 1.5 days and involves discussion with staff during their work, review of documents/records and completion of a detailed checklist relating to the standards. The inspectors write a report in English, noting any areas of non-compliance with the standards, which is reviewed by the JACIE Office, the medical director and the accreditation committee. 3.3. Report and correction phase Based on the inspectors’ findings, a summary report is prepared making specific recommendations for corrections and improvements. Between 3 and 12 months is allowed for the centre to correct deficiencies, depending on the amount of work required. The centre must indicate acceptance of the findings and then in due course submit documentary evidence to confirm corrections or amendments. The original inspectors review the documentation. Review by the inspectors rather than by the Accreditation Committee, MD or JACIE office alone is required because of language issues. In some cases a limited revisit may be the best way to show that deficiencies have been remedied. The inspectors confirm to JACIE that all necessary corrections have been made or indicate that there are still outstanding areas for completion. 102
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CHAPTER 4 • JACIE accreditation
3.4. Accreditation The Accreditation Committee reviews all the reports and relevant documentation and if satisfied that all previous deficiencies have been corrected, makes a recommendation to the JACIE Board that the centre be awarded accreditation. If approved, accreditation is awarded, valid for 3 years, subject to an annual report from the centre noting any significant changes in personnel or procedures and including annual activity figures. After 3 years reaccreditation is necessary, which follows exactly the same process as the initial inspection.
4. Outcome of inspections 4.1. Inspections to date To date, over 125 facilities have formally applied for accreditation. Between January 2004 and January 2008, 86 centres were inspected. Of these, the majority (70 centres) applied for accreditation for a combination of clinical, collection and processing facilities. Seven centres applied for clinical and collection only, 5 centres applied for collection and processing only, 3 for apheresis collection only, 29 for the clinical programme only, and 11 for processing only. Almost all were found to be functioning at a high level of excellence. However some deficiencies in compliance were found in all programmes inspected so far, varying from minor non-compliances, e.g. in formatting of SOPs, to major deficiencies e.g. lack of reliable alarm systems for products stored in liquid nitrogen. In almost all cases correction of deficiencies was verified by submission of documentation by the centre. The time taken for correction of deficiencies has varied from 3 to 16 months. In 3 cases a limited reinspection was required to demonstrate correction of deficiencies. At the time of reporting 45 inspected centres have completed correction of all deficiencies and achieved accreditation. These are listed on the JACIE website (3). 4.2. Common deficiencies The most common deficiencies are in documentation, labelling and in the quality management programme (Figure 3). This is consistent with the initial experience of the FACT accreditation programme in the United States (10). 4.2.1. Quality management programme (QMP) Deficiencies in QPM were by far the most common cause of failure of compliance with the standards and, including problems with policies and procedures (SOPs), accounted for 35% (201 of 570) of total cited deficiencies in the initial 2 years of inspections (11). HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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Figure 3: Common Deficiencies at Inspection Facilities Communication Safety Staffing Training QM general SOP format/review Missing SOP(s) Donor selection policy Donor evaluation Donor consent Testing
Lab accreditation Therapy Clinical-other Procedures (collection) Equipment, supplies & reagents Procedures (processing) Labelling Storage Transport Issue/disposal Records 0%
2%
4%
6%
8%
10% 12% 14% 16% 18% 20%
Areas of deficiencies as % of total deficiencies. Based on analysis of 570 deficiencies encountered in 35 inspections
Deficiencies in QMP included: - Problems with the formatting and content of SOPs, e.g. • missing examples of worksheets/forms/labels 104
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CHAPTER 4 • JACIE accreditation
missing references (where relevant) failure to include range of expected results (where relevant) Lack of procedure for documenting deviations from SOPs Lack of regular review process for SOPs No SOPs for critical procedures e.g. bone marrow collection Inadequate document control procedure Lack of validation of equipment/procedures in collection and processing facilities Inadequate audit activity e.g. • no SOP for audit • no written programme for planned audits • no documentation of results of audits • no formal process for disseminating results Inadequate adverse event (AE) reporting/reviewing. Centres often used a hospitalbased incident reporting system, but in many cases it appeared from the number of reported AEs that this was not adequate to meet the needs of the HSCT programme. Often it was not clear that all AEs were reviewed by the programme director and/or that a report was issued to the patient’s physician. • •
-
-
4.2.2. Patient/donor issues Deficiencies in documentation were frequent: - No verification of patient’s initial diagnosis from primary records - No formally documented criteria for defining a suitable donor - Failure to document vaccination history, transfusion history or travel history - Pregnancy not always assessed in female donors of childbearing age 4.2.3. Labelling of components JACIE standards for labelling of components require that essential information including a unique alphanumeric identifier, name of the product and the recipients name are shown. The label must be securely attached, of a size that allows the contents of the pack to be inspected and clearly labelled using indelible ink. In the majority of centres it was found that labelling of products during collection and processing was not compliant with the standards in a variety of ways and this accounted for 15% of all cited deficiencies (83 of 570), e.g. - time of end of collection/processing missing - volume of anticoagulant missing - unique alphanumeric identifier not used at collection. Many centres have a common problem in designing JACIE-compliant labels. This has been addressed by an international collaboration – the Cellular Therapy Coding and Labeling Advisory Group (CTCLAG) – which has designed HPC and TC component labels
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based on the ISBT 128 system for stem cell component identification (7). Their proposals are currently being discussed by a workshop of European Union member states so that a single coding system for cellular therapy products as required by the ED Directive on Tissues and cells (2004/23/EC) can be agreed. A date of March 2008 has been agreed as the date by which to complete its final documents and recommendations to the EU. Further information on the work of the CTCLAG and EU workshop is available on the ICCBBA and CEN websites (12, 13). 4.3. Experience of centres implementing JACIE It was anticipated that implementation of the JACIE standards would pose some difficulties for applicant centres, particularly in relation to establishing a Quality Management system (QMS). It was also anticipated that there would be resource implications in terms of staff time because of the amount of detailed documentation that is required to demonstrate compliance with the standards. An initial survey was designed to assess the difficulties experienced by centres in preparing for accreditation. The results showed that the most difficult part of preparation was implementing the QM system, adverse event reporting system and other documentation. Lack of a culture of QM was cited as an important problem. The extra resources most frequently required were a quality manager (62% centres) and a data manager (35%). Only 19% needed to improve their physical facilities. There is clearly an important need for training of clinical staff (doctors and nurses) in quality management. It is also important for centres to have a designated quality manager who has appropriate experience in quality management systems. A second survey was designed to assess the effect of implementation of JACIE. Centres were asked to complete this survey 6 months after inspection. All responding centres indicated that they had benefited from implementing the JACIE standards. As shown in Table 3, the areas of greatest perceived benefit were in procedure and practices, staff motivation, control of adverse events, and co-ordination between different areas of the programme. Significant benefits were also perceived in patient satisfaction, facilities, patient care and safety and training of new and existing staff. Improvements clearly depend on the level of existing services, so that failure to demonstrate improvement in, for example, facilities or data management may reflect good pre-existing resources. In other areas, e.g. adverse event reporting, the systems for monitoring performance were only set up as part of implementing JACIE, so that it is difficult to monitor improvements without an established baseline for comparison. Indeed implementation of JACIE may have the paradoxical effect of seeming to increase adverse events because these were not previously adequately reported. 106
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CHAPTER 4 • JACIE accreditation
Table 3: Effect of implementation of JACIE Level of improvement
High
Medium
Low
None
N/A
Control of incidents, events and adverse reactions
41%
59%
-
-
-
Data management
23%
64%
9%
-
5%
Internal coordination
50%
32%
9%
-
9%
Patient satisfaction
23%
36%
18%
14%
9%
Staff motivation
55%
45%
-
-
-
Costs
9%
18%
23%
45%
5%
Facilities
-
50%
18%
32%
Patient safety and care
27%
55%
9%
5%
5%
Procedures and practices
50%
45%
5%
-
-
Training of new/existing staff
41%
45%
9%
-
5%
Compliance with health insurers'/social security demands
14%
9%
23%
45%
9%
Government recognition
18%
27%
27%
23%
5%
Level of improvements in programme functioning experienced by centres, as reported in Survey 2. Results are expressed as percentage of responding centres. N/A: this question not answered
Eighty-one percent of the centres reported that implementation of the QM system had highlighted a need for changes in the running of the transplant programme. The most common areas cited were a need for improved co-ordination between the different facilities, i.e. clinical, collection and processing, and a need for more systematic audit and adverse event reporting. Other items were a need to improve patient medical records and to improve donor evaluation procedures. All centres felt that accreditation was worth the effort invested. In addition, with the implementation of the EU Directive on Safety of Tissues and Cells (Directive 2004/23/EC) it is likely that collection and processing facilities will increasingly view compliance with JACIE standards as important in providing evidence that they are complying with the requirements of the directive.
5. Conclusion The JACIE accreditation system is now firmly established in Europe and the experience of centres that have been inspected is that implementation of the JACIE standards has led to significant improvements in different aspects of their transplant
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programmes. JACIE has further assisted with a number of training courses for preparing centres for accreditation and is currently working on a practical guide to quality management in HPC units. JACIE has also developed a close working relationship with other organisations involved in cellular therapy, which will form the basis for a new global approach to harmonisation of standards and accreditation systems worldwide. This collaboration represents an innovative and proactive approach to solving the problems of international exchange of tissues and cells as these relate to the stem cell transplant community.
References 1. Kvalheim G. EBMT and ISHAGE-Europe create a foundation for inspection and accreditation in Europe. Editorial. Cytotherapy 1999; 1: 363-364. 2. FACT: www.factwebsite.org 3. JACIE: www.jacie.org 4. Directive 2004/23/EC of the European Parliament and of the Council of 31 March 2004 on setting standards of quality and safety for the donation, procurement, testing, processing, preservation, storage and distribution of human tissues and cells: http://ec.europa.eu/health/ph_threats/human_substance/legal_tissues_cells_en.htm 5. Commission Directive 2006/17/EC implementing Directive 2004/23/EC of the European Parliament and of the Council as regards certain technical requirements for the donation, procurement and testing of human tissues and cells: http://ec.europa.eu/health/ ph_threats/human_substance/legal_tissues_cells_en.htm 6. Commission Directive 2006/86/EC implementing Directive 2004/23/EC of the European Parliament and of the Council as regards traceability requirements, notification of serious adverse reactions and events, and certain technical requirements for the coding, processing, preservation, storage and distribution of human tissues and cells: http://ec.europa.eu/health/ph_threats/human_substance/legal_tissues_cells_en.htm 7. NETCORD: www.Netcord.org 8. FACT-Netcord: Details of Cord Blood Bank accreditation process: http://www.factwebsite.org/ main.aspx?id=100 9. JACIE: Requirements for inspectors: www.jacie.org/portal/jacie/inspectors 10.Warkentin PI, Nick L, Shpall EJ. FAHCT accreditation: Common deficiencies during on-site inspections. Cytotherapy 2000; 2: 213-220. 11.Samson D, Slaper-Cortenbach I, Pamphilon D, et al. Current status of JACIE accreditation in Europe: a special report from the Joint Accreditation Committee of the ISCT and the EBMT (JACIE). Bone Marrow Transplantation 2007; 39: 133-140. 12.International Cell Therapy Coding and Label Advisory Group: www.iccbba.org 13.CEN workshop: www.cen.eu/cenorm/businessdomains/technicalcommitteesworkshops/workshops/guideline s_cen_workshops.asp 14.AHCTA: www.ahcta.org
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which of the following statements about JACIE accreditation is not correct? a) JACIE accreditation is voluntary. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) JACIE accredits Collection and Processing facilities . . . . . . . . . . . . . . . . . . . . . . . . c) JACIE accredits Cord Blood Banks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) An inspection visit usually lasts 1.5 days . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which of the following statements about required clinical activity is correct? a) Programmes performing allogeneic and autologous transplants must perform 10 transplants of each type every year . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Programmes performing only autologous transplants must transplant 10 patients every year. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Programmes performing allogeneic and autologous transplants must perform 10 allogeneic transplants but there is no minimum requirement for autograft numbers. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Programmes performing transplants on 2 sites must transplant 10 patients at each site every year . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which of the following statements about donor evaluation is correct? a) There must be written criteria for donor eligibility . . . . . . . . . . . . . . . . . . . . . . . . . b) HTLV 1 and 2 testing is mandatory for all donors . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Pregnancy assessment is not required for autologous donors . . . . . . . . . . . . . . d) Infectious disease markers must be tested within 10 days prior to collection of cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which of the following statements about collection and processing facilities is correct? a) The minimum number of marrow harvests that must be carried out each year by a marrow collection facility is 5 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A collection facility must have SOPs in place covering transport of components only if products are transported outside of the facility . . . . . .
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c) All products must be processed in an environment providing Grade A quality air in a grade B background . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Every individual donation must be identified with a unique alphanumeric number . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which of the following statements about JACIE inspection is correct? a) All deficiencies found at inspection must be corrected within 6 months . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Accreditation is valid for 3 years . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Reaccreditation does not necessarily require a reinspection visit . . . . . . . . . d) Accreditation is awarded by the inspectors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 5
Sources and procurement of stem cells
J. Larghero, J. Garcia, E. Gluckman
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CHAPTER 5 • Stem cell procurement
1. Introduction The source and procurement of haematopoietic stem cells (HSC) has varied and diversified over time. Nowadays, there are three possibilities: bone marrow (BM) HSC, granulocyte-colony stimulating factors (G-CSF) mobilised peripheral blood HSC and cord blood (CB). Each of them seems to give similar clinical results although the use varies according to age of the donor and the recipient, the indication and the preference of the centres and the donors. Usually BM collection is preferred in children, for adults it can be either BM or PBSC, CB depends on the availability of a suitable unit in banks and the absence of an HLA identical donor.
2. Use of different cell sources for HSCT in Europe An EBMT survey on HSC transplantation (HSCT) activity in 2005 reported that 24,168 first HSCT were performed in Europe by 597 centres (1). Among these transplants, 15,278 were autologous and 8,890 allogeneic. Of the 8,890 allo-HSCT, 4,702 were provided by an HLA identical sibling donor, 514 by an HLA-mismatched family donor, and 57 by a homozygous (syngeneic) twin. An UD was used in 3617 patients (41%). The main indications were leukaemia (6,107 patients), lymphoproliferative diseases (1,520 patients), solid tumours (130 patients) and nonmalignant diseases (1,048 patients). The number of allo-HSCT increased by 20% from 2004 to 2005, whereas numbers of auto-HSCT remained stable (1).
3. Choice of stem cell source Traditionally, HSC were harvested from the iliac crests under general anaesthesia. Thereafter, mobilised PBSC have been increasingly used in both auto- and allo-HSCT. Mobilisation of HSC in sufficient number in peripheral blood can be achieved by the classical administration of growth factor such as G-CSF (allo-HSCT) and/or myelosuppressive chemotherapy (auto-HSCT). In the 1990s, unmanipulated CB cells collected and cryopreserved at birth have been used both in related and unrelated HLA matched and mismatched allogeneic transplants in children, and more recently in adults. In 2005, the stem cell source for auto-HSCT was from peripheral blood in 98% of the 15,278 transplants and from BM in 2%. In the allogeneic setting, BM was used in 2,331 of 8,890 transplants (21%) and PBSC in 74%, confirming the increasing use of this stem cell source. CB was used in 395 allogeneic HSCT compared to 281 in 2004. It has become evident that there are many differences, both quantitative and qualitative, between these cell sources (1). The main differences between cell sources are:
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Bone marrow
Collection under general anaesthesia Limited number of haematopoietic stem cells Median number of nucleated cells: 2 x 108/kg Median number of CD34+ cells: 2.8 x 106/kg Median number of T-cells: 2.2 x 107/kg
G-CSF mobilised PBSC
Collection easy No requirement for general anaesthesia Side effects of G-CSF High number of cells Median number of nucleated cells: 9 x 108/kg Median number of CD34+ cells: 7 x 106/kg Median number of T-cells: 27 x 107/kg
Cord blood
Collection easy and harmless Immediate availability of cryopreserved units and lower risk of transmissible diseases Acceptable partial HLA mismatches Number of cells is the limiting factor Median number of nucleated cells: 0.3 x 108/kg Median number of CD34+ cells: 0.2 x 106/kg Median number of T-cells: 0.4 x 107/kg
4. Donor work-up (2–5) (Tables 1–3) 4.1. Donor risk from the procedure The risks for the healthy donor must be minimised. A careful inquiry before donation must be performed. It is recommended that two separate physicians examine the donor prior to the procedure. Detailed information must be given and signed consent must be obtained from the donor and also from both parents in the case of donors under the age of legal consent. In some European countries, when the donor is <16 years old, the consent must be signed in front of a judge and both the family and the child must be seen by a committee of 3 independent experts. The presence of any risk from general anaesthesia for BM aspiration or from cardiac disease for PBSC collection is an absolute contraindication to the donation. The situation is of course completely different for CB, which represents a harmless and easily accessible source of haematopoietic stem cells without any risk to donors.
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CHAPTER 5 • Stem cell procurement
Table 1: Potential transmission of donor diseases to recipients
Infectious diseases
Transmission of
Donor disease
Viruses
HIV / Hepatitis B and C / HTLV-1 / CMV / EBV / Parvovirus B19 / West Nile virus
Bacteria
Contaminants(a)/ Brucellosis
Parasites
Toxoplasmosis / Malaria / Leishmania / Babesia
Fungi
Candida / Aspergillus
Prions
Creutzfeld-Jakob disease(b)
Congenital disorders
Enzyme deficiencies
Gaucher’s disease
Haemoglobinopathies
Thalassemia / Sickle cell anaemia
Acquired disorders
Autoimmune diseases
Myasthenia gravis / Atopy / Lupus erythematosus / Thyrotoxicosis / Diabetes mellitus type I / Sarcoidosis / Coeliac disease / Autoimmune thrombocytopenia
Haematological malignancies
AML / CML / T-cell lymphoma
Non-haematological malignancies (c)
Small-cell lung cancer from renal transplantation / Glioblastoma multiforme from liver transplantation
(a) Contamination rates of 1 in 3000 units in platelet concentrates. No information available on stem cell grafts. (b) No known cases in HSCT and blood transfusions. (c) Not reported in HSCT
Table 2: Recommendations for donor work-up Test
WMDA
NMDP (a)
JACIE (b)
Proposal
Medical history
Yes
Infectious (including risk for infections), pregnancy, blood donation history
Vaccination, travel, blood transfusion history, questions to identify persons at high risk of trasmitting infectious, inherited, haematological or immunological disease
Family history, travel, vaccination, smoking, blood transfusion and donation, infectious, pregnancy, aller-gy, autoimmune, vaccination and tumour history
Physical examination
Yes
Yes
Yes
Particular attention to cardiovascular, bleeding and malignant diseases
ECG
Yes
Yes
Not specified
Yes
Chest X-ray
Yes
Yes
Not specified
Yes
continue
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Test
WMDA
NMDP (a)
Blood counts
Full blood count Complete blood count and differential w. blood film
Blood group
Yes with antibody screen
Coagulation screen
JACIE (b)
Proposal
Not specified
Complete blood count, manual differential on blood film
Yes
ABO group and Rh type ABO Rh type and appropriate red cell compatibility with the recipient No No
ABO group Rh type and appropriate red cell compatibility with the recipient PT, PTT, fibrinogen
HLA-typing
Yes
Yes
Yes
A, B, C, DRB1, DQB1
Biochemical profile
Urea, electrolytes, creatinine, liver function tests, blood sugar
Urea, electrolytes, creatinine, bilirubin, serum protein electrophoresis, Hb S for PBSC donors only
Not specified in detail
Urea, electrolytes, creatinine, liver function tests (bilirubin, AST, ALT, AP, g-GT), LDH, blood sugar, protein electrophoresis, urine analysis
Infectious disease markers (within 30 days prior to collection)
Syphilis, HBsAg, HIV Ag + Ab, HB core Ag, HCV, HTLV-1, HSV, CMV, VZV, EBV serology
HIV 1 and 2, HBV, HCV, HTLV 1 and 2, Treponema pallidum (syphilis), CMV
HIV 1 and 2, HBV, HCV, HTLV 1 and 2, Treponema pallidum and CMV
HIV 1 and 2, HBV (HBs Ag, HBsAb, and HBcAb), HCV (HCVAb), HTLV type II, Treponema pallidum, CMV (IgG, IgM), Toxoplasmosis (IgG, IgM) and EBV
Yes
Yes
Gynaecological visit + pregnancy test + physical breast exam
Pregnancy test Yes (when indicated)
Other proposals: - Check for malignant diseases in donors >55 yrs (applies to related HSCT) Æ PSA in males, physical examination, occult blood in stool. BM aspiration if medical history or tests are abnormal. CT-chest scans in case of a long smoking history. - Congenital disorders Æ Testing for congenital disorders of planned recipient within family donors; testing for other congenital disorders of the family.
(a) 18th Edition Standards/ September 2002. (b) www.jacie.org
Table 3: Donor selection in case of donors with abnormal findings Family Donors Abnormal finding
Absolute contraindications
Relative contraindications
Specific consideration
Infectious
HIV HTLV-1
HBV, HCV, malaria within past 3 yrs, parvovirus B19
EBV, CMV, toxoplasmosis,
Congenital
Thalassaemia major Thalassaemia minor Combined immune deficiency Gaucher’s disease Congenital disease with severely reduced life expectancy
Malignancies
Any malignancy except in situ cancer
Targeted CB should be tested for congenital disease of planned recipient
Skin cancer removed in toto
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CHAPTER 5 • Stem cell procurement
Family Donors Abnormal finding
Absolute contraindications
Pregnancy
Relative contraindications
Specific consideration
BM donation and G-CSF stimulated or unstimulated apheresis
Unrelated Donors (a) Infectious
As for blood donation
Congenital
As for blood donation
Malignancies
Any malignancy except in situ cancer
Pregnancy
Any donation
Parvovirus B19, if known after collection: Gram-positive or Gramnegative bacterial graft infection
EBV, CMV, toxoplasmosis
Skin cancer removed in toto
Unrelated Cord Blood (b) Infectious
As for blood donation. Gram-positive or Gramnegative contamination
Congenital
As for blood donation Exclude, if congenital diseases known in family
Malignancies
Any, in child
EBV, CMV, toxoplasmosis
(a) Not enough information is available for West Nile virus. Contamination of stem cell graft with epidermal bacteria might be a relative contraindication
4.2. Recipient risk from a particular donor (2) Potentially transmittable diseases from donors include infections, congenital disorders, and acquired diseases such as malignancies or autoimmune diseases.
5. Comparison of BMT and PBSCT (5–12) In 1995, 3 pivotal studies demonstrated the safety and feasibility of using G-CSF mobilised PB allografts (Table 4). Patients experienced prompt engraftment with an incidence of GvHD similar to that of BM recipients (Table 5). In addition, no serious short-term complications of G-CSF mobilised PB harvesting were observed in the donors. Direct comparison of PB and BM in allogeneic sibling donor transplantation has been reported in at least 8 randomised trials. Most of them did not show a benefit in overall
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Table 4: Collection protocols for PBSC allografts in 4 randomised trials Study
G-CSF dose µg/kg
Days G-CSF
No of aphereses
106/kg 106/kg 106/kg Ttarget actual cell dose CD34+ dose CD34+ dose
Schmitz Blood 108: 4288, 2006
10
4
1 (1-3)
4
5.8 (1.5-68)
300 (16-2123)
Couban BBMT 10: 624, 2004
5
4
2 (1-2)
2.5
6.4 (0.7-32)
370 (120-3080)
Bensinger 16 NEJM 344: 175, 2001
5
1 (1-4)
5
7.3 (1-30)
279 (143-788)
Blaise JCO 18: 537, 2000
5
2 (1-3)
4
6.6 (1.5-19)
356 (131-754)
10
Table 5: Neutrophil and platelet recovery and incidence of aGvHD and cGvHD after allo-PBSCT compared to allo-BMT in different randomised studies Reference
Source
n
Neutrophils (median days)
Platelets (median days)
aGvHD (%)
Extensive cGvHD (%)
Bensinger
BM PBSC
91 81
21 16
19 13
57 64
35 46 p=ns
Blaise
BM PBSC
52 48
21 15
21 13
42 44
7.5 31 p=0.03
Schmitz
BM PBSC
166 163
15 12
20 15
37 50 p=0.01
11.5 25 p=0.0009
Table 6: Relapse incidence (RI) and survival after allo-PBSCT compared to alloBMT in different randomised studies Reference
Source
n
RI
Survival
Bensinger
BM PBSC
91 81
25% 14% p=0.04
54% 66% p=0.06
Blaise
BM PBSC
52 48
ns
65% 67% p=ns
BM PBSC
166 163
ns
ns
Schmitz
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- Early status: 72 vs. 75% p=ns - Advanced: 33 vs. 57% p=0.04
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survival (OS). The incidence of aGvHD was the same in all but one of the studies, but an increase (statistically significant or a trend) in the incidence of overall and extensive cGvHD was demonstrated in recipients of PB allografts (Table 6). The magnitude of this observation and its effect on relapse, OS and recipient quality of life is less clear. In unrelated transplant recipients, matched cohort comparisons of unrelated (UD) BMT and PBSCT reported faster haematological recovery among PB recipients with no difference in either acute or chronic GvHD. While results of randomised studies are pending, the use of PB allografts in UD HSCT has varied among transplant centres and countries. Some unrelated Donor Registries have permitted the collection of allografts from PB whereas others have not. Transplant centres may request a PB or a BM graft but the collection centre and wishes of the donor also determine which product is ultimately collected. Because of the absence of definitive data comparing both sources of cells, there is no indication to prefer either source of cells except perhaps in patients with advanced disease where chronic GvH and subsequent GvL might decrease relapse and improve OS or, in a situation where a high number of cells is necessary for engraftment, for example after non-myeloablative conditioning or if TCD is planned for a HLA mismatched transplant.
6. Cord blood collection and banking The first allogeneic CB transplantation (CBT) was successfully performed in 1988 in a child with Fanconi’s anaemia; the CB donor was his HLA identical sister. Fifteen years later, this patient is doing well with full donor haematopoietic and lymphoid reconstitution. This first success showed that a single CB unit contained enough HSC to reconstitute definitely the host lympho-haematopoietic compartment, that a CB unit could be collected at birth without any harm to the new-born infant and finally that CB HSC could be cryopreserved and transplanted in a myeloablated host after thawing without losing their repopulating abilities. CB has many theoretical advantages due to the immaturity of newborn cells. HSC from CB are enriched in primitive haematopoietic stem cells, which are able to produce in vivo long-term repopulating SC. The properties of CB cells should compensate for the relatively low number of cells contained in a single CB unit and, through rapid expansion reconstitute myeloablated patients. In spite of the capacity of CB cell expansion, clinical results showed that haematopoietic recovery is delayed after CBT engraftment but improves if a higher number of nucleated and CD34+ cells are infused. In the long term, a higher frequency of reconstitution of early and committed haematopoietic progenitors was observed in children receiving a CBT compared to BMT suggesting that the delay in engraftment may reflect the difficulty
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of CB progenitors to reprogram themselves toward differentiation. The second advantage of CBT relates to the immaturity of the immune system at birth. This property should decrease the alloreactive potential of lymphocytes and as a consequence reduce the incidence and severity of GvHD after an HLA matched or mismatched transplant. These properties should lead to less stringent criteria for HLA donor recipient selection (Table 7). Recent studies have shown that the cell dose is the most important factor for donor selection: a minimum cell dose of 3 x 107 nucleated cells/kg is required for successful engraftment. More recently the use of two CB units seems to improve the engraftment rate.
Table 7: Advantages and disadvantages in the search and identification process of BM and CB unrelated donor
Information on A+B+DRB1(DNA) type Median search time Donors identified but not available Rare haplotypes represented (a) Major limiting factors to graft acquisition Ease of arranging date of cell infusion Potential for second HSC graft or DLI Potential - for viral transmission - for congenital diseases Risk to donor
Bone marrow
Cord blood
16–56% 3–6 months 30% 2% HLA match Difficult Yes Yes No Yes
50–80% <1month < 1% 29% Cell dose Easy No from the same donor No Yes No
(a) Data of the ethnic distribution of CB (n=4577) and BM donors registered (n=68487) in the British Bone marrow Registry (Cristina Navarrete, personal communication)
7. Cord blood banking for unrelated CBT In order to organise the development of CB banks, Netcord, an International Foundation promoting high quality cord blood banking, currently includes more than twenty experienced large CB banks in the USA, Europe, Japan and Australia. Netcord has developed a detailed set of standards for CB banking, the third edition of which has been recently released. These include respective national and international regulatory aspects and, with the collaboration of the Foundation for Accreditation of Hemopoietic Cell Therapy (FAHCT) (USA), an international accreditation program was started in 2002, with more than 15 accredited CBBs up to date. Furthermore, a joint allocation system employing most recent Internet technology has been implemented to facilitate the rapid allocation of CB units 120
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according to histocompatibility and number of nucleated cells within an average time of 48 hours. The number of CB transplants from UD has increased dramatically, and more than 5,800 patients have undergone CBT from UD thus far (www.netcord.org). 7.1. Practical aspects of cord blood banking 7.1.1 Informed consent CB is a discarded product that, up to some years ago, could be used without permission; however, today it is considered a transplantable product (cells, tissue or pharmaceutical drug according to the regulations in different countries) and informed consent must be obtained for its donation and for carrying out tests on the mother and the CB. Careful questioning about medical history allows the opportunity of excluding donors who have a high risk of transmitting any infectious or genetic diseases. The families must be informed about the genetic and infectious tests performed. 7.1.2. Collection techniques Two techniques are used to collect CB: - One is collection in the delivery room while the placenta is still in utero, and - The other is collection of the CB in an adjacent room after the delivery. In the first instance, the collection can be done in the delivery room by an obstetrician or a midwife. The advantage of this procedure is that the volume of cells collected is usually higher if the cord is clamped early and the collection is begun immediately; however, this can disrupt the normal process of delivery and is not always feasible. The collection of CB after delivery is easier and it can be performed by designated personnel; however, fewer cells may be collected and there may be an increase in the risk of bacterial contamination or clotting. 7.1.3. Infectious disease testing Tests for syphilis and for viral infections (including those for HIV, HBV and HCV, and CMV) are performed on the mother’s blood. In some countries, the mother is also tested for HTLV-I/II and toxoplasmosis or other transmissible diseases according to local prevalence. Most often, virology tests are not performed immediately; rather, a separate frozen aliquot is kept so that these tests can be performed before a transplant procedure. CB should be quarantined until a confirmatory test is performed on the mother 3–6 months after delivery. This should decrease the risk of virus transmission considerably. These risk estimates may be higher in CB donors because, unlike regular whole blood
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donors, CB donors are recruited only once. Bacterial infection is also a major issue, but it seems that the incidence of bacterial contamination diminishes with the expertise of the staff in charge of the collection. In all cases, a bacterial culture for anaerobic and aerobic bacteria must be performed. 7.1.4. Genetic disease testing The decision to perform tests for genetic diseases should be directed by the family medical history, its ethnic background and the follow-up of the donor. Tests on cord blood are expensive and there is no real consensus on the type and number of tests that should be performed. There are also some concerns about notifying the family of the test results. 7.1.5. HLA typing HLA typing is performed on an aliquot of CB. In collaboration with the Eurobank project, Eurocord/Netcord collects DNA from the donor and recipient to perform molecular typing for HLA-C, -DQB1, -DPB1, some HLA-A and -B antigens, and other markers. Some banks type routinely the mother for HLA in order to have information on the haplotype and to control the accuracy of CB typing. 7.1.6. Cell processing Because of the small volume of CB collected, which usually ranges from 40–150 mL, there is some concern that any attempt at cell manipulation and concentration may result in a considerable cell loss that could impair engraftment. Many banks, specially the larger ones, use advanced GMP procedures for CB volume reduction, freezing and storage, which have minimised cell loss. The thawing technique is well established and aims at removing red residual cells and cryoprotectants. Evaluation of HSC content is very important. Several studies have shown that the number of nucleated cells infused correlates with engraftment. A correlation has also been demonstrated between placental weight, time of clamping, speed of processing, volume collected, and progenitor cell content. Quantification of the cellular content of cord blood is not always easy. Most studies refer to nucleated or mononuclear cells infused per kilogram of body weight before and after thawing of cells. Enumeration of CD34+ cells by flow cytometric analysis is performed routinely by most laboratories, but these results are not always comparable. Others count the number of CFU-GM in clonogenic assays. There is a large variation of methodology among laboratories, which explains why quantification has been a problem. In the European study, a correlation was found between the number of nucleated cells infused and engraftment, but this correlation was not found when the number of CD34+ cells and CFU-GM were evaluated.
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7.1.7. Ethical and legal aspects CB can be collected for infusion into an unrelated or a related recipient and, more hypothetically, for autologous use. For unrelated CBT, the mother must be aware that the donation is anonymous and free, and that there will be no guarantee that it will be possible to retrieve the cord blood if it is needed later for family or autologous use.
8. Comparison of UD CBT and UD BMT 8.1. Children with acute leukaemia In 2001, a comparison between UD CBT and UD BMT was carried out by Eurocord and EBMT in 515 children with AL, transplanted either with an UD CBT (n=99) or an UD BMT (n=416) (13). Recipients of CB were younger and had more pre-transplant adverse disease factors. Most of the BM donors were HLA matched or had a maximum of 1 HLA difference while most of the CB donors had 2 to 3 HLA differences. The median number of nucleated cells infused was 4 x 108/kg in UD BMT and 0.38 x 108/kg in UD CBT. In unrelated BMT, 262 children received an unmanipulated BMT and 180 a TCD BMT using Campath-1M as the most frequent mean of TCD. The median followup was 29 months. During the first 100 days after HSCT, while unmanipulated and TCD BMT did not differ in haematopoietic recovery and TRM, the main findings that emerged from these adjusted comparisons were the poor results of CBT in relation to neutrophil and platelet recovery and early TRM. In contrast CBT and TCD BMT gave less aGvHD than the unmanipulated BMT group. Finally the unmanipulated BMT and CBT groups had less leukaemic relapse than the T-cell depleted group. In the long term, the unmanipulated BMT group had more cGvHD than the TCD BMT and the CBT groups. Furthermore, while the outcome of the 3 groups were comparable in terms of long term relapse, mortality after day 100 was increased in the TCD BMT group, mainly because of the occurrence during the first 100 days after BMT of early relapse, GvHD and lack of engraftment. In summary, the main differences in adjusted outcomes between the three unrelated transplant groups appeared in the first 100 days post transplant but without giving an advantage to any group. Indeed, the delay of engraftment and increased TRM observed after UD CBT must be balanced against the higher risk of aGvHD after unmanipulated UD BMT and to the higher risk of relapse after TCD. In contrast, after day 100 posttransplant, the 3 groups achieved similar results in terms of relapse but cGvHD occurred more frequently with unmanipulated UD BMT and death with TCD UD BMT. Recently, these results have been reinforced by the report of CIBMTR and New York cord blood bank, comparing the outcomes of 503 children with acute leukaemia transplanted with UD mismatched CB with 282 UD BM recipients (of which 116 were
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8/8 antigen HLA matched) (14). While they showed that TRM was increased in children receiving a low CB cell dose (less than 3 x 107/kg) and one HLA-mismatched CB, or two HLA-mismatched CBT independently of cell dose, this study also demonstrated that leukaemia-free survival (LFS) was not statistically different from one or two HLA-mismatched CBT compared with a HLA-allele-matched UD BMT. These results justify the simultaneous search of UD CB donors and UD BM in children with AL. The decision to perform CBT will be based on the cell content of the graft the number of HLA disparities and the urgency of the transplant. 8.2. Adults with acute leukaemia (14, 15) In 2004, two groups reported the transplant outcomes of adults with leukaemia transplanted with either UD CB or UD BM (15, 16). In both studies, CB recipients were younger and had more advanced disease at the time of transplantation. In the study comparing CB with HLA-matched UD BM, multivariate analysis showed lower acute GvHD ≥ grade II after CB transplantation but delayed neutrophil recovery. The 2-yrs cumulative incidence of cGvHD, TRM, leukaemia relapse, and the 2-yrs probability of OS and LFS were not significantly different. The second report compared transplant outcomes between patients receiving UD CB (one or two mismatch) with patients grafted with UD BM (HLA-matched or one mismatch). UD CB and mismatched BM transplantation were associated with a higher risk of cGvHD and aGvHD, respectively. Interestingly, the incidence of TRM, treatment failure, and overall mortality were similar in patients who received mismatched BM or mismatched CB transplants. These data suggest that, despite increased HLA disparity, CB from UD offers comparable results to matched UD BM in adults with AL leading to the conclusion that the donor search process for BM and CB from UD should be started simultaneously in adults especially in patients with AL where the time factor is very important.
9. Algorithm for donor search High resolution HLA typing of patient and family No donor: Alternative donor search
124
BM donor registries
Cord blood banks
HLA identical A, B, C, DR, DQ 10 /10 or 9/10
Cell dose >2 x 108/kg 1 or 2 HLA mismatches A, B, DR
Transplant
Transplant
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CHAPTER 5 • Stem cell procurement
10. HSC and regenerative medicine The need for an alternative source of pluripotent stem cells for tissue engineering that does not require the use of human embryos or therapeutic cloning has resulted in increased interest in the use of adult stem cells, although they are likely more limited in potential. However, stem cells (haematopoietic, mesenchymal, endothelial, etc.) isolated from either BM or CB represent an ethically uncontroversial source of cells for clinical applications. The experimental and first clinical data reported with such stem cells herald new applications in cellular therapy and tissue regeneration, ranging from neurodegenerative, muscular, brain or bone diseases to myocardial infarction, liver cirrhosis, or vascular disorders.
References 1. Gratwohl A, Baldomero H, Frauendorfer K, et al. Results of the EBMT activity survey 2005 on haematopoietic stem cell transplantation: Focus on increasing use of unrelated donors. Bone Marrow Transplant 2007; 39: 71-87. 2. Niederwieser D, Gentilini C, Hegenbart U, et al. Transmission of donor illness by stem cell transplantation: Should screening be different in older donors? Bone Marrow Transplant 2004; 34: 657-665. 3. Cleaver SA, Warren P, Kern M, et al. Donor work-up and transport of bone marrowrecommendations and requirements for a standardized practice throughout the world from the Donor Registries and Quality Assurance Working Groups of the World Marrow Donor Association (WMDA). Bone Marrow Transplant 1997; 20: 621-629. 4. Favre G, Beksac M, Bacigalupo A, et al. for the European Group for Blood and Marrow Transplantation (EBMT). Differences between graft product and donor side effects following bone marrow or stem cell donation. Bone Marrow Transplant 2003; 32: 873-880. 5. Couban S, Barnett M. The source of cells for allografting. Biol Blood Marrow Transplant 2003; 9: 669-673. 6. Champlin RE, Schmitz N, Horowitz MM, et al. Blood stem cells compared with bone marrow as a source of hematopoietic cells for allogeneic transplantation. Blood 2000; 95: 37023709. 7. Bensinger WI, Martin PJ, Storer B, et al. Transplantation of bone marrow as compared with peripheral blood from HLA identical relatives in patients with hematologic cancers. N Engl J Med 2001; 344: 175-181. 8. Blaise D, Kuentz M, Fournier C, et al. Randomized trial of bone marrow versus lenogastrimprimed blood cell allogeneic transplantation in patients with early stage leukemia: A report from the Société Française de Greffe de Moelle. J Clin Oncol 2000; 18: 537-546. 9. Gorin NC, Labopin M, Rocha V, et al. for the Acute Leukemia Working Party (ALWP) of the European Cooperative group for Blood and Marrow Transplantation (EBMT). Marrow versus peripheral blood for geno-identical allogeneic stem cell transplantation in acute myelocytic leukemia: Influence of dose and stem cell source shows better outcome with rich marrow. Blood 2003; 102: 3043-3051.
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10.Ringden O, Remberger M, Runde V. Peripheral blood stem cell transplantation from unrelated donors: A comparison with marrow transplantation. Blood 1999; 94: 455-464. 11.Remberger M, Ringden O, Blau IW, et al. No difference in graft versus host disease, relapse and survival comparing peripheral blood stem cells to bone marrow using unrelated donors. Blood 2001; 98: 1739-1745. 12.Ringden O, Labopin M, Bacigalupo A, et al. Transplantation of peripheral blood stem cells as compared with bone marrow from HLA identical siblings in adult patients with acute myeloid leukemia and acute lymphoblastic leukemia. J Clin Oncol 2002; 20: 4655-4664. 13.Rocha V, Cornish J, Sievers EL, et al. Comparison of outcome of unrelated bone marrow and umbilical cord blood transplants in children with acute leukemia. Blood 2001; 97: 2962-2971. 14.Eapen M, Rubinstein P, Zhang MJ, et al. Outcomes of transplantation of unrelated donor umbilical cord blood and bone marrow in children with acute leukaemia: A comparison study. Lancet 2007; 369: 1947-1954. 15.Laughlin MJ, Eapen M, Rubinstein P, et al. Outcomes after transplantation of cord blood or bone marrow from unrelated donors in adults with leukaemia. N Engl J Med 2004; 351: 2265-2275. 16.Rocha V, Labopin M, Sanz G, et al. Transplants of umbilical-cord blood or bone marrow from unrelated donors in adults with acute leukaemia. N Engl J Med 2004; 351: 22762285.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Compared to allogeneic bone marrow cells, peripheral blood HSC give: a) Less GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Accelerated neutrophil recovery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Higher relapse rate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Better overall survival . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. The best criterion for cord blood search is: a) Number of nucleated cell dose >3 x 107/kg . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) ABO compatibility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) HLA identity determined by allele typing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Donor sex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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3. Quality of unrelated matched HSCT collection relies on which of the following criteria? a) Total volume . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Number of CD34 cells/kg . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Number of viable cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Number of CFU-GM . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which of the following viral infections in the unrelated donor does contraindicate the use of the donor? a) HIV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) CMV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Hepatitis B . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Hepatitis C . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. The main complication of collection of PBSC is: a) Hypotension . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Bleeding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Flu like symptoms due to G-CSF . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 6
Principles of conditioning
A. Gratwohl
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CHAPTER 6 • Principles of conditioning
1. Introduction Conditioning plays a central part in HSCT. It is the source of the inherent dilemma in HSCT: how to get rid of the disease but without toxicity. Conditioning is a key cause of early mortality but holds out the prospect of long-term disease control or even cure. Errors in its application can have serious, even immediately fatal consequences. Quality management issues (see below) therefore have a central role in all aspects of conditioning. It is of importance to be familiar with all aspects of conditioning. The term “conditioning” in HSCT means to “condition”, e.g. to prepare the patient for its transplant. Conditioning is given with three main objectives: "creation of space", immunosuppression and disease eradication. 1.1. Creation of space This is a somewhat controversial concept, which originated from the belief that immature progenitor cells occupy defined niches within the marrow stroma in order to obtain the necessary support for proliferation and differentiation. According to this theory, existing host stem cell cells must be eradicated in order for donor stem cells to obtain access to these niches and for engraftment to occur. Experimental and clinical data are controversial about the need to create space. There are indications that engraftment after RIC HSCT is more rapid in patients with an “empty” bone marrow compared to a “full” bone marrow. 1.2. Immunosuppression to prevent a host-versus-graft reaction Immunosuppression is required to prevent rejection of the incoming donor cells by host immune cells. This is clearly not required in autologous or syngeneic HSCT. The need for immunosuppression increases with the increase of disparity in major histocompatibility antigens (HLA). The risk of rejection is increased in situations where the recipient has been "pre-sensitised" against minor histocompatibility antigens, e.g. by the administration of multiple blood products prior to HSCT. Rejection is also increased in TCD HSCT. Vice versa, graft acceptance is increased by high stem cell dose and high T-cell dose. 1.3. Disease eradication The key goal of the conditioning regimen is long-term disease control. This is the main objective for patients with malignancies, but it is also of vital importance in diseases characterised by hyperplastic marrows, e.g. thalassaemia. Partial engraftment may be sufficient in situations where only a "specific product" is required, e.g. Bcells in some immunodeficiency states or when an enzyme produced by haematopoietic cells is required, e.g. in mitochondrial neurogastrointestinal encephalopathy.
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2. Historical perspective In order to understand the diversity and multitude of today's conditioning regimens, they have to be seen in their historical context. Bone marrow transplantation was initially investigated as a tool to protect survivors of accidental high dose irradiation due to nuclear accident or atomic bomb exposure. At the same time, it was realised that TBI could be used as a tool to eliminate a leukaemic haematopoietic system without causing irreversible damage to other organs, similar to surgical removal of a diseased organ in solid organ transplantation. “Condition” meant the preparation of the recipient to accept a new organ in place of the diseased and eradicated haematopoietic system. TBI proved to be sufficient for engraftment but insufficient for long-term disease control when used on its own. The addition of cyclophosphamide (Cy) to TBI and transplantation at an early stage of the disease were the main elements for successful BMT in the late seventies. At that time attempts were already being made to replace TBI with “radiomimetic” drugs or high dose “leukaemia specific” chemotherapy, using drugs including Cy, busulfan (Bu), etoposide (VP16), cytosine arabinoside (ARA-C), carmustine (BCNU) and melphalan (MEL). The combinations Cy/TBI, Bu/Cy alone or BACT (BCNU, ARA-C, VP16 and 6TG) were the main conditioning regimens for HSCT at this time. In the 1980’s, emphasis was on dose intensification and exploring alternatives to Cy in combination with TBI. New conditioning regimens were designed with the aim of reducing the risk of relapse and rejection. A number of studies defined maximum tolerable doses for single drugs in combination with TBI. Dose escalation studies suggested that up to 60 mg/kg of VP16, 110-180 mg/m2 of MEL and 36 g/m2 of ARA-C could be combined with TBI. Dose limiting toxicities were interstitial pneumonitis (IP) for TBI, stomatitis and veno-occlusive disease (VOD) for MEL/TBI and VP16/TBI, and CNS and skin toxicity for ARA-C/TBI. Several centres evaluated the use of TBI plus more than one drug, but no consensus was reached regarding the maximal tolerated doses. None of these regimens made any significant improvements to clinical outcome. Any reduction in relapse or rejection risk was usually accompanied by an increase in TRM. OS and DFS remained unchanged or worsened. Similarly, no study with “leukaemia specific agents” such as ARA-C or VP16, showed superiority to Cy, a drug that is not used in the conventional treatment of leukaemia. So far, no single study has shown any conditioning regimen to provide better long-term survival than Cy/TBI or Bu/Cy. In the 1990’s the emphasis switched to concepts focused on reducing TRM and morbidity with improvements in QoL. Better understanding of GvL effects, the recognition of the capacity of DLI to shift the balance between donor and recipient in a predictable way and preclinical experiments which defined the minimal requirements for stable engraftment, led to the introduction of the concept: 130
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“reduced intensity conditioning”. Most of the conditioning regimens (standard, intensified or reduced) were developed as phase I/II trials and then adopted by individual institutions without formal phase III studies. There are a few exceptions. Standard conditioning with Cy/TBI was compared with Bu/Cy in several studies in haematological malignancies. There were substantial differences in toxicities (more VOD, more permanent alopecia with Bu/Cy) but little differences in long-term survival. In ALL there is probably an advantage at long term with TBI containing regimens. Current strategies focus on a new concept. Until recently, most patients were given the same conditioning regimen. Today, it is believed that patients with high risks for TRM and low disease risk should receive a different conditioning regimen from patients with low risk for TRM and high-risk disease. The approach should integrate conditioning, graft product and GvHD prophylaxis and treatment from the very beginning (see Tables 1 and 2).
3. RIC HSCT and the concept of “immunoablation versus myeloablation” The concept of RIC HSCT is based on the idea of circumventing the high morbidity and mortality associated with standard conditioning in patients with advanced age or comorbidities. It evolved from preclinical studies which defined the minimal requirements for engraftment. They also made use of the potential of DLI to shift mixed chimerism to full chimerism. The goal of RIC HSCT is not tumour eradication or destruction of host haematopoiesis by cytotoxic therapy but via immune mediated effects. The graft-versus-tumour potential of the approach is based on several components: initial conditioning, graft composition, prevention of post-transplant rejection and use of DLI in case of incomplete chimerism at specified time points. Many of these regimens include double immunosuppression with both CsA and mycophenolate mofetil (MMF) post-transplant. Initially, emphasis was given to the idea that “immunoablative” drugs, e.g. fludarabine or anti-T-cell antibodies, were more important than “myeloablative” drugs, such as MEL or Bu. This idea has been largely abandoned. Most drugs as well as TBI have profound effects on lymphoid as well as myeloid elements and the total doses of drugs and TBI appear to have more influence on toxic side effects than on immunosuppression. Isolated antilymphoid antibodies might be the only exception. RIC HSCT are an established component of transplant strategies today with emphasis on patients with advanced age. Despite many studies, none has shown unequivocal benefit in long term survival compared to standard conditioning. Early organ toxicity can be reduced but impact on GvHD and infectious complications is much less and RIC HSCT is always associated with an increased risk of relapse during follow-up (see Figure 1).
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Table 1: Conditioning regimens Total Dose Regimen • Conventional “old” regimens Cy/TBI 120 mg/kg Cyclophosphamide 12–14.4 Gy Total Body Irradiation Bu/Cy 16 mg/kg Busulfan 200 mg/kg Cyclophosphamide BACT 200 mg/m2 BCNU ARA-C 800 mg/m2 Cyclophosphamide 200 mg/kg 6-Thioguanine 800 mg/m2 • Alternative “standard” regimens TBI/VP Total Body Irradiation 12–13.2 Gy Etoposide 60 mg/kg AC/TBI ARA-C 36 g/m2 Total Body Irradiation 12 Gy MEL/TBI Melphalan 110–140 mg/m2 Total Body Irradiation 10–14.85 Gy Bu/Cy Busulfan 16 mg/kg Cyclophosphamide 120 mg/kg Bu/MEL Busulfan 16 mg/kg Melphalan 140 mg/m2 • Intensified regimens Cy/VP/TBI Cyclophosphamide 120 mg/kg Etoposide 30–60 mg/kg Total Body Irradiation 12–13.75 Gy TBI/TT/Cy/ATG Total Body Irradiation 13.75 Gy Thiotepa 10 mg/kg Cyclophosphamide 120 mg/kg ATG** 120 mg/kg Bu/Cy/MEL Busulfan 16 mg/kg Cyclophosphamide 120 mg/kg Melphalan 140 mg/m2 • Reduced intensity regimens TBI/Fluda Total Body Irradiation 2 Gy Fludarabine 90 mg/m2 Fluda/Bu/ATG Fludarabine 180 mg/m2 Busulfan 8 mg/kg ± ATG** 40 mg/kg
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Daily Dose
Administration
Days
60 mg/kg 2–2.4 Gy (2x/day)
IV in 1 hour
-6, -5 -3, -2, -1
4 mg/kg* 50 mg/kg**
p.o. q 6 hour IV in 1 hour
-9, -8, -7, -6 -5, -4, -3, -2
200 mg/m2 200 mg/m2 50 mg/kg 200 mg/m2
IV in 2 hours IV in 2 hours IV in 1 hour p.o
-6 -5, -4, -3, -2 -5, -4, -3, -2 -5, -4, -3, -2
2–2.5 Gy (2x/day) 60 mg/kg
IV in 2 hours
-7, -6, -5, -4 -3
3 g/m2 2 Gy (2x/day)
IV q 12 hours in 2 h -9, -8, -7, -6, -5, -4 -3, -2, -1
110–140 mg/m2 2 Gy (2x/day)
IV in 1 hour
-3 -2, -1, 0
4 mg/kg* 60 mg/kg
p.o. q 6 hours IV in 1 hour
-7, -6, -5, -4 -3, -2
4 mg/kg* 140 mg/m2
p.o. q 6 hours IV in 1 hour
-5, -4, -3, -2 -1
IV in 1hour 60 mg/kg IV in 2 hours 30–60 mg/kg 2–2.25 Gy (2x/day)
-6, -5 -4 -3, -2, -1
1.25 Gy (3x/day) 5 mg/kg 60 mg/kg 30 mg/kg
p.o. q 6 hours IV in 1 hour IV in 5–6 hours
-9, -8, -7, -6 -5, -4 -3, -2 -5, -4, -3, -2
4 mg/kg* 60 mg/kg 140 mg/m2
orally every q 6 hours -7, -6, -5, -4 -3, -2 IV in 1 hour -1 IV in 1 hour
2 Gy 30 mg/m2
IV in 30 min
0 -4, -3, -2
30 mg/m2 4 mg/kg* 10 mg/kg
IV in 30 min p.o. q 6 hours IV in 8–10 hours
-10 to -5 -6, -5 -4, -3, -2, -1
* Aternatively 3.2 mg/kg as IV formulation (Busulfex®); **ATG: specific dose depends on brand of ATG
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Table 2: Conditioning regimen by diseases Total Dose Regimen • Severe Aplastic Anaemia Cy 200 mg/kg Cyclophosphamide Cy/ATG 200 mg/kg Cyclophosphamide 90 mg/kg ATG** • Lymphoma BEAM 300 mg/m2 BCNU Etoposide 400–800 mg/m2 ARA-C 800–1600 mg/m2 Melphalan 140 mg/m2 CBV BCNU 300–600 mg/m2 Etoposide 750–2400 mg/m2 Cyclophosphamide 4.8–7.2 g/m2 • Solid Tumours LACE CCNU 200 mg/m2 Etoposide 1 g/m2 ARA-C 4 g/m2 Cyclophosphamide 5.4 g/m2 CCB Cyclophosphamide 5625 mg/m2 Cisplatin 165 mg/m2 BCNU 600 mg/m2 CTCb (Stamp) Thiotepa 500 mg/m2 Cyclophosphamide 6 g/m2 Carboplatin 800 mg/m2 ICE (high dose) Ifosfamide 16 g/m2 Carboplatin 1.8 g/m2 Etoposide 1.5 g/m2 Cy/Fluda/ATG Cyclophosphamide 120 mg/kg Fludarabine 125 mg/m2 ATG** (ATGAMR) 200 mg/kg • Fanconi Fluda/Bu/Cy/ATG Fludarabine 100 mg/m2 Busulfan 6 mg/kg Cyclophosphamide 40 mg/kg ATG** 6 mg/kg • Thalassaemia Bu/Cy Busulfan 14–16 mg/kg Cyclophosphamide 200 mg/kg
Daily Dose
Administration
Days
50 mg/kg
IV in 1 hour
-5, -4, -3, -2
50 mg/kg 30 mg/kg
IV in 1 hour IV in 8–10 hours
-5, -4, -3, -2 -5, -4, -3
300 mg/m2 150–200 mg/m2 200–400 mg/m2 140 mg/m2
IV in 2 hours IV in 2 hours IV in 2 hours IV in 1 hour
-6 -5, -4, -3, -2 -5, -4, -3, -2 -1
100-200 mg/m2 250-800 mg/m2 1.2–1.8 g/m2
IV in 2 hours IV in 2 hours IV in 1 hour
-8, -7, -6 -8, -7, -6 -5, -4, -3, -2
200 mg/m2 1 g/m2 2 g/m2 1.8 g/m2
p.o. IV in 2 hours IV in 2 hours IV in 1 hour
-7 -7 -6, -5 -4, -3, -2
1406.3 mg/m2 41.2 mg/m2 600 mg/m2
IV in 1 hour IV in continuous inf. IV in 2 hours
-6, -5, -4, -3 -6, -5, -4, -3 -3
125 mg/m2 1.5 g/m2 200 mg/m2
IV in continuous inf. IV in continuous inf. IV in continuous inf.
-7, -6, -5, -4 -7, -6, -5, -4 -7, -6, -5, -4
4 g/m2 600 mg/m2 500 mg/m2
-6, -5, -4, -3 IV in 2 hours IV in continuous inf. -6, -5, -4 IV in 2 hours (2x day) -6, -5, -4
60 mg/kg 25 mg/m2 40 mg/kg
IV in 1 hour IV 30 min IV in 8–10 hours
-7, -6 -5, -4, -3, -2 -5, -4, -3, -2
25 mg/m2 1.5 mg/kg* 10 mg/kg 1.5 mg/kg
IV 30 min p.o. q 6 hours IV in 1 hour IV in 8–10 hours
-5, -4, -3, -2 -9, -8, -7, -6 -5, -4, -3, -2 -4, -3, -2, -1
3.5–4 mg/kg 50 mg/kg
p.o. q 6 hours IV in 1 hour
-9, -8, -7, -6 -5, -4, -3, -2
* Aternatively 0.8 mg/kg as IV formulation (Busulfex®); ** ATG: specific dose depends on brand of ATG HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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Figure 1: Survival and cause of death after HSCT for early stage leukaemia with standard conditioning 1a: Overall survival and cumulative incidence of main cause of death Other causes 0.10 Infections 0.20 GvHD 0.30 Cumulative incidence
Relapse 0.40 0.50 0.60 0.70 0.80 0.90
12.00 24.00 36.00 48.00 60.00 72.00 84.00 96.00 108.00 Months since transplantation
1b: Relative contribution in cumulative incidence of main cause of death. Cause of death over time is allocated for deceased patients only
Relative contribution to cumulative incidence
100%
Relapse 75%
GvHD 50%
Infections 25% Other causes 0% 12.00 24.00 36.00 48.00 60.00 72.00 84.00 96.00 108.00 Months since transplantation
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4. Special considerations for specific conditioning regimens 4.1. TBI TBI requires specific knowledge and should only be applied at an experienced institution under the guidance of a specialist radiophysicist or radiation oncologist. It has the specific advantage of having access to so-called "sanctuary sites" of malignancies, e.g. CNS or gonads, where some drugs may not penetrate. TBI is most commonly delivered to the patient using a linear accelerator. Patients are placed within a single irradiation field and lie on their side or in lateral position at a specified distance from the radiation source. Irradiation is given with horizontal beams using the anterior-posterior or lateral-lateral technique. Effects of TBI depend on total dose, dose rate and fractionation. TBI is usually prescribed in one of three ways: - as a single dose (sTBI) from 1 or 2 Gy up to 7.5–8 Gy total dose, - as a fractionated dose (fTBI) in 5–6 fractions over 3 days to a total of 10–14 Gy or - as "hyperfractionation" TBI consisting of 10–12 fractions over 4 days, usually to a total of 14–15 Gy. Dose rates differ considerably between machines and have different radiobiological effects. Lower total doses must be applied when higher dose rates are used. Fractionation reduces the incidence and severity of acute and late complications of normal tissue. As such TBI is similar to conditioning with chemotherapy. Low total doses, e.g. 6 Gy, may decrease toxicity but increase the risks of graft failure and disease recurrence. Very high doses, e.g. 15.75 Gy, may reduce the incidences of graft rejection and disease relapse, but at the expense of increased morbidity and mortality. Radiation dose is normally calculated as midline tissue dose at the level of the umbilicus but other strategies, e.g. targeting maximum lung dose, are possible. The received radiation doses to various parts of the body are monitored by in vivo dosimetry using diodes and thermoluminescent detectors placed anteriorly and posteriorly on various parts of the body. The doses given using standard techniques are relatively uniform and most parts of the body receive within ±10% of the dose delivered to the abdomen. In some institutions parts of the body, usually the lungs or the eyes, are protected with lead blocks during part of the application to "shield" this organ and to reduce organ specific toxicity. Other centres avoid shielding since leukaemic cells could also be protected by the shields. Dosimetry is still a difficult undertaking and measured dose is less exact than calculated delivered dose. Monitoring during TBI is essentially required to detect major
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technical errors, e.g. too high or too low an applied dose. Immediate effects of TBI include a transient increase in granulocyte numbers. Simultaneously, lymphocyte numbers fall. Changes in the serum concentration of various hormones are observed. Cortisol, adrenocorticotropic and corticotropin releasing factor levels increase a few hours after the completion of TBI with a concomitant increase in IL-6 and TNF levels. Some of these findings may be explained by activation of the hypothalamic-pituitary-adrenal axis. The mechanism by which TBI kills leukaemic (and normal) cells is still a matter of debate. TBI induces DNA and cell membrane damage, which triggers apoptosis. Data suggest that there may be a correlation between the magnitude of the immediate apoptotic response and "radiocurability". Similarly, "survival factors" for specific cells, e.g. cytokines, such as flt-3 ligand can protect stem cells from the toxic effects of TBI by their anti-apoptotic effects. Immediate side effects of TBI are nausea, vomiting and, typically, parotid swelling. Adequate preventive measures are recommended. Pilocarpin p.o. is used to reduce parotid swelling. 4.2. TCD and conditioning T-cells are an essential component of the graft product and for engraftment. Graft facilitating cells in the T-cell fraction enhance engraftment via soluble factors and by their direct effect on residual host T-cells. Simultaneously, T-cells are responsible for acute and chronic GvHD and for graft versus leukaemia (GvL) effects. Therefore, TCD of the stem cells prior to infusion is highly effective in the prevention of acute and chronic GvHD, but at the expense of an increased risk of graft rejection and disease recurrence. TCD is essential in haploidentical transplants to prevent lethal GvHD. The incidence of graft failure and graft rejection in TCD HLA-identical sibling HSCT is in the order of 10–20% with standard conditioning, and is higher in haploidentical, UD HSCT or RIC HSCT. Several approaches have been used to overcome the increase in graft failure associated with TCD. Some centres have intensified CT/RT by increasing dosages or adding TLI or TAI (total lymphoid/abdominal irradiation). This approach can effectively overcome graft failure but is associated with increased morbidity and mortality, resulting in similar OS rates. Because TCD is so effective at preventing GvHD, no further GvHD prevention e.g. with CsA and/or MTX is given after HSCT. The re-introduction of CsA to these protocols can reduce the rate of graft failure/rejection. Other centres combine TCD with additional immunosuppressive therapy for patients prior to stem cell infusion, e.g. in vivo Campath-1H, ATG or MoAb to OKT-3 for several days before HSCT. It may also be possible to overcome graft failure by increasing the SC dose as has 136
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been shown not only in murine models of transplant but also in haplo-identical family member transplants for acute leukaemia. Addition of T-cells which are depleted of alloreactive T-cells or selected NK cells are novel approaches under current investigation. 4.3. Paediatric transplants There are two main differences between adult and paediatric patients in relation to conditioning. Children in general tolerate side effects better than older patients and higher total doses may be applied. In contrast, conditioning affects growth and endocrine development. Retarded growth and failed or retarded puberty are main late sequelae in paediatric HSCT patients. There is no general consensus between the various paediatric groups about the best conditioning. This reflects the uncertainty about the optimal regimen. Nevertheless, a general outline of the strategies used in children can be given. Since many reports comparing TBI containing conditioning regimens with those containing only CT show similar outcomes, TBI should be avoided in small children, whenever possible. It should probably never be given to children below 2 years of age. TBI (2 doses per day) in combination with Cy and/or VP16 is commonly used in children undergoing HSCT for ALL. These children have usually already received all the drugs useful in this disease and TBI is thought to have an additional antileukaemic effect in ALL. Probably the most frequently used conditioning protocols in paediatric HSCT are: 1. Fractionated TBI + Cy and/or VP in ALL (>2 years of age) 2. Cy + Bu (+ MEL) in AML, CML and MDS.
5. Disease specific aspects 5.1. Inborn errors of the lymphohaematopoietic system In patients with profound deficiencies of the lymphoid immune system, such as SCID, the capacity for graft rejection is extremely low or absent. Preparative regimens are therefore unnecessary but HSCT results in an unusual form of lympho-haematopoietic chimerism. Donor cell engraftment after HLA identical sibling HSCT is usually restricted to the lymphocytes, whilst the myeloid system remains of host cell origin. The same approach in TCD HLA-mismatched HSCT has led to more variable results. Here, complete or partial graft failure, in particular failure of B-cell reconstitution, has been observed. This experience has resulted in the use of CT based preparative
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regimens in patients transplanted for SCID from HLA-mismatched donors. The regimen most frequently employed is that of Bu 2 mg/kg/day for 4 d + Cy 50 mg/kg/d for 4 d. In the presence of serious clinical conditions, e.g. pulmonary infections prior to the transplant, the use of chemotherapy may greatly increase the morbidity and mortality of the procedure. In these cases, HSCT may be performed without conditioning, as even partial immunological reconstitution may be of therapeutic benefit to the child. In less severe variants of lymphoid deficiencies and in all other inborn errors, conditioning is required for myelo-ablation and to prevent graft rejection and disease recurrence. Some young patients come to transplant with a number of complications related to their underlying disease, in particular poorly controlled chronic infections. The use of non-CT based conditioning protocols which allow stable engraftment remains a long-term goal. 5.2 Severe aplastic anaemia The marrow is empty in patients with SAA and the sole aim of the conditioning protocol is to provide immunosuppression is. Cy 50 mg/kg for 4 successive days is an appropriate conditioning regimen. It is very immunosuppressive and its use involves few serious long-term side-effects. Rejection remains a concern, especially in patients sensitised to their donors via blood products. The addition of ATG has reduced risk of rejection in one pilot study but prospective randomised and retrospective multicentre analyses have failed to confirm its value in nonsensitised recipients. Still, it is the most frequently used approach at the present time. The optimal conditioning regimen for patients with SAA and alternative donor transplants still needs defining. The addition of Fluda or low dose TBI (2 Gy) is under current investigation. All long-term follow up studies have shown a higher incidence of late malignancies in SAA patients given TBI or TAI during conditioning. 5.3. Fanconi’s anaemia Fanconi’s anaemia is a genetic disorder associated with diverse congenital abnormalities, progressive BM failure and increased risk of leukaemia and other cancers. HSCT is an effective treatment. The underlying molecular defects are heterogeneous but all result in defective DNA repair mechanisms. These patients are therefore extremely susceptible to the effects of CT/RT, in particular to irradiation and alkylating agents. Conditioning regimens must be reduced in intensity. Most regimens contain Cy 5–10 mg/kg/d for 4 days. Some centres add low dose TBI (400–450 cGy) or TAI or TLI (500 cGy) to overcome graft rejection. More recent approaches investigate the use of Bu or the addition of Fluda. 138
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However, there is concern that the addition of TBI may increase the incidence of secondary malignancies in a group of patients already susceptible to further malignant disease. 5.4. Lymphoma The conditioning regimens for high dose therapy in lymphoma have been based on custom and practice rather than proof or confirmation by randomised trials. Most frequently used regimens avoid inclusion of TBI. Auto-HSCT: BEAM (BCNU, VP, ARA-C and MEL) has been the most commonly used regimen both for NHL and HD. Originally described jointly by groups in Lyon and London, it has become standard treatment both in Europe and, to a lesser extent, in the United States. The standard BEAM uses a total dose of VP of 800 mg/m2/d for 4 days. Several groups have attempted to change the individual components. None has proved superior and there is no evidence that any other regimen has more anti-tumour effect than BEAM. Allo-HSCT: BEAM and CBV have both been used in allografting but neither regimen is fully myeloablative. Fluda based RIC HSCT have been proposed. TBI plus CT is considered as the standard preparative regimen for allo-HSCT. The incidence of TRM is clearly associated with the status of the patients at the time of transplant. If transplant is performed for refractory HD or NHL, the TRM reaches 30–40%. Bu and Cy have also been used but Bu is not in common use in the treatment of lymphoma. There are no comparative data for Bu/Cy versus TBI containing regimens or for RIC versus standard conditioning HSCT. 5.5. Myeloma Relatively clear data are available concerning the best conditioning regimen for MM. Auto-HSCT: retrospective data on several thousand autologous HSCT for MM show clear results. Survival is best with a conditioning using MEL 200 mg/m2. No other regimen has shown better results. Allo-HSCT: standard conditioning with Cy TBI shows best long-term survival. RIC HSCT reduces early mortality but retrospective comparative data show inferior survival at +5 years. RIC HSCT may be used in planned double autologous allogeneic HSCT programs. The best approach is not known. 5.6. Solid tumours Dose intensification was the basic concept when auto-HSCT became an investigational tool for treatment of solid tumours. Initial results appeared promising and there was huge interest in the mid-nineties. Conditioning regimens were developed with "tumour specific" drugs in mind and a multitude of regimens appeared. None has
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proved to be superior to any other and HSCT in solid tumours with very few exceptions is still considered investigational. Conditioning regimens should follow those specified in the clinical protocols. They are discussed in the Chapter on solid tumours. Allo-HSCT is being investigated in more recent years as a tool for promoting graft vs. tumour effects. Hence, primarily RIC regimens are chosen with the aim of establishing donor type engraftment. Any RIC regimen can be used without a need for tumour specific therapy. RIC HSCT for solid tumours is still to be considered investigational.
6. Conditioning induced organ damage The desirable effects of conditioning regimens are offset by their highly predictable toxicities, which are responsible for considerable morbidity and mortality (see Table 3). These involve the GI, renal, hepatic, pulmonary and cardiac systems and include cardiac toxicity (Cy >1.5 g/m2/d; BCNU), pulmonary toxicity (TBI >8 Gy), mucositis, hepatic toxicity, bladder toxicity, renal toxicity and neurological toxicity. Prevention and treatment of early and late complications are discussed in detail in Chapters 9 and 12.
7. Quality management issues Application of high dose therapy in the context of HSCT is a complex and challenging therapy. It is prone to errors and all strategies should be undertaken to minimise such risks. This includes specifically the following points, as indicated in the JACIE standards and as outlined in Chapter 4. Each institution administering high dose CT in the context of HSCT must establish a quality management system for administration of this therapy. Such a system must be established in close cooperation with the institutional pharmacy and the nursing team. Conditioning has to be based on pre-printed orders. Regular check points at different levels and immediately prior to administration have to safeguard that the right patient is given the specified drugs at the correct dose and appropriate timing. All check points include the “4 eyes” principle. Similar considerations apply to the administration of TBI.
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CHAPTER 6 • Principles of conditioning
Table 3: Factors influencing the outcome of HSCT* Disease factors Stage: increasing stage Chronic phase patients only: Basophil count (>3%) Sokal/Hasford score (higher) Leukocyte count (higher) Time interval (> 12 m) Patient-related factors Age (higher) Sex (male) Race Viral status (CMV positivity) Donor-related factors Histocompatibility (vs. HLA-id sibling) Identical twin Unrelated Mismatched Sex (female D for male R) Viral status (CMV positivity) Peri-transplant factors Conditioning (intensified) (reduced) GvHD prevention (intensified) (reduced) Cell content (CD34) high low Stem cell source: BM vs. PB Post-transplant factors Acute GvHD (increasing grade) Chronic GvHD (none vs. any)
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LFS
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D: donor; R: recipient * These factors are confirmed for CML but apply in principle to most other situations
References 1. Thomas et al. A history of bone marrow transplantation. In: Thomas' Hematopoietic Cell Transplantation, 3rd Edition (Blume KG, Forman SJ, Appelbaum FR Eds.), Blackwell Publishing Oxford, Malden, Victoria, 2004, pp 4-8. 2. Storb RF, Lucarelli G, McSweeney, Childs PW. Hematopoietic stem cell transplantation for benign hematological disorders and solid tumors. Hematology 2003; 372-397. 3. Santos GW, Tutschka PJ, Brookmeyer R. Marrow transplantation for acute non-lymphocytic
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leukemia after treatment with busulfan and cyclophosphamide. N Engl J Med 1983; 309: 1347-1353. 4. Ridell S, Appelbaum FR, Buckher CD, et al. High-dose cytarabine and total body irradiation with or without cyclophosphamide as a preparative regimen for bone marrow transplantation for acute leukemia. J Clin Oncol 1988; 6: 576-582. 5. Gore ME, Selby PJ, Viner C, et al. Intensive treatment of multiple myeloma and criteria for complete remission. Lancet 1989; ii: 879-882. 6. Philips G, Shepherd J, Bernett M, et al. Busulfan, cyclophosphamide and melphalan as a conditioning regimen for bone marrow transplantation in children with myelodysplastic syndrome. Leukemia 1994; 8: 1880-1888. 7. Slavin S, Nagler A, Naparstek E, et al. Non-myeloablative stem cell transplantation and cell therapy as an alternative to conventional bone marrow transplantation with lethal cytoreduction for the treatment of malignant and non-malignant hematologic diseases. Blood 1998; 3: 756-763. 8. Giralt S, Estey E, Albitar M, et al. Engraftment of allogeneic hematopoietic progenitor cells with purine analog-containing chemotherapy: Harnessing graft-versus-leukemia without myeloablative therapy. Blood 1997; 89: 4531-4536. 9. Copelan EA. Hematopoietic stem-cell transplantation. N Engl J Med 2006; 354: 1813-1826. 10.Appelbaum FR. Hematopoietic-cell transplantation at 50. N Engl J Med 2007; 357: 14721475. 11.Armand P, Antin JH. Allogeneic stem cell transplantation for aplastic anemia. Biol Blood Marrow Transplant 2007; 13: 505-516. 12.Blaise D, Vey N, Faucher C, Mohty M. Current status of reduced-intensity-conditioning allogeneic stem cell transplantation for acute myeloid leukemia. Haematologica 2007; 92: 533-541. 13.Attal M, Moreau P, Avet-Loiseau H, Harousseau JL. Stem cell transplantation in multiple myeloma. Hematology (Am Soc Hematol Educ Program) 2007; 311-316. 14.Hegenbart U, Niederwieser D, Sandmaier BM, et al. Treatment for acute myelogenous leukemia by low-dose, total-body, irradiation-based conditioning and hematopoietic cell transplantation from related and unrelated donors. J Clin Oncol 2006; 24: 444-453. 15.Secondino S, Carrabba MG, Pedrazzoli P, et al.; European Group for Blood and Marrow Transplantation Solid Tumors Working Party. Reduced intensity stem cell transplantation for advanced soft tissue sarcomas in adults: A retrospective analysis of the European Group for Blood and Marrow Transplantation. Haematologica 2007; 92: 418-420. 16.Satwani P, Cooper N, Rao K, et al. Reduced intensity conditioning and allogeneic stem cell transplantation in childhood malignant and nonmalignant diseases. Bone Marrow Transplant 2007; 26: in press. 17.Barrett AJ, Savani BN. Stem cell transplantation with reduced-intensity conditioning regimens: A review of ten years experience with new transplant concepts and new therapeutic agents. Leukemia 2006; 20: 1661-1672.
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CHAPTER 6 • Principles of conditioning
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Total body irradiation (TBI) is frequently used for conditioning in HSCT. Its biological effects depend primarily on: a) Radiation source . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Total radiation dose applied . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Combination of dose rate and fractions of doses . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Combination of total dose, dose rate and fractions of doses . . . . . . . . . . . . . . 2. The type of conditioning regimen is more important in autologous than in allogeneic HSCT, because: a) Graft versus tumour reaction is the only element in allogeneic HSCT for tumour control . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Dose intensification of tumour specific therapy is the key element of autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) There is no specific chemotherapy in allogeneic HSCT. . . . . . . . . . . . . . . . . . . . . . d) Monoclonal antibodies, such as CD20+ antibodies can only be integrated in the conditioning in autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . 3. Which of the following statements about reduced intensity conditioning (RIC) is correct? a) RIC has a specific advantage in patients with advanced malignancies . . . b) RIC has a specific advantage in patients with non-malignant conditions where no graft-versus-disease effect is required . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) RIC has a specific advantage in CMV positive patients with CMV negative donors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Peripheral blood as a stem cell source is associated with less rejection than bone marrow in patients with RIC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. There is an inherent dilemma in allogeneic HSCT: Increasing conditioning decreases risk of rejection and risk of relapse at the expense of increased toxicity; vice versa, decreased conditioning decreases direct regimen related toxicity at the expense of increased risk of rejection and relapse. Which of the following statements about this dilemma is true?
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a) Different regimens should be employed for patients with high-risk disease and low transplant risk compared to patients with low-risk disease but high transplant risk . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) T-cell depletion can abolish this dilemma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The statement does not hold true for unrelated cord blood transplants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The statement is not correct for non-malignant disorders . . . . . . . . . . . . . . . . . 5. The following conditioning regimens can be generally accepted as “established”; e.g., novel regimens need to be evaluated against this standard: a) Cyclophosphamide +/- ATG for severe aplastic anaemia . . . . . . . . . . . . . . . . . . . . b) Cyclophosphamide + busulfan for Hodgkin’s lymphoma . . . . . . . . . . . . . . . . . . . . c) Cyclophosphamide + TBI for multiple myeloma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Fludarabine for myeloid malignancies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 7
Transfusion policy
D.H. Pamphilon
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CHAPTER 7 • Transfusion policy
1. Introduction Haematopoietic stem cell transplant (HSCT) patients often require intensive blood component support. Transfusion may be complicated by transfusion transmitted infection (TTI) – both viral and bacterial, transfusion-associated (TA)-GvHD, febrile non-haemolytic transfusion reactions (FNHTR) and transfusion-related acute lung injury (TRALI). Alloimmunisation (AI) to red cell antigens may cause difficulties in selecting compatible blood whilst AI to the human leukocyte antigens (HLA) present on platelets may cause refractoriness to subsequent transfusions of randomly-selected platelets. It is therefore essential to define robust transfusion policies and procedures and these should be regularly audited. This Chapter describes the blood components available for transfusion including granulocytes and their clinical use in the setting of HSCT. The impact of reduced-intensity conditioning (RIC) transplantation on transfusion requirements is highlighted. Amongst infectious agents transmissible by blood components, cytomegalovirus (CMV) is particularly important in BMT patients and strategies to minimise CMV transmission in susceptible recipients are described.
2. General policies for the selection of high quality, appropriate transfusions The European Union Directive 2002/98/EC sets standards for the collection, testing, processing, storage and distribution of human blood and blood components (1). It requires that Blood Establishments should be licensed and this is of importance for both Blood Centres in EU countries that undertake these activities as well as hospitals that collect and issue e.g. granulocytes for transfusion. The most important aspects of the Directive are: • The fate of each unit of all blood components should be recorded and this record kept for 30 years, i.e. donor to recipient traceability • Robust Quality Systems should be in place • The processing of blood and blood components should be undertaken by licensed blood establishments (see above) • Training should be provided for hospital transfusion laboratory staff • Haemovigilance systems should be established to include the reporting of adverse events. Establishments are licensed by the Competent Authorities in EU Member States following inspection by a regulatory body – in the UK this is the Medicines and Healthcare Products Regulatory Authority (MHRA). Reports of compliance must be submitted.
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2.1. Testing of donated blood for infectious disease markers (IDM) A number of microbial agents may be transmitted by blood transfusion. These include hepatitis B and C, HIV-1 and -2, HTLV-1 and 2, CMV and syphilis. Blood Services routinely test blood for the following: • Hepatitis B: Hepatitis B surface antigen (HbsAg)* • Hepatitis C: Hepatitis C antibodies (anti-HCV)* • Human immunodeficiency virus 1 and 2: HIV-1 + 2 antibodies (anti-HIV 1+2)* • Human T-lymphotropic virus 1 and 2: HTLV-1 + 2 antibodies (anti-HTLV-1+2) • Syphilis. * These tests are mandated by the EU Blood Directive (2002/98/EC) (1) In addition, donations may be tested for: • anti-HBc, i.e. anti-hepatitis B core antigen • alanine aminotransferase (ALT): A surrogate marker of hepatitis C • HCV-RNA by PCR for hepatitis C • p 24 antigen for HIV-1. There is a variation from country to country in the number of tests performed on each blood donation. Testing for anti-CMV antibody to identify CMV seronegative donors is done on a proportion of blood donations, sufficient to identify enough CMV seronegative components for transfusion to those patients for whom it is appropriate. Bacterial contamination is a relatively common occurrence with an incidence estimated at 0.05–0.5% of components (2). The sources of bacteria are donor bacteraemia and contamination with bacteria present on the skin at the time of donation or present in blood packs. The organisms that most frequently contaminate red cell and platelet transfusions are shown in Table 1. Screening tests for bacteria in platelet concentrates (PCs) using automated blood culture systems e.g. BacT/ALERT have been evaluated (2) and are now used routinely by some transfusion services. PCs are not issued until at least 48 hours
Table 1: Bacteria that most frequently contaminate blood components Red cells Yersinia enterocolitica Pseudomonas fluorescens Other species
51% 27% 22%
Platelets Staph. epidermidis Staphylococcus aureus Salmonella choleraesuis Serratia marcescens Bacillus cereus Others
Figures are percentage of total contaminants for red cells and platelets respectively 148
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25% 6% 14% 10% 6% 39%
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CHAPTER 7 • Transfusion policy
after collection but the storage period may be extended to 7 days once sterility has been evaluated. The risk of bacterial transmission is also minimised by careful donor selection, meticulous attention to sterility during venepuncture, diversion of the first 30 mL of blood collected (contains most of the bacteria) away from the primary collection pack and sterility during preparation of blood components. Bacterial contamination should be suspected in any patient who develops a febrile reaction characterised by fever, chills ± hypotension. Microbiological testing does not completely remove the risk of TTI, although the chance of infection in the UK after transfusion of screened blood components from known/previously tested donors is estimated to be less than 1 in 2 x 106 for HIV1, HBV and HCV. This risk will vary somewhat according to the donor selection and testing policies that are operative within a Blood Service. 2.2. Blood grouping and antibody testing The ABO and Rhesus D types of all donated blood are determined by standard techniques. This is a requirement of the EU Blood Directive (1). All donations are tested to exclude the presence of immune IgG antibodies that are reactive with common blood groups and which occur after an immunising stimulus such as pregnancy or transfusion. Selected units of red cells may be more extensively phenotyped (Kell, Duffy, Kidd, MNSs antigens) for patients who develop red cell alloantibodies. 2.3. Prevention of CMV transmission A proportion of HSCT patients are CMV seropositive pre-transplant or have seropositive donors. They require regular screening by PCR and antigenaemia testing together with ganciclovir therapy where appropriate to minimise the impact of virus reactivation and prevent clinical infection post-transplant. All CMV seronegative HSCT patients with CMV seronegative donors (neg/neg) and CMV seronegative patients with haematological and other disorders who are likely to proceed to a transplant should receive blood components that have a minimal risk of causing CMV acquisition (3). Studies show that the use of CMV seronegative components is associated with a less than 3% incidence of CMV infection and/or disease in neg/neg HSCT. CMV is transmitted via leukocytes, and leukodepletion also minimises the risk of CMV transmission. CMV seronegative and leukodepleted blood components are probably of equivalent efficacy but this view is not generally accepted (4, 5). Further evidence from prospective randomised controlled studies (PRCT) using pre-storage leukodepleted blood components is required. Centres must establish their own policies.
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2.4. Leukodepleted blood components Transfused leukocytes cause alloimmunisation (AI) to HLA Class 1 antigens (HLA AI) in a proportion of patients. This may be manifested clinically as FNHTRs, although these may also be caused by antibodies to neutrophils, platelets or plasma proteins and by cytokines such as interleukin (IL)-1, IL-6, IL-8 and tumour necrosis factor (TNF)-a which accumulate in stored blood components, especially PCs. HLA AI may cause accelerated destruction of transfused platelets that are HLA incompatible. This is clinically manifest as a failure to achieve a satisfactory increment after platelet transfusion (refractoriness). A summary of the adverse effects of transfused leukocytes is shown in Table 2. Donor dendritic cells (DC) which are present in red cell and platelet transfusions appear to be responsible for sensitisation to HLA. Studies show that removal of leukocytes to less than 5 x 106 per blood component prevents primary HLA AI in >97% of patients with haematological malignancies. The use of leukodepleted components also reduces secondary AI and refractoriness to platelet transfusion. Refractoriness is not always prevented since in >50% of cases it results from increased platelet destruction due to non-immune causes which include fever, splenomegaly, DIC and amphotericin therapy. AI is also associated with a higher incidence of graft failure in patients with severe aplastic anaemia. Filtration of blood or its components is best performed in Blood Centres and hospital blood banks. Data from studies where leukocytes were filtered from blood components at the bedside show that this may not be effective in preventing or reducing FNHTR, AI and refractoriness.
Table 2: Adverse effects of transfused leukocytes
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HLA alloimmunisation causing
- FNHTR - Refractoriness to random donor platelets - Graft rejection - Shortened red cell survival
Transmission of microorganisms
- CMV - HTLV-1/11 - Toxoplasma gondii - Yersinia enterocolitica
Immunomodulation
- GvHD - Activation of viruses in host cells e.g. HIV-1 - Immune suppression of T- and NK-cell functions
Affecting the quality of stored blood
- Microaggregate formation - Metabolic deterioration during storage
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2.4.1. Indications for leukodepleted blood components (6) • Pre-HSCT in patients with SAA to reduce the likelihood of graft failure; • Pre- and post-HSCT to prevent recurrent FNHTR; • Pre-and post-HSCT to minimise HLA AI and platelet refractoriness. This is optional since there is no evidence of a significant impact on important clinical outcome measures such as survival post-HSCT except in patients with SAA. Nonetheless many Blood Services have implemented leukodepletion of a large proportion or, in some cases, all of their blood components. In the UK universal leukodepletion was implemented in 1999 with the aim of minimising the risk of transfusionassociated transmission of the causative agent of variant Creutzfeld-Jakob disease (vCJD); • As an alternative to CMV seronegative components. 2.5. Gamma-irradiation of blood components and TA-GvHD HLA incompatible third party leukocytes contained in donated blood components can engraft and initiate an alloreactive response after transfusion. This can cause TA-GvHD, manifest clinically by fever, rash, diarrhoea, jaundice and pancytopenia, and this is fatal in >90% of cases, so prevention is essential. Donor leukocytes are inactivated by gamma-irradiation of 2500 cGy and all components for HSCT recipients should be irradiated from the time that conditioning therapy is started and continued until 6 months post-transplant or until the lymphocyte count is 1 x 109/L in the absence of chronic GvHD. In addition, HLA matched PCs should be irradiated, as should those from family members, since HLA haplotype sharing may result in TA-GvHD even in immunocompetent patients. A summary of the indications for blood component irradiation is shown in Table 3. Platelets show normal functional characteristics through 5 days storage after irradiation with doses up to 5000 cGy. Red cells leak potassium during storage and
Table 3: Indications for irradiated blood components • • • • • • •
Allo-HSC recipients from time of conditioning therapy for 6 months or until the lymphocyte count is 1 x 109/L in the absence of cGvHD Allo-HSC donors Auto-HSC recipients (from 7 days before harvest until 3 months post transplant) All donations from HLA-matched donors or 1st or 2nd degree relatives All patients with Hodgkin disease at any stage of therapy All patients treated with purine analogues e.g. fludarabine All patients with congenital immunodeficiency states
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this is worsened by irradiation. Therefore, storage is limited to 14 days after irradiation with 2500 cGy. TA-GvHD has been shown to occur after 1500–2000 cGy and this dose range is not recommended (7).
3. Pre-transplant transfusions The following provisions apply: • Red cell transfusions for patients with sickle cell disease are initially matched for ABO, Rhesus D and Kell antigens but additional matching for the Rhesus CcEe and for Duffy (Fya Fyb), Kell (Kk), Kidd (Jka Jkb) and MNSs antigens may be required if the patient has developed alloantibodies; • Leukodepleted blood components should be transfused to all patients with aplastic anaemia (6); • Either CMV seronegative or leukodepleted blood components should be transfused to susceptible patients to prevent CMV acquisition (3); • Blood components should be gamma-irradiated for PBSC transplant patients during stem cell mobilisation and collection since transfused leukocytes might be captured in the PBSC harvest and subsequently induce TA-GvHD (7); • Blood components transfused to allogeneic marrow donors immediately pre- or intra-operatively should also be irradiated (7).
4. Blood component transfusions The following definitions for red cells, PCs, FFP, cryoprecipitate and granulocyte products were derived from the Guidelines for UK Blood Services. The requirement for red cell and platelet transfusions is decreased in the setting of RIC transplantation (8). 4.1. Red cells Red cells, usually suspended in an optimal additive solution (OAS) are transfused to correct anaemia due to marrow failure, haemorrhage or haemolysis, aiming to keep the haemoglobin or packed cell volume (PCV) above predefined levels to ensure good tissue oxygenation. Reduced intensity conditioning (RIC) transplants require fewer red cell transfusions. Transfusions may be: • Suspended in OAS, usually a combination of saline, adenine, glucose and mannitol (SAG-M): PCV 50–70%; volume 220–420 mL. This is the product of choice; • Derived from whole blood from which a proportion of the plasma has been removed – plasma reduced blood (PRB): PCV 50–60%; volume 200–450 mL; • Unmodified whole blood: volume 420–520 mL. This last term is misleading since platelets and labile coagulation factors deteriorate rapidly in stored blood. 152
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The storage period is 35–42 days at 4 ± 2ºC. Transfusion policy • Red cells should be matched for ABO and Rhesus D type (1). • Extended phenotyping may be necessary in patients, e.g. those with sickle cell disease, who have formed red cell alloantibodies after previous transfusions. • Red cells should be cross-matched against the patient’s serum by standard techniques prior to transfusion. • Thresholds should be defined for haemoglobin and PCV below which red cell transfusions are always given. Suggested arbitrary cut off points are Hb less than 8.0 g/dL and PCV less than 25%. • In adults 1 unit of red cells raises the Hb by 1.0 g/dL whereas in children the volume of blood to be transfused is derived from the formula: Volume = Increase in Hb (g/dL) required x 4 x weight (kg) 4.2. Platelet transfusions 4.2.1. Manufacture Platelet concentrates (PCs) are made: • From whole blood by centrifuging units in a “top – top” pack format to obtain platelet rich plasma (PRP), which is then further concentrated to give a PC. PRPPCs may be transfused individually or pooled in multiples – usually 6; • From whole blood by centrifuging units in a “bottom & top“ pack format to separate the buffy coat (BC), pooling 4 BCs and recentrifuging to separate PRP which is then expressed into a secondary storage container for PC preparation; • By collecting PCs directly on a cell separator. Dual arm, continuous flow apheresis is preferred and some cell separators collect PCs with an inherently low WBC content (Table 4) (9).
Table 4: Platelet content and WBC contamination of different types of platelet concentrates PRP-PC
BC-PC
Apheresis PC
Mean platelet Content x 1011/unit
3.4
3.2
3.15
Mean WBC Contamination x 106/unit
365
5.7
0.3
PRP-PC: platelet concentrate prepared from whole blood; BC-PC: platelet concentrate prepared from buffy coat; Apheresis PC: platelet concentrate obtained directly by apheresis. Data from the National Blood Service, Bristol Centre (9)
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The storage period is 5 days at 22 ± 2ºC unless bacterial screening has been carried out in which case it may be extended to 7 days. Transfusion policy Current practice, based on the results of randomised studies, is to transfuse PC prophylactically when the platelet count is less than 10 x 109/L. A recent Cochrane Systematic Review concluded that, whilst there is no reason to change current practice, blood products may become scarcer and further trials should be undertaken to compare prophylactic versus therapeutic platelet transfusion – i.e. PC given only when there is clinical bleeding (10). In autologous PB HSCT this has been found to be safe (11). Fewer PC transfusions are required in RIC allografted patients compared to those who receive full myeloablative conditioning (8). Best current practice is that: • PCs should be ABO and Rh compatible wherever possible since ABO incompatibility may reduce the expected count increment (CI) by 10–30%; • Group O PCs should be tested for high titre anti-A, B and if positive should only be transfused to group O recipients to avoid haemolysis caused by passive administration of antibody; • If Rh D positive platelets are given to an Rh D negative patient then give 250 IU polyclonal anti-Rh (D) immunoglobulin. Since the chance of Rh immunisation is probably less than 5% this may be omitted and the patients serum screened for immune red cell antibodies, or prior to a red cell transfusion; • Studies show that a threshold of 10 x 109/L in stable thrombocytopenic patients is optimal for prophylactic platelet transfusion; • A higher threshold of 20 x 109/L should be used in patients with fever, sepsis, splenomegaly and other well-established causes of increased platelet consumption; • If an invasive procedure is planned, e.g. central line insertion, the platelet count should be >50 x 109/L; • PCs should be transfused when there is significant clinical bleeding, irrespective of the platelet count; • PCs are contraindicated in patients with TTP; • In adults the usual dose of platelets is 3 x 1011 (an adult therapeutic dose – ATD) in a volume of 200–300 mL; • Children >30 kg receive one ATD. Children <30 kg are given 10 mL/kg; • Rate of transfusion: adults 1 ATD is given in less than 60 minutes children e.g. 2–5 mL/kg/hr. The outcome of platelet transfusions can be monitored by: • Looking for cessation of bleeding; • Measuring the platelet count the following day. A persistent value <20 x 109L suggests refractoriness; 154
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•
Measuring the platelet count at between 10–60 minutes post-transfusion – the CI. A corrected (C) CI is calculated as follows: CCI =
CI x 109/L x surface area (m2) Platelets transfused x 1011
The CCI should be more than 7.5. If the patient is refractory to transfusion of PCs, samples should be taken to test for HLA antibodies. If these are detected, HLA-matched platelets collected by apheresis of HLA-typed donors should be used in these patients. If the CCI is less than 7.5 following transfusion of HLA matched PCs and the patient is not bleeding then withhold platelet transfusions. If the CCI using well-HLA-matched PCs is less than 7.5 and/or bleeding persists then: • Check for non-immune causes of refractoriness. If refractoriness is due to nonimmune causes, particularly if there is significant clinical bleeding then either give 2 or 3 ATD or give 1 ATD twice or three times daily; • Look for platelet-specific antibodies – this is a rare cause of refractoriness in HSCT patients; • Consider using cross-matched platelets. HLA-typed or random units or platelets are cross-matched against the patient’s serum usually by an immunofluorescent technique and non-reactive units selected if possible. 4.3. FFP and cryoprecipitate transfusion Fresh frozen plasma (FFP) may be given to correct the abnormalities of coagulation that are observed where there is, for example, liver disease resulting from graft versus host disease (GvHD) or veno-occlusive disease (VOD), thrombotic thrombocytopenic purpura (TTP) or disseminated intravascular coagulation (DIC). • FFP is made by centrifuging whole blood and freezing separated plasma within 6 hours of collection. The volume is 200–340 mL and the Factor VIII level should be greater than 70 IU/mL. • Cryoprecipitate is made by thawing FFP at 4ºC and collecting the precipitate that forms by further centrifugation in a volume of approximately 20–30 mL. This is then refrozen. The fibrinogen content should be greater than 140 mg/dL and the Factor VIII level greater than 70 IU/mL. FFP and cryoprecipitate have a storage period of 12 months at –30ºC. Transfusion policy FFP transfusion (at a volume of 10–15 mL/kg) is indicated after HSCT: • As replacement fluid in TTP where plasma exchange is undertaken;
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In the presence of liver disease causing significant defects of coagulation factors; In severe DIC. FFP transfusion may also be indicated after HSCT where a large volume blood transfusion, e.g. after haemorrhage, has caused a dilutional coagulopathy. Cryoprecipitate transfusion is indicated in severe DIC when the fibrinogen is <100 mg/dL. The outcome of FFP and cryoprecipitate transfusion should be monitored by measuring the prothrombin time (PT) and activated partial thromboplastin time (APTT). The ratios compared to control should correct to less than 1.5. In DIC the fibrinogen should be greater than 100 mg/dL. •
•
4.4. Granulocyte transfusions Granulocyte transfusions (GT) are prepared by pooling buffy coats from e.g. 10–20 whole blood donations and then reducing the red cell content further by sedimentation using starch or dextran. They may also be collected by the apheresis of steady state healthy donors who may be family members or unrelated volunteers. The granulocyte content is in the range 5–10 x 109/unit for both these preparations. Mobilised granulocytes are collected from donors who receive G-CSF (5–10 micrograms/kg) and/or dexamethasone (8 mg) – both given 12–24 hours before to increase the number that can be collected during a standard apheresis procedure. This strategy gives a granulocyte yield of between 10–100 x 109 per unit and data available so far indicates that significant granulocyte increments e.g. 1–2 x 109/L can be obtained. By contrast it is unusual to observe such increments with buffy coat or unmobilised granulocytes. All granulocyte products must be irradiated prior to transfusion to prevent TA-GvHD. Cross-matching is also required. A recent Cochrane Systematic Review indicated that there is currently inconclusive evidence from PRCTs to support or refute the use of GT in neutropenic patients. Further PRCT are required before definitive recommendations can be made (12). Transfusion policy • There is recent anecdotal evidence that prophylactic administration of granulocytes may reduce the incidence of severe fungal infections after BMT but currently few centres use such transfusions and further studies are needed (13). Furthermore, granulocyte transfusions increase the likelihood of HLA immunisation and platelet refractoriness. • Granulocyte transfusions are probably best reserved for patients with granulocyte counts less than 0.2 x 109/L and documented bacterial or fungal infections not responding to at least 3 days of appropriate antimicrobial therapy, in situations where the granulocyte count is not expected to recover within 7 days. 156
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5. Donor/recipient ABO incompatibility and transfusion support 5.1. Background Approximately 15–25% of HLA identical sibling donor/recipient pairs are ABO incompatible. The figure is higher in alternative donor transplants. In myeloablative transplants ABO incompatibility is associated with an increased risk of delayed red cell engraftment, pure red cell aplasia (PRCA), haemolysis and increased transfusion requirements. There are some reports of increased platelet transfusion requirements. ABO mismatch does not affect neutrophil engraftment, the incidence of graft rejection, GvHD, disease progression or overall survival (14). In RIC transplants it was demonstrated by chimerism studies that early erythroid progenitors engrafted as promptly as myeloid progenitors (15). However, as with myeloablative HSCT, engraftment of mature red cells is delayed, cases of PRCA have also been reported and ABO mismatch is associated with increased red cell transfusion requirements (16). Recipient plasma cells produce anti-donor ABO alloagglutinins and after RIC HSCT the rate of decline of anti-donor alloagglutinins takes twice as long as after myeloablative conditioning, and this can lead to more haemolysis (17). In one report of 40 patients who had RIC HSCT, ABO mismatch was associated with one death due to haemolysis, 3 cases of PRCA, 6 cases of thrombotic microangiopathy (3 fatal), an increase in rehospitalisation days, relapse or disease progression and higher TRM (5, 18). By contrast other reports do not show an inferior outcome (15–17). The same authors also reported in 2007 on 32 RIC HSCT; in 10 cases there was a minor or major plus minor (bidirectional) mismatch and in 5 this caused severe haemolysis leading to death in 3 patients (19) (see also 5.3 below). 5.2. Definitions • Major ABO incompatibility is defined as the presence in the recipient's plasma of anti-A, -B or -A,B alloagglutinins reactive with the donor’s red cells, e.g. donor group A and recipient group O. • Minor ABO incompatibility is defined as the presence of anti-A, -B or -A,B alloagglutinins in the donors plasma reactive with the recipient’s red cells, e.g. donor group O and recipient group A. • Major plus minor ABO (bidirectional) incompatibility is defined as the presence in both the donor and recipients plasma of anti-A, -B or -A,B alloagglutinins reactive with recipient and donor cells respectively, e.g. donor group A and recipient group B. 5.3. Incompatible stem cell graft infusion If the alloagglutinin titre is less than 1:64 unmodified bone marrow or PBPC grafts
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may be infused. At higher titres red cells should be removed from the graft. Marrow processing or PBPC collection on certain apheresis machines e.g. the GAMBRO Spectra, usually results in red cell contamination of less than 5 mL and ABO incompatibility may be ignored. Likewise, stem cells that are separated on density gradients and washed can also be infused without regard to ABO incompatibility. Plasma may be removed from the transplant in cases of minor ABO mismatch where the e.g. anti-A titre is high to avoid acute haemolysis in the recipient. Delayed haemolytic transfusion reactions may follow the infusion of donor HSC where there is a minor ABO mismatch. This is called Passenger Lymphocyte Syndrome (PLS) and occurs because of a secondary (anamnestic) immune response mediated via memory B-cells in the graft against recipient ABO antigens. A rise in anti-A, -B or -A,B titre is seen together with anaemia and jaundice. This phenomenon is rarely, if ever, seen when bone marrows are depleted of alloreactive T-lymphocytes using strategies such as alemtuzumab (Campath-1) antibody or CD34 positive cell selection since B-cells are also depleted. In RIC HSCT minor or bidirectional ABO mismatch can be associated with severe and even fatal haemolysis and in one study recipient red cell exchange was performed pre-transplant. This reduced TRM from 53 to 16% and overall survival was improved (65 vs. 40%; both p<0.05) (19). 5.4. Blood groups used for transfusion support Pre-transplant, recipient-type red cells and platelets should be given. Post-transplant (see Figure 1): • For major ABO mismatch use group O red cell products, irrespective of ABO group of recipient or donor until recipient ABO antibodies are undetectable and the antiglobulin test is negative. Give platelets and plasma from donors of the recipient’s ABO type until recipient red cells are no longer detected; • For minor ABO mismatch use red cells of the donor type, i.e. group O throughout. Give platelets and plasma of recipient type until recipient-type red cells are no longer detected; • For major and minor ABO mismatch use group O red cells until recipient ABO antibodies are undetectable and the antiglobulin test is negative and then switch to donor type. For platelets and plasma use group AB until recipient red cells are undetectable; • Following graft rejection, revert to recipient-type red cells and platelets.
6. Conclusion Transfusion support in BMT patients requires special consideration and carefully defined policies. The use of high quality blood components which have a high degree 158
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Figure 1: Strategy for the provision of blood components in ABO mismatched HSCT Recipient group
Group 0
Group AB
Donor group
Major ABO incompatibility
Red cells Plasma/platelets Minor ABO incompatibility
Red cells Plasma/platelets Major and minor ABO incompatibility
Red cells Plasma/platelets 1 1 2 3
2
3
HSCT ABO antibodies to donor RBC not detected. Direct antiglobulin test negative RBC of recipient group no longer detected
of microbiological safety and which are also gamma-irradiated and, in addition, may be CMV seronegative and leukodepleted provides optimum transfusion support and minimises the chance of adverse effects.
References 1. European Union Directive 2002/98/EC http://eur-lex.europa.eu/LexUriServ/site/en/oj/2003/l_033/l_03320030208en00300040.pdf 2. Ramirez-Arcos S, Jenkins C, Dion J, et al. Canadian experience with detection of bacterial contamination in apheresis platelets. Transfusion 2007; 47: 421-429. 3. Pamphilon DH, Rider JR, Barbara JA, Williamson LMP. Prevention of transfusion-transmitted cytomegalovirus infection. Transf Med 1999; 9: 115-123. 4. Narvios AB, de Lima M, Shah H, et al. Transfusion of leukoreduced cellular blood components from cytomegalovirus-unscreened donors in allogeneic hematopoietic transplant recipients: Analysis of 72 recipients. Bone Marrow Transplantation 2005; 36: 499-501. 5. Nichols WG, Price TH, Gooley T, et al. Transfusion-transmitted cytomegalovirus infection after receipt of leukoreduced blood products. Blood 2003; 101: 4195-4200. 6. BCSH Blood Transfusion Task Force. Guidelines on the clinical use of leucocyte-depleted
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blood components. Transf Med 1998; 8: 59-71. 7. BCSH Blood Transfusion Task Force. Guidelines on gamma irradiation of blood components for the prevention of transfusion-associated graft-versus-host disease. Transf Med 1996; 6: 261-271. 8. Weissinger F, Sandmaier BM, Maloney DG, et al. Decreased transfusion requirements for patients receiving nonmyeloablative compared with conventional peripheral blood stem cell transplants from HLA-identical siblings. Blood 2001; 98: 3584-3588. 9. Anderson NA, Gray S, Copplestone JA, et al. A prospective randomised study of three types of platelet concentrates in patients with haematological malignancy: Corrected platelet count increments and frequency of nonhaemolytic febrile transfusion reactions. Transf Med 1996; 7: 33-39. 10.Stanworth SJ, Hyde C, Heddle N, et al. Prophylactic platelet transfusion for haemorrhage after chemotherapy and stem cell transplantation. Cochrane Database Syst. Rev. 2004 Oct 18; (4): CD004269. 11.Wandt H, Schaefer-Eckart K, Frank M, et al. A therapeutic platelet transfusion strategy is safe and feasible in patients after autologous peripheral blood stem cell transplantation. Bone Marrow Transplantation 2006; 37: 387-392. 12.Stanworth SJ, Massey E, Hyde C, et al. Granulocyte transfusions for treating infections in patients with neutropenia or neutrophil dysfunction. Cochrane Database Syst. Rev. 2005 Jul 20; (3): CD005339. 13.Kerr JP, Liakopoulou E, Brown J, et al. The use of stimulated granulocyte transfusions to prevent recurrence of past severe infections after allogeneic stem cell transplantation. British Journal of Haematology 2003; 123: 114-118. 14.Helbig G, Stella-Holowiecka B, Wojnar J, et al. Pure red-cell aplasia following major and bi-directional ABO-incompatible allogeneic stem cell transplantation: Recovery of donorderived erythropoiesis after long-term treatment using different therapeutic strategies. Ann Hematol 2007 May 8 (Epub ahead of print). 15.Maciej Zaucha J, Mielcarek M, Takatu A, et al. Engraftment of early erythroid progenitors is not delayed after non-myeloablative major ABO-incompatible haematopoietic stem cell transplantation. British Journal of Haematology 2002; 119: 740-750. 16.Canals C, Muniz-Diaz E, Martinez C, et al. Impact of ABO incompatibility on allogeneic peripheral blood progenitor cell transplantation after reduced intensity conditioning. Transfusion 2004; 44: 1603-1611. 17.Griffith LM, McCoy JP, Bolan CD, et al. Persistence of recipient plasma cells and anti-donor isohaemagglutinins in patients with delayed donor erythropoiesis after major ABO incompatible non-myeloablative haematopoietic cell transplantation. British Journal of Haematology 2005; 128: 668-675. 18.Worel N, Kalhs P, Keil F, et al. ABO mismatch increases transplant-related morbidity and mortality in patients given nonmyeloablative allogeneic HPC transplantation. Transfusion 2003; 43: 1153-1161. 19.Worel N, Greinix HT, Supper V, et al. Prophylactic red cell exchange for prevention of severe immune hemolysis in minor ABO – mismatched allogeneic peripheral blood progenitor cell transplantation after reduced-intensity conditioning. Transfusion 2007; 47: 1494-1502.
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Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The European Union (EU) Blood Directive (2002/98/EC) requires that all hospital blood banks should: a) Keep a record of all blood components for 10 years to ensure donor-recipient traceability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Be licensed by EU competent authority following inspection or completion of a compliance report every 3 years . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Establish a haemovigilance system which includes optional reporting of adverse events . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Provide training for all hospital transfusion laboratory staff . . . . . . . . . . . . . . 2. The EU Blood Directive (2002/98/EC) requires that all donated blood for transfusion should be tested for all of the following except one. Which one? a) Hepatitis B (HbsAg) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Hepatitis C (Anti-HCV) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) HIV 1 and 2 (Anti-HIV 1 + 2) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Syphilis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which of the following statements about leukodepleted blood components is correct? a) Routinely contain less than 5 x 106 leuckocytes . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Their use in transplant patients prevents the development of platelet transfusion refractoriness in >80% of the patients who receive them . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Their use in transplant patients prevents CMV acquisition in all allogeneic SCT where both the donor and recipient are CMV negative . . . . d) Leukodepletion is best done at the patient’s bedside . . . . . . . . . . . . . . . . . . . . . . 4. Gamma irradiation of blood components is indicated: a) For all autologous SCT patients until 1 year post-transplant . . . . . . . . . . . . . . b) After allogeneic SCT until the lymphocyte count is >0.5 x 109/L in the absence of chronic GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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c) In all patients with non-Hodgkin’s lymphoma . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) In all patients treated with purine analogues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Platelet transfusions given to allograft recipients should always be: a) ABO and Rh compatible . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Given when the platelet count falls below 20 x 109/L . . . . . . . . . . . . . . . . . . . . . c) Tested for high titre anti-A,B if they are to be transfused to non-group O recipients. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Transfused to patients with TTP when the platelet count falls below 50 x 109/L . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 8
Supportive care
T. Masszi
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CHAPTER 8 • Supportive care
1. Introduction Supportive care in HSCT is essential in optimising the outcome of the treatment process. This Chapter will focus on nutritional support, central venous catheter related problems, the use of haematopoietic growth factors, the management of mucosal injury and anti-emesis strategies.
2. Nutritional support Patients with haematological malignancies can be well nourished at the time of transplantation, whereas malnutrition is more frequent in patients with solid tumours. Impaired nutritional status before transplantation is a negative prognostic factor for outcome after HSCT and better-nourished patients have a shorter time to engraftment. Irrespective of nutritional status, however, parenteral nutritional support is commonly administered prophylactically after HSCT until patients are able to maintain an adequate oral nutritional intake, usually following BM recovery. Although enteral nutrition is theoretically possible, total parenteral nutrition (TPN) is largely favoured in HSCT patients mainly because nausea, vomiting and orooesophageal mucositis prevent the insertion and subsequent tolerability of nasogastric tubes. Moreover virtually all patients undergoing HSCT have a CVC in situ through which TPN can be easily administered. Finally, parenteral nutrition probably allows better modulation of fluid, electrolyte, and nutrient administration which can be of critical importance when complications such as GvHD or VOD arise. 2.1. Total parenteral nutrition The routine use of TPN during transplantation is based on a study performed twenty years ago that randomly assigned 137 previously well nourished patients undergoing BMT for different malignancies (mainly acute leukaemia) to either prophylactic TPN or intravenous maintenance fluids (dextrose, electrolytes, minerals, trace elements and vitamins). Treatment was started during conditioning, and continued for 4 weeks following transplantation. Compared to the control arm, patients receiving TPN had significantly better OS and DFS, and a longer time to relapse. Notably, 61% of control patients eventually required TPN because of a decline in their nutritional status (1). In contrast other studies have not provided strong support for the routine use of TPN in patients HSCT. Moreover, the care of recipients of HSCT has changed significantly over the last twenty years, e.g. hospital stays are shorter due to the increasing use of PBSCT and the use of recombinant haematopoietic growth factors, and a number of questions remain unanswered regarding nutritional support in current management. For instance, in a prospective randomised trial, 57 patients undergoing BMT received either prophylactic TPN or an enteral feeding program (2). TPN was
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associated with significantly more days of diuretic use, more frequent hyperglycaemia and more CVC related complications. Although the patient cohort was small this study suggests that the role of TPN in patients undergoing HSCT deserves further investigation. In study, which was double-blinded, 258 patients were randomly assigned to receive either TPN or hydration in an outpatient setting. Patients who received TPN had a delay in the resumption of 85% of their caloric requirement (16 vs. 10 days), suggesting that the administration of TPN may suppress normal appetite (3). There were no effects of TPN on hospital readmission, disease relapse, or survival. Although this study did not specifically examine whether prophylactic TPN is indicated for patients who receive HSCT, it suggests that after hospital discharge, TPN may not be necessary for all patients. One further study attempted to define subgroups of patients with haematological malignancies receiving intensive chemotherapy (with or without transplantation), who were likely to require TPN (4). They used the following three criteria to define the need for TPN: severe malnutrition at admission, a prolonged (at least 7–10 days) period of minimal oral intake or clinical weight loss exceeding 10% during treatment. Any one of these criteria was sufficient. According to these definitions TPN was found to be necessary in only 55% of patients undergoing HSCT with a range from 37% of autologous recipients to 92% of those receiving an HLA mismatched allograft. 2.2. Caloric, protein and other necessities It is clear that energy expenditure differs between auto- and allo-HSCT recipients. Also, there is a consensus that energy requirements in such patients may exceed 130–150% of the estimated basal energy expenditure. Protein needs are also elevated. The usual amino acid dose in TPN is 1.5–2 g/kg/day. The use of fat as an energy source is safe especially as fat-free TPN can be complicated by hepatic steatosis and glycogenosis. Thus a balanced caloric intake with both fat and carbohydrate is recommended. Long chain triglycerides (LCT), containing saturated fatty acid moieties of 20–24 carbons or a mixture of long chain and medium chain triglycerides (MCT, 6–12 carbons) should provide 30% of caloric intake. MCT seem to be advantageous in TPN over LCT because they are more water soluble, more rapidly cleared from plasma, have protein sparing effects, do not accumulate in the liver or adipose tissue and also do not serve as precursors of eicosanoids (e.g. prostaglandins and leukotrienes) and so might improve immune function and decrease inflammation. However, clinical studies in patients with respiratory illnesses, hepatic dysfunction, and sepsis with multi-system organ failure have suggested that mixtures of MCT and LCT are advantageous compared to LCT alone (5). Electrolytes, minerals, vitamins and trace elements (chromium, zinc, copper, manganese, selenium) 166
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are added to the TPN according to the recommended daily amount. Glutamine supplementation of TPN has been addressed recently, as glutamine is an important precursor for nucleotide synthesis and thus can be a source for rapidly dividing cells, such as gastrointestinal epithelial cells. Although several studies have evaluated the effect of enteral or parenteral administration of glutamine on gastrointestinal toxicity, none have shown a clear preventative or therapeutic effect on intestinal mucositis. On the other hand, prospective studies have suggested positive effects of glutamine administration after HSCT on nitrogen balance, infectious complications and length of hospital stay (6). Preliminary data support the concept that parenteral glutamine may preserve hepatic function in HSCT recipients by maintaining hepatic glutathione concentrations which protect hepatocytes from the oxidant stress of the conditioning regimen. Glutamine supplementation may also have a role in preventing and possibly treating VOD (7). 2.3. Evaluation of nutritional status and monitoring nutritional support Nitrogen balance is considered the most accurate way of assessing nutritional status in HSCT recipients, as it is the direct expression of the imbalance existing between protein breakdown and synthesis. From studies published of TPN in HSCT some kind of consensus can be derived concerning what parameters should be used to monitor nutritional status/support. Daily monitoring of weight (primarily to judge hydration status) is essential, together with electrolytes, BUN/urea, creatinine, and glucose. Liver function tests should be measured at least twice per week as abnormalities can result from TPN. Finally, weekly serum albumin, transferrin (reflective of amino acid intake for visceral protein synthesis) triglyceride and nitrogen balance are also helpful (Table 1). 2.4. Timing of nutritional support Current practice in the timing of TPN is heterogeneous. In the study of Weisdorf et
Table 1: Monitoring of nutritional support during the in-patient stay Daily
Two times a week
Once a week
- weight (fluid balance) - blood glucose - serum electrolytes - BUN - serum creatinine - calorie and protein intake
- liver function tests - serum calcium - serum magnesium - serum phosphorus
- nitrogen balance - serum transferrin - serum albumin - serum triglyceride - serum zinc
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al. (1) (which became the scientific basis of routine administration of TPN in HSCT patients), parenteral nutrition was started before the conditioning regimen and continued to day 28 following HSCT. However in many centres TPN is started only when severe mucositis develops. Others initiate TPN on day 1 post transplant and continue for 2–3 weeks according to the intensity and duration of mucositis. TPN is usually not administered routinely to recipients of auto-HSCT unless prolonged mucositis occurs (8). Simultaneous use of parenteral feeding and oral intake is also contradictory. According to one theory oral intake should be avoided during TPN to minimise the risk of gut contamination. However most studies consider concomitant oral ingestion useful as even small amounts of enteral intake can reduce infection rates or hepatic complications of TPN by avoiding the enteric stasis that results in high levels of endotoxins in the portal venous blood. 2.5. Complications of TPN Complications essentially can be divided into two types: metabolic complications or those connected with the central venous catheter. 2.5.1. Metabolic complications Abnormal liver function is the major metabolic complication of TPN. The initial manifestation is usually an elevation of transaminases, occurring 1–2 weeks after the start of TPN. Increases in serum bilirubin and alkaline phosphatase generally occur 1 or 2 weeks later. These changes often resolve spontaneously without longterm consequences especially if the period of administration of TPN is not longer than 3 months. However there are many other possible causes of elevated liver enzymes after HSCT and the differential diagnosis can be difficult. Drug toxicity, infections, GvHD, VOD, or recurrence of the original malignancy should also be considered (Table 2). If all these possible causes are excluded various other measures can be tried e.g. shortening the period of infusion of TPN from 24 hours to 12–20 hours, introducing some oral feeding, decreasing the non-protein caloric intake and commencing treatment with ursodeoxycholic acid (Table 3). Table 2: Common causes of elevated liver enzymes in the post transplant period Side effects of drugs (methotrexate, cyclosporin-A) Infections (bacterial, fungal, viral) • Veno-occlusive liver disease • Graft versus host disease • Relapse of malignancy • Parental nutrition • •
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Table 3: Treatment modalities in elevated liver enzymes during TPN Search for causes other than TPN (see Table 2) Shorten TPN cycle to 12-20 from 24 hours/day • Reduce the non-protein caloric intake by 10-15% of the total daily calories • Initiate some oral intake if possible • Treat with ursodeoxycholic acid • Utilise oral metronidazole to decrease enteral endotoxin formation • •
2.5.2. Complications related to the central venous catheter Although CVC are indispensable in HSCT, they also represent a significant source of complications including infections, venous thromboembolism, mechanical obstruction, dislodgment and leakage. The most important of these complications are infections, the incidence of which remains high despite the use of aseptic techniques. In a prospective study of 111 HSCT recipients, representing 143 Hickman catheter placements, 44% of patients had positive blood cultures. Most of these (40/63) were coagulase-negative Staphylococci suggesting a primary line infection rather than a secondary contamination from a blood borne source. 2.6. Prevention of central venous catheter related infections A catheter related blood stream infection is defined as bacteraemia or fungaemia in a patient with an intravascular catheter with at least one positive blood culture obtained from a peripheral vein, clinical manifestation of infections (i.e. fever, chills, and/or hypotension) and no apparent source for the blood stream infection except for the catheter. One of the following should be present: • A positive semi-quantitative (more than 15 CFU/catheter segment - i.e. piece of catheter sent for culture) or quantitative (more than 103CFU/catheter segment) culture where the same organism (species and antibiogram) is isolated from the catheter segment and peripheral blood. • Simultaneous quantitative blood cultures with a ratio higher than 5:1 comparing CFU from the CVC sample vs. a peripheral blood sample. • A differential period of CVC culture vs. peripheral blood culture positivity of greater than 2 hours. There are four main possible mechanisms for developing a CVC related infection: • Migration of skin organisms at the insertion site into the cutaneous catheter tract with colonisation of the catheter tip. • Contamination of the catheter hub leading to intraluminal colonisation. • Occasionally, catheters might be haematogenously seeded from another focus of infection and rarely. • Infusate contamination. HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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The most important recommendations concerning the prevention of CVC related blood stream infections are listed in Table 4 based on the guidelines developed in the USA by a working group led by the Infectious Disease Society of America (9).
Table 4: Recommendations for prevention of CVC related blood stream infections
170
Education
Health-care worker education and training for the insertion and maintenance of CVCs is essential. Moreover, periodic assessment of their knowledge of and adherence to is strongly recommended. Trained personnel for the insertion and maintenance of CVCs should be designated.
Catheters and materials
An important pathogenetic determinant is the material from which the device is made. In vitro studies demonstrate that CVCs made of polyethylene or polyvinyl chloride are less resistant to adhesion of micro-organisms than are CVCs made of teflon, silicone or polyurethane. The number of ports must be kept to the minimum required for the patient's management. Cuffed and tunnelled CVCs should be employed if their use is to be prolonged (e.g. allogeneic transplant).
Site of catheter insertion
In adults, a subclavian site is preferred as lower extremity sites are associated with a higher risk of infection (and deep venous thrombosis). Subclavian sites also reduce the risk of infection compared to jugular sites.
Maximal sterile barrier precautions during insertion
Full aseptic techniques should be used at the time of insertion. 2% aqueous chlorhexidine gluconate (preferably), tincture of iodine, or 70% alcohol can be used to prepare the skin before CVC insertion. Organic solvents (e.g. acetone and ether) should not be applied.
Catheter and catheter site care
One port should be designated exclusively for hyperalimentation if a multilumen catheter is used. The routine use of prophylactic intranasal or systemic antibiotics before insertion or during the use of the CVC and antibiotic lock solutions are not recommended. The catheter site can be covered by sterile gauze or a sterile, transparent semi-permeable dressing and the dressings should be replaced whenever they become damp or loosened. Gauze dressings should be replaced at least every two days and transparent dressings every 7 days. The catheter sites must be monitored visually or by palpation regularly and if patients have tenderness at the insertion site and/or fever without obvious source, the dressing should be removed for thorough examination.
Replacement of catheter
Catheters that are no longer essential should be removed promptly. However routine replacement of CVCs to prevent catheter related infections is not advised. CVCs must not be removed on the basis of fever alone. Clinical judgement is required to assess the appropriateness of catheter removal if infection is evidenced elsewhere or if a non-infectious cause of fever is suspected.
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2.7. Conclusions Nutritional support is an integral part of the supportive care of patients receiving HSCT and the main tool remains TPN. Further studies will be required to determine the exact role of TPN in the course of HSCT. Until then, it seems to be prudent to administer TPN to patients undergoing HSCT if they have severe mucositis or gastrointestinal manifestations of GvHD, when a long period of insufficient oral intake is anticipated. The most common complications of TPN are CVC related.
3. Haematopoietic growth factors 3.1. G-CSF and GM-CSF The application of recombinant human haematopoietic growth factors such as G-CSF and GM-CSF have resulted in clear clinical benefits in the post-transplantation setting, especially in auto-BMT and PBSCT. GM-CSF is no longer readily available in Europe but many of the results from clinical trials of GM-CSF can be extrapolated to G-CSF, hence its mention here. Growth factor therapy is usually started 1–3 days after transplantation and continued until the white blood count reaches 5.0–10 x 109/L. 3.1.1. Autologous transplantation Both G-CSF and GM-CSF have been shown to shorten the time to engraftment in randomised phase III trials. Some studies have also demonstrated a reduction in the number of days of antibiotic therapy and length of hospitalisation. In one study of recipients of auto-BMT, GM-CSF therapy reduced both the time to reach an absolute neutrophil count (ANC) of 0.5 x 109/L and the incidence of bacterial infections (15 vs. 28 days and 36 vs. 63% respectively) compared to placebo. Similarly, in another controlled trial GM-CSF therapy was associated with a shorter median time to an ANC of 0.5 x 109/L (14 vs. 21 days) and a shorter duration of hospitalisation (23 vs. 28 days). However, the positive effects in these trials were limited to neutrophil engraftment and neutropenia-related complications. None of the studies have demonstrated enhanced platelet or red blood cell engraftment and none of them showed a survival advantage. The value of colony stimulating factors following auto-PBSCT is less impressive. The addition of G-CSF may further accelerate neutrophil engraftment, with 3–6 fewer days to achieve an ANC of 0.5 x 109/L. There have been no comparative studies of different colony stimulating factors (10). 3.1.2. Allogeneic transplantation In a randomised, controlled trial, 109 patients who underwent allo-BMT received either GM-CSF or placebo following the transplantation of SC. GM-CSF was associated
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with a significant shortening of the time to engraftment (ANC above 0.5 x 109/L 13 vs. 17 days) and a reduction in infectious complications. No differences could be detected in platelet or erythrocyte recovery (11). Another study randomised 54 patients undergoing allo-PBSCT to receive either G-CSF or placebo. The G-CSF arm was associated with significant acceleration of engraftment (median time to ANC above 0.5 x 109/L was 11 vs. 15 days p = 0.008). No differences were found in time to platelet engraftment, red blood cell transfusion independence, incidence of aGvHD or 100 day mortality (12). However the long term benefit of colony stimulating factor therapy following allo-HSCT remains controversial, as it may be associated with an increased risk of GvHD, especially cGvHD. In a retrospective study of 155 recipients sibling transplants at a single centre, patients treated with G-CSF after the transplant (43%) had a significantly higher incidence of grades II-IV acute GvHD (13). Moreover, a recent retrospective EBMT study of 1789 acute leukaemia patients concluded that G-CSF should probably not be used after HSCT from HLA identical sibling donors because it was associated with increased risks of GvHD and TRM (14). In the haplo-identical setting, post-transplant administration of G-CSF was found to impair functional immune recovery by promoting T-helper (Th)-2 immune responses which, unlike Th-1 responses, do not protect against intracellular pathogens and fungi. 3.2. Erythropoietin (EPO) EPO has been used to accelerate the recovery of red blood cells. This idea was supported by the observation that EPO levels after transplantation were lower than calculated for the degree of anaemia. However studies of its usage in this situation have not shown any major benefits. In allo-HSCT some studies have shown a reduction in the median time to transfusion independence but transfusion requirements were not different. There were however subsets of patients at high risk for transfusion in which EPO reduced transfusion needs. Because of the modest and mixed results of the studies, most centres do not use EPO in the early posttransplant setting. A few randomised trials have addressed the efficacy of EPO following autologous transplantation. None found a significant reduction of transfusion requirements. The lack of benefit is particularly true with PBSCT. The use of thrombopoietin for accelerating platelet recovery is still under investigation. 3.3. Conclusion Evidence exists in favour of the use of G-CSF (and GM-CSF) in some transplantation settings. Besides stem cell mobilisation, these include their use for accelerating engraftment following autologous stem cell transplantation (bone marrow and 172
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peripheral blood as well). The prophylactic use of G-CSF or GM-CSF following allogeneic stem cell transplantation is generally not recommended except for the cases of delayed neutrophil engraftment or secondary neutropenia due to infection or drug toxicities.
4. Oral mucositis, mouth care and pain relief Despite the benefits of TPN, mucositis per se remains an important clinical problem. Mucositis is usually managed by the application of topical and systemic pain medications. Many patients require narcotic pain medication and the length of time of intravenous narcotic need is one of the best indicators of the degree of mucositis. Routine mouth care typically consists of oral rinses performed every four hours with sodium bicarbonate or chlorhexidine. Patients should cease using a toothbrush (even a soft brush) while neutropenic and should not restart until the neutrophil count exceeds 1.0 x 109/L. Dentures are generally removed for the duration of mucositis because of discomfort. A variety of mucosal coating agents have been used with or without analgesics: diphenhydramine, corticosteroids, antacids, and sodium hyaluronate. Unfortunately, the evidence supporting benefit for any of these interventions is weak. Analgesia may necessitate systemic narcotics. In a study of 119 BMT recipients patient controlled analgesia with IV morphine, hydromorphone, or sufentanil resulted in nearly equivalent pain relief but side effects were lowest in the morphine group. Thus, morphine was recommended as the opioid of choice for patient controlled analgesia. Mucositis is associated with an increased risk of systemic infection resulting from bacteraemia associated with the breakdown of mucosal barriers. The severity of mucositis and the associated infection risk is a significant cause of morbidity and mortality, and more effective treatment modalities are therefore needed (15). 4.1. Keratinocyte growth factor (KGF, palifermin, Kepivance®) One new approach is the use of KGF. The efficacy of prophylactic intravenous palifermin was demonstrated in a double-blind multicentre trial of 212 patients undergoing HSCT for haematological malignancies. Significantly fewer patients receiving palifermin had grade 3 or 4 mucositis (63 vs. 98 percent with placebo) and the duration of mucositis was shorter (median 6 vs. 9 days). These benefits were associated with significantly less use of opioid analgesics and less frequent requirement for TPN support. On the basis of this study palifermin was approved for the reduction of mucositis following HSCT. However an unsettled issue is the effect of palifermin on GvHD; in an animal model it seemed to decrease the risk of GvHD but in a human clinical trial KGF did not show any effect on GvHD (16).
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4.2. Amifostine Amifostine is a phosphorylated aminothiol, which has a protective effect on normal tissues against radiation and alkylating agent toxicity. In a retrospective study of 68 patients receiving high dose melphalan prior to auto-HSCT those who received amifostine (35 pts) prior to conditioning had a significantly lower median number of days of analgesic therapy (0 vs. 6 days) and a lower incidence of severe mucositis (21 vs. 51%). There was no difference in haematological recovery between the two groups. The results of other attempts such as the application of TGF-b, IL-11, or laser therapy are even less mature. 4.3. Conclusion Several approaches have been tried for the prophylaxis or therapy of mucositis. However the problem has not yet been solved. A number of new agents promise clinical benefit, either singly or in combination therapy. Further studies are warranted.
5. Prevention and treatment of chemotherapy induced nausea and vomiting The objective of antiemetic treatment in the transplant setting is the complete prevention of nausea and vomiting caused by the conditioning regimen. For the classification of CT induced vomiting and the emetogenic potential of cytotoxic therapy (see Tables 5 and 6). Table 5: Classification of CT induced emesis Acute emesis
Delayed emesis
Anticipatory emesis
Occurs during the first 24 h following CT
Occurs later than 24 h
Conditioned response of patients who developed significant nausea and vomiting during previous CT
Table 6: Classification of emetogenic potential of CT Emetogenic potential High Moderate Low
Probability of vomiting (a) 60–99% 30–59% 1–29%
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The high dose CT/RT used in standard conditioning regimens (see Chapter 6) has itself high emetic potential, and there are some additional factors (consecutive day administration, prior cytotoxic treatment, and other medication e.g. opiate analgesics, which may cause emesis), which can increase the risk of vomiting. 5.1. Prophylaxis and treatment There are few randomised trials specifically studying the issue of nausea/vomiting in the transplant setting. There is a general consensus that a combination of a serotonin receptor antagonist (ondansetron, granisetron, tropisetron) and dexamethasone (8–20 mg IV) should be the standard prophylaxis on days of the conditioning. In case of failure of the prophylaxis adding of further dexamethasone (max 20 mg/day) and/or a benzodiazepine (e.g. lorazepam max 4 mg IV) may help to counter increased patient anxiety and possible anticipatory emesis. An alternative is to switch to a different serotonin antagonist, since there is an incomplete cross-resistance between agents (Figure 1).
Figure 1: Prevention and treatment of conditioning induced nausea/vomiting
On days of conditioning serotonin antagonist od-bd IV + dexamethasone 12 mg od IV
Does prophylactic regimen control emesis?
Yes
Continue
No
1. Rule out anticipatory emesis, consider benzodiazepine (e.g. lorazepam max 4 mg/d IV) 2. Add dexamethasone max 20 mg/d IV 3. Consider switch to another serotonin antagonist 4. Exclude other aetiologies
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Neurokinin–1 receptor antagonist aprepitant represents a new class of antiemetic drugs. When used in combination with serotonin antagonists and corticosteroids, aprepitant appears to provide better protection against both acute and delayed emesis in very highly emetic CT.
References 1. Weisdorf SA, Lysne J, Wind D, et al. Prophylactic total parenteral nutrition on long-term outcome of bone marrow transplantation. Transplantation 1987; 48: 833-838. 2. Szeluga DJ, Stuart RK, Brookmeyer R, et al. Nutritional support of bone marrow parenteral nutrition to en enteral feeding program. Cancer Res 1987; 47: 3309-3316. 3. Charuhas PM, Fosberg KL, Breummer B, et al. A double blind randomized trial comparing outpatient parenteral nutrition with intravenous hydration: Effect on resumption of oral intake after marrow transplantation. J Parenter Enteral Nutr 1997; 21: 157-161. 4. Iestra JA, Fibbe WE, Zwinderman AH, et al. Parenteral nutrition following intensive cytotoxic therapy: An exploratory study on the need for parenteral nutrition after various treatment approaches for haematological malignancies. Bone Marrow Transplant 1999; 23: 933-939. 5. Ulrich H, Pastores SM, Katz DP, et al. Parenteral use of medium-chain triglycerides: A reappraisal. Nutrition 1996; 12: 231-238. 6. Ziegler TR, Young LS, Benfell K, et al. Clinical and metabolic efficacy of glutamine supplemented parenteral nutrition after bone marrow transplantation. A randomized, double-blind, controlled study. Ann Intern Med 1992; 116: 821-828. 7. Brown SA, Goringe A, Fegan C, et al. Parenteral glutamine protects hepatic function during bone parrow transplnatation. Bone Marrow Transplant 1998; 22: 281-284. 8. Muscaritoli M, Grieco G, Capria S, et al. Nutritional and metabolic support in patients undergoing bone marrow transplantation. Am J Clin Nutr 2002; 75: 183-190. 9. O’Grady NP, Alexander M, Patchen Dellinger E, et al. Guidelines for the prevention of intravascular catheter-related infections. CID 2002; 35: 1281-1307. 10.Dekker A, Bulley S, Beyene J, et al. Meta-analysis of randomized controlled trials of prophylactic granulocyte colony-stimulating factor and granulocyte-macrophage colonystimulating factor after autologous and allogeneic stem cell transplantation. J Clin Oncol 2006; 24: 5207-5215. 11.Nemunaitis J, Rosenfeld CS, Ash R, et al. Phase III randomized, double-bind placebo controlled trial of rhGM-CSF following allogeneic bone marrow transplantation. Bone Marrow Transplant 1995; 15: 949-954. 12.Bishop MR, Tarantolo SR, Geller RB, et al. A randomized double-blind trial of filgastrim (granulocyte colony-stimulating factor) versus placebo following allogeneic blood stem cell transplantation. Blood 2000; 96: 80-85. 13.Remberger M, Naseh N, Aschan J, et al. G-CSF given after haematopoietic stem cell transplantation using HLA identical sibling donors is associated to higher incidence of acute GvHD II-IV. Bone Marrow Transplant 2003; 32: 217-223. 14.Ringden O, Labopin M, Gorin NC, et al. Treatment with granulocyte colony-stimulating 176
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factor after allogeneic bone marrow transplantation for acute leukemia increases the risk of graft-versus-host disease and death: A study from the acute leukemia working party of the European Group for Blood and Marrow Transplantation. J Clin Oncol 2004; 22: 416423. 15.Fanning SR, Rybicki L, Kalaycio M, et al. Severe mucositis is associated with reduced survival after autologous stem cell transplantation for lymphoid malignancies. Br J Haematol 2006; 135: 374-381. 16.Blazar BR, Weisdorf DJ, Defor T, et al. Phase 1/2 randomized, placebo-control trial of palifermin to prevent graft-versus-host disease (GvHD) after allogeneic hematopoietic stem cell transplantation (HSCT). Blood 2006; 108: 3216-3222.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which of the following is the most accurate for assessing nutritional status in HSCT recipients? a) Serum cholesterol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Serum glucose. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Nitrogen balance. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Anthropometric measurements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. What is the most common complication of TPN? a) Elevated liver enzymes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Hypertriglyceridaemia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Elevated serum creatinine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Catheter-related complications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which of the following is not considered a standard indication for the use of G-SF? a) Stem cell mobilisation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Accelerate engraftment following allo-HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Accelerate engraftment following auto-HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Treat secondary neutropenia following transplant due to drug toxicity . . .
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4. Which statement is not true regarding the CVC? a) Health-care worker education and training for the insertion and maintenance of the CVC is essential . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) In adults, a subclavian site is preferred . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Catheters that are no longer essential should be removed promptly . . . . . . d) Polyvinyl chloride catheters are more resistant to adhesion of micro-organisms than teflon, silicone, or polyurethane catheters . . . . . . . . 5. Which statement is not true concerning oral mucositis? a) Palifermin is especially indicated in allo-HSCT as it decreases the risk of GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Oral mucositis per se is a significant cause of morbidity and mortality of HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Patients should stop using a toothbrush while neutropenic . . . . . . . . . . . . . . . d) Routine mouth care typically consists of oral rinses performed every four hours with chlorhexidine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 9
Early complications after HSCT
E. Carreras
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CHAPTER 9 • Early complications after HSCT
1. Introduction The high dose of RT and/or CT included in conditioning regimens (see Chapter 6) affects all organs and tissues of the recipient, producing several early and late secondary effects of variable intensity. The most common early effects like nausea, vomiting, mucositis and pain are treated in Chapter 8, and late effects are covered in Chapter 12. In this Chapter we summarise some early complications that, despite not being very frequent, are an important cause of morbidity and mortality.
2. Haemorrhagic cystitis (HC) (1) Pathogenesis HC is produced by direct toxicity of the conditioning regimen on the urothelium or by viral infections affecting the urinary tract. Usually, HC secondary to conditioning appears early after HSCT (several days after receiving CT agents) and can be produced by Cy (or ifosfamide), Bu (especially if combined with Cy), VP or TBI (both uncommon). Viral HC appears later (usually after day +30) and can be due to human polyomavirus type BK or JC, adenovirus type 11 (less frequent) or CMV (exceptional). Incidence HC secondary to CT: 1 to 25%, depending on the preventive measures adopted. The incidence of viral HC is not well established; 5 to 25%, according to the degree of immunosuppression of the recipient. Prophylaxis Continuous irrigation of the bladder during conditioning is effective but is no longer used. Nowadays, HC prophylaxis is based on hyperhydration and Mesna administration. The recommended daily dose of hydration is 3 L/m2. The usual daily dose of Mesna is 1.0–1.5 x daily dose of Cy (e.g. daily dose of Cy: 4.2 g Æ daily dose of Mesna: 4.2–6.3 g) administered IV as: a) continuous infusion in 1 L of 0.9% saline over 12–24 hrs, beginning 4 hours prior to the 1st dose of Cy and ending 12–24 hrs after the last Cy dose; or b) bolus injections, 20% of daily dose of Cy dose administered as a bolus 1/2–1 h before Cy and the remaining daily dose divided into bolus injections q 2–3 h, maintained up to 12–24 after ending Cy. Despite these classical forms of administration, some pharmacodynamic studies have shown that the best method of administering Mesna is to combine a continuous infusion with intermittent bolus injections during the 6 hrs after Cy dose. The presence of acrolein in the bladder 24 h after Cy makes it advisable to prolong Mesna administration 24 hrs after the last Cy dose. Treatment Treatment should be based on a three step approach: 1. Forced hydration plus intensive platelet support. The use of procoagulant agents
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like aminocaproic acid is contraindicated because they favour clot formation in the bladder. 2. Continuous bladder irrigation with saline solution. Some success has been reported with bladder instillation of alumina 1% (risk of encephalopathy if associated renal failure), prostaglandin E2 or E1, GM-CSF or cidofovir (all with limited experience). Similarly, hyperbaric oxygen or oestrogens (2–4 mg q 8 h p.o., Odermann et al., BMT 2000) have also been reported as effective measures. 3. If the previous measures do not solve HC other salvage approaches can be considered: selective embolisation of bladder arteries (one of the simplest and effective measure in the hands of an expert angioradiologist) (2); suprapubic cystotomy; cystoscopy + installation of formalin (very painful, risk of scars and bladder contraction, requires anaesthesia); catheterisation of both urethers to rest the bladder; hypogastric bond (can produce sexual impotence); and, as a last resort, cystectomy.
3. Early complications of vascular origin Injury of the vascular endothelium seems to be the most important initial event in a variety of complications with imprecise diagnostic criteria and overlapping clinical features, which are observed within the first 30–60 days after HSCT. The best defined syndromes resulting from this endothelial injury are: 1) Veno-occlusive disease of the liver; 2) Capillary leak syndrome; 3) Engraftment syndrome; 4) Diffuse alveolar haemorrhage; 5) Thrombotic microangiopathy; 6) Idiopathic pneumonia syndrome; 7) Multiple-organ dysfunction syndrome. Figure 1 shows their common pathogenesis. 3.1. Hepatic veno-occlusive disease (VOD) Definition VOD is the term used to designate the symptoms and signs that appear early after HSCT as a consequence of the conditioning regimen-related hepatic toxicity. This syndrome is characterised by jaundice, fluid retention and tender hepatomegaly appearing in the first 35–40 days after HSCT (3–8). Pathogenesis (3, 6–8) The hepatic metabolism of certain drugs (e.g. Cy) by the cytochrome P-450 enzymatic system produces several toxic metabolites (e.g. acrolein). These toxic metabolites are converted to stable (non-toxic) metabolites by the glutathione enzymatic system (GSH) and eliminated. When this process occurs in patients with a reduced GSH activity, due either to pre-existing liver disease or to the action of agents as Bu, BCNU or TBI that reduce GSH levels, toxic metabolites are not 182
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Figure 1: Common pathogenesis of early complications of vascular origin after HSCT
VEGF: vascular endothelial growth factor; Tx: thromboxane; PG: prostaglandin; TM: thrombomodulin; PAI-1: plasminogen activator inhibitor type 1
metabolised. Toxic metabolites are predominantly located in area 3 of the acinus (around centrilobular veins) because this area is rich in P-450 and poor in glutathione. Consequently, damage to hepatocytes and sinusoidal endothelium occurs predominantly in this anatomic zone. The remaining factors mentioned in Figure 1 can also contribute to the endothelial injury. Experimental models show that the first events after endothelial injury by toxic metabolites are loss of sinusoidal endothelial cell (SEC) fenestrae, formation of gaps within and between SEC, and rounding up or swelling of SEC. Consequently, red blood cells penetrate into space of Disse and dissect off the sinusoidal lining, which embolises downstream and blocks the sinusoids reducing the hepatic venous outflow and producing post-sinusoidal hypertension. In the light of all these observations some authors have proposed the term of sinusoidal obstruction syndrome (SOS) for this complication. Clinical features (3, 4, 6, 8) Classical VOD. Occurs within days after conditioning (from day -1 to +14) and is characterised by presence of: jaundice (in almost 100% of cases), hepatomegaly and/or
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right upper quadrant pain, and weight gain (not attributable to excessive fluid administration) with oedemas and ascites. Late VOD. The same clinical manifestations as classical VOD but developing late after HSCT (one third of cases occur after patient discharge). Mainly observed after conditioning including several alkylating agents in combination (busulfan, melphalan, thiotepa). One third have a biphasic course with an initial transitory peak followed by a definitive late phase (5) VOD with multiorgan failure. The same clinical manifestations as previously described plus: thrombocytopenia (refractoriness to platelet transfusions); pleural effusion / pulmonary infiltrates; progressive renal, cardiac, pulmonary failure; confusion, encephalopathy and coma. Incidence The incidence of VOD ranges from 3 to 54% in the largest series. This variability is the consequence of the presence or absence of a number of well-known risk factors for this complication (see Table 1). In the only prospective multicentre study published the incidence of VOD was 8% in allo-HSCT and 3% in auto-HSCT (6). Diagnostic criteria As for any syndrome the diagnosis of VOD must be established clinically. All HSCT teams use one of the following sets of clinical criteria (3, 4, 6 ,8): Seattle criteria: In the first 20 days after HSCT presence of two or more of the following: bilirubin > 2 mg/dL; hepatomegaly or pain in the right-upper quadrant; weight gain (>2% basal weight). Baltimore criteria: In first 21 days after HSCT, presence of bilirubin >2 mg/dL plus ≥2 of the following: painful hepatomegaly; ascites; weight gain (>5% basal weight). In both, other possible causes of these clinical features should be excluded before accepting the diagnosis of VOD (see differential diagnosis). Additionally, it is necessary to remember that some VOD cases can appear late after HSCT. Additional investigations Other complementary studies that can aid diagnosis are: Haemodynamic study of the liver carried out through the jugular or femoral veins (9): Despite its usefulness, this is only indicated to confirm the diagnosis of VOD before adopting a therapeutic approach that may be potentially hazardous for the patient. An hepatic venous gradient pressure (HVGP) ≥10 mmHg in a patient without previous liver disease allows a precise differential diagnosis with a high degree of specificity. However, a normal HVGP does not exclude the diagnosis of VOD. Liver biopsy: Thrombocytopenia usually present in this phase of HSCT precludes a transparietal liver biopsy; consequently hepatic tissue can only be obtained by means of a transvenous biopsy in the course of a haemodynamic study. In addition to the 184
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Table 1: Risk factors (3, 4, 6–8) Risk
Lower risk < Higher risk
Transplant type Donor type HLA compatibility Stem cell origin T-cell depletion Diagnosis Status of the disease
Syngeneic or autologous < allogeneic Sibling < another relative < unrelated HLA match < any mismatch Peripheral blood < bone marrow With TCD < without TCD non malignant disease < malignant disease Remission < relapse
Conditioning - Intensity - TBI
- Busulfan - Timing Age / Sex Karnofsky index ASAT/ALAT before HSCT Transplant number Previous hepatic irradiation Previous Mylotarg Status of the liver CMV serological status Fever in conditioning Hepatotoxic drugs Genetic predisposition
Cy alone < Cy + TBI < BVC (a) Fractionated TBI < single dose TBI Less than 12 Gy < more than 12 Gy Low dose rate < high dose rate IV BU < adjusted oral Bu < non adjusted oral Bu Interval Cy – TBI 36 hours < 12 hours Younger < older / men < women 100–90 < lower than 90 Normal < high First < second No < yes No < yes (b) Normal < fibrosis < cirrhosis or infiltration Negative < positive Absent < present Progestogens, ketoconazole, CsA, methotrexate, amphotericin B, vancomycin, acyclovir, IV Ig (c) GSTM1 positive < GSTM1 null genotype (d)
The most important risk factors are indicated in bold type. (a) BVC (BCNU, VP, Cy). (b) VOD incidence up to 64% (Wadleigh et al. Blood 2003). (c) Higher incidence of VOD with high-dose IVIg. (d) Srivastava et al., Blood 2004
classical histological changes of VOD (concentric non-thrombotic narrowing of the lumen of small intrahepatic veins) other less specific abnormalities can be observed in patients with a VOD syndrome, including eccentric narrowing of the venular lumen; phlebosclerosis; sinusoidal fibrosis and hepatocyte necrosis. Due to patchy nature of VOD a normal biopsy does not exclude the diagnosis. Ultrasound: A variety of abnormalities can be observed; gallbladder wall thickening, ascites, hepatomegaly and attenuate or reversed portal flow, but all of them are nonspecific. Biological markers: Although the serum of patients with VOD shows an increase in HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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levels of plasminogen activator inhibitor-1 (PAI-1) (marker with the highest specificity and sensitivity for VOD), aminopropeptides of type III collagen, and hyaluronic acid, all these measurements are of little utility in routine clinical practice. Differential diagnosis To accept the diagnosis of VOD all the following possible causes of similar clinical features should be excluded as far as possible, including: • Infections: Cholangitis lenta (sepsis of liver) / fungal infection / viral hepatitis • Immune dysfunctions: Acute GvHD of the liver • Drug toxicity: CsA, azoles, MTX, progestogens, trimethoprim-sulphamethoxazole, TPN, among others • Reduction of venous outflow / increased volume: Constrictive pericarditis / congestive heart failure / fluid overload / renal failure • Others: Pancreatic ascites / chylous ascites / infiltration of the liver. Prophylaxis of VOD (6, 8) (Table 2) Table 2: Prophylaxis of VOD Avoidance of risk factors • When possible delay HSCT if an acute hepatitis exists; adjust Bu dose or use IV Bu; fractionate TBI; avoid hepatotoxic drugs, etc. • In high risk patients, consider allo-RIC HSCT (lower incidence of VOD) Pharmacological The following drugs have been used to prevent VOD from the beginning of conditioning until day +21–30: • Sodium heparin: 100 U/kg/day by continuous infusion. Two randomised studies showed a beneficial effect but others have suggested that it is ineffective and dangerous • Prostaglandin E1: 0.3 µg/kg/h by continuous infusion. Evaluated in several clinical trials usually combined with heparin. When administered alone no beneficial effect was observed • Ursodeoxycholic acid: 600–900 mg/day p.o. Four randomised trials and 2 historically controlled studies have shown a reduction in incidence of VOD and in TRM • N-acetylcysteine. Very limited experience • Low molecular weight heparin: Enoxaparin 40 mg/day or fraxiparin 5000 U/day subcutaneously seem to be relatively safe and may have some effect but a large randomised study is needed to confirm these results • Pre-emptive ATIII replacement: Ineffective • Defibrotide. Several preliminary reports have shown encouraging results
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Treatment of established VOD (6, 8) (Table 3) Table 3: Treatment of established VOD First line therapy Symptomatic (a)
-
Specific (b)
-
Restriction of salt and water intake ± diuretics Maintain intravascular volume and renal perfusion by means of albumin, plasma expanders and transfusions (haematocrit >30%) Defibrotide 6.25 mg/kg IV in 2 h infusion q 6 h IV during 14 days (1) (c) (d) rt-PA 0.05 mg/kg/h during 4 hours (maximum 10 mg/day) for 2–4 days ± sodium heparin 20 U/kg as a bolus (maximum 1000 U) followed by 150 U/kg/day by continuous infusion for 10 days (e)
Other measures Symptomatic (a)
-
Specific
-
Low dose dopamine (effectiveness not demonstrated) Analgesia Paracentesis / thoracocentesis Haemodialysis / haemofiltration Mechanical ventilation TIPS (transvenous intrahepatic portosystemic shunt) (f) Surgical shunt Liver transplantation
rt-PA: recombinant tissue plasminogen activator. (a) Symptomatic treatment should be established first, reserving specifies measures for most severe cases. (b) Although other agents have been used (antithrombin III, prostaglandin, corticosteroids, glutamine/vitamin E, N-acetylcysteine, etc.) the only ones occasionally effective are those mentioned. (c) Defibrotide permits the resolution of 50–55% of severe VOD with multiorgan dysfunction and a 47–60% of survival at day +100 with no secondary effects in adults and children (8, 10). (d) In a randomised study defibrotide at 25 mg/kg/day has shown similar effectiveness to the classical dose of 40 mg/kg /d (8). (e) rt-PA has been shown to be effective only in patients with a non-advanced VOD. Its use is contraindicated in patients with multi-organ dysfunction syndrome (MODS), haemorrhages or severe hypertension. (f) Despite improvement in portal hypertension and ascites, long term efficacy and survival are extremely poor
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VOD evolution (3, 4, 6 ,7) (Table 4) Table 4: VOD evolution Complete resolution on day +100 w/o treatment Complete resolution on day +100 with treatment Non resolution before death (c) or on day +100 Mortality attributable to VOD by day +100 (e)
Classification (a) Frequency (b) Mild VOD 8–23% Moderate VOD 48–64% Severe VOD (d) 23–28% 1–3% of all HSCT 18–28% of all VOD 75–95% of severe VOD (f)
(a) Classification described by Seattle group for retrospective evaluation of VOD. (b) Values observed in two large series (3, 6); (c) In many cases VOD is not the direct cause of death but contributes to it. (d) The severity of VOD can be predicted by means of a mathematical model (Bearman et al., JCO 1993). (e) Data from pre-defibrotide era. (f) The equivalent predicted mortality in non-severe VOD cases ranges between 10 and 20%
3.2. Capillary leak syndrome (CLS) (11) Pathogenesis The injury to the capillary endothelium produces a loss of intravascular fluids into interstitial spaces and the clinical manifestations. Incidence The absence of well-established clinical criteria for its diagnosis precludes an accurate estimation of its incidence. Additionally, the differential diagnosis with VOD, ES or IPS can be very difficult. Clinical features Development, in the first 15 days after HSCT, of: • Weight gain (>3% in 24 hours), and • Generalised oedemas (ascites, pleural effusion, pericarditis) that characteristically does not respond to frusemide treatment. Other features occasionally observed are: tachycardia, hypotension, renal insufficiency of pre-renal origin and hypoalbuminaemia. Differential diagnosis From engraftment syndrome (ES): Its earlier development, the absence of skin rash and the poor response to corticosteroids. From VOD: The absence of jaundice and painful hepatomegaly, and the poor response to furosemide. From IPS: The presence of generalised oedema. Risk factors The use of G-CSG, GM-CSF or K-CSF; high cumulative dose of CT in the pre-HSCT phase; 188
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unrelated or HLA mismatched donor grafts. Treatment To withdraw growth factors. Despite being systematically used the response to corticosteroids is poor. There is no other specific treatment. Evolution There is a high mortality if it progresses to MODS. 3.3. Engraftment syndrome (ES) (12–14) Pathogenesis Massive release of pro-inflammatory cytokines by tissues injured by intensive conditioning and by recovering neutrophils has been hypothesised to play a role. Incidence Variable depending on the diagnostic criteria used. After auto-HSCT: from 5 to up to 25% in patients with breast cancer or autoimmune diseases. After conventional allo-HSCT: only occasionally described (possibly because of the difficult differential diagnosis from GvHD). After allo-RIC: 10% in a recent series (14). Clinical features Development, within 72 hours of the start of neutrophil recovery, of the following major clinical criteria: • High fever of a non-infectious origin (unresponsive to antibiotics and negative cultures); • Skin rash affecting >25% body surface and not attributable to an allergic reaction; • Lung infiltrates or hypoxia not attributable to fluid overload, lung embolism, or congestive heart failure. Other symptoms occasionally observed are diarrhoea, weight gain and liver, kidney or CNS dysfunction (minor criteria). Diagnosis There are no well-established criteria for its diagnosis. Spitzer (12): 3 major criteria or 2 major and one or more minor criteria; Majolino (13): fever with either skin rash, pulmonary infiltrates or diarrhoea; Gorack (14): ≥2 major criteria plus weight gain. Risk factors Most cases of ES have been described since the introduction of growth factors and use of PBSCT. For this reason, a high number of CD34+ cells, faster engraftment and use of growth factors (especially GM-CSF) are considered to be the main risk factors as well as the underlying disease (breast cancer, multiple sclerosis, POEMS syndrome). Treatment MethylPDN 1 mg/kg q 12 h (3 days) with progressive tapering over one week. An appropriate empiric antibiotic treatment should always be maintained due to the difficulty of excluding an infectious origin of the fever.
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Evolution Complete resolution in 1–5 days in >80% of cases if steroids are introduced early. 3.4. Diffuse alveolar haemorrhage (DAH) (15, 16) Pathogenesis Very similar to VOD pathogenesis but affecting the lungs. Incidence The reported incidence ranges from 1 to 5% in auto-HSCT and from 3 to 7% in alloHSCT. Some authors consider that underlying undetected infections can play a role in DAH pathogenesis and postulate that infection-associated alveolar haemorrhage and DAH should be considered as equivalents. Clinical features Despite some cases of late-onset DAH, it is usually diagnosed within the first 30 days after HSCT. The main manifestations are: • Dyspnoea, non productive cough, tachypnoea • Hypoxaemia that can require oxygen-therapy • Chest X-ray or CT with focal or diffuse interstitial or alveolar infiltrates located in middle and inferior lung fields • Bronchoalveolar lavage (BAL) progressively bloodier, and not attributable to infection (absence of pathogens in BAL), thrombocytopenia, fluid overload or heart failure. Successive aliquot of 20 mL, in at least three segmentary bronchi, become pro-gressively more bloodstained (indicating blood in the alveoli). Risk factors DAH is not related to low platelet counts. Factors that favour this complication are older age, previous thoracic radiation, allogeneic donor, myeloablative conditioning, and severe acute GvHD. Treatment After publication of some small retrospective series high-dose methylPDN (250–500 mg q 6 h, 4–5 days and tapering in 2–4 weeks) was considered the treatment of choice. However, many other authors have not observed that corticosteroids modify the poor outcome associated with DAH. Recombinant FVIIa has been used with success in some cases. The possible role of cytokine antagonists and antiinflammatory agents should be evaluated. Evolution The overall mortality rate at 60 days from the onset of the haemorrhage is around 75% despite in many patients the death is not directly related to the haemorrhage. 3.5. Thrombotic microangiopathy (TMA) (17–20) TMA is the term used to describe haemolytic uraemic syndrome (HUS) and thrombotic 190
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thrombocytopenic purpura (TTP) associated with HSCT. Pathogenesis Conditioning regimen related toxicity, together with other triggering factors that are not clearly understood, produces a generalised endothelial dysfunction with intravascular platelet activation and formation of platelet-rich thrombi within the microcirculation. In contrast to classical TTP, ADAMTS13 activity very rarely falls below 10%. Incidence Less than 4% in auto-HSCT. Up to 15% in allo-HSCT (7% in an EBMT survey). Clinical manifestations Usually develop around day +60 but early (day +4) and late (2 years) episodes have been described. Characterised by: • Microangiopathic haemolytic anaemia (MHA) (anaemia, >2–5% schistocytes, LDH and other markers of haemolysis) • Thrombocytopenia or increase in transfusional requirement • Fever of non-infectious origin • Renal dysfunction and/or neurological abnormalities (cortical blindness, seizures, typical images in CNS CT-scan). Diagnostic criteria for HSCT-associated TMA (Table 5) Table 5: Diagnostic criteria for HSCT-associated TMA Blood & Marrow Transplant Clinical Trials Network consensus (18) 1) RBC fragmentation and 2 schistocytes per high-power field on PB smear 2) Concurrent increased serum LDH 3) Concurrent renal (a) and/or neurologic dysfunction w/o other explanations 4) Negative direct and indirect Coombs test International Working Group (19) 1) Increased percentage (>4%) of schistocytes in the blood 2) De novo, prolonged or progressive thrombocytopenia 3) Sudden and persistent increase in LDH 4) Decrease in Hb concentration or increased RBC transfusion requirement 5) Decrease in serum haptoglobin concentration (a) Doubling of serum creatinine from baseline
Risk factors A higher incidence has been observed in patients receiving TBI, calcineurin inhibitors (CNI), sirolimus, urelated or HLA-mismatched donor grafts, or developing GvHD or CMV/fungal infections.
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Clinical forms Two main forms of TMA can be observed: 1. CNI-associated nephrotoxicity (or neurotoxicity) with MHA: Classically develops early after HSCT, is related to toxic levels of CNI, and is reversible after stopping its administration. Usually has a favourable evolution if it improves quickly after stopping CNI. 2. Not associated with CNI toxicity, with two clinical forms: a. Conditioning associated HUS: TMA primarily affecting the kidney, often causing oliguric or anuric renal failure with hypertension, MHA and thrombocytopenia, and b. Fulminating multifactorial TMA: Early after HSCT, renal failure, CNS disturbances, hypertension, MHA and thrombocytopenia associated with GvHD, viral or fungal infection. Most cases have a fatal evolution and do not respond to CNI suppression, plasma exchange or other treatments (see below). Prevention The only reasonable measure is to have a close control (2–3 times per week) of CNI, LDH and creatinine levels. If any of them increase peripheral blood smear, haptoglobin and CNI metabolites should be tested. Treatment The only effective measure in some cases is to inmediately stop CNI, adding another agent for GvHD prophylaxis/treatment (corticosteroids, mycophenolate, azathioprine). Plasma exchange cannot be currently considered the standard of care despite some success (less than 50% of responses and 70–90% of mortality in published series possibly due to selection bias). Some authors have reported successful results with anti-TNF MoAb (etanercept/infliximab), defibrotide, daclizumab, rituximab, and eicosapentaenoic acid. 3.6. Idiopathic pneumonia syndrome (IPS) (21) Pathogenesis The formerly used term of interstitial pneumonia has been progressively abandoned because it does not correspond to the real pathologic findings. Apparently, this syndrome is the result of a diversity of lung insults, including the toxic effects of conditioning, immunologic cell-mediated injury, inflammatory cytokines and, probably, occult pulmonary infections. Incidence As a consequence of the improvement of diagnostic methods the incidence of IPS has reduced from more than 20% in earlier series of allo-HSCT to less than 10% at present time (8.4 and 2.2% after conventional and allo-RIC, respectively, in a recent series). It is uncommon in auto-HSCT setting. 192
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Clinical features Development around day +21 of: • Fever, non-productive cough • Tachypnoea, hypoxaemia • Diffuse alveolar or interstitial infiltrates on X-ray or scan. Diagnosis The diagnosis is confirmed when the previous clinical manifestations are associated with: • Absence of infectious pathology or DAH in FBS, BAL or lung biopsy, and • Absence of other possible pathogens (lung oedema, lung haemorrhage, fat embolism, leukaemic infiltration, or lung toxicity due to leucoagglutinins). Risk factors Myeloablative conditioning, age older than 40, and grade III–IV acute GvHD. With myeloablative HSCT, the use of TBI, especially in patients older than 40 years is an additional risk factor. Treatment Supportive care combined with prophylaxis and treatment of infections. Some patients improve with methylPDN and some successes have been described with antiTNF MoAb (etanercept/infliximab). Evolution Up to 50–70% of patients will die due to a progressive impairment of respiratory function. This percentage reaches 97% if mechanical ventilation is required. 3.7. Multiple-organ dysfunction syndrome (MODS) (22) Pathogenesis All mechanisms previously mentioned. Incidence Unknown, due to the difficulty in differentiating this from the syndromes already described. Clinical features This diagnosis should be considered when early after HSCT a patient presents two or more of the following: • CNS dysfunction (>4 points on the scale of Folstein) • Lung dysfunction (O2sat < 90% in two occasions separated by more than 2 hr in the same day) • Renal dysfunction (creatinine >1.5 mg/dL [>133 mmol/L]) • Hepatic dysfunction (VOD criteria, see 3.1.). Treatment There is no effective treatment and this syndrome is irreversible.
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References 1. Sencer SF, Haake RJ, Weisdorf DJ. Hemorrhagic cystitis after bone marrow transplantation. Risk factors and complications. Transplantation 1993; 56: 875-879. 2. Gine E, Rovira M, Real I, et al. Successful treatment of severe hemorrhagic cystitis after hemopoietic celltransplantation by selective embolization of the vesical arteries. Bone Marrow Transplant 2003; 31: 923-925. 3. McDonald GB, Hinds MS, Fisher LD et al. Veno-occlusive disease of the liver and multiorgan failure after bone marrow transplantation: A cohort study of 355 patients. Ann Intern Med 1993; 118: 255-267. 4. Carreras E, Bertz H, Arcese W, et al. Incidence and outcome of hepatic veno-occlusive disease (VOD) after blood or marrow transplantation (BMT): A prospective cohort study of the European group for Blood and Marrow Transplantation (EBMT). Blood 1998; 92: 3599-3604. 5. Lee JL, Gooley T, Bensinger W, et al. Veno-occlusive disease of the liver after busulfan, melphalan, and thiotepa conditioning therapy: Incidence, risk factors, and outcome. Biol Blood & Marrow Transplant 1999; 5: 306-315. 6. Carreras E. Veno-occlusive disease of the liver after hematopoietic cell transplantation. Eu J Haematol 2000; 64: 281-291. 7. DeLeve LD, Shulman HM, McDonald GB. Toxic injury to hepatic sinusoids: Sinusoidal obstruction syndrome (veno-occlusive disease). Semin Liver Dis 2002; 22: 27-42. 8. Ho VT, Linden E, Revta C, Richardson PG. Hepatic veno-occlusive disease after hematopoietic stem cell transplantation: Review and update on the use of defibrotide. Semin Thromb Hemost 2007; 33: 373-388. 9. Carreras E, Gra_ena A, Navasa M, et al. Transjugular liver biopsy in BMT. Bone Marrow Transplant 1993; 11: 21-26. 10.Richardson PG, Murakami C, Jin Z et al. Multi-institutional use of defibrotide in 88 patients after stem cell transplantation with severe veno-occlusive disease and multisystem organ failure: response without significant toxicity in a high-risk population and factors predictive of outcome. Blood 2002; 100: 4337-4343. 11.Nürnberger W, Willers R, Burdach S, Göbel U. Risk factors for capillary leak-age syndrome after bone marrow transplantation. Ann Hematol 1997; 74: 221-224. 12.Speizer TR. Engraftment syndrome following hematopoietic stem cell transplantation. Bone Marrow Transplant 2001; 27: 893-898. 13.Maiolino A, Biasoli I, Lima J, et al. Engraftment syndrome following autologous hematopoietic stem cell transplantation: Definition of diagnostic criteria. Bone Marrow Transplant 2003; 31: 393-397. 14.Gorak E, Geller N, Srinivasan R, et al. Engraftment syndrome after nonmyeloablative allogeneic hematopoietic stem cell transplantation: Incidence and effects on survival. Biol Blood Marrow Transplant 2005; 11: 542-550. 15.Afessa B, Tefferi A, Litzow MR et al. Diffuse alveolar hemorrhage in hematopoietic stem cell transplant recipients. Am J Respir Crit Care Med 2001; 166: 641-645. 16.Majhail NS, Parks K, Defor TE, Weisdorf DJ. Diffuse alveolar hemorrhage and infectionassociated alveolar hemorrhage following hematopoietic stem cell transplantation: 194
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Related and high-risk clinical syndromes. Biol Blood Marrow Transplant 2006;12: 10381046. 17.Daly AS, Xenocostas A, Lipton JH. Transplantation-associated thrombotic microangiopathy: Twenty-two years later. Bone Marrow Transplant 2002; 30: 709-715. 18.Ho VT, Cutler C, Carter S, et al. Blood and marrow transplant clinical trials network toxicity committee consensus summary: Thrombotic microangiopathy after hematopoietic stem cell transplantation. Biol Blood Marrow Transplant 2005;11: 571-575. 19.Ruutu T, Barosi G, Benjamin RJ, et al. European Group for Blood and Marrow Transplantation; European LeukemiaNet. Diagnostic criteria for hematopoietic stem cell transplantassociated microangiopathy: Results of a consensus process by an International Working Group. Haematologica. 2007; 92: 95-100. 20.Batts ED, Lazarus HM. Diagnosis and treatment of transplantation-associated thrombotic microangiopathy: Real progress or are we still waiting? Bone Marrow Transplant 2007 Jul 2; [Epub ahead of print]. 21.Fukuda T, Hackman RC, Guthrie KA, et al. Risks and outcomes of idiopathic pneumonia syndrome after nonmyeloablative and conventional conditioning regimens for allogeneic hematopoietic stem cell transplantation. Blood 2003; 102: 2777-2785. 22.Gordon B, Lyden E, Lynch J, et al. Central nervous system dysfunction as the first manifestation of multiorgan dysunction syndrome in stem cell transplant patients. Bone Marrow Transplant 2000; 25: 78-83.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Late-onset haemorrhagic cystitis usually is produced by: a) The direct action of cyclophosphamide on the bladder. . . . . . . . . . . . . . . . . . . . . b) The sum of several toxic factors that produce a bladder damage . . . . . . . . . . c) A polyomavirus infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) A bacterial infection of the urinary tract . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) The neutropenia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which of the following complications could not be attributed to an endothelial dysfunction? a) Engraftment syndrome. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Veno-occlusive disease of the liver . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Haemorrhagic cystitis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Leak capillary syndrome . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) Thrombotic microangiopathy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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3. Which of the following is not a clinical manifestation of VOD? a) Weight gain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Painful hepatomegaly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Ascites . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Platelet refractoriness . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) Diarrhoea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. All but one of the following are classical manifestations of engraftment syndrome, which one? a) Skin rash . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Back pain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Fever . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Hypoxaemia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) Diarrhoea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which is the main cause of thrombotic microangiopathy after HSCT? a) Bacterial infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Graft allo-reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Immunological phenomena . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Cyclosporin toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) Renal failure. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 10
Infections after HSCT
C. Cordonnier
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1. Introduction Despite considerable progress in the management of the complications of HSCT, infection remains an important cause of post-transplant morbidity and mortality, mainly after allogeneic HSCT. The major advances in the management of infectious complications have come from better understanding of the mechanisms of the complex depression of immunity observed during the first months after transplant and their role in the predisposition to given infections, and also from well-designed therapeutic trials. Although the proportion of infectious deaths after allogeneic HSCT has decreased over the last two decades (1), much remains to be done to further decrease this risk and implement more efficient preventive and prophylactic strategies adapted to this high-risk population. Even though the risk of infectious deaths is much lower after autologous transplant, the risks of the procedure are greater than those of conventional chemotherapy, and preventive policies should be implemented in any transplant program.
2. The timing of immune reconstitution determines the timing of infections Stem cell transplantation offers a unique model of gradual immune reconstitution, and illustrates perfectly the relationship between the type of immune deficiency and the occurrence of infection due to special pathogens. Immune reconstitution after HSCT is the topic of another Chapter of this book, and will not be detailed here. However, some basic observations deserve to be emphasised. After allogeneic HSCT following conventional (i.e. myeloablative) conditioning regimens, the sequence of infections can be divided into three periods: 1. The first is the aplastic phase following the conditioning regimen until neutrophil recovery from the donated marrow. During this phase, the infectious complications of HSCT patients are not very different from those encountered in other profoundly neutropenic patients such as acute leukaemia patients, except, in most cases, for more severe mucosal damage, especially after total body irradiation. This is also the beginning of the at-risk period for fungal infections, mainly aspergillosis. Viral infections, especially HSV, are also common. Infection-related mortality at this time is mainly due to severe bacterial sepsis, pneumonia, and fungal infections. 2. The second phase corresponds to the period from initial marrow engraftment to at least the third or fourth month, and is characterised by cell-mediated immune deficiency with decreased number and function of specific and non-specific
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cytotoxic cells. For many years CMV infection, which is mainly due to reactivation, was the greatest problem during this phase. However, the routine practice of early diagnosis through PCR or antigenaemia and prophylactic or pre-emptive therapy have both been shown to decrease mortality due to CMV disease. Other viral diseases, less frequent than CMV, have also been described during this phase, especially those due to adenovirus and enteric and respiratory viruses. The occurrence and severity of GvHD is the main factor delaying immune recovery and favouring infections. 3. The third phase, beginning at the fourth month, is considered to be the late posttransplantation period. Here again, immune reconstitution is mainly influenced by the presence and severity of chronic GvHD. Most patients have immunoglobulin deficiency, particularly of IgG2, which is responsible for a decrease in the response to polysaccharide antigens. Allogeneic HSCT recipients are particularly vulnerable to encapsulated bacteria (e.g. S. pneumoniae and H. influenzae). In the absence of chronic GvHD, this deficiency will often be transient and will resolve over time. In other cases, it may persist indefinitely. However, even in these cases, active immunisation may be beneficial, especially with conjugate vaccines. Although most infectious mortality is observed during the first 6 months after transplant, late infections may be life-threatening (particularly S. pneumoniae) and should not be neglected, especially in patients with chronic GvHD. The development of non-myeloablative approaches has modified the timing and features of the main infectious complications after transplant, although their overall incidence is not greatly changed. Neutropenia almost disappears or is significantly reduced, as is mucositis, and therefore the risk for early bacterial or fungal infections decreases (2). However, patients who undergo reduced intensity conditioning (RIC) regimens are usually older than those who receive conventional conditionings. As the main risk factor for severe infections is GvHD, and as, until now, the risk of GvHD is roughly comparable whatever the conditioning regimen, patients who receive reduced intensity conditioning remain at risk of infections, especially viral and fungal infections. Actually, although we may agree that RIC regimens decrease the overall transplant-related mortality, there is no clear evidence from the literature that RIC decreases infection-related mortality (3, 4). After autologous HSCT, morbidity and mortality related to infection is much lower. Since more than 95% of autologous HSCT are performed with peripheral blood stem cells, neutropenia generally lasts no more than 14 days, and the risk of bacterial sepsis is close to that observed in similar patients after chemotherapy. Fungal risk is extremely low and one may consider that these patients are not at risk for 200
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Aspergillus, and in standard situations they do not need rooms with filtered air. However, in addition to the risk of the autologous HSCT procedure itself, these patients may also be at risk due to previous immunosuppressive factors, such as prolonged steroid therapy, or hypogammaglobulinemia, which may favour other, non neutropenia-related, infections. Late infections may also occur after autologous transplant, but are exceptional without total body irradiation.
3. Clinical management of fever and infectious complications Because allogeneic transplant patients are those at higher risk for opportunistic infections and severe outcome, most of the recommendations given below mainly apply to these patients, but autologous transplant recipients should benefit from similar approaches, at least for the first 2 months after transplant. Two special clinical settings are taken as examples of the management of these patients when infection is probable, or even just possible, because they require special attention with essential diagnostic and therapeutic procedures. 3.1. Fever Febrile neutropenia is a special situation where bacterial infection must be the main target of empirical anti-infective therapy. There are not many differences between febrile neutropenia in patients at the neutropenic phase of stem cell transplant, and febrile neutropenia following chemotherapy, except that patients conditioned with total body irradiation are more likely to suffer from mucositis which is one of the main factors for streptococcal bacteraemia. Only 30% of febrile neutropenic episodes are microbiologically documented, with 20% due to Gram-positive cocci - mainly coagulase negative staphylococci - and 10% due to Gram-negative bacteria. As in any febrile neutropenia, broad-spectrum antibiotics should be administered promptly after blood cultures and sampling of any clinical site of infection whenever possible. Guidelines from the recent ECIL 1 (European Conference on Infections in Leukaemia) have clarified the need for aminoglycosides (5) and glycopeptides (6) in febrile neutropenia: the group considers that in standard situations betalactam monotherapy is as efficacious as betalactam plus aminoglycoside combination, both as initial empirical therapy and in case of persistent fever. Therefore, considering their potential toxicity, the use of aminoglycosides is not recommended, except in case of severe sepsis or septic shock where aminoglycosides are an initial option for first line antibiotic therapy, but where they should be quickly reconsidered according to the clinical outcome (5). Similarly, the use of glycopeptides as initial antibacterial therapy - combined with a betalactam - should be restricted to patients with hypotension or shock, skin or soft tissue infections (including high suspicion of central catheter infection) or in patients
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with known colonisation with methicillin - resistant S. aureus (6). If fever persists after 3 to 5 days of empirical antibacterials, fungal and viral infection must be considered, and an empirical antifungal treatment must be administered (7). Several antifungals may be used in this indication, but liposomal amphotericin B and caspofungin are recommended (8). Routine screening for fungal infection by galactomannan antigenaemia and lung CT-scan offers new hope of moving from the classical empirical strategy to a more risk-targeted, so-called “pre-emptive”, strategy (9, 10), but this strategy is, until now, not standardised for allogeneic HSCT recipients. When a transplant patient is febrile but not neutropenic, any infection should be considered and possible diagnoses should be made according to the clinical presentation and timing of fever after transplant. Although fever can be a feature of GvHD, most febrile episodes in transplant patients are due to infection, and diagnostic methods should be used to identify the responsible pathogen. The main difference between neutropenic and non-neutropenic patients is that in nonneutropenic patients, it is usually possible to investigate the patient properly before starting anti-infective drugs. For example, when an episode of pneumonia occurs within the first 6 months of transplant, and has developed over several days or weeks, it is usually possible to wait for fibre-optic bronchoscopy and BAL, whenever possible within 24h, before giving antibiotics. However, it is not always advisable to wait even in non-neutropenic patients, e.g. those with hypoxemic pneumonia, or severe sepsis. 3.2. Pneumonia Pneumonia complicates the course of half of allogeneic HSCT recipients treated with conventional conditioning regimens. There are many possible causes of pulmonary infiltrates observed after transplant. However, two thirds are due to infection. Pulmonary oedema, pulmonary embolism, alveolar haemorrhage, and alveolar proteinosis, may also occur, especially within the first 3 months after transplant. Fibre-optic bronchoscopy with BAL is the main diagnostic tool. It must be performed early in the course of pneumonia, before administration of antibacterials whenever possible, and a protected sample should be obtained for quantitative microbiological assessment, by either brushing or aspiration. BAL has a limited diagnostic yield for fungal infections. However, despite this limitation, it is the procedure with the highest risk/benefit ratio for first-line investigation. Since the use of prophylactic or preemptive treatment for CMV infection, which makes the occurrence of CMV pneumonia very rare, special attention must be paid to the possibility of respiratory virus pneumonia. Transbronchial lung biopsy is associated with a greater risk of pneumothorax and bleeding, and no greater diagnostic benefit for most causes of pneumonia in the transplant setting, except for mycobacterial and fungal infections. Open lung biopsy should be considered in sub-acute pneumonia when BAL does not 202
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provide a clear diagnosis. Although there is not much available data about the value of indirect non-invasive procedures in such patients (i.e., legionella, pneumococcal, cryptococcal antigens in blood and/or urine), such procedures deserve to be used since they may easily provide indirect evidence of the cause of pneumonia.
4. Bacterial infections Two important points should be remembered: - Timing and immune reconstitution are key points in making a possible diagnosis at the bedside of a transplant patient suspected of bacterial infection. - Additionally, HSCT patients are among the most immunodepressed patients in the hospital. Therefore, they are at risk for nosocomial infections, depending on the environmental measures and the epidemiology of the centre. Bacterial infections occurring during the neutropenic phase are usually the same as those observed during any chemotherapy-induced neutropenia, i.e. streptococci and Gram-negative bacteria. Neutrophil recovery usually marks the end of the bacterial risk for most autologous HSCT patients, but not for allogeneic HSCT recipients who remain at high risk for nosocomial infection for many weeks after neutrophil recovery, especially when they stay in the hospital for the treatment of severe GvHD with multiple complications, and keep their central IV line. After 2 months, the risk for encapsulated bacteria, mainly for S. pneumoniae and H. influenzae, appears and is strongly - but not exclusively - linked to chronic GvHD. Specific deficiencies in anti-S. pneumoniae and anti-PRP antibodies - the main H. influenzae type B capsular polysaccharide - have been noted, which may predispose the patient to these infections. S. pneumoniae may cause bacteraemia and/or pneumonia or sinusitis. Fulminant fatal outcomes may be observed, even years after transplant. S. pneumoniae infections are mainly observed in patients with GvHD and in those who have received TBI (11). Although it rarely causes fulminant disease, H. influenzae may be responsible for upper and lower respiratory tract infection, and bacteraemia. Other rare bacterial infections have been reported after transplant, i.e., mycobacterial or legionella infection. However, due to the rarity of these cases, the timing and risk factors are not well identified. 4.1. Prevention of bacterial infections During the early phase of transplant, three preventive measures must be considered: a) Because of the hospital environment and its resistant bacteria, the physical environment of transplant patients should aim to decrease the risk of nosocomial infection, as in any immunocompromised patient. Different measures can be implemented. The easiest is simple protection including mask, gloves, and
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gowns, which is usually enough for autologous transplant patients. Although there is no clear demonstration in the literature that such measures are superior to hand washing alone in preventing cross-transmission, this kind of isolation is usually easier to control enforce than hand washing alone, and such visible protection usually discourages the staff from making unnecessary approaches to the patient during the neutropenic phase. The control of room air quality through filtration (HEPA) is the main measure that can be taken to decrease the risk of aspergillosis but there is no demonstration that such air treatment has any impact on the incidence of bacterial pneumonia. Water control is also necessary, in any units caring for high-risk patients, to avoid contamination of the patients with Legionella sp. or P. aeruginosa. b) The second measure is the prevention of bacterial translocation by oral antibacterials - absorbable such as quinolones, or non-absorbable such as nonabsorbable gut decontamination - and a diet low in microbes. The gut is the main reservoir for Gram-negative bacteria, and a source of subsequent entry into the body. A diet low in microbes is generally recommended for these patients, but the measures vary from one centre to another. The minimal measures are to avoid fresh vegetables and fruits, and any food suspected to content high quantities of bacteria. This is a logical recommendation, although its benefit alone has never been clearly shown. Gut decontamination is differently approached in different centres and countries. Quinolones have been shown, in comparative trials, to decrease the risk for Gram-negative bacteraemia and the number of days of fever in patients with prolonged neutropenia. They are widely used for allogeneic HSCT patients in Europe, especially ciprofloxacin. In acute leukaemia patients and autologous HSCT recipients, a large trial comparing levofloxacin to placebo has shown that the use of levofloxacin during the neutropenic phase significantly reduces the risk of fever, bacterial infection and bacteraemia, and especially of Gram-negative infections, reducing the cost of intravenous antibacterials for febrile neutropenia (12). So far, there are no data on levofloxacin in allogeneic HSCT patients. Once started, quinolones should be given until the first febrile episode, or until the recovery of neutropenia in the absence of fever. However, quinolones may select Gram-negative quinolone-resistant bacteria. Therefore, their use is not recommended in units with a high level of quinolone resistance among Gramnegative bacteria, and should be associated with periodic monitoring of the local epidemiology (13). The other option, widely used in some European countries, is the use of non-absorbable antibiotics (i.e. colimycine and aminoglycosides). They similarly reduce the risk for Gram-negative bacteraemia, but historical trials failed to show any survival benefit. Non-absorbable gut decontamination 204
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has constraints, especially the need to be combined with sterile food and with frequent microbiological controls of the stools to look for the possible selection of resistant strains. Whatever the option (quinolones, or non-absorbable gut decontamination), an antifungal prophylaxis must also be given to decrease the risk of overgrowth of yeasts in the gut. c) The third measure is the management of central IV lines. Catheters may be the source of bacteraemia, with significant morbidity and potential mortality. During neutropenia, it is controversial whether a catheter may be left in situ when blood cultures have documented a pathogen, except in the case of methicillin resistant S. aureus, Candida sp., Bacillus sp. and Corynebacterium JK, and any hospitalacquired resistant pathogen, such as P. aeruginosa or Acinetobacter sp, for which there are clear recommendations to take the catheter out (2). Central catheter management should be formalised in any transplant unit in written procedures. From the third month after transplant, transplant patients may be profoundly hypogammaglobulinaemic (immune globulin levels below 3 g/L). These patients should receive IV immunoglobulin replacement to maintain immunoglobulin serum levels over 3 g/L, especially when they have chronic GvHD and are receiving immunosuppressive therapy. The most logical and cost-effective approach to prevent S. pneumoniae and H. influenzae infections is active immunisation. For S. pneumoniae, the historical trials with polysaccharide vaccines have been disappointing since they show that patients with GvHD, or receiving steroids, those with the higher risk of infection, have a poor antibody response, especially before 6 months post-transplant. Therefore, long-term antibiotic prophylaxis with penicillin is still the practice in most centres for allogeneic transplant recipients, but the development of penicillin resistance makes this policy unsatisfactory. The recent availability of a conjugate pneumococcal vaccine offers new hopes of improving the protection of HSCT patients since the conjugation of the pneumococcal polysaccharides to proteins elicits a T-cell dependent immune response. A first study shows that 3 doses of the heptavalent pneumococcal conjugate vaccine, given at 3, 6 and 12 months after allogeneic HSCT induces protective IgG concentrations in 60% of the patients regardless of donor immunisation (14). Another study in children showed that 3 doses of the heptavalent vaccine given from 6 months allows protective antibody levels in more than two thirds of the patients (15). A prospective EBMT trial recently completed shows that the administration can be given as soon as 3 months post-transplant. Similarly, the H. influenzae type-B tetanus conjugate vaccine has been shown to be highly effective, giving a satisfactory response in 85% of immunised HSCT
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patients, and should be administered from 4 months. Probably because of the high immunogenicity of such conjugate vaccines, the response does not seem to be influenced by the presence of GvHD. Therefore, these vaccines are recommended in all patients, and even more in those with GvHD who are at higher risk of infection. The main recommendations of the IDWP for active immunisation in transplant patients are summarised in Table 1 (16).
5. Fungal infections Aspergillus is the most serious fungal infection after allogeneic stem cell transplant (17), and is the main cause of infectious death (18). Reported incidences vary from 0 to 20% of transplants. The most common site is the lung and GvHD is the main risk factor. A first peak of incidence occurs during the neutropenic period, particularly in leukaemic patients who may have been previously colonised. A second wave of aspergillosis occurs during the 2nd and 3rd months after transplant and is strongly linked to GvHD; this is now more important than the first peak and late cases are more and more frequent. In patients previously infected by Aspergillus during induction or consolidation treatment phases, fungal recurrence may occur in roughly one third of the patients (19) and the use of a secondary antifungal Aspergillus prophylaxis is widely admitted, although the optimal choice of the drug is not fixed. Despite major improvements in the treatment of fungal infections (20), the mortality of Aspergillus in allogeneic HSCT patients remains over 50% in recent series. PCR and galactomannan antigenaemia may help in the early detection of Aspergillus infections. Candida infection is more rare and has no special clinical presentation in transplant patients when compared to other haematology patients. More and more nonAspergillus, non-Candida infections are reported in HSCT patients. Pneumonias due to endemic fungi, such as histoplasmosis or coccidioidomycosis, particularly in North America, must be considered in these patients, as well as the emerging fungi, including Trichosporon, Alternaria, Fusarium, and the Mucorales. Any of these fungi can cause pneumonia in transplant patients, as in similarly immunocompromised hosts, and their management needs a close collaboration between the mycologist and the clinician. 5.1. Prevention of fungal infections The most efficient protection from fungal air-born infections during the neutropenic phase of allogeneic transplant is the use of air filtration with positive pressure. This kind of isolation has been shown to decrease the early risk of Aspergillus in 206
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Table 1: EBMT recommendations for immunisations of long-term survivors after allogeneic and autologous stem cell transplantation (16) Available forms
Available data in HSCT patients
Recommended after HSCT
Strength No of of recommen- doses dation+
Time Improved after HSCT by donor (months) vaccination
BACTERIAL VACCINES S. pneumoniae
PS Conjugate PS
Yes Yes
Yes AII Yes (subgroups) CII
1 3
12 Unclear
No Yes
H. influenzae type B
Conjugate PS
Yes
Yes
BII
3
6
Yes
N. meningitidis types A and C N. meningitidis type C
PS
Yes
Individual assessment
CII
1
6-12
Unknown
Conjugate PS
No
1
6
Unknown
BCG
Live
No
Contra-indicated EII
NA
NA
Unknown
Tetanus
Toxoid
Yes
Yes
BII
3
6-12
Yes
Diphtheria
Toxoid
Yes
Yes
BII
3
6-12
Likely
B. pertussis#
Acellular, toxoid +/- other antigens
Yes
See text
CIII
3
6-12
Unknown
Inactivated
Yes
Yes, yearly
AII
1
4-6
Unknown
Inactivated polio Inactivated
Yes
Yes
BII
3
6-12
Unknown
VIRAL VACCINES Influenza
Hepatitis B
Inactivated plasma Yes or recombinant DNA derived
See text
BII
3
6-12
Yes
Hepatitis A virus
Inactivated
No
In endemic areas CIII and in travellers
3
6-12
Unknown
Measles§
Live
Yes
Individual assessment
BII
1
24*
Unknown
Rubella§
Live
Yes
Individual assessment
BIII
1
24*
Unknown
Mumps§
Live
Yes
Individual assessment
CIII
1
24*
Unknown
Varicella
Live
Limited
Individual assessment
CIII
Unclear Before HSCT Unknown or at 24*
Yellow fever
Live
Limited
Individual asses- CIII sment, travellers
1
24*
Unknown
+The recommendations are graded according to the CDC system; *Not in patients with chronic GvHD or ongoing immunosuppression; #Combination vaccines including also tetanus, diphtheria, and pertussis with or without HIB and poliovirus components are available; §Usually combined in a MMR vaccine; BCG: Bacille Calmette-Guérin; HIB: Haemophilus influenzae type b; PS: Polysaccharide
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allogeneic HSCT. Good management of air filters, and effective collaboration with the infection control unit of the hospital are of paramount importance for allogeneic transplant units, and for haematology departments in general. Epidemics of mould infections have been reported in stem cell transplant units and have led to interruption of transplant programs until the infections can be controlled (21). However, air filtration, of course, does not provide any protection from yeast infections. In two randomised trials, fluconazole prophylaxis at the dose of 400 mg/day has been shown to reduce the risk for invasive fungal infections, especially Candida infections, and is associated with a long-term survival benefit. However, because non-albicans Candida are more and more frequent in haematology patients, the benefit of fluconazole prophylaxis is probably lower than ten years ago, and anyway unsatisfactory considering the main concern of the transplant community is mould infections. Recently, Ullmann et al. have shown that posaconazole (600 mg/d), when given from the onset of GvHD, significantly decreases the incidence of proven and probable invasive fungal infection, especially Aspergillus (22). In autologous HSCT, except in centres with a high incidence of candidaemia, the risk of invasive Candida infection is low, and therefore the use of fluconazole in this setting is optional (23). 5.2. Treatment of fungal infections Voriconazole, a triazole antifungal agent, is fungicidal active against Aspergillus and Candida species, including non-albicans Candida, and other rare fungal infections such as Fusarium or Scedosporium species which occur in stem cell transplant patients. In the first-line treatment of aspergillosis, it has been shown to be more effective and better tolerated than amphotericin B (20). Despite a substantial number of side effects (including visual disturbances, confusion, skin reactions, and liverfunction abnormalities), and potential drug interactions, especially with cyclosporin, voriconazole is widely used after stem cell transplant. A recent, large prospective study comparing 3 vs. 10 mg/kg/d of liposomal amphotericin B in first line treatment of Aspergillus infections did not find any advantage to the 10 mg dose, but showed comparable results of the 3 mg/kg/d arm with the voriconazole study (24). Echinocandins have been studied in refractory aspergillosis with encouraging results but no study of first-line therapy has been published so far. Echinocandins are excellent candidates for combinations with either azoles, or polyenes. However, there are, until now, no prospective data published on first line therapy of fungal infections with echinocandin combinations. These combinations will be extremely expensive, and there is no evidence as yet that they will provide better results than 208
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single drug regimens. Prospective, comparative trials need to be done before any conclusion can be drawn. The ECIL 1 has recently published treatment guidelines for Candida and Aspergillus infections (25).
6. Viral infections Viral infections are frequent after HSCT. They may be life threatening, especially when affecting lung, liver, or central nervous system in allogeneic HSCT recipients. The availability of new antiviral agents in the last 2–3 years, together with data from comparative trials, have allowed a better control of herpes virus infections. However, due to the subsequent decrease in CMV infection and CMV disease, new viral infections have emerged, especially due to respiratory viruses and adenovirus. Herpes simplex virus (HSV) infections are extremely common, due to reactivation in seropositive patients. The main early manifestation is mucous lesions, difficult to distinguish from chemotherapy-induced mucositis in the absence of viral documentation. These lesions are painful and maybe the portal of entry of bacteria from the gut. In seropositive patients, prophylaxis with acyclovir or valaciclovir is recommended to decrease the risk of reactivation during the early phase of transplant. In case of HSV infection, IV acyclovir is usually effective. Acyclovir resistance is rare in HSCT patients, but this possibility must be considered in case of HSV disease documented during prophylaxis. Due to common mechanisms at the cellular level (thymidine-kinase dependent), resistance to acyclovir is associated with resistance to ganciclovir and famciclovir. The best choice in case of acyclovir resistance of HSV is foscavir. Cytomegalovirus (CMV) disease has historically been a main cause of death in allogeneic transplant patients, except where both donor and recipient are seronegative. Since the demonstration that CMV infection usually precedes CMV disease, and considering the poor prognosis of CMV disease even when treated, two strategies - prophylactic and pre-emptive - have been developed in order to reduce the risk of CMV disease. First, prophylactic trials comparing IV ganciclovir versus placebo showed that ganciclovir prevents the risk of CMV infection and disease, but does not improve survival and additionally favours the delay of specific immune reconstitution, and consequently, the occurrence of late CMV infection. This prophylactic strategy was finally not more effective than a pre-emptive strategy. Therefore, prophylactic strategies are usually reserved for patients with high-risk of CMV disease, such as mismatched transplant recipients. For other patients, a preemptive strategy is more cost-effective. Controlled trials are needed to know
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whether the oral pro-drug of ganciclovir, valganciclovir, will be as effective and no more toxic than the IV ganciclovir. First experiences are encouraging but suggest caution is needed because of the different pharmacokinetics of the two drugs (26). The clinical impact of theses differences should be explored further. Although most first line pre-emptive strategies have used ganciclovir, foscavir has been shown, in an EBMT comparative trial, as effective and no more toxic than ganciclovir (27). Both can be used as first-line treatment of CMV infection, for an initial duration of 2 weeks. If CMV is still detected after 2 weeks of therapy, an additional course of 2 weeks should be given. Cidofovir has been studied only in uncontrolled trials and because of its toxicity profile; its use should be reserved for second line preemptive therapy. These strategies must be based on sensitive diagnostic methods of CMV infection. Allogeneic HSCT recipients must be monitored for CMV in peripheral blood at least weekly with a sensitive method, until day 100 after transplant, and longer in case of prolonged GvHD or previous CMV reactivation. Antigenaemia testing detects CMV antigen pp65 in leukocytes by immune staining with monoclonal antibodies. This test is semiquantitative and rapid. PCR is also widely used. The quantification of viral load seems to be important since higher levels of CMV DNA are indicators for a higher risk of CMV disease. The availability of Real Time or Light Cycler technologies has improved the quantitative evaluation of the viral load. Detection of mRNA by nucleic acid sequence-based amplification (NASBA) is also available and has been shown to be similarly effective as pp65 antigenaemia or detection of DNA by PCR for the early detection of CMV infection. Because the risk of CMV disease is extremely low after autologous transplant, systematic CMV screening is not recommended in this population, except in high-risk patients, like those receiving CD34-selected grafts, or patients who have recently received fludarabine, 2-CDA, or alemtuzumab. In additionally to antiviral strategies, CMV prophylaxis must include transfusion policies to avoid acquisition of CMV through blood products, especially for CMV seronegative recipients of seronegative donors. These patients must receive blood products either from CMV seronegative donors exclusively, or leukocyte-depleted products. Although less established by prospective trials, the same policy is logical for CMV seronegative recipients of autologous transplants. Varicella-zoster virus (VZV) infections occur both after allogeneic and autologous transplants. Primary varicella may be severe in both populations. IV acyclovir is the therapy of choice. Valaciclovir can also be used, especially in late infections, and in patients who do not receive high-dose immunosuppressive drugs. These patients, when given valaciclovir, must be correctly managed in case of rash extension or 210
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systemic manifestations, and informed of the possible need for switching to the IV form of the antiviral. Human herpes virus 6 (HHV6) infection after allogeneic HSCT has been associated with pneumonia, delayed marrow engraftment, and particularly with prolonged thrombocytopenia and encephalitis. HHV6-DNA is frequently detected in blood during the first months after transplant in asymptomatic patients, so that its implication in clinical symptoms is usually difficult to establish, except in encephalitis when HHV6-DNA is detected in CSF. Epstein-Barr virus associated lymphoproliferative disease (EBV-LPD) is a lifethreatening complication occurring after allogeneic HSCT. Monitoring of the EBV viral load by quantitative PCR permits the early detection of EBV reactivation that may lead to EBV-LPD. Recipients of a T-cell-depleted HSCT, or patients conditioned with antithymocyte globulins are at higher risk of EBV-LPD. In these patients, preemptive therapy of EBV reactivation with rituximab has been shown to improve outcome. Infusion of EBV-specific cytotoxic T-cells has also been studied in highrisk patients with elevated EBV-DNA levels. However, the exact indications for preemptive therapy based on EBV-viral load for preventing EBV-LPD are not yet clear and should highly depend on the transplant population. Prospective trials are needed. 6.1. Respiratory virus infections Respiratory viruses, including respiratory syncytial virus (RSV), parainfluenza virus, rhinovirus, and influenza virus, appear to be now more frequent than CMV pneumonia. A prospective study from the EBMT showed an incidence of respiratory virus pneumonia of 2.1% in allogeneic, and 0.2% in autologous transplant patients (28). Most cases in this series were due to RSV or influenza A. The mortality of these infections also varies among series, and with the time after transplant and the degree of immunosuppression, but it may be as high as 80% in RSV pneumonia. Few data are available in the literature on the efficacy of antivirals in RSV pneumonia. Due to the rarity of the disease, and the poor prognosis of the lower tract infections, prospective trials are extremely difficult to perform. In the European experience, there was no clear advantage or disadvantage of adding IV ribavirin to either aerosolised ribavirin or to IV immune globulins. The best therapeutic option for RSV pneumonia is not established. Due to the risk of spread in the transplant unit, it is important to diagnose these patients very early, and to prevent transmission in the ward. Oseltamivir has been used in an open study to control an influenza A outbreak and appears to be safe in allogeneic transplant recipients (29).
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6.2. Adenovirus infections Adenovirus infections can be a cause of severe disseminated infections in allogeneic HSCT recipients. Patients receiving mismatched or unrelated donor transplants, patients with severe acute GvHD, patients showing isolation from multiple sites or from blood are at high risk of developing organ involvement due to adenovirus. There are currently no established strategies for prophylaxis or treatment of adenovirus disease. Spontaneous recovery may be observed. However, in high-risk patients such as children receiving mismatched transplants, adenovirus infection usually develops into adenovirus disease, which has a high mortality rate, and pre-emptive treatment should be logical but has not been studied extensively. A recent retrospective study by the IDWP of the EBMT has showed that cidofovir was effective in 10/16 patients with invasive adenovirus disease (30). Ribavirin has been used in case reports with varying outcome. Both drugs - cidofovir and ribavirin - may be effective in preemptive therapy. Prospective controlled studies are needed to define the best strategy.
7. Other infections Toxoplasmosis occurring after HSCT has been mainly investigated in Europe, due to a higher seroprevalence of the disease when compared to US. Patients at risk are those who are seropositive for toxoplasmosis before transplant, irrespectively of the serology of the donor. Blood PCR allows early detection of toxoplasma reactivation. A prospective study from the IDWP on 106 allogeneic toxoplasma seropositive recipients, screened weekly by blood PCR, showed an incidence of PCR-documented infection of 16%, and an incidence of toxoplasma disease of 6% up to 6 months post-transplant. All patients developing disease were previously or simultaneously PCR-positive in blood (31). Most of these reactivations occur in patients with GvHD, while trimethoprim-sulfamethoxazole has been stopped for side effects, and replaced by aerosolised pentamidine for P. jiroveci prophylaxis. Whether asymptomatic toxoplasma infection documented by blood PCR should be treated is not yet clear. However, there are reports in the literature where high-risk patients, with severe GvHD, developed toxoplasma infection which evolved, in the lack of specific therapy, to toxoplasma disease with a high mortality rate. P. jiroveci pneumonia must be prevented in allogeneic stem cell transplant recipients from engraftment to at least 6 months, even longer in case of prolonged immunosuppression. The best option is trimethoprim-sulfamethoxazole. In case of intolerance, alternatives are dapsone or aerosolised pentamidine, but the latter does not provide optimal protection (32). 212
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8. Other preventive strategies Many issues regarding infectious complications in HSCT patients remain to be explored, especially in allogeneic HSCT recipients. Active immunisation against tetanus, poliovirus, and diphtheria is highly recommended in all transplant populations, due to the usual loss of specific immunity both after autologous and allogeneic stem cell transplantation. This is the only way to allow leukaemic patients to have similar levels of immunity to these pathogens - even rarely encountered - as the normal population. Immunisation with live vaccines is classically prohibited in immunocompromised patients. However, transplant patients may require active immunisation with vaccines which only exist in a live form, i.e. measles, mumps, or yellow fever. Ljungman et al. have showed that live vaccine to measles, mumps and rubella may be safely administered to transplant children, as far as they are at more than 2 years after transplant, have no GvHD, and are not receiving immunosuppressive drugs. Immune globulins have been administered for years to allogeneic HSCT recipients to try to prevent the occurrence of GvHD and to reduce infectious mortality. A French double-blind study comparing 3 doses of IVIg to placebo in recipients of HLA-identical sibling donors shows that IVIg administration did not affect the incidence of infections over the first 6 months after transplant, the occurrence of GvHD, or survival. On the other hand, severe veno-occlusive disease was significantly more frequent in patients receiving high doses of IVIG (250 mg/kg and 500 mg/kg/weekly) (18). Similar findings have been reported in another randomised trial in unrelated transplant patients. Consequently, considering the availability of other strategies now available for infection prophylaxis, and considering the high cost of IVIg, prophylactic administration of IVIg is not recommended in allogeneic transplant recipients.
References 1. Gratwohl A, Brand R, Frassoni F, et al. Cause of death after allogeneic haematopoietic stem cell transplantation in early leukaemias: An EBMT analysis of lethal infectious complications and changes over calendar time. Bone Marrow Transplant 2005; 36: 757769. 2. Junghanss C, Marr KA, Carter RA, et al. Incidence and outcome of bacterial and fungal infections following nonmyeloablative compared with myeloablative allogeneic hematopoietic stem cell transplantation: A matched control study. Biol Blood Marrow Transplant 2002; 8: 512-520. 3. Diaconescu R, Flowers C, Storer B, et al. Morbidity and mortality with nonmyeloablative compared to myeloablative conditioning before hematopoeitic cell transplantation from HLA matched related donors. Blood 2004; 104: 1550-1558. 4. Scott BL, Sandmaier BM, Storer B, et al. Myeloablative vs nonmyeloablative allogeneic
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transplantation for patients with myelodysplastic syndrome or acute myelogenous leukemia with multilineage dysplasia: A retrospective analysis. Leukemia 2006; 20: 128-135. 5. Drgona L, Paul M, Bucaneve G, et al. The need for aminoglycosides in combination with betalactams for high-risk, febrile neutropenic patients with leukaemia. Eur J Cancer 2007; Suppl. 5: 13-22. 6. Cometta O, Marchetti O, Calandra T. Empirical use of anti-Gram positive antibiotics in febrile neutropenic cancer patients with acute leukaemia. Eur J Cancer 2007; Suppl. 5: 23-31. 7. Hughes WT, Armstrong D, Bodey GP, et al. 2002 guidelines for the use of antimicrobial agents in neutropenic patients with cancer. Clin Infect Dis 2002; 34: 730-751. 8. Marchetti O, Cordonnier C, Calandra T. Empirical antifungal therapy in neutropenic cancer patients with persistent fever. Eur J Cancer 2007; Suppl. 5: 32-42. 9. Maertens J, Theunissen K, Verhoef G, et al. Galactomannan and computed tomographybased pre-emptive antifungal therapy in neutropenic patients at high risk for invasive fungal infection: A prospective feasibility study. Clin Infect Dis 2005; 41: 1242-1250. 10.Cordonnier C, Pautas C, Maury S, et al. Empirical versus pre-emptive antifungal strategy in hig-risk febrile neutropenic patients: A prospective randomized study. 48 th ASH Annual meeting, Orlando, Florida, 9-12 December 2006. 11.Engelhard D, Cordonnier C, Shaw PJ, et al. Early and late invasive pneumococcal infection following bone marrow and stem cell transplantation. Br J Haematol 2002; 117: 444-450. 12.Bucaneve G, Micozzi A, Menicheti F, et al. Levofloxacin to prevent bacterial infection in patients with cancer and neutropenia. N Engl J Med 2005; 353: 977-987. 13.Bucaneve G, Castagnola E, Viscoli C, et al. Quinolone prophylaxis for bacterial infections in afebrile high risk neutropenic patients. Eur J Cancer 2007; Suppl. 5: 5-12. 14.Molrine DC, Antin JH, Guinan EC, et al. Donor immunization with pneumococcal conjugate vaccine and early protective antibody responses following allogeneic hematopoietic cell transplantation. Blood 2003; 101: 831-836. 15.Meisel R, Kuypers L, Dirksen U, et al. Pneumococcal conjugate vaccine provides early protective antibody responses in children after related and unrelated allogeneic hematopoietic stem cell transplantation. Blood 2007; 109: 2322-2326. 16.Ljungman P, Engelhard D, Locasciulli A, et al. Vaccination of stem cell transplant recipients. Recommendations of the Infectious Diseases Working Party of the EBMT. Bone Marrow Transplant 2005; 35:737-746. 17.Marr KA. Epidemiology and outcome of mold infections in hematopoietic stem cell transplant recipients. Clin Infect Dis 2002; 34: 909-917. 18.Cordonnier C, Chevret S, Legrand M, et al. Should immunoglobulin therapy be used in allogeneic transplantation? A randomized, double-blind, dose-effect, placebo-controlled, multicenter trial. Ann Intern Med 2003; 139: 8-18. 19.Offner F, Cordonnier C, Ljungman P, et al. Impact of previous aspergillosis on the outcome of bone marrow transplantation. Clin Infect Dis 1998; 26: 1098-1103. 20.Herbrecht R, Denning DW, Patterson TF, et al. Voriconazole versus Amphotericin B for primary therapy of invasive aspergillosis. New Engl J Med 2002; 347: 408-415. 21.Gratwohl A, McCann S, Byrne JL, et al. Outbreaks of infectious disease in stem cell transplant units: A silent cause of death for patients and transplant programmes. Bone Marrow 214
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Transplant 2002; 29: S 53. 22.Ullmann A, Lipton J, Vesole DH, et al. Posaconazole or Fluconazole for prophylaxis in severe graft-versus-host disease. N Engl J Med 2007; 356: 335-347 23.Maertens J, Frere P, Lass-Flörl C, et al. Primary antifungal prophylaxis in leukaemia patients. Eur J Cancer 2007; Suppl. 5: 43-48. 24.Cornely OA, Maertens J, Bresnik M, et al. Liposomal amphotericin B as initial therapy for invasive mold infection: A randomized trial comparing a high-loading dose regimen with standard dosing. Clin Infect Dis 2007; 44: 1298-1306. 25.Herbrecht R, Flückiger U, Gachot B, et al. Treatment of invasive Candida and Aspergillus infections in adult haematological patients. Eur J Cancer 2007; Suppl. 5: 49-59. 26.Einsele H, Reusser P, Bornhaüser, et al. Oral valganciclovir leads to higher exposure to ganciclovir than intravenous ganciclovir in patients following allogeneic stem cell transplantation. Blood 2006; 107: 3002-3008. 27.Reusser P, Einsele H, Lee J, et al. Randomized multicenter trial of foscarnet versus ganciclovir for preemptive therapy of cytomegalovirus infection after allogeneic stem cell transplantation. Blood 2002; 99: 1159-1164. 28.Ljungman P, Ward KN, Crooks BNA, et al. Respiratory virus infections after stem cell transplantation. A prospective study from the Infectious Diseases Working Party of the European Group for Blood and Marrow Transplantation. Bone Marrow Transplant 2001; 28: 479-484. 29.Vu D, Peck AJ, Nichols WG, et al. Safety and tolerability of oseltamivir prophylaxis in hematopoietic stem cell transplant recipients: A retrospective case-control study. Clin Infect Dis 2007; 45: 187-193. 30.Ljungman P, Ribaud P, Eyrich M, et al. Cidofovir for adenovirus infection after allogeneic stem cell transplantion (HSCT). A retrospective survey of the Infectious Diseases Working Party of the European Group for Blood and Marrow Transplantation. Bone Marrow Transplant 2003; 31: 481-486. 31.Martino R, Bretagne S, Einsele H, et al. Early Detection of Toxoplasma Infection by Molecular Monitoring of Toxoplasma gondii in Peripheral Blood after Allogeneic Stem Cell Transplantation. Clin Infect Dis 2005; 40: 67-78. 32.Center for Disease Control. Guidelines for preventing opportunistic infections among hematopoeitic stem cell transplant recipients. Recommendations of CDC, the Infectious Diseases Society of America, and the American Society of Blood and Marrow Transplantation. October 20, 2000 / 49 (RR10); 1-128. (www.cdc.gov).
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Among these statements comparing RIC regimens and conventional regimens, one is true. Which one? a) RIC decreases the incidence of encapsulated bacterial infections . . . . . . . . . HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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b) RIC increases the incidence of early bacterial infections . . . . . . . . . . . . . . . . . . . c) RIC decreases the incidence of fungal infections . . . . . . . . . . . . . . . . . . . . . . . . . . . d) RIC delays the occurrence of CMV infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Among these statements about S. pneumoniae vaccination, one is true. Which one? a) The conjugate pneumococcal vaccine is a live vaccine . . . . . . . . . . . . . . . . . . . . . b) The conjugate pneumococcal vaccine includes more antigens than the olysaccharide vaccine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The polysaccharide vaccine is recommended in allogeneic SCT recipients from 3 months after transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The conjugate pneumococcal vaccine is more immunogenic because it induces a T-cell response . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Among these statements about the use of antibacterials in febrile neutropenic patients, one is true. Which one? a) The use of aminoglycosides is not recommended in febrile neutropenic patients without severe sepsis or septic shock. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The use of glycopeptides in combination with a betalactam is recommended as first line treatment of febrile neutropenic patients in standard situations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) In neutropenic patients, the risk of S. aureus bacteraemia is increased in the presence of severe mucositis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) In neutropenic patients, the risk of streptococcal bacteraemia is increased in the presence of catheter infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Among these statements about antifungal strategies in allo-HSCT, one is true. Which one? a) Posaconazole prophylaxis decreases the incidence of invasive fungal infection when given from transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Posaconazole prophylaxis decreases the incidence of invasive fungal infection when given from the onset of GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Increasing the dose of liposomal Amphotericin B from 3 to 10 mg/kg in first line treatment of aspergillosis improves the clinical response . . . . d) It has been shown in a prospective study that the combination of 216
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caspofungin with voriconazole is more efficient than voriconazole alone in first line treatment of aspergillosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Among these statements about toxoplasmosis after allo-HSCT, one is true. Which one? a) Toxoplasmosis after allo-HSCT occurs almost exclusively in patients who were seropositive for toxoplasma before transplant . . . . . . . . . . . . . . . . . . . . . . . . b) Reactivation documented by blood PCR is observed in approximately 30–40% of seropositive allogeneic transplant recipients. . . . . . . . . . . . . . . . . . . c) Asymptomatic toxoplasma infection never develops into toxoplasma disease. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Aerosolised pentamidine prophylaxis is effective against both P. jiroveci and toxoplasma infection. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 11
Graft versus host disease
A. Devergie
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CHAPTER 11 • Graft vs. host disease
Graft versus host disease (GvHD) is the most frequent complication after allogeneic haematopoietic stem cell transplantation (HSCT). GvHD can occur despite aggressive immunosuppressive prophylaxis even when the donor is a “perfectly” matched (HLA-identical) sibling. It is a consequence of interactions between antigen (Ag)presenting cells of the recipient and mature T-cells of the donor.
1. History When the first allografts were performed, it was shown that patients given marrow from donors other than monozygotic twins were likely to develop what was then called “secondary disease”. Clinical manifestations involved the skin, intestinal tract and liver. They were similar to those observed in mice neonatally transplanted with allogeneic spleen cells (runt disease) and in some immunodeficient children who had received blood transfusions. Patients given allo-HSCT fulfilled the three conditions necessary for the development of GvHD, as defined by Billingham in 1966 (1): • Administration of immunocompetent cells • Histo-incompatibility between donor and recipient • Inability of the recipient to destroy or inactivate the transfused or transplanted cells.
2. Pathophysiology A 3-step process reflects the current view of the development of acute GvHD (Table 1) (2). Phase 1: Effect of conditioning: Underlying malignancy, effects of previous therapies and conditioning-related tissue damage lead to generation of large amounts of inflammatory cytokines such as TNF-a and IL-1 that enhance early activation of host Ag-presenting cells (APCs). Translocation of lipopolysaccharide (LPS) across damaged intestinal mucosa activates the innate immune system and promotes the inflammatory cytokine cascade.
Table 1: Immunobiology of aGvHD as a 3-step process Phase
Cells
Cytokines, chemokines
1
Effect of conditioning
Host APCs (dendritic cells) Epithelial cell damage
TNF-a, IL-1 Adhesion molecules, LPS
2
T-cell activation
Donor T-cells (mainly CD4+) Host APCs
IL-2 IFN-g
3
Cellular and inflammatory effector phase
CTLs NK cells
“Cytokine storm” TNF-a / IL-1 / nitric oxide
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Phase 2: T-cell activation: In this milieu, transplanted naïve donor T lymphocytes (and other cellular compartments) interact with host APCs, are activated and express cytokines such as IFN-g, IL-2 and TNF-a (among others), leading to T-cell expansion. Phase 3: Cellular and inflammatory effector phase: This phase is a complex cascade of multiple effectors including cytotoxic effector cells (CTLs), using Fas- and perforin-mediated mechanisms, natural killer (NK) cells and large granular lymphocytes (LGLs), followed by the generation of inflammatory cytokines, such as TNF-a, IL-1 and macrophage-derived nitric oxide. Interactions of innate (LGL) and adaptive (alloreactive T-cells) immune responses lead to organ damage. Additional complexity has been added by the recent description of regulatory cell populations including regulatory T-cells (Treg), regulatory APC populations and perhaps mesenchymal stem cells (3).
3. Classification of GvHD The traditional definition of acute (aGvHD) or chronic GvHD (cGvHD) was based on the time of onset after transplantation (less or more than 100 days after HSCT). This distinction is no longer tenable. For instance, aGvHD may present beyond 3 months in patients who have received reduced-intensity conditioning (RIC), and symptoms characteristic of cGvHD may occur before D 100. Furthermore, some signs and symptoms are common to both cGvHD and aGvHD. The recent National Institutes of Health (NIH) Consensus Conference proposed a definition of aGvHD or cGvHD, each with 2 subcategories, based on the specificity of signs and symptoms rather than the criterion of time of onset (Table 2) (4).
4. Incidence and risk factors The median incidence of clinically significant (grade II-IV) aGvHD is about 40% but
Table 2: Definition of acute and chronic GvHD Category
Time of manifestation
aGvHD features
cGvHD features
Classic
£ 100 days
Yes
No
Persistent, recurrent, late onset
> 100 days
Yes
No
Classic
No time limit
No
Yes
Overlap syndrome
No time limit
Yes
Yes
aGvHD
cGvHD
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ranges from 10 to 80% according to a number of risk factors (Table 3). The risk of aGvHD increases with the use of unrelated donors, multiparous female donor, older age of the recipient, graft type (cord blood has a lower rate and peripheral blood stem cells have a higher rate than marrow), and certain conditioning regimens. Although the central role of HLA matching in HSCT is well established, it has become apparent that other genetic systems affect the development of GvHD. Minor histocompatibility antigens (mHA) are peptides from polymorphic cellular proteins, encoded by regions outside the major histocompatibility complex (MHC). Interestingly, some mHA are expressed mainly by malignant cells and represent attractive target for the induction of Graft-versus-Leukaemia (GvL) effect without promoting GvHD. Genes controlling inflammatory processes, such as cytokines, chemokines and their receptors, can modulate GvHD. Gene polymorphisms affecting IL-1, IL-6, IL-10, TNF, TGF-b, and IFN-g have all been implicated in the incidence and severity of GvHD, both in experimental models and immunogenetic analysis of retrospective clinical data (5). These findings suggest that, in the future, the genotyping of patients and/or donors with respect to a panel of cytokines, chemokines, pharmacogenes, etc. will possibly complement histocompatibility typing and increase our ability to predict the risk of transplant-related toxicity (6).
Table 3: Risk factors for the development of GvHD Donor
Recipient
HLA compatibility (related/unrelated) Sex mismatched (FÆM) Alloimmunisation (parity, transfusions) Source of SC (PBSC > BM > CB)
Age Conditioning regimen Prevention of GvHD
5. Histopathology Apoptosis of cells in the tissue layer responsible for proliferation and regeneration is a typical histologic feature of GvHD and is the final event of the allogeneic reaction (Phase 3: Cellular and inflammatory effector phase). The targets are epithelial cells, including basal and suprabasal cells of the epidermis, the intestinal epithelium and the biliary duct epithelium. In the three target organs, the characteristic lesion of GvHD is the same, with infiltrating immune cells in the vicinity of the apoptotic cell giving a feature classically termed “satellite cell necrosis”. 5.1. Skin biopsies Skin biopsies are widely used in the diagnosis of a GvHD. The dermoepidermal junction
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is most severely affected: There is epidermal and basal cell vacuolar degeneration, disorganisation of epidermal cell maturation, eosinophilic body formation and melanocyte incontinence. In the first 3 weeks after transplant, the microscopic lesions induced by the conditioning regimen, i.e. lymphocytic infiltrates and epithelial damage, can be very similar to early lesions of GvHD. When the histological lesions are mild, it can be useful to look for satellite cell necrosis by electron microscopy, or to use an anti-TNF antibody to stain activated intraepithelial lymphocytes. In very severe forms of aGvHD with epidermal necrolysis (Lyell syndrome) clinical diagnosis is obvious. Skin biopsy would show satellite cell necrosis of all the keratinocytes of the basal layer leading to the dissociation of the upper part of the epidermis from the basal layer. 5.2. Gut biopsies Gut biopsies are performed as a diagnostic procedure to explore gastro-intestinal (GI) tract symptoms, mainly nausea or diarrhoea. Rectal or colon biopsies have been generally replaced by gastro-duodenal biopsies. The target cells for GvHD are basal epithelial cells localised in the epithelium of the crypts, which undergo apoptosis in the close vicinity of intraepithelial lymphocytes. In the acute phase of GvHD, it is important to distinguish lesions of GvHD from those induced by virus, particularly CMV. Classically, epithelial cells infected by CMV have voluminous clear inclusions and become necrotic. However, in GI lesions of recent onset, viral inclusions can be difficult to see and, in some cases, CMV and GvHD can be associated. Therefore, the possibility of CMV infection should be excluded using antibodies directed against CMV early proteins. 5.3. Liver biopsies Liver biopsies are performed only to rule out viral infection or drug toxicity when isolated hepatic GvHD is suspected. The target is the epithelium of the biliary canal and the lymphocytic infiltrates are mainly localised in portal tracts. Foci of necrotic hepatocytes can be observed, but there is no sclerosis in the acute phase of GvHD. Hepatic small bile ducts show segmental disruption, injury to the periductular epithelium, bile duct atypia and cellular degeneration. Cholestasis may be present.
6. Acute GvHD 6.1. Clinical manifestations and grading 6.1.1. Skin A maculo-papular rash, often involving the palms and soles usually marks the onset 222
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of aGvHD. Lesions may be pruritic and/or painful. The rash then spreads and can involve the entire body surface. In more severe cases, bullae can form and surface areas can desquamate, leading to extremely painful denudation associated with protein loss. 6.1.2. Liver Liver involvement results in cholestatic hepatopathy, with or without jaundice, in which the cholestatic enzymes are substantially raised whilst the transaminases show only non-specific changes. The clinical diagnosis of aGvHD of the liver is difficult since distinguishing liver impairment due to therapy associated hepatotoxicity, infection, veno-occlusive disease or GvHD is not always possible. 6.1.3. Gastro-intestinal Involvement of the GI tract primarily manifests as nausea and green watery diarrhoea. The enteral fluid loss is used as a measure of gut involvement. Severe abdominal pain, bloody diarrhoea and massive enteral fluid losses accompany advanced disease. A variant of mild enteric GvHD involving only upper GI tract has been described. Symptoms include anorexia and nausea without diarrhoea and this usually responds well to immunosuppressive therapy. 6.1.4. Concomitant signs of acute GvHD These include fever, decrease in performance status and weight loss. Other tissues such as lymphoid organs, mucous membranes, conjunctivae, exocrine glands and the bronchi may also be involved, but these are not included in the clinical staging and grading established by Glucksberg and more recently by the International Bone Marrow Transplant Registry (IBMTR). 6.1.5. Grading In 1974, Glucksberg published the first aGvHD classification (7). Each organ is staged from 0 to 4 (Table 4). These stages are combined to calculate an overall grade including objective assessment of organ function and subjective assessment of performance status (Table 5). It is routine practice to dichotomise GvHD severity into clinically insignificant grades 0–I and clinically significant grades II–IV. In recognition of the system’s limitations, a Consensus Workshop was held in 1995 and a modified grading was system proposed (Table 6) (8). The latter system retained the objective organ staging criteria of the Glucksberg system but excluded the subjective criteria of clinical performance. Thus, a revised system was developed by the IBMTR (Table 7) (9). To compare prospectively the Glucksberg and IBMTR classifications, a prospective
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multicentre study was conducted in 607 patients receiving T-cell replete allografts in 18 transplantation centres (10). Patients were scored weekly for aGvHD. The 2 classifications performed similarly in explaining variability in survival by aGvHD score, although the Glucksberg classification predicted early survival better. There was less physician bias or error in assigning grades with the IBMTR scoring system. With either system, only the maximum observed grade had prognostic significance for survival.
Table 4: Grading system: Stage for each organ Stage
Skin / Maculo-papular rash
Liver / Bilirubin
GI / Diarrhoea
+
<25% of body surface
34-50 mmol/L
> 500 mL
++
25-50% of body surface
51-102 mmol/L
> 1000 mL
+++
Generalised erythroderma
103-255 mmol/L
> 1500 mL
++++
Generalised erythroderma with bullae formation and desquamation
> 255 mmol/L
Severe abdominal pain with or without ileus
Table 5: Overall grading system (Glucksberg) Grade of aGvHD
Degree of organ involvement
I
Skin: + to ++
II
Skin: + to +++ Gut and/or liver: + Mild decrease in clinical performance
III
Skin: ++ to +++ Gut and/or liver: ++ to +++ Marked decrease in clinical performance
IV
Skin: ++ to ++++ Gut and/or liver: ++ to ++++ Extreme decrease in clinical performance
Table 6: Consensus Conference on aGvHD grading
224
Grade
Skin
Liver
Gut
I
Stage 1–2
0
0
II
Stage 3 or
Stage 1 or
Stage 1
III
–
Stage 2–3 or
Stage 2–4
IV
Stage 4 or
Stage 4
–
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Table 7: IBMTR Severity Index for aGvHD Skin
Liver
GI
INDEX
Stage (max)
Extent of rash
Stage (max)
Bilirubin (µmol/L)
Stage (max)
Diarrhoea (mL/d)
A
1
<25%
0
<34
0
<500
B
2
25–50% or
1–2
34–102
1–2
550–1500
C
3
>50% or
3
103–255 or
3
>1500
D
4
Bullae or
4
>255 or
4
Pain, ileus
Thus, the authors failed to demonstrate a clear advantage of one system for grading aGvHD. 6.2. Prevention and treatment Acute GvHD is the major cause of early TRM. Mortality is due not only to GvHD itself but also to treatment related complications, both being responsible for a profound immune deficiency with frequent opportunistic infections. A major goal is prevention of GvHD. However, to date, it has not been possible to separate the deleterious effect of GvHD from the beneficial GvL effect. This explains why it is difficult to define the best strategy to prevent GvHD. Once GvHD occurs, the most important predictor of long-term survival is the primary response to therapy, as results with secondary treatments have been disappointing. 6.2.1. Prevention of GvHD a) Post transplant immunosuppressive treatment of the recipient (Table 8) Currently, most centres use a combination of a calcineurin inhibitor (cyclosporin (CsA) or tacrolimus) with “short course” methotrexate (MTX). Although other regimens are being explored, this standard regimen, described twenty years ago (11), has been shown repeatedly to result in a reasonable balance of GvHD and GvL in matched sibling transplants after ablative conditioning regimen. For higher risk groups or groups receiving non-conventional grafts (such as mismatched donors, older patients, reduced intensity regimen etc.), the best prophylaxis is less clearly established and other immunosuppressive drugs have been used: Sirolimus has been used in combination with tacrolimus and low dose (5 mg/m2) short MTX or with tacrolimus alone with prompt engraftment and minimal TRM (12). The efficacy of mycophenolate mofetil (MMF) associated with CsA has been studied mainly after RIC regimens (13). MMF may substitute MTX in the standard CsA combination because of less mucositis and overall good tolerance.
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Table 8: GvHD prophylaxis agents and mechanisms of action Mechanism of action
Dose
Cyclosporin (CsA)
Calcineurin inhibitor: blockade of T-cell activation
3 mg/kg IV
Tacrolimus
Calcineurin inhibitor: blockade of T-cell activation
0,02 mg/kg IV
Methotrexate (MTX)
Antimetabolite, folic acid analogue
15 mg/m2 D+1, 10 mg/m2 D+3, +6 and +11
Methylprednisolone (MP)
Receptor-mediated lympholysis + additional mechanisms
0,5–1 mg/kg
Mycophenolate mofetil (MMF)
Inhibition of DNA synthesis: lymphocyte apoptosis
1.5 to 3 g/d
Sirolimus (Rapamycine)
Macrolide antibiotic: blockade of T- and B-cell activation
12 mg D–3 then 4 mg/d
Antithymocyte globulin (ATG)
Rabbit or equine antibodies against human T-cells
2,5 mg/kg/d x 4
Alemtuzumab (Campath-1H)
Humanised monoclonal antibody to CD52
MethylPDN is the first line treatment for aGvHD, but its role in the prevention of GvHD is controversial. Several combination using prednisone in combination with MTX, CsA or both have been reported without substantial benefit. b) In vitro or in vivo T-cell depletion (TCD) of the transplant For patients receiving mismatched grafts, more intensive immunosuppression is usually needed. Methods of ex vivo TCD as well as pharmacologic in vivo TCD (anti-thymocyte globulin, alemtuzumab) have been used. In general, these methods reduce aGvHD but increase the incidence of infection (due to delayed reconstitution of the immune system) and the incidence of relapse (due to a decreased GvL effect). Several randomised or comparative studies have been performed comparing in vitro TCD to CsA + MTX, but so far, it has not been conclusively established whether TCD can improve LFS (14). 6.2.2. Treatment of aGvHD a) Primary treatment The most important predictor of long-term survival is the primary response of GvHD to therapy. Methyl-prednisolone (MP) at a dose of 2 mg/kg/d is the best initial therapy for aGvHD. This treatment, associated with a calcineurin inhibitor, is given for 7 to 14 days, and then tapered slowly if there is a complete response to therapy. 226
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Complete responses occur in 25 to 40% of patients with grade II to IV aGvHD. However, the likelihood of response decreases with increasing severity of the disease and the dynamic of responses may differ between target organs and between patients. Failure of therapy is usually defined as: • Progression after 3 days, or • No change after 7 days, or • Incomplete response after 14 days. b) Secondary treatment Patients in whom initial therapy has failed will receive a secondary treatment. Table 9 provides a summary of available agents. Numerous strategies have been used but only very few controlled studies have been conducted and there are no criteria to identify patients who are likely to respond to any second line treatment. Overall, results have been disappointing. The rate of partial and complete response to second line therapy varies from 35 to 70%, but the 6–12 months survival is low, in the order of 30%, in most large trials, because of high incidence of infectious complications or recurrence of GvHD. Treatment of steroid-refractory aGvHD has not made major advances in the last decade, as shown in several reviews recently published (15–17). In patients with predominant GvHD of the skin, which is refractory to steroids, the use of phototherapy may be an attractive and non-toxic strategy. In case of visceral
Table 9: Second line treatment of steroid-refractory aGvHD Methylprednisolone (2–5 mg/kg) Immunosuppressive drugs: - Tacrolimus, MMF, sirolimus (if not used for prophylaxis) Oral non-absorbable steroids (in case of GI involvement) Anti-thymocyte globulin Monoclonal antibodies: - Anti-IL-2 receptor (CD25) antibody: Inolinomab, basiliximab, daclizumab, denileukin difitox - Anti-TNFa antibody: Infliximab, etanercept - Anti-CD52 antibody (broad specificity T-cell antibody): Alemtuzumab (Campath 1H) - Anti-CD147 antibody (anti activated T- and B-cells): ABX/CBL - Anti-CD3 (broad specificity T-cell antibody): Visilizumab, OKT3 Pentostatin: - Inhibitor of adenosine-deaminase Extracorporeal photopheresis: - Suppression of T-cell reactivity and cytokine release; induction of regulatory T-cells Mesenchymal stem cells: - Immunomodulatory and tissue repairing effect
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GvHD, despite concerns about toxicity, second line treatment can be either polyclonal antibodies (i.e. ATG) or monoclonal antibodies. Other approaches, such as the use of mesenchymal stem cells, need to be explored in prospective randomised trials. There is agreement that efforts must be aimed at reducing steroid exposure and protecting the patient against infectious complications. c) Supportive care Supportive measures are very important. They include GI rest, parenteral hyperalimentation, replacement therapy of enteral losses of fluids, pain control, and infectious prophylaxis. Nevertheless, despite all preventive measures, viral, bacterial and mycotic infections are common and are the most frequent causes of death in patients with severe aGvHD. Ultimately, the outcome of aGvHD depends on two main factors: the overall grade of aGvHD and the response to treatment.
7. Chronic GvHD 7.1. Clinical manifestations and grading Chronic GvHD is the primary cause of late morbidity and non-relapse mortality in transplant survivors. Chronic GvHD has features resembling autoimmune and other immunologic disorders such as scleroderma, Sjögren syndrome, primary biliary cirrhosis, wasting syndrome, bronchiolitis obliterans (BO), immune cytopenias and chronic immunodeficiency. Symptoms usually present within 3 years after allogeneic HCT and are often preceded by a history of aGvHD. Manifestations of cGvHD may be restricted to a single organ or tissue or may be widespread. Chronic GvHD can lead to debilitating consequences, e.g. joint contractures, loss of sight, end-stage lung disease and profound chronic immunosuppression. Risk factors for cGvHD have been identified, including prior aGvHD, older patient age, the use of female donors for male recipients, use of donor lymphocyte infusion, use of unrelated or HLA-mismatched donors, and, more recently, the use of PBSC as a source of stem cells. 7.1. Diagnosis and staging The list of signs and symptoms of cGvHD, as established by the NIH working group report on diagnosis and staging (4), is shown in Table 10, with a distinction between diagnostic signs (that establish the diagnosis of cGvHD without the need for further testing or symptom), distinctive signs (not found in aGvHD but not sufficient to establish the diagnosis of cGvHD), other features of cGvHD (non-specific) 228
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Table 10: Signs and symptoms of cGvHD Organ/site
“Diagnostic”
Skin
Poikiloderma Depigmentation Lichen planuslike features
Nails
Other
Both acute and chronic Erythema, maculopapular rash, pruritus
Dystrophy
Scalp Mouth
“Distinctive”
Alopecia Lichen planus
Eyes
Xerostomia
Mucositis
Keratoconjunctivitis Photophobia, sicca blepharitis
Genitalia
Lichen planus
GI tract
Strictures of oesophagus
Exocrine pancreatic insufficiency
Liver
Anorexia, weight loss Bilirubin or alkaline phosphatase > 2 x upper limit of normal
Lung
Bronchiolitis obliterans
Muscles, fascia, joints
Fasciitis, joint contractures
Myositis
Cramps, arthralgias
Haematopoietic and immune
Thrombocytopenia eosinophilia, lymphopenia
Other
Ascites, pericardial or pleural effusions
and common signs or symptoms (found in both aGvHD and cGvHD). The diagnosis of cGvHD requires at least one diagnostic manifestation of cGvHD or at least one distinctive manifestation, with the diagnosis confirmed by pertinent biopsy or laboratory test. Historically, cGvHD was classified as limited or extensive on the basis of a small retrospective study (18). The NIH working group proposed a new global scoring system, which includes both the number of organs or sites involved and the severity within each affected organ: score 0 = no symptoms, score 1 = mild symptoms, score 2 = moderate symptoms and score 3 = severe symptoms. Mild cGvHD involves only 1 or 2 organs (except lung) with a maximum score of 1. Moderate cGvHD involves at least 1 organ with score 2, or 3 or more organs with score 1 (or lung score 1). Severe cGvHD indicates a score of 3 in any organ (or score 2 in the lung) (Table 11). HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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Table 11: Global scoring of cGvHD, according to the number of sites and the severity for each organ or site Number of organs
Mild cGvHD
Moderate cGvHD
Severe cGvHD
1 organ or site
Score 1
Score 2
Score 3
2 organs or sites
Score 1
Score 2
Score 3
3 or more organs
Score 1
Score 3
Lung involvement
Score 1
Score 2
7.3. Prevention and treatment Despite the fact that significant aGvHD is one of the strongest predictors for the development of cGvHD, successful efforts to reduce aGvHD with combinations of immunosuppressive agents, such as tacrolimus or MMF, have not resulted in a reduced incidence of cGvHD (19). Other methods, such as in vivo or in vitro TCD have been studied but the benefits due to a decreased incidence of acute and cGvHD need to be balanced against the consequences of delayed immune reconstitution. The role of prolonged administration of CsA for cGvHD prophylaxis has been studied and has shown only a non-significant reduction in the risk of extensive cGvHD with no difference in overall survival or disease free survival (20). 7.3.1. Primary treatment A combination of CsA and prednisolone has been the standard frontline therapy for cGvHD for almost 20 years (21). The initial dose of steroid ranges from 1 to 1.5 mg/kg/day for at least 2 weeks then the dose is slowly tapered, according to response. Duration of therapy is also determined by response, but is prolonged, usually continuing for close to 12 months, even in patients achieving complete resolution. The morbidity associated with steroid therapy is significant, including avascular necrosis, glucose intolerance requiring administration of insulin, infections, hypertension, changes in body habitus, cutaneous atrophy, cataracts, osteoporosis, emotional lability, interference with sleep, and growth retardation in children. So far, the addition of other immunosuppressive drug to standard upfront therapy, such as thalidomide, has not led to any significant difference in cGvHD response rate or survival (22). There is a need for randomised prospective trials to investigate the addition of other immunosuppressive drugs to upfront therapy in an effort to improve outcomes and ameliorate steroid-related side-effects. Such a trial, sponsored by the EBMT, has been activated in 2007 (double-blind placebo-controlled trial comparing CsA plus steroids with or without myfortic (the biologically active component of MMF) as primary treatment for extensive cGvHD). 230
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CHAPTER 11 • Graft vs. host disease
7.3.2. Salvage therapy There is no standard second-line therapy for patients who have failed primary treatment. Various approaches have been developed, including low dose total lymphoid irradiation, PUVA therapy, extracorporeal photochemotherapy, MMF, tacrolimus, and thalidomide. All these treatments have been reported to improve clinical manifestations (23). Recent publications have described encouraging responses following therapy with rituximab (anti-CD20 monoclonal antibody) (24). The efficacy of sirolimus (25) or of pentostatin (26) has also been investigated and some objective responses were described. 7.3.3. Antimicrobial prophylaxis and supportive care Infection is the leading cause of death among patients with cGvHD. Antimicrobial prophylaxis is a very important aspect of the treatment of these patients. Infections with Streptococcus pneumoniae and Haemophilus influenzae are frequent and must be prevented with prophylactic antibiotics and/or vaccination. Pneumocystis pneumonia occurring late after HSCT is strongly associated with cGvHD and prophylaxis must be given to patients receiving long-term immunosuppressive treatment. Additional treatments include ursodeoxycholic acid in patients with hepatic GvHD, anti-osteoporosis agents, and artificial tears in patients with eye involvement. A multidisciplinary approach to the management of patients with cGvHD is essential, including dermatologists, ophthalmologists, stomatologists, gynaecologists and others. In retrospective series of patients with extensive cGvHD, only 10 to 30% became long-term survivors. Late mortality is related not only to opportunistic infections but also to an increased incidence of secondary cancers.
8. Conclusion In conclusion, early institution of effective immunosuppressive therapy has changed the outcome for patients with GvHD, but larger studies demonstrating safety and efficacy of second line treatments for patients non-responsive to standard therapy with steroids are mandatory.
References 1. Billingham RE. The biology of graft versus host reactions. Harvey lectures 1966-1967; 62: 71-78. 2. Reddy P, Ferrara JLM. Immunobiology of acute graft-versus-host disease. Blood Reviews 2003; 17: 187-194. HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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3. Tse WT, Pendleton JD, Beyer Wm, et al. Suppression of allogeneic T-cell proliferation by human marrow stromal cells: Implication in transplantation. Transplantation 2003; 75: 389-397. 4. Filipovich AH, Weisdorf D, Pavletic S, et al. National Institutes of Health consensus development project on criteria for clinical trials in chronic Graft-versus-host Disease: I. Diagosis and Staging Working group report. Biol Blood Marrow Transplant 2005; 11: 945956. 5. Dickinson AM, Charron D. Non-HLA immunogenetics in hematopoietic stem cell transplantation. Curr opin Immunol 2005; 17: 517-525. 6. Baron C, Somogyi R, Greller LD, et al. Prediction of graft-versus-host disease in humans by donor gene-expression profiling. PLoS Med 2007; 4: e23. 7. Glucksberg H, Storb R, Fefer A, et al. Clinical manifestations of graft versus host disease in human recipients of marrow from HLA-matched sibling donors. Transplantation 1974; 18: 295-304. 8. Przepiorka D, Weisdorf D, Martin P, et al. Consensus conference on acute GvHD grading. Bone Marrow Transplant 1995; 15: 825-828. 9. Rowlings PA, Przepiorka D, Klein JP, et al. IBMTR severity index for grading acute GvHD: retrospective comparison with Glucksberg grade. Br J Haematol 1997; 97: 855-864. 10.Cahn JY, Klein JP, Lee SJ, et al. Prospective evaluation of 2 acute GvHD grading system: A joint Société française de Greffe de Moelle et Thérapie Cellulaire (SFGM-TC), Dana Farber Cancer institute (DFCI), and International Bone Marrow Transplant Registry (IBMTR) prospective study. Blood 2005; 106: 1495-1500. 11.Storb R, Deeg HJ, Whitehead J, et al. Methotrexate and cyclosporine compared with cyclosporine alone for prophylaxis of acute graft versus host disease after marrow transplantation for leukaemia. N Engl J Med 1986; 314: 729-735. 12.Cutler C. Li S, Ho VT, et al. Extended follow-up of methotrexate-free immunosuppression using sirolimus and tacrolimus in related and unrelated donor peripheral blood stem cell transplantation. Blood 2007; 109: 3108-3114. 13.Niederwieser D, Maris M, Shizuru JA, et al. Low-dose total body irradiation and fludarabine followed by hematopoietic cell transplantation from HLA-matched or mismatched unrelated donors and post grafting immunosuppression with cyclosporine and mycophenolate mofetil can induce durable complete chimerism and sustained remission in patients with haematological diseases. Blood 2003; 101: 1620-1629. 14.Wagner JE, Thompson JS, Carter S, et al. Effect of GvHD prophylaxis on 3-year DFS in recipients of unrelated donor bone marrow (T-cell Depletion Trial): A multi-centre, randomised phase II-III trial Lancet 2005; 366: 733-741. 15.Kim SS. Treatment options in steroid-refractory acute GvHD following hematopoietic stem cell transplantation. The Annals of Pharmacotherapy 2007; 41:1436-1444. 16.Bacigalupo A. Management of acute GVHD. Br J Haematol 2007; 137: 87-98. 17.Deeg JH. How I treat refractory acute GVHD. Blood 2007; 109: 4119-4126. 18.Shulmann H, Sullivan KM, Weiden PL, et al. Chronic graft-versus-host disease in man: A clinic pathologic study of 20 long term Seattle patients. Am J Med 1980; 69: 204-217. 19.Fraser CJ, Baker KS. The management and outcome of chronic graft-versus-host disease. Br J Haematol 2007; 138: 131-145. 232
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20.Kansu E, Gooley T, Flowers ME, et al. Administration of cyclosporine for 24 months compared with 6 months for prevention of chronic graft-versus-host disease: A prospective randomized clinical trial. Blood 2001; 98: 3868-3870. 21.Sullivan KM, Witherspoon RP, Storb R, et al. Alternating day cyclosporine and prednisone for treatment of high-risk chronic graft-versus-host disease. Blood 1988; 72: 555-561. 22.Arora M, Wagner JE, Davies SM, et al. Randomized clinical trial of thalidomide, cyclosporine and prednisone versus cyclosporine and prednisone as initial therapy for chronic GvHD. Biol Blood Marrow Transplant 2001; 7: 265-273. 23.Vogelsang GB. How I treat chronic graft-versus-host disease. Blood 2001; 97: 1196-1201. 24.Cutler C, Miklos D, Kim HT, et al. Rituximab for steroid-refractory chronic graft-versushost disease. Blood 2006; 108: 756-762. 25.Jurado M, Vallejo C, Perez-Simon JA, et al. Sirolimus as part of immunosuppressive therapy for refractory chronic graft-versus-host disease. Biol Blood Marrow Transplant 2007; 13: 701-706. 26.Jacobsohn DA, Chen AR, Zahurak M, et al. Phase II study of pentostatin in patients with corticosteroid-refractory chronic graft-versus-host disease. J Clin Oncol 2007; 25: 42554261.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The prognosis of aGvHD is mainly related to: a) The diagnosis of the initial disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The age of the donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The % of body surface involved . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The response of GvHD to initial therapy with 2 mg/kg methyl prednisolone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. One factor is not significantly associated with an increased risk of cGvHD: a) The use of a male donor for a female recipient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The previous occurrence of an aGvHD grade ≥ I . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The use of PBSC as a source of stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The age of the recipient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. T-cell depletion of the transplant is: a) Associated with a better long term survival in all patients when compared with a non T-cell depleted transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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b) Associated with an increased risk of cGvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) An effective prophylaxis of aGvHD but is associated with an increased risk of graft failure/relapse/opportunistic infections . . . . . . . . . . . . . . . . . . . . . . . d) Associated with an increased risk of secondary solid tumour . . . . . . . . . . . . . . 4. The “classical” prevention of GvHD is: a) A combination of methylprednisolone 1 mg/kg/d and a calcineurin inhibitor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A combination of a calcineurin inhibitor and short course methotrexate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Mycophenolate mofetil alone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) A course of ATG during 10 days . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Chronic GvHD is: a) Associated with an increased risk of relapse of the leukaemia . . . . . . . . . . . . b) Associated with an increased risk of secondary cancer . . . . . . . . . . . . . . . . . . . . . c) Associated with an increased risk of veno-occlusive disease . . . . . . . . . . . . . . d) Observed only after a grade II to IV aGvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 12
Long term survivorship, general health status, quality of life and late complications after HSCT
A. Tichelli, C-P. Schwarze, G. Socié
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CHAPTER 12 • Late effects
1. Introduction Large numbers of patients now survive long term following haematopoietic stem cell transplantation (HSCT). The aim of HSCT is not only to cure the patients from their primary disease, but also to allow the long-term survivor to obtain a normal health status, to return to work or to school and to have a normal social life. Immediate survival is therefore no longer the sole concern. Still, HSCT remains associated with considerable morbidity and mortality. Long-term health status and the development of late events related to HSCT have therefore gained increasing interest. Secondary malignant diseases are of particular clinical concern as more patients survive the early phase after transplantation and remain free of their original disease. Nonmalignant late effects are heterogeneous, and although often non-life threatening they significantly impair the quality of life of long-term survivors. This Chapter presents an overview of these malignant and non-malignant late complications, describes the consequences of these late events on the general health status, social integration and quality of life in long-term survivors, and provides recommendations regarding their prevention and early treatment (1, 2). Readers will find extensive literature summary and references in recently published reviews (3–5).
2. Non malignant late complications (Table 1) (2) 2.1. Late ocular effects 2.1.1. Ocular complications of the posterior segment These can be divided into microvascular retinopathy, optic disk oedema, haemorrhagic complications and infectious retinitis. Fungal infections typically occur within 120 days of HSCT, while Herpes zoster, CMV and Toxoplasma retinitis occur later. Ischaemic retinopathy with cotton-wool spots and optic-disk oedema has been described in 10% of patients following HSCT. Microvascular retinopathy occurs mainly after TBI conditioned allogeneic HSCT in patients receiving cyclosporin as GvHD prophylaxis. In most cases, the retinal lesions resolve with withdrawal or reduction of immunosuppressive therapy. 2.1.2. Ocular complications of the anterior segment The two most common late complications affecting the anterior segment are cataract formation and kerato-conjunctivitis sicca syndrome. Posterior subcapsular cataracts have long been recognised in recipients of HSCT as one of most frequent late complications of TBI. After single dose TBI almost all patients develop cataracts within 3 to 4 years and most if not all require surgical repair. Although the probability of developing cataracts after fractionated TBI is
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around 30% at 3 years the incidence may be more than 80% 6 to 10 years post HSCT (6, 7). Use of TBI in general and, single fractionations in particular and the use of
Table 1: Non-malignant late tissue and organ toxicity in long-term survivors Organ
Clinical manifestation
Risk factors
Monitoring
Intervention
Eye
Cataracts
Radiation Steroids
Split lamp examination
Fractionation of TBI Surgical repair
Keratoconjunctivitis Radiation GvHD
Schirmer test
Treatment of GvHD Topical lubricants Topical steroids
Restrictive or dilated Anthracyclines cardiomyopathy Mediastinal Arrhythmia radiotherapy Autonomic neuropathy
LVEF 24-hour ECG
Treatment of cardiac insufficiency Pace maker
Infection GvHD Smoking
Pulmonary function testing Chest radiographic testing if clinically indicated
Treatment of infection Immune globulin replacement Treatment of GvHD
Restrictive lung disease
Radiation Chemotherapy Infection
Pulmonary function testing Chest radiographic testing if clinically indicated
Fractionation of radiation Lung shielding Treatment of infection Steroids
Chronic hepatitis C
HVC infection Iron overload
Liver tests Hepatitis serologies Viral load if positive
Treatment of hepatitis C Treatment of iron overload (phlebotomy or chelation therapy)
Liver cirrhosis
HCV infection Iron overload
Ferritin
Heart
Respira- Chronic obstructive tory tract lung disease Bronchiolitis obliterans
Liver
Chronic GvHD of the liver Kidney
Nephropathy
TBI Chemotherapy (platinum) Cyclosporin
Liver biopsy if indicated
Treatment of GvHD
Renal function tests
Control of hypertension
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Organ
Clinical manifestation
Skeletal
Monitoring
Intervention
Avascular necrosis Steroids of the bone Radiation
Radiographic testing
Avoidance of long term treatment with steroids Symptomatic relief of pain Orthopoedic measures Surgical repair
Osteoporosis
Dual photon densitometry
Sex hormone replacement Treatment of osteoporosis (bisphosphonates, calcium, vitamin D)
Chronic stomatitis Chronic GvHD Radiation
Oral inspection
Oral hygiene Treatment of GvHD
Dental late effects
Radiation Chronic GvHD
Dental inspection
Prophylaxis: oral hygiene, instruction for dental care, brushing teeth, application of fluoride Treatment of caries
Thyroid gland
Hypothyroidism
Radiation
TSH, T4 annually
Thyroid hormone replacement
Gonadal function
Gonadal failure
Radiation Chemotherapy
FSH, LH, testosterone (males), oestradiol (females)
Hormone replacement Sperm banking in males
Fertility
Infertility
Radiation Chemotherapy
FSH, LH, testosterone (males), oestradiol (females) Sperm analysis
Cryopreservation of sperm fluid before HSCT
Nervous system
Leukoencephalopathy
Cranial radiation Intrathecal chemotherapy
Evaluation according to symptoms
Peripheral neuropathy
Chemotherapy GvHD
Atherosclerosis Cerebrovascular events Cardiovascular events Peripheral vascular events
Established Cardiovascular risk cardiovascular risk factors factors GvHD (?) Radiation (?)
Oral
Vascular compartment
Risk factors
Steroids, cyclosporin, tacrolimus Hypogonadism, TBI, chemotherapy Immobility
Correction of cardiovascular risk factors
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steroid treatment for longer than 3 months are associated with a significant risk of cataract development. The development of cataracts is also more likely if the TBI is delivered at a high dose rate. Finally, in prospective studies of the incidence of cataracts according to various risk factors, patients who receive cyclophosphamide and TBI (Cy/TBI) have a higher incidence of cataract formation than those treated with busulfan and cyclophosphamide (BuCy). The only treatment for cataract is to surgically remove the opacified lens from the eye to restore transparency of the visual axis. Today, cataract surgery is a low risk procedure and improves visual acuity in 95% of eyes that have no other pathology. Keratoconjunctivitis sicca syndrome is usually part of a more general syndrome with xerostomia, vaginitis and dryness of the skin. All these manifestations are closely related to cGvHD, which may lead in its most extensive forms to a Sjögren like syndrome. The ocular manifestations include reduced tear flow, keratoconjunctivitis sicca, sterile conjunctivitis, corneal epithelial defects, and corneal ulceration. The incidence of late-onset keratoconjunctivitis sicca syndrome may reach 20% fifteen years after HSCT, but reaches nearly 40% in patient with cGvHD, compared to less 10% in those without GvHD. Risk factors for late-onset keratoconjunctivitis include cGvHD, female sex, age >20 years at HSCT, single dose TBI, and the use of methotrexate for GvHD prophylaxis. Treatment is based on the management of cGvHD with regular use of topical lubricants. Topical corticosteroids may improve symptoms but can cause sight-threatening complications if used inappropriately in Herpes simplex virus or bacterial keratitis. 2.2. Pulmonary late effects 2.2.1. Restrictive lung disease Restrictive lung disease is frequently observed 3 to 6 months after HSCT in patients conditioned with TBI and/or receiving an allogeneic HSCT but in most cases it is not symptomatic. Restrictive disease is often stable and may recover, partially or completely, within 2 years. However, some patients do develop severe late restrictive defects and may eventually die from respiratory failure. 2.2.2. Chronic obstructive lung disease Chronic obstructive pulmonary disease can be detected in up to 20% of long-term survivors after HSCT. It has been mainly associated with cGvHD, but other potential risk factors including TBI, hypogammaglobulinemia, GvHD prophylaxis with methotrexate, and infections have been described. Mortality is high among these patients, particularly in those with an earlier onset and rapid decline of FEV1. Symptoms consist of nonproductive cough, wheezing and dyspnoea; chest radiography 240
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is normal in most cases. High-resolution CT scanning may reveal non-specific abnormalities. Symptomatic relief can be obtained in some patients with bronchodilators; however in most cases obstructive abnormalities are not improved by this treatment. Patients with low IgG and IgA levels should receive immunoglobulins to prevent infections, which may further damage the airways. Immunosuppressive therapy may be of benefit but typically improvements occur in less than 50% of cases. Asymptomatic patients with abnormal pulmonary function tests (PFTs) should be closely monitored for the development of respiratory symptoms; an early recognition of airflow obstruction allows the initiation of treatment at a potentially reversible stage. Obliterative bronchiolitis (OB), the best characterised obstructive syndrome, has been reported in 2 to 14% of allogeneic HSCT recipients and carries a mortality rate of 50%. OB is strongly associated with cGvHD and low levels of immunoglobulins. Initial symptoms often resemble those of recurrent upper respiratory tract infections, and then persistent cough, wheezing, inspiratory rales and dyspnoea appear. PFTs gradually deteriorate with severe and non-reversible obstructive abnormalities. Chest radiographs and CT scanning may reveal hyperinflation with or without infiltrates and vascular attenuation; however radiological findings do not correlate with lung function changes. Bronchoscopy with transbronchial biopsy can help to rule out infection and may reveal obliteration of bronchioles with granulation tissue, mononuclear cell infiltration or fibrosis. It is not clear to what extent combined immunosuppressive treatment can be effective in the treatment of this disease, which typically does not respond to treatment with steroids. Azathioprine and mycophenolate mofetil may lead to improved symptoms in some cases. Prophylaxis and prompt treatment of infections are the most important elements of clinical management and may help to alter the clinical course of the disease. 2.3. Late complications of bones and joints 2.3.1. Avascular necrosis of bone (AVN) The incidence of AVN varies from 4% to more than 10%. The mean time from transplant to AVN is 18 months and the first clinical manifestation is usually pain. Early diagnosis can rarely be made using standard radiography alone and magnetic resonance imaging is the investigation of choice. The hip is the affected site in over 80% of cases with bilateral involvement occurring in more than 60% cases. Other sites include the knee (10% of patients with AVN), the wrist and the ankle. Symptomatic relief of pain and orthopoedic measures to decrease the pressure on the affected joints are of value, but most adult patients with advanced damage will require surgery. The probability of total hip replacement following a diagnosis of
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AVN is approximately 80% at 5 years. While short-term results of joint surgery are excellent in the majority (>85%) of cases, it is clear that long term follow-up of the prostheses are needed in young patients who have a long life expectancy. Studies evaluating risk factors for AVN have clearly identified the use of steroids (both total dose and duration) as the strongest risk factor. Thus, unnecessary long-term low dose steroids for non-active chronic GvHD should be avoided. The second major risk factor for AVN is TBI, the highest risks being associated with receipt of single doses of 10 Gy or higher or >12 Gray in fractionated doses. 2.3.2. Osteoporosis (8) The degree of reduction in bone mass can be quantified on dual photon densitometry. The cumulative dose and number of days of glucocorticoid therapy and the number of days of cyclosporine or tacrolimus therapy showed significant associations with loss of bone mass. Non-traumatic fractures may occur in 10% of patients. Using WHO criteria, nearly 50% of the patients have low bone density, a third have osteopenia and roughly 10% have osteoporosis, 12–18 months post transplant. Preventative measures of osteoporosis must include sex-hormone replacement in patients with gonadal failure; the efficacy of new treatments for osteoporosis in long-term survivors of HSCT requires further evaluation. 2.4. Endocrine function after HSCT 2.4.1. Thyroid dysfunction Seven to 15.5% of patients will develop sub-clinical hypothyroidism (slightly-high serum TSH and normal free-T4 levels) in the first year post HSCT. It is not yet clear if patients who develop sub-clinical hypothyroidism should be treated with Lthyroxine since the majority of these cases are mild, compensated and may resolve spontaneously. One possible approach is to monitor TSH and free-T4 levels twice yearly and to consider L-thyroxine treatment only if the TSH concentration remains high or is increasing. The frequency of hypothyroidism requiring L-thyroxine replacement therapy is highly variable depending to a large extent on the type of pre-transplant conditioning applied: nearly 90% in patients who have received 10 Gy single-dose TBI, 14–15% of patients following fractionated TBI, and smaller numbers after conditioning with BuCy. Treatment with L-thyroxine is indicated in all cases of frank hypothyroidism (elevated TSH with low free-T4 blood levels). Thyroid hormone levels should be measured 4–6 weeks after commencement of replacement therapy, and dosage should be tailored thereafter to the individual patient and adjusted according to thyroid function evaluation every 6 months. Elderly patients 242
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should have an ECG prior to commencing treatment to exclude associated ischaemic heart disease and/or arrhythmias. Autoimmune thyroiditis, presumably transferred via donor cells, has also been reported. 2.4.2. Gonadal failure Gonadal failure (both testicular and ovarian) is a common long-term consequence of the chemotherapy given prior to HSCT, and of the pre-transplant conditioning. The major cause of gonadal damage leading to hypergonadotrophic-hypogonadism is irradiation. Similar damage can also be caused by busulfan. In males, the testicular germinal epithelium (Sertoli-cells), the site of spermatogenesis, is more vulnerable to radiation and chemotherapy than the testicular Leydig-cell component, which is involved testosterone secretion. Therefore, testosterone levels are usually normal even where spermatogenesis is reduced or absent. The serum FSH is typically elevated while LH levels may remain in the normal range. The great majority of patients will not, therefore, require testosterone replacement to ensure sexual activity, libido, erection and ejaculation. Sex-hormone replacement therapy (SHRT) with testosterone derivatives in males is indicated in patients with severe uncompensated hypogonadism. In females, hypergonadotrophic-hypogonadism is almost inevitable as the ovaries are more vulnerable to irradiation and chemotherapy than the testes. Busulfan is one of the most gonadotoxic agents while cyclophosphamide is usually associated with only minor effects on gonadal function. The majority of adult females will need SHRT in order to maintain menstruation and bone turnover/mineralisation. In prepubertal girls who do not undergo puberty spontaneously post-HSCT, oestrogen treatment should be started at the age of 12–13 years to promote breast and uterine development and the pubertal growth spurt. The dose of oestrogen treatment will need to be gradually increased and a combination of cyclical oestro-progesterone treatment introduced after 1–2 years to initiate menstruation and to reduce the risk of future osteoporosis. SHRT can be interrupted once every 2–3 years, for a period of six months, to evaluate possible spontaneous recovery of ovarian activity, which occurs in the minority of women. Due to the high incidence of gonadal dysfunction and early menopause in patients after HSCT, an annual clinical and biological gynaecological assessment is recommended. 2.4.3. Special considerations in puberty Hypogonadism occurs in up to 70% of paediatric patients after HSCT. Male patients are more likely than females to enter and progress through puberty. During spontaneous puberty, measurement of testosterone is recommended if the pubertal growth spurt is blunted. A high percentage of female patients will need SHRT. The
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probability of ovarian recovery after fractionated TBI is higher in younger patients but after busulfan hypogonadism prevails. Baseline measurements of LH and FSH are recommended at 8 yrs in females and 9 yrs in males. The timing and progression of puberty have a major influence on growth and final height. This must be considered when initiating replacement therapy. Delaying puberty beyond the ages of 13 and 14 yrs is usually not advisable. Temporary cessation of replacement therapy after completion of puberty and growth should be considered in order to evaluate spontaneous recovery. Precocious puberty occurs in a few cases and necessitates gonadotrophin releasing hormone (GnRH)-agonist treatment. Central/hypogonadotrophic hypogonadism can be diagnosed by a GnRH test. 2.4.4. Fertility following stem cell transplantation The overall incidence of pregnancy is low (<2%) except in patients transplanted for SAA. However, not all patients wish to become parents after HSCT. Some have completed their family prior to transplantation, the proportion of long-term survivors living with a fixed partner is significantly lower compared with siblings and finally, the diagnosis of a potentially life threatening illness makes patients and their partners reluctant to parent children (9). Fertility following HSCT for non-malignant disease Return of gonadal function following cyclophosphamide conditioning without TBI for SAA is reported in half of adult female survivors and roughly 25% subsequently conceived. In adult male survivors 60% have return of sperm production and a quarter subsequently fathered children. Gonadal recovery is usual in women less than 25 years at the time of transplant but sharply decreases thereafter. In thalassaemia, gonadal failure is common as a result of both transfusional hemosiderosis and conditioning with BuCy and pregnancies are very rare. Fertility following HSCT for malignant disease The majority of patients given TBI conditioning experience gonadal failure. Recovery of gonadal function occurs in 10% of the women and the incidence of pregnancy is less than 3% (9, 10). In men, recovery of gonadal function has been reported in less than 20% patients and use of increasing doses of TBI may be associated with considerably lower recovery. Parenting a child following administration of TBI is a rare event in men. However, with increasing follow-up, the impact of TBI on gonadal damage after allogeneic transplantation becomes more attenuated. In a recent study on male fertility, younger age at transplantation, a longer interval between HSCT and the seminal fluid analysis and the absence of cGvHD were associated with a higher incidence of spermatogenesis production (11). BuCy is also associated with a high incidence of gonadal failure in women and there have been no pregnancies reported using BuCy for patients with leukaemia. In men, this 244
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preparative regimen appears to be associated with return of gonadal function in approximately 15% cases. Few patients have subsequently fathered children naturally; 20 such patients were identified by the EBMT Late Effects Working Party (LEWP). Fewer studies have evaluated gonadal function following autologous HSCT and most of these have involved small numbers of patients. The use of BEAM conditioning for autografts may be associated with a high incidence of gonadal recovery in women but in men azoospermia is almost always the rule. 2.4.5. Pre-transplant counselling and treatment options The pre-transplant counselling process should include information about the chance of gonadal failure and should assess the relevance of this to the patient. This should include discussion of assisted conception techniques and in women management of a premature menopause. In women with no residual ovarian function following HSCT, implantation of embryos cryopreserved prior to HSCT is currently the only option for parenting her own genetic child. This, however, requires that prior to HSCT the underlying disease can tolerate a minimum of 4–6 weeks delay in treatment so as to undertake controlled stimulation of the ovary and egg collection. A sperm donor must also be available. In situations where treatment cannot be delayed or there is no sperm donor, consideration can be given to freezing ovarian tissue prior to HSCT. Centres in some countries will provide this service for young women. However the patients must be aware that currently this is a research rather than a clinical option as to date there have been no successful pregnancies in human subjects using this methodology. Sperm cryopreserved prior to HSCT may be used post HSCT for artificial insemination, in vitro fertilisation and embryo transfer or for in vitro injection into the cytoplasm of the oocyte. Post-transplant management should routinely include symptomatic and biochemical monitoring of gonadal function. Distressing vasomotor symptoms may commence acutely post HSCT but this may be prevented by initiating SHRT. While the patient should be prepared for infertility a possible need for contraception soon after HSCT must also be emphasised, particularly in women who resume menstruation or in patients who do not wish to become parents. Patients have conceived within 6 months of HSCT and unexpected pregnancy may result in requests for termination. 2.4.6. Growth With respect to failure of growth after HSCT, it has been difficult to separate the adverse contributions of the numerous contributory factors. Chemotherapy has an additive negative effect when combined with RT. In general, chemotherapy with Bu/Cy alone does not seem to cause growth impairment but there have been reports of growth hormone deficiency (GHD) after such a combination. In contrast, patients
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with neuroblastoma generally do not reach their target height. Cranial (CI), craniospinal (CSI) and TBI can result in growth failure. Single dose TBI (7–8 Gy) causes a loss in final height of –2.0 SD (standard deviations) for boys and –1.0 SD for girls. Much of this loss occurs during puberty. Fractionated TBI (12 Gy) does not cause substantial loss of height with final heights being above –2.0 SD. However, these data were derived from patients who were a median of 10.8 yrs old at the time of transplant, whereas in children younger than 6 yrs the final height was markedly reduced at –3.5 SD. CI prior to HSCT and TBI aggravates the negative effect of TBI. Growth is also disproportionate, with a reduced growth of the spine, which is most severe in CSI. The incidence of GHD following TBI and high dose Cy occurs in 20–70% of children. The diagnosis of GHD may be difficult to establish and repetitive testing may be necessary, as the GH response to stimuli may be discordant. Data on spontaneous GH secretion are scarce. IGF-1 (insulin growth factor 1) and IGF BP-3 (IGF binding protein 3) are unreliable markers for GHD. GH replacement may be started 2 yrs after HSCT on an individual basis.
3. Malignant late complications 3.1. Introduction It has been usual to divide the problem of secondary malignancies following haematopoietic stem cell transplantation into three groups, i.e. leukaemia, lymphoma and solid tumours. The occurrence of second malignancies after HSCT has been extensively reviewed, in particular the critical problem of myelodyplastic syndrome (MDS) and acute leukaemia following autologous HSCT, and rare occurrence of leukaemia in donor cells after allogeneic HSCT (not reviewed in this Chapter; (see ref. 3 and 12 for additional reading and references). The main risk factors are summarised in Table 1. 3.2. Post-transplant lymphoproliferative disorders (PTLD) (13) Most cases of lymphoproliferative disorders after HSCT have been observed in allogeneic transplant recipients. B-cell PTLD are clinically and morphologically heterogeneous, and are usually associated with T-cell dysfunction and the presence of EBV. The mean interval from transplantation to the development of B-cell PTLD is between 5 to 6 months. In the largest series involving 18,014 patients who underwent allogeneic bone marrow transplantation at 235 centres worldwide, PTLD developed in 78 recipients with 64 cases occurring less than a year after transplantation (9). The cumulative incidence of PTLD was 1.0±0.3% at 10 years. The incidence was highest 1–5 months post-transplant (120 cases/10,000/year) 246
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followed by a steep decline to less than 5/10,000/yr among 1 year-plus survivors. B-cell PTLD are almost always of donor origin, and are associated with EBV-genomic DNA integration. Biopsies reveal monomorphic or polymorphic, diffuse large-cell lymphoma of B-cell origin. The most frequent presenting features of PTLD are fever, with or without nodal and extra-haematopoietic organ involvement. Today, the availability of quantitative polymerase chain reaction (PCR) for EBV DNA has dramatically changed the ability to make an early and accurate diagnosis. Using EBV viral load, patients are now frequently diagnosed with “PTLD” at the time of isolated fever with (or even without) low tumour burden and a monoclonal gamma-globulin. Quantitative PCR techniques allow frequent monitoring of the DNA load to predict the development of PTLD. The presence of EBV-specific T-lymphocytes can also be monitored using tetramer technology, which allows detection of minute numbers of antigen-specific T-cells. The risk of early-onset PTLD (less than a 1 year) is strongly associated with unrelated or HLA mismatched related donors, T-cell depletion of donor marrow or use of anti-thymocyte globulin for prophylaxis or treatment of aGvHD. Methods of T-cell depletion that selectively target T-cells or T+NK cells are associated with markedly higher risks of PTLD than methods that removed both T and B-cells, such as the Campath-1 monoclonal antibody or elutriation. Most recently the efficacy of anti-CD20 monoclonal antibody (rituximab) in the treatment of PTLD after HSCT has been reported. Currently many groups consider pre-emptive treatment based on increased EBV load in high-risk patients. Whether this approach is safe and efficacious requires further longitudinal multi-centre studies. The use of cellular therapy has also been advocated in this setting. However, it may induce GvHD if non-EBV-specific CTL are used and still requires high-level biotechnology laboratories to provide either EBV-specific CTL clones or T-lymphocytes transduced with suicide genes that can be activated in the event of any harmful effects. 3.3. Solid tumours The median elapsed time from HSCT to the presentation with a solid tumour is between 5 to 6 years. The cumulative incidence of invasive solid cancers is in the order of 8% at 20 years. The risk increases with time since transplantation with no evidence of a plateau. In a collaborative study, Curtis et al. (14) analysed the results from 19,220 patients transplanted between 1964 and 1992. There were 80 solid tumours giving an observed/expected ratio of 2.7. In patients surviving at least 10 years after transplantation, the risk was increased 8 fold. The risk was increased significantly for melanoma, cancers of the buccal cavity, liver, CNS, thyroid, bone,
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and connective tissue. The risk was highest for the youngest patients and declined with age. Most striking was the link of squamous cell carcinoma with cGvHD and male gender. The underlying diagnosis was important in so far as the risk of solid tumours was higher for patients with leukaemia and lower in patients with lymphoma or aplastic anaemia. The risk associated with TBI declined if irradiation was given with a fractionation regimen but increased with the total cumulative dose. This analysis strongly suggests that reduced doses of TBI, the omission of limited field irradiation, and the prevention of GvHD, in particular cGvHD, should reduce the risk of post-transplant solid tumours. The main features of solid tumours occurring after HSCT include: - The incidence of the second malignancy continues to increase with prolonged follow-up without any evidence of a plateau - Irradiation is the strongest risk factor but recent data suggest that solid cancers can also occur after non-irradiation based conditioning regimens - Chronic GvHD and/or its treatment is strongly associated with the occurrence of squamous cell carcinoma - Studies to try to identify cancer-specific risk factors are on-going (see Table 2) - Biological data are required to investigate the oncogenic process in recipients of HSCT - Most patients who have undergone allogeneic HSCT had already received substantial amounts of chemotherapy and radiation before transplant. The role of those pre-transplant treatments on the occurrence of solid tumours clearly needs evaluation. The treatment of patients with second solid tumours is unclear. There are few data to answer this critical and practical question. Patients with SCC of the head and neck
Table 2: Risk factors for second malignancies according to tumour type
248
Risk factor
Second malignancies
TBI
Melanoma, thyroid, CNS tumour
Limited field irradiation
SCC, head and neck
T-cell depletion
Melanoma, PTLD
Chronic GvHD
SCC, head and neck, skin
HLA-mismatch
PTLD
ATG, OKT3
PTLD
Acute GvHD
PTLD
Oncogenic viruses
PTLD, SCC head and neck
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have a bad prognosis using conventional therapy with surgery and/or radiotherapy. With this exception patients with solid tumours can generally be treated safely with conventional treatment.
4. Health status, social integration and quality of life after HSCT Patients who survive more than 5 years after HSCT are generally in good health and are socially reintegrated (15). Most of them regain normal or minimally impaired clinical performance and full social activity in work and school. The clinical performance status of more than 90% of patients is compatible with normal activity (Karnofsky scores of 90 to 100%). More than 90% of patients return to full-time work or school. However, the risks for illness and death are increased in long-term survivors. Patients who survive disease-free for 2 years after allogeneic HSCT have a 10-fold increased risk of mortality compared with an age-matched general population. The mortality continues to be significantly increased even for those who have survived 15 years after transplantation. The major causes of death are late recurrence of the original malignant disease, cGvHD, late infections without GvHD, secondary malignancies, pulmonary complications and cardiac complications. Despite the fact that functional well-being is good, there are still considerable differences compared to their sibling donors. As compared with sibling donors, fewer long-term survivors are currently married, and they have more difficulty holding jobs and obtaining health/life insurance (16). Quality of life (QoL) refers to every dimension of life except for its length, and includes physical abilities, symptoms, social well-being, psycho-emotional status, and spiritual/existential qualities. It reflects how well people feel, what they can accomplish, how satisfied they are with their lives, and whether their lives have meaning and purpose. Following HSCT, QoL can range from perfect, with no physical, emotional or social sequelae and a greater appreciation for life, to severely compromised with physical disability, pain and psychological despair. However, although long-term survivors report many specific symptoms and limitations in their daily activities, almost all patients indicate they would undergo the procedure again given similar circumstances. Fatigue and sleep disturbances are the most common complains for recipients of both, autologous and allogeneic grafts. Many survivors continue to experience the negative effects of the cancer diagnosis and its treatment on their daily lives, resulting in decreases in their QoL well beyond the completion of therapy. However, positive coping strategies and enhanced quality of life in long-term survivors of cancer have been described. Positive psychological effects of the cancer and its treatment include feelings of being grateful and fortunate to be alive, an enhanced appreciation of life, and increased self-esteem
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and sense of mastery. Documented negative psychosocial effects include concerns about the future, heightened sense of vulnerability (my body has let me down), a sense of loss for what might have been (e.g., loss of fertility) and increased health worries or hypervigilance. Large HSCT survivor studies conclude that less than half of survivors report normal functional status in most domains. Many specific physical symptoms and limitations persist, particularly for recipients of allogeneic transplant. Continued improvement over time with re-adaptation and return to some form of normalcy has been observed. Within the 3 first years of HSCT the quality of life is significantly worse compared to the general population, while those surviving in excess of 3 years form transplant claim improvements particularly in social functioning, mental health and vitality (17). However, many long-term survivors still have residual deficits, and approximately 5% of very long-term survivors rate their health as poor. Long-term HSCT survivors report poorer physical, psychological and social functioning, but conversely, more psychological and interpersonal growth, differences that appeared to persist years after transplantation (18). In a prospective, longitudinal cohort study, function was assessed from immediately post transplantation to 5-years later. Physical recovery occurred earlier than either psychological well-being or return to work. The proportion without limitations increased over time. Patients with cGvHD, with less social support before transplantation and women were more depressed after HSCT. Full recovery was a process demanding 3 to 5 years. It is possible that this might be accelerated by effective interventions. At a minimum, screening for depression is recommended. Clinical assessment for psychological symptoms should be maintained annually, with mental health professional counselling for those with recognised deficits. Attention should also be given to the partners of patients, who often report loneliness and to the children, who may suffer from separation from one of the parents.
5. Conclusions Individuals undergoing allogeneic HSCT, even when cured, will never become nonpatients. Allogeneic HSCT is a lifelong commitment for all – the patient, his or her family, the primary care physician, the transplantation team, and the healthcare providers – and structures have to be created to assure long-term follow-up of survivors after HSCT.
References 1. Rizzo JD, Wingard JR, Tichelli A, et al. Recommended screening and preventive practices for long-term survivors after hematopoietic cell transplantation: Joint recommendations of the European Group for Blood and Marrow Transplantation, Center for International Blood 250
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and Marrow Transplant Research, and the American Society for Blood and Marrow Transplantation (EBMT/CIBMTR/ASBMT). Bone Marrow Transplant 2006; 37: 249-261. 2. Rizzo JD, Wingard JR, Tichelli A, et al. Recommended screening and preventive practices for long-term survivors after hematopoietic cell transplantation: Joint recommendations of the European Group for Blood and Marrow Transplantation, the Center for International Blood and Marrow Transplant Research, and the American Society of Blood and Marrow Transplantation. Biol. Blood Marrow Transplant 2006; 12: 138-151. 3. Ades L, Guardiola P, Socié G. Second malignancies after allogeneic hematopoietic stem cell transplantation: New insight and current problems. Blood Rev 2002; 16: 135-146. 4. Socie G, Salooja N, Cohen A, et al. Nonmalignant late effects after allogeneic stem cell transplantation. Blood 2003; 101: 3373-3385. 5. Tichelli A, Socié G. Considerations for adult cancer survivors. Hematology (Am. Soc. Hematol. Educ. Program.) 2005; 22: 516-522. 6. Benyunes MC, Sullivan KM, Deeg HJ, et al. Cataracts after bone marrow transplantation: Long-term follow-up of adults treated with fractionated total body irradiation. Int J Radiat Oncol Biol Phys 1995; 32 :661-670. 7. Tichelli A, Gratwohl A, Egger T, et al. Cataract formation after bone marrow transplantation. Ann Intern Med 1993; 119: 1175-1180. 8. Schimmer AD, Minden MD, Keating A. Osteoporosis after blood and marrow transplantation: clinical aspects. Biol Blood Marrow Transplant 2000; 6: 175-181. 9. Salooja N, Szydlo RM, Socié G, et al. Pregnancy outcomes after peripheral blood or bone marrow transplantation: A retrospective survey. Lancet 2001; 358: 271-276. 10.Sanders JE, Hawley J, Levy W, et al. Pregnancies following high-dose cyclophosphamide with or without high-dose busulfan or total-body irradiation and bone marrow transplantation. Blood 1996; 87: 3045-3052. 11. Rovo A, Tichelli A, Passweg JR, et al. Spermatogenesis in long-term survivors after allogeneic hematopoietic stem cell transplantation is associated with age, time interval since transplantation, and apparently absence of chronic GvHD. Blood 2006; 108: 1100-1105. 12.Deeg HJ, Socié G. Malignancies after hematopoietic stem cell transplantation: Many questions, some answers. Blood 1998; 91: 1833-1844. 13.Curtis RE, Travis LB, Rowlings PA, et al. Risk of lymphoproliferative disorders after bone marrow transplantation: A multi-institutional study. Blood 1999; 94: 2208-2216. 14.Curtis RE, Rowlings PA, Deeg HJ, et al. Solid cancers after bone marrow transplantation. N Engl J Med 1997; 336: 897-904. 15 Duell T, Van Lint MT, Ljungman P, et al. Health and functional status of long-term survivors of bone marrow transplantation. EBMT Working Party on Late Effects and EULEP Study Group on Late Effects. European Group for Blood and Marrow Transplantation. Ann Intern Med 1997; 126: 184-192. 16.Bhatia S, Francisco L, Carter A, et al. Late mortality after allogeneic hematopoietic cell transplantation and functional status of long-term survivors: Report from the Bone Marrow Transplant Survivor Study. Blood 2007; 110: 3784-3792. 17.Sutherland HJ, Fyles GM, Adams G, et al. Quality of life following bone marrow transplantation: A comparison of patient reports with population norms. Bone Marrow
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Transplant 1997; 19: 1129-1136. 18.Andrykowski MA, Bishop MM, Hahn EA, et al. Long-term health-related quality of life, growth, and spiritual well-being after hematopoietic stem-cell transplantation. J Clin Oncol 2005; 23: 599-608.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Many late effects in allogeneic haematopoietic stem cell transplantation are related to chronic GvHD. One of the late effects below is clearly unrelated to GvHD: a) Chronic obstructive lung disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Cataract formation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Male infertility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Squamous cell carcinoma. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2. Total body irradiation (TBI) is not directly involved in: a) Secondary tumours after HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Gonadal insufficiency . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Osteoporosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Thyroid dysfunction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. Which of the following response concerning lymphoproliferative disorders (PTLD) after HSCT is not correct? a) Appear usually within the first 6 months after transplantation . . . . . . . . . . . b) Risk factors for PTLD are mismatched unrelated donors, T-cell depletion or ATG prophylaxis for GvHD. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Can be treated with anti-CD20 monoclonal antibody (rituximab) . . . . . . . . . d) B-cell PTLD are almost always of recipient origin . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. In long-term survivors after allogeneic HSCT, which of the following statements is true? a) Hypothyroidism is seen in nearly 90% of patients conditioned with BuCy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 252
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b) The major risk factor for avascular necrosis of bone is TBI, mainly when applied as single dose of 10 Gy or higher . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Younger age at HSCT, a longer interval between HSCT and seminal fluid analysis and the absence of chronic GvHD are associated with a higher recovery of spermatogenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Chronic obstructive pulmonary disease is mainly associated with chronic GvHD and can be efficiently treated with bronchodilatators. . . . . . . . . . . . . . . 5. Which sentence concerning general health status, social integration or quality of life is correct? a) 15 years after allogeneic HSCT the risk of late mortality is no longer increased when compared to a general population . . . . . . . . . . . . . . . . . . . . . . . . . b) 75% of patients who survive more than 5 years after HSCT return to full work or school after allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Fatigue and sleep disturbances are the most common complains after autologous and allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Psychological well-being occurs earlier than physical recovery in long-term survivors after allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 13
The value of molecular monitoring in haematological malignancy; minimal residual disease (MRD), relapse and chimerism
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1. Introduction It is estimated that patients with leukaemia have 1012 malignant cells at presentation. Following chemotherapy and/or HSCT, patients who are in complete remission by conventional analyses may still have 108/106 malignant cells which are below the detection limit of standard clinical evaluation. More sophisticated techniques are therefore necessary to detect this minimal level of disease. The detection of minimal amounts of residual disease (MRD) in acute lymphoblastic leukaemia (ALL) in children, ostensibly in complete remission (CR) following induction/consolidation therapy, has provided us with proof of principle of the clinical relevance of MRD. In such children, the detection and in particular the kinetics of MRD are clearly associated with disease relapse. The principle of MRD detection can be extended to other leukaemias in which there is either a demonstrable fusion gene e.g. BCR-ABL in chronic myeloid leukaemia (CML), aberrant antigen expression e.g. HNG2 in B-ALL with t(4:11) or immunoglobulin and/or T-cell receptor (Ig-TCR) gene rearrangements e.g. B-ALL. Methods to predict disease relapse after chemotherapy or HSCT allow early intervention and may result in “salvage” of many patients and improvement in the probability of long-term disease free survival (DFS). In addition to MRD analysis the determination of the chimeric status of the patients post allo-HSCT (the contribution of donor and recipient cells to post-transplant haematopoiesis) has assumed increased importance. In the rare event of graft rejection, the detection of emerging recipient cells may also facilitate early clinical intervention. Refinement of the technologies involved has allowed molecular monitoring to move from the research laboratory to the clinic.
2. Detection of Minimal Residual Disease (MRD) Cytogenetic analysis has identified a range of non-random chromosomal abnormalities (e.g. fusion genes, deletions), which are identifiable in malignant cells. Many of these changes involve translocation events, e.g. t(9,22) in CML. Molecular approaches have identified the genes involved in these translocation events, which in turn have allowed the development of specific molecular assays. While these assays have helped in defining a molecular classification of leukaemias, they have also permitted post treatment monitoring of residual disease. MRD may be defined as the presence of a small number of cells bearing molecular markers of the disease, but detected at a threshold far below that which can be achieved with conventional approaches such as morphological or classical karyotypic analysis (Figure 1). Serial monitoring of MRD status can provide an “early warning system” for disease relapse, thus permitting therapeutic intervention.
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Figure 1: Minimal residual disease
At diagnosis, the leukaemia blast load is approximately 1012. Pre transplant conditioning and alloHSCT reduces this leukaemic burden, leading to the concept of disease remission. However classical morphological remission in fact may only represent a 2-log reduction in leukaemia burden. Given the risk of disease relapse following allo-HSCT, more sensitive techniques to detect leukaemia cells may allow sub microscopic numbers of leukaemia cells to be detected. Use of molecular techniques has given rise to the concept of MRD where small numbers of leukaemia cells may either escape the conditioning regimen or re-appear post allo-HSCT giving rise to relapse. The GvL effect may also act to control low levels of MRD, thus reducing risk of leukaemia relapse. Detection of molecular evidence of relapse may provide a window of opportunity to intervene with approach such as DLI prior to evidence of clinical relapse
2.1. Methodology Optimal methods for MRD detection must have specificity for the leukaemia cell population and be sensitive enough to detect small numbers of leukaemia cells in a background of normal cells. 256
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2.1.1. Cytogenetic analysis If a karyotypic abnormality is detected at the time of diagnosis, cytogenetic analysis can be used to evaluate remission samples. However, the sensitivity is low (10-2) and the requirement for dividing cells to generate metaphase spreads may lead to a high failure rate of this technique, particularly in the immediate post treatment phase when obtaining sufficient metaphases may prove technically difficult. 2.1.2. Fluorescent in situ hybridisation Fluorescent in situ hybridisation technology (FISH), particularly when applied to interphase nuclei, is a more sensitive technique (10-3) than conventional cytogenetics. The lack of requirement for dividing cells means that the failure rate is lower than standard karyotyping in post-treatment marrows. 2.1.3. Immunophenotyping Leukaemia cells commonly express aberrant or unusual patterns of antigen expression when compared to their normal counterparts. This leukaemia-associated phenotype (LAP) allows improved classification of leukaemias and also provides a marker for MRD analysis. Immunophenotyping by multiparameter (3–4 colour) flow cytometry detects both surface antigen and intracellular markers and sensitivities range from 10-3 to 10-5. This approach has been particularly relevant in MRD analysis in childhood ALL and is the preferred technique in many clinical laboratories. The use of bench top flow cytometers which allow >6 colour flow to be performed will ensure high quality, sensitive and specific monitoring of MRD. Critical to the success of multiparameter flow cytometry is the requirement for inter-laboratory standardisation and this is currently being addressed by the Euroflow Consortium, a European Commission funded study to standardise fast and sensitive diagnosis and follow up of haematological malignancies (1). 2.1.4. Polymerase chain reaction analysis PCR based approaches have particular relevance in the detection of MRD due to their high specificity and sensitivity. In patients where a defined chromosomal lesion is present, primers are designed to bind to the nucleic acid of each of the gene partners in the translocation, thus allowing specific detection of the chromosomal lesion. In many cases, RNA will be the primary target but as PCR does not amplify RNA, the RNA must first be converted into cDNA using the enzyme reverse transcriptase (RT). When a chromosomal lesion is not present, MRD analysis relies on detection of clone-specific rearrangements such as the rearranged immunoglobulin (Ig) or Tcell receptor (TCR) genes (Ig-TCR-gene-rearrangements). These are particularly
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relevant for MRD detection in ALL, as many cases, even those without chromosomal aberrations, have rearranged IgH or TCR genes. Sensitivities of RT-PCR range from (10-4) to (10-6) depending on the type of leukaemia. Standardisation of detection of clonal IgH and TCR fingerprints and of the most frequent fusion transcripts in ALL (E2A-PBXI, MLL-AF4, BCR-ABL, TEL-AML) and the intrachromosomal microdeletion generating the SIL-TALI fusion gene, by the European Biomed and Europe Against Cancer Networks has allowed robust MRD detection to be available for single centre and multicentre studies (2). While semi-quantitative MRD studies yielded interesting results, emphasising the relevance of MRD detection, the introduction of real time quantitative PCR approaches (RQ-PCR) has empowered the clinical application of the results of MRD studies (3, 4). The European Study Group of MRD detection in ALL (ESG-MRD-ALL) has produced excellent standards and guidelines, harnessing the work of 30 European laboratories in concert to establish reproducible approaches for detecting, quantifying and assessing the relevance of MRD detection (4). To avoid contamination of PCR assays with old PCR products, a number of suggestions have been made, including use of dUTP and uracil N glycosylase (UNG), UV irradiation of surfaces, materials and reagents. However, physical separation of the individual part of the PCR assay into sample preparation, pre-PCR and post-PCR areas should be the central part of any contamination control strategy. Although carryover of amplified sequences contributes to the majority of the false positives, sampleto-sample contamination can also be a factor. Consequently, precautions must be taken in all aspects of sample handling, including use of plugged pipettes and tips at all stages of the process.
3. MRD detection in acute leukaemia Immunophenotyping and PCR based systems are currently the most relevant methods for detecting MRD, as they are highly sensitive and can be employed in the analysis of virtually all cases of acute leukaemia. With respect to immunophenotyping the most frequently expressed aberrant phenotypes in B-cell precursor ALL involve TdTdim/CD10+ and CD38dim/CD34+. CD19 is a useful additional gating marker. A panel of markers is employed at diagnosis, allowing selection of the most appropriate LAPs for MRD analysis in individual patients (Figure 2). In T-ALL, the employment of Tcell markers, particularly TdT and cytoplasmic CD3 with CD7 as an additional gating marker allows the majority of T-ALL to be detected. Judicious choice of the marker panel and its evaluation in individual patients allows the selection of robust MRD LAPs.
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Figure 2: MRD detection by immunophenotyping
MRD analysis was performed using the combination of CD66 and CD10 as a potential LAP. CD10 is labelled with PE and CD 66 with FITC. This combination does not appear on normal B-cells. By day 28 post therapy, leukaemia cell load has been reduced but MRD can still be detected
3.1. Methods 3.1.1. PCR of chromosomal aberrations RT-PCR can be applied in 40-45% of childhood precursor B-cell ALLs where 5 main targets for PCR analysis are used (Figure 3) and in 35–40% of adult precursor B-ALL, but is only relevant in 15–25% of T-ALLs. An example of an RT-PCR screen is shown in Figure 4. While standard RT-PCR is useful in establishing the appropriate target for MRD detection, RQ-PCR is the “gold standard” for MRD detection and evaluation. TaqMan probes designed for each of the 5 translocations most commonly seen in childhood ALL allow the kinetics of MRD to be determined by sequential RQ-PCR (Figure 5). 3.1.2. RQ-PCR of junctional regions of Ig and TCR gene rearrangements The variable (V), diversity (D) and junctional (J) regions of both the Ig and TCR genes undergo rearrangement to generate the primary antigen specific repertoire of the immune system. In normal lymphoid cells, there is a huge diversity of possible rearrangements whereas in lymphoid malignancies, the clonal nature of the disease means that cells derived from the precursor leukaemic clone will carry the same Ig or TCR rearrangement. Thus the junction of either region can be used as a target
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Figure 3: Frequency of chromosomal translocations observed in childhood B-cell ALL
The chromosomal translocation and genes involved in each translocation are indicated in the key legend
Figure 4: Molecular screening for translocations in childhood B-ALL
At diagnosis the sample is screened to ensure that the correct translocation as judged by karyotyping is detected. Two primer sets (A, B) which flank the fusion breakpoint are used to amplify material from the diagnostic sample (in the example shown the sample is positive for the E2A-PBX1 fusion gene). In order to confirm positivity, a confirmatory PCR is performed using a nested set of primers (C) located internal to the original PCR primers. Once the confirmatory screen has been performed, samples can be requested for subsequent MRD analysis 260
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Figure 5: Methodology of real-time PCR
Real time quantitative PCR (RQ-PCR) for MRD analysis for TEL-AML1. Amplification of different dilutions of fusion transcript (A) and calculation of the threshold cycle (Ct) for each dilution allows a standard curve to be generated (B) which will permit quantification of MRD levels in the follow-up samples. Equal amounts of RNA are examined at each time point allowing changes in leukaemia burden to be determined (C). Control gene amplification of a house-keeping gene ensures that equal amounts of RNA are being analysed (D)
for PCR based detection systems. Use of consensus PCR primers and subsequent DNA sequencing identifies patient specific sequences, allowing the design of TaqMan probes which can be employed in RQ-PCR assays to assess MRD quantitatively during treatment (Figure 6). As clonal evolution of Ig/TCR gene rearrangements between diagnosis and relapse has been described in 5–30% of cases in different studies, it is essential to employ at least two Ig/TCR targets for each patient being monitored by MRD. The employment of rigorous controls and the use of standardisation protocols (ESG-MRD-ALL) ensure that correct interpretation of MRD data will be performed in different laboratories and allows appropriate comparison
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Figure 6: Design of patient specific clonality marker
g
d
Schema for MRD analysis using clonality makers. The example shown is screening for an IgH clonality marker
of MRD data from different clinical studies, thus allowing robust incorporation of MRD based stratification into clinical protocols. 3.2. MRD detection in ALL Following the initial seminal papers on MRD in childhood leukaemia (5, 6), MRD analysis has been demonstrated to have clinical utility in a number of studies, both in de novo and relapsed ALL as well as in ALL patients undergoing allogeneic HSCT. This has led to the incorporation of MRD analysis into risk stratification in a number of childhood ALL protocols (7). MRD detected at early stages of treatment (< 3 months) reflect the patient’s response to the treatment regimen. A rapid clearance of residual disease is considered predictive of a more favourable outcome. Stratification based on MRD kinetics may allow (i) identification of high-risk disease and patients may require more intensive treatment (ii) identification of patients at low risk of relapse who might benefit from a reduction in treatment, thus reducing toxicity. 262
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3.3. MRD detection in acute myeloid leukaemia (AML) Protocols involving all trans-retinoic acid (ATRA) combined with anthracycline-based chemotherapy lead to cure in approximately 70% patients with PML-RARa-associated acute promyelocytic leukaemia (APL). PCR of the PML-RARa rearrangement can help identify patients who may require additional therapy or patients at low risk who may be spared unnecessary toxicity. As a consequence, molecular monitoring at 3 monthly intervals has been incorporated into a number of treatment algorithms. Based on samples acquired from patients entered in the Spanish PETHEMA protocols RQPCR has also been used to refine the significance of MRD positivity by stratifying patients into those at high, intermediate and low risk of relapse (8). Although the data is not as mature, MRD analysis in patients with t(8;21) and inv16 rearrangements employing both flow cytometry and RQ-PCR in tandem has demonstrated that mean differences in MRD levels between relapsing and non-relapsing patients are statistically significant and cut off levels could be assigned to risk of relapse (9). A number of other potential markers including WTI and nucleophosmin have been identified in acute leukaemia but their clinical relevance requires further study.
4. MRD detection in chronic leukaemias 4.1. MRD detection in chronic myeloid leukaemia (CML) Over 90% of cases of CML are associated with the chromosomal translocation t(9;22), and virtually all patients with CML express BCR-ABL fusion transcripts (e13a2 or e14a2) In initial studies, BCR-ABL fusion transcripts were detected by RTPCR based assays. Nested PCR (where a second round of PCR is performed using primers internal to the primer set used in the first round of PCR) can allow detection of a single BCR-ABL expressing leukaemia cell in 106 normal cells. The quality of RNA is assessed by amplification of a housekeeping gene transcript (e.g. ABL, BCR, Gus, etc). 4.1.1. Predictive value of early and late RT-PCR positivity Until the year 2000, allo-HSCT was the standard method of treating many patients with CML. Early RT-PCR positivity can be detected up to 9 months post allo-HSCT in patients receiving unmanipulated stem cell products in first chronic phase but over time patients usually revert to RT-PCR negativity. Early RT-PCR positivity is not associated with an increased risk of relapse. However early positive RT-PCR results may be relevant for patients who received a T-cell depleted allo-HSCT, and may be used to initiate donor lymphocyte infusions and/or tyrosine kinase inhibitors to prevent frank relapse. Late RT-PCR positivity (≥1 year post HSCT) in both unmanipulated and T-cell depleted allo-HSCT identifies patients at high risk of relapse.
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Serial testing is a prerequisite for using the results of RT-PCR to help guide therapy, as the kinetics of MRD evolution are more relevant than individual positive or negative results. Because of this, quantitative PCR approaches have now become the gold standard in assessing BCR-ABL transcripts in CML (10). Although originally designed to detect early relapse after allo-HSCT these techniques are now being used to assess the depth of response to the tyrosine kinase inhibitors. Quantitation of mRNA by RQ-PCR has been rigorously evaluated in a number of laboratories using either fluorescent-based Taqman or Light Cycler approaches. These approaches, which monitor product accumulation during the extension phase of PCR, are now being used for molecular quantification of MRD in many laboratories (Figure 7). Rising numbers of BCR-ABL transcripts following allo-HSCT precede cytogenetic and haematological relapse and are used as indicators for intervention with approaches such as DLI, which result in long-term disease free and RT-PCR negative remissions. Recent attempts to standardise mRNA quantitation by RQ-PCR for BCR-ABL have involved a multi-centre study which employed a lyophilised preparation of a K562 cell line as a potential quality control reagent (11). Vials were sent to 22 laboratories in 4 continents and results indicated that this QC control could be successfully employed in different laboratories with different PCR instruments, a first step in providing a universal QC reagent for BCR-ABL, RQ-PCR analysis. More recently there has been a worldwide effort to harmonise the methodology and reporting of RT–PCR for BCR-ABL transcripts (12).
Figure 7: Real-time quantification of BCR-ABL expression
Real time (RQ) PCR utilises a DNA hybridisation probe (e.g. a TaqMan probe) in addition to a set of fusion gene specific primers. The probe is constructed such that a fluorescent and a quencher moiety are linked together. In this configuration, no fluorescence is generated. During PCR, as the Taq polymerase extends the annealed primers during the extension phase of PCR, its exonuclease activity cleaves the bond between the reporter and the quencher, thus allowing the fluorescent moiety to be released into solution. During the linear phase of PCR, there is a direct correlation between the amount of target and the amount of fluorescence generated. Collection of this real time PCR data allows accurate quantification of the target, allowing MRD levels to be accurately quantified 264
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4.2. MRD detection in chronic lymphocytic leukaemia (CLL) Until relatively recently research into cellular abnormalities in CLL was hampered by the inability to make human B cells divide. Newer techniques such as FISH molecular analysis and 3–4 colour flow cytometry have allowed the establishment of a number of potential prognostic markers (CD38, Zap 70, somatic hypermutation) and to begin to look for residual disease following treatment. Treatment, with the exception of the few patients receiving allo HSCT, was until recently confined to palliation. With the advent of newer therapeutic regimens including purine analogues (fludarabine), cyclophosphamide and the monoclonal antibody Rituximab, complete response (CR) has become a possibility. The absence of identifiable CLL cells in the blood or bone marrow by flow cytometry confirms a CR and serial examinations for the re-emergence of MRD in the guise of CD5/CD19 + CD20/CD79b neg cells may serve as an indication for further therapy or a change in strategy (Figure 8). Figure 8: B-CLL MRD analysis by flow cytometry
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5. Chimerism following allogeneic HSCT The term chimerism following HSCT refers to the proportion of donor and recipient cells found in the host at any given time following the transport procedure. Proportions of each may change over time and a schema for definitions is given in Table 1.
Table 1: The four distinct categories of Chimerism in the human following allogeneic HSCT Donor Chimerism (DC)
All cells are of donor origin following allogeneic HSCT
Transient Mixed Chimerism (TMC)
A mixed profile with a proportion of cells (typically 1–5%) post HSCT of recipient origin is detected in the first 6 months post transplant; subsequently the patient reverts to donor chimerism
Stable Mixed Chimerism (SMC)
A mixed profile with a proportion of cells (usually 1–20% recipient cells) detected post allogeneic HSCT and remain at a constant level over time
Progressive Mixed Chimerism (PMC)
A mixed profile with a proportion of recipient cells. These recipient cells increase to >10% recipient cells over time
5.1. Techniques to measure chimerism 5.1.1. Cytogenetics Cytogenetic analysis can be performed, provided there is a marker which distinguishes between donor and recipient cells, such as the presence of the Y chromosome in sex mismatched transplants. In addition the presence of a karyotypic abnormality can also serve as a marker. However chimerism studies using cytogenetic analyses are compromised by low sensitivity and the need for dividing cells. Use of fluorescent in situ hybridisation (FISH) analysis can increase sensitivity and eliminates the need for dividing cells but is probably only of real benefit in the sex mismatched transplant setting. 5.1.2. DNA based techniques Southern blotting techniques using either single copy or minisatellite probes were initially used to detect post-transplant chimerism. While these techniques allow some degree of assessment of virtually all allo-HSCT, the relative lack of sensitivity and the laborious nature of the techniques has prevented their widespread use. Most 266
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laboratories now use PCR based techniques, exploiting polymorphisms in DNA to distinguish between donor and recipient cells. Variable number tandem repeats (VNTRs) can be amplified to allow size differences between donor and recipient cells to be used in PCR-based post-transplant chimerism analysis but the most widely used approach involves PCR of short tandem repeats (STR) (Figure 9). The Eurochimerism Concerted Action, a consortium of leading laboratories in this field from 10 European Countries, has developed a Eurochimerism marker panel (EUR-STR) and
Figure 9: STR DNA polymorphisms to evaluate chimerism after allo-HSCT -HSCT
Select informative polymorphism(s)
Study post-HSCT pattern
Short tandem repeat PCR (STR-PCR) exploits the presence of small repeated sequences of DNA which can vary in the number of repeats between donor and recipient. An example of 2 types of STR is shown (A) a dinucleotide CA repeat and (B) a tetranucleotide TTTA repeat Variation in repeat length between donor and recipients allows post transplant chimerism to be assessed. A panel of STR markers is first used to identify informative polymorphisms (C) which are then used to monitor samples from the recipient post HSCT (D). In this example an initial period of donor chimerism is followed by re-emergence of recipient cells and disease relapse. Samples can be electrophoresed through a gel matrix as indicated or by capillary electrophoresis. Activation of the fluorescent primers by an argon laser using for example a 310 or 3100 DNA sequence detector (Applied Biosystems) allows quantitation of donor and recipient profiles. D: Donor Profile; R: Recipient profile; 1-5: Serial chimerism profiles post-HSCT; O: Operator profile (to control for contamination)
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has shown its application to quantitative chimerism analysis. Furthermore it has established the allele frequencies of the EUR-STR panel in different European populations, has standardised the nomenclature and has developed DNA extraction methodologies for chimerism analysis (www.eurochimerism.org). Chimerism techniques employ a fluorescent based PCR methodology providing a suitable format for application in clinical laboratories. 5.2. Significance of chimerism analysis following allo-HSCT “Classic” allo-HSCT is preceded by myeloablative conditioning regimens, the aim of which is the complete destruction of host haematopoiesis. In patients with leukaemia, the re-emergence of cells of host origin frequently heralded leukaemia relapse, although rarely normal host cells may partially repopulate host haematopoiesis. Transient mixed chimerism (TMC) may occur in the first 6–9 months post transplant, particularly following matched unrelated donor HSCT (MUD-HSCT) and may be associated with a lower risk of GvHD. Since the advent of reduced intensity stem cell transplantation (RIC-HSCT) the relevance of serial chimerism analysis has been re-stabilised. The philosophy of RICHSCT is to reduce the potency of the conditioning therapy to reduce toxicity and not to attempt myeloablation. This theoretically results in a period of mixed haematopoietic chimerism in the host following infusion of donor stem cells. Over time, donor cells will predominate and exercise their graft versus leukaemia effect (GvL) (Figure 10). In the presence of persistent or progressive MC, DLI is performed in an attempt to establish donor chimerism, albeit at the risk of precipitating GvHD. Chimerism in these recipients is best evaluated by the use of lineage specific analysis. Selection of appropriate cell populations (T-cells, myeloid cells etc) is first performed and the individual populations are subsequently subjected to chimerism analysis. Achievement of complete donor chimerism in the T-cell lineage may be of particular relevance to the long-term success of this approach. Chimaeric analysis therefore is useful to follow the success of donor lymphocyte infusions (DLI) for relapsed leukaemia (especially CML) or when DLI is used to convert mixed chimaeras into donor chimaeras following RIC-HSCT (13). 5.2.1. Chimerism analysis in severe aplastic anaemia (SAA) Although an extremely successful therapy for SAA, allo HSCT in this setting is significantly different from acute or chronic leukaemia. Current conditioning regimens are not myeloablative (cyclophosphamide alone or with ATG). The presence of mixed chimerism therefore is common and stable mixed chimerism (SMC) is associated with a good outcome and absence of GvHD. Emergence of progressive mixed chimerism (PMC) usually precedes disease relapse/graft rejection and carries 268
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Figure 10: Chimerism after a non-myeloablative stem cell transplant
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Shows recipient (a) and donor (b) STR profiles prior to NST. Following reduced intensity conditioning a mixed chimaeric state is common (c). Overtime, this may progress to full donor chimerism (d). Persistent mixed chimerism is an indication for DLI (e) to convert the patient to a full donor chimaera (d)
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a poor prognosis. In contrast to the leukaemias, allo HSCT for SAA may be followed by graft rejection. This may be early, when there is no evidence of donor engraftment, or may occur later following transient donor engraftment with a subsequent rise in host cells and graft rejection which may be interpreted as disease relapse. Reduction in exposure to multiple red cell and platelet transfusions and newer treatment regimens have reduced the graft rejection rate to <10%. A small number of patients (1.0–4.0%) will achieve complete autologous recovery. A number of studies have indicated that SMC is associated with excellent long-term disease free survival and absence of GvHD while complete donor chimerism (DC) is associated with an increase risk of GvHD. The detection of rising numbers of host haematopoietic cells prior to or at the time of withdrawal of immune suppression (usually after one year) may be a warning of impending graft rejection, which carries a poor prognosis. Therefore careful monitoring of the percentage of host haematopoietic cells should be undertaken prior to the discontinuation of immunosuppressive therapy. A detailed chimeric analysis of a subset of these patients has shown that stable mixed chimerism is associated with little or no GvHD (14). In sickle cell disease and in a number of genetic disorders that are amenable to allogeneic HSCT (e.g. Hurlers syndrome), achievement of mixed haematopoietic chimerism is associated with a significant clinical improvement in the underlying condition.
6. Discussion and future directions Initially, the detection of MRD in CML following HSCT was extremely helpful in identifying patients at risk of relapse as early intervention with approaches such as DLI was associated with prolonged disease free survival. The introduction of TKIs in the treatment of CML has opened a new era in the use of molecularly targeted therapy for the treatment of malignant disease. Subsequently, the application of MRD techniques in paediatric patients with ALL treated with chemotherapy has allowed stratification of patients into risk groups and consequently identified those who might benefit from early intervention with HSCT. A similar approach has proved successful in adult patients with APL. Monitoring of chimerism status following allo-HSCT may help predict graft rejection (in SAA) or disease relapse (in leukaemia). It is of particular value In the setting of RIC conditioning regimens, as the indication for use of DLI will depend on the kinetics of chimerism. Monitoring the response to DLI or other immuno-modulatory approaches is also made possible by serial monitoring of chimerism status. The concept of molecular monitoring of malignant haematological disorders following either allo HSCT or innovative drug therapies has proven to be of great value. With 270
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the proliferation of small molecules in the treatment of various leukaemias and other haematological disorders, the use of molecular monitoring will become a standard method for determining the timing of intervention and the outcome for patients with these disorders. The standardisation of techniques and strict laboratory controls are extremely important if these results are to be used by clinicians in treatment decisions. Efforts to standardise these techniques have been successful (BIOMED, EAC, EUROCHIMERISM ESG-MRD-ALL, EUROFLOW) and it is hoped that co-operation between different laboratories will allow standardisation to become a universal phenomenon. MRD detection and its application in haematological malignancies represent an excellent example of the application of translational medicine in clinical management of leukaemia patients and may serve as a model for other cancers.
Acknowledgments We wish to acknowledge the support of the Bone Marrow for Leukaemia Trust, the Health Research Board, the Children’s Leukaemia Research Project and Cancer Research Ireland.
References 1. Orfa A, Lopez A, Flores J, et al. Diagnosis of haematological malignancies; New applications for flow cytometry. Haematology (EHA Edu Program) 2006; 2: 6-13. 2. Van Dongen JJ, Macintyre EA, Gabert JA, et al. Standardized RT-PCR analysis of fusion gene transcripts from chromosome aberrations in acute leukemia for detection of minimal residual disease. Report of the BIOMED-1 Concerted Action: Investigation of minimal residual disease in acute leukemia. Leukemia 1999; 13: 1901-1928. 3. Gabert J, Beillard E, van der Velden VH, et al. Standardization and quality control studies of ‘real-time’ quantitative reverse transcriptase polymerase chain reaction of fusion gene transcripts for residual disease detection in leukemia - a Europe Against Cancer program. Leukemia 2006; 20: 886-888. 4. Van der Velden VH, Cazzaniga G, Schrauder A, et al. European Study Group on MRD detection in ALL (ESG-MRD-ALL). Analysis of minimal residual disease by Ig/TCR gene rearrangements: Guidelines for interpretation of real-time quantitative PCR data. Leukemia 2007; 21: 604611. Epub 2007 Feb 8. 5. Cavé H, van der Werff ten Bosch J, Suciu S, et al. Clinical significance of minimal residual disease in childhood acute lymphoblastic leukemia. European Organization for Research and Treatment of Cancer – Childhood Leukemia Cooperative Group. N Engl J Med 1998; 339: 591-598. 6. Van Dongen JJ, Seriu T, Panzer-Grümayer ER, et al. Prognostic value of minimal residual disease in acute lymphoblastic leukaemia in childhood. Lancet 1998; 352: 1731-1738. 7. Van der Velden VH, Cazzaniga G, Schrauder A, et al; European Study Group on MRD detection in ALL (ESG-MRD-ALL). Analysis of minimal residual disease by Ig/TCR gene rearrangements:
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Guidelines for interpretation of real-time quantitative PCR data. Leukemia 2007; 21: 604611. Epub 2007 Feb 8. 8. Santamaría C, Chillón MC, Fernández C, et al. Using quantification of the PML-RARalpha transcript to stratify the risk of relapse in patients with acute promyelocytic leukemia. Haematologica 2007; 92: 315-322. 9. Perga G, Lasa A, Aventin A, et al. Minimal residual disease in AML; prognostic value of minimal residual disease (MRD) in acute myeloid leukaemia with favourable cytogenetics et {(8;21 and inv (16)}. Leukaemia 2006; 20: 87-94. 10.F Lin, F van Rhee, JM Goldman, NC Cross. Kinetics of increasing BCR-ABL transcript numbers in chronic myeloid leukemia patients who relapse after bone marrow transplantation. Blood 1996; 87: 4473-4478. 11.Saldanha J, Silvy M, Beaufils N, et al. Characterization of a reference material for BCRABL (M-BCR) mRNA quantitation by real-time amplification assays: Towards new standards for gene expression measurements. Leukemia 2007; 21: 1481-1487. Epub 2007 May 3. 12.Hughes T, Deininger M, Hochhaus A, et al Monitoring CML patients responding to treatment with tyrosine kinase inhibitors: Review and recommendations for harmonizing current methodology for detecting BCR-ABL transcripts and kinase domain mutations and for expressing results. Blood 2006; 108: 28-37. 13.Peggs KS, Thomson K., Hart DP, et al. Dose escalated donor lymphocyte infusions following reduced intensity transplantation: Toxicity, chimerism, and disease responses. Blood 2004; 103: 1548-1556. Epub 2003 Oct 23. 14.McCann S, Passweg J, Bacigalupo A, et al. The influence of cyclosporine alone or cyclosporine and methotrexate on the incidence of mixed haemopoietic chimerism following allogeneic bone marrow transplantation for severe aplastic anaemia. Bone Marrow Transplant 2007; 39: 109-114.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which of the following is most likely to occur after non-myeloablative stem cell transplantation? a) All patients become complete donor chimaeras . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) No patients become complete donor chimaeras . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Mixed chimerism is a common finding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Donor lymphocyte infusions are always required. . . . . . . . . . . . . . . . . . . . . . . . . . . .
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2. The sensitivity of detection of the leukaemia associated phenotype in childhood ALL detected by flow cytometry is: a) 10-1–10-2 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 10-2–10-3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 10-3–10-4 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 10-3–10-5 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which of the following is true of the clinical evaluation of chimerism following HSCT? a) Always involves the use of radioactivity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Can be carried out with the use of fluorescent primers . . . . . . . . . . . . . . . . . . . . c) Fluorescent primers are inadequate for clinical use . . . . . . . . . . . . . . . . . . . . . . . . . d) Tetranucleotide repeats cannot be used . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. In CML which of the following statements about complete molecular response after imatinib therapy is correct? a) Complete molecular response is accepted at transcript levels 3 logs less than at the beginning of treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Complete molecular response is accepted at transcript levels 2 logs less than at the beginning of treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Complete molecular response is never achieved . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Complete molecular response is achieved in 90% of patients treated in CP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Following treatment for APML molecular monitoring should be carried out: a) Every month . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Every 3 months . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Every 6 months . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Is of no value . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 14
Immunotherapy post-transplant
*
14.1
Immunotherapy post-transplant for infections
M. Kapp, G.U. Grigoleit, H. Einsele
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CHAPTER 14.1 • Immunotherapy post-transplant for infections
1. Introduction The main issue regarding immunotherapy of infections after allogeneic haematopoietic stem cell transplantation (HSCT) is human cytomegalovirus (HCMV) infection. Therefore, this Chapter will focus on the experiences made with immunotherapy for HCMV infection. However, the techniques described here have been applied to immunotherapy of other viral infections such as adenovirus and EBV infection.
2. HCMV infection after allogeneic stem cell transplantation HCMV infection continues to be one of the most important and life threatening complications after allogeneic HSCT. To improve T-cell immunity against HCMV and other infections in bone marrow transplant patients different strategies have been explored. In general, HCMV-specific T-cells can be selected from the donor, and be transferred to the patient either with or without in vitro expansion, or the HCMV-specific T-cells can be activated and expanded in vivo by stimulation with antigen presenting cells (APCs) loaded with specific proteins or peptides.
3. Isolation and infusion of HCMV specific CD8+ T-cells Riddell et al. and Walter et al. have demonstrated that adoptive immunotherapy by transfer of HCMV-specific CD8+ T-cell clones into patients at risk of HCMV disease protected the patients from HCMV-related complications (1, 2). 1-2 x 109 HCMVspecific CD8+ T-cells were administered and these cells were detectable in patients’ blood for at least 8 weeks. Patients were protected against HCMV disease although HCMV-specific CTLs declined progressively in patients who did not develop a concomitant HCMV-specific CD4+ TH response. Analyses using the new technologies developed for determining antigen-specific Tcells (e.g. intracellular cytokine staining or staining with tetrameric HLA Class I /peptide complexes) have contributed to a substantial improvement in our understanding of the role and function of immune responses in vivo. The use of peptide–HLA multimers facilitates the visualisation and isolation of antigen-specific CTLs (3). CD8+-T-cells that bind multimeric HLA complexes can be isolated to high purity using magnetic beads or FACS sorting (4–6). Newly developed multimeric HLA complexes, binding reversibly to the T-cell receptor, offer the opportunity of selecting unmanipulated antigen specific CTLs (7). Thus, phenotypical analysis with MHC-peptide multimers, functional assays as well as multimer-based enrichment protocols can now be used in the setting of adoptive T-cell therapy. The transfer of HCMV-specific CTLs freshly isolated from peripheral blood might be superior to
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the in vitro expansion and manipulation of T-cells. The in vitro expansion may increase the expression of the pro-apoptotic FAS molecule (CD95) and reduce telomere length of specific T-cells, leading to a shorter survival of the adoptively transferred T-cells. In addition, the prolonged in vitro culture of T-cells is cumbersome when performed under GMP conditions (requirements for sterility, media, cytokines, serum, etc.).
4. Adoptive transfer of HCMV-specific CD4+ TH-cells for the treatment of HCMV infection The safety and efficacy of cellular therapy with HCMV-specific CD8+ lymphocytes in immunocompromised patients has been documented. These studies demonstrated the role of HCMV-specific CD4+ cells in maintaining an HCMV-specific CTL response. Several studies have outlined the significance of antiviral effector functions of TH cells in maintaining CTL responses after adoptive transfer and their capacity to produce antiviral cytokines (8). We therefore evaluated the infusion of HCMV-specific CD4+ TH-cells to treat patients with HCMV viraemia resistant to antiviral chemotherapy after allogeneic HSCT (9). The patients enrolled in this study showed a documented lack of an HCMV-specific CD4+ TH and an HCMV-specific CTL response. We generated HCMV-specific T-cell lines for adoptive transfer by 4 repetitive weekly stimulations of donor lymphocytes with HCMV lysate in vitro. No side effects occurred during and after infusion of the cells, even in patients receiving their graft from a donor, mismatched in 1 to 3 HLA antigens with the stem cell graft recipient. Reconstitution of HCMV-specific CTL responses could be demonstrated following the transfusion of HCMV-specific CD4+ T-cell lines. Five out of 8 patients cleared the viral infection following a single infusion of HCMVspecific CD4+ T-cells, one other patient after a second infusion. These findings show that HCMV-specific adoptive immune transfer is a therapeutic option in patients with reactivated HCMV-infection after HSCT. Even the infusion of low numbers of CD4+ HCMV-specific T-cell lines was found to be successful in some of these patients. Another issue which has to be addressed is the role of CD4+ T-cells in the direct control of infection. It has been shown that highly differentiated CD4+ T-cells, specific for HCMV pp65, mediate antiviral effector functions (10): HCMV-specific CD27- CD4+ Tcells degranulate when they encounter a cognate antigen and the number of CD4+ T-cells which contain granzyme A, granzyme B and perforin raises during the maturation process of these cells. Furthermore, target cells bearing a distinct HCMV pp65-derived MHC Class II-restricted epitope were killed by CD4+ T-cells from an individual in which degranulation occurred in a subset of cells with a high frequency of perforin. Overall, it could be demonstrated that mature HCMV-specific
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CD4+ T-cells have functional features which equal antiviral CD8+ T lymphocytes. Taken together, the transfer of HCMV-specific CD4+ T-cells is not only a promising option for providing helper cell support for pre-existing HCMV-specific CTLs but may also exert effector functions which directly contribute to a persistent virus control.
5. Infusion of HCMV-specific CD4+ and CD8+ T-cells The opportunity to generate cellular products which contain both CD4+ and CD8+ arises from the provision of selection protocols based upon cytokine secretion. Our group has performed a study which avoids the restrictions in the generation of virusspecific T-cell lines for adoptive transfer into allogeneic HSCT recipients without affecting the function of the generated T-cell lines (11). The IFN-g secretion assay, which conforms to current GMP-regulations, was used for the enrichment of virusspecific CD4+ and CD8+ T-cells. One important step to stimulate both cytotoxic and helper cells in one procedure was to evaluate the optimal stimulus. First, we compared HCMV-Ag only in eliciting a combined HCMV-specific CD4+ and CD8+ Tcell response to concomitant usage of HLA matched HCMV-specific MHC-I epitopes and HCMV-Ag. Stimulation of HCMV-specific CD4+ and CD8+ T-cells was similarly effective when using HCMV-antigen compared to the stimulation with HCMVantigen and peptides, as assessed by intracellular cytokine staining (ICC). The number of HCMV-specific T-cells required for an effective adoptive transfer and their composition in respect to CD4+/CD8+ ratio required for either prevention and/or treatment of HCMV-viraemia after allogeneic stem cell transplantation is not yet defined. The Seattle group and others demonstrated that adoptive transfer of up to 5 x 109/m2 CD8+ T-cell clones could reconstitute HCMV-specific immune responses (1, 2). In our hands, the combined generation of HCMV-specific CD4+ and CD8+ cells resulted in an average of 1.3 x 108 HCMV-specific stimulated, selected and expanded T-cells from 8/8 randomly selected HCMV-seropositive donors in 10 days from one single 500 mL collection of blood, utilising feeder cells, media and cytokines compatible with current GMP regulations. The specificity of this GMP-grade product could be clearly demonstrated by detection of intracellular cytokine production after antigen-specific stimulus and lysis of HCMV-infected target cells by the HCMV-directed CTL lines. Furthermore, we were able to show that the generated HCMV-specific T-cells do not represent terminally differentiated CD4+ and CD8+ T-cells as stimulation led to several cell divisions as demonstrated by dilution of CFSE dye and a corresponding cell expansion. Thus, adoptive transfer of the generated T-cells into HSCT recipients may allow further in vivo expansion, if T-cells are stimulated by HCMV-Ag presenting cells in vivo.
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6. Vaccination with HCMV peptide loaded DCs Mature dendritic cells (DCs) are the most potent APCs, with the ability to initiate and boost primary immune responses. Mature DCs can be generated in vitro from peripheral blood monocytes by culturing in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 4 (IL-4) and tumour necrosis factor a (TNF-a). Vaccination with peptide pulsed DCs was shown to be feasible and effective in inducing tumour-specific T-cell responses and was able to induce regression of metastatic disease in a minority of patients. In our trial (12) HCMV seropositive patients who underwent an allogeneic-HSCT with a graft from a HCMV seronegative donor received a vaccination with HCMV peptide loaded mature DCs in addition to the routinely applied pre-emptive antiviral chemotherapy. The DCs were pulsed with nonamer peptides from the HCMV protein pp65, restricted by the HLA-class I elements A1, A2, A3, A11, A68 and B7. The DC vaccination study was designed as a phase I/II trial. The primary objectives were to evaluate tolerability and safety. Therefore, the acute local and systemic side effects of the vaccination procedure were analysed as well as the induction or aggravation of acute or chronic graft versus host disease (GvHD) or any other autoimmune phenomenon. As a secondary objective, the efficacy of the DC vaccination was documented by analysing the HCMV specific T-cell response and the long-term control of HCMV infection. Because all the patients with HCMV infection or reactivation at the time of vaccination received additional anti-viral chemotherapy, the short-term control could not be analysed. Our DC vaccination trial recruited HCMV seropositive HSCT recipients, who are at a high risk for HCMV disease without a HCMV specific CTL response. Patients at high risk for HCMV disease were defined as patients with prolonged (more than 4 weeks) antiviral chemotherapy prior to day 100, patients receiving a T-cell depleted (in vivo by ATG or ex vivo by CD34 selection) graft and HCMV seropositive patients with a seronegative stem cell donor. In all patients, DC vaccination was well tolerated without any local or systemic side effects. No induction or aggravation of acute or chronic GvHD was observed in any of the patients receiving DC vaccination for chemotherapy refractory HCMV infection. DC vaccination was shown to induce HCMV specific T-cell responses with antiviral activity even in a patient who was infected by a HCMV strain resistant to ganciclovir, foscavir and cidofovir. In conclusion, HCMV specific DC vaccination is a feasible and effective immunotherapy among recipients of an allogeneic haematopoietic stem cell transplant and will be further tested in a multicentre study.
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7. Vaccination with recombinant modified vaccinia Ankara (MVA) The usage of full-length antigens expressed from MVA is a powerful immunotherapeutic tool applicable regardless of an individual’s HLA type. It has been shown that dual HCMV antigen pp65/pp150 expressing MVA can be strongly recognised in vitro by PBMC from CMV positive healthy subjects with HLA A*1101, HLA A*6801, HLA A*0301 and HLA B*0702 haplotypes. Furthermore, evaluation of immunogenicity indicates that pp65-IE1-MVA additionally can induce robust primary immune response to both antigens in HLA A2.1 transgenic mice (13, 14). Since MVA has an exceedingly large capacity for foreign DNA, potentially multiple disease targets could be incorporated into one vaccine, thereby reducing manufacturing costs and dosing to patients.
Table 1: Treatment options for HCMV infection after allogeneic HSCT Adoptive immunotherapy with Vaccination Based on Dendritic cells HCMV-specific T-cells
Vaccination Based on Modified Vaccinia Virus (MVA)
Adoptive immunotherapy with HCMV specific cytotoxic T-cells has shown to be effective in improving the elimination of HCMV. A detailed summary of different strategies when performing adoptive immunotherapy is provided in Table 2
Preclinical studies have been performed and demonstrate the feasibility and efficacy of vaccinia-based HCMV vaccines in mice (13, 14)
Study using HCMV-peptide pulsed dendritic cells to treat HCMV infections after allogeneic HSCT. This strategy is feasible and improves control of HCMV in allografted patients (12)
8. Conclusions Antigen specific T-cells are essential to the control of reactivation or primary infection with HCMV or other viral infections. Immunotherapy offers an attractive tool to improve immune reconstitution in these patients, leading to control of viral replication without apparent side effects. This may reduce the usage of potentially toxic antiviral chemotherapy (adverse effects such as myelo- or nephrotoxicity) and circumvent the increasingly reported problems of the development of antiviral drug resistance. Stimulation and expansion conditions have to be improved to generate T-cell lines containing not only terminally differentiated effector cells but also central-memory T-cells, which are essential to build up a memory T-cell response in the recipient (15). Further controlled trials with adoptive transfer of virus-specific T-cells versus
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Table 2: Clinical application of HCMV-specific immunotherapy
280
Year (Ref.)
Strategy
Results
Riddell S. 1992 (2)
Infusion of HCMV-specific cytotoxic T-cell clones
First trial that demonstrated a successful transfer of HCMV-specific CTL isolated from bone marrow donors, propagated in vitro, and adoptively transferred to immunodeficient bone marrow transplant recipients. The transfer was safe, and reconstitution of HCMV-specific CTL responses could be documented.
Riddell S. 1995 (1)
Infusion of HCMV-specific cytotoxic T-cell clones
Fourteen patients received intravenous infusions of HCMV-specific T-cell clones from their donors. In total, 56 infusions (4/patient) of HCMVspecific cytotoxic-T-lymphocyte clones were performed without any major side effects and no viraemia or HCMV disease was observed in any patients receiving adoptive immunotherapy.
Einsele H. 2002 (8)
Infusion of HCMV-specific polyclonal CD4+ T-cell lines
HCMV load dropped significantly in all 7 evaluable patients, with a maximal reduction after a median of 20 days (range, 5-31 days). Anti-HCMV cellular therapy was successful in 5 of 7 patients, whereas in 2 of 7 patients, who received an intensified immune suppression at the time of or after T-cell therapy, only transient reductions in virus load were obtained.
Mackinnon S. 2003 (6)
Adoptive cellular therapy with virus-specific T-cell lines
Polyclonal HCMV-specific T-cell lines were generated for the treatment of 16 patients. A massive in vivo expansion of HCMV-specific CTLs was observed leading to a recovery of virusspecific T-cell responses. 50% of the treated patients (8/16) did not require a further treatment with antiviral drugs.
Moss P. 2005 (5)
Direct isolation and infusion of HCMV specific CTL using HLA peptide multimeric complexes
Reduction of HCMV viraemia in all treated patients and a complete clearance of HCMV infection in 8 patients, including one patient who had a prolonged history of HCMV infection that was refractory to antiviral therapy. HCMV-specific CD8+ T-cells were detected in all patients within 10 d of infusion.
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Table 3: Current projects “Adoptive immune therapy of chemotherapy-refractory CMV-infection with Streptamerselected T-cells after allogeneic bone marrow- or peripheral blood stem cell transplantation” Correspondence to: Dr. Götz Ulrich Grigoleit Medizinische Klinik und Poliklinik II Building C11 Josef-Schneider-Strasse 2 97080 Würzburg, Germany E-Mail:
[email protected]
Although the adoptive transfer of in vitro expanded T-cells is effective its application has several drawbacks. It is very time consuming and laborious to prepare the high cell numbers needed for transplantation, especially since patients with an acute CMV-infection may not allow to wait for the preparation of the cells. In addition the costs of such a treatment are extraordinary high, raising the question if such a medication could be broadly introduced into clinical practice. Due to these drawbacks multimer-based direct isolation of antigen-specific T-cells has evolved. The use of low cell numbers without the need of time consuming in vitro expansion greatly improves the adoptive transfers of T-cells. Even though this approach has advanced adoptive transfer protocols, multimer-based T-cell isolation still suffers from several disadvantages. The multimer reagents bind to the T-cell receptor during the isolation procedure and thus stimulate the respective T-cell. This stimulus is likely to cause apoptosis in the isolated cells, interfering with an effective T-cell transplantation. In addition the multimer reagents are transferred together with the T-cells to the patient, which may cause toxic or immunogenic side effects. Recently the Streptamer technology was developed to overcome these problems (7). Streptamers are reversible multimers which are unlikely to interfere with T-cell function since Streptamer reagents can be rapidly dissociated from the T-cell receptor. In addition adoptively transferred cells do not contain Streptamer. Primary endpoints of this study are acute infusion related toxicities, such as anaphylactic reaction, and the development of GvHD. Secondary endpoints are the HCMV-load measured by quantitative PCR and the reconstitution of HCMV specific cellular immunity. To assess the side effects (primary endpoint) of the treatment, all patients undergo the following examinations before and 1, 2, 4 weeks then monthly (up to 6 month) after transplantation: physical examination, Karnofsky-score, bodyweight, body size, surface area, clinical grading of GvHD (Seattle-scheme), serum chemistry, coagulation tests, and differential blood count. To assess the efficacy of the adoptive T-cell therapy (secondary objective) the viral load and the frequency of the HCMV specific T-cells will be monitored before and 1, 2, 4 weeks then monthly (up to 6 month) after adoptive T-cell transfer. continued
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“Adoptive immunotherapy of chronic HCMV infection post allogeneic HSCT – DC vaccination” Correspondence to: Dr. Götz Ulrich Grigoleit Medizinische Klinik und Poliklinik II Building C11 Josef-Schneider-Strasse 2 97080 Würzburg, Germany E-Mail:
[email protected]
Dendritic cell (DC) vaccination is a pathogen specific and highly immunostimulatory approach to fight different infections and malignancies, but has never been evaluated in the setting of allogeneic HSCT. The risk of GvHD still inhibits the use of immunostimulatory therapies such as DC vaccination. We completed a phase I/II study including 24 allogeneic HSCT recipients at high risk for HCMV disease to analyse feasibility and efficacy of vaccination with HCMV peptide loaded DC. No acute side effects were observed and we could demonstrate a significant clinical benefit in comparison to our control group. Furthermore, an induction or expansion of HCMV-specific CTL could be observed in 5 patients after DC vaccination. To evaluate efficacy of this vaccination strategy, we set up a consecutive study with the goal to vaccinate another 50 patients. “Adoptive immunotherapy of chemotherapy-refractory CMV or EBV infection using CD4+ and CD8+ T-cells selected by cytokine-secretion assay” Correspondence to: Dr. Max S. Topp Medizinische Klinik und Poliklinik II Röntgenring 11 97070 Würzburg, Germany E-Mail:
[email protected]
Goals: To assess toxicity and efficacy of cytokine-capture based selection of CMV- or EBV-specific CD4+ and CD8+ T cells in patients with refractory CMV/EBV infection after allo-HSCT.
pre-emptive or prophylactic antiviral drug administration are needed to define the role of cellular immunotherapy in the treatment algorithms of infections in the immunocompromised host. Although availability of clinical grade reagents for the selection of antigen specific T-cells has largely improved in the last few years, we still need to learn more about the best composition of a cellular product used for adoptive transfer. There is still controversy about the benefit of transferring different T-cell subsets for adoptive immunotherapy: namely, should we transfer only CD4+ T-cells or CD8+ T-cells or a combination of both. 282
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In addition, various methods of cell isolation and/or expansion result in different stages of T-cell senescence. Freshly isolated and specifically selected T-cells have greater expansion potential in vivo when compared to repetitive in vitro stimulated T-cells. In contrast specific stimulation ex vivo might be more effective in depleting alloreactive T-cells from the T-cell product. In view of the many different methods of isolating and generating T-cells, future studies have to define the best isolation procedures, the optimal T-cell subpopulations to be used for antiviral T-cell therapy and the differentiation/activation stage or stages of specific T-cells to be preferentially applied. In addition we will have to define the optimal cell dose depending on viral load and immunosuppression of the patient for each of these different cell populations. According to established protocols for the generation of sufficient numbers of donorderived virus-specific T-cells for adoptive immunotherapy, the donor has to be antigenexperienced. Thus, alternative strategies for priming virus-specific T-cell responses are highly warranted. Because no relevant adverse effects were observed in our first DC vaccination trial in allogeneic HSCT recipients and because induction and expansion of HCMVspecific T-cell responses leading to viral control was observed in some of the patients, we will further evaluate and try to improve the efficacy of DC vaccination post allogeneic HSCT in a larger cohort of patients.
References 1. Walter EA, Greenberg PD, Gilbert MJ, et al. Reconstitution of cellular immunity against cytomegalovirus in recipients of allogeneic bone marrow by transfer of T-cell clones from the donor. N Engl J Med 1995; 333: 1038-1044. 2. Riddell SR, Watanabe KS, Goodrich JM, et al. Restoration of viral immunity in immunodeficient humans by the adoptive transfer of T-cell clones. Science 1992; 257: 238241. 3. Altman JD, Moss PA, Goulder PJ, et al. Phenotypic analysis of antigen-specific T lymphocytes. Science 1996; 274: 94-96. 4. Keenan RD, Ainsworth J, Khan N, et al. Purification of cytomegalovirus-specific CD8 T cells from peripheral blood using HLA-peptide tetramers. Br J Haematol 2001; 115: 428434. 5. Cobbold M, Khan N, Pourgheysari B, et al. Adoptive transfer of cytomegalovirus-specific CTL to stem cell transplant patients after selection by HLA-peptide tetramers. J Exp Med 2005; 202: 379-386. 6. Peggs KS, Verfuerth S, Pizzey A, et al. Adoptive cellular therapy for early cytomegalovirus infection after allogeneic stem-cell transplantation with virus-specific T-cell lines. Lancet 2003; 362: 1375-1377. 7. Knabel M, Franz TJ, Schiemann M, et al. Reversible MHC multimer staining for functional
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isolation of T-cell populations and effective adoptive transfer. Nat Med 2002; 8: 631-637. 8. Riddell SR, Greenberg PD. Principles for adoptive T cell therapy of human viral diseases. Annu Rev Immunol 1995; 13: 545–586. 9. Einsele H, Roosnek E, Rufer N, et al. Infusion of cytomegalovirus (CMV)-specific T-cells for the treatment of CMV infection not responding to antiviral chemotherapy. Blood 2002; 99: 3916-3922. 10.Casazza JP, Betts MR, Price DA, et al. Acquisition of direct antiviral effector functions by CMV-specific CD4+ T lymphocytes with cellular maturation. J Exp Med 2006; 203: 2865–2877. 11.Rauser G, Einsele H, Sinzger C, et al. Rapid generation of combined CMV-specific CD4+ and CD8+ T-cell lines for adoptive transfer into recipients of allogeneic stem cell transplants. Blood 2004; 103: 3565-3572. 12.Grigoleit GU, Kapp M, Hebart H, et al. Dendritic cell Vaccination in allogeneic stem cell recipients: induction of HCMV-specific CTL responses even in patients receiving a transplant from an HCMV-seronegative donor. J Infect Dis 2007; 196: 699-704. 13.Wang Z, La Rosa C, Mekhoubad S, et al. Attenuated poxviruses generate clinically relevant frequencies of CMV-specific T cells. Blood 2004; 104: 847-856. 14.Wang Z, La Rosa C, Li Z, et al. Vaccine properties of a novel marker gene-free recombinant modified vaccinia Ankara expressing immunodominant CMV antigens pp65 and IE1.Vaccine 2007; 25: 1132-1141. 15.Berger C, Jensen MC, Lansdorp PM, et al. Adoptive transfer of effector CD8 T cells derived from central memory cells establishes persistent T cell memory in primates. J Clin Invest 2008; 118: 294-305.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Why might the transfer of “unmanipulated” virus-specific T-cells (without in vitro expansion) be more efficient in control of viral infection when compared to in vitro expanded cells? a) The in vitro expansion decreases the expression of the pro-apoptotic FAS molecule (CD95) and therefore leads to a shorter survival of the transferred cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The telomere length of specific T-cells after in vitro expansion is reduced, possibly leading to a shorter survival of the adoptive transferred T-cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) In vitro expanded cells have a greater risk of inducing GvHD compared to directly selected cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The TCR of the selected cells is blocked by the selecting agent . . . . . . . . . . . 284
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2. Which risk of adoptive immunotherapy post allogeneic SCT is potentially reduced by long term ex-vivo culture? a) Acute transfusion reactions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Contamination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Costs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which statement is correct regarding an MVA-based vaccines? a) Multiple antigens can be included in one single vaccine . . . . . . . . . . . . . . . . . . . b) MVA soon will be the standard in immunotherapy post-transplant due to a high evidence level based on multiple randomised controlled trials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) These vaccines could be offered only to a limited number of patients because only patients with distinct HLA haplotypes can be vaccinated . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) MVA has a small capacity for foreign DNA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. What is the relevance of HCMV-specific TH cells in immunotherapy post-transplant? a) HCMV-specific TH cell responses are not necessary for the maintenance of an adequate pathogen-specific immune responses. Transfer of a CTL response is always sufficient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Mature HCMV-specific CD4+ T-cells have functional features which equal antiviral CD8+ T-lymphocytes and are important for maintenance of transferred CMV-specific CD8+ T-cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) TH infusions have been demonstrated to cause severe transfusion related reactions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Generation of TH cells is much more expensive and time consuming than generation of CD8+ clones . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which is not a potential risk of the transfer of CMV-specific T-cells? a) Alloreactivity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Increase in viral load . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Transfer of fungal infections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Development of antiviral drug resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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Adoptive cellular immunotherapy to harness post-transplant alloreactivity E. Nadal
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CHAPTER 14.2 • Adoptive cellular immunotherapy
1. Introduction The bidirectional allorecognition responses taking place after HSCT between donor and recipient are responsible for the major complications of the procedure: graft failure and graft versus host disease (GvHD), but also for one of its unique advantages, the graft versus leukaemia (GvL) effect (1). During the past decades the evidence of this anti-leukaemia effect exerted by donor lymphocytes and responsible in part for the success of allogeneic transplantation has led to a change in the classic definition of HSCT as rescue treatment after myeloablation to the more refined one of cellular immunotherapy. This has led in turn to the introduction of reduced intensity conditioning (RIC) regimens as platforms for host versus graft tolerance and subsequent use of donor lymphocytes to exploit the GvL effect, aiming to achieve long-term remissions with low transplant related toxicity. But unfortunately even using reduced intensity conditioning regimens, HSCT is still associated with severe complications due to alloimmune reactions, precluding a wider use of this technique.
2. Alloreactions post-HSCT Alloreactivity after HSCT is a complex process that involves donor T-cells and NK cells interacting with specific recipient target-tissues. This immune response is mediated both by direct lymphocyte-target cell interaction and by cytokines. Alloimmune responses after HSCT are responsible for three major transplant events that determine success or failure of the transplant: engraftment, GvHD, and GvL effects. 2.1. Engraftment/rejection In allogeneic HSCT, graft rejection is predominantly mediated by residual host Tcells with anti-donor specificity. In conventional HSCT donor T-cells usually counteract this host-versus graft effect. However in T-cell depleted HSCT and in nonmyeloablative transplants the incidence of graft failure is increased. In order to achieve a long-lasting engraftment in these settings significant immunosuppression is needed to suppress the recipient immune system and allow the incoming donor cells to grow and eventually to predominate. The use of highly lymphotoxic conditioning regimens (including anti-CD52, fludarabine or anti-thymocyte globulin, amongst others), better post-HSCT immunosuppression and better graft quality have much reduced the incidence of graft failure. 2.2. Graft versus host disease (GvHD) GvHD remains the major cause of mortality and morbidity after allogeneic HSCT and precludes the more extensive use of this procedure. Acute GvHD (aGvHD) is primarily
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a T-cell mediated event where the infused donor T-cells recognise recipient antigens presented by recipient-APCs, leading to tissue damage. This systemic alloimmune response is amplified by the conditioning regimen, especially radiation-based ones, which initiate a proinflammatory cytokine storm including TNF-a and IL-1. A decrease in aGvHD incidence can be obtained eliminating the donor T-cells from the graft, but this approach is associated with a higher incidence in relapse and recurrent life-threatening infections. Chronic GvHD (cGvHD) is a pleiotropic syndrome with similarities to autoimmune diseases such as Sjogren syndrome or scleroderma. The pathophysiology of cGvHD is fundamentally distinct from that of aGvHD and still poorly understood, in part due to a lack of a good animal model. Although donor T-cells are critical in the initiation of cGvHD, it is now established that the effector pathways involve cells of myeloid lineage and fibrogenic cytokines such as TGF-b. 2.3. GvL/graft-versus-tumour effect The graft versus leukaemia or tumour effect (GvL/GvT) refers to the donor anti-host response directed against the leukaemia/tumour cells remaining after the conditioning treatment. GvL accounts for the main advantage of allogeneic HSCT over autologous HSCT. The first evidence for GvL in humans was a report of increased risk of relapse in those patients not developing GvHD after HSCT (2). The separation of these two mirrored graft versus host responses, GvHD and GvL, still remains the holy grail of stem cell transplantation. The GvL reaction may be directed against a leukaemiaspecific targets or minor histocompatibility antigens (mHag) differentially expressed on haematopoietic cells such as HA-1, HA-2, HB-1 and BCL2A1 (3). Donor T-cells are the primary effectors targeting mHags expressed by leukaemia cells as well as normal tissues, thus contributing to both GvHD and GvL. Although some experimental data suggested that CD8+ cells were the major effectors of GvHD, some recent studies imply that CD4+ cells alone are sufficient to induce a GvL response (4). Recently, alloreactive NK cells have also emerged as effectors of GvL in haploidentical transplants. The mechanism by which allorecognition occurs during the GvL response has not been extensively investigated, although it has been reported that donor APC are not required for the GvL response whereas host APC are necessary (5, 6). Several studies have suggested a differential use of cytolytic pathways by GvHD and GvL effectors. The former would preferentially use the Fas-FasL pathway (specially in target-organ GvHD) whereas GvL activity by CD8+ T-cells is mainly dependent on perforin-mediated cytotoxicity. Alternative GvL cytotoxic pathways like TRAIL (TNFrelated apoptosis-inducing ligand) with selective activity for malignant target cells have also been proposed. 288
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3. Adoptive cellular immunotherapy post-transplant 3.1 Donor lymphocyte infusions The best demonstration of the immunotherapeutic effect of alloreactive T-cells is the successful use of DLI for the treatment of relapsed leukaemia after allogeneic HSCT. This is particularly true in the case of CML, whereby post-HSCT relapses can be effectively treated with escalating DLI. Using this approach, around 60–70% of patients re-enter even molecular remission. Other haematological diseases e.g. myeloma and lymphoma, have also been shown to have DLI sensitivity, although to a lesser extent. Variability in the susceptibilities of different types of leukaemia is thought to relate, at least in part, to differences in leukaemic-cell growth kinetics and in GvL effector mechanisms. Unfortunately DLI still involves a risk of GvHD in up to 50% of patients, which can be reduced using an escalating dose approach, delaying the DLI or eliminating CD8 cells from the infusion (7). Another less common complication after DLI is pancytopenia, mainly reported in patients treated in full-blown relapse and with very reduced donor haematopoiesis. The role of DLI not only as a treatment for relapse but also as pre-emptive treatment in RIC transplants is being investigated. 3.2. Suicide-gene-transfected donor T-cells Another potential solution to alloreactivity from unmanipulated T-cells is to transduce the donor T-cells with a suicide gene, so that they can be eliminated if GvHD occurs. The herpes simplex virus 1–thymidine kinase (HSV–TK) gene has been used in several clinical trials without significant acute toxicity and showing dissociation of GvL from GvHD. However, the results are still conflicting and some safety issues need to be addressed before this strategy is consolidated in clinical practice. Other gene modification approaches like timed induction of genes involved in T-cell apoptosis, like FAS or caspase 9, to treat GvHD are currently being investigated. 3.3. Regulatory T-cells Among the major mechanisms involved in transplant tolerance, i.e. central deletion, clonal anergy and suppression/regulation of donor-reactive T-cell clones, the latter has proven to be pivotal in the maintenance of tolerance. Several types of regulatory T-cells have been described, such as agT-cells, NKT-cells, CD8+ and CD4+ T-cells. The later can be divided into two groups, those CD4+ regulatory cells that exert their action secreting inhibitory cytokines like IL-10 or TGF-b such as Tr1 and Th3 cells, and the recently described subset of CD4+ regulatory T-cells expressing the IL-2
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receptor a-subunit (CD25), which exert their suppressive function in a cell-to-cell contact manner. The CD4+CD25+ or regulatory T-cells (Treg) subset is considered to be a crucial population in preserving peripheral tolerance not only to autoantigens but also to foreign antigens, and their role in transplantation tolerance is being increasingly acknowledged. 3.3.1. Naturally occurring CD4+CD25+ Tregs CD4+CD25+ regulatory T-cells were first described in murine systems in 1995 by Sakaguchi et al. (8). They demonstrated that this small thymic derived subset was crucial in preventing autoreactivity, as those mice depleted from these cells developed severe systemic autoimmune disease. In humans Tregs account for 1–2% of circulating CD4+ lymphocytes. In contrast with their murine counterpart human Tregs account mainly for those CD4 with intense CD25 expression (CD4+CD25high). Tregs co-express preferentially other surface markers like GITR (glucocorticoidinduced TNFR-related protein), CD62-L, CTLA-4 or CD152 or CD45RO. However, none of these markers is specific of Tregs and can be also up-regulated in activated T-cells. The most specific Treg marker available to date is the forkhead transcription factor FOXP3, which has proven to be central in the development and function of CD25+ Tregs in both mice and humans. Although it is known that Tregs exert their suppressive activity in a cell-to-cell contact manner, the exact mechanism remains to be elucidated. In mouse models Tregs have shown to prevent and control GvHD in HLA-mismatched transplants when co-infused with conventional T-cells in a 1:1 ratio. In tumourbearing mice the infusion of allogeneic Tregs alone was unable to exert a GvL effect and notwithstanding the absence of GvHD those animals died of disease progression. However some studies have demonstrated that co-infusion of allogeneic Tregs and effector cells prevented GvHD while preserving GvL, suggesting a distinctive pathway of cell killing for those populations (9). The role of Tregs in clinical HSCT remains unclear and trials to address this are ongoing. If as in mouse models they demonstrate to be crucial in preventing GvHD then the infusion of ex vivo expanded Tregs could be a plausible treatment for this life threatening condition, although recent publications indicate that in humans Tregs may have a short life-span after infusion. If on the other hand Tregs are found to suppress anti-tumour immune responses in the transplant setting then their depletion from the DLI product could be advisable to reach higher remission rates. 3.4. NK cells Both clinical and experimental evidence support a role for NK cells as GvL effectors. Human leukocyte antigen (HLA)-C is now known to be the most important MHC class 290
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I subgroup involved in NK inhibition through killer inhibitory receptors (KIRs). In the setting of HLA-mismatched haploidentical HSCT for AML, donor NK clones fail to encounter their class I inhibitory KIR ligands, resulting in the killing of host leukaemic cells. An advantage of using alloreactive NK cells as anti-tumour effectors is that, unlike T-cells, they do not cause GvHD. Clinical trials to address the role of NK-cell-mediated anti-tumour reactivity in the context of KIR-ligand-mismatched allo-HSCT have obtained promising results (review in (10)). 3.5. Mesenchymal stem cells (MSCs) MSCs are multipotent cells with capacity to differentiate in vitro and in vivo into several mesenchymal tissues, such as bone, cartilage and fat. Although bone marrow is the main source for MSCs they can also be isolated from blood, adipose tissue, foetal tissue and cord blood. They play a crucial role maintaining the marrow stroma and they have shown in animal models to enhance engraftment after autologous and allogeneic HSCT. Several studies also support their role in restoring myeloid, lymphoid and megakaryocytic lineages. Importantly, MSCs also disclose strong inhibition of T-cell proliferation to alloantigens in vitro. Autologous and allogeneic MSCs disclosed low immunogenecity and are safe to infuse in humans. Thus, MSCs may be used to enhance haematopoietic three-lineage engraftment and to prevent graft rejection and GvHD in HSCT. Their exact mechanism of action is unclear; however, a handful of reports already support the use of MSC to treat refractory GvHD and recurrent graft failure; trials to confirm their role in this setting are ongoing (review in (11)). 3.6. Cellular vaccines The lymphopenic environment and “enhanced” cytokine milieu that immediately follows HSCT favours the expansion of T-cell clones, supporting the use of post HSCT vaccination. Using specific CTLs directed against tumour proteins (e.g. BCR–ABL, PR1 and WT1) or minor antigens is a plausible way of enhancing the GvL effect after transplant for haematologic malignancies. However, before vaccination can be applied effectively in HSCT recipients it will be necessary to develop methods for selectively preventing aGvHD and thus eliminating the need for post-transplant immunosuppression. The combination of the potent GvL effect of the allograft with a vaccine boost for leukaemia-specific T-cells could prove to be a highly effective strategy to control refractory leukaemias. Moreover adoptive immunotherapy with donor-derived HA-1 CTLs in combination with HSCT could also become an attractive treatment for solid tumours. Another vaccination approach is DC-based immunotherapy after allogeneic stem cell transplantation, used to enhance GvL reactions without aggravation of GvHD. The
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experience so far from phase I/II clinical trials shows the feasibility of DC vaccination using various target antigens like bcr/abl or patient-specific idiotypic proteins, but the clinical benefit is still unclear.
4. Future directions Adoptive T-cell immunotherapy with DLI is already an established and efficacious treatment of post-HSCT relapse in certain settings. New strategies aiming at more specific responses against relapsed or persistent leukaemias and tumours following HSCT and at inhibiting GvHD are being explored. These include depletion or infusion of selected cell populations, genomic modification and vaccination. However, several studies have already identified difficulties with implementing such strategies including inadequate cell persistence or expansion in vivo. A better understanding of the mechanisms involve in the GvL/GvHD immunobiology postHSCT together with an improve methodology for T-cell expansion and manipulation is crucial to eventually harnessing post-transplant alloreactivity and improving patient outcome.
References 1. Horowitz MM, Gale RP, Sondel PM, et al. Graft-versus-leukemia reactions after bone marrow transplantation. Blood 1990; 75: 555-562. 2. Weiden PL, Flournoy N, Thomas ED, et al. Antileukemic effect of graft-versus-host disease in human recipients of allogeneic-marrow grafts. N Engl J Med 1979; 300: 1068-1073. 3. Marijt WA, Heemskerk MH, Kloosterboer FM, et al. Hematopoiesis-restricted minor histocompatibility antigens HA-1- or HA-2-specific T cells can induce complete remissions of relapsed leukemia. Proc Natl Acad Sci USA 2003; 100: 2742-2747. 4. Alyea EP, Soiffer RJ, Canning C, et al. Toxicity and efficacy of defined doses of CD4(+) donor lymphocytes for treatment of relapse after allogeneic bone marrow transplant. Blood 1998; 91: 3671-3680. 5. Shlomchik WD, Couzens MS, Tang CB, et al. Prevention of graft versus host disease by inactivation of host antigen-presenting cells. Science 1999; 285: 412-415. 6. Mapara MY, Kim YM, Wang SP, et al. Donor lymphocyte infusions mediate superior graftversus-leukemia effects in mixed compared to fully allogeneic chimeras: A critical role for host antigen-presenting cells. Blood 2002; 100: 1903-1909. 7. Porter D, Levine JE. Graft-versus-host disease and graft-versus-leukemia after donor leukocyte infusion. Semin Hematol 2006; 43: 53-61. 8. Sakaguchi S, Sakaguchi N, Asano M, et al. Immunologic self-tolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J Immunol 1995; 155: 1151-1164. 9. Edinger M, Hoffmann P, Ermann J, et al. CD4+CD25+ regulatory T cells preserve graft-versustumor activity while inhibiting graft-versus-host disease after bone marrow transplantation. 292
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Nat Med 2003; 9: 1144-1150. 10.Ruggeri L, Mancusi A, Burchielli E, et al. Natural killer cell recognition of missing self and haploidentical hematopoietic transplantation. Semin Cancer Biol 2006; 16: 404–411. 11.Dazzi F, Ramasamy R, Glennie S, et al. The role of mesenchymal stem cells in haemopoiesis. Blood Rev 2006; 20: 161-171.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. One of the following is considered to date the most specific marker for human regulatory T-cells: a) GITR. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) CTLA-4 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) FR4 (folate receptor 4) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) FoxP-3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which of the following is true for mesenchymal stem cells (MSC)? a) MSC can only be isolated form cord blood . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) MSC are highly immunogenic . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) MSC inhibit T-cell proliferation to alloantigens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) MSC have shown to promote graft failure in murine systems . . . . . . . . . . . . . . 3. One of the following minor histocompatibility Ag (mHAg) expression is restricted to haematopoietic cells: a) HA-3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) HA-2 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) B60/HY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) A1/HY. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which subset of lymphocytes has demonstrated to enhance GvL reactions in haploidentical SCT? a) Recipient CD8+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) ag lymphocytes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Alloreactive NK cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Tregs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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5. One of the following strategies would not be useful to enhance GvL responses: a) Infusion of KIR-mismatch alloreactive NK cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Infusion of ex vivo expanded Tregs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) DC vaccination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) WT-1 peptide vaccination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 15
Immune reconstitution after allogeneic HSCT
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1. Introduction Assessment of the host immune status is becoming a key issue in allo-HSCT, especially in the long-term follow-up of these patients, because severe posttransplant infections, relapse or secondary malignancies may be directly related to persistent immune defects. Immune deficiency leading to an increased susceptibility to infections lasts for more than a year. In relation to the occurrence of infections, the post-transplant period is subdivided in different phases (see Figure 1 and Chapter 10). Although infections that occur in the first month mostly result from a deficiency in both granulocytes and mononuclear cells (MNC), later post-engraftment infections are due to a deficiency in MNC subsets, primarily CD4 T-cells and B-cells. T-cell reconstitution has been extensively studied because of the central role of Tcells in mediating both GvHD, evidenced by the reduced incidence of this complication following TCD, and a GvL effect as shown by DLI. DLI may cure 20–80% of patients with post-transplant relapsed leukaemia and lymphoma depending on the type and
Figure 1: The time course of infections after allogeneic HSCT
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extent of the disease. This is one of the most important breakthroughs in HSCT in the last years illustrating the powerful anti-leukaemia effect mediated by allogeneic lymphocytes and the potential of immunotherapy in the treatment of malignant diseases.
2. General principles In transplants performed following myeloablative conditioning regimens, immune reconstitution (IR) will depend upon the ability of the haematopoietic graft to generate de novo lymphoid and myeloid lineage cells and on the function of mature cells contained in the graft. Post-transplantation, the different MNC populations reconstitute at different tempos. The first cells to reconstitute (within first 100 days) are those of the innate immune response, granulocytes, monocytes, macrophages and NK cells. In contrast, T and B-lymphocytes remain severely reduced and their function is impaired for several months or years after HSCT. IR of these various lymphocyte populations will be analysed separately with an emphasis on T-cell reconstitution.
3. Main factors affecting IR Host factors
Age, sex, conditioning regimen, initial pathology
Genetic differences
The degree of genetic differences between donor and recipient including HLA, minor histocompatibility Ag and genes associated with immune responses to microorganisms (see Chapter 3).
Source of HSC The type of HSC, either unmanipulated or TCD BM, PB or CB has an impact on IR. This is an important parameter, for instance recipients of PBSC, who receive at least 10 times more lymphocytes than recipients of BM, have higher lymphocyte-subset counts and fewer infections (1). IR in CB transplants is slow but progressively reaches in the longterm (>2 yrs) even better values than after BM grafts (2). Post-HSCT events
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May in turn have a worsening effect on IR, especially aGvHD and cGvHD, relapse and infectious complications (EBV or CMV viruses, fungal infections, toxoplasmosis) either directly or through drugrelated side effects.
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4. Assessment of B-cell reconstitution 4.1. B-lymphocyte phenotyping with B-lineage markers (CD19, CD20, CD21) and activation or differentiation markers (CD5, CD27) CD19+ B-cells normalise by one year after transplant. B-cell regeneration may be associated with transient appearance of monoclonal B-cell expansions. 4.2. Quantification of serum total IgG, IgM and IgA and of IgG subclasses After a decline in the first few months after HSCT, levels of specific antibodies to protein Ag frequently encountered after transplantation (e.g. CMV) return to pretransplantation levels within 1 yr. In contrast, antibodies to protein Ag that are unlikely to be encountered after HSCT (e.g. tetanus, measles, polio) continue to decline. This supports the recommendation of post-HSCT vaccination. Antibody levels in the first year, are affected primarily by pre-HSCT antibody levels in the recipient (3). A persistent defect in IgA, especially in patients with cGvHD explains mucosal infections of the respiratory and digestive tracts. IgG2 and IgG4 subclasses are also deficient in the case of GvHD, accounting for the increased susceptibility to infections, primarily those due to encapsulated bacteria (e.g. Streptococcus pneumoniae or Haemophilus influenzae). PBSC recipients do not have higher antibody levels than BM recipients. 4.3. Vaccinations Vaccinations with inactivated or conjugated vaccines (see Chapter 10) should be initiated when CD4 and B-lymphocyte counts are sufficient to expect efficacy, usually from 6 months post-transplant onwards.
5. NK-cell reconstitution NK-cells are lymphocytes that act early in the immune response against infection and tumour-transformed cells. Based on phenotyping (CD16 and CD56), they are the first lymphocyte subpopulation to be reconstituted in all graft settings, usually within 3 months. The genetic organisation and function of NK receptors, either inhibitory or activating, has been unravelled in the past few years. NK-cell receptors are encoded by 2 structurally distinct families of molecules: The killer immunoglobulin-like receptors (KIR) and the lectin-like CD94:NKG2 heterodimers. Every NK-cell expresses at least one inhibitory receptor specific for autologous HLA Class I, thereby ensuring selftolerance. As KIR and HLA segregate independently and as unrelated individuals almost always have different KIR genotypes, we predict that approximately 25% of
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transplants between HLA-identical siblings involve KIR identity and approximately 75% KIR disparity. For transplantation with an HLA-MUD, the frequency of KIR incompatibility approaches 100%. In the haploidentical TCD graft setting, a beneficial effect of KIR disparity on GvL has been evidenced (4). This is an important finding which needs to be extended to other types of grafts and which may change our current criteria of donor/recipient matching based on HLA compatibility (see Chapter 3). There are still few studies directly assessing NK reconstitution at the level of KIR and lectin-like NK receptors expression level and function (5). This study and others showed that CD94:NKG2A may be expressed earlier than KIR and that most patients reconstitute a donor-type NK repertoire depending on their KIR genotype. Different NK subsets are now more precisely defined and especially the CD56brightCD16and CD56dimCD16+, respectively prone to cytokine production or cytotoxicity. The rapid recovery of NK-cells after graft is due to an expansion of CD56brightCD16-. The precise role of this NK subset in GvHD and GvL is a key issue in HSCT.
6. T-cell reconstitution 6.1. Naïve and memory T-cells Memory T-cells are the first to expand after HSCT; they may be either of donor origin in the case of a non-TCD BM or, in the case of a TCD, originate from host T-cells that have survived the conditioning regimen (6). They respond quickly to previously encountered pathogens, are easier to trigger, faster to respond and enter tissues more readily than naïve T-cells. They are frequently directed towards periodically reactivated herpes viruses, CMV or EBV, which they keep under control. They constitute the majority of oligoclonal T-cell expansions found in healthy adults, especially in the CD8+ population. They are also less dependent than naive T-cells upon recognition of self MHC-peptide complexes in their survival and expansion in the periphery. In the long term, broad immune responses need the reconstitution of a naïve T-cell repertoire able to respond to a broad range of pathogens encountered by the host and to tumour antigens. Reconstitution of this compartment is an ongoing process which requires a functional thymus for the recovery of a complete T-cell ontogeny. The thymus itself may be a target of the alloreactive immune attack with possible consequences on thymic selection, escape of self-reactive T-cell clones and perpetuation of GvHD. This has been well documented in animal models (7) and deserves more insight in humans. 6.2. How to evaluate naïve and memory T-cell populations? The current immunological tests assess naïve and memory lymphocyte populations: 300
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CD45RO for memory T-cells and CD45RA or CD62L for naïve T-cells are the most usual markers. However, naïve T-cells may undergo expansion without phenotypic changes and have a long lifespan, up to 20 yrs. In addition, memory CD45RO+ T-cells may revert to a naive CD45RA+ phenotype, especially in case of persistent infection with herpes viruses. Therefore, other markers should be added to definitely assess naïve T-cells and the different categories of memory T-cells. CCR7, a molecule involved in the homing of T-cells to lymph nodes is especially valuable. A combination of these markers allows the definition of: - Naïve T-cells: CD45RAhighCD45RO-CCR7+CD28+ - 2 populations of CD8+CD45RA- memory cells: - CCR7+ “central memory”, expressing L-selectin (CD62L) - CCR7- “effector memory”, L-selectin-, IL-2 dependent, which migrate to inflammatory sites and secrete IFN-g. T-cell diversity (“T-cell repertoire”) and thymic function can be directly evaluated. The size of the T-cell repertoire and the extent of T-cell diversity has been measured only recently, the value of about 25 x 106 different TCR ab complexes being lower than that was previously estimated. T-cell diversity is contributed by the naïve population and in healthy adults memory T-cells, which account for approximately 1/3 of the total T-cells, contribute to less than 1% of the abT-cell diversity. A practical consequence for HSCT is that evaluation of T-cell repertoire diversity reflects the extent of the naïve T-cell compartment. Various approaches known as “Immunoscope” or “spectratyping” may be used. They are based on the size diversity analysis of the CDR3 b-chain region as an index of the diversity of the whole abT-cell population. T-cell repertoire diversity in allo-HSCT recipients is a function of: - Number and diversity of infused T-cells with the graft (TCD, age of the donor, source of graft) - Residual T-cells present in the recipient - Thymic pathway of regeneration, for which the age of the recipient is the main factor - Immunosuppressive treatment and complications (GvHD, viral infections). Overall, early after HSCT (within 6 months after graft) many abnormalities of the T-cell repertoire are demonstrable but are difficult to correlate with the clinical status of the patient. Conversely, later after the graft (after 1 yr at least) and ongoing for at least 2 to 3 yrs post-transplant, it is possible to correlate repertoire disturbance with the occurrence of GvHD, severe infectious complications or relapse. T-cell repertoire reconstitution is delayed in case of TCD or in CD34+ purified grafts and is improved where there is full donor haematopoiesis. Techniques of TCR b-chain sequencing have clearly separated T-cell clones mediating GvHD and GvL and could be used in the future to monitor GvHD-causing clones in HSCT recipients (8). HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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The reconstitution and maintenance of a diverse repertoire in the peripheral lymphocyte pool is dependent on the generation of functional thymocytes throughout adult life. These recent thymic emigrants can now be evaluated by measuring the episomal DNA excision circles of the TCR d locus deleted during recombination of the a locus in all functional ab T-cells (known as “TREC” for T-cell receptor rearrangement excision DNA circles). This ex vivo marker of thymic function has been used in allo-HSCT monitoring: - TREC levels are low until 3–6 months after allo-BMT. Low TREC values are associated with increasing patient age and TCD but mainly with GvHD (9), leukaemia relapse or opportunistic infections - High TREC levels and a broad T-cell repertoire have been associated with an efficient IR after CB transplantation in the long term (2) although there is some delay in IR in that setting (10). The thymic function of the recipient before graft could be associated with a more favourable outcome in terms of survival, GvHD and bacterial or viral infections (11). It could be a valuable prognostic factor predicting IR after transplant. 6.3. How is the Ag-specific immune response reconstituted after allo-HSCT? Naïve T-cell reconstitution is a key issue for the long-term recovery of immune responses but memory T-cells are also needed for an efficient and timely response towards pathogens. Therefore, especially in some graft settings (TCD, CBT, HLA mismatch UD) adoptive immunotherapy can be used in an attempt to compensate for the lack of Ag specific immunocompetent T-cells. In order to do this, it is necessary to evaluate patients at risk and to be able to monitor Ag specific immune responses towards pathogens. Herpes viruses (CMV, EBV) are of primary importance in HSCT because reactivation of EBV can result in potentially fatal EBV-associated lymphoproliferative disease and because of the frequency of late CMV reactivation in the host even under pre-emptive therapy. It is possible: - To monitor EBV and CMV-specific cytotoxic responses by Elispot functional assays or intracellular cytokine staining which are easier to perform than the conventional 51Cr release cytotoxic assay in a routine laboratory - To use the tetramer technology to stain directly ex vivo CD8+ T-cells reactive with peptide/HLA complexes and to characterise these cells in terms of phenotype and function. This is a very sensitive method which can stain 1/5000 CD8+ T-cells (or 1/5 x 104 PBMC). It may also be used to isolate Ag-specific T-cells by cell sorting and to expand them in vitro. It is becoming a routine laboratory analysis for CMV (12) and EBV (13) specific CD8+ T-cell responses.
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7. From monitoring to immune intervention The combination of these various structural (Immunoscope for TCR diversity analysis, TREC for thymic function) and functional approaches (Elispot, tetramer staining) should enable more precise immune monitoring of patients at risk of relapse or persistent severe infectious complications, particularly in the context of adoptive immunotherapy. The tetramer approach has direct benefits for the identification of patients with impaired CD8+ specific cytotoxic responses who may be eligible for cellular adoptive immunotherapy against CMV (14) or for CD20-specific MoAb treatment (rituximab) to prevent post-HSCT lymphoproliferative disorder during EBV reactivation (13). The transfer of viral-specific CTL is possible with tetramer-sorted CTL without culture (15). Based on experimental models, other attempts could be pursued to improve IR and graft outcome: - Improve thymic function recovery, by growth factors such as KGF, Flt3l or androgen blockade - Selective depletion of alloreactive T-cells from donor lymphocytes (16) - Use of minor histocompatibility Ag as tumour Ag to mediate GvL effect, as described for HA-1 minor antigen (17) - Genetic modification of T-lymphocytes with TK suicide gene (18) - Haploidentical NK immunotherapy (19) - Immune modulation through alternative stem cell sources (mesenchymal stem cells), dendritic cells or Treg manipulation. Manipulation of immune system homeostasis to facilitate the emergence of regulatory CD4+CD25high T-cells (or Treg) which have been shown in animal models to control GvHD without impairing GvL (20). In humans, although evidence for the role of Treg has been less clear, it appears that an in situ defect in these populations could be associated with aGvHD (21). Importantly, Treg could be expanded in vitro and keep their functional properties, thus being an approach of choice in the future for controlling GvHD.
Acknowledgments This work was supported by research grants from the Cancéropôle Ile-de-France, EC programs EUROBANK, EUROCORD and FP6 ALLOSTEM (#503319).
References 1. Storek J, Dawson MA, Storer B, et al. Immune reconstitution after allogeneic marrow transplantation compared with blood stem cell transplantation. Blood 2001; 97: 3380-3389.
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2. Talvensaari K, Clave E, Douay C, et al. A broad T-cell repertoire diversity and an efficient thymic function indicate a favorable long-term immune reconstitution after cord blood stem cell transplantation. Blood 2002; 99: 1458-1464. 3. Storek J, Viganego F, Dawson MA, et al. Factors affecting antibody levels after allogeneic hematopoietic cell transplantation. Blood 2003; 1001: 3319-3324. 4. Farag SS, Fehniger TA, Ruggeri L, et al. Natural killer cell receptors: New biology and insights into the graft-versus-leukemia effect. Blood 2002; 100: 1935-1947. 5. Shilling HG, McQueen KL, Cheng NW, et al. Reconstitution of NK cell receptor repertoire following HLA-matched hematopoietic cell transplantation. Blood 2003; 101: 3730-3740. 6. Roux E, Dumont-Girard F, Starobinski M, et al. Recovery of immune reactivity after T-celldepleted bone marrow transplantation depends on thymic activity. Blood 2000; 96: 2299-2303. 7. Hauri-Hohl MM, Keller MP, Gill J, et al. Donor T-cell alloreactivity against host thymic epithelium limits T-cell development after bone marrow transplantation. Blood 2007; 109: 4080-4088. 8. Michalek J, Collins RH, Hill BJ, et al. Identification and monitoring of graft-versus-host specific T-cell clone in stem cell transplantation. The Lancet 2003; 361: 1183-1185. 9. Weinberg K, Blazar BR, Wagner JE, et al. Factors affecting thymic function after allogeneic hematopoietic stem cell transplantation. Blood 2001; 97: 1458-1466. 10.Komanduri KV, St John LS, de Lima M, et al. Delayed immune reconstitution after cord blood transplantation is characterized by impaired thymopoiesis and late memory T cell skewing. Blood 2007; 110: 4543-4551. 11.Clave E, Rocha V, Talvensaari K, et al. Prognostic value of pretransplantation host thymic function in HLA-identical sibling hematopoietic stem cell transplantation. Blood 2005; 105: 2608-2613. 12. Aubert G, Hassan-Walker AF, Madrigal JA, et al. Cytomegalovirus-specific cellular immune responses and viremia in recipients of allogenic stem cell transplants. J Infect Dis 2001; 184: 955-963. 13.Clave E, Agbalika F, Bajzik V, et al. Epstein-Barr virus (EBV) reactivation in allogeneic stemcell transplantation: Relationship between viral load, EBV-specific T-cell reconstitution and rituximab therapy. Transplantation 2004; 77: 76-84. 14.Peggs KS, Verfuerth S, Pizzey A, et al. Adoptive cellular therapy for early cytomegalovirus infection after allogeneic stem-cell transplantation with virus-specific T-cell lines. The Lancet 2003; 362: 1375-1377. 15.Cobbold M, Khan N, Pourgheysari B, et al. Adoptive transfer of cytomegalovirus-specific CTL to stem cell transplant patients after selection by HLA-peptide tetramers. J Exp Med 2005; 202: 379-386. 16.Amrolia PJ, Muccioli-Casadei G, Huls H, et al. Adoptive immunotherapy with allodepleted donor T-cells improves immune reconstitution after haploidentical stem cell transplantation. Blood 2006; 108: 1797-1808. 17. Mutis T, Blokland E, Kester M, et al. Generation of minor histocompatibility antigen HA-1specific cytotoxic T cells restricted by nonself HLA molecules: A potential strategy to treat relapsed leukemia after HLA-mismatched stem cell transplantation. Blood 2002; 100: 547-552. 304
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18.Bondanza A, Valtolina V, Magnani Z, et al. Suicide gene therapy of graft-versus-host disease induced by central memory human T lymphocytes. Blood 2006; 107: 1828-1836. 19.Ruggeri L, Capanni M, Urbani E, et al. Effectiveness of donor natural killer cell alloreactivity in mismatched hematopoietic transplants. Science 2002; 295: 2097-2100. 20.Edinger M, Hoffmann P, Ermann J, et al. CD4+ CD25+ regulatory T cells preserve graftversus-tumor activity while inhibiting graft-versus-host disease after bone marrow transplantation. Nature Medicine 2003; 9: 1144-1150. 21.Rieger K, Loddenkemper C, Maul J, et al. Mucosal FOXP3+ regulatory T cells are numerically deficient in acute and chronic GvHD. Blood 2006; 107: 1717-1723.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1.
After allo-HSCT the earliest lymphocyte population(s) to recover: a) NK lymphocytes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) T CD4+ naïve T-cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) T CD8+ memory T-cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All at the same time . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.
Among the following lymphocyte phenotypic marker(s), which one is the most precise to define memory T-cells: a) CD45RA+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) CD45RO+ CCR7- . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) CD45RA+ CCR7+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) CD16+ CD56+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.
Which is the main factor directly affecting thymic recovery after allo-HSCT? a) Sex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) CMV infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) HLA mismatch . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4.
After allogeneic HSCT, the risk of EBV-induced proliferative disease (PTLD) is especially increased in case of which one of the following: a) T-cell depletion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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b) Genoidentical sibling donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Sex-mismatch between donor and recipient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Aplastic anaemia as primary disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5. Among the following lymphocyte phenotypic marker(s), which is the most precise to define naïve T-cells: a) CD45RA+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) CD45RO+ CCR7- . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) CD45RA+ CCR7+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) CD16+ CD56+ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 16
Psychosocial aspects of HSCT
A. Kiss, M. Kainz
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CHAPTER 16 • Psychosocial aspects of HSCT
1. Introduction HSCT has moved from an experimental treatment to become an accepted therapy. At the same time, the primary focus on increasing survival in previously lethal diseases has been enlarged to encompass psychosocial issues such as quality of life. In this Chapter, specific psychosocial aspects of the patient, the donor, the family, and the transplant team will be discussed in relation to the time trajectory of HSCT shown in Figure 1.
Figure 1: Trajectory of HSCT
Diagnosis Decision to transplant Donor search
HSCT In-hospital treatment Side effects, Toxicity Engraftment
Short-term follow-up Frequent controls Tx-related mortality GvH disease
Long-term follow-up Quality of life Return to "normal life" Relapse
2. Psychosocial aspects - the patient 2.1. Psychosocial morbidity The diagnosis of a deadly disease and the option of a treatment that may cure but which also has potentially lethal side effects puts the patient under heavy pressure and adjustment is difficult. Psychosocial morbidity is frequent, particularly adjustment disorders with symptoms of depression and anxiety. Approximately one third of patients report significant symptoms of intrusive and avoidance stress responses (1). The search for a suitable related or UD in allo-HSCT is a hard time for patients, for fear of not finding a donor. Coping mechanisms vary according to individual patients from fighting spirit to hopelessness and helplessness. Psychosocial evaluation systems such as the Transplant Evaluation Rating Scale (TERS) (2) have been proposed for assessing the psychosocial functioning of HSCT recipients, which could then allow early intervention in case of poor functioning. Psychiatric morbidity prolongs hospital stay independently of in-hospital somatic risk factors (1) and pre-transplant physical and mental functioning is strongly associated with self-reported recovery from stem cell transplantation (3). The greatest emotional distress occurs after admission to hospital and before the transplantation. Anxiety and depression decrease one week after the transplant. Psychosocial well-being after the transplant is heavily influenced by mucositis, toxicity, and other side effects, but conversely psychological factors such as anxiety, BMT-related distress, and social support also have a significant impact
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on how severely patients experience their mouth pain. Beside the usual medical treatments, psychosocial interventions such as hypnosis and muscle relaxation can substantially reduce nausea and pain. After discharge, many patients are disappointed by their low energy level (fatigue), high susceptibility to infections and the very slow return to normal life. In the first year after transplant psychological distress declines (4). However, elevated levels of anxiety and depression prior to transplantation predict more anxiety and depression in the follow-up period (4). Somatic risk factors for outcome after HSCT are well described, and include factors relating to the patient, the donor and the type of transplant. However coping strategies assessed pre-transplant such as emotional support, acceptance, taking control, and compensation also seem to have an influence. Compensation is reported to be associated with shorter, the other strategies with longer survival. Replication of the data is essential before clinical recommendations can be made (5). Patients suffering from cGvHD have lower QoL and may therefore be more vulnerable to depression and anxiety disorders. At long-term follow-up, some survivors still have to cope with a low energy level, some with fear of losing their job, and all have to deal with infertility and the fear of relapse and secondary malignancies. At 3 yrs post transplantation 80% of women and 29% of men report sexual problems (6). 2.2. Quality of life (QoL) Good QoL in HSCT has been reported repeatedly. However, it should be kept in mind that QoL is a poorly defined concept and instruments professing to measure QoL measure different things. The approaches that are most frequently used are shown in Table 1.
Table 1: Different approaches to the measurement of QoL in HSCT Approach
Examples
Generic
SF-36 (Medical Outcomes Survey - Short Form 36; EORTC QLQ-C30 (European Organisation for Research and Treatment of Cancer Core Quality of Life questionnaire), FACT (Functional Assessment of Cancer Therapy)
Disease-specific Leukaemia/BMT module of EORTC QLQ-C30, FACT-BMT (Bone Marrow Transplantation)
310
Utilitarian
TTO (Time Trade-Off)
Individualised
SEIQoL-DW (Schedule for the Evaluation of Individual QoL - Direct Weighting)
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A standardised multidimensional questionnaire (physical, psychological, functional and social dimensions) is often used in combination with a disease-specific module (7). Recently, a high-dose chemotherapy questionnaire module to supplement the European Organisation for Research and Treatment of Cancer Core Questionnaire (EORTC QLQ-C30) has been developed (8). The advantage of a generic approach is to obtain a numeric figure for a specific item, which can be used to assess the impact on QoL across different diseases or to assess changes in the same patient over time. However, the individual perception of a given symptom is neglected by these questionnaires: e.g. not being able to climb two flights of stairs may represent a substantial impairment for one patient but may have no impact on another patient. The utilitarian approach has only been used in solid organ transplantation but not for HSCT, as far we know. In this approach it is left to the patient to decide the areas in which the health-related QoL is compromised, as well as the severity of impairment. However, no information about the characteristics of the compromised dimension is given. An individualised approach not only lets the patient define the specific domains which are most important for his or her QoL, but also allows the patient to weight the importance of these domains individually (9). Such areas included positive aspects, e.g. a changed view of life and oneself (10). Cancer-related fatigue is one of the biggest constraints of QoL after HSCT. The majority of studies are cross-sectional and restricted to short follow-up. Compared to normal controls, HSCT survivors at a mean of 7 years after transplantation reported poorer physical, psychological, and social functioning but, conversely, more psychological and interpersonal growth, differences that appeared to persist many years after HSCT (11). In a study in survivors 10 years after HSCT, health problems were not focused on specific diseases or limited to survivors with readily identifiable risk factors. Musculoskeletal problems were frequent. Survivors require screening for sexual problems, urinary frequency, mood and need for antidepressants or benzodiazepines (12). However, it should be kept in mind even if QoL is measured more frequently nowadays, the data are often not understandable for the physician involved and therefore not used in the decision-making process with the patient (13). 2.3. Neglected issues 2.3.1. The patient with delirium Half of patients who undergo HSCT experience an episode of delirium during the 4 weeks post transplantation. Symptoms of delirium are poorly recognised by professionals because of the variability in the symptoms of confusion over the 24-
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hour period and also the patients’ tendency to dissimulate, as they do not want to be identified as “mad”. If symptoms are recognised there is a tendency to label them as difficulty in coping with an unbearable situation rather than a diagnosis of delirium. Transplant physicians are often inexperienced in the treatment of delirium, which can be successfully managed with a low dose of narcoleptics. Compared to patients without delirium, patients who experience delirium during myeloablative HSCT showed impaired neurocognitive abilities and persistent distress 80 days after transplantation (14). The long-term effect of delirium post HSCT remains to be determined. 2.3.2. The dying patient A substantial number of patients die despite HSCT and they represent the primary sources of stress in nurses and doctors working on a transplant unit (15). In a recent study about Advanced Care Planning (ACP) in HSCT patients, those patients least likely to have planned for poor outcomes were the ones most likely to face them (16). 2.3.3. The non-compliant patient Non-compliance in patients transplanted for solid organs is one the main factors of transplant failure. The prevalence of non-compliance in HSCT recipients is unknown (17). Given the complex medical regimen and the strict dietary and behavioural rules after transplantation, the medical team may incorrectly ascribe poor therapeutic outcomes to inadequacies in the regimen instead of non-compliance of the patients. They may prescribe more potent medicine with the potential for greater adverse reactions (17). Poverty is a potential factor in non-compliance that is often neglected: In the US underinsured/poor HSCT recipients have the same mortality during inpatient treatment as the non-poor, but a significantly higher mortality in the following 100 days. The authors hypothesise that the higher mortality is due to the poor patients’ inability to comply with or seek medical care because of deficient socioeconomic resources (18).
3. Psychosocial aspects - the donor 3.1. Related donor In contrast to living solid organ donors little is known about the decision-making process of HSC donors. Most donors do not even recognise a decision-making process, they have “no choice”, they just want to help the recipient (19). They believe the psychological aspects of the procedure outweighs the physical aspects of 312
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CHAPTER 16 • Psychosocial aspects of HSCT
donation (19). Siblings of unsuccessful transplant recipients may feel guilty for their “bad marrow”, some donors have problems in coping, and difficulties within the affected families are not uncommon. 3.2. Unrelated donors Medical advances have made possible HSCT from UD. Many donor registries have been established in recent years. The motivations of UD donors may be different from related donors, as they do not know the recipients. Higher levels of ambivalence about BM donation are associated with joining during a recruitment drive for a specific patient, perceiving the recruitment staff as less informative, being discouraged from joining by others and not having an intrinsic commitment to donate (20). Unrelated female donors are mostly motivated by positive feelings, empathy and the desire to help someone.
4. Psychosocial aspects - the family The most significant social support for HSCT recipients comes from their family. Family members experience the similar distress as do the patients and report more impairments in family relationship than patients (21). Physical and emotional recovery after HSCT depends on the quality of family relationships as perceived by the recipient. They seem to have the most important role as a filter for stress. In a recent study in HCT survivor/partner pairs (n=177) spouses/partners experience similar emotional and greater social long-term costs of cancer and HCT than survivors without the potential compensatory benefits of post-traumatic growth (22).
5. Psychosocial aspects - the transplant team 5.1. Present and future of the transplant team There is great economic pressure for transplant teams to carry out more transplant procedures in less time with the same or even reduced staff. Earlier discharge and frequent unscheduled readmission may result. 5.2. Psychosocial well-being of team members The very nature of HSCT is characterised by the dominance of technology and the rapidity of decision-making and practice. Death is considered as a failure and is often due to the toxicity of the procedure itself or infections (23). Both doctors and nurses consider regular work with dying patients as the primary source of stress, besides other stressors such as interpersonal staff conflicts, excessive responsibilities and
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highly demanding patients and families (15). The death of a patient may be considered as a mismatch of high investment (high costs, time, emotional labour) and low returns (causing suffering without survival). Burnout in nursing and medical staff is high, half of them being emotionally exhausted, and 80% reporting feelings of low personal accomplishment (15). Doctors and nurses reported that the most frequent effects from prolonged stress were increased illness, reduced productivity and increased clinical errors. 5.3. Care for/of the team As the care of the patient is the primary focus for transplant teams, their own needs for care is are often denied and seldom addressed. Burnout must be considered as a permanent danger and not as an individual problem of the team member concerned. Time and resources have to be devoted to the care of the team as a prophylactic measure and not only after a team member has cracked under the pressure.
6. Personal conclusions and practical applications Psychosocial issues in patients, donors, families and transplant teams are not “soft data” but have a substantial impact on the morbidity of all persons involved and probably on the survival of patients. Communication and psychosocial skills are core competencies for doctors and nurses. They cannot be delegated to mental health professionals. Evidence-based training in these skills is available but seldom used by transplant teams. “The availability of specialised psychosocial care is necessary and, as with medical treatment, it should be carried out by specially trained staff” (24, 25). Every transplant unit must have a mental health professional who is a team member of the transplant team. A consulting psychiatrist coming solely on request as demanded by the present JACIE accreditation procedure is not sufficient. The tasks of a mental health professional regularly working in the transplant unit and its outpatient department is to care for individual patients and their families, and to support the transplant team.
References 1. Prieto JM, Blanch J, Atala J, et al. Psychiatric morbidity and impact on hospital length of stay among hematologic cancer patients receiving stem-cell transplantation. J Clin Oncol 2002; 20: 1907-1917. 2. Hoodin F, Kalbfleisch KR. Factor analysis and validity of the Transplant Evaluation Rating Scale in a large bone marrow transplant sample. J Psychosom Res 2003; 54: 465-473. 3. Andorsky DJ, Loberiza FR, Lee SJ. Pre-transplantation physical and mental functioning is strongly associated with self-reported recovery from stem cell transplantation. Bone 314
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Marrow Transplantation 2006; 37: 889-895. 4. Hjermstad MJ, Loge JH, Evensen SA, et al. The course of anxiety and depression during the first year after allogeneic or autologous stem cell transplantation. Bone Marrow Transplantation 1999; 24: 1219-1228. 5. Grulke N, Bailer H, Hertenstein B, et al. Coping and survival in patients with leukemia undergoing allogeneic bone marrow transplantation - long term follow-up of a prospective study. J Psychosom Res 2005; 59: 337-346. 6. Syrjala KL, Roth-Roemer SL, Abrams JR, et al. Prevalence and predictors of sexual dysfunction in long-term survivors of marrow transplantation. J Clin Oncol 1998; 16: 31483157. 7. Kopp M, Schweigkofler H, Holzner B, et al. EORTC QLQ-C30 and FACT-BMT for the measurement of quality of life in bone marrow transplant recipients: a comparison. Eur J Haematol 2000; 65: 97-103. 8. Velikova G, Weis J, Hjermstad MJ, et al. The EORTC QLQ-HDC29: a supplementary module assessing the quality of life during and after high-dose chemotherapy and stem cell transplantation. Eur J Cancer 2007; 43: 87-94. 9. Frick E, Borasio GD, Zehentner H, et al. Individual quality of life of patients undergoing autologous peripheral blood stem cell transplantation. Psychooncology 2004; 13: 116-124. 10.Wettergren L, Sprangers M, Bjorkholm M, Langius-Eklöf A. Quality of life before and one year following stem cell transplantation using an individualized and a standardized instrument. Psychooncology 2007 Jul 5 [Epub ahead of print]. 11.Andrykowski MA, Bishop MM, Hahn EA, et al. Long-term health-related quality of life, growth, and spiritual well-being after hematopoietic stem-cell transplantation. J Clin Oncol 2005; 23: 599-608. 12.Syrjala KL, Langer SL, Abrams JR, et al. Late effects of hematopoietic cell transplantation among 10-year adult survivors compared with case-matched controls. J Clin Oncol 2005; 23: 6596-6606. 13.Lee SJ, Joffe S, Kim HT, et al. Physicians’ attitudes about quality-of-life issues in hematopoietic stem cell transplantation. Blood 2004; 104: 2194-2200. 14.Fann JR, Alfano CM, Roth-Roemer S, et al. Impact of delirium on cognition, distress, and health-related quality of life after hematopoietic stem-cell transplantation. J Clin Oncol 2007; 25: 1223-1231. 15.Molassiotis A, van den Akker OB, Boughton BJ. Psychological stress in nursing and medical staff on bone marrow transplant units [published erratum appears in Bone Marrow Transplant 1995 Aug; 16: 328]. Bone Marrow Transplantation 1995; 15: 449-454. 16.Ganti AK, Lee SJ, Vose JM, et al. Outcomes after hematopoietic stem-cell transplantation for hematologic malignancies in patients with or without advance care planning. J Clin Oncol 2007; 25: 5643-5648. 17.Bishop MM, Rodrigue JR, Wingard JR. Mismanaging the gift of life: Noncompliance in the context of adult stem cell transplantation. Bone Marrow Transplantation 2002; 29: 875-880. 18.Selby GB, Ali LI, Carter TH, et al. The influence of health insurance on outcomes of relateddonor hematopoietic stem cell transplantation for AML and CML. Biol Blood Marrow
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Transplant 2001; 7: 576. 19.MacLeod KD, Whitsett SF, Mash EJ, Pelletier W. Pediatric sibling donors of successful and unsuccessful hematopoietic stem cell transplants (HSCT): A qualitative study of their psychosocial experience. J Pediatr Psychol 2003; 28: 223-230. 20.Switzer GE, Myaskovsky L, Goycoolea JM, et al. Factors associated with ambivalence about bone marrow donation among newly recruited unrelated potential donors. Transplantation 2003; 75: 1517-1523. 21.Siston AK, List MA, Daugherty CK, et al. Psychosocial adjustment of patients and caregivers prior to allogeneic bone marrow transplantation. Bone Marrow Transplantation 2001; 27: 1181-1188. 22.Bishop MM, Beaumont JL, Hahn EA, et al. Late effects of cancer and hematopoietic stemcell transplantation on spouses or partners compared with survivors and survivor-matched controls. J Clin Oncol 2007; 25: 1403-1411. 23.Futterman AD, Wellisch DK. Psychodynamic themes of bone marrow transplantation. When I becomes thou. Hematol Oncol Clin North Am 1990; 4: 699-709. 24.Fallowfield L, Jenkins V, Farewell V, et al. Efficacy of a Cancer Research UK communication skills training model for oncologists: A randomised controlled trial. Lancet 2002; 359: 650-656. 25.Gratwohl A, Apperley JF, Gluckman E, (eds). The EBMT Handbook. Blood and Marrow Transplantation. European School of Haematology; European Blood and Marrow Transplantation, 2000.
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CHAPTER 17
Indications and current practice for allogeneic and autologous HSCT for haematological diseases, solid tumours and immune disorders
P. Ljungman, A. Gratwohl for the European Group for Blood and Marrow Transplantation
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CHAPTER 17 • Indications and current practice
1. Introduction The EBMT has since 1996 regularly published special reports on the indications for and current practice of haematopoietic stem cell transplantation (HSCT) in Europe for congenital or acquired haematological diseases, solid tumours, immune disorders and inborn errors of metabolism (1–4). Major changes have occurred since the first report was published. Today, approximately 25,000 transplants (10,000 allogeneic and 15,000 autologous) are performed yearly by teams reporting to the annual EBMT survey (Table 1). Autologous and allogeneic HSCT are now established treatment options and have been incorporated into the treatment algorithms for many disorders. In addition, potential new indications have emerged such as autoimmune disorders and AL amyloidosis for autologous transplants and solid tumours for allogeneic transplants. Carefully conducted prospective studies hopefully will define the role of HSCT in these situations. On the other hand alternative non-transplant based treatment options are also making major progress, thus influencing the practice of HSCT. For example the number of transplants for previously important indications such as chronic myeloid leukaemia (CML) has been reduced with the introduction of the tyrosine kinase inhibitors. Furthermore, the technical developments made during the last decade have been impressive. Unrelated donor pools have expanded and alternative donors are now more extensively used. Stem cell sources include bone marrow, peripheral blood and cord blood. It is evident that recommendations are based on an ever-changing field. Long-term follow up is lacking for recently introduced methods, while available long term observations may relate to technologies no longer in use today. Still, a few principles remain valid.
2. Risk factors for transplant outcome The main risk factors for outcome can be defined today. They are based on stage of the disease, age of the patient, time interval from diagnosis to transplant and, for allogeneic HSCT, donor-recipient histocompatibility and donor-recipient sex combination. These risk factors are cumulative and can be modified by additional particularly good or poor prognostic features as exemplified in Table 2. The importance for outcome of this type of risk assessment has been shown for CML (5) and similar evaluations are in process for other diseases. In general, transplant related mortality increases and survival rates decrease with advanced disease stage, increasing age, increasing time from diagnosis to transplant, increasing histoincompatibility and in grafts involving male recipients with a female donor. All components have to be integrated into the risk assessment and the decision whether or not to perform a transplant. These factors are never absolute, for example, the age of an individual patient remains one of the most important determinants of
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Table 1: Number of patients treated with a first HSCT by indication, donor type and stem cell s
D Allogeneic TEAMS = 605
Leukaemias Acute myeloid leukaemia 1st complete remission not 1st complete remission Acute lymphatic leukaemia 1st complete remission not 1st complete remission Chronic myeloid leukaemia chronic phase not 1st chronic phase MDS incl. Sec AL MPS Chronic lymphatic leukaemia Lymphoproliferative disorders Plasma cell disorders - MM Plasma cell disorders - other Hodgkin's lymphoma Non Hodgkin lymphoma Solid tumors Neuroblastoma Soft tissue sarcoma Germinal tumours Breast cancer Ewing Renal cancer Melanoma Colon cancer Other solid tumours Non malignant disorders Bone marrow failure - SAA Bone marrow failure - other Haemoglobinopathies - thal Haemoglobinopathies - other Immune deficiencies Inherited disorders of metabolism Auto immune disease Others TOTAL 320
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BM 793 275 198 77 310 189 121 99 73 26 83 21 5 88 20 3 14 51 13 7 1
2
HLA-id PBPC 2449 1248 834 414 433 271 162 237 163 74 286 93 152 790 255 7 120 408 29 1 1 1 8 2 6 2
3 399 164 32 92 26 64 19 2 17
8 195 100 21 52 7 9 3 3 20
1310
3483
Cord 14 4 2 2 6 3 3 0
2 2 0
2 2
29 3 3 19 1 2 1
45
Family non-id BM PBPC 46 300 20 166 8 48 12 118 17 67 5 17 12 50 3 16 2 4 1 12 36 5 10 1 5 4 48 1 8 1 1 15 1 25 1 22 1 9 7 1 4
19 11
1 64 6 4 12 1 35 6
1
2
99
436
47 6 4 7
twin Cord 2
BM 5 0
2 2
2 1 1 0
0
PB 2 1 1 5 5 2 3 2 2
1 2
1
0
2 2
8 3
0
3 2 1
2
1 1
1 1
11
3
5 0
2
4
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and stem cell source in Europe in 2006 DONOR SOURCE No. of patients
neic
d
Autologous
Total
Unrelated twin BM 5 0
2 1 1 0
PBPC 23 15 10 5 5 2 3 2 2
1 2
1
2 2
8 3
3 2 1
5 0
BM 646 237 111 126 255 125 130 57 40 17 63 24 10 69 13 17 39 4 3 1
1 1
11
1 1
32
PBPC 2181 913 379 534 464 249 215 196 87 109 363 130 115 554 184 4 85 281 10
Cord 325 142 54 88 129 56 73 10 2 8 31 7 6 34 3 8 23 1
1 3 3 2
BM only 78 64 49 15 6 3 3 0
9 1
BM + PBPC 1101 747 632 115 145 93 52 13 4 9 37 9 150 12477 5918 251 1742 4566 1402 295 63 299 134 237 7
1 7 95 20 1 28 46 77 37 9 7
195 59 21 19 4 69 18 5 18
1 93 36 16 6
1 89 14 15 1
14 5
5 362 122
1
2
23 9 3 13
35 23 1 9
3 1 1
2 118 31
932
2851
458
256
15133
Cord 0 0
0
0
0
0
0
0
Allo
Auto
Total
6784 3020 1644 1376 1690 920 770 620 373 247 866 294 294 1597 489 15 260 833 85 25 10 4 15 7 8 2 0 14 1115 390 116 208 39 258 90 14 80
1179 811 681 130 151 96 55 13 4 9 38 9 157 12572 5938 252 1770 4612 1479 332 72 306 134 246 8 0 5 376 127 0 0 3 0 3 2 119 32
7963 3831 2325 1506 1841 1016 825 633 377 256 904 303 451 14169 6427 267 2030 5445 1564 357 82 310 149 253 16 2 5 390 1242 390 116 211 39 261 92 133 112
9661
15389
25050
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Table 2: Quantification of risk of transplant-related mortality Disease stage - Early (e.g. AML CR1) - Intermediate (e.g. AML CR2) - Advanced (e.g. refractory disease)
0 1 2
Age of patient - <20 yr - 20-40 yr - >40 yr
0 1 2
Time interval diagnosis to transplant - >12 months - >12 months (does not apply to patients in CR1)
0 1
Histocompatibility - HLA-identical sibling - Other donor
0 1
Gender combination - Other - Female donor for male recipient
0 1
Additional elements - Comorbidity / Karnofsky <80 add - Donor >50 years add - CMV not -/add - Syngeneic twin - Unrelated donor 10/10 high resolution matched
+1 +1 +1 -1 -1
outcome following both allogeneic and autologous HSCT-procedures. Generally, HSCT in children gives better results than in adults. Age cannot be seen as a single risk factor but must be taken together with other factors in the decision-making regarding HSCT. It should, however, be recognised that biological rather than chronological age is the more important determining factor for outcome and with reduced intensity conditioning regimens in allogeneic transplantation, the age limit has increased, permitting the inclusion of older patients.
3. The EBMT recommendations The EBMT recommendations are based on existing prospective clinical trials, EBMT registry data, and expert opinions. They are not formal evidence-based documents. Many potential indications are rare and will never be supported by evidence from an adequately powered randomised controlled trial. In addition, since thousands of patients survive long-term, issues of quality of life and late side effects are becoming increasingly important. This is especially 322
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important in children for whom late effects, such as growth retardation, sterility, impairment of intellectual ability, and secondary tumours have an even larger impact than in adults. It is, therefore, important when recommendations are made to integrate the possible survival gain from HSCT, the risk for late complications and the quality of life into the risk assessment strategy. Some recommendations were made based upon analogy, inference, and expertise. However, there is increasing knowledge from a number of well-designed prospective studies that have used standard randomisation procedures or the so called genetic randomisation technique. In the future, EBMT recommendations will incorporate evidence assessed in a more formal way. The EBMT recommendations are not meant to decide whether a transplant is the correct choice of procedure or not for an individual patient. They give guidance which must be considered together with the risk of the disease, the risk of the transplant procedure and the chances of strategies in the same situation. 3.1. Conditioning regimens Conditioning regimens vary in their intensity and are classified as standard intensity conditioning, reduced intensity conditioning, or intensified conditioning regimens. Reduced intensity conditioning (RIC) regimens can be used in the allogeneic setting with the intention of shifting the balance between risk of transplantrelated mortality and risk of relapse. During recent years approximately one quarter of all allogeneic HSCT were performed with RIC regimens. A wide variety of RIC regimens have been described in publications and RIC HSCT should preferably be performed with a previously published protocol to gain adequate experience with a few protocols. Extensive feasibility studies have been published and short-term results show that RIC HSCT can lower the risk for early transplant related mortality. This has been used as the main argument to use RIC HSCT for older patients and for patients with co-morbidities. Results have been published for related donor HSCT up to 75 years and for unrelated donor HSCT up to 70 years. The preferred stem cell source has been peripheral blood (90%). Experience with unrelated donors has been published with results comparable to those with related donors. No formal prospective or retrospective studies however, have so far shown superior long-term results with RIC HSCT compared to standard HSCT. A conventional transplant remains the therapy of choice for younger patients without co-morbidities in the absence of results from prospective, controlled trials. 3.2. Classification of indications An important aim of the indication documents has been to classify indications and to give advice about the settings in which these various types of transplants should be performed. They have been classified as “standard of care”, “clinical option”,
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“developmental” or “generally not recommended”. Respective examples are given in Table 3.
Table 3: Examples of transplant categories Standard of care
- AML CR2, allogeneic HSCT - Multiple myeloma, autologous HSCT
Clinical option
- Non-Hodgkin lymphoma, allogeneic HSCT
Developmental
- Autoimmune disease, autologous HSCT
Not recommended
- CML, blast crisis
3.2.1. “Standard of care” “Standard of care” transplants may be performed in any specialist centre with experience with HSCT procedures provided they have an appropriate infrastructure as defined by the EBMT and JACIE guidelines (6). The results of such transplants are reasonably well defined and compare favourably (or are superior to) results of non-transplant treatment approaches. Reporting of data to international transplant registries is considered as mandatory for EBMT members. Defining a transplant as the standard of care does not mean that it is necessarily the optimal therapy for a given patient in all clinical circumstances. 3.2.2. “Clinical option” The next category is transplants classified as a “Clinical option”. This is the most difficult category. It encompasses many rare diseases and the paucity of data relating to transplant outcome, the variability in transplant techniques and the contribution of patient factors such as age and co-morbidity makes the assessment of indications for transplantation much more complex. Our current interpretation of existing data for indications in this category supports HSCT as a valuable option for individual patients after careful discussions of risks and benefits with the patient. However the value of HSCT for patients included in this category needs further evaluation. Furthermore, it is necessary to carefully consider the potential impact of various prognostic factors such as the nature of the donor, the stem cell source, and the conditioning regimens used, since outcome is likely to vary depending on these choices. We believe that transplants for indications under the “Clinical option” heading should be performed in a specialist centre with major experience with HSCT procedures, with an appropriate infrastructure as defined by EBMT guidelines and, optimally, should meet JACIE standards (6, 7). It is also important 324
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CHAPTER 17 • Indications and current practice
that data from these procedures are reported to the international transplant registries in more detail (preferably on MED-B forms) that additional knowledge can be gained and used to further assess the value of HSCT in these indications. 3.3.3. “Developmental” The third category is “Developmental” and indications have been classified in this category if there is little experience with this particular type of transplant and when additional research is needed to define the role of HSCT. These transplants should be done within the framework of a clinical protocol in which patients are offered the opportunity to undergo allogeneic or autologous HSCT in the context of a study that has been designed for individuals who satisfy defined diagnostic criteria. The protocol may be performed in a single institutional study or may reflect national or international multi-centre collaboration. Protocols for “Developmental” transplants will have been approved by local research ethics committees and must be performed according to current international standards. It is implied that the results of the study are intended for presentation to and/or publication for the medical community at large. Centres performing transplants under the category of “Developmental” should meet JACIE standards (5). The reporting of MED-B data to the international transplant registries is a prerequisite to allow further assessment of the value of HSCT in these indications. 3.3.4. “Generally not recommended” Finally, we have also defined a “Generally not recommended” category. This category includes HSCT in early disease stages when results of conventional treatment do not normally justify the additional risk of transplant related mortality, or when the disease is so advanced that the chance of success is so small that the risk of the harvest procedure for the normal donor is difficult to justify. This grading may not apply to specific situations, e.g. where a syngeneic donor exists. This category also includes HSCT for a disease in a phase or status in which patients are conventionally not treated by HSCT. Therefore, there will be some overlap between “Generally not recommended” and “Developmental”. “Generally not recommended” does not exclude the possibility that centres with a focus on a certain disease can investigate HSCT in these situations. If a HSCT is performed for a “Generally not recommended” indication, the reporting of MED-B data is strongly recommended to allow further assessment of the value of HSCT in these indications.
4. Conclusion It is beyond the scope of this Chapter to discuss the specific indications and their grading. Instead, we recommend consulting the latest published version of the HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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indications document and the accompanying disease specific Chapters in this handbook. However, based on these recommendations and on individualised risk assessments, a risk adapted strategy should be developed at diagnosis. Depending on the assessment, the transplant can be planned as initial treatment, as rescue therapy or be rejected. With better knowledge of the disease risks and the transplant risks, those algorithms will become more and more refined.
References 1. Schmitz N, Gratwohl A, Goldman J. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Current practice in Europe in 1996 and proposals for an operational classification. Bone Marrow Transplant 1996; 17: 471-477. 2. Goldman JM, Schmitz N, Niethammer D, Gratwohl A. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Current practice in Europe in 1998. Accreditation Sub-Committee of the European Group for Blood and Marrow Transplantation. Bone Marrow Transplantation 1998; 21: 1-7. 3. Urbano-Ispizua A, Schmitz N, de Witte T, et al. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Definitions and current practice in Europe. Bone Marrow Transplant 2002; 29: 639-646. 4. Ljungman P, Urbano-Ispizua A, Cavazzana-Calvo M, et al. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Definitions and current practice in Europe. Bone Marrow Transplant 2006; 37: 439-449. 5. Gratwohl A, Hermans J, Goldman JM, et al. Risk assessment for patients with chronic myeloid leukaemia before allogeneic blood or marrow transplantation. Chronic Leukemia Working Party of the European Group for Blood and Marrow Transplantation. Lancet 1998; 352: 1087-109. 6. Link H, Schmitz N, Gratwohl A, Goldman JM. Standards for specialist units undertaking blood and marrow stem cell transplants - recommendations from the EBMT. Bone Marrow Transplant 1995; 16: 733-763. 7. Joint Accreditation Committee. EBMT-EuroISHAGE (JACIE) Accreditation Manual (www.jacie.org).
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CHAPTER 18
Statistical evaluation of HSCT data
R.M. Szydlo
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CHAPTER 18 • Statistical evaluation of HSCT data
1. Introduction Stem cell transplantation (HSCT) is a widely accepted treatment modality, with both allogeneic and autologous HSCTs offering effective options for a number of diseases (eg. some leukaemias and severe aplastic anaemia) and curative potential for others (e.g. thalassaemia and CML). However, there is still much to be learnt, and the analysis of data generated from a stem cell transplant programme is not only fundamental to assessing the effectiveness of the treatment, but can provide invaluable information on the prognostic role of disease and patient factors. Thus, the appropriate analysis of such data is of paramount importance.
2. Outcomes Patients who undergo a HSCT procedure require considerable support and supervision, which in turn, allows the treatment modality to be reviewed in a variety of ways. Key events are assessed at varying times post-HSCT and these can be used to calculate a number of outcomes defined below: • Survival - the probability of survival irrespective of disease state • Disease-free survival (DFS) - the probability of being alive and free of disease (in leukaemia this outcome could also be termed leukaemia-free survival (LFS) • Graft vs. host disease (GvHD) - the probability of developing GvHD (the severity of disease being estimated would need to be clearly stated) • Graft failure (GF) - the probability of primary graft failure • Transplant related mortality (TRM) - the probability of dying without recurrence of disease • Relapse - the probability of disease recurrence • Progression-free survival (PFS) - the probability of being alive and with a disease stage not advanced of that at the time of transplantation • Neutrophil engraftment - defined as the first of 3 consecutive days post HSCT where values above a specified level are achieved (e.g. ≥0.5 x 109/L) • Platelet engraftment - defined as the first of 3 consecutive days post HSCT where values above a specified level are achieved (e.g. ≥50 x 109/L) Probability curves describing these outcomes fall into two categories: Survival, DFS/LFS and PFS involve events with decreasing cumulative probabilities over time, whilst GvHD, TRM, GF and relapse involve events that result in increased cumulative probabilities over time.
3. Survival analysis The outcomes outlined above require careful consideration before a statistical analysis can be considered. Each event of interest may occur at variable times post transplant,
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so in statistical terms it has two components - whether it occurs at all and, if it does, the length of time from transplant to the event. However, inherent in many studies is the problem that the event of interest is seldom observed in all of the patients. Thus, a patient who has not yet had the event of interest at the time of analysis, or who is lost to follow-up, would be “censored” at the time of last contact. The inclusion of data that is censored precludes the use of simple statistical methods such as chi-squared analysis or rank methods and requires a statistical treatment known as survival analysis, which can be applied to a variety of end points. 3.1. Kaplan-Meier method There are a number of methods for analysing survival data, and though these depend on the precision of the recorded time interval, are usually summarised as survival or Kaplan-Meier (1) curves which are derived from calculated tables commonly known as life tables (constructed on the basis of a series of conditional probabilities). The term life table is also frequently used to describe data where the results are grouped into time intervals, often of equal length, and this method of calculation is described as actuarial. In fact the terms “actual” and “actuarial” are often used mistakenly to describe survival probabilities generated by Kaplan-Meier methods. Figure 1 shows typical survival data from fifteen consecutive patients transplanted at a single centre. Four have died and eleven were still alive at various time points post-transplant. If the data are rearranged in order of time, then a life-table can be calculated by the method of Kaplan-Meier as shown in Table 1.
Patient number
Figure 1: Survival data from fifteen patients who received a haematopoietic stem cell transplant
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15 14 13 12 11 10 9 8 7 6 5 4 3 2 1 0,0
Alive Dead
0
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40
80
120 160 200 240 280 320 Days post BMT
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CHAPTER 18 • Statistical evaluation of HSCT data
Table 1: Life table for fifteen patients who received an allogeneic haematopoietic stem cell transplant Time (days)
Status (0=alive, 1=dead)
Number at risk
Probability of survival
Standard error
0 1 1 0 1 0 1 0 0 0 0 0 0 0 0
15 14 13 12 11 10 9 8 7 6 5 4 3 2 1
1.00 0.93 0.86
0.069 0.094
0.78
0.113
0.69
0.129
16* 26 66 69* 74 82* 88 89* 117* 133* 144* 172* 252* 291* 305* * censored observation
The data presented in Table 1 can be used to produce a survival curve, also known as a cumulative survival rate, or a survival function (Figure 2). Vertical tick marks on the curve represent censored individuals who make no contribution to the curve after that particular time point. The curve is an estimated probability of survival, and using appropriate methods to compute the standard error, 95% confidence intervals (95%CIs) can be calculated. In common with many analyses of small data sets, the standard error calculated from day 88 post BMT has yielded a large 95% CI, and so the survival curve must therefore be interpreted with some caution. 3.2. Cumulative incidence procedure The following outcomes–relapse, TRM, GvHD and GF are subject to the problem of “competing risks” (for example, in the case of calculating a relapse probability, a patient who dies in remission cannot relapse), and so the most appropriate method of analysing such data is to produce a cumulative incidence curve (2). Although this methodology is not included in most commercial statistical packages, it is present in the statistical package NCSS (Statistical analysis & data analysis software), and
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Figure 2: Probability of survival following a haematopoietic stem cell transplant (n=15)
Probability of survival %
100 80
69%
60 40 95% confidence interval 20 0
0
28
56
84
112 140 168 196 224 252 280 308
Days post BMT
macros are available to allow such curves to be calculated using the statistical packages SAS and R (3). Although it is possible to use the Kaplan-Meier method for each of these outcomes, this is likely to produce an overestimate of the true probability as calculated by the cumulative incidence approach (4). This discrepancy will be largest where the event of interest occurs later after HSCT (TRM, relapse and chronic GvHD - see Figure 3) and may be negligible with early outcomes (acute GvHD and graft failure).
4. Other methods for describing outcomes Recent advances in statistical methodology have enabled LFS and DFS curves to be modified to take into account durable remissions achieved after relapse (5). The generation of current leukaemia-free survival (CLFS) curves does however require detailed follow-up data and the use of macros designed for the statistical software package SAS. If the exact times to the onset of GvHD are not known, then simple proportions of grades of disease can be presented for those patients who survived long enough potentially to develop the disease (thus patients who died within 100 days postHSCT are not eligible for chronic graft versus disease). Engraftment times can either be described with a median and range, or with cumulative probability curves. Comparisons between GvHD groups should be made using the chi-squared test or chi-squared trend test, whilst the Mann-Whitney or Kruskal-Wallis test are applicable for engraftment data. An example of data presentation from 45 consecutive patients is illustrated in Figure 3.
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CHAPTER 18 • Statistical evaluation of HSCT data
Figure 3: Probability curves of 6 different possible outcomes
5. Composite outcome diagrams An interesting new way of graphically representing how outcome probabilities change with time post-HSCT has been developed by Ronald Brand and is included in a recent paper (Figure 4) (6). Cumulative incidences of relapse, and of non-relapse
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Figure 4: Composite outcome diagram 1.00 0.90 0.80 Disease Free Survival
0.70
Overall Survival
0.60 Alive after relapse
0.50
Relapse Incidence
0.40 Dead after relapse
0.30 0.20
NRM
0.10 0
350
720
Days following transplantation
death are estimated and simultaneously plotted with the Kaplan-Meier survival probability. The resulting diagram thus provides the 4 possible patient states after a HSCT - alive without relapse of disease (white), alive after relapse (light blue, an outcome not normally calculated), dead after relapse (dark blue) and non-relapse death (grey). In addition, the proportion of patients relapsing (relapse incidence) is provided by the sum of the horizontal and diagonal hashed groups, the interface between the alive and dead components represents overall survival, and the interface between alive with and without relapse – relapse-free survival. One is thus able to view in one diagram the relative importance of all these possible outcomes. This is especially useful for illustrating differences between groups identified from univariate or multivariate analyses as being of prognostic significance. Specialist software is not required to create such diagrams, as macros are available for the statistical package SPSS-14.
6. Comparison of survival curves Survival curves provide a visual assessment for disease course and/or outcome of a particular treatment or disease course. In order to establish whether there is a survival advantage between, for example, two treatments, it is necessary to perform a statistical test to compare the two life tables. This is achieved using the logrank or Mantel-Cox test (7). In this test each observation is given an equal “weight”. However, in the transplantation setting, where there may be considerable early mortality, it may be more useful to “weight” early observations and in this context the Breslow test (8) may be more appropriate (this test is also less sensitive to late 334
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events when few subjects may be present in the study). The Tarrone-Ware test (9) is a compromise between the Mantel-Cox and Breslow tests. An example of the relative merits of these tests is provided in Figure 5. Here survival data from two hypothetical groups are presented. The proportion of survivors could be compared using a standard chi-squared test (treatment A 20/30 vs. treatment B 12/30), yielding a significant result (p=0.04), and suggesting treatment A to be the superior. However, this analysis ignores the time to an event, and gives a misleading result as exemplified by Figure 5. There is a clear survival advantage in the first four years post BMT for treatment A, but for long-term survival, treatment B may be better. The curves presented in Figure 5 do however highlight another analytical problem. In order to perform a log-rank test, the groups being tested should run in parallel and not cross over. A more sophisticated approach for analysing such data should therefore be undertaken (4). In addition to comparing treatments, the log-rank test can be used to compare selected sub-groups within one treatment or disease category e.g. males vs. females, patient age <30y vs. patient age ≥30y, early vs. late stage of disease, etc. As with all statistical comparisons of subgroups, a more stringent criterion for significance needs to be predetermined in accordance with the number of tests to be performed. If one or more prognostic variables are known, then a stratified log-rank analysis can be undertaken to look at the influence of these factors on outcome. Thus, for example, in acute myeloid leukaemia where disease sub-type is an important prognostic indicator, the effect of patient sex could be investigated with a stratified log-rank test using disease sub-type as the stratified variable. Figure 5: Probability of survival following BMT for two hypothetical groups
Probability of survival %
100
p1
= Log-rank test
p2 = Tarrone-Ware test p3 = Breslow test
p1 = 0.05 p2 = 0.003 p3 = 0.0004
80
Treatment A (n=30)
60 40
38%
Treatment B (n=30)
20 0
12%
0
12
24
36
48
60
72
84
Months post BMT
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7. Presentation of survival data In quoting survival rates, probabilities at specific points in time should be indicated together with a confidence interval or standard error (e.g. the probability of survival at 3 years was 59% (95% CI’s 44–73%)). Although 95% CI’s can be calculated by taking the survival rate ± 1.96*standard error, if the survival rate is close to 100% or 0%, this can lead to confidence intervals greater than 100% or less than 0%. To avoid this, asymmetrical confidence intervals can be calculated by the method of Rothman (10). Median survival times can be derived from survival curves, and correspond to the time at which the survival proportion reaches 0.5, but this is not always possible (as demonstrated in Figure 2) if the survival curve reaches a plateau above this point. Guidelines for the presentation of results of transplantation data have been suggested by Klein (4) and Labopin (3). The presentation of survival curves from univariate analyses of prognostic factors should be viewed with extreme caution, as adequate control of other potential prognostic factors or biases cannot be guaranteed. A multivariate approach is therefore to be recommended.
8. Proportional hazards regression analysis The log-rank test enables the survival experience of two or more groups to be compared but in order to investigate a number of possible prognostic variables simultaneously, a regression method introduced by Cox and known as proportional hazards regression analysis has to be employed (11). The special nature of survival data as outlined previously, makes the use of usual regression methods (e.g. linear regression, logistic regression) inappropriate. The use of the Cox model allows the identification of prognostic factors that are related to the outcome. In addition, the search for variables of unknown prognostic significance can be performed after adjusting for variables of known prognostic significance. Thus for example, in chronic myeloid leukaemia where the stage of disease at transplant is a major factor in survival, the influence of other factors would be investigated having taken into account disease stage. This approach also allows for the generation of survival curves for a given factor that are adjusted for the influence of other factors. Such curves are likely to be much more informative that simple univariate analysis curves. The use of the Cox model does require a sound statistical knowledge, as there are many potential difficulties with the method both in application, and interpretation of results. Several reviews of the subject have been published and are recommended (3, 4).
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9. Conclusions The analysis and presentation of survival data can provide important information on the effectiveness of transplantation in treating a particular disease, and with sufficient numbers of patients, subtle differences between patient groups can be identified. The now routine availability of computers and statistical software enables the analysis of complex data sets to be carried out with relative ease, but the importance of statistical advice at all stages of data analysis should not be underestimated.
References 1. Kaplan EL, Meier P. Non-parametric estimation from incomplete observations. J Am Stat Assoc 1958; 53: 457-481. 2. Gooley TA, Leisenring W, Crowley JA, Storer BE. Estimation of failure probabilities in the presence of competing risks: New representations of old estimators. Statistics in Medicine 1999; 18: 695-706. 3. Labopin M, Iacobelli S. Statistical guidelines for EBMT. 2003; http://www.ebmt.org/1WhatisEBMT/Op_Manual/OPMAN_StatGuidelines_oct2003.pdf 4. Klein JP, Rizzo JD, Zhang M-J, Keiding N. Statistical methods for the analysis and presentation of the results of bone marrow transplants. Part I: Unadjusted analysis. Bone Marrow Transplant 2001; 28: 909-915. Part II: Regression modeling. Bone Marrow Transplant 2001; 28: 1001-1011. 5. Craddock C, Szydlo RM, Klein JP, et al. Estimating leukemia-free survival after allografting for chronic myeloid leukemia: a new method that takes into account patients who relapse and are restored to complete remission. Blood 2000; 96: 86-90. 6. Lim ZY, Ingram W, Brand R, et al. Clonal gammopathies following alemtuzumab-based reduced intensity conditioning haematopoietic stem cell transplantation: Association with chronic graft-versus-host disease and improved overall survival. Bone Marrow Transplant, Epub 2007, Aug 20: 747-752. 7. Mantel N. Evaluation of survival data and two new rank order statistics arising in its consideration. Cancer Chemother Rep 1966; 50: 163-170. 8. Breslow N. A generalised Kruskal-Wallis test for comparing k samples subject to unequal patterns of censorship. Biometrika 1974; 57: 579-594. 9. Tarone RE, Ware J. On distribution-free tests for equality of survival distributions. Biometrika 1977; 64: 156-160. 10. Rothman KJ. Estimation of confidence limits for the cumulative probability of survival in life table analysis. J Chron Dis 1978; 31: 557-560. 11.Cox DR. Regression models and life tables. Journal of the Royal Statistical Society 1972; 34, Series B: 187-220.
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Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Who are censored patients? a) Those who have experienced the event of interest early after SCT . . . . . . . . b) Those who have not experienced the event of interest and are alive . . . . . c) Those who have experienced the event of interest but are lost to follow-up . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Those who failed to engraft. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2. What is the correct test for comparing two Kaplan-Meier survival curves? a) Chi-squared test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Mann-Whitney test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) T-test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Log-rank test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. What is the cumulative incidence procedure used for? a) To calculate survival curves where there is a competing risk to the event of interest . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) To compare survival curves where the event of interest occurs early after SCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) To compare survival curves where the event of interest occurs late after SCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) To identify censored patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. What proportional hazards regression analysis is used for which of the following: a) To calculate survival curves with more than one event of interest . . . . . . . . b) To help identify prognostic factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) To identify patients lost to follow-up . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) To censor events of interest that are competing risks . . . . . . . . . . . . . . . . . . . . . .
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5. How do cumulative incidence curves differ from Kaplan-Meier curves: a) Only when the event of interest is time independent . . . . . . . . . . . . . . . . . . . . . . b) By taking into account competing risks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) By providing the correct estimate of the competing risk . . . . . . . . . . . . . . . . . . d) Because they don’t take into account censored events . . . . . . . . . . . . . . . . . . . .
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CHAPTER 19
Methodology of conducting academic clinical trials in Europe
Z. Doran
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CHAPTER 19 • Methodology of conducting academic trials in Europe
1. Introduction In the era of evidence based medicine, it is ever more important that changes in clinical practice are developed through the mechanism of prospective clinical trials. Clinical trials allow us to develop therapy in a structured fashion in which the discipline of protocol driven management not only increases exposure to experts but also improves standards and encourages adherence to “Good Clinical Practice” (GCP). What do we mean by the term methodology in relation to clinical trials? In simple terms the methodology of clinical trials would describe the methods, rules and assumptions that are employed to explore or examine a research question. In a broader sense any discussion on the methodology of clinical trials should also characterise the philosophical principles that guide us in their conduct. This Chapter provides: - A brief review of the history of research ethics - A description of the current EU Directives governing the conduct of clinical trials and the associated challenges for academic research - A description of the different types and phases of trials - Guidance on the design and composition of a protocol - An overview of approval process, both at the national and local level.
2. History of research ethics Until the early 1900s there were no regulations whatsoever addressing the ethics of involving human subjects in research. This started to change with the advent of various governing bodies and legislation over the first decades of the 20th century. However, the philosophies governing the conduct of modern clinical trials owe their origin to more recent history, such as the Nuremberg Trials, the catastrophic results of the use of the non-licensed thalidomide in pregnant women and the lesserknown Tuskegee Syphilis study in the US. The Nuremberg Trials In 1946 an American Tribunal brought criminal proceedings against a number of leading German physicians and scientists who had conducted medical research studies on thousands of men, women and children in the concentration camps, without their consent. As a direct result of the trials, the Nuremberg Code was established in 1948. Although this was not implemented into law, it was the first international agreement advocating voluntary participation after informed consent. It also stated that the perceived benefits of the research must always outweigh the possible risks to the subjects. Its basic premise was that “voluntary consent of the human subject is absolutely essential”.
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Declaration of Helsinki To further protect subjects and to give guidance to researchers, the World Medical Association established recommendations for the conduct of clinical research in the Declaration of Helsinki in 1964 (http://www.wma.net/e/policy/b3.htm). “It is the duty of the physician to promote and safeguard the health of the people. The physician’s knowledge and conscience are dedicated to the fulfilment of this duty.” “The health of my patient will be my first consideration”. International Conference on Harmonisation – Good Clinical Practice (ICH-GCP) In more recent years guidelines have been developed by the International Conference on Harmonisation, whose first conference was held in 1991. It is an ongoing process and subsequent meetings have refined the guidelines. ICH-GCP is the accepted international standard for designing, conducting, recording and reporting trials and it is mandatory that all personnel who participate in clinical trials are trained in GCP (http://www.ich.org/cache/compo/276-254-1.html).
3. Current legislation for clinical trials in Europe In the EU Member States the conduct of clinical trials is now governed by the various Directives issued by the European Union (EU). Whilst the objectives of implementing such Directives were laudable, the primary goal of achieving harmonisation has not been met, as each Member State has interpreted the Directives within the framework of their own national legislation and has thereby introduced inconsistencies. Conducting a multi-centre, multi-national study has consequently become a challenge, but it is important to remember that although it is difficult, it is not impossible. This section describes the major challenges and provides suggestions for various strategies to minimise the impact of the paperwork and fulfil the requirements of the Directives in an academic setting. It is important to know that these Directives exist and that the interpretations of each Directive may vary slightly from country to country. It should be noted that most of the EU legislation on the conduct of clinical trials is focused on Investigational Medicinal Products (IMPs) or new medical devices (not addressed in this Chapter). Discussion of each Directive in detail is not possible in this format. Those with the most universal application to academic research in the field of transplant are listed in Table 1, together with a brief overview of their content. The full directives can be found on the European Commission website at http://ec.europa.eu/enterprise/pharmaceuticals/eudralex/index.htm.
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Table 1: European Regulations and Directives affecting the conduct of clinical trials in the field of stem cell transplantation Regulations & Directives
Date of issue
Content
Directive 2000/70/EC of the European Parliament and of the Council
16 November 2000 Amends Council Directive 93/42/EEC as regards medical devices incorporating stable derivates of human blood or human plasma
Directive 2001/20/EC of the European Parliament and of the Council
4 April 2001
Lays down the laws, regulations and administrative provisions of the Member States relating to the implementation of good clinical practice in the conduct of clinical trials on medicinal products for human use
Directive 2001/83/EC of the European Parliament and of the Council
6 November 2001
Describes the Community code relating to medicinal products for human use
Directive 2002/98/EC of the European Parliament and of the Council
7 January 2003
Sets standards of quality and safety for the collection, testing, processing, storage and distribution of human blood and blood components and amending Directive 1/83/EC
Directive 2004/23/EC of the European Parliament and of the Council
31 March 2004
Sets standards of quality and safety for the donation, procurement, testing, processing, preservation, storage and distribution of human tissues and cells
Commission Directive 2005/28/EC
8 April 2005
Lays down principles and detailed guidelines for GCP as regards investigational medicinal products for human use, as well as the requirements for authorisation of the manufacturing or importation of such products
Commission Directive 2006/86/EC
24 October 2006
Implements Directive 2004/23/EC of the European Parliament and of the Council as regards traceability requirements, notification of serious adverse reactions and events and certain technical requirements for the coding, processing, preservation, storage and distribution of human tissues and cells
Regulation (EC) No 1901/2006 of the European Parliament and of the Council
12 December 2006
Addresses medicinal products for paediatric use and amends Regulation (EEC) No 1768/92, Directive 2001/20/EC, Directive 2001/83/EC and Regulation (EC) No 726/2004
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3.1. Challenges for the academic community Academic groups have found it difficult to access resources to fulfill the requirements imposed by the Directives and this has prevented some studies from taking place. To address this issue, a draft Directive modifying EU 2001/20/EC and EU 2005/28/EC for the conduct of non-commercial trials had been released for public consultation. The extensive responses to this have led to a delay in the issue of any amended Directive as the EU legislators consult with experienced trials staff from both the commercial and academic sectors. It is easy to be overwhelmed by the number of Directives but in reality most academic research is now conducted within academic cooperative groups with an administrative infrastructure that should provide guidance on managing the mass of legislation. The main issues that should be considered are: - Sponsorship - Registration with the European Union Drug Regulatory Authorities (Eudra) - Clinical Trials Authorisation (CTA) - Study conduct in compliance with ICH-GCP compliance - Indemnity - Safety reporting - Registration with the International Committee of Medical Journal Editors. 3.1.1. Sponsorship Prior to the implementation of the Directives all participating institutions took responsibility for their own patients on a study. Now all studies require a single “Sponsor”. This Sponsor is responsible for ensuring that the protocol is conducted appropriately according to EU Law and ICH-GCP, including indemnification. Identifying a Sponsor for an academic study can prove to be a challenge. Possible solutions include: 1. Sponsorship undertaken by an academic cooperative group e.g. EBMT, EORTC, etc. 2. Sponsorship undertaken by a national body e.g. UK Medical Research Council 3. Sponsorship undertaken by a university 4. Sponsorship undertaken by the lead institution. The last option is probably the least likely as most institutions are reluctant to take overall responsibility for other institutions that are not under their management, particularly those in another country working within a different system of healthcare provision. If it proves impossible to find an overall Sponsor then there is another potential strategy. Some or all of the responsibilities of Sponsorship can be delegated to other(s). This can enable a single entity or an academic group to be a “figurehead” Sponsor. Using this model it is possible that each hospital (or perhaps each 344
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country) could look after all the local aspects themselves, just as they used to in the pre-EU Directive academic studies. However although the Sponsor may delegate they remain legally responsible, so such delegation must be formally described in a contract detailing the arrangements for the conduct of the study. In practice we will probably witness a spectrum of solutions as shown in Figure 1.
Figure 1: Spectrum of organisation of the responsibilities on the Sponsor
Well funded/resourced Sponsor undertakes all roles
Poorly resourced/non funded Sponsor legally responsible Sponsor delegates all roles under contract
3.1.2. EudraCT registration Obtaining a EudraCT number for a study is a very simple online procedure, which takes only a few minutes (http://eudract.emea.europa.eu). Basic information only is required for the registration but this must include the name of the Sponsor. If a Clinical Trials Authorisation (CTA) is required for the study (see below), this can be done via the same website. All trials conducted within the EU must apply for a EudraCT number, regardless of whether or not they are investigating an Investigational Medicinal Product (IMP). 3.1.3. Clinical Trials Authorisation Some trials require a Clinical Trials Authorisation, particularly if the involves an IMP or a new device. The definition of an IMP is critical and seems to have caused particular confusion in the fields of haematology and oncology. It would seem logical that new and experimental therapies are IMP, but the definition also includes licensed medications being used for unlicensed indications. Developing a new drug or therapy is a very expensive operation and takes many years. Consequently most IMPs undergo investigation to obtain a Marketing Authorisation (MA) or license in a limited number of indications. The use of the product in other situations is generally driven by the medical profession and is described as “off-label” use. In the field of transplant this has become a problem, as most of the drugs used in the preparative regimens and/or in supportive care are not licensed for transplant. As a consequence they must be considered as IMPs. The legislation does not accommodate the realities of drug
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development or standard medical practice in this regard, and has proven a challenge for both the legislators and those conducting clinical trials. It was obviously not the intent to consider well-established therapies as IMPs but it has been one of the end results. As a result some Competent Authorities (CA) (the bodies designated in each country to ensure the correct implementation of the Directives) provide conflicting advice on this subject. A reasonable approach would be to apply for a CTA if in any doubt, and ensure that exemption from the requirement for a CTA is provided in writing by the CA. The current legislation does not consider blood and blood products to be IMPs although this decision may be overturned in the future. 3.1.4. Study conduct in compliance with ICH-GCP compliance Under current legislation the conduct of all trials must be comply with the principles of ICH-GCP. GCP is an international ethical and scientific quality standard that addresses all aspects of clinical trials from design through to the reporting of the results. The guidelines define the roles of all participants, describe the essential elements of a protocol and the investigator’s brochure as well as detailing the standards for the conduct of a study. All personnel involved in the conduct of clinical trials should complete an initial ICH-GCP course and attend regular updates. 3.1.5. Indemnity Insurance must be available for each study. Prior to the implementation of the Directives most academic studies were conducted under the insurance of the participating institution and in some instances with additional cover from the clinician’s medical negligence insurance and the manufacturer’s product liability insurance. It is still possible to conduct studies in the same manner, but the requirement for a single overall sponsor under EU law would mean that the overall Sponsor would delegate insurance cover to the participating Institutions. If funding allows it is simpler and safer to take out a study indemnification policy. 3.1.6. Safety reporting (Pharmacovigilance) Safety reporting is a mandatory component of study conduct and is onerous on the part of all concerned. Serious Adverse Events (SAEs) Serious adverse events are adverse events that result in one or more of the following: - Hospitalisation or prolongation of hospitalisation - Death - Life-threatening - Results in persistent or significant disability/incapacity 346
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- Congenital abnormality or birth defect - Is otherwise medically significant in the opinion of the investigator. In order to minimise the paperwork, the adverse events that are normal and expected for the population should be identified, detailed in the protocol and specifically excluded from the necessity of reporting. For instance the most common cause of an SAE is hospitalisation so if hospitalisation is expected, for instance if patients are given, in a an out-patient setting, chemotherapy that is likely to induce neutropenia and therefore febrile events necessitating in-patient care, this should be identified in the protocol. Suspected Unexpected Serious Adverse Reaction (SUSAR) This is defined as an adverse reaction, the nature or severity of which is not consistent with the known study treatment information. A serious event or reaction is not defined as a SUSAR when: - It is serious but expected - It does not fit the definition of an SAE, whether expected or not. Requirements for reporting adverse events All adverse events that are defined as reportable in the protocol should be recorded in the case record forms (CRFs). If an adverse event fulfils the criteria of a SAE or SUSAR then additional reporting requirements must be followed, as specified in Table 2.
Table 2: Reporting requirements for adverse events, serious adverse events and SUSARs Report from
Report to
Timeframe
AEs
Site
Sponsor
Trial Specific
SAEs
Site
Sponsor
24 hrs
SUSARs
Sponsor
CAs / ECs
Expedited
PIs / MAHs
Trial Specific
ASR
Sponsor
CAs / ECs
From 1 year anniversary of CTA
MAH: Marketing Authorisation Holder; ECs: Ethical Committees; CA: Competent Authorities; EMEA: European Medicines Evaluation Agency; C-PI: Coordinating Principle Investigator; PI: Principle Investigator (each institution); ASR: Annual Safety Report
3.1.7. Registration with the International Committee of Medical Journal Editors Registration with the International Committee of Medical Journal Editors is required for publication. This additional requirement is a relatively quick process.
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4. Clinical trials The scope of a clinical trial varies depending on the topic, ranging from the laboratory, through screening and prevention to testing of new therapeutic strategies. There are many trial designs and some of the options are listed in Table 3.
Table 3: Types of clinical trials Scope
Design
Diagnostic Prevention Screening Supportive Care (Quality of Life) Therapeutic Genetic Technical Laboratory Pharmacodynamic/Pharmacokinetic Bioequivalence
Randomised Non Randomised, Registration or Cohort Single Blind Double Blind Crossover Controlled – Placebo, Comparator Parallel Open Label
4.1. The stages of study development The development of new medicinal products follows a clearly defined pathway. Trials of blood and marrow transplantation and other cellular therapies may have a slightly different sequence with the Phase I stage being replaced by a Pilot study in which the safety, feasibility and initial indications of efficacy are studied. 4.1.1. Preclinical Studies In vitro As the name suggests, these are pre-clinical studies conducted in the laboratory. Animal Studies The fist clinical studies may be conducted in animals, notably rodents and primates although this is not an absolute requirement. 4.1.2. Phase I/Pilot Studies Phase I studies are the first to be conducted in humans and are designed to determine the safety and feasibility of the study. They include dose-finding and the identification of toxicities. They aim to identify the maximum tolerated dose (MTD), defined as the dose at which subjects experienced a dose limiting toxicity (DLT). The DLT is itself defined as the dose at which a toxicity that is considered unacceptable is experienced by a proportion or all of the patients in that cohort.
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By identifying the MTD, the Phase I study can then help to establish the recommended dose for any Phase II study. This is, defined as the dose level below the MTD. These studies enter cohorts of patients, usually 3–5 individuals, at a particular dose level. The more accurate the estimation of the probable dose from the evidence produced in pre-clinical/animal studies, the shorter the study. In the case of biologics a different approach has evolved as many of these compounds have proven to be non-toxic so that a DLT is not seen. In these cases it is necessary it is necessary to establish the minimum therapeutic dose, i.e. that at which the desired efficacy is achieved. Phase I studies may also give some insight as to efficacy and selection of the most appropriate patient population for further product development. 4.1.3. Phase II Studies The target population for Phase II studies is identified by the pre-clinical studies and from preliminary efficacy data from Phase I studies. The primary goal of Phase II studies is to determine the efficacy of the new therapeutic strategy. They can also be termed “therapeutic exploratory studies”. In general 25–50 patients are required to answer the question of efficacy. The design often includes an early stopping rule to ensure that a study is stopped promptly if the investigational approach is deemed too dangerous or to have insufficient efficacy. Phase II studies should further define the patient population for the novel therapeutic approach. 4.1.4. Phase III Studies Phase III studies are required to establish a new therapeutic strategy as the standard of care. These require a much larger sample size and the number of patients required to meet the statistical end points of a study may be hundreds or even thousands. The smaller the improvement in efficacy over previous best therapy, the larger the number of patients required for statistical proof. 4.1.5. Phase IV Studies Phase IV studies are designed to expand the knowledge and experience of the new approach in the tested and in other patient populations. 4.2. Designing a new study protocol Gaining experience in the field of clinical research is an important part of the career path of an academic clinician. After the implementation of the EU Directives, it has become logistically far more difficult to initiate a new study within the realm of academia, and it is ever more important to follow a measured and structured approach is followed.
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4.2.1. Step 1: Identify and describe the clinical question to be explored The initial step is obviously to identify the clinical question to be studied. Current practice and literature should be explored and discussed with peers and mentors. Deficits in knowledge and/or treatment options must be considered in order to identify an appropriate and relevant research proposal. 4.2.2. Step 2: Feasibility check A preliminary feasibility study is essential and should address issues such as: - Is the potential study of interest and potential value to the blood and stem cell transplant community at large? - Is the potential study statistically sound? For instance how many patients would be required to answer the question with an acceptable level of confidence? If there a suitably sized target population available? - In which environment should/can the study be conducted? Will the study take place in a single centre, several centres nationally or internationally? - Is it feasible to complete the study in a reasonable time frame? - Are the resources available to conduct the study and to collect and analyse the data? - Are there potential funding options? - Who/what will be the potential Sponsor under EU Law? 4.2.3. Step 3: Prepare the proposal A synopsis of the study should be generated and should contain: - Background and rationale for the study - Study design and schema - Study endpoints - Eligibility criteria - Statistical considerations. At this time a draft budget will be required. Help with this task may be available from a local institution (research department or clinical trials office), European research groups (EBMT, EORTC, etc.), and/or national research groups (ALFA, Hovon, IFM, GITMO, GOELAMS, etc.). All the requirements for study conduct in accordance with EU law should be considered and these include insurance, data management, data monitoring, submission fees (ethics committees and competent authorities), translations, per patient fee to provide resources at the local institutions, data analysis, statistical support and publication costs. An accurate budget is essential to adequately inform and thereby obtain a Sponsor. If the proposal is accepted after peer review, then the full protocol is developed. The essential elements of a protocol are detailed in Table 4. To facilitate comprehension in a multi-lingual group, ever effort should be made to maximise the use of schemas, flow-charts and tables in preference to prose (see Table 5, Figure 2). 350
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Table 4: Essential elements of a protocol Element
Description
Title page
Title of study, main study personnel, Sponsor and EudraCT number
Synopsis
An overview of the major elements of the study, include the sections market with an * below
Introduction
Reviews the disease, target patient population, treatment options, published data, the background for the study
Study rationale
Justification for the therapeutic approach to be taken in the study
Primary and Secondary Endpoints*
Dependent on the type and phase of the study. Common endpoints include MTD, determination of therapeutic dose, dose limiting toxicity, response rate, relapse or progression free survival, overall survival, safety data
Study design and schema
Details in prose and in diagrammatical form the design of the study
Eligibility criteria*
Details the characteristics of the patient population, ensures patients are suitable and fit enough to enter the study and have no confounding conditions/medications that would prevent their evaluation
Study treatment plan*
Details all therapeutic options of the study, including doses, routes of administration, treatment schedules etc. Describes the criteria which require any therapeutic option to be either decreased or increased, stopped, re-started etc. Describes any supportive medications that are either mandated, recommended or not allowed. Defines response (criteria for complete response, partial response, stable disease and disease progression, minor response). Defines conditions for continuing or discontinuing study e.g. response, progression, failure to achieve a response, patient request, unacceptable toxicity. Use schemas where possible (Figure 2)
Study investigations
All investigations to monitor the subjects’ general health, response, disease evaluation and toxicity. Use a table for easy comprehension either in the body of the protocol or as an appendix (Table 5)
Safety reporting
Standard definitions of adverse events, and reporting requirements
Statistical considerations*
Develop with professional advice
Study monitoring
Basic requirements, start up (initiation), during the study and close of study
Quality assurance and audit
Should be done to validate results as well as satisfying ICH-GCP
Ethical and Regulatory Considerations
Details the approvals to be sought
Study administration
Details of registration and/or randomisation, data collection and indemnity
Publication Policy References
Reference all published data used in the introduction
Appendices
Sample patient information leaflet and consent form, table of study investigations and procedures, any guides for response evaluation, grading etc, any questionnaires, including quality of life
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Table 5: Example of a table of investigations Pre study
Treatment all cycles End of cyles 2, 4, 6, 8, 10, 12
End of treatment
Post-treatment follow-up
Evaluation
Day-14 Day Day Day Day to-1 1 4 8 11
Medical History
X
Clinical Status
X
X
X
X
Karnofsky performance status
X
X
X
X
Pregnancy test (1)
X
Informed Consent
X
Randomisation
X
Weight and Height
X
Bone marrow aspirate and/or biopsy
X
Cytogenetics
X
Serum Protein Electrophoresis
X
X
X
X
X
Serum and urine protein immuno-fixation
X
X(6)
X(6)
X(6)
X(6)
Serum biochemistry (2)
X
X
X(4)
X
X
Haematology (3)
X
X(4) X(4) X(4) X(4)
X
X
24 hour urine for protein and creatinine clearance
X
X
X
X
Skeletal survey (plain films)
X
Suggested repeat to confirm diagnosis of CR
EMG
X
FACT/GOG-Neurotoxicity Questionnaire
X
Adverse Events Assessment
Repeat to confirm diagnosis of CR X(5)
X
X
Repeat every 3 months during the study X
X
X
X
1: In women of child bearing potential; 2: To include AST, ALT, bilirubin (total), total protein, serum calcium, creatinine, BUN, LDH, CRP, b2 microglobulin, IgG, IgA, IgM, and if possible Serum free light chain, not mandatory; 3: Complete blood count with differential; 4: Must be within 24 hours before every dose; 5: To document complete response and to repeat in patients in complete responses; 6: If patient has no more abnormal fraction in serum protein electrophoresis or negative proteinuria
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Figure 2: Examples of study schema Cycles 1-8 (21 day cycle) 1
2
3
4
Drug A
X
Drug B
X
X
X
X
Drug C
X
X
X
X
5
6
7
X
8
9
10 11 12 13 14 15 16 17 18 19 20 21
X
X
X X
X
X
X
X X
X
X
X
X
X
X
X
X
X
X
Prednisone Week
Dose (mg/kg/day)
0
1.5
2
1.0/0.5 alternate days
4
1.0/0.25 alternate days
6
Response Assessment CR/PR
SD/PD
1.0 every other day 8
Off-study
CR
PR
0.75 every other day
1.0 every other day
10
0.55 every other day
maintain dose
12
0.45 every other day
14
0.35 every other day
16
0.25 every other day
18
0.20 every other day
20
0.15 every other day
22
discontinue
4.3. The Approval Process The approval process of a new study protocol can be a long and involved process, starting with peer review, progressing through national regulatory and ethical approvals and finally local approval. A schematic of a broad overview of the general process can be found in Figure 3.
5. Summary The new Directives and subsequent regulations have undoubtedly increased the HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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Figure 3: Schematic Flow-Chart of approval processes Peer review: scientific Sponsor review: science/resources/finance National approval Competent authority* National ethics
Local/regional approval Regional/Federal approval* Local ethics Institutional review Pharmacy *
Essential Documents in Place Copy of CA approval * Copy of ethics approval** Name and contact details of local ethics committee Trial agreement Signed and dated protocol signature page** Staff delegation log** Laboratory reference ranges** CVs of all investigators Financial disclosure Form** Any other local requirements**
* if required; ** for each Institution
complexity of the design and conduct of prospective clinical studies. However translational studies are essential to the progress of medicine and improvements in the health of individuals and must not be abandoned because the task of complying with the regulatory authorities appears too daunting. The new requirements have generated major challenges to the conduct of multinational multicentre prospective studies and the past few years have witnessed a sharp learning curve for academic clinicians. This Chapter has summarised the recent changes and the major challenges and has provided some guidance for the development of new protocols. Stem cell transplantation has been at the forefront of translational haematological research for many years and the Board of the EBMT has committed the organisation to the continuation of this tradition. Within the EBMT, clinical studies are generated by the individual Working Parties and the EBMT Clinical Trials Office now provides the infrastructure for the regulatory process. EBMT members are encouraged to make use of this expertise to ensure the continued development of our speciality. 354
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. What was the first international agreement on the ethical conduct of clinical trials? a) Declaration of Helsinki . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) International Conference on Harmonisation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The Nuremberg Code . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The Clinical Trials Directive 2001/20/EC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Who has legal responsibility for a clinical trial in a participating Institution? a) Funding body . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The Party that insures the study . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The Institution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The Sponsor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Who/what issues a Clinical Trials Authorisation? a) European Medicines Evaluation Agency (EMEA) . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The Competent Authority of each country . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The National Ethics Committee . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The EudraCT Registration Database . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. What Phase of Study is also referred to as a Therapeutic Exploratory Study? a) Phase I . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Phase II . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Phase III . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Phase IV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. What is the timeline for reporting a life-threatening SUSAR to the Competent Authorities? a) 24 hrs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 2 days . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 5 days . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 7 days . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 20
HSCT for acute myeloid leukaemia in adults
R. Varaldo, F. Frassoni
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CHAPTER 20 • AML in adults
1. Introduction Acute myeloid leukaemia (AML) is a heterogeneous clonal disorder of haematopoietic progenitor cells which lose the ability to differentiate normally and to respond to normal regulators of proliferation. AML is the most common adult leukaemia, with an annual incidence of approximately 2.4 cases per 100,000 population up to the sixth decade, rising to 16–18 cases in the seventh decade (1). The median age at presentation is approximately 68 years but the majority of data on AML treatment refer to younger patient (<60 years). Thus, 50% or more patients are generally neglected by the majority of clinical trials and reports in the journals and at meetings. AML is initially with chemotherapy (CT) to induce remission. The treatment plan involves a remission induction phase aimed to establishing a complete remission (CR) and a post-induction phase aimed to eradicating/reducing residual disease. Combination CT induces CR in an average of 60 to 80% of adults aged less than 60 years. Once CR has been achieved, further intensive therapy is needed to prevent relapse. Post-remission therapy for patients under the age 60 includes CT or HSCT. HSCT is one type of post-remission therapy. In general, HSCT are performed after 2 or 3 courses of chemotherapy. When HSCT is performed in remission, the leukaemic mass has been reduced by least of two logs with respect to the onset of the disease. Who should be transplanted for AML? In 2007 this is still an open question. To find the answer to this question, one of the most important areas of interest in the treatment of AML over the past decade has been the clarification of prognostic factors, leading to much-improved risk determination. All major cooperative groups that have studied the treatment of AML have identified cytogenetic abnormalities at presentation and time to achieve remission as the major disease-specific prognostic factors (2). Currently AML patients are stratified into 3 main risk categories based on cytogenetics, molecular biology and response to initial CT. The difficulty with using cytogenetic grouping as a risk factor is that most patients fall in the intermediate risk group, and the majority of these have a normal karyotype. Between 40 and 50% of young adult AML patients in Europe and the USA have been found to have a normal karyotype. The intermediate group, unlike the favourable or the unfavourable cytogenetics group, comprises a very heterogeneous mix of patients, with some having a poor prognosis and others having a better prognosis. However, molecular genetics is continuously defining new biological entities. This, in turn, identifies new prognostic factors and often moves the new AML subset into another prognostic group, particularly within the intermediate and favourable groups. Unfavourable prognosis is associated with several of these entities, including: - the Wilms tumour gene, WT1
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- the genes for the apoptosis regulators B-cell lymphoma protein 2 (BCL-2): BCL2, and BCL-2–associated X protein, BAX - the brain and acute leukaemia cytoplasmic gene, BAALC - the ecotropic viral integration site 1 gene, EVI1 - the FMS-like tyrosine kinase type 3 gene, FLT3 - KIT, ERG, and the mixed-lineage leukaemia gene, MLL. Some mutations of specific genes confer a more favourable prognosis; most notably, mutations in the genes for CCAAT enhancer binding protein-a (C/EBP-a), CEBPA, and nucleophosmin, NPM1 (3–5).
2. Autologous haematopoietic stem cell transplantation The available data indicate that better outcome (less relapse) is achieved when cells are harvested after the second or third course of chemotherapy. In principle, autologous HSCT has several potential advantages over allogeneic HSCT: - a matched donor is not needed - there is no risk of complications such as graft-versus-host disease (GvHD) - the convalescence period is much shorter and simpler - the immunological reconstitution is much faster and more complete - the work of nurses and physicians is much easier. However, there are three major important disadvantages: - the relapse rate is much higher, probably due to lack of graft-versus-leukaemia (GvL) effect and the contamination of the graft with leukaemic cells - there are often difficulties in collecting sufficient HSC - in high-risk leukaemia the role of autologous HSCT is minimal since the final outcome is dismal. In general, the results of autograft for AML have remained constant over the last 10 years. There has been a reduction of transplant related mortality (TRM) probably due to better supportive therapy. The leukaemia free survival (LFS) ranges from 40 to 50% at 3 years. The data of the Acute Leukaemia Working Party (ALWP) registry from 1996 to 2001, encompassing 2100 patients autografted for AML in 1CR indicate the following outcomes at 5 years: LFS = 43%, OS = 51%, RR = 53%, TRM = 9% (unpublished data from ALWP-EBMT). This is an important achievement, especially considering that autologous HSCT can be now offered to patients up to 65 years and beyond. Auto-HSCT produces durable second remission in 25–30% of patients with relapsed AML who achieved a second CR, but its low TRM is offset by a high relapse rate (6). For a number of reasons auto-HSCT has become less appealing in recent years. From the scientific point of view the reason lies in the fact that there has not been any 358
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improvement in methods of reducing relapse. There are centres which continue to perform auto-HSCT using purged graft. Their results are generally superior to those reported with the unpurged marrow. However, since there has never been a randomised study, the majority of AML experts remain sceptical about this manoeuvre. In the autografting setting, it is still unclear whether a better outcome can be obtained with PB compared with BM. How many patients reach the possibility of undergoing auto-HSCT? One of the major drawbacks of autografting studies in AML has been the scarce accrual of patients. In general no more than 50 to 60% of patients who in principle were allocated to auto-HSCT actually reach the transplant. This has mainly been caused by early relapse of leukaemia, or the harvest of an insufficient haematopoietic cell graft. Currently, autografting is restricted to older patients, patients with APL in second molecular remission and younger patients lacking a sibling or matched unrelated donor.
3. Allogeneic haematopoietic stem cell transplantation Allo-HSCT has had a tremendous relevance in AML. In the last 20 years, thousands of AML patients have been cured by allo-HSCT. These patients would have otherwise died of their disease. In the last five years a variety of data have emerged which clearly indicate that: - the results of MUD have improved and are still improving - the results of cord blood transplant are equivalent to the results of MUD transplants and seem to have much more scope for improvement in the near future - the age limit remains but older patients are increasingly considered for a transplant and several strategies have been designed for reducing transplant-related toxicity. A transplant consists of several procedures. Several tasks need to be accomplished for a successful transplant: - reducing the tumour burden (it is believed that the conditioning regimen reduces the tumour burden by 3-4 logs) - immunosuppression; without this the risk that the graft is rejected is very high - the prevention of GvHD - the prevention and treatment of the infections (bacterial and fungal) linked to neutropenia in the first phase of transplant and the later viral and fungal infections related to the slow recovery of T-cells and immune function in general. The wide variation in the preparative regimens used is one of the major causes of the differences in outcome in different studies. Besides the considerable variation in the regimens utilised to prepare the patient for HSCT, there are also different sources of haematopoietic cells that can be used. The goal of performing a
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transplant which maintains the positive effect of allo-reactivity while removing the detrimental effect is being approached but a true “intelligent manipulation” has yet to come. The HLA barrier remains the most important factor. The best results are those using an HLA compatible family donor. Most patients do not have a compatible HLA family donor. Thus, the use of partially matched family donors and matched unrelated donors is increasing. 3.1. HLA compatible sibling transplant This type of HSCT is considered the standard reference for any other transplant. There has been a considerable improvement of the results starting from 1990 compared with the antecedent period, and there is evidence of ongoing improvement. The current results for AML patients transplanted in CR1 give a probability of DFS ranging from 45 to 70% and the EBMT registry data of the last 5–7 years indicate approximately 55–60% DFS. With respect to the past, centres are now transplanting older patients. There is a variation among centres depending on the type of patients transplanted and on the experience and skill of the Centre (7). The relapse rate continues to be approximately 20–25% for patients with intermediate risk. The current results from the ALWP-EBMT registry are shown in Figure 1.
Figure 1: Myeloablative HSCT in AML - HLA compatible sibling donor 2000-2007 ALWP-EBMT Leukaemia Free Survival 1.0
0.8
CR1 patients (n=2808) 55 ± 1% 0.6
0.4
CR2 patients (n=451)
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0
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CHAPTER 20 • AML in adults
3.2. Matched unrelated donor transplants This type of HSCT, utilising the worldwide donor registry network, represents the major effort of transplant centres in the last decade. There has been a significant effort to create a network from which matched unrelated donors can be available and the network encompasses now more than 10 million potential donors. Several studies have shown that the results of matched unrelated donor (MUD) transplants can approach those of transplant using a HLA compatible sibling donor. The continuous improvement in HLA typing is associated with improved results. However, while the outcome is improving, this type of transplant is still associated with considerable morbidity and mortality. New and interesting aspects of HLA matching are also emerging (8, 9) which may be useful in guiding donor selection. At the same time the high resolution HLA typing restricts the number of suitable donors. In addition, although the time from the beginning of search has shortened, a number of patients with AML often need a transplant in a very short period of time. These are some of the reasons why alternative source such as Cord Blood Unit is emerging as an alternative strategy. The current results from the ALWP-EBMT registry since the year 2000 are shown in Figure 2.
Figure 2: Myeloablative HSCT in AML - matched unrelated donors 2000-2007 ALWP-EBMT Leukaemia Free Survival 1.0 0.8
0.6
CR1 patients (n=708) 46 ± 3%
0.4
CR2 patients (n=424) 37 ± 4% 0.2
0.0
0
1
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3.3. Source of haematopoietic stem cells In the past decade, the use of peripheral blood (PB) as a source of haematopoietic stem cells for allogeneic transplantation has increased considerably. A large number of retrospective studies and also several prospective randomised trials have compared the outcome of patients receiving allografts with bone marrow (BM) versus PB, using an identical sibling; in all studies PB has resulted in faster engraftment and shorter hospital stay. In most studies the incidence and severity of acute GvHD (aGvHD) (10, 11) has been similar with PB and BM; PB on the other hand has been associated with more chronic GvHD (cGvHD). In the same period, other studies on BM and PB transplantation have drawn attention to the importance of the dose of stem cells infused; a lower transplantrelated mortality (TRM) and, in some studies, a lower relapse rate after transplantation have been observed in patients undergoing allografting who receive a higher stem cell dose (12); however, this holds only for BM transplant recipients. Finally, the available data suggests that with standard conditioning the outcome for patients with low-risk disease might be better with BM; for patients with high-risk disease it might be better with PB grafts. 3.4 Umbilical cord blood transplants (CBT) Two relatively recent publications have shown that CBT are not inferior to matched unrelated transplants in acute leukaemias (13, 14). In addition, if CBT could overcome the problem of delayed/failure to engraft it is likely that it could become the best choice for unrelated transplants (9, 13–15). In fact compared with MUD several advantages exist: - less restrictive HLA compatibility requirements - time to transplant is far more rapid - absence of any risk for the donor. Currently, several means to improve engraftment are being evaluated. They include: cell expansion, double cord transplant, co-infusion of haploidentical donor CD34 selected cells, co-infusion of MSC, and direct intra-bone transplant. In addition, there are several reports indicating that high resolution matching might improve the outcome of UCB transplants (9); this would mean there would be a need to expand the number of banked units. 3.5. Haplo-identical transplants Haplo-identical transplant has been developed by the Perugia group. The approach is based on the concept of complete T-cell depletion and the use of megadoses of stem cells (16, 17). No GvHD prophylaxis is given. The most recent publication reports
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CHAPTER 20 • AML in adults
an event-free survival rate was 48 ± 8% and 46 ± 10%, respectively, for the 42 AML and 24 ALL patients receiving transplantation in remission. Most patients with advanced disease simply do not make it to transplantation. In other words, those patients who do receive MUD transplants for advanced AML are a select group who survived long enough to undergo transplantation. In comparison, haploidentical transplant recipients represent a group of patients who are transplanted much sooner; many of them would never have survived long enough to undergo an MUD transplantation. Finally, a relevant impact of NK allo-reactivity in controlling AML is increasingly being shown (16, 18). From these studies the possibility of utilising NK allo-reactivity outside the transplant setting has been derived. 3.6. Reduced-intensity conditioning Reduced-intensity conditioning has become an accepted treatment modality. Despite its wide use, to date there have been no prospective comparative studies. Available information is based on data from phase II clinical studies (19) and some retrospective comparative data. The best data in patients with AML have come from a multi-centre study in the USA and Europe, referred to as the Consortium Study (20). One hundred and twenty-two AML patients, with approximately equal numbers of related and unrelated donor-recipient pairs, were included in this study. The most important reason for reduced-intensity conditioning was age or significant comorbidity. Engraftment was prompt in all patients. The overall survival rate at 2 years was 48%, and patients receiving transplantation during CR1 had 2-year overall survival rates of 44% (related HSCT) and 63% (unrelated HSCT). Cumulative incidences of acute GvHD (grades 2–4) were 35% at 180 days after related HSCT and 42% after unrelated HSCT. The probability of one of the most important complications, chronic GvHD, was 36% at 2 years (20). The preponderance of data has established reduced-intensity conditioning as a feasible option, but how feasible and how important this is in the management of AML, and how much of a difference it will make, still needs to be established in prospective studies. The only solid fact which argues in favour of reduced-intensity conditioning transplants (RIC) may be that with RIC one can offer a transplant to patients who otherwise would not have any other valid therapeutic options. The current results of RIC from the ALWP-EBMT registry are shown in Figures 3 and 4. EBMT is currently investigating in a prospective randomised trial the role of RIC HSCT in elderly patients with AML.
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Figure 3: RIC HSCT in AML - HLA compatible sibling donor 2000-2007 ALWP-EBMT Leukaemia Free Survival 1.0
0.8
0.6
CR1 patients (n=953) 41 ± 1% 0.4
CR2 patients (n=210) 37 ± 5% 0.2
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0
1
2
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Figure 4: RIC HSCT in AML - matched unrelated donors 2000-2007 ALWP-EBMT Leukaemia Free Survival 1.0
0.8
0.6
CR1 patients (n=228) 41 ± 5% 0.4
CR2 patients (n=157) 42 ± 5% 0.2
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0
1
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3
4
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3.7. Comparison of results of different strategies and the place of allogeneic HSCT in the management of AML There have been four large studies that have attempted to address which modality is better – allogeneic HSCT, autologous HSCT, or chemotherapy alone. These studies have all identified patients at diagnosis and then assigned patients in first remission to allogeneic transplantation if they have a matched sibling donor, randomising the remaining patients to consolidation chemotherapy or autologous transplantation (21–23). In recent years, a popular method for assessing the role of allo-HSCT in acute leukaemia has been the evaluation of whether having a family HLA matched donor was associated with an improvement in survival: the so call donor versus no-donor analysis. This method has produced conflicts in the interpretation of the results. The most recent large and important study analysing the advantage of having a family donor refers, however, to acute lymphoblastic leukaemia. A donor versus no donor analysis showed that the survival difference was significant in standard-risk patients, but not in high-risk patients (24). Recently data from some reported studies were subjected to a meta-analysis (25). The analysis was designed to include studies that had evaluated the outcome of patients with AML in CR1 by availability of a related donor using an intention to treat approach. Four of the studies had demonstrated a benefit with respect to DFS in favour of patients with a donor but as single studies this was not translated in a benefit in OS; however, an advantage in OS was demonstrated for the whole cohort of patients with donors included in the meta-analysis. When analysed by cytogenetic risk group, a significant advantage was observed for poor-risk patients with donors. For the favourable and intermediate risk group patients a survival benefit was not observed. Most experts today consider that the effect of allogeneic stem cell transplantation needs to be re-assessed in specific biological subgroups taking account, as well as “classic “ cytogenetic stratification risk, a variety of new molecular biological data (mainly based on minimal residual disease assays) with the aim of defining a tailored therapy as has already been demonstrated in the in paediatric acute leukaemia setting. In any case, for patients with high-risk features and beyond CR1, allo-HSCT remains the sole possibility of surviving.
4. Current strategy for patients with AML Taking account the above limitations, at present most experts in the field of AML treatment would suggest the following strategy:
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- For good-risk patients in first CR chemotherapy seems not inferior to transplant strategies. However, one must be sure that the schedule and protocols have been proven to provide a DFS above 60%. This because the results of HSCT now approach that outcome - Acute promyelocytic leukaemia (APL) is never recommended for a transplant if the PCR is negative because the results of CT alone are excellent. If APL patients are not in molecular remission or are in overt relapse auto or allo-BMT are an option - For intermediate risk patients, who represent the majority of patients with AML, allo-HSCT probably remains the best option if a HLA matched sibling donor is available. If a HLA compatible sibling is unavailable, autograft may be a possible option; however, this choice is increasingly challenged by the results of MUD and especially of CBT - For poor-risk patients in CR1 and for all other patients who relapse during and following the planned therapy the chance of surviving without a transplant is very low. For those patients an early transplant strategy should be organised. In this area, the use of an alternative donor (see MUD or cord blood or haplo-identical transplant) represents the sole possibility of survival. In this respect, the early identification of high-risk patients is very important since the timing of a search for an alternative source of haematopoietic stem cell may be crucial for the life of the patient.
5. Conclusion There has been a paradigm shift in the field of AML treatment. Disease risk can be assessed much better than ever before and a classification into low, intermediate and high-risk should become possible for all patients. In parallel, the risks of the transplant can also be better assessed, based on patient and donor characteristics as outlined in above. In order to implement future strategies, every patient with newly diagnosed AML should have a complete assessment of disease risk and a search for a donor at diagnosis. Depending on disease risk and transplant risk, algorithms for treatment after induction and consolidation might appear as follows and be separated into planned HSCT in CR1 or delayed HSCT (see Table 1) (26).
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Table 1: Transplant algorithm for AML Transplant risk according to extended EBMT risk score (see indications)
Disease risk Transplants in 1st CR Low risk Intermediate risk High risk
risk score 0-1 risk score 0-2 risk score 0-5
Transplants for refractory diseases
risk score 0-6
Transplants in Low risk High risk No transplant
2nd
CR risk score 0-3 risk score 0-6 risk score >6
References 1. Ries L, Eisner M, Kosary C, et al. SEER Cancer Statistics Review, 1975-2001, National Cancer Institute. http://seer.cancer.gov/csr/1975_2001/ Bethesda, MD: National Cancer Institute 2004. 2. Wheatley K, Burnett AK, Goldstone AH, et al. A simple, robust, validated and highly predictive index for the determination of risk-directed therapy in acute myeloid leukaemia derived from the MRC AML 10 trial. Brit J Haematol 1999; 107: 69-79. 3. Kottaridis PD, Gale RE, Frew ME, et al. The presence of a FLT3 internal tandem duplication in patients with acute myeloid leukemia (AML) adds important prognostic information to cytogenetic risk group and response to the first cycle of chemotherapy: Analysis of 854 patients from the United Kingdom Medical Research Council AML 10 and 12 trials. Blood 2001; 98: 1752-1759. 4. Preudhomme C, Sagot C, Boissel N. Favorable prognostic significance of CEBPA mutations in patients with de novo acute myeloid leukemia: A study from the Acute Leukemia French Association (ALFA). Blood 2002; 100: 2717-2723. 5. Falini B, Nicoletti I, Martelli MF, Mecucci C. Acute myeloid leukemia carrying cytoplasmic/ mutated nucleophosmin (NPMc+ AML): Biologic and clinical features. Blood; 109: 874-885. 6. Breems DA, Löwenberg B. Acute myeloid leukemia and the position of autologous stem cell transplantation. Semin Hematol 2007; 44: 259-266. 7. Frassoni F, Labopin M, Powles, et al. Acute Leukaemia Working Party of the European Group for Blood and Marrow Transplantation. Effect of centre on outcome of bone-marrow transplantation for acute myeloid leukaemia. Lancet 2000; 355: 1393-1398. 8. Shaw BE, Gooley TA, Malkki M, et al. The importance of HLA-DPB1 in unrelated donor hematopoietic cell transplantation. Blood 2007; 110: 4560-4566. 9. Eapen M, Rubinstein P, Zhang MJ, et al. Outcomes of transplantation of unrelated donor umbilical cord blood and bone marrow in children with acute leukaemia: A comparison study. Lancet 2007; 369: 1947-1954.
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10.Flowers M, Parker P, Johnston L, et al. Comparison of chronic graft versus host disease after transplantation of peripheral blood stem cells versus bone marrow in allogeneic recipients: Long term follow up of a randomized trial. Blood 2002; 100: 415-419. 11.Schmitz N, Eapen M, Horowitz MM, et al. Long-term outcome of patients given transplants of mobilized blood or bone marrow: A report from the International Bone Marrow Transplant Registry and the European Group for Blood and Marrow Transplantation. Blood 2006; 108: 4288-4290. 12.Gorin NC, Labopin M, Rocha, et al. European Cooperative Group for Blood and Marrow Transplantation Acute Leukaemia Working Party. Marrow versus peripheral blood for geno-identical allogeneic stem cell transplantation in acute myelocytic leukaemia: Influence of dose and stem cell source shows better outcome with rich marrow. Blood 2003; 102: 3043-3051. 13.Laughlin MJ, Barker J, Bambach B, et al. Hematopoietic engraftment and survival in adult recipients of umbilical-cord blood from unrelated donors. N Engl J Med 2001; 344: 1815-1822. 14.Rocha V, Labopin M, Sanz G, et al. Transplants of Umbilical-Cord Blood or Bone Marrow from Unrelated Donors in Adults with Acute Leukemia. N Engl J Med 2004; 351: 22762285. 15.Barker JN, Weisdorf DJ, DeFor TE, et al. Transplantation of 2 partially HLA-matched umbilical cord blood units to enhance engraftment in adults with hematologic malignancy. Blood 2005; 105: 1343-1347. 16.Ruggeri L, Capanni M, Urbani E, et al. Effectiveness of donor natural killer cell alloreactivity in mismatched hematopoietic transplants. Science 2002; 295: 2097-2100. 17.Aversa F, Terenzi A, Tabilio A, et al. Full haplotype-mismatched hematopoietic stem-cell transplantation: A phase II study in patients with acute leukemia at high risk of relapse. J Clin Oncol 2005; 23: 3447-3454. 18.Miller JS, Cooley S, Parham P, et al. Missing KIR ligands are associated with less relapse and increased graft-versus-host disease (GvHD) following unrelated donor allogeneic HCT. Blood 2007; 109: 5058-5061. 19.McSweeney PA, Niederwieser D, Shizuru JA, et al. Hematopoietic cell transplantation in older patients with hematologic malignancies: Replacing high-dose cytotoxic therapy with graft-versus-tumor effects. Blood 2001; 97: 3390-3400. 20.Hegenbart U, Niederwieser D, Sandmaier BM, et al. Treatment for acute myelogenous leukemia by low-dose, total-body, irradiation-based conditioning and hematopoietic cell transplantation from related and unrelated donors. J Clin Oncol 2006; 24: 444-453. 21.Suciu S, Mandelli F, de Witte T, et al. Allogeneic compared with autologous stem cell transplantation in the treatment of patients younger than 46 years with acute myeloid leukaemia (AML) in first complete remission (CR1): An intention-to-treat analysis of the EORTC/GIMEMAAML-10 trial. Blood 2003; 102: 1232-1240. 22.Zittoun RA, Mandelli F, Willemze R, et al. Autologous or allogeneic bone marrow transplantation compared with intensive chemotherapy in acute myelogenous leukaemia. N Engl J Med 1995; 332: 217-223. 23.Burnett AK, Goldstone AH, Stevens R, et al. Randomised comparison of addition of 368
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autologous bone-marrow transplantation to intensive chemotherapy for acute myeloid leukaemia in first remission: Results of MRC AML 10 trial. Lancet 1998; 351: 700-708. 24.Goldstone AH, Richards SM, Lazarus HM, et al. In adults with standard-risk acute lymphoblastic leukemia (ALL) the greatest benefit is achieved from a matched sibling allogeneic transplant in first complete remission (CR) and an autologous transplant is less effective than conventional consolidation/maintenance chemotherapy in ALL patients: Final results of the international ALL trial (MRC UKALL XII/ ECOG E2993). Blood 2007 Nov 29; [Epub ahead of print]. 25.Yanada M, Matsuo K, Emi N, Naoe T. Efficacy of allogeneic hematopoietic stem cell transplantation depends on cytogenetic risk for acute myeloid leukemia in first disease remission: A metaanalysis. Cancer 2005; 103: 1652-1658. 26.Breems DA, Van Putten WL, Huijgens PC, et al. Prognostic index for adult patients with acute myeloid leukemia in first relapse. J Clin Oncol 2005; 23: 1969-1978.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. For intermediate risk AML patients in CR1 what is the probability of relapsing after HLA identical sibling transplant? a) 30–40% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 20–25% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 40–50% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 20% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. The recent meta-analysis of studies comparing chemotherapy, allo-transplant, and auto-transplant has shown better OS for allo-transplant in: a) Poor-risk cytogenetic patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Intermediate risk patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Favourable risk patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) a+b . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Good risk AML is characterised by involvement of which of the following genes? a) FLT3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) EV1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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c) WT1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) None of the above . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. In RIC unrelated transplant the incidence of chronic GvHD at 180 days is: a) 20–25% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 40–45% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 10–20% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 55–60% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. What is the current actuarial DFS at 3 years for AML patients autografted in CR1? a) 20–30% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 70–80% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 40–50% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 10–20% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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*
CHAPTER 21
HSCT for acute lymphoblastic leukaemia in adults
N. Gökbuget, D. Hoelzer
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CHAPTER 21 • ALL in adults
1. Introduction Haematopoietic stem cell transplantation (HSCT) has gained an increasingly important role in the treatment of adult ALL. The majority of large prospective studies in adult ALL have addressed the issue of HSCT, but indications for HSCT in first CR, scheduling and procedures are still not satisfactorily defined (1, 2). The advantages of HSCT (short treatment duration, favourable outcome in some trials) must be compared to the disadvantages (mortality and morbidity, late complications) and assessed in relation to the outcome of increasingly effective conventional and targeted chemotherapy regimens. The focus of debate is still the question whether all adult ALL patients with a sibling donor should proceed to HSCT, or only those with specific risk factors.
2. Indications for transplantation Most European ALL study groups define an indication for HSCT in patients with unfavourable prognostic factors associated with a survival probability of less than 40% with chemotherapy alone (3). Both matched related (SIB) and matched unrelated (MUD) HSCT are considered as alternatives. The prognostic models differ slightly as well as the upper age limits. The presence of detectable minimal residual disease (MRD) beyond first consolidation chemotherapy (CT) provides an important new indication for HSCT. Two major questions can be raised about MRD based indications for HSCT: - Is HSCT is really a recommendable option in patients with high MRD, since they are prone to a higher relapse rate after HSCT? - Is HSCT justified in patients with conventional high-risk (HR) features but negative MRD status? Some study groups define an indication for SIB-HSCT in all patients with a sibling donor. Most recently this recommendation was reconsidered and SIB-HSCT particularly suggested for young standard risk (SR) patients (4). This is in contrast to the strategy of adopting paediatric intensive chemotherapy regimens, rather than HSCT, in adolescents and young adults, in order to avoid acute mortality and long-term effects. There is general agreement that all patients in 2nd or later remission are candidates for HSCT. This includes molecular relapse, defined as reappearance of MRD above 10-4–10-3. In advanced disease, depending on donor availability and general condition, experimental HSCT procedures (see below) may be considered, ideally within clinical trials. 2.1. Evidence-based recommendations for HSCT in adult ALL An evidence based review underlined that HSCT offers an advantage compared to
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chemotherapy in HR patients and in 2nd remission (5). A meta-analysis of 7 studies showed a broad variation in terms of the proportion of patients allocated to transplant who actually received a transplant, which varied between 68–96% for allo- and 9–81% for auto-HSCT. Outcome in patients with a donor was correlated with the proportion who actually received an allo-HSCT. Again the survival for HSCT was superior to chemotherapy; the advantage was particularly evident in HR patients (6). However the role of allo-HSCT in SR ALL remains unclear. Table 1 outlines the indications for HSCT in the German Multicenter Study Group for adult ALL (GMALL) as an example of a risk adapted approach.
Table 1: Indications for HSCT in the GMALL trials Indication
Priorities *
Controversies
High Risk
All patients within 3–4 months from diagnosis
1. Allo sibling 2. Allo MUD**
No HSCT in MRD negative pts
Standard Risk
Molecular non-responders
See above
Allo sibling HSCT in all young SR pts with donor
Relapse (including molecular relapse)
All patients in 2nd CR (if necessary in good PR or early relapse)
See above (consider cord blood or haploidentical HSCT if no donor available)
*Decision depends on age, patient’s general condition and donor availability. **Matched or one antigen mismatch
3. Role and outcome of HLA-identical sibling transplant According to the EMBT and CIBMTR registries, survival after SIB-HSCT in 1st CR is about 48–49% in adults (7, 8). Relapse rate (RR) and transplant related mortality (TRM) range both between 25–30%. Although TRM is strongly correlated with age, the upper age-limit for SIB-HSCT has increased continuously up to 50–55 years. Survival after HSCT is poorer for patients in 2nd remission (29–34%) and with advanced disease (15–18%) (7, 8). This is mainly due to an increasing RR.
4. Role and outcome of matched unrelated (MUD) allogeneic transplant The role of MUD becomes more important since only 1/3 of patients have a matched sibling donor. The survival for MUD-HSCT in 1st CR is around 42–45%, with a lower RR and higher TRM (around 32%) compared to SIB-HSCT (7, 8). It has to be borne 374
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in mind that MUD series generally comprise selected HR patients. In 2nd remission MUD-HSCT results in a long term survival of 28% and in advanced disease of around 11%, due to a higher RR but particularly to a higher TRM of 43–48% (7). Due to improved supportive care, better donor selection and extension of indications beyond very high-risk patients, the results of MUD-HSCT are nowadays almost equivalent to those of SIB-HSCT.
5. Role and outcome of autologous transplant According to published studies the overall survival of patients receiving autoHSCT in 1st CR is 42%. The major problem is the high RR of 50% or more. The intensity of pre-treatment has an important impact on outcome of auto-HSCT, since it leads to a reduction of tumour load. Thus auto-HSCT may be an option in HR patients with negative MRD status. Maintenance therapy after HSCT e.g. with mercaptopurine and methotrexate, or imatinib in Ph-positive ALL – particularly in MRD positive patients – is also an effective approach. Several randomised studies showed no difference for the comparison of chemotherapy versus auto-HSCT or even a significantly inferior outcome for auto-HSCT (4). Most comparisons of allo- and auto-HSCT have shown an inferior outcome of auto-HSCT.
6. Comparisons of chemotherapy and HSCT based approaches To circumvent the problem with comparability of HSCT and chemotherapy, several groups have designed prospective trials with a “genetic” randomisation offering SIBHSCT in CR1 to all patients with donor. Results certainly strongly depend on the compared “conventional” treatment approach and on whether those patients with a donor actually receive HSCT. The majority of studies did not show an overall advantage for patients with donor; however in the HR group generally patients with a donor had a superior outcome (1). Recently the ECOG/MRC group reported their results comparing patients with a donor (SIB-HSCT) to those without donor (randomised comparison of chemotherapy and auto-HSCT). The special feature of this trial was the use of age (< or >35 years) as a prognostic factor. SR patients were by definition younger than 35 years without adverse prognostic factors. Phnegative patients with a donor had a superior OS (53%) compared to those without a donor (45%), mainly due to a lower RR. The difference was particularly evident in SR (OS 62 vs. 52%) but not in HR (OS 41 vs. 35%) patients (4). The TRM reached 20% even in the young SR patients and 39% in the older HR patients. Two conclusions can be drawn: - The outcome of HSCT is better in younger patients - In older HR patients the outcome is similarly poor with chemo and HSCT.
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It is likely that outcome with optimal chemotherapy is similarly good or even better than HSCT in young SR patients, as demonstrated in paediatric studies. For older HR patients the results underline the need to improve conditioning regimens and reduce pre-transplant morbidity and TRM.
7. Role and outcome of haploidentical transplant For haploidentical HSCT experience is restricted mainly to paediatric patients, where it may be considered in patients without donor and with urgent need of HSCT. In adults haploidentical HSCT is an experimental approach and should be restricted to specialised centres and later stage of disease, and should preferably be performed within clinical trials.
8. Role and outcome of cord blood transplant (UCB) The experience with UCB transplantation in ALL mainly comes from paediatric patients. In adults the limited cell dose is one of the major obstacles. Initial registry results for younger adults with acute leukaemia indicate however that UCB (single or double) can be considered as an alternative source, if available (9).
9. Conditioning There is also no standard for the conditioning regimen before HSCT in ALL. Most regimens are based on total body irradiation (TBI). The usual dose is 12 Gy. TBI is most frequently combined with cyclophosphamide (Cy) or VP16. A recent analysis of EBMT registry patients showed no difference in outcome for SIB-HSCT with TBI/VP16 or TBI/Cy in CR1. In second remission the TBI/VP16 combination was associated with a lower relapse risk (10). Clearly inferior results were reported for busulfan-based preparative regimens compared to TBI based regimens. With RICHSCT one third of patients may survive if transplant is performed in 1st CR (11).
10. Nature and role of MRD monitoring after transplant After transplantation regular evaluation of chimerism and MRD shows the individual course of disease. The outcome after HSCT is influenced by the status of MRD before and after HSCT. In Ph-positive ALL it has been demonstrated that in patients with MRD after HSCT the survival was significantly superior in those who rapidly responded to imatinib compared to those who did not respond (12). The outcome of patients with a high level of MRD before HSCT is poor. Therefore MRD status must be considered both before and after HSCT, to decide on additional treatment either before HSCT in order to reduce tumour load or after HSCT to prevent relapse.
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10.1. Nature and role of any additional cellular or chemotherapy post-transplant In early relapse, preferably molecular relapse, immunologic treatments such as reduction of GvHD prohylaxis and/or application of donor lymphocytes are promising approaches for preventing overt relapse. In Ph-positive ALL post-transplantation treatment with imatinib – either up-front or after detection of MRD – appears to be a very successful approach.
11. Stem cell transplantation in Ph+ ALL Due to the poor outcome with intensive chemotherapy, HSCT has always been the treatment of choice for Ph+ ALL. The survival after allo-HSCT in first CR ranges between 27–65% (13). The RR is higher than in Ph-negative ALL and the outcome is compromised by TRM due to the higher median age of Ph+ ALL patients. Nowadays the majority of patients with Ph+ ALL receive imatinib as front-line therapy. Apparently there is no increase in TRM if HSCT is performed thereafter. The outcome of HSCT in Ph-positive ALL is strongly correlated with the level of MRD before and after HSCT and with the use of imatinib as part of the post-transplantation strategy (see above).
12. Prognostic factors Prognostic factors are not only used to identify HR patients who could benefit from HSCT. To an increasing extent prognostic factors have been described which help to estimate the risks of the HSCT itself, such as patient age, donor characteristics, degree of matching etc. Bringing both risk estimates together will in future allow a more accurate definition of indications for HSCT. Besides age, scoring of comorbidities may assist decision-making regarding indication for HSCT and the intensity of conditioning.
13. Future options in HSCT Large national and even international study groups are committed to the development of chemotherapy schedules and optimal integration of HSCT in front-line therapy. One important point is the balance between efficacy and toxicity of pre-transplant treatment and preparative regimens, in order to reduce TRM. Also the optimal timing of HSCT has to be defined. For an improvement of outcome after allogeneic HSCT a reduction in both RR and TRM is required. High resolution HLA typing and improved infection prophylaxis are important in this respect. For patients without a donor or with a contraindication to conventional HSCT alternative approaches need to be explored. MRD evaluation before and after HSCT is of interest, particularly in order to decide
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on maintenance therapy and immunotherapy such as donor-lymphocyte infusions. The prophylactic application of donor lymphocytes may be considered in patients with no or low GvHD.
References 1. Gökbuget N, Hoelzer D. Treatment of adult acute lymphoblastic leukemia. Hematology Am Soc Hematol Educ Program 2006: 133-141. 2. Bachanova V, Weisdorf D. Unrelated donor allogeneic transplantation for adult acute lymphoblastic leukemia: A review. Bone Marrow Transplant 2007 Oct 29; [Epub ahead of print]. 3. European Leukemia Study Registry. www.leukemia-net.org 2008. 4. Goldstone AH, Richards SM, Lazarus HM, et al. In adults with standard-risk acute lymphoblastic leukemia (ALL) the greatest benefit is achieved from a matched sibling allogeneic transplant in first complete remission (CR) and an autologous transplant is less effective than conventional consolidation/maintenance chemotherapy in All patients: Final results of the international ALL trial (MRC UKALL XII/ ECOG E2993). Blood 2007 Nov 29; [Epub ahead of print]. 5. Hahn T, Wall D, Camitta B, et al. The role of cytotoxic therapy with hematopoietic stem cell transplantation in the therapy of acute lymphoblastic leukemia in adults: An evidence-based review. Biol Blood Marrow Transplant. 2006; 12: 1-30. 6. Yanada M, Matsuo K, Suzuki T, Naoe T. Allogeneic hematopoietic stem cell transplantation as part of postremission therapy improves survival for adult patients with high-risk acute lymphoblastic leukemia: A metaanalysis. Cancer 2006; 106: 1657-1663. 7. Chaidos A, Kanfer E, Apperley JF. Risk assessment in haemotopoietic stem cell transplantation: Disease and disease stage. Best Pract Res Clin Haematol. 2007; 20: 125154. 8. Loberiza F. Summary Slides 2003 - part III. IMBTR/ABMTR Newsletter 2006; 10: 6-9. 9. Rocha V, Labopin M, Sanz G, et al. Transplants of umbilical-cord blood or bone marrow from unrelated donors in adults with acute leukemia. N Engl J Med 2004; 351: 2276-2285. 10.Marks DI, Forman SJ, Blume KG, et al. A comparison of cyclophosphamide and total body irradiation with etoposide and total body irradiation as conditioning regimens for patients undergoing sibling allografting for acute lymphoblastic leukemia in first or second complete remission. Biol Blood Marrow Transplant 2006; 12: 438-453. 11.Arnold R, Massenkeil G, Bornhauser M, et al. Nonmyeloablative stem cell transplantation in adults with high-risk ALL may be effective in early but not in advanced disease. Leukemia 2002; 16: 2423-2428. 12.Wassmann B, Pfeifer H, Stadler M, et al. Early molecular response to posttransplantation imatinib determines outcome in MRD+ Philadelphia-positive acute lymphoblastic leukemia (Ph+ ALL). Blood 2005; 106: 458-463. 13.Fielding AK, Goldstone AH. Allogeneic haematopoietic stem cell transplant in Philadelphiapositive acute lymphoblastic leukaemia. Bone Marrow Transplant. 2007 Oct 29; [Epub ahead of print].
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which post-transplantation strategy is most successful in Ph/BCR-ABL positive ALL a) Imatinib . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Mercaptopurine/methotrexate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) No treatment (to avoid toxicities) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) DLI only . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Is non-myeloablative HSCT an option in adult ALL? a) Not at all . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Generally preferable due to lower toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Within studies focussed on older and/or patient with comorbidities . . . . . d) Based on individual decision and patients wish . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. What are the results of allo sibling HSCT compared to matched unrelated SCT (MUD) in adult ALL? a) Sibling SCT clearly superior . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) MUD SCT clearly superior . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Similar overall results with higher relapse rate and lower TRM for sibling SCT. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Similar for matched or mismatched donors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. What are the results of autologous HSCT compared to chemotherapy? a) Similar in most studies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Inferior compared to chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Superior compared to chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Independent of minimal residual disease and post-transplant maintenance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which is the standard for conditioning in adult ALL? a) TBI based regimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Busulfan based regimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Non-myeloablative regimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Use of ATG and prophylactic DLI. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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CHAPTER 22
HSCT for myelodysplasia in adults
T. de Witte, G. Sanz
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CHAPTER 22 • Myelodysplasia in adults
1. Introduction Myelodysplasia (MDS) consists of a heterogeneous group of clonal stem cell disorders. The spectrum of MDS varies from a disease with an indolent course over several years to a form with rapid progression to acute myeloid leukaemia (AML). Since 1982, myelodysplasia has been classified according to the French–American–British (FAB) criteria (1). The new World Health Organization (WHO) classification system for MDS corrected several limitations of the original FAB classification in 1997 (2). An international workshop generated an International Prognostic Scoring System (IPSS) in 1997 (3). MDS usually occurs without a preceding provoking event (primary MDS), but treatment with radiotherapy and certain chemotherapeutic agents promotes the development of therapy-related (t-)MDS. Both alkylating substances and drugs targeted at topoisomerase II are capable of inducing t-MDS and t-AML. The majority of MDS patients are older than 60 years. For these patients supportive care, including new biologic response modifiers, is the mainstay of therapy. Allogeneic haematopoietic stem cell transplantation (HSCT) is usually considered the treatment of choice for most young MDS patients who have a histocompatible donor. Long-term disease-free survival (DFS) can be attained by these patients. For patients who lack a compatible donor, the outcome with autologous HSCT appears comparable with allogeneic SCT. Many clinicians consider allogeneic HSCT as the only curative treatment option. However, a retrospective study evaluating intensive chemotherapy alone versus chemotherapy followed by HSCT did not show a clear benefit for either chemotherapy alone or chemotherapy followed by HSCT.
2. Role of reduced-intensity conditioning (RIC) regimens in myelodysplasia The general age of patients with MDS is higher than 60 years and co-morbidity is rather common. For these reasons, RIC regimens have recently been increasingly used in MDS. The initial reports in MDS showed an encouraging low TRM compared with conventional conditioning. Kröger et al. reported on 37 MDS patients who were ineligible for conventionally conditioned HSCT. The actuarial DFS rate at 3 years was 38%, with a median follow-up of 20 months (4). More favourable results were reported following conditioning with fludarabine, busulfan, and alemtuzumab in 62 MDS patients. The 1-year DFS rates were 61 and 59% in patients with matched sibling donors (n=24) and unrelated donors (n=38), respectively (5). The EBMT analysed the outcomes of 215 RIC patients, and standard myeloablative conditioning (SMC) in 621 patients. In a multivariate analysis, the 3-year relapse rate was significantly increased after RIC (hazard ratio [HR] 1.61, P=0.001), but the 3-year non-relapse mortality (NRM) was decreased after RIC (0.61, P=0.015) (6).
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It is difficult to evaluate the contribution of RIC regimens to the improved outcome of allogeneic HSCT for MDS patients in view of the recently improved outcomes of HSCT with marrow ablative regimens and the heterogeneity of the patient populations (age, comorbidity, stage of disease). Therefore, the EBMT has launched a prospective randomised study comparing RIC regimens with standard conditioning regimens (for details, see: http://www.ebmt.org/5WorkingParties/CLWP/clwp8.html).
3. Role and outcome of autologous stem cell transplantation In view of the high relapse rate after chemotherapy alone, transplantation with autologous stem cells has been applied in an attempt to intensify post-remission therapy. One of the first reports by the EBMT on autologous HSCT in MDS showed a 2-year DFS of 34%. In a prospective study, 24 of the 39 candidates received an autologous HSCT, resulting in a median DFS of 29 months from transplantation (7). A European study compared the results of 100 patients who had entered CR after remission–induction chemotherapy and who were candidates for allogeneic and autologous HSCT, depending on the availability of an HLA-identical sibling. The 4year DFS rates in the group of patients with or without a donor were 31 and 27%, respectively (HR 0.93, 95% CI 0.57–1.52). This outcome suggested that patients with high-risk MDS might benefit from either allogeneic or autologous HSCT (8). A successful autograft is theoretically restricted to patients who achieve CR following induction chemotherapy, and in whom a suitable autologous harvest can be collected. Stem cell mobilisation was feasible in only 44/102 patients (43%) in the recovery phase after chemotherapy with G-CSF (9). This relatively low yield of a sufficient number of stem cells might reflect the low number of residual normal stem cells or the damage to the bone marrow stroma caused by pro-apoptotic cytokines produced by the MDS clone. It is clear that better mobilisation schedules and approaches should be developed before autologous peripheral HSCT can be recommended as part of the post-remission treatment of MDS patients treated with intensive anti-leukaemic therapy.
4. Role and outcome of HLA-identical sibling transplants Results of allogeneic HSCT have improved over time. The International Bone Marrow Transplant Registry (IBMTR) reported a 3-year DFS rate of 40% for 452 patients who underwent HLA-identical sibling HSCT for MDS performed between 1989 and 1997 (10). Deeg et al. reported favourable results in MDS patients treated with a busulfan-based regimen in which the busulfan dosage was adjusted to maintain blood levels at 800–900 ng/mL. The 3-year non-relapse mortality (NRM) rate was 28% with related donors (11). 382
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Cytogenetic abnormalities have a major influence on the outcome after HSCT. Using cytogenetic risk categories defined by the IPSS, the event-free survival rates for the poor-risk, intermediate-risk, and good-risk groups were 6, 40, and 51%, respectively (12). More advanced age (continuous variable) and long disease duration (>12 months) before HSCT were associated with an increased risk for treatment-related death after HSCT. This mandates consideration of HSCT early in the disease course. However, Cutler et al. showed that delayed HSCT results in maximised overall survival (OS) for low and intermediate-1 IPSS groups. They hypothesised that the optimal timing of HSCT in this cohort is at the time of development of a new cytogenetic abnormality, the appearance of a clinically important cytopenia, or an increase in the percentage of marrow blasts (13). Whether patients with advanced stage MDS should receive remission-induction chemotherapy prior to HSCT conditioning remains a point for debate. Retrospective analyses have reported conflicting data. Interpretation of the data is hampered by different selection biases in the two treatment approaches. Details regarding the type of chemotherapy administered are lacking in most studies. Preliminary analysis of the Criant study presented in 2005 showed that the majority of patients with an identified donor who were treated with remission-induction and consolidation chemotherapy received the planned HSCT (47/50). The 4-year DFS rate of the donor group was 46% – encouraging when compared with large-registry data (9). However, only prospective randomised studies can prove the benefit of remissioninduction therapy prior to transplant conditioning. The EBMT launched such a study in December 2006 (see http://www.ebmt.org/5WorkingParties/CLWP/clwp8.html for full details).
5. Role and outcome of matched unrelated allogeneic transplants Several reports have demonstrated that transplantation from matched unrelated donors (MUD) is a feasible and curative strategy for MDS patients. The National Marrow Donor Program (NMDP) reported on 510 MDS patients, transplanted between 1988 and 1998. At 2 years, the probability of DFS was 29%, and the cumulative incidence of TRM was 54% (14). A more recent report described the results obtained with a conditioning regimen of oral busulfan targeted to plasma concentrations of 800–900 ng/mL plus cyclophosphamide. The 3-year relapse-free survival of 64 MDS patients who underwent HSCT from MUDs was 59% and the NRM was 30% (11). The status of the disease at transplantation and the time from diagnosis to transplantation have shown close relationships with DFS after MUD transplantation for MDS. Patients with MDS secondary to chemo/radiotherapy have poorer DFS rates than patients with de novo MDS. There also appears to be an improvement in DFS
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rates in patients who have been transplanted in recent years (>1992). Other characteristics that have been associated with better DFS rates in some series are younger recipient age (continuous variable), greater cell dose, CMV seronegativity, and grades 0–I acute GvHD.
6. Role and outcome of cord blood transplants Experience with CB transplantations from unrelated donors for MDS patients is still very scarce. Ooi et al. from the University of Tokyo have published the outcomes of 13 patients with advanced MDS with a median age of 40 years (15). Despite the Figure 1: Algorithm for the management of MDS patients candidates for intensive therapy A
Marrow blasts <10% Hypocellular marrow/ Marrow fibrosis With matched donor
Without matched donor
<60–70 years Immediate alloSCT
<50 years Alt. donor alloSCT
50–65 years chemotherapy
CR AutoSCT
B
No CR Alt. Donor alloSCT
Marrow blasts >10% Chemotherapy CR
With matched donor# alloSCT
No CR No matched donor# With matched donor** autoSCT
alloSCT
No matched donor# If <50 years Alt. donor alloSCT
* Matched donor: HLA-identical sibling or HLA-matched unrelated donor; Alt. donor: all stem cell sources except stem cells from HLA-identical sibling or HLA-matched unrelated donor; # exception: good risk cytogenetics; ** exception: progressive disease after chemotherapy 384
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very advanced disease status at transplantation, 10/13 patients were alive and in CR, with an estimated 76% DFS rate at 2 years. The Eurocord Cooperative Group has reported their preliminary experience with CB transplantation in 50 MDS patients. With a median follow-up of 21 months, 2-year probability of DFS is 29%. These preliminary results suggest that CB transplantation could constitute an alternative approach to transplantations from MUD in MDS patients.
7. Conclusions Allogeneic HSCT is the treatment of choice for the majority of young patients with MDS who have a histocompatible donor (sibling or unrelated). For patients who lack an HLA-compatible sibling donor, the outcomes with autologous HSCT or chemotherapy appear comparable. These might be good alternatives for MDS patients with good-risk cytogenetic characteristics. Achievement of CR and the harvest of a sufficient number of autologous stem cells are prerequisites for autologous SCT.
Reference 1. Bennett JM, Catovsky D, Daniel MT, et al. Proposals for the classification of the myelodysplastic syndromes. Br J Haematol 1982; 51: 189-199. 2. Harris NL, Jaffe ES, Diebold J, et al. The World Health Organization classification of hematological malignancies report of the Clinical Advisory Committee Meeting, Airlie House, Virginia, November 1997. Mod Pathol 2000; 13: 193-207. 3. Greenberg P, Cox C, LeBeau MM, et al. International scoring system for evaluating prognosis in myelodysplastic syndromes. Blood 1997; 89: 2079-2088. 4. Kroger N, Bornhauser M, Ehninger G, et al. Allogeneic stem cell transplantation after a fludarabine/busulfan-based reduced-intensity conditioning in patients with myelodysplastic syndrome or secondary acute myeloid leukemia. Ann Hematol 2003; 82: 336-342. 5. Ho AY, Pagliuca A, Kenyon M, et al. Reduced-intensity allogeneic hematopoietic stem cell transplantation for myelodysplastic syndrome and acute myeloid leukemia with multilineage dysplasia using fludarabine, busulphan, and alemtuzumab (FBC) conditioning. Blood 2004; 104: 1616-1623. 6. Martino R, Iacobelli S, Brand R, et al. Retrospective comparison of reduced-intensity conditioning and conventional high-dose conditioning for allogeneic hematopoietic stem cell transplantation using HLA-identical sibling donors in myelodysplastic syndromes. Blood 2006; 108: 836-846. 7. de Witte TM, van BA, Hermans J, et al. Autologous bone marrow transplantation for patients with myelodysplastic syndrome (MDS) or acute myeloid leukemia following MDS. Chronic and Acute Leukemia Working Parties of the European Group for Blood and Marrow Transplantation. Blood 1997; 90: 3853-3857. 8. de Witte TM , Suciu S, Verhoef G, et al. Intensive chemotherapy followed by allogeneic or autologous stem cell transplantation for patients with myelodysplastic syndromes (MDSs) and acute myeloid leukemia following MDS. Blood 2001; 98: 2326-2331.
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9. de Witte TM, Hagemeijer A, Suciu S, et al. Allogeneic (Allo-) or autologous (Auto-) peripheral blood stem cell transplantation (SCT) randomized versus a second intensive consolidation course in patients with bad prognosis myelodysplastic syndromes and acute myelogenous leukemia following MDS of more than 6 months duration: The final analysis of a joint study (Criant) of the EORTC, EBMT, SAKK, HOVON, and GIMEMA Leukemia groups. Blood 2005; 106: #794. 10.Sierra J, Perez WS, Rozman C, et al. Bone marrow transplantation from HLA-identical siblings as treatment for myelodysplasia. Blood 2002; 100: 1997-2004. 11.Deeg HJ, Storer B, Slattery JT, et al. Conditioning with targeted busulfan and cyclophosphamide for hemopoietic stem cell transplantation from related and unrelated donors in patients with myelodysplastic syndrome. Blood 2002; 100: 1201-1207. 12.Nevill TJ, Fung HC, Shepherd JD, et al. Cytogenetic abnormalities in primary myelodysplastic syndrome are highly predictive of outcome after allogeneic bone marrow transplantation. Blood 1998; 92: 1910-1917. 13.Cutler CS, Lee SJ, Greenberg P, et al. A decision analysis of allogeneic bone marrow transplantation for the myelodysplastic syndromes: Delayed transplantation for low-risk myelodysplasia is associated with improved outcome. Blood 2004; 104: 579-585. 14.Castro-Malaspina H, Harris RE, Gajewski J, et al. Unrelated donor marrow transplantation for myelodysplastic syndromes: Outcome analysis in 510 transplants facilitated by the National Marrow Donor Program. Blood 2002; 99: 1943-1951. 15.Ooi J, Iseki T, Takahashi S, et al. Unrelated cord blood transplantation for adult patients with advanced myelodysplastic syndrome. Blood 2003; 101: 4711-4713.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. A male patient, 40 years age old, diagnosis: refractory anaemia without multilineage dysplasia, normal cytogenetics, platelet count 130 x 109/L, neutrophil count, 1.2 x 109/L. Transfusion need: two units every 4 weeks. What is the best treatment approach? a) Immediate allogeneic stem cell transplantation if a histocompatible sibling has been identified . . . . . . . . . . . . . . . . . . . b) Supportive care with transfusion and iron chelation . . . . . . . . . . . . . . c) Treatment with lenalinomide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Other treatment options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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2. Allogeneic stem cell transplantation with reduced intensity conditioning regimens is usually applied in: a) Patients >55 years . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Patients with advanced stages of myelodysplasia . . . . . . . . . . . . . . . . c) Both patient categories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Neither of these patient categories . . . . . . . . . . . . . . . . . . . . . . . . . 3. Autologous haematopoietic stem cell transplantation is a reasonable treatment option for patients: a) With refractory anaemia or refractory anaemia with ring sideroblasts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) In complete remission with high risk cytogenetics before treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) In complete remission with low risk cytogenetic characteristics before treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All 3 patient categories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. The optimal timing of allogeneic stem cell transplantation in a patient with refractory anaemia and excess of blasts (RAEB), 7% marrow blasts and a platelet count of 15 x 109/L is: a) Immediate allogeneic stem cell transplantation with histocompatible sibling donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Allogeneic stem cell transplantation after intensive remission induction therapy (AML-type) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Supportive care followed by allogeneic stem cell transplantation if there is disease progression. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Alternative options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. The outcome of matched unrelated donor transplantation is comparable with that of transplantation with a histocompatible sibling in: a) Patients <50 years . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Patients <60 years . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) All ages. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) None of these three . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 23
HSCT for chronic myeloid leukaemia in adults
D. Niederwieser
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CHAPTER 23 • CML in adults
1. Introduction CML is a rare disease, since only 1 to 2 patients with CML are diagnosed per 100,000 population each year. In many aspects, however, CML is a disease with considerable historic significance in haematology, and in medicine in general. The discovery of the Philadelphia (Ph) chromosome in more than 90% of affected individuals led to the molecular characterisation of the disorder. The t(9;22) chromosomal abnormality results in the creation of the BCR-ABL fusion gene and the production of a deregulated tyrosine kinase. This in turn resulted in the identification of small molecules designed to inhibit the kinase activity and subsequently highly effective targeted therapy.
2. Role of imatinib therapy Previously the only curative approach for patients with CML was allogeneic HSCT. For patients unsuitable for HSCT, interferon (IFN)-a with or without cytosine arabinoside (Ara-C) was considered the treatment of choice and was therefore selected as the standard arm in a phase III randomised study of the first tyrosine kinase inhibitor (TKI), imatinib (the IRIS study). Imatinib was superior to the combination of IFN+Ara-C in terms of haematological and cytogenetic responses, tolerability, and the likelihood of progression to accelerated phase (AP) or blast crisis (BC). After five years of follow-up only 7% of patients had progressed to acceleratedphase or blast crisis. Patients who had a complete cytogenetic response had a significantly lower risk of disease progression than did patients without a complete cytogenetic response. Grade 3 or 4 adverse events diminished over time, and there was no clinically significant change in the profile of adverse events (1). As a result imatinib has replaced IFN-a as first line therapy for CML (2). As the haematological community has gained experience with imatinib, it has become clear that the majority of patients will have long-term benefit from imatinib alone. However a significant minority of patients (approximately 15–20%) will fail to achieve complete cytogenetic responses and a further proportion (15–20%) will lose these excellent responses (3). It would seem reasonable that allogeneic HSCT would be second line therapy for individuals intolerant to or failing imatinib. However the mechanisms of resistance to imatinib are currently the subject of considerable research activity and at least one of these has been elucidated. It would seem that leukaemic cells bearing point mutations in the tyrosine kinase domain can be present at diagnosis and/or emerge during treatment due to genomic instability. These mutations interfere with imatinib binding rendering the drug less effective. Several second generation tyrosine kinase inhibitors have subsequently being developed that are capable of inhibiting the great majority of the mutated kinases, including
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dasatinib, nilotinib, bosutinib and Inno 406 (4). Phase I and II studies have confirmed the efficacy of these new TKIs in many patients with resistance or intolerance to imatinib, particularly if resistance occurs when the patients are still in the chronic phase (CP). The role of dasatinib and nilotinib in the management of CML has still to be defined and will be addressed in the next version of the recommendations of the European Leukaemia Net (ELN) and EBMT.
3. Role of HSCT Any decision regarding the advisability of allogeneic HSCT must now take into consideration not only the factors long recognised to have prognostic value, i.e. age, disease phase, duration of disease, nature of stem cell donor and the genders of recipient and donor, but also the likelihood of response to first (and second) generation of TKIs. Within the EBMT recommendations for transplantation (5), allogeneic HSCT from sibling and well-matched unrelated donors remains a standard of care for the chronic and accelerated phases of the disease. In the past allografting has not been recommended for patients with blast crisis. However the introduction of imatinib has resulted in patients being referred for transplant at later stages in their disease and it is likely that this negative recommendation might have to be reviewed. It is possible that allogeneic HSCT in combination with second-generation drugs may offer these individuals better outcome. Autologous transplantation seemed to offer valuable prolongation of survival for selected groups of patients and was under investigation in randomised phase III studies at the time that imatinib became readily available. These studies were then unable to recruit sufficient numbers of patients and were closed prematurely. Appropriately this approach remains developmental according to EBMT recommendations (5). 3.1. Autologous HSCT The frequencies of autologous HSCT for CML have decreased considerably during recent years and currently fewer than 50 transplants per year are being reported to EBMT. Most of these patients are in acceleration or blast crisis. The rationale for autografting is to delay progression of disease and restore susceptibility to imatinib and/or IFNa. In the past autologous transplant has involved the use of stem cell products heavily contaminated with leukaemic cells. The ability to achieve complete cytogenetic remission on imatinib has permitted the collection of stem cells with much reduced quantities of tumour and autologous transplant might find a role in disease management in the future. Stem cells harvested from patients in cytogenetic remission have been stored but few transplants have so far been performed.
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3.2. Syngeneic HSCT The results of syngeneic HSCT were evaluated recently by the EBMT. Excellent results were observed with an OS of 62% and a DFS of 32% at 20 years. 3.3. Allogeneic HSCT HSCT remains the only curative treatment for CML. Since the advent of the TKIs, transplant rates have understandably decreased during recent years. However the outcome of allogeneic HSCT has in general improved over the same period of time (6). Patients with EBMT risk assessment scores of 0-2 (Table 1) (7) can expect 5 year overall survivals in excess of 85% so transplant remains an acceptable choice as upfront treatment for young patients with high Sokal and/or Hasford scores who are, in general, less likely to do well with imatinib. However the real controversy about allogeneic HSCT is whether this approach or a second generation TKIs should be the management choice for patients with imatinib resistance. At present the balance is in favour of HSCT for younger patients with HLA-identical sibling transplants but for older patients and younger patients without a donor the situation is less clear. On the one hand, the results of unrelated donor HSCT have improved considerably, reaching survivals similar to those seen in HSCT with related donors. On the other hand, the second generation TKI have reasonable efficacy and relatively low toxicity. It is entirely reasonable to attempt a short (say 3 months)
Table 1: Risk factors for overall survival and transplant related mortality in CML Risk factor
Score
Donor type
HLA-identical sibling Unrelated
0 1
Disease stage
First chronic phase Acceleration Blast crisis
0 1 2
Age of recipient at HSCT
<20 years 20–40 years >40 years
0 1 2
Gender of donor and recipient
Other Male recipient/female donor
0 1
Time from diagnosis to HSCT
<12 months >12 months
0 1
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trial of the second generation TKIs and to recommend transplant for those patients who have not shown an improvement in their cytogenetic responses. The method of transplant at this point remains wherever possible myeloablative. In elderly patients and in patients with concomitant diseases, reduced intensity conditioning transplantation offers an acceptable alternative. Reduced intensity conditioning procedures, perhaps with maintenance with a TKI and/or pre-emptive use of donor lymphocyte infusions, reduce the transplant related toxicity with compromising long term disease free survival (8), but are not ready yet to substitute conventional stem cell transplantation in eligible patients. To date, pre-transplant imatinib treatment has not been shown to have a negative effect on outcome after allo-HSCT.
4. Monitoring response after allogeneic HSCT Early data relating to the incidence and risk of relapse after allografting were derived from qualitative RT-PCR assays for BCR-ABL transcripts. However minimal residual disease (MRD) can be detected by RT-PCR for years post-transplant without progression and suggests that low levels of BCR-ABL transcripts identified some years after transplant for CML may not always herald relapse. The predictive value of MRD detection is strengthened by BCR-ABL quantification. Several studies have demonstrated that the molecular burden of BCR-ABL mRNA, and the kinetics of increasing BCR-ABL, predict relapse. A recent study from the Hammersmith has further attempted to quantify the risk of relapse (9). This group has defined disease recurrence as requiring intervention if the BCR-ABL/ABL ratio exceeded 0.02% on three occasions or reached 0.05% on two occasions. 243 patients were by serial quantitative RT-PCR and classified into 4 groups: 1) 36 patients were “durably negative” or had a single low level positive result, 2) 51 patients had more than one positive result but never more than two consecutive positive results (“fluctuating positive”, low level (BCR-ABL/ABL ratio not satisfying definition of relapse) 3) 27 patients had persisting low levels of BCR-ABL transcripts but never more than three consecutive positive results (“persistently positive, low level”) and 4) 129 patients relapsed. In 107 of these 129 relapse was based initially only on molecular criteria; in 72 (67.3%) patients the leukaemia progressed to cytogenetic or haematologic relapse either prior to or during treatment with donor lymphocyte infusions. The study not only confirmed the value of their definition of relapse but also indicated that the probability of disease recurrence was 20% and 30% in groups 2 and 3 respectively. In view of the importance of the detection of residual disease at a time of low tumour burden when DLI are likely to be most effective, monitoring by RT-PCR is recommended at intervals no greater than three monthly for the first two to three 392
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years post transplant, six monthly until 5 years after grafting and annually thereafter. Any patient with a positive result should be monitored more frequently (approximately 4 weekly) until the course of their disease can be defined more precisely. It is essential for each laboratory to establish their own quantitative definition of relapse as it is not possible to extrapolate the results achieved in one institution with those obtained elsewhere. This situation is unsatisfactory but will hopefully benefit from a global attempt at harmonisation of RT-PCR standards for the detection of BCRABL transcripts (10).
5. Treatment of relapse post-transplant Donor lymphocyte infusions (DLI) have become the treatment of choice for patients who relapse after allogeneic SCT and durable molecular remissions are achieved in the majority of patients relapsing into chronic phase. In an EBMT study, survival after relapse was related to 5 factors: time from diagnosis to transplant, disease phase at transplant and at relapse, time from transplant to relapse and donor type (11). The effects of individual adverse risk factors were cumulative so that patients with 2 or more adverse features had a significantly reduced survival (35 vs. 65% at 5 years). Furthermore, DLI was less effective in patients who developed GvHD after transplant. However for patients transplanted in and relapsing in chronic phase the efficacy of escalating dose DLI was exceptionally high at >90% with a 5% procedural related mortality, rendering DLI the gold standard for the management of relapse in this group. GvHD and marrow aplasia remain the two most important complications of DLI but when an escalating dose schedule is used these problems are greatly reduced. This has been shown recently in a large retrospective study conducted by the Chronic Leukaemia Working Party (CLWP) on 344 patients at 51 EBMT centres (12). Patients starting DLI with a dose of 2 x 107 mononuclear cells/kg or less followed by escalating doses had less GvHD, less myelosuppression, the same response rate, better OS, better EFS, and less DLI-related mortality. Imatinib is now an alternative to donor lymphocyte infusions as it could potentially be used to achieve remission without the risk of GvHD. It could also be effective when DLI has failed, and could be used in combination with lower doses of DLI to maximise responses whilst minimising the risk of GvHD. A number of groups have now used imatinib in the management of patients relapsing after allogeneic transplantation. Most of these patients were treated for relapse into advanced phase disease, as DLI are of limited value in this situation. Other patients were treated for cytogenetic relapse or haematological relapse into chronic phase, often in the presence of on-going immunosuppression for GvHD and/or after failure of DLI.
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Recently, the Chronic Leukemia Working Party of the EBMT has reported a retrospective analysis of 128 patients treated with imatinib for relapse after allogeneic transplant (13). The overall haematological response rate was 84% (98% for patients in chronic phase (CP)). The complete cytogenetic response (CCR) was 58% for patients in CP, 48% for accelerated phase (AP) and 22% for patients in blast crisis (BC). Complete molecular responses were obtained in 25 patients (26%) of whom 21 were in CP or AP. With a median follow up of 9 months the estimated two-year survivals for CP, AP and BC patients were 100, 86 and 12% respectively. However Weisser et al. have recently compared the use of DLI or imatinib in 31 patients (14). 21 were treated for disease recurrence with DLI and 10 received imatinib because of lack of availability of the original donor. Molecular remissions were observed in 20 of the 21 patients (95%) who received DLI and 7 of 10 (70%) who were given imatinib. However 6 of the 10 patients treated with imatinib lost their best response whilst receiving the drug. Imatinib had been discontinued in 4 patients with confirmed molecular remission and disease recurred 2–4 months later in all but one of these individuals. 7 of the patients who failed imatinib subsequently received DLI and 6 achieved a molecular remission. Only 3 of the 20 patients who initially responded to imatinib experienced disease relapse. The authors concluded that imatinib, unlike DLI, cannot induce durable response in the majority of patients. Of course future practice will involve patients who have received allo-SCT largely as a consequence of having failed imatinib and perhaps also second line TKIs. This will mean that the efficacy of targeted therapy for disease recurrence will have to be re-established.
References 1. Druker BJ, Guilhot F, O’Brien SG, et al. Five-year follow-up of patients receiving imatinib for chronic myeloid leukemia. N Engl J Med 2006; 355: 2408-2417. 2. Baccarani M, Saglio G, Goldman J, et al. Evolving concepts in the management of chronic myeloid leukemia. Recommendations from an expert panel on behalf of the European Leukemia Net. Blood 2006; 108: 1809-1820. Epub 2006 May 18. 3. Apperley JF. Mechanisms of resistance to imatinib in chronic myeloid leukaemia. Lancet Oncol 2007; 8: 1018-1029. 4. Apperley JF. Management of resistance to imatinib in chronic myeloid leukaemia. Lancet Oncol 2007; 8: 1116-1128. 5. Ljungman P, Urbano-Ispizua A, Cavazzana-Calvo M, et al. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Definitions and current practice in Europe. Bone Marrow Transplant 2006; 37: 439-449. 6. Gratwohl A, Brand R, Apperley JF, et al. Allogeneic hematopoietic stem cell transplantation for chronic myeloid leukemia in Europe 2006: Transplant activity, long-term data and current results. Haematologica. 2006; 91: 513-521. Epub 2006 Mar 1. 394
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7. Gratwohl A, Hermans J, Goldman JM, et al. Risk assessment for patients with chronic myeloid leukaemia before allogeneic blood or marrow transplantation. Chronic Leukemia Working Party of the European Group for Blood and Marrow Transplantation. Lancet 1998; 352: 1087-1092. 8. Olavarria E, Siddique S, Griffiths MJ, et al. Post-transplantation imatinib as a strategy to postpone the requirement for immunotherapy in patients undergoing reduced-intensity allografts for chronic myeloid leukemia. Blood 2007; 110: 4614-4617. Epub 2007 Sep 19. 9. Kaeda J, O’Shea D, Szydlo RM, et al. Serial measurement of BCR-ABL transcripts in the peripheral blood after allogeneic stem cell transplant for chronic myeloid leukemia: An attempt to define patients who may not require further therapy. Blood 2006; 107: 41714176. Epub 2006 Jan 31 10.Hughes T, Deininger M, Hochhaus A, et al. Monitoring CML patients responding to treatment with tyrosine kinase inhibitors: Review and recommendations for harmonizing current methodology for detecting BCR-ABL transcripts and kinase domain mutations and for expressing results. Blood 2006; 108: 28-37. 11.Guglielmi C, Arcese W, Hermans J, et al. Risk assessment in patients with Ph+ chronic myelogenous leukemia at first relapse after allogeneic stem cell transplant: An EBMT retrospective analysis. The Chronic Leukemia Working Party of the European Group for Blood and Marrow Transplantation. Blood 2000; 95: 3328-3334. 12. Guglielmi C, Arcese W, Brand R, et al. Donor lymphocyte infusion for relapsed chronic myelogenous leukemia: Prognostic relevance of the initial cell dose. Blood 2002; 100: 397-405. 13.Olavarria E, Ottmann OG, Deininger M, et al. Response to imatinib in patients who relapse after allogeneic stem cell transplantation for chronic myeloid leukemia. Leukemia 2003; 17: 1707-1712. 14.Weisser M, Tischer J, Schnittger S, et al. A comparison of donor lymphocyte infusions or imatinib mesylate for patients with chronic myelogenous leukemia who have relapsed after allogeneic stem cell transplantation. Haematologica 2006; 91: 663-666.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. What is the only curative treatment for CML? a) IFN-a . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Imatinib . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) IFN-a and Ara-C . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Haematopoietic stem cell transplantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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2. Which factors are important for overall survival and transplant related mortality after SCT in CML? a) Donor type, disease stage, age of recipient at HSCT, gender of donor and recipient and time from diagnosis to HSCT . . . . . . . . . . b) Donor type, age of donor at HSCT, gender of donor and recipient and time from diagnosis to HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Imatinib pre-treatment, disease stage, age of recipient at HSCT, gender of donor and recipient and time from diagnosis to HSCT . . . . . . . . . . d) Age of donor and recipient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. If a patient with CML in chronic phase has a syngeneic donor, would you consider: a) A related non-syngeneic transplant, because of the unsatisfactory results of syngeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A syngeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) An unrelated HSCT with an unrelated donor having a 10/10 match . . . . . . d) A cord blood HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. At which time intervals and which response should be monitored in patients with CML after allogenic HSCT? a) Cytogenetics no greater than three monthly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) RT-PCR yearly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) RT-PCR no greater than three monthly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) RT-PCR no greater than three monthly and in case of positivity 4 weekly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5. What is today the treatment of choice (gold standard) for patients who relapse after allogeneic HSCT? a) Imatinib . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Donor lymphocyte infusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Dasatinib . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Nilotinib . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 24
HSCT for myeloproliferative disorders in adults
P. Guardiola
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CHAPTER 24 • MPD in adults
1. Introduction This Chapter will consider the indications for, and the results of, haematopoietic stem cell transplant (HSCT) in myeloproliferative disorders (MPD) other than chronic myeloid leukaemia (CML). These include myeloid metaplasia with myelofibrosis (MMM), myelofibrosis secondary to polycythemia vera (PV) and essential thrombocythopenia (ET).
2. Indications for transplantation Factors that have to be considered for choosing the optimal timing of transplantation in non CML-MPD, are the prognostic factors for disease outcome when nontransplant approaches are used, and those associated with a worse outcome following allogeneic transplantation. In non-transplanted patients with MMM, the main prognostic factors identified are: leukocytes <4 x 109/L or >30 x 109/L haemoglobin level <10 g/dL, abnormal karyotype, peripheral blood blasts, night sweats and weight loss. In low-risk patients with none of these prognostic factors, the expected median survival is longer than 5 years. Therefore, allogeneic transplantation should only be considered in patients with either intermediate- or high-risk prognostic scores (according to the classifications of Dupriez et al. or Cervantes et al. (1)) since their expected median survival is less than 2 years. In patients with PV or ET, occurrence of marrow fibrosis is the main criterion that should lead to consideration of allogeneic transplantation. In MMM, factors associated with a worse outcome following allogeneic transplantation are the same as those that predict poor outcome in non-transplanted patients, i.e. a low haemoglobin level, the need for red blood cell transfusions, a low platelet count, presence of severe marrow fibrosis and abnormal karyotype, and also older age at transplantation (in the setting of myeloablative preparative regimens only) (2–5). On the other hand patients without any of these poor-risk features can survive many years without any treatment and the risks of transplant would not be justified in early disease. Therefore the most appropriate time for HSCT is when one of these poor prognosis features appears, i.e. when life expectancy with classical treatments is usually around 2 years or less, Presence or new occurrence of any of these poor-prognostic factors, and especially inefficient erythropoiesis, should lead to consideration of allogeneic transplantation in patients who have a suitable donor and can tolerate the procedure.
3. Disease specific preparative regimens In the setting of myeloablative preparative regimens, retrospective studies favour non-total body irradiation-based regimens, since they are associated with a lower
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incidence of grade II-IV acute graft-versus host disease (GvHD) and less transplantrelated mortality (TRM) (3–5). The fact that patients with MMM are most often diagnosed at an age greater than 50 years, and that a high TRM has been reported with myeloablative regimens, suggests that reduced-intensity preparative regimens might be appropriate for a majority of MMM patients. However, data about this approach are limited and the follow-up is still too short to give “a firm recommendation” regarding the type of reduced-intensity preparative regimen to use.
4. Role and outcome of autologous transplantation The role of autologous transplantation in patients with MMM or myelofibrosis secondary to PV or ET is not clearly defined. The main objective of this non-curative strategy is to restore efficient haematopoiesis while reducing the tumour burden, i.e. splenomegaly and/or hepatomegaly, with an acceptable toxicity. Anderson et al. in 2001 reported the largest series of autologous transplants performed in patients with advanced MMM or myelofibrosis secondary to PV or ET (6). Since this publication, which included 27 patients, only limited data have been reported. From the Fred Hutchinson Cancer Research Center study, it appeared that G-CSF mobilisation of haematopoietic progenitor cells and leukapheresis were feasible and safe. Toxicity of the oral busulfan-only preparative regimen was acceptable (n=21 patients undergoing the transplant procedure), with a 3-year survival of 61% and three patients dying of non-relapse causes. Haematopoietic recovery was delayed as compared to autologous PBPC transplantation performed in lymphoma or myeloma patients (median time to neutrophil recovery 21 day; range: 10–96), five patients requiring subsequent back-up PBPC infusions. More than 50% of the patients experienced improvement in either erythroid (10 of 17 anaemic patients) or platelet (4 of 8 patients) response, and a reduction in symptomatic splenomegaly (n=7 of 10 patients). Despite these first encouraging results, development of reduced-intensity allogeneic transplantation and the risk of pancytopenia after autologous transplantation have made this approach less attractive.
5. Role and outcome of HLA-identical sibling transplantation with myeloablative preparative regimens Guardiola et al. reported the first large retrospective collaborative study of allogeneic transplantation using myeloablative regimens, initially including 55 patients with MMM, then 66 patients (2, 3). They showed that this strategy could lead to full engraftment and long-term complete histo-haematological remissions in a majority of patients (>15 years of follow-up), with complete disappearance 400
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of marrow fibrosis during the first year after transplantation in responding patients (2, 4). Median time to achieve neutrophil recovery was 20 days (range, 11–50). Pre-transplant splenectomy, absence of osteomyelosclerosis, and a high number of nucleated cells infused were associated with faster neutrophil recovery. In patients receiving T-cell replete grafts from HLA identical related donors, the 5-year overall and event-free survivals were 54 and 48%, respectively. For patients transplanted since 1977, the 1-year transplant-related mortality was 22%, and the 5-year incidence of treatment failure, 28%. Factors associated with an improved outcome were: absence of severe marrow fibrosis, haemoglobin level >10 g/dL, no red blood cell transfusion, normal karyotype, and recipient age <45 years at transplantation. In patients who had haemoglobin level >10 g/dL and who had never received red blood cell transfusion before transplantation, the 5-year overall survival rate was greater than 75%, whereas it was below 25% in other patients. Similar results were reported by Deeg et al. (4), who also observed that patients prepared with a non-TBI regimen - targeted busulfan + cyclophosphamide - had an improved outcome, whereas those with low pre-transplant platelet counts had a worse survival. In this later study, no difference in term of outcome was observed according to the type of donor. Of note, Daly et al., in a series of 25 patients, reported a high TRM, i.e., 48%, which could be explained by the fact that 23 of these patients were prepared with a combination of total body irradiation and cyclophosphamide (7). More recently, Kerbauy et al., reported the Seattle experience with myeloablative regimens (n=95) or a fludarabine plus 2 Gy total body irradiation regimen (n=9), in patients with either MMM (n=62) or myelofibrosis secondary to PV or ET (n=30) transplanted with HLA identical sibling (n=52), HLA matched unrelated donors (n=36) or other donor type (n=16) (5). Using a multivariable regression model, they found that use of targeted-busulfan plus cyclophosphamide as preparative regimen, high platelet count at transplantation for myelofibrosis secondary to PV or ET, younger patient age, and decreased comorbidity score were significantly associated with an improved survival. For instance, patients transplanted with targeted-busulfan plus cyclophosphamide had a 7-year survival of 68%.
6. Role and outcome of HLA matched unrelated transplantation with myeloablative preparative regimens Because of the limited number of patients transplanted all over the world for non CML-MPD, reported studies so far have combined results from HLA identical siblings and alternative donors (2–5, 7). Deeg et al. reported a significantly increased risk of failure of sustained engraftment in patients transplanted with alternative donors
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(n=25 of 56 patients), 14 being transplanted with HLA matched unrelated donors (4). Of note, all these patients received a bone marrow graft rather than PB, which was also identified as a risk factor for engraftment failure. However, the outcome of patients transplanted with alternative donors was not significantly different from that of patients receiving grafts from HLA matched related donors; acute GvHD incidence and severity as well as overall mortality rates being similar for HLA-identical related and alternative donor transplants. Similar results regarding overall survival were observed in a subsequent study from the Seattle group (5).
7. Role and outcome of allogeneic transplantation with reduced-intensity preparative regimens Rondelli et al. reported a retrospective series of 21 MMM patients transplanted with reduced-intensity preparative regimens (intermediate risk, n=13; high-risk, n=8) (8). Median age was 54 years, with a range of 27 to 68 years. Eighteen of 21 received PBPC grafts from HLA matched related donors. Various preparative regimens were used. Full engraftment was observed in all but one case. Grade II to IV acute GvHD was observed in seven cases, and extensive chronic GvHD in about 50% of evaluable patients. With a median follow-up of 31 months, 17 patients were alive in remission. Encouraging results were also reported by Kroger et al. in a prospective trial including 21 patients, of whom 13 received PBPC grafts from unrelated donors (9). Full donor cell engraftment was observed in 20 cases. Grade II to IV acute GvHD occurred in about 50% of the cases, ands chronic GvHD in 55%. Transplant-related mortality was 16% by 1 year after transplantation. Complete histopathological remission was confirmed in 75% of the patients. With a median follow-up of 22 months, the 3-year overall and disease-free survivals were 84%. These data suggest that in older patients with MMM, reduced-intensity preparative regimens could result in prolonged survival with a low TRM. However, incidence of either grade II-IV acute GvHD or chronic extensive GvHD remains high, and the followup of these studies is still short to confirm that this strategy will lead to persisting complete disease remission. Patients to be transplanted with reduced-intensity preparative regimens should be included in prospective trials, so that it will become possible to properly analyse results of this strategy.
8. Post-transplant treatment and monitoring 8.1. Nature and role of cellular or chemotherapy post-transplantation Data from retrospective studies and short reports suggest that a graft-versusdisease effect could be induced in MMM, via either donor lymphocyte infusions (1) 402
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or the occurrence of GvHD, which has been associated with a reduced incidence of treatment failures (2, 3). 8.2. Nature and role of minimal residual disease monitoring after transplantation Using a real-time PCR approach, Kroger et al. have shown that quantification of the JAK2-V617F mutation, which occurs in about 50% of patients with myelofibrosis, was feasible after allogeneic transplantation, and that it could be used to monitor minimal residual disease in the setting of transplantation prepared with reducedintensity regimens (10). In all but one responder (n=17 of 22 patients), PCR negativity was achieved during the first six months following transplantation. This real-time PCR has also been used as a decision-making analysis tool to guide immunomodulation in the post-transplant period, especially regarding timing of donor lymphocyte infusion. In this setting, this highly sensitive PCR (10-4) was also useful to monitor minimal residual disease following donor lymphocyte infusions and assess their efficacy.
References 1. Cervantes F, Rovira M, Urbano-Ispizua A, et al. Complete remission of idiopathic myelofibrosis following donor lymphocyte infusion after failure of allogeneic transplantation: Demonstration of a graft-versus-myelofibrosis effect. Bone Marrow Transplant 2000; 26: 697-699. 2. Guardiola Ph, Anderson JE, Bandini G, et al. Allogeneic stem cell transplantation for agnogenic myeloid metaplasia: A European Group for Blood and Marrow Transplantation, Société Française de Greffe de Moelle, Gruppo Italiano per il Trapianto del Midollo Osseo, and Fred Hutchinson Cancer Research Center collaborative study. Blood 1999; 93: 28312838. 3. Guardiola Ph, Anderson JE, Gluckman E. Myelofibrosis with myeloid metaplasia. N Engl J Med 2000; 343: 659-660. 4. Deeg HJ, Gooley TA, Flowers MED, et al. Allogeneic hematopoietic stem cell transplantation for myelofibrosis. Blood 2003; 102: 3912-3918. 5. Kerbauy DMB, Gooley TA, Sale GE, et al. Hematopoietic cell transplantation as curative therapy for idiopathic myelofibrosis, advanced polycythemia vera, and essential thrombocythemia. Biol Blood Marrow Transplantation 2007; 13: 355-365. 6. Anderson JE, Tefferi A, Craig F, et al. Myeloablation and autologous peripheral blood stem cell rescue results in hematologic and clinical responses in patients with myeloid metaplasia with myelofibrosis. Blood 2001; 98: 586-593. 7. Daly A, Song K, Nevill T, et al. Stem cell transplantation for myelofibrosis: A report from two Canadian centers. Bone Marrow Transplantation. 2003; 32: 35-40. 8. Rondelli D, Barosi G, Bacigalupo A, et al. Allogeneic hematopoietic stem-cell transplantation with reduced-intensity conditioning in intermediate- or high-risk patients with myelofibrosis with myeloid metaplasia. Blood 2005; 105: 4115-4119.
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9. Kroger N, Zabelina T, Schieder H, et al. Pilot study of reduced-intensity conditioning followed by allogeneic stem cell transplantation from related and unrelated donors in patients with myelofibrosis. Br J Haematol 2005; 128: 690-697. 10.Kroger N, Badbaran A, Holler E, et al. Monitoring of the JAK2-V617F mutation by highly sensitive quantitative real-time PCR after allogeneic stem cell transplantation in patients with myelofibrosis. Blood 2007; 109: 1316-1321.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which one of the following criteria does not help in deciding the optimal timing for allogeneic transplantation in non CML-MPD? a) Haemoglobin level . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) White blood cell count . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Spleen size . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Abnormal karyotype . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which one of the following criteria helps in deciding the optimal timing for allogeneic transplantation in non CML-MPD? a) Presence of dacryocytes on blood smear . . . . . . . . . . . . . . . . . . . . . . b) Presence of erythroblasts in the peripheral blood . . . . . . . . . . . . . . . c) Haemoglobin level . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Gender of the patient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. In retrospective studies assessing the results of allo-HSCT in non-CML MDP patients, splenectomy performed before the transplantation procedure has been associated with which one of the following? a) Faster neutrophil recovery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Improved overall survival . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Reduced transplant-related mortality . . . . . . . . . . . . . . . . . . . . . . . . d) Decreased post-transplant incidence of disease relapses . . . . . . . . . . .
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4. In retrospective studies assessing the results of allo-HSCT in non-CML MPD patients, myelo-ablative conditioning regimens using total body irradiation have been associated with which one of the following? a) An improved overall survival. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A reduced risk of disease relapse after transplantation . . . . . . . . . . . . c) A decreased risk of grade II to IV acute graft-versus-host disease . . . . . . d) An increased risk of transplant-related mortality . . . . . . . . . . . . . . . . 5. Which is one of the following sentences is wrong? a) Marrow fibrosis is a reversible process that can disappear following allo-HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Detection of the JAK2 mutation by quantitative PCR can be used to monitor minimal residual disease in non-CML MPD disorders after allo-HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Spleen size before transplantation is correlated with the outcome of patients undergoing allo-HSCT . . . . . . . . . . . . . . . . . . . . d) A “graft-versus leukaemia” effect has been observed in non-CML MPD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 25
HSCT for chronic lymphocytic leukaemia in adults
P. Dreger
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CHAPTER 25 • CLL in adults
Efforts to develop curative treatment strategies for chronic lymphocytic leukaemia (CLL) have focussed on autologous and allogeneic HSCT during recent years.
1. Indications In CLL, allogeneic HSCT (allo-HSCT) is a possible treatment option for eligible patients who have poor-risk disease as defined by the EBMT CLL Transplant Consensus (Table 1) (1). Although controlled trials are lacking, available evidence strongly suggests that allo-HSCT is the only therapy with curative potential in CLL. In contrast to conventional treatment, it can provide long-term disease control even in patients with a very unfavourable biological and clinical risk profile. Thus, alloHSCT may be offered to suitable patients as a standard procedure if disease-specific prospective clinical trial protocols are not available (2). Table 1: Criteria for poor-risk disease according to the EBMT CLL Transplant Consensus (1) • • •
Non-response or early relapse (within 12 months) after purine analogue-containing therapy Relapse (within 24 months) after purine analogue combination therapy or treatment of similar efficacy (i.e. autologous stem cell transplantation) p53 deletion /mutation (del 17p13) requiring treatment
Autologous HSCT (auto-HSCT) is a clinical option for consolidation treatment of patients in first or second remission who are able to withstand high-dose therapy. In the absence of evidence for superiority over effective non-transplant regimens, however, auto-HSCT should be performed in the context of a clinical trial (2).
2. Conditioning regimens There is no doubt that the crucial therapeutic principle of allo-HSCT in CLL is graftversus-leukaemia (GvL) activity. Evidence for this derives from the observation that even in patients with poorest-risk disease long-term clinical remissions can be observed after allo-HSCT but not with any other treatment modality, and from the fact that - in contrast to auto-HSCT or other intensive therapies – the relapse incidence seems to decrease over time (Table 2). In addition, GvL effectiveness in CLL is indicated by a reduced relapse risk in the presence of chronic graft-versushost disease (GvHD) (3), an increased relapse risk associated with the use of T-cell depletion (TCD) (4), efficacy of DLI (4), and post allo-HSCT minimal residual disease (MRD) kinetics (see below). Altogether, there appears to be sound evidence that GvL activity represents the main contributor to durable disease control after allo-HSCT even in poor-risk CLL.
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Table 2: Prospective trials of T-replete RIC allo-HSCT in CLL (all phase II) Study
Houston (11)
Boston (12)
Seattle (10)
DKTSG (9)a
DCLLSG (1)
n Age (years)
39 57 (34–70)
46 53 (35–67)
44 b 20 c 56 (44–69)
30 50 (12–63)
44 53 (27–63)
Median number n.a. of pretreatment lines
5
4
3
4
Refractory at HSCT
28%
57%
45% 57%
46%
20%
Alternative donor b
18%
67%
100% 0%
57%
48%
Conditioning
FluCy-Rituximab
FluBu6.4
FluTBI2
FluBu8
FluCy
TRM
30%
17%
22%
20% (2y)
15% (5y)
7% (3y)
Relapse rate
n.a.
48% (2y)
34% 5% (2y)
30% (4y)
33% (3y)
Late relapses (>2 yrs)
0
0
0
0
4
2
EFS
44% (2y) d
34% (2y)
44%
75% (2y)
58% (4y)
62% (3y)
OS
64% (2y)
54% (2y)
56%
74% (2y)
69% (4y)
79% (3y)
Follow-up (mo) 27 (4–80)
20 (6–48)
24 (3–63)
44 (25–67)
29 (5–81)
3.5
EFS: event-free survival; FluBu6.4: fludarabine 120mg/m2 + busulfex 6.4mg/kg; Flubu8: fludarabine 150mg/m2 + busulfan 8mg/kg; FluCy: fludarabine 150mg/m2 + cyclophosphamide 2,5g/m2; FluTBI2: fludarabine 90mg/m2 + total body irradiation 2Gy; n.a.: not available; OS: overall survival; TRM: treatmentrelated mortality. a and unpublished data; b matched unrelated donors or mismatched related donors; c matched related donors; d current progression-free survival
Accordingly, long-term disease control can be achieved with a broad range of conditioning intensities. Current evidence is not sufficient to identify a generally superior conditioning regimen. Considering the quality of trials performed, it appears that the most convincing data supporting allo-HSCT in CLL comes from RIC studies rather than from trials with traditional myeloablative allo-HSCT. However, impaired disease control associated with RIC cannot be ruled out (3). Thus, according to the individual situation, the optimum choice of conditioning regimens may vary: Whereas in the presence of comorbidity and sensitive disease RIC appear to be more appropriate, high-intensity regimens might be preferable in younger patients with good performance status but poorly controlled disease (1). Prospective clinical trials should help to guide the choice of conditioning intensity in allo-HSCT for CLL. Evidence for clinical benefit of T-cell depletion in CLL is lacking. In the majority of published data on autologous stem cell transplants for CLL the
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CHAPTER 25 • CLL in adults
myeloablative regimen contained TBI. The rationale behind this is that CLL cells similar to other indolent lymphatic neoplasms - are sensitive to irradiation. In the absence of prospective comparisons, however, the best preparative regimen for autografting patients with CLL is still unknown.
3. Outcome and role of autologous transplants in CLL Auto-HSCT for CLL was pioneered by the Dana-Farber Cancer Center. Patients with advanced CLL underwent myeloablative therapy (TBI/CY) followed by reinfusion of autologous bone marrow (BM) purged with anti-B-cell monoclonal antibodies and complement. Recently updated results on 137 patients show that relapses continued to occur over 10 years of follow-up, translating into a progression-free survival of 30% at 6 years post transplant (4). Similarly, no plateau in the survival curve was evident in registry analyses, such as those performed by the EBMT (5). Moreover, two large uncontrolled trials on auto-HSCT as part of first-line therapy of high-risk CLL (MRC pilot trial and CLL3 study of the German CLL Study Group (GCLLSG)) were characterised by continuous relapses which could occur even after 5 years of follow-up (5, 6). Finally, failure to achieve durable MRD negativity after auto-HSCT implies that complete disease eradication is not possible by this intensive approach in the vast majority of patients (7, 8). Similar to other diseases, such as multiple myeloma, auto-HSCT could confer a substantial therapeutic benefit in CLL even without being curative. In the MRC and GCLLSG prospective trials, median progression-free survival (PFS) was rather long with 54 and 59 months from study entry, respectively (5, 6). However, similar disease control seems to be possible with modern purine analogue-based combinations. Therefore, a reliable evaluation of the impact of HSCT on the prognosis of CLL requires prospective randomised studies comparing autografting with conventional treatment. Such a trial has been performed as a European intergroup effort coordinated by the EBMT. Patients in first or second remission after conventional chemotherapy for symptomatic CLL were randomised to receive a consolidating auto-HSCT or just observation. This trial has recently finished accrual after randomisation of more than 220 patients, but final results will not be available before 2009. Although there is no evidence that the incidence of treatment-related myelodysplastic syndromes and acute myeloblastic leukaemia (t-MDS/AML) after auto-HSCT for CLL exceeds the range reported previously for B-cell lymphoma, and also fludarabinealkylator combinations are associated with an increased risk of t-MDS/AML, this serious complication has to be taken into account when weighing the benefits and risks of auto-HSCT versus alternative modalities.
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4. Outcome and role of HLA-identical sibling transplants in CLL The risks of allo-HSCT in patients with CLL are mostly the general risks of allogeneic HSCT and are basically due to GvHD. Toxicity and mortality seem to be strongly influenced by the type of conditioning regimen employed (3). As pointed out previously, long-term disease control due to a low rate of late recurrences has been observed in all published series (excluding those employing in vivo or ex vivo T-cell depletion) irrespective of donor source and conditioning regimens used. Accordingly, a considerable proportion of patients survive leukaemia-free after allo-HSCT, as illustrated by 5-year EFS and OS rates ranging from 30 to 70% in the prospective RIC T-replete studies published to date (Table 2). In summary, cure seems to be possible in one to two thirds of patients undergoing allo-HSCT for poor-risk CLL.
5. Outcome and role of unrelated and alternative transplants in CLL In prospective studies including both matched unrelated donors (MUD) and sibling donors significant outcome differences did not become evident (Table 2). Therefore MUD allo-HSCT is regarded as standard treatment similar to sibling transplants in poor-risk CLL (1, 2). Transplants from mismatched donors should be restricted to clinical trials. Due to the rarity of the disease and the high average age, in CLL experience with haploidentical transplants and cord blood transplants is very sparse, and it is unlikely that disease-specific evidence for benefit of these procedures can ever be obtained.
6. Post-transplant minimal residual disease monitoring and immune intervention in CLL In CLL, sensitive (i.e. 1 cell in 10,000 or below) MRD quantification can be obtained by PCR- or flow cytometry-based assays and has strong prognostic impact after both auto- and allo-HSCT. The generally delayed decline of the MRD level and its close correlation with immune-relevant events strongly supports the assumption that GvL activity is the crucial contributor to tumour control in allo-HSCT. GvL-induced MRD negativity after allo-HSCT is sustained in the vast majority of cases and highly predictive of freedom from relapse, whereas in auto-HSCT MRD negativity is achieved only infrequently and is generally short-lived (7, 8). Furthermore, in CLL quantitative MRD monitoring seems to be a valid instrument for sensitive guidance of immune interventions directed at disease eradication after allo-HSCT. In contrast to DLI upon clinical relapse which often shows only limited benefit in CLL (9, 10), MRD-triggered pre-emptive DLI can be highly effective (7). However, the best approach to post-transplant immunotherapy (including monoclonal antibodies (11)) in CLL needs further study. 410
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7. Summary and perspectives To date, there is only limited hope that autotransplantation can cure CLL. Nevertheless, the results of prospective trials suggest that auto-HSCT is capable of exerting profound disease control, especially if employed early. However, until the results of the randomised EBMT Intergroup trial are available, auto-HSCT in CLL has to be considered as an experimental procedure which should not be performed outside of a clinical trial protocol. The combination of auto-HSCT with alternative innovative approaches, such as alemtuzumab in vivo purging or rituximab-purine analogue combinations, might open new perspectives for long-term and eventually durable disease control. Allo-HSCT from matched related or unrelated donors can be highly effective in otherwise resistant CLL. Therefore it is regarded as a standard treatment option for eligible patients who fulfil accepted criteria for poor-risk disease. Due to the absence of controlled trials in defined disease settings, however, it is unclear what the real impact of allo-HSCT in the treatment of CLL might be, and whether it can change the natural history of poor-risk CLL. The German CLL Study Group is currently conducting a randomised phase-III trial addressing these issues as well as optimum timing of transplantation.
References 1. Dreger P, Corradini P, Kimby E, et al. Indications for allogeneic stem cell transplantation in chronic lymphocytic leukaemia: The EBMT transplant consensus. Leukaemia 2007; 21: 12-17. 2. Ljungman P, Urbano-Ispizua A, Cavazzana-Calvo M, et al. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Definitions and current practice in Europe. Bone Marrow Transplant 2006; 37: 439-449. 3. Dreger P, Brand R, Milligan D, et al. Reduced-intensity conditioning lowers treatmentrelated mortality of allogeneic stem cell transplantation for chronic lymphocytic leukaemia: A population-matched analysis. Leukaemia 2005; 19: 1029-1033. 4. Gribben JG, Zahrieh D, Stephans K, et al. Autologous and allogeneic stem cell transplantation for poor risk chronic lymphocytic leukaemia. Blood 2005; 106: 4389-4396. 5. Dreger P, Brand R, Michallet M. Autologous stem cell transplantation for chronic lymphocytic leukaemia. Semin Hematol 2007; 44: 246-251. 6. Milligan DW, Fernandes S, Dasgupta R, et al. Autografting for younger patients with chronic lymphocytic leukaemia is safe and achieves a high percentage of molecular responses. Results of the MRC Pilot Study. Blood 2005; 105: 397-404. 7. Ritgen M, Stilgenbauer S, von Neuhoff N, et al. Graft-versus-leukaemia activity may overcome therapeutic resistance of chronic lymphocytic leukaemia with unmutated immunoglobulin variable heavy chain gene status: Implications of minimal residual disease measurement with quantitative PCR. Blood 2004; 104: 2600-2602. 8. Moreno C, Villamor N, Esteve J, et al. Clinical significance of minimal residual disease,
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as assessed by different techniques, after stem cell transplantation for chronic lymphocytic leukaemia. Blood 2006; 107: 4563-4569. 9. Schetelig J, Thiede C, Bornhauser M, et al. Evidence of a graft-versus-leukaemia effect in chronic lymphocytic leukaemia after reduced-intensity conditioning and allogeneic stemcell transplantation: The Cooperative German Transplant Study Group. J Clin Oncol 2003; 21: 2747-2753. 10.Sorror ML, Maris MB, Sandmaier BM, et al. Hematopoietic cell transplantation after nonmyeloablative conditioning for advanced chronic lymphocytic leukaemia. J Clin Oncol 2005; 23: 3819-3829. 11. Khouri IF, Saliba RM, Admirand J, et al. Graft-versus-leukaemia effect after nonmyeloablative haematopoietic transplantation can overcome the unfavourable expression of ZAP-70 in refractory chronic lymphocytic leukaemia. Br J Haematol 2007; 137: 355-363. 12.Brown JR, Kim HT, Li S, et al. Predictors of Improved Progression-Free Survival After Nonmyeloablative Allogeneic Stem Cell Transplantation for Advanced Chronic Lymphocytic Leukaemia. Biol Blood Marrow Transplant 2006; 12: 1056-1064.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Female patient, 53 years old, first diagnosis of CLL after known “elevated white blood count” over years, WBC 120 x 109/L with no other laboratory abnormalities, no symptoms, no lymphadenopathy, stage Binet A. Which one of the following strategies should be recommended? a) Further diagnostic procedures, e.g. bone marrow biopsy . . . . . . . . . . . . . . . . . . . b) No immediate intervention, observation of course . . . . . . . . . . . . . . . . . . . . . . . . . c) Chlorambucil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Fludarabine-cyclophosphamide followed by auto-HSCT . . . . . . . . . . . . . . . . . . . . . 2. In which one of the following situations allo-HSCT is not worth being considered? a) Diagnosis of CLL with deletion 17p13 without symptoms in a 55-year old patient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Non-response of CLL to fludarabine in a 60-year old patient . . . . . . . . . . . . . . c) CLL progression 2 years after fludarabine-cyclophosphamide-rituximab in a 62-year old patient without siblings. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) CLL relapse in a 49-year old patient 18 months after auto-HSCT . . . . . . . . . .
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CHAPTER 25 • CLL in adults
3. Which one of the following situations might be an indication for auto-HSCT? a) Diagnosis of symptomatic CLL with deletion 17p13 in a 55-year old patient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Non-response of CLL to fludarabine in a 60-year old patien . . . . . . . . . . . . . . . c) CLL progression 2 years after fludarabine-cyclophosphamide-rituximab in a 62-year old patient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) First remission of ZAP70-positive CLL after 3 cycles of fludarabine-cyclophosphamide in a 49-year old patient within a clinical protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which one of the following statements is not correct? a) In CLL, the sensitivity of MRD assessment using 4-colour flow cytometry cannot be higher than 1 tumour cell in 10,000 normal cells . . b) MRD negativity is frequently achieved after auto-HSCT for CLL and indicates cure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) MRD negativity after allo-HSCT for CLL often occurs only after immunomodulating manoeuvres, such as withdrawal of systemic immunosuppression or DLI . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) MRD negativity after allo-HSCT for CLL occurring after immunomodulating manoeuvres is generally not durable . . . . . . . . . . . . . . . . . . 5. Female patient, 53 years old, refractory after salvage treatment with fludarabine, stage Binet C with 95% BM infiltration, night sweats, moderate lymphadenopathy. FISH karyotype del 11q22, del 17p13. Recommended strategy: a) Further diagnostic procedures, e.g. mutational status, ZAP70 . . . . . . . . . . . . . b) Salvage fludarabine-cyclophosphamide-rituximab . . . . . . . . . . . . . . . . . . . . . . . . . . c) Salvage alemtuzumab, followed by allo-HSCT only in case of response . . d) Salvage alemtuzumab, followed by allo-HSCT also in case of refractory disease. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 26
HSCT for multiple myeloma in adults
J.A. Pérez-Simón, J. San Miguel
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CHAPTER 26 • MM in adults
Multiple myeloma (MM) is the most common plasma cell disorder with an incidence of 4–5 new cases per 100.000 individuals/year. Unfortunately, MM remains incurable with conventional chemotherapy. Nevertheless the availability of new drugs, which target not only the PC but also the microenvironment, together with the well established use of high dose chemotherapy is changing the prognosis of these patients.
1. Indications for transplant High dose therapy followed by auto-HSCT is considered the standard of care for patients diagnosed with multiple myeloma. Accordingly, MM is currently the most common indication for auto-HSCT in North America and Europe. As far as allogeneic transplantation is concerned, it remains as the only curative therapeutic approach in MM patients. However, it is associated with a high mortality and morbidity (mainly due to GvHD). Accordingly, it should be used in carefully defined situations and, preferably, within the context of clinical trials.
2. Specific conditioning Melphalan 200 mg/m2 is considered the gold standard conditioning regimen prior to auto-HSCT. The IFM randomised trial (1) confirmed that patients receiving melphalan 200 displayed a better median overall survival as compared to patients treated with melphalan 140 mg/m2 in combination with TBI (65 vs. 45% survival at 45 months). Other studies using intensification of the doses or addition of alkylating agents have not demonstrated significant improvements either in terms of response rate nor in outcome. As far as allogeneic transplantation is concerned, there is a marked heterogeneity in the type of conditioning regimen used: varying from myeloablative conditioning (MC) regimens to a variety of reduced intensity non-myeloablative regimens (RIC). Moreover, within each modality of conditioning the dose and type of drugs or radiotherapy is highly variable.
3. Role and outcome of autologous transplant High dose therapy followed by auto-HSCT prolonged overall survival as compared to standard dose therapy (SDT) in prospective randomised trials conducted by the French (IFM) and English (MRC) groups (2, 3) and has provided evidence for >10 year survivorship at least in a subset of patients. Nevertheless, the US study (SWOG 9321), the French MAG91 study and the Spanish PETHEMA group, although they confirmed the benefit of auto-HSCT in terms of response rate and EFS, did not find superiority in terms of survival as compared to SDT (4, 5, 6). These discrepancies
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can be, at least in part, explained by: 1) differences in the studies’ design (the Spanish study randomised patients responding to initial therapy while, in the others, randomisation was performed up-front), 2) differences in the conditioning regimens and, particularly, 3) differences in the intensity and duration of the chemotherapy arm (the dose of alkylating agents and steroids were higher in the SWOG and Spanish trials, which may explain why overall survival for conventionally treated patients was longer in these two studies as compared to IFM and MRC trials). Finally, two recent meta-analyses did not provide evidence of an overall survival advantage for HDT as compared to SDT (7, 8). In spite of these discrepancies, HDT is currently considered as standard of care for younger patients with multiple myeloma, mainly based on the benefit on response rate and EFS. Nevertheless, the availability of highly efficient new drugs may challenge this statement. Thus, novel drugs combinations based on thalidomide, bortezomib or lenalidomide, have shown to be superior to VAD-like regimens as debulking agents prior to auto-HSCT, with response rates >80%, including up to 10–30% CR rates. The next question is whether or not auto-HSCT is able to up-grade the response obtained with novel agents. In six pilot studies based on bortezomib-induction regimens, it was observed that the CR rate was improved following auto-HSCT, which suggests that the two approaches (induction with novel agents and auto- HSCT) are complementary rather than alternative. Regarding tandem auto-HSCT, its use will decrease for two reasons: 1) according to IFM (9) and Italian (10) experience only patients achieving a very good partial response with the first transplant benefit from the second and 2) a similar benefit is obtained upon using thalidomide as consolidation/maintenance therapy (11). In contrast, second transplant at relapse may be increasingly used, providing that the duration of the response to first transplant has lasted for more than 2–3 years.
4. Role and outcome of HLA-identical sibling transplant Although HDT followed by autologous transplantation allows long-term survival, at least in a subset of patients, there is no clear plateau on the survival curve. Moreover, patients displaying poor prognostic features such as IgH translocations plus Rb or p53 deletions and advanced stage according to the International Staging System display poor prognosis after HDT. On the contrary, allogeneic transplantation remains the only curative therapeutic approach which may offer long-term disease free survival. Unfortunately, although TRM has been
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CHAPTER 26 • MM in adults
reduced in the last years, conventional allogeneic transplantation is still associated with a high TRM. In the SWOG 9321 randomised trial, patients with a suitable donor received allogeneic transplantation after conditioning with melphalan 140 mg/m2 plus TBI and the arm was closed due to a 1 year TRM of 53%. Interestingly, 7 year estimated progression free survival in this subset of patients was 39%, similar to that reported for patients receiving autologous transplant or SDT, and this was due to a low relapse rate among patients receiving allogeneic transplant. In order to decrease TRM different RIC regimens have been developed (allo-RIC). In a prospective randomised trial, the French group compared double auto-HSCT to auto-HSCT followed by allo-RIC among patients displaying poor prognostic features (high B2microglobulin and monosomy of chromosome 13). Unfortunately, there were no event free survivors at 5 years either after double auto-HSCT or after auto-HSCT followed by allo-RIC (12). By contrast, the Italian group, using a similar approach, has recently described an improvement in terms of overall survival among patients receiving auto-HSCT followed by allo-RIC as compared to double auto-HSCT (13). In addition to differences in patient characteristics, the different GvHD prophylaxis and conditioning regimens used could explain these differences. Currently available data suggest that the use of RIC rather than myeloablative conditioning regimens may decrease TRM, but no prospective comparison is so far available. In a retrospective study from the EBMT comparing MC vs. allo-RIC, TRM was 37 vs. 24% and cumulative incidence of disease progression were 27 vs. 54% for MC vs. allo-RIC, respectively, which resulted in similar overall survival (51 vs. 38%). In high risk MM patients, including those displaying poor cytogenetic features (t(4;14), t(14;16) and t(14;20), Chr 13 deletion by conventional cytogenetics, p53 deletion, complex karyotype or hypodiploidy) or those with progressive disease during induction therapy, the use of novel agents as induction therapy followed by a tandem transplant, auto-HSCT and allo-RIC, represents an attractive approach, although these type of studies should be conducted within well controlled clinical trials. Regarding rescue therapy, in a series of 54 patients, 14 patients obtained complete remission or partial response out of 19 patients with refractory disease undergoing auto-HSCT followed by RIC allogeneic transplant (14). Unfortunately, a significant proportion of these patients finally relapse, leading to a poor event free survival, especially among patients who had relapsed after a prior autograft or with active disease at the time of allogeneic transplantation (15) (Table 1).
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Table 1: Results of randomised trials Trials (Ref.)
EFS / PFS
OS
p
Standard dose chemotherapy (SDT) vs. autologous HSCT IFM(2) SDT Auto-HSCT
18 months (10% 5 years) 27 months (28% 5 years)
37 months (12% 5 years) not reached (52% 5 years)
S
MRC(3) SDT* Auto-HSCT *
19 months 31 months
42 months 54 months
SWOG(4) SDT* Auto-HSCT *
14% 7 years 17% 7 years
39% 7 years 38% 7 years
PETHEMA(5) SDT* Auto-HSCT *
33 months 42 months
66 months 61 months
MAGG(6) SDT* Auto-HSCT *
19 months 25 months
47 months 47 months
S
NS
NS
NS
Single vs. double autologous HSCT IFM(9) Single* Double*
25 months (10% 7 years) 30 months (20% 7 years)
48 months (21% 7 years) 58 months (42% 7 years)
S
Cavo(10) Single Double
23 months 35 months
46 months 43 months
NS
Autologous vs. allogeneic HSCT IFM(12) Auto-HSCT (double) 30 months (0% 5 years) Auto-HSCT plus RIC-allo 25 months (0% 5 years)
41 months (44% 5 years) 35 months (33% 5 years)
NS
Bruno(13) Auto-HSCT (double)* 29 months Auto-HSCT plus RIC-allo * 35 months
54 months 80 months
SWOG(4) Auto-HSCT Allo-HSCT (MC)
38% 7 years 39% 7 years
S
NS 17% 7 years 22% 7 years
(p) differences for OS: (NS) non significant; (S) significant differences. (*) results expressed as median OS: overall survival; EFS: event-free survival; PFS: progression-free survival; MC: myeloablative conditioning
5. Role and outcome of matched unrelated donor (MUD) allogeneic transplant In the unrelated donor setting, the use of fludarabine and melphalan as conditioning regimen plus ATG as graft-versus-host disease prophylaxis was associated with a 90% 418
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CHAPTER 26 • MM in adults
response rate (40% complete and 50% partial) in a series of 21 MM patients. TRM was 26% at 1 year and 2 year overall and progression free survival were 74% and 53% (16). In another series of 17 patients, receiving MUD after fludarabine plus 2 Gy TBI, including 71% with chemotherapy resistant disease, 42% achieved complete remission and 17% partial response after transplant. Outcome was significantly better among those receiving tandem auto-HSCT followed by allo-RIC as compared to those who directly proceeded to allogeneic transplantation (51 vs. 11% progression free survival, respectively) (17).
6. Other sources: role and outcome of haploidentical transplant and cord blood transplant Data using alternative sources of progenitor cells in MM patients are too limited to draw any firm conclusion.
7. Nature and role of any additional cellular or chemotherapy posttransplant Donor lymphocyte infusions (DLI) given for relapsed myeloma following allogeneic transplantation induce responses in 30–50% of patients, the most common approach being the use of escalating dose at a usual starting dose of 1 x 107 CD3/kg (106 in the unrelated setting). In a recent multicentre analysis, the most important prognostic factors for response to DLI after RIC were the development of acute and chronic GvHD. Interestingly, the combination of DLI with thalidomide or bortezomib may improve the response rate and contribute to modulate the immune response, although further studies are required to confirm these data (18).
8. Nature and role of minimal residual disease monitoring after transplant A high relapse rate has been reported among MM patients receiving transplantation even in the allogeneic setting. For this reason, MRD monitoring might allow individualised treatment strategies. In this regard, PCR may contribute to identify patients at high risk of relapse; thus, in a series of MM patients undergoing transplantation, among 16 PCR negative patients no relapses were observed as compared to 100% among 13 patients with positive PCR (19). Unfortunately, a high proportion of patients develop extramedullary relapses without bone marrow involvement (20) indicating that, although the disease may be under control in the bone marrow milieu, extramedullary spread may occur. For this reason, MRD monitoring in bone marrow may not allow an early identification of all patients at risk of relapse and other tools, such as PET/MRI should be considered for a better follow-up of these patients.
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References 1. Moureau P, Facon T, Attal M, et al. IFM. Comparison of 200 mg/m2 melphalan and 8 Gy total body irradiation plus 140 mg/m2 melphalan as conditioning regimens for peripheral blood stem cell transplantation in patients with newly diagnosed multiple myeloma: Final analysis of the IFM 9502 randomized trial. Blood 2002; 99: 731-735. 2. Attal M, Harousseau JL, Stoppa AM, et al. A prospective randomised trial of autologous bone marrow transplantation and chemotherapy in multiple myeloma. Intergroupe Francais du Myelome. N Eng J Med 1966; 335: 91-97. 3. Child JA, Morgan GJ, Davies FE, et al. Medical Research Council adult Leukemia Working Party. High dose chemotherapy with hematopoietic stem cell rescue for multiple myeloma. N Eng J Med 2003; 348: 1875-1883. 4. Barlogie B, Kyle RA, Anderson KC, et al. Standard chemotherapy compared with high dose chemoradiotherapy for multiple myeloma: Final results of a phase III US intergroup trail S9321. J Clin Oncol 2006; 24: 929-936. 5. Blade J, Rosignol L, Sureda A, et al. PETHEMA. High dose therapy intensification compared with continued standard chemotherapy in multiple myeloma patients responding to the initial chemotherapy: Long term results from a prospective randomised trial from the Spanish cooperative group PETHEMA. Blood 2005; 106: 3755-3759. 6. Fermand JP, Katsahian S, Divine M, et al. Group Myeloma Autogreffe. High dfose therapy and autologous blood stem cell transplantation compared with conventional treatment in myeloma patients aged 55 to 65: Long term results of a randomised control trial from the MAG. J Clin Oncol 2005; 23: 9227-9233. 7. Levy V, Katsehian S, Fermand JP, et al. A meta-analysis on data from 575 patients with multiple myeloma randomly assigned to either high-dose therapy or conventional therapy. Medicine (Baltimore) 2005; 84: 250-260. 8. Koreth J, Cutler CS, Djulbegovic B, et al. High dose therapy with single autologous transplantation versus chemotherapy for newly diagnosed multiple myeloma. A systematic review and meta-analysis of randomised controlled trials. Biol Blood Marrow Transplant 2007; 13: 183-196. 9. Attal M, Harousseau JL, Facon T, et al. IFM. Single versus double autologous stem cell transplantation for multiple myeloma. N Eng J Med 2003; 349: 2495-2502. 10.Cavo M, Tosi P, Zamagni E, et al. Prospective, randomized study of single compared with double autologous stem-cell transplantation for multiple myeloma: Bologna 96 clinical study. J. Clin Oncol 2007; 25: 2434-2441. 11.Attal M, Harouseau JL, Leyvraz S, et al. Maintenance therapy with thalidomide improves survival in patients with multiple myeloma. Blood 2006; 108: 3289-3294. 12.Garban F, Attal M, Michallet M, et al. Prospective comparison of autologous stem cell transplantation followed by dose-reduced allograft (IFM99-03 trial) with tandem autologous stem cell transplantation (IFM99-04 trial) in high-risk de novo multiple myeloma. Blood 2006; 107: 3474-3480. 13.Bruno B, Rotta, M, Patriarca F, et al. A Comparison of Allografting with Autografting for Newly Diagnosed Myeloma. N Engl J Med 2007; 356: 1110-1120. 14.Maloney D, Molina A, Sahebi F, et al. Allografting with nonmyeloablative conditioning 420
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following cytoreductive autografts for the treatment of patients with multiple myeloma Blood 2003; 102: 3447-3454. 15.Kröger N, Perez-Simon J, Myint H, et al. Influence of timing allogeneic stem cell transplantation after dose-reduced melphalan/fludarabine conditioning in multiple myeloma. Biol Blood and Marrow Transplant 2004; 10: 698-708. 16.Kröger N, Sayer H, Schwerdtfeger R, et al. Unrelated stem cell transplantation in multiple myeloma after a reduced-intensity conditioning with pretransplantation antithymocyte globulin is highly effective with low transplantation-related mortality. Blood 2002; 100: 3919-3924. 17.Georges G, Maris M, Maloney D, et al. Nonmyeloablative unrelated donor hematopoietic cell transplantation to treat patients with poor-risk, relapsed, or refractory multiple myeloma. Biology of Blood and Marrow Transplantation 2007; 13: 423-432. 18.Van de Donk N, Kröger N, Hegenbart U, et al. Prognostic factors for donor lymphocyte infusions following non-myeloablative allogeneic stem cell transplantation in multiple myeloma. Bone Marrow Transplant 2006; 37: 1135-1141. 19.Corradini P, Cavo M, Lokhorst H, et al. Molecular remission after myeloablative allogeneic stem cell transplantation predicts a better relapse-free survival in patients with multiple myeloma. Blood. 2003; 102: 1927-1929. 20.Pérez-Simón J, Sureda A, Fernández-Avilés F, et al. Reduced intensity conditioning allogeneic transplantation is associated with a high incidence of extramedullary relapses in multiple myeloma patients. Leukemia 2006; 20: 542-545.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Regarding auto-HSCT, which of the following sentences is wrong? a) Patients receiving melphalan 200 mg/m2 display a better median overall survival as compared to patients treated with melphalan 140 mg/m2 in combination with TBI. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Two recent meta-analyses have not provided evidence of an overall survival advantage auto-HSCT as compared to standard dose therapy . . . . c) Novel drugs combinations based on thalidomide, bortezomib or lenalidomide, have shown to be superior to VAD-like regimens as debulking treatment prior to auto-HSCT, with response rates >80%, including up to 10–30% CR rates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Auto-HSCT does not improve the response rate obtained with bortezomib-based induction regimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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2. Regarding auto-HSCT, one of the following answers is incorrect: a) MM is currently the most common indication for auto-HSCT in North America and Europe . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) In the Spanish (PETHEMA) and American (SWOG) trials, the dose of alkylating agents and steroids used in the chemotherapy arm were higher than in the French (IFM) and UK (MRC) trials, which may explain why the survival was similar to that obtained with auto-HSCT . . . c) According to two randomised trials, patients achieving complete remission with the first auto-HSCT do benefit from the second transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Second transplant in relapsing patients offers no benefit to those patients in whom the duration of the response to first transplant has lasted less than 1 year . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Choose the correct answer: a) In the SWOG 9321 randomised trial, patients with a suitable donor received allogeneic transplantation after conditioning with melphalan 140 mg/m2 plus TBI and the arm was closed due to a 1 year TRM of 53% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Seven year estimated overall survival in this subset of patients was 39%, similar to that reported for patients receiving auto-HSCT or standard dose chemotherapy, and this was due to a low relapse rate among patients receiving allogeneic transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The IFM compared double auto-HSCT to auto-HSCT followed by RIC-allo among patients displaying poor prognostic features (high B2microglobulin and monosomy of chromosome 13). There were no event free survivors at 5 years in either arm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) In the IFM trial, the conditioning regimen among patients receiving allo-RIC consisted of fludarabine and melphalan . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Regarding allogeneic transplantation one of the following answers is incorrect: a) Median overall survival among patients receiving double auto-HSCT was 54 months as compared to 80 months among those receiving auto followed by allo-RIC in a multicentre prospective randomised Italian trial conducted by Bruno et al. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 422
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CHAPTER 26 • MM in adults
b) This study included only patients displaying poor cytogenetics . . . . . . . . . . . c) In a retrospective study from the EBMT comparing myeloablative conditioning vs. allo-RIC or RIC, the TRM was lower with RIC-allo but the cumulative incidence of disease progression was higher, which resulted in similar overall survival in both subgroups . . . . . . . . . . . . . . d) In that study, the use of Campath in a subset of patients receiving RIC-allo was associated with a significant increase in the risk of relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Regarding allogeneic transplantation which of the following answers is correct: a) Response rates ranging from 70 to 90% has been reported even among patients with refractory MM undergoing auto followed by RIC-allo transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) DLI given for relapsed myeloma following allogeneic transplantation induce responses in >80% of patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The most common approach is the use of escalating dose at a usual starting dose of 1 x 108 CD3/kg (107 in the unrelated setting) . . . . . . . . . . d) The combination of DLI plus thalidomide or bortezomib may improve the response rate as compared to DLI alone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 27
HSCT for primary amyloidosis in adults
J. Esteve
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CHAPTER 27 • Amyloidosis in adults
1. Introduction Primary systemic amyloidosis (AL) is a misfolding protein disease which leads to the extracellular deposition of abnormal protein fibrils in various tissues, including kidney, heart, liver, gastrointestinal tract, and peripheral nervous system, causing organ dysfunction. Amyloid fibrils in AL are constituted by a insoluble monoclonal light chain (LC) which aggregates forming b-pleated sheets. Diagnosis of AL is based on the recognition of amyloid substance in an appropriate tissue (subcutaneous fat tissue, rectum, bone marrow, involved organ), revealed by the characteristic staining pattern with Congo red dye, and further identification of amyloid fibril composition. Adequate identification of fibril precursor protein, a monoclonal LC, allows differentiation from other types of amyloidoses (familial type, secondary amyloidosis, dialysis-associated, senile). In most patients a monoclonal gammopathy is detected in serum and/or urine. Since AL is a clonal plasma cell disorder responsible for the synthesis of amyloidogenic LC, therapeutic agents used for AL therapy are similar to those used against multiple myeloma. Despite the usual low tumour burden characteristic of this disorder, AL is a poor-prognosis disease, with only a modest response pattern to standard chemotherapy, which cannot prevent progression of tissue damage. In contrast, intensive therapy with high-dose melphalan with auto-HSCT produces a high proportion of responses, followed by significant amelioration of organ damage in most responding patients. Unfortunately, this procedure is associated with an exceedingly high toxicity, reflecting the underlying organ damage secondary to amyloid deposits (1).
2. Indications Treatment with high-dose melphalan followed by peripheral blood stem cell rescue (auto-HSCT) should be considered in younger patients, up to 65–70 years, diagnosed with systemic AL, with no limiting organ damage, i.e., in the absence of uncompensated cardiac failure, severe renal failure or marked increase in bilirubin, and suitable for such an intensive procedure.
3. Stem cell mobilisation and collection, and conditioning regimen Stem cell mobilisation and leukapheresis in patients with AL is associated with unusual morbidity and with some reports of fatal events, due to the impaired organ and cardiovascular reserve of these patients. Thus, a syndrome of hypoxia and hypotension has been described both during mobilisation with G-CSF and during the leukapheresis procedure itself, probably as a result of diverse causes such as a capillary leak syndrome triggered by G-CSF, platelet activation during SC collection, and the release of inflammatory cytokines. Therefore, use of reduced doses of G-
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CSF (5–6 mg/kg ever 12 hours) and careful monitoring during the leukapheresis procedure is highly encouraged, with admission if necessary to an Intensive Care Unit, in order to avoid or correct immediately any sudden volume imbalance (hypovolemia or fluid overload) that may arise during the SC collection process. Conditioning regimens in AL are based on high-dose melphalan. The usual melphalan dose is 200 mg/m2, although a “risk-adapted” approach, with dose reduction to 140 mg/m2, has been proposed in higher risk patients in order to decrease transplantrelated toxicity (2). Proposed criteria for defining high risk patients are age >60, increased creatinine level, performance status (PS) 2 or compensated cardiac failure. Reduced melphalan dose has been associated with a decreased response rate in some studies (2, 3), although this observation has not been confirmed in other studies (4).
4. Results: toxicity, response, and long-term outcome Auto-HSCT is associated with a remarkably high risk of morbidity and mortality in patients with AL, with a TRM ranging from 11 to 43% (Table 1). Cardiogenic shock, fatal arrhythmias, gastrointestinal bleeding, and infections are the most frequent complications involved in procedure-related deaths during this phase. Infrequent causes such spontaneous splenic rupture or DMSO-triggered cardiac arrest have also been reported in this setting. Furthermore, renal insufficiency develops frequently after auto-HSCT, occurring in approximately 20% of patients. As regards activity against AL, high-dose melphalan results in significant responses in 50–60% of cases, with complete responses in about one third of patients. CR is defined, in this setting, by a negative immunofixation and normal free Table 1: Summary of the outcome of patients with primary amyloidosis undergoing autotransplant according to largest series Source
No. of pts
Overall response TRM (%) rate (CR) %
Overall survival (%)
Boston (US) (3) CIBMTR (multicenter) (11)
205 107
NR (40) 32 (16)
13 (100-day) 18 (30-day) 27 (1-yr)
60 (3-yr) 66 (1-yr) 56 (3-yr)
UK (multicenter) (12)
92
64 (35)
23 (100-day)
50 (5-yr)
Mayo Clinic (7)
282
71 (33)
11 (100-day)
60 (5-yr)
French Intergroup (MAG-IFM) (9)
50
49 (30)
24 (100-day)
45 (3-yr)
CR: complete response; TRM: transplant-related mortality; NR: not reported; CIBMTR: Center for International Blood and Marrow Transplant Research; UK: United Kingdom; MAG-IFM: Myélome Autogreffe-Intergroupe Francophone du Myélome 426
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immunoglobulin light-chain (FLC). Interestingly, haematologic responses are followed by organ responses, i.e. improvement of involved organ function, in most cases. Although median time to achieve a response is between 3–4 months, responses can take several months, up to 2 years, in some patients. On the other hand, some relapses are observed during follow-up, although the overall incidence of relapses is relatively low, especially among patients who achieve CR after autoHSCT. Thus, the relapse incidence at 10-years among CR patients was 21% in the Boston series (3). These combined results translate into a long-term survival between 45–60%, according to larger series (Table 1). Guidelines for an adequate evaluation of potential candidates to an autotransplant, recommended clinical care following transplant and basic criteria for assessment of response after auto-HSCT are summarised in Table 2.
5. Prognostic factors Several factors have been related to transplant outcome in AL. Thus, cardiac involvement, as assessed by several methods (congestive heart failure, thickened intraventricular wall on ultrasonography), is invariably identified as an adverse prognostic factor. In this regard, measurement of cardiac troponins (cTnT, cTnI) and pro-brain natriuretic peptide (NT-proBNP) provides a refined surrogate marker of myocyte damage in AL and these cardiac biomarkers are strong predictors of survival after auto-HSCT, with a significant shortened life expectancy among patients with increased levels (5). Concurrent renal failure is also associated with shorter survival after transplant and, in general terms, auto-HSCT is contraindicated in patients with severe renal failure. Variables related to disease extent also predict outcome after transplant. Thus, involvement of more than two organs correlates with an unfavourable prognosis (6). More recently, baseline level of FLC has been identified as a prognostic factor in this setting, with an increased risk of TRM in patients with higher pre-transplant levels (7). Of note, the degree of response achieved after transplant showed a striking correlation with long-term outcome, with the most favourable outcome observed in patients who achieved CR (8). On the contrary, patients who fail to achieve a significant response show a poor outcome, with rapid disease progression.
6. Role of auto-HSCT in the management of the disease: comparison with other treatment approaches Long-term survival after auto-HSCT appears to be prolonged, especially in patients who achieve CR after transplant, and compares favourably with historic controls. This observation, however, should be interpreted with caution as it might reflect
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Table 2: Specific considerations regarding evaluation of auto-HSCT in patient with primary amyloidosis Diagnostic accuracy
1. Demonstration of amyloid deposition in an appropriate tissue (Congo Red staining) 2. Confirmation of primary origin AL: - Presence of monoclonal light chain in amyloid fibril - (or) Detection serum/urine monoclonal light-chain
Is the patient a candidate to an autotransplant?
1. Age up to 65–70 years 2. Adequate performance status (£2) 3. Absence of limiting organ damage: - No compensated cardiac failure - Severe renal failure - Markedly increased bilirubin
Assessment of risk factors
1. Assessment of cardiac involvement: - Determination of cardiac biomarkers (cTnT, cTnI, NT-proBNP) - Echocardiogram (interventricular wall thickness) 2. Assessment of amyloid deposition: - Number of involved organs (renal, cardiac, hepatic, gastrointestinal, peripheral & autonomic neuropathy) - Serum FLC measurement
Recommended care during procedure
1. Monitor stem cell mobilisation and collection. Consider admission to an Intensive Care Unit 2. Adequate risk-adapted conditioning: - Standard dose: melphalan 200 mg/m2 - Reduced dose (if concurring risk factors): melphalan 140 mg/m2 3. Careful post-transplant management: - Close monitoring of cardio-vascular function - Prevention of mucosal & gastrointestinal bleeding: specific platelet transfusion policy (maintain > 50 x 109/L)
Adequate assessment of post-transplant response
1. Haematologic response: - CR: negative serum & urine immunofixation; normal k/l ratio & absolute value of FLC - PR: 50% reduction of serum M component, urine light chain & FLC 2. Organ response. Re-assessment of specific parameters of pre-transplant involved organ: - Renal (24-hour urinary protein, creatinine level) - Heart (functional class, septal thickness, ejection fraction) - Liver (alkaline phosphatase, liver size) - Nerve (nerve conduction)
merely a selection bias, since candidates for auto-HSCT constitute a selected population with better prognosis. Therefore, the precise impact of auto-HSCT in the management of the disease remains controversial; there are only a few studies 428
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CHAPTER 27 • Amyloidosis in adults
comparing auto-HSCT with standard dose chemotherapy and the results of these studies are conflicting. Thus, on the one hand, a case control-study conducted by the Mayo Clinic, which compared the outcome of 63 patients who had received an auto-HSCT with that of 63 patients treated with standard therapy, showed a better outcome in the intensively treated subgroup, with a 4-year survival of 71 vs. 41% (9). This study, despite matching for main variables, has some limitations derived from the retrospective nature of the study and therapy administered in the control group, which mostly consisted of melphalan and prednisone. In contrast, recently published results of the only randomised trial comparing high-dose melphalan with standard dose melphalan and dexamethasone showed a better outcome in patients not randomised to high-dose melphalan, when analysed on an intentionto-treat basis (10). This study, however, also has some limitations, such as the relatively small number of patients included, 50 per arm, and the low compliance with the assigned therapy, since only two-thirds of patients randomised to intensive therapy finally underwent auto-HSCT. When the analysis was restricted to patients who actually received the assigned therapeutic option, no major differences were observed between the arms. Therefore, the potential benefit of high-dose melphalan and the exact target population of AL patients remain to be clarified in further studies. Moreover, the potential improvement in the “control arm” (i.e., based on nonintensive therapy) with the introduction of newer agents such as immunomodulators (thalidomide, lenalidomide) or the proteasome inhibitor bortezomib, should be considered while assessing the therapeutic role of auto-HSCT in this disease.
7. Future perspective and conclusions Given the relevance of achieving a response after transplant, several approaches for increasing the proportion of responses after auto-HSCT have been proposed. Thus, the Boston group is conducting a trial of tandem transplants, with performance of a second transplant with melphalan at a dose of 140 mg/m2, in patients not achieving CR after first transplant. The administration of post-transplant maintenance therapy with newer agents is another potential strategy for improving control of the disease. Finally, an allogeneic procedure using reduced-intensity conditioning has been performed in a minority of patients, with the aim of exploiting a possible “graft-versus-amyloidosis” effect. In conclusion, auto-HSCT results in haematological responses and organ improvement in a significant proportion of patients with AL. Moreover, responding patients show a relatively prolonged survival. Nonetheless, this procedure is associated with a exceedingly high mortality, with a TRM of at least 10%, reflecting the fragile condition of patients due to multiorgan damage caused by amyloid deposition. Therefore, careful selection of patients for transplant is critical. In this regard, a
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more refined assessment of cardiac involvement or the extent of the disease, by means of measurement of cardiac biomarkers and quantification of serum FLC, might contribute to a more accurate evaluation of patients prior to auto-HSCT. Nonetheless, the long-term effectiveness of high-dose melphalan for the management of the disease is currently unclear, and comparison with standard-dose agents shows conflicting results. Finally, development of new strategies to intensify and/or prolong response after auto-HSCT might improve the outcome. In this regard, further prospective studies are required to elucidate the role of auto-HSCT in AL.
References 1. Comenzo RL. AL amyloidosis. In: Multiple Myeloma and Related Disorders. Gahrton G, Durie BGM, Samson DM (eds), London, UK: Arnold; 2004: 400-419. 2. Gertz MA, Lacy MQ, Dispenzieri A, et al. Risk-adjusted manipulation of melphalan dose before stem cell transplantation in patients with amyloidosis is associated with a lower response rate. Bone Marrow Transplant 2004; 34: 1025-1031. 3. Skinner M, Sanchorawala V, Seldin DC, et al. High-dose melphalan and autologous stem cell transplantation in patients with AL amyloidosis: An 8-year study. Ann Int Med 2004; 140: 85-93. 4. Perfetti V, Siena S, Palladini G, et al. Long-term results of a risk-adapted approach to melphalan conditioning in autologous peripheral blood stem cell transplantation for primary (AL) amyloidosis. Haematologica 2006; 91: 1635-1643. 5. Dispenzieri A, Gertz M, Kyle RA, et al. Prognostication of survival using cardiac troponins and N-terminal pro-brain natriuretic peptide in patients with primary systemic amyloidosis undergoing peripheral blood stem cell transplantation. Blood 2004; 104: 1881-1887. 6. Gertz MA, Lacy MQ, Dispenzieri A, et al. Stem cell transplantation for the management of primary systemic amyloidosis. Am J Med 2002; 113: 549-555. 7. Dispenzieri A, Lacy MQ, Kartzmann JA, et al. Absolute values of immunoglobulin free light chains are prognostic in patients with primary systemic amyloidosis undergoing peripheral blood stem cell transplantation. Blood 2006; 107: 3378-3383. 8. Gertz MA, Lacy MQ, Dispenzieri A, et al. Effect of hematologic response on outcome of patients undergoing transplantation for primary amyloidosis: Importance of achieving a complete response. Haematologica 2007; 92: 1415-1418. 9. Dispenzieri A, Kyle RA, Lacy MQ, et al. Superior survival in primary systemic amyloidosis patients undergoing peripheral blood stem cell transplantation: A case-control study. Blood 2004; 103: 3960-3963. 10.Jaccard A, Moreau P, Leblond V, et al. High-dose melphalan versus melphalan plus dexamethasone for AL amyloidosis. N Engl J Med 2007; 357: 1083-1093. 11.Vesole DH, Pérez WS, Akasheh M, et al. High-dose therapy and autologous hematopoietic stem cell transplantation for patients with primary systemic amyloidosis: A Center for International Blood and Marrow Transplant Research Study. Mayo Clin Proc 2006; 81: 880-888. 12.Goodman HJ, Gillmore JD, Lachmann HJ, et al. Outcome of autologous stem cell transplantation for AL amyloidosis in the UK. Br J Haematol 2006; 134: 417-425. 430
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Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Regarding diagnosis of primary AL, indicate the correct answer: a) Diagnosis of AL relies exclusively on Congo Red positivity . . . . . . . . . . . . . . . . b) A monoclonal light chain is rarely detected in serum . . . . . . . . . . . . . . . . . . . . . . c) Congo Red staining must be performed in the involved organ . . . . . . . . . . . . d) AL is generally a low burden monoclonal gammopathy, with a low level of plasma cell bone marrow involvement and M spike. . . . . . . . . . . . . . . .
2. All of the following statements about TRM in patients with AL undergoing autotransplantation are true, except one. Indicate the incorrect answer: a) Is exceedingly high, between 10–20%. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Is related to increased level of pro-brain natriuretic peptide (NT-proBNP) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Is correlated with the heavy chain immunoglobulin idiotype of associated monoclonal gammopathy (IgG, IgA, IgM) . . . . . . . . . . . . . . . . . . . . . . d) Correlates with the number of involved organs . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. Concerning response after autotransplant, which of the following is the correct answer? a) Haematologic response assessment is based on serum and urine immunofixation and serum free light chain measurement . . . . . . . . . . . . . . . . . b) Organ responses can be observed in many patients who do not achieve a significant haematologic response. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Long-term survival is similar in patients achieving a complete response and those who obtain a partial response . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Organ response are always observed in the early period post-transplant, i.e., no longer than 3 months . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. Which of the following complications can be observed in AL patients undergoing autotransplant? a) Frequent gastrointestinal bleeding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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b) DMSO-triggered cardiac arrest . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) G-CSF induced respiratory failure during mobilisation . . . . . . . . . . . . . . . . . . . . . . d) All the previous have been reported in this setting . . . . . . . . . . . . . . . . . . . . . . . . 5. Which of the following answers is not appropriate for describing outcome after autotransplant in AL patients? a) Long-term follow-up shows a significant proportion of long-term survivors, between 50–60%. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Relapse rate in patients achieving CR is relatively low, <25% at 5 years . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Prospective randomised studies comparing the outcome of patients undergoing autotransplant with standard therapy demonstrate a clear benefit in patients receiving intensive therapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Outcome of patients that do not achieve at least a partial response after transplant is poor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 28
HSCT for high-grade non-Hodgkin’s lymphoma in adults
A. Johnston, B. Coiffier
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CHAPTER 28 • High-grade NHL in adults
1. Introduction This review will discuss the indications, role and outcome of haematopoietic stem cell transplantation in patients with aggressive non-Hodgkin’s lymphoma (NHL). The specific disease entities which we will consider are diffuse large B-cell lymphoma (DLBCL) and peripheral T-cell lymphoma (PTCL). The outcome of patients with high-intermediate and high-risk aggressive NHL is unsatisfactory with standard treatment approaches. In DLBCL approximately half of patients with high-intermediate and high-risk disease are cured with immunochemotherapy, usually R-CHOP, but the results are worse for patients with PTCL. Thus these patients may be candidates for front-line high-dose therapy (HDT) and autologous stem cell transplantation (ASCT). In younger patients with relapsed or refractory chemosensitive aggressive lymphoma HDT can be curative in a significant subset. However, many areas of uncertainty remain, such as the effectiveness of these strategies in patients with DLBCL who receive new standard therapies containing rituximab and the role of allogeneic transplantation in aggressive NHL.
2. High-dose therapy and autologous stem cell transplantation in firstline therapy of aggressive non-Hodgkin’s lymphoma HDT has been used as part of front-line therapy in an effort to improve results in young patients with aggressive NHL. Comparison between the various trials is difficult due to the inclusion of disparate patient groups (in terms of definition of risk, remission status and histology) and different therapeutic strategies in the transplantation and standard therapy arms. Four randomised trials have demonstrated a benefit of this approach in terms of an improvement in event-free survival (EFS) or overall survival (OS) in patients less than 60 years with aggressive NHL and high-risk features (e.g. age-adjusted International Prognostic Index (aaIPI) score of 2 or 3). All these studies included DLBCL and PTCL and were carried out before the era of monoclonal antibodies. For example, the LNH 87-2 trial randomised 236 patients with aaIPI of 2 or 3 who achieved complete remission (CR) after induction therapy to HDT or sequential consolidation and found a 8-year OS rate of 64% in the HDT arm compared with 49% in the sequential chemotherapy arm (p=0.04). However, a number of other trials, including the GELA trial LNH 93-3, the study of the German High-Grade NHL Study Group and the EORTC study, have not demonstrated a benefit of this approach in front-line therapy. Many, but not all, of the negative trials have utilised an abbreviated induction regime in the transplant arm, suggesting they may not have had adequate dose-intensity before HDT. The majority of patients in the EORTC trial belonged to favourable IPI risk groups. A multicentre European
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study (MISTRAL study) which compared 6–8 cycles of CHOP-21 with sequential HDT found no benefit and increased toxicity in the sequential HDT arm. A recent meta-analysis of 15 randomised trials was not conclusive but suggested a benefit of front-line HDT in poor risk patients with aggressive lymphoma (1). Immunochemotherapy with rituximab is now considered the standard of care in DLBCL. No randomised comparison exists of HDT after immunochemotherapy with rituximab in DLBCL. The results of trials that evaluate the relevance of this approach in the rituximab era are eagerly awaited. The role of HDT as consolidation therapy for patients with PTCL in first CR has not been defined. The majority of studies in this area are small, retrospective and heterogeneous. Some are confounded by the inclusion of patients with ALK-positive anaplastic large cell lymphoma which has a favourable prognosis when treated with chemotherapy alone. In subgroup analyses of PTCL included in the two GELA studies that examined this approach, there did not appear to be a benefit of HDT/ASCT over sequential chemotherapy for these patients (2, 3). A number of other retrospective analyses suggest a benefit of HDT over conventional chemotherapy. In a prospective study currently ongoing, a significant minority of patients (28%) were not transplanted due to early progression before myeloablative therapy (4). Due to the very poor prognosis of these patients with conventional chemotherapy, further prospective studies of the potential benefit of early HDT are needed.
3. High-dose therapy and autologous stem cell transplant in relapsed and refractory aggressive non-Hodgkin’s lymphoma The PARMA study established HDT as the standard of care in patients with chemosensitive relapsed aggressive NHL. In this study, 109 patients who demonstrated responsiveness to salvage chemotherapy with DHAP were randomly assigned to receive four further cycles of DHAP or intensive chemotherapy. Survival at five years was superior in the transplantation arm compared to the conventional chemotherapy arm, 53 versus 32% (p=0.038). Subsequent analyses of this cohort have demonstrated that time to relapse, less than 12 months versus greater than 12 months, was the most important prognostic factor influencing OS after relapse. At 8 years 13% of patients who relapsed early were projected to be alive compared with 29% of those who relapsed late (p<0.00001). The second-line aaIPI as well as the quality of response to salvage chemotherapy (CR versus PR) have also been demonstrated to be important prognostic factors in patients with relapsed and refractory aggressive lymphoma. A number of studies have demonstrated a similar benefit of HDT in patients with primary refractory aggressive lymphoma compared to those with relapsed disease, provided they remain chemosensitive (5). By contrast, patients with chemotherapy 436
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resistance do very poorly with HDT with long-term EFS in the order of 10% or less and thus are not considered candidates for this therapy. The prognosis of transformed low-grade B-cell lymphomas is generally poor. Although the data on HDT in this setting is limited and retrospective, it appears that approximately one-third of patients remain disease-free at five years (6). Thus it should be considered in eligible patients with chemosensitive disease. Available evidence suggests that patients with chemosensitive relapsed and refractory PTCL have similar outcomes with HDT to patients with relapsed aggressive B-cell lymphoma, despite the fact that the T-cell phenotype is demonstrated to be an adverse prognostic factor for primary therapy (7). However this is unlikely to hold true with the addition of rituximab to salvage chemotherapy and transplant protocols for DLBCL. As discussed above, the quality of the response to salvage therapy is an important prognostic factor, thus attempts have been made to improve results of ASCT by increasing the rate of CR to salvage therapy. The benefit of the addition of rituximab to salvage chemotherapy in DLBCL has been confirmed in a recent randomised trial of the HOVON group which compared salvage treatment with DHAP to that with RDHAP and found a significant advantage in terms of failure-free and OS (8). It is currently unknown which salvage protocol is the most efficacious. The ongoing Collaborative Trial in Relapsed Aggressive Lymphoma (CORAL) trial compares R-DHAP and R-ICE as salvage regimens prior to HDT and aims to answer this question in patients with relapsed DLBCL. A second randomisation looks at the role of rituximab maintenance therapy in these patients. Other strategies to improve the outcome of these patients are currently being explored. In DLBCL the addition of rituximab to pre- and post-transplantation therapy appears to be feasible and a phase III Intergroup trial is currently evaluating this approach. The incorporation of radioimmuoconjugates targeting the CD20 antigen in DLBCL is another promising approach under evaluation.
4. Allogeneic stem cell transplantation and aggressive non-Hodgkin’s lymphoma The role of allogeneic transplantation in aggressive NHL is uncertain. Theoretical advantages over autologous stem cell transplantation include a pure stem cell source free of tumour contamination and the possibility of a graft-versus-lymphoma effect. High treatment-related mortality of conventional myeloablative allogeneic transplantation (33% for high-grade lymphoma in an EBMT registry study) has hindered further development of this modality (9). Reduced-intensity conditioning (RIC) approaches can reduce treatment-related mortality but rely on a graft-versus-
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lymphoma effect. The evidence for a significant graft-versus-lymphoma effect in DLBCL is lacking. There are few data available on the role of allogeneic stem cell transplantation in PTCL. A small prospective series of RIC allogeneic transplant in relapsed or refractory PTCL suggested the existence of a graft-versus-T-cell lymphoma effect with promising 3-year OS rates. Further studies of this approach in this poorprognosis group are warranted. In certain subtypes of PTCL with an abysmal prognosis (e.g. hepatosplenic T-cell lymphoma) allogeneic stem cell transplantation may be considered to consolidate first-line therapy. There are reports of long-term disease-free survival after allogeneic transplantation in patients with chemorefractory aggressive NHL (10), thus this approach may be considered in suitable patients with aggressive NHL not responding to salvage chemotherapy.
5. Conclusions HDT may have a role in improving the prognosis of young patients with poor risk aggressive lymphoma in front-line therapy and prospective randomised studies comparing HDT to new rituximab-containing standard therapies in DLBCL are needed. In PTCL prospective randomised trials are required to evaluate the benefit of HDT in the front-line setting and this will require international collaboration. Where possible these high-risk patients should be enrolled in clinical trials. In patients with relapsed and refractory chemosensitive aggressive lymphoma HDT is demonstrated to improve outcome compared to conventional chemotherapy. Current efforts are focused upon identifying the optimal salvage regimen, defining the role of rituximab pre- and post-ASCT and optimally incorporating radioimmunoconjugates in transplant protocols (Figure 1).
Figure 1: Summary algorithm for treatment of aggressive NHL At diagnosis PTCL
1) Enrol in clinical trial if possible 2) If no clinical trial available consider HDT as consolidation in young transplant eligible patient with high-risk features (e.g. aaIPI 2 or 3) and chemosensitive disease (except ALK-positive ALCL)
DLBCL
In young patient (≤60 years) eligible for transplant with 2-3 factors of aaIPI; 1) Enrol in clinical trial if possible 2) If no clinical trial, consider HDT in patients with chemosensitive disease after discussion of risks and benefits continue
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At relapse PTCL Salvage chemotherapy
If chemosensitive, patient eligible for transplant
If disease progression, depending on clinical circumstance
HDT
1) experimental protocol 2) non-cross resistant salvage 3) allogeneic transplant 4) palliation DLBCL Salvage chemotherapy plus rituximab
If chemosensitive, patient eligible for transplant
If disease progression, depending on clinical circumstance
HDT
1) experimental protocol 2) non-cross resistant salvage 3) allogeneic transplant 4) palliation
References 1. Koreth J, Cutler CS, Djulbegovic B, et al. High-dose therapy with single autologous transplantation versus chemotherapy for newly diagnosed multiple myeloma: A systematic review and meta-analysis of randomized controlled trials. Biol Blood Marrow Transplant 2007; 13: 183-196. 2. Mounier N, Gisselbrecht, C, Briere, J et al. All aggressive lymphoma subtypes do not share similar outcome after front-line autotransplantation: A matched-control analysis by the Groupe d’Etude des Lymphomes de l’Adulte (GELA). Ann Oncol 2004; 15: 1790-1797. 3. Mounier N, Simon D, Haioun C, et al. Impact of high-dose chemotherapy on peripheral T-cell lymphomas. J Clin Oncol 2002; 20: 1426-1427. 4. Reimer P, Ruediger T, Schertlin T, et al. Autologous Stem Cell Transplantation as FirstLine Therapy in Peripheral T-Cell Lymphomas. A Prospective Multicenter Study. ASH Annual Meeting Abstracts 2005; 106: 2074.
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5. Vose JM, Zhang M-J, Rowlings PA, et al. Autologous Transplantation for Diffuse Aggressive Non-Hodgkin’s Lymphoma in Patients Never Achieving Remission: A Report from the Autologous Blood and Marrow Transplant Registry. J Clin Oncol 2001; 19: 406-413. 6. Lerner RE, Burns LJ. Transformed lymphoma: An Achilles’ heel of non-Hodgkin’s lymphoma. BM Transplant 2003; 31: 531-537. 7. Song KW, Mollee P, Keating A, Crump M. Autologous stem cell transplant for relapsed and refractory peripheral T-cell lymphoma: Variable outcome according to pathological subtype. Br J Haematol 2003; 120: 978-985. 8. Vellenga E, van Putten WLJ, van ‘t Veer MB, et al. Rituximab improves the treatment results of DHAP-VIM-DHAP and ASCT in relapsed/progressive aggressive CD20+ NHL. A prospective randomized HOVON trial. Blood 2007: blood-2007-08-108415. 9. Peniket AJ, de Elvira MCR, Taghipour G, et al. An EBMT registry matched study of allogeneic stem cell transplants for lymphoma: Allogeneic transplantation is associated with a lower relapse rate but a higher procedure-related mortality rate than autologous transplantation. Bone Marrow Transplant 2003; 31: 667-678. 10.Doocey RT, Toze CL, Connors JM, et al. Allogeneic haematopoietic stem-cell transplantation for relapsed and refractory aggressive histology non-Hodgkin lymphoma. Br J Haematol 2005; 131: 223-230.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. For which of these patients has high-dose therapy followed by autologous transplant been proven to increase the PFS? a) Patient 45 years old with peripheral T-cell lymphoma in CR after first line of chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Patient 45 years old with diffuse large B-cell lymphoma in PR after first line of chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Patient 45 years old with diffuse large B-cell lymphoma, IPI score of 1, in CR after first line of chemotherapy . . . . . . . . . . . . . . . . . . . . . . d) Patient 45 years old with diffuse large B-cell lymphoma, IPI score of 4, in CR after first line of chemotherapy . . . . . . . . . . . . . . . . . . . . . . 2. Which one of these propositions is not true? a) High-dose therapy followed by autologous transplant is the standard treatment for a young patient with a positive PET scan after 6 cycles of R-CHOP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The efficacy of R-CHOP in patient with diffuse large B-cell 440
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CHAPTER 28 • High-grade NHL in adults
lymphoma has decreased the indications of high-dose therapy with autologous transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The best treatment for a relapsing patient with diffuse large B-cell lymphoma responding to salvage chemotherapy is high-dose therapy with autologous transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) A patient with diffuse large B-cell lymphoma in partial response may be treated with rituximab maintenance with same benefit and less toxicity than high-dose therapy with autologous transplant . . . . . . . . . 3. Which salvage regimen has been proven to be the best before high-dose therapy with autologous transplant in a patient with diffuse large B-cell lymphoma in first relapse? a) ESHAP combined with rituximab . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) DHAP combined with rituximab . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) ICE combined with rituximab . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All three of them . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. What is the best strategy for a patient aged 45 years who progressed during treatment with R-CHOP? a) Salvage with another chemotherapy regimen combined with rituximab . . b) Salvage with another chemotherapy regimen combined with rituximab followed by autologous transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Salvage with another chemotherapy regimen combined with rituximab followed by an allogeneic transplant. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) There is no good strategy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which one of these propositions is not true? a) A combination of zevalin plus BEAM has not yet proven its efficacy compare to BEAM alone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Outcome of a patient treated with high-dose therapy with autologous transplant is better if the patient reached CR with salvage regimen . . . . . . c) Outcome of a patient treated with high-dose therapy with autologous transplant is better if the patient relapsed more than 12 months after the first line treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Other anti-CD20 monoclonal antibodies may replace rituximab with the same efficacy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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*
CHAPTER 29
HSCT for low-grade non-Hodgkin’s lymphoma in adults
J.G. Gribben
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CHAPTER 29 • Low-grade NHL in adults
1. Introduction The classification of the non-Hodgkin’s lymphomas (NHLs) has been a challenge for pathologists as well as practising physicians. The World Health Organization (WHO) lymphoma classification is based upon cell of origin and pathophysiology of the lymphoma and does not include the terminology “low-grade lymphoma”. This is a clinical and not pathologic term, and defines those lymphomas which tend to grow and spread slowly and produce few symptoms. Follicular lymphoma (FL) is by far the most common of the low-grade lymphomas and is the second most common subtype of lymphoma worldwide, accounting for approximately 20% of malignant lymphomas in adults, but 40% of all lymphomas diagnosed in Western Europe and the USA. Follicular lymphoma affects predominantly adults, with a median age of 59 years and rarely occurs in individuals under age 20 years. Although low-grade NHLs are incurable, patients may live for many years. Ten-year survival for Stages I and II is between 60 and 80%. Five- and ten-year median survival for those diagnosed with Stage III disease is 60 and 40% respectively. Five- and ten-year median survival for those diagnosed with Stage IV disease is 50 and 10% respectively. Until recently there was little evidence that the natural history of FL had changed over the last 30 years from the median survival of 10 years from diagnosis, but this may be changing with the introduction of monoclonal antibodies in combination with chemotherapy, with more recent data suggesting that with improvements in treatment the median survival is now 12–14 years. The clinical course is extremely variable, with some patients having an extremely aggressive course and death within one year, while others may live for more than 20 years and never require therapy. The follicular lymphoma international prognostic index (FLIPI) is a five factor prognostic index based upon the clinical characteristics age, stage, number of nodal sites, haemoglobin and LDH level, and defines three prognostic risk groups of almost equal numbers of patients (1). This tool is useful in assessing the likely need for early treatment of patients and potential outcome, as well as in comparing the outcomes of different clinical trials. Multiple treatment approaches exist for advanced stage low-grade lymphomas, and since there is no clearly defined treatment algorithm for most patients with indolent lymphomas optimal care is that eligible patients should be included whenever possible in clinical trials. Patients remaining on an expectant course (watch and wait) should be followed every three months for history, physical examination and blood tests, including LDH, and special attention should be paid to any change in symptoms that might be suggestive of histologic transformation. Once transformation occurs, these patients should be treated as a high-grade lymphoma. Expectant management is the treatment of choice for asymptomatic patients with
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low bulk disease until clear indications for initiation of treatment are seen, except for those patients enrolled in clinical trials assessing the impact of early therapy. This approach is based upon the demonstration of no survival advantage for institution of immediate compared to deferred treatment until time of progression, a finding confirmed by three randomised trials. A major clinical trial question is whether identification of clinical or molecular risk factors can identify which patients are candidates for early therapy. A survival predictor score has also been developed from gene expression profiling studies. A major component of the gene expression prognostic signature is related to immune cells in the tumour microenvironment. Future guidelines for treatment may well be based upon clinical staging systems, genetic profiles and immune response signatures, but these factors do not yet help us to decide who should have immediate therapy, and decisions to treat with approaches including transplant should not be based on such measures except in the clinical trial setting. Treatment is indicated in patients with symptomatic disease, bulky lymphadenopathy and/or splenomegaly, risk of local compressive disease, marrow compromise or rapid disease progression. Once indicated, numerous treatment approaches are available. Options range from a watch and wait expectant management approach, to single agent chemotherapy or monoclonal antibody therapy with rituximab, to combination chemo-immunotherapy, with use of autologous or allogeneic transplant. Table 1 summarises the currently accepted indications for HSCT in low-grade lymphoma.
Table 1: Indications for transplantation in low-grade lymphoma Disease
Low-grade NHL
Disease status
Allo
Auto
Sibling donor
Alternative donor
CR1
NR
NR
CP
Relapse, CR2, CR3
CP
D
S
S: in standard use for selected patients; CP: to be undertaken in approved Clinical Protocols; D: developmental or pilot studies can be approved in specialist units; NR: not generally recommended
2. The role of autologous transplant in low-grade lymphomas 2.1. Relapsed disease (Table 2) The use of high-dose chemotherapy with autologous transplant in the treatment of low-grade lymphomas has not yet been fully established. The rational for considering transplantation is that the disease is incurable using standard approaches and young patients with indolent lymphomas will die of their disease. Promising results have 444
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CHAPTER 29 • Low-grade NHL in adults
Table 2: Autologous transplant for relapsed low-grade lymphoma Author
N
Follow-up (months)
OS (95% CI)
EFS (95% CI)
Freedman
153
96
66 (57–74)
42 (30–51) 1
(2)
Cortelazzo
103
36
47 (36–59) 61 (48–72) 64 (50–74) 1
(3)
Bierman
100
48
65 (54–75)
44 (33–55) 6
(4)
Colombat
42
60
83
7
(5)
Schouten
46
69
58
PFS (95% CI)
TRM %
Reference
(6)
22 unpurged
71 (52–91)
58 (37–79) 12.5
24 purged
77 (60–95)
55 (34–74) 8
been observed in a number of phase II studies (2–5). The EBMT sponsored CUP study (conventional chemotherapy, unpurged, purged autograft) is the only prospective randomised trial to assess the role of autologous HSCT in patients with relapsed FL (6). Purging was examined because patients with follicular lymphoma, frequently have significant bone marrow involvement. The results of the study demonstrated a PFS advantage and suggest an OS advantage of autologous transplant over conventional chemotherapy, with 4 year OS of 46% for the chemotherapy arm, versus 71% for the unpurged and 77% for the purged autologous transplant arms, with no benefit observed for those patients who underwent purging. There is some concern that the study was closed early because of slow accrual, with 140 of the planned 250 patients accrued and only 89 randomised. However, the results of the CUP trial are generally in line with that observed in the phase II studies, which include larger numbers and for which longer follow up is available. A major concern relates to the risk of development of secondary myelodysplasia/acute myeloid leukaemia. This complication appears to be decreased with the use of chemotherapy-only conditioning regimens without TBI. Based upon the results of the CUP trial and the encouraging results of the phase II studies, autologous transplant has become standard treatment (S) for patients with relapsed follicular lymphoma who are deemed to be high-risk, although precise criteria that define such “high-risk” patients are lacking. Patients with low-grade lymphomas who are potentially suitable candidates for transplantation should be referred to a transplant centre early to discuss the potential role and timing of transplantation. Best results are seen when transplantation is considered before the disease become chemo-refractory since high dose therapy and autologous transplant
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is an effective treatment approach for younger patients with chemo-responsive relapsed disease. Autologous transplant approaches in this disease setting must always be considered in the context of the improving results that are being seen with salvage therapy alone. 2.2 Autologous transplant as consolidation of first remission The role of high dose therapy and autologous HSCT in FL patients during first remission has been examined in three phase III randomised trials (7–9). The German low-grade study group (GLSG) trial (7) recruited 307 patients up to 60 years of age and patients who responded after induction chemotherapy with 2 cycles of CHOP or mitoxantrone-chlorambucil-prednisone (MCP) were randomised to autologous SCT or interferon (IFN)-alpha maintenance. Among 240 evaluable patients, the 5year PFS was 64.7% after autologous transplant, and 33.3% in the IFN-alpha arm (p< 0.0001). Acute toxicity was higher in the autologous transplant group, but early mortality was below 2.5% in both study arms. Longer follow-up is necessary to determine the effect of autologous transplant on OS. In the Groupe Ouest Est des Leucemies Aigues et des Maladies du Sang (GOELAMS) study, 172 newly diagnosed advanced FL patients were randomised either to cyclophosphamide, doxorubicin, teniposide, prednisone, (CHVP) and IFN-alpha or to high-dose therapy followed by purged autologous transplant (8). Patients treated with high-dose therapy and autologous transplant had a higher response rate than patients who received chemotherapy and IFN-alpha (81 versus 69%, p=0.045) and a longer median PFS (not reached versus 45 months), but this did not translate into a better OS due to an excess of secondary malignancies after autologous transplant. A subgroup of patients with a significantly higher event-free survival rate could be identified using the FLIPI. The Groupe Etude Lymphoma Folliculaire (GELF) 94 study enrolled 401 previously untreated advanced stage FL patients who were randomised to receive CHVP plus IFN-alpha compared with four courses of CHOP followed by high dose therapy with total body irradiation (TBI) and autologous HSCT. Response rates were similar in both groups (79 and 78% respectively) and 87% of eligible patients underwent autologous HSCT. Intent-to-treat analysis after a median follow-up of 7.5 years showed no difference between the two arms for OS (p=0.53) or PFS (p=0.11). Long-term follow-up demonstrated no statistically significant benefit in favour of first-line auto-HSCT in patients with follicular lymphoma, which they conclude should be reserved for relapsed patients. In view of these results, autologous HSCT should be used in first remission only in the setting of clinical trials (CP).
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CHAPTER 29 • Low-grade NHL in adults
3. Role and outcome of allogeneic HSCT for low-grade lymphoma 3.1. HLA identical sibling transplant and unrelated donors There is a trend towards increasing use of allogeneic transplant in the management of low-grade lymphomas. There have been no randomised controlled trials in this disease setting, but long term PFS has been observed in patients with low-grade NHLs after allogeneic transplant (10–12) as outlined in Table 3. In a report of the International Bone Marrow Transplant Registry (IBMTR), results are described for 904 patients with FL, 176 of whom underwent allogeneic transplant from HLA matched sibling, 131 patients underwent autologous transplant using purged stem cells and 597 using unpurged autologous stem cells (13). The treatment related mortality (TRM) in these three groups was 30, 14 and 8% respectively, disease recurrence in 21, 43 and 58% and 5 year overall survival was 51, 62 and 55% respectively. The use of TBI containing regimens was associated with increased TRM but decreased risk of relapse. The use of HLA identical sibling allogeneic transplant was associated with increased TRM - compared to autologous transplantation - but significantly lower risk of disease recurrence in keeping with a graft versus lymphoma effect in this disease. Long term PFS has been observed after allogeneic SCT even in patients with refractory FL (12). In 29 FL patients, 11 of whom had refractory disease, the nonrelapse mortality was 24% and there was a 23% incidence of relapse. Twenty of these patients underwent allogeneic transplantation from HLA identical siblings. The five year OS was 58% with 53% event free survival. A group of patients with very poor outcome are those patients who have relapsed after previous autologous transplant. The outcome following myeloablative allogeneic transplant of 114 such patients have been reported from the IBMTR (14). The treatment related mortality was 22% and
Table 3: Allogeneic transplant for low-grade lymphoma Histology
No. of Median age Status patients (range)
Conditioning Outcome
TRM Reference (%)
FL 6 SLL 4
10
42 (31–55)
3 sensitive 8 refractory
CY/TBI
68% PFS at 2 years
30
(10)
FL 93 SLL 20
113
38 (15–61)
66 sensitive 39 refractory
TBI 93 Non TBI 20
49% PFS at 3 years
40
(11)
FL 29
29
42 (20–53)
6 induction failure 18 sensitive 6 refractory
27 TBI
53% PFS at 5 years
24
(12)
FL: follicular lymphoma; SLL: small lymphocytic lymphoma
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the probability of disease progression was 52% at 3 years. The use of TBI conditioning regimens and achievement of CR at the time of allogeneic SCT were associated with improved outcome. In view of the high TRM and the long disease course allogeneic transplant from HLA matched siblings is not recommended in first CR or PR (NR). 3.2. Reduced intensity HSCT The use of reduced intensity conditioning regimens results in decreased TRM and appears to be associated with improved outcome. There have been no randomised clinical trials, but a number of phase II studies have clearly demonstrated evidence of a graft versus lymphoma effect that can be exploited in low-grade NHLs, as outlined in Table 4 (15–23). In many studies results are combined together for patients with low-grade and high-grade NHL, and for HLA matched siblings and unrelated donors, making full comparison of outcomes in specific subgroups impossible. The outcome following reduced intensity conditioning transplant regimen a immunosuppressive
Table 4: Reduced intensity conditioning allogeneic transplant for low-grade lymphoma No. of patients
Prior treatment Median (range)
Conditioning
GvHD (%)
Graft Outcome failure
23
3 (2–6)
FLU/BU/ATG
Acute 34
0
40% PFS at 3 yrs (15)
20
2 (1–5)
FLU/CY/rituximab Acute 20 Chronic 64
0
84% PFS at 2 yrs (16)
13
3 (1–7)
200 cGY TBI
Acute 54 Chronic 62
1
7 CR
(17)
44
3 (0–6)
FLU/MEL/ alemtuzumab
Acute 16 Chronic 2
1
22 CR 11 PR
(18)
88 (mixed histologies)
4 (2–6)
FLU/MEL/ alemtuzumab
Acute 30
4
73% OS at 3 yrs (low-grade)
(19)
65
2 (1–6) 11% prior autologous SCT
BEAM/ alemtuzumab
Acute 17 Chronic 17
3
69% PFS at 2 yrs (20)
47
62% prior autolo- FLU/MEL/ gous transplant alemtuzumab
Acute 23 Chronic 6
2
75% OS at 1 yr 61% PFS at 1 yr
(22)
188
3 Various 48% prior autologous transplant
Acute 37 Chronic 17
6
25% TRM 50% OS at 2 yrs
(23)
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therapy has been reported for 81 patients with lymphoma including 41 with lowgrade NHL (19). Patients received a conditioning regimen consisting of alemtuzumab, fludarabine and melphalan, and received short course cyclosporin as GvHD prophylaxis. The use of this conditioning regimen was associated with a low incidence of GvHD and the treatment related mortality was decreased in patients with low-grade compared to higher grade histology. The three-year progression free survival was 65% for patients with low-grade lymphoma. 3.3. Role of haploidentical donors and cord blood transplant in low-grade lymphoma The use of alternative donor sources including haploidentical transplant and cord blood transplant are considered developmental and should be considered for selected patients only in specialised centres.
4. Post-transplant management 4.1. Nature and role of cellular or chemotherapy post-transplant The use of monoclonal antibodies to attempt to eradicate any residual lymphoma cells after autologous or allogeneic transplant is being examined in ongoing clinical trials. Donor lymphocyte infusion (DLI) are effective in treating relapse after allogeneic transplant provides very strong evidence for a graft versus lymphoma effect that can be exploited in indolent lymphomas. 4.2. Nature and role of minimal residual disease monitoring after transplant Detection of MRD has been a useful surrogate marker for tracking long-term PFS in patients examining the autologous stem cells or serial samples after both autologous and allogeneic transplant (24–27). The increasing use of therapeutic monoclonal antibodies may make the use of peripheral blood rather than bone marrow a less useful cell source for monitoring MRD, since monoclonal antibodies appear to clear peripheral blood very successfully. There is concern using detection of the BCL2/IgH rearrangement as a marker for MRD detection in FL since this rearrangement can be found in the blood of many healthy individuals, but these cells seem to be cleared by chemotherapy so that when MRD is detected after chemotherapy, it is usually from the malignant clone (28). Monitoring and treatment of MRD in lowgrade lymphomas is indicated in the setting of clinical trials.
References 1. Solal-Celigny P, Roy P, Colombat P, et al. Follicular lymphoma international prognostic index. Blood 2004; 104: 1258-1265.
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2. Freedman AS, Neuberg D, Mauch P, et al. Long-term follow-up of autologous bone marrow transplantation in patients with relapsed follicular lymphoma. Blood 1999; 94: 3325-3333. 3. Cortelazzo S, Rambaldi A, Rossi A, et al. Intensification of salvage treatment with highdose sequential chemotherapy improves the outcome of patients with refractory or relapsed aggressive non-Hodgkin’s lymphoma. Br J Haematol 2001; 114: 333-341. 4. Bierman PJ, Vose JM, Anderson JR, et al. High-dose therapy with autologous hematopoietic rescue for follicular low-grade non-Hodgkin’s lymphoma. J Clin Oncol 1997; 15: 445-450. 5. Colombat P, Donadio D, Fouillard L, et al. Value of autologous bone marrow transplantation in follicular lymphoma: A France Autogreffe retrospective study of 42 patients. Bone Marrow Transpl 1994; 13: 157-162. 6. Schouten HC, Qian W, Kvaloy S, et al. High-dose therapy improves progression-free survival and survival in relapsed follicular non-Hodgkin’s lymphoma: Results from the randomized European CUP trial. J Clin Oncol 2003; 21: 3918-3927. 7. Lenz G, Dreyling M, Schiegnitz E, et al. Myeloablative radiochemotherapy followed by autologous stem cell transplantation in first remission prolongs progression-free survival in follicular lymphoma: Results of a prospective, randomized trial of the German Low-Grade Lymphoma Study Group. Blood 2004; 104: 2667-2674. 8. Deconinck E, Foussard C, Milpied N, et al. High-dose therapy followed by autologous purged stem-cell transplantation and doxorubicin-based chemotherapy in patients with advanced follicular lymphoma: A randomized multicenter study by GOELAMS. Blood 2005; 105: 38173823. 9. Sebban C, Mounier N, Brousse N, et al. Standard chemotherapy with interferon compared with CHOP followed by high-dose therapy with autologous stem cell transplantation in untreated patients with advanced follicular lymphoma: The GELF-94 randomized study from the Groupe d’Etude des Lymphomes de l’Adulte (GELA). Blood 2006; 108: 2540-2544. 10.Verdonck LF, Dekker AW, Lokhorst HM, et al. Allogeneic versus autologous bone marrow transplantation for refractory and recurrent low-grade non-Hodgkin’s lymphoma. Blood 1997; 90: 4201-4205. 11.van Besien K, Sobocinski KA, Rowlings PA, et al. Allogeneic bone marrow transplantation for low-grade lymphoma. Blood 1998; 92: 1832-1836. 12.Toze CL, Barnett MJ, Connors JM, et al. Long-term disease-free survival of patients with advanced follicular lymphoma after allogeneic bone marrow transplantation. Br J Haematol 2004; 127: 311-321. 13.van Besien K, Loberiza FR Jr, Bajorunaite R, et al. Comparison of autologous and allogeneic hematopoietic stem cell transplantation for follicular lymphoma. Blood 2003; 102: 3521-3529. 14.Freytes CO, Loberiza FR, Rizzo JD, et al. Myeloablative allogeneic hematopoietic stem cell transplantation in patients who experience relapse after autologous stem cell transplantation for lymphoma: a report of the International Bone Marrow Transplant Registry. Blood 2004; 104: 3797-3803. 15.Nagler A, Slavin S, Varadi G, et al. Allogeneic peripheral blood stem cell transplantation using a fludarabine-based low intensity conditioning regimen for malignant lymphoma. Bone Marrow Transplant 2000; 25: 1021-1028. 450
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16.Khouri IF, Saliba RM, Giralt SA, et al. Nonablative allogeneic hematopoietic transplantation as adoptive immunotherapy for indolent lymphoma: Low incidence of toxicity, acute graftversus-host disease, and treatment-related mortality. Blood 2001; 98: 3595-3599. 17.McSweeney PA, Niederwieser D, Shizuru JA, et al. Hematopoietic cell transplantation in older patients with hematologic malignancies: Replacing high-dose cytotoxic therapy with graft-versus-tumor effects. Blood 2001; 97: 3390-3400. 18.Kottaridis PD, Milligan DW, Chopra R, et al. In vivo CAMPATH-1H prevents graft-versushost disease following nonmyeloablative stem cell transplantation. Blood 2000; 96: 2419-2425. 19.Morris E, Thomson K, Craddock C, et al. Outcomes after alemtuzumab-containing reducedintensity allogeneic transplantation regimen for relapsed and refractory non-Hodgkin lymphoma. Blood 2004; 104: 3865-3871. 20.Faulkner RD, Craddock C, Byrne JL, et al. BEAM-alemtuzumab reduced-intensity allogeneic stem cell transplantation for lymphoproliferative diseases: GVHD, toxicity, and survival in 65 patients. Blood 2004; 103: 428-434. 21.Khouri IF, Lee MS, Saliba RM, et al. Nonablative allogeneic stem cell transplantation for chronic lymphocytic leukemia: Impact of rituximab on immunomodulation and survival. Exp Hematol 2004; 32: 28-35. 22.Chakraverty R, Peggs K, Chopra R, et al. Limiting transplantation-related mortality following unrelated donor stem cell transplantation by using a nonmyeloablative conditioning regimen. Blood 2002; 99: 1071-1078. 23.Robinson SP, Goldstone AH, Mackinnon S, et al. Chemoresistant or aggressive lymphoma predicts for a poor outcome following reduced-intensity allogeneic progenitor cell transplantation: an analysis from the Lymphoma Working Party of the European Group for Blood and Bone Marrow Transplantation. Blood 2002; 100: 4310-4316. 24.Gribben JG, Freedman AS, Neuberg D, et al. Immunologic purging of marrow assessed by PCR before autologous bone marrow transplantation for B-cell lymphoma. N Engl J Med 1991; 325: 1525-1533. 25.Gribben JG, Neuberg D, Freedman AS, et al. Detection by polymerase chain reaction of residual cells with the bcl-2 translocation is associated with increased risk of relapse after autologous bone marrow transplantation for B-cell lymphoma. Blood 1993; 81: 3449-3457. 26.Corradini P, Ladetto M, Zallio F, et al. Long-term follow-up of indolent lymphoma patients treated with high-dose sequential chemotherapy and autografting: Evidence that durable molecular and clinical remission frequently can be attained only in follicular subtypes. J Clin Oncol 2004; 22: 1460-1468. 27.Ladetto M, Vallet S, Benedetti F, et al. Prolonged survival and low incidence of late toxic sequelae in advanced follicular lymphoma treated with a TBI-free autografting program: updated results of the multicenter consecutive GITMO trial. Leukemia 2006; 20: 18401847. 28.Ladetto M, Drandi D, Compagno M, et al. PCR-detectable nonneoplastic Bcl-2/IgH rearrangements are common in normal subjects and cancer patients at diagnosis but rare in subjects treated with chemotherapy. J Clin Oncol 2003; 21: 1398-1403.
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which statement is not true regarding the CUP trial? a) The study demonstrated a PFS advantage of ASCT over chemotherapy . . . . b) Purging was beneficial . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) There is a concern that the study was closed early because of slow accrual . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) This is the only prospective randomised trial to assess ASCT in relapsed FL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which statement is not true regarding auto-HSCT for FL? a) Auto-HSCT has become standard treatment for pts with relapsed FL . . . . . . b) Best results are seen when auto-HSCT is considered before the disease becomes chemo refractory . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Auto-HSCT should be used in first remission only in clinical trials . . . . . . . . d) Secondary MDS/AML is less frequent if the conditioning contains TBI . . . 3. Which process results in the highest treatment related mortality in relapsed FL? a) Allogeneic transplant from with standard conditioning . . . . . . . . . . . . . . . . . . . . b) Autologous transplant using purged stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Autologous transplant using unpurged stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . d) Allogeneic transplant with reduced intensity conditioning . . . . . . . . . . . . . . . . 4. Which sentence is true? a) Randomised trials demonstrated long term PFS following allo-SCT in FL . b) Reduced intensity conditioning does not decrease TRM . . . . . . . . . . . . . . . . . . . c) There is a strong graft versus low-grade lymphoma effect . . . . . . . . . . . . . . . . . d) DLI is not effective in FL. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which of the following transplant should be considered standard of care in FL? a) Autologous transplant without purging in chemosensitive relapse . . . . . . . . 452
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CHAPTER 29 • Low-grade NHL in adults
b) Autologous transplant without purging in CR1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Alternative donor transplant in CR2 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Sibling donor transplant in CR1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 30
HSCT for Hodgkin’s lymphoma in adults
A. Sureda
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CHAPTER 30 • HL in adults
1. Introduction Newly diagnosed patients with advanced stage Hodgkin’s lymphoma (HL) have an excellent prognosis and the vast majority of them can be cured with initial treatment. In contrast, the prognosis of most patients relapsing after first-line therapy remains poor. In patients in sensitive relapse or in second complete remission (CR) high-dose chemotherapy and auto-HSCT is nowadays considered to be the treatment of choice. Auto-HSCT is also an option for those patients with primary refractory disease (PRD). Allo-HSCT is still considered an experimental procedure for patients with relapsed or refractory HL (1).
2. Auto-HSCT in refractory/relapsed Hodgkin’s lymphoma 2.1. Conditioning regimens in the auto-HSCT setting There has never been a randomised trial comparing preparative regimens for transplant for relapsed HL. The only recent study addressing this question was a retrospective review by investigators at the Fred Hutchinson Cancer Research Center. Between 1990 and 1998, 92 patients with relapsed HL were transplanted with either a TBI - based regimen or busulfan/melphalan/thiotepa. There was no difference in 5-year OS (57 vs. 52%) or EFS (49 vs. 42%) rates for patients treated with TBI or chemotherapy only. Given the reports of increased risk of second cancers and myelodysplasia following TBI, a chemotherapy-only preparative regimen is currently favoured by most transplant centres. 2.2. Auto-HSCT for relapsed Hodgkin’s lymphoma The use of auto-HSCT is now considered the standard of care for relapsed HL patients (1). The first randomised trial of transplant for relapsed disease was a small trial from the British National Lymphoma Investigation (BNLI) comparing auto-HSCT with BEAM to mini-BEAM without auto-HSCT (2) in patients with active HL, for whom conventional therapy had failed. Twenty patients were assigned to treatment with BEAM plus auto-HSCT and 20 to mini-BEAM. All had been followed up for at least 12 months (median 34 months). Five BEAM recipients died compared with 9 miniBEAM recipients. That difference was not significant (p = 0.318) and there was no difference in OS. However, both 3-year EFS and PFS showed significant differences in favour of BEAM plus auto-HSCT (p = 0.025 and p = 0.005, respectively). In the second randomised trial, which was performed by investigators of the German Hodgkin’s disease Study Group and the Lymphoma Working Party (LWP) of the EBMT, 161 patients with relapsed HL were randomly assigned two cycles of DexaBEAM and either two further courses of Dexa-BEAM or high-dose BEAM and auto-
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HSCT (3). Of the 117 patients with chemosensitive relapse there was a significant improvement in 3-year freedom from treatment failure (FFTF) for patients undergoing auto-HSCT compared to 4 cycles of Dexa-BEAM (55 vs. 34%, p = 0.019). With a median follow up of 39 months, the 3-year FFTF was significantly better for patients treated with BEAM. No significant improvement of auto-HSCT over conventional salvage chemotherapy (CT) in terms of FFTF could be observed in the small sub-group of patients treated for multiple-relapsed disease (n = 24). There was no statistically significant difference in OS for any sub-group of patients. 2.3. Auto-HSCT for patients with primary refractory disease The prognosis of patients with PRD, defined as progression during first-line CT or within 3 months after the end of therapy, is extremely poor. Nevertheless as opposed to non-Hodgkin’s lymphoma there seems to be a general consensus that even patients who fail first- and second-line CT may still enjoy a 20%–30% chance of cure with auto-HSCT. In the EBMT registry analysis (4), 175 HL patients with PRD were reviewed. Actuarial 5-year PFS and OS were 32 and 36%, respectively. In the Autologous Bone Marrow Transplant Registry (ABMTR) analysis on 122 patients undergoing auto-HSCT after an induction failure (IF) (5), actuarial probabilities at 3 years were 38 and 50% for PFS and OS, respectively. Lazarus et al. (5) found that the presence of B symptoms at diagnosis as well as Karnofsky status at auto-HSCT correlated with survival. In the EBMT analysis (4), patients receiving more than one line of CT before transplantation did worse, both in terms of OS and PFS. The long-term outcome of 75 consecutive patients with biopsy-confirmed HL at the completion of primary CT has been summarised by the Memorial Sloan Kettering Cancer Center group (6). All patients underwent standard-dose salvage therapy followed by involved field radiotherapy (RT). Patients without progression went on to receive high-dose etoposide, cyclophosphamide and either total lymphoid irradiation or carmustine followed by bone marrow or peripheral stem cell rescue. Patients who had shown less than a 25% decrease in tumour burden with standard salvage therapy (n = 27) prior to auto-HSCT had a 10-year EFS of 17 versus 60% for those with at least a 25% decrease on standard salvage therapy (n = 48).
3. Allo-HSCT in refractory/relapsed Hodgkin’s lymphoma 3.1. Myeloablative conditioning and allo-HSCT in Hodgkin’s lymphoma The first reports on allo-HSCT in patients with HL appeared in the mid-eighties. Two larger registry-based studies published in 1996 gave disappointing results. Gajewski 456
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et al. analysed 100 HL patients allografted from HLA-identical siblings (7). The 3year rates for OS, DFS, and the probability of relapse were 21, 15 and 65%, respectively. The major problems after transplantation were persistent or recurrent disease, or respiratory complications, which accounted for 35 to 51% of deaths. Acute and/or chronic GvHD did not significantly reduce the risk of relapse. A casematched analysis including 45 allografts and 45 autografts reported to the EBMT (8) did not find significant differences in actuarial probabilities of OS, PFS, and relapse rates between allo-HSCT and auto-HSCT (25, 15, 61% vs. 37, 24, 61%, respectively at 4 yrs). The actuarial TRM at 4 years was significantly higher for allografts than for autografts (48 vs. 27%, p = 0.04). Acute GvHD ≥ grade II was associated with a significantly lower risk of relapse, but also with a lower survival rate. 3.2. Reduced intensity conditioning and allo-HSCT in Hodgkin’s lymphoma Since the first clinical experiences that suggested that allo-HSCT after a nonmyeloablative conditioning (RIC allo-HSCT) might represent an interesting alternative to classical allo-HSCT, a number of reports have addressed the question whether RIC allo-HSCT might also work for patients with HL (Figure 1). Several groups have already published their results in small groups of patients with a relative short follow-up (9–12) (Table 1). The largest cohort of patients treated with RIC allo-HSCT in HL was recently reported by the LWP of the EBMT (13) and
Figure 1: Allo-HSCT for relapsed and refractory Hodgkin’s lymphoma. Comparison between conventional conditioning and RIC regimens RIC Conventional 100%
% of HSCT
80% 60% 40% 20% 0%
1994
1996
1998
2000
2002
2004
2006
Experience of the LWP of the EBMT (with permission)
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Table 1: Clinical characteristics and outcome of patients with relapsed or refractory Hodgkin’s lymphoma treated with a RIC allo-HSCT BBMT 2004 (12)
BMT 2005 (9)
Lancet 2005 (11) BBMT 2006 (10)
N. of patients
27
58
49
40
Sex (M:F)
–
–
25 / 24
24 / 16
Age [median (range)] in years
37 (21–65)
32 (19–59)
32
31 (16–53)
Previous lines of CT 5 (2–9) [median (range)]
5 (2–9)
5 (3–8)
4 (2–6)
Prior RT (%)
25 (92)
44 (75)
–
23 (58)
Prior auto-HSCT (%)
24 (89)
48 (83)
44 (89)
29 (73)
Dx to RIC-allo [median (range)] in months
–
23 (9–145)
4.8 (0.6–14.8)
37 (11–300)
Auto-HSCT to 16 (2–78) RIC-allo [median (range)] in months
5 (1–34)
–
17 (4–146)
Disease status at RIC-allo (sensitive/refractory)
20 / 7
30 / 28
36 / 13
20 / 2
Type of donor (MRD / UD)
18 / 9
25 / 33
31 / 18
38 / 2
aGvHD (grades II-IV)
47% (MRD) / 55% (UD)
28%
16%
45%
cGvHD
50% (MRD) / 60% (UD)
74%
14%
45%
100-day TRM
7%
7%
4.1%
12%
1-year TRM
35%
15% (24 months)
16% (2-year)
25%
PFS
11% (MRD) / 32% (24 months) 35% (UD) (1-year)
32% (4-year)
32% (2-year)
OS
39% (MSD) / 64% (24 months) 75% (UD) (1-year)
56% (4-year)
48% (2-year)
M: Male; F: Female; CT: Chemotherapy; RT: Radiotherapy; auto-HSCT: Autologous stem cell transplantation; Dx: Diagnosis; RIC-allo: Reduced intensity allogeneic stem cell transplantation; MRD: Related donor; UD: Unrelated donor; aGvHD: Acute graft versus host disease; cGvHD: Chronic graft versus host disease; TRM: Transplant related mortality; PFS: Progression free survival; OS: Overall survival 458
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CHAPTER 30 • HL in adults
included 374 patients. Patients had received an average of four lines of prior therapy (1–8) and 288 patients (77%) had failed one or two auto-HSCT. At the time of allo-HSCT, 79 patients (21%) were in CR, 146 patients (39%) had chemosensitive disease and 149 patients (40%) had chemoresistant disease. Two hundred and thirtyfour patients (63%) were allografted from a matched sibling donor, 112 (30%) from a matched unrelated donor, and 28 from a mismatched donor (7%). Grade II-IV acute GvHD (aGvHD) was reported in 27% of patients, chronic GvHD (cGvHD) in 40% of patients at risk. The 100-day TRM was 12% but increased to 20% at 12 months, and to 22% at three years; it was significantly worse for patients with chemoresistant disease. Two-year PFS was 29% and again significantly worse for patients with chemoresistant disease (p < 0.001). In a landmark analysis the development of either acute or chronic GvHD by 9 months post transplant was associated with a significantly lower relapse rate. 3.3. Comparison of myeloablative and reduced-intensity conditioning prior to allo-HSCT in relapsed and refractory Hodgkin’s lymphoma The LWP of the EBMT has performed the only analysis reported so far which compares outcomes after reduced-intensity (n = 97) or myeloablative conditioning (n = 93) and allo-HSCT in patients with HL (14). Non-relapse mortality was significantly decreased in the RIC allo-HSCT group [HR 2.43 (95% CI 1.48–3.98), p < 0.001]. PFS and OS were also better in the reduced intensity group [HR 1.28 (95% CI 0.92–1.78), p = 0.1 and HR 1.62 (95% CI 1.15–2.28), p = 0.005]. The development of cGvHD significantly decreased the incidence of relapse after transplantation, which translated into a better PFS. This analysis indicates that RIC allo-HSCT is able to significantly reduced TRM after transplantation and improves the long-term outcome of relapsed and refractory patients treated with an allograft. 3.4. Graft-versus-Hodgkin effect The significant reduction of the TRM observed in the RIC allo-HSCT has been able to put in evidence the existence of a graft-versus-HL effect. Direct evidence of a graftvs-HL effect comes from two main sources: the demonstration that the development of acute or chronic GvHD after allo-HSCT is associated to a lower relapse rate and the clinical information coming from donor lymphocyte infusions (DLIs). Relapse rate is significantly lower in those patients developing GvHD after transplantation. Additionally, reported disease responses to DLIs range between 30 to 55% (9–11, 13).
4. Conclusions The use of auto-HSCT is now considered the standard of care for relapsed HL patients. Two randomised trials showed significant benefit in FFTF for auto-HSCT HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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over conventional chemotherapy for relapsed disease. The results of these trials have resulted in the recommendation of auto-HSCT at time of first relapse for even the most favourable patients. Results of auto-HSCT in PRD are poor and new therapeutic alternatives should be sought for these patients. Allo-HSCT has been increasingly used in relapsed or refractory HL patients with the introduction of RIC protocols. RIC allo-HSCT significantly decreases TRM in relation to conventional protocols and improve long-term outcome of these patients. The significant reduction of the TRM observed in the RIC allo-HSCT setting has been able to demonstrate the existence of a graft-versus-HL effect mostly associated to the development of GvHD after transplantation. Nevertheless, allo-HSCT in HL is still considered an experimental procedure and patients should be included in prospective clinical trials.
References 1. Ljungman P, Urbano-Ispizua A, Cavazzana-Calvo M, et al. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: definitions and current practice in Europe. Bone Marrow Transplant 2006; 37: 439-449. 2. Linch DC, Winfield D, Goldstone AH, et al. Dose intensification with autologous bonemarrow transplantation in relapsed and resistant Hodgkin’s disease: Results of a BNLI randomized trial. Lancet 1993; 341: 1051-1054. 3. Schmitz N, Pfistner B, Sextro M, et al. Aggressive conventional chemotherapy compared with high-dose chemotherapy requiring autologous haemopoietic stem cell transplantation for relapsed chemosensitive Hodgkin's disease: A randomised trial. Lancet 2002; 359: 20652071. 4. Sweetenham JW, Carella AM, Taghipour G, et al. High dose therapy and autologous stem cell transplantation for adult patients with Hodgkin’s disease who fail to enter remission after induction chemotherapy: Results in 175 patients reported to the EBMT. J Clin Oncol 1999; 17: 3101-3109. 5. Lazarus HM, Rowlings PA, Zhang M-J, et al. Autotransplants for Hodgkin’s disease in patients never achieving remission: A report from the Autologous Blood and Marrow Transplant Registry. J Clin Oncol 1999; 17: 534-545. 6. Moskowitz CH, Kewalramani T, Nimer SD, et al. Effectiveness of high dose chemoradiotherapy and autologous stem cell transplantation for patients with biopsy-proven primary refractory Hodgkin’s disease. Br J Haematol 2004; 124: 645-652. 7. Gajewski JL, Phillips GL, Sobocinski KA, et al. Bone marrow transplants from HLAidentical siblings in advanced Hodgkin’s disease. J Clin Oncol 1996; 14: 572-578. 8. Milpied N, Fielding AK, Pearce RM, et al. Allogeneic bone marrow transplant is not better than autologous transplant for patients with relapsed Hodgkin’s disease. J Clin Oncol. 1996; 14: 1291-1296. 9. Anderlini P, Saliba R, Acholonu S, et al. Reduced-intensity allogeneic stem cell transplantation in relapsed and refractory Hodgkin’s disease: Low transplant-related 460
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mortality and impact of intensity of conditioning regimen. Bone Marrow Transplant 2005; 35: 943-951. 10.Alvarez I, Sureda A, Caballero MD, et al. Non-myeloablative stem cell transplantation is an effective therapy for refractory or relapsed Hodgkin’s lymphoma: Results of a Spanish prospective cooperative protocol. Biol Blood Marrow Transplant 2006; 12: 172-183. 11.Peggs KS, Hunter A, Chopra R, et al. Clinical evidence of a graft-versus-Hodgkin’slymphoma effect after reduced-intensity allogeneic transplantation. Lancet 2005; 365: 1934-1941. 12.Burroughs LM, Maris MB, Sandmaier BM, et al. HLA-matched related (MRD) or unrelated donor (URD) nonmyeloablative conditioning and hematopoietic cell transplant (HCT) for patients with advanced Hodgkin disease (HD) [abstract]. Biol Blood Marrow Transplant 2004; 10 (Suppl. 1): 73-74. 13.Robinson S, Taghipour G, Sureda A, et al. Reduced intensity allogeneic stem cell transplantation for Hodgkin’s disease. Outcome depends primarily on disease status at the time of transplantation. Blood 2004; 104: 639a. 14.Sureda A, Robinson S, Canals C, et al. Reduced-Intensity Conditioning Compared With Conventional Allogeneic Stem-Cell Transplantation in Relapsed or Refractory Hodgkin's Lymphoma: An Analysis From the Lymphoma Working Party of the European Group for Blood and Marrow Transplantation. J Clin Oncol 2008; 26: 455-462.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. In terms of indications for auto-HSCT in patients with HL in 1st CR, choose the correct answer: a) Nowadays auto-HSCT is not indicated for patients with HL in 1st CR. . . . . . b) Autografting should be considered for patients in 1st CR with at least 3 adverse prognostic factors at diagnosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Auto-HSCT can be done, but we have to take into consideration the excessive risk of secondary malignancies after auto-HSCT . . . . . . . . . . . . . . . . . d) The published phase II trials indicate a higher than usual TRM in this subgroup of patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which do you consider the best therapeutic strategy for a 40-year old female with relapsed HL? The duration of the 1st CR was 10 months and the Ann Arbor stage at relapse was IVB: a) Collection of peripheral blood stem cells and auto-HSCT without prior salvage chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . HAEMATOPOIETIC STEM CELL TRANSPLANTATION
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b) Consider an allo-HSCT as the best therapeutic option. Look for a matched unrelated donor if the patient does not have a matched sibling donor available . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Treat the patient with a salvage chemotherapy protocol, collect peripheral blood stem cells and proceed to auto-HSCT . . . . . . . . . . . . . . . . . . . . . d) If the patient has not received RT before, consider RT in all involved areas as a curative treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. In patients with primary refractory HL, all of the following statements are true except one. Indicate the incorrect answer: a) The TRM of an autologous procedure is similar to that experienced by patients autografted in sensitive relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Response to second line chemotherapy is one of the best prognostic indicators of the long-term outcome of an auto-HSCT . . . . . . . . . . . . . . . . . . . . . c) New therapeutic strategies need to be developed. The results obtained with an auto-HSCT are clearly inferior to those in relapsed patients . . . . . d) There is no indication for an auto-HSCT in primary refractory patients due to the extremely poor long-term outcome of this subgroup of patients with the autologous procedure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Indicate the correct answer in relation to the conditioning regimens used in patients with HL: a) In the allogeneic setting, the use of low dose TBI seems to improve the outcome after transplant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The use of TBI in the autologous setting seems to be associated with an increased risk of secondary malignancies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The protocol CBV (cyclophosphamide, BCNU and VP-16) is superior to the BEAM regimen (BCNU, VP-16, ARA-C and melphalan) for HL patients in the autologous setting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Myeloablative conditioning regimens are increasingly being used in the allogeneic setting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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5. We have a 50-year old female patient with HL who relapses 13 months after an auto-HSCT with advanced stage disease and we want to consider an allo-HSCT as a therapeutic option for her. Indicate the correct answer: a) There is no indication for an allo-HSCT in HL nowadays . . . . . . . . . . . . . . . . . . . b) There is no indication for an unrelated donor search in this patient. Results of allo-HSCT from matched unrelated donors have been demonstrated to be inferior to those with HLA matched sibling donors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The use of RIC regimens has not allowed us to reduce the high TRM associated with conventional conditioning regimens . . . . . . . . . . . . . . . . . . . . . . d) It has been demonstrated that the best GvHD prophylaxis needs to include Campath 1H . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 31
HSCT for solid tumours in adults
M. Bregni
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CHAPTER 31 • Solid tumours in adults
1. Introduction In the years 2005–2007, 3631 autologous and 192 allogeneic transplants for solid tumours in adults were reported to the EBMT. Most transplants (3596) were performed with PBPC, a small minority (215) with BM cells, and 7 with cord blood. Diseases treated by transplant can be roughly categorised as: sarcoma and primitive neuroectodermal tumour (pNET) (891), central nervous system (CNS) tumours, including neuroblastoma (1041), germ cell tumours (934), epithelial tumours (624). These numbers are probably an underestimate because not all transplants are currently registered with the EBMT. However, some general conclusions can be drawn: 1. Compared with previous years, the number of transplants is: - increasing for sarcoma/pNET; - stable for CNS and germ cell tumours; - decreasing for epithelial tumours. 2. The existence of a dose-response effect in epithelial tumours (breast, ovarian, small cell lung cancer) is still a matter of investigation. The benefit of autologous transplant in breast cancer has not been confirmed by randomised studies; however, an appreciable number of transplants is still being performed for breast cancer (206 in 2005, 116 in 2006). At the 2007 San Antonio Breast Cancer Symposium, the results of the EBMT-MD Anderson meta-analysis on 15 randomised phase III trials comparing high-dose with conventional-dose chemotherapy have been reported (1). These results, while not supporting a survival advantage for high-dose chemotherapy (HDCT), suggest that selected subsets of patients may profit from HDCT. In 2007 the report was published by AGO-OVAR/AIO and EBMT of the randomised phase III study for first-line treatment of advanced ovarian cancer in which high-dose sequential chemotherapy with peripheral blood stem cell support was compared with standard dose chemotherapy (2). No statistically significant difference in progression-free survival or OS was observed, and the authors concluded that HDCT does not appear to be superior to conventional dose chemotherapy. Small-cell lung cancer is the third epithelial tumour for which a phase III trial has been organised by EBMT. The results of this randomised study will be published shortly (3). 3. Allografting for solid tumours decreased in the period 2005–2007, for all indications. The transplant-related mortality attributable to the conditioning regimen and to GvHD (around 10% in most studies) makes it difficult to offer allografting as a salvage treatment to patients with metastatic disease and poor prognosis. Attempts to improve the therapeutic index of allogeneic transplantation in solid tumours by innovative clinical strategies are underway.
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In order to highlight changes in the clinical indications for transplant in solid tumours in adult population, we will concentrate on three issues: germ cell tumours, breast cancer, and allogeneic transplant.
2. Indications for transplant 2.1. Germ cell tumours In 2006, autologous transplantation for germ cell tumours was considered a standard therapy for sensitive relapse, and a clinical option for refractory disease, according to the EBMT indications (4). An authoritative confirmation of these guidelines came in 2007 from the publication of Einhorn et al. in the New England Journal of Medicine (5). They conducted a retrospective review of 184 consecutive patients with metastatic testicular cancer who had progressed after receiving cisplatin-containing combination chemotherapy. One hundred seventy-three patients received two consecutive courses of HDCT consisting of 700 mg of carboplatin/m2 and 750 mg of etoposide/m2 each for 3 consecutive days, and each course followed by an infusion of autologous peripheral-blood haematopoietic stem cells; the other 11 patients received a single course of this treatment. Of the 184 patients, 116 had complete remission of disease without relapse during a median follow-up of 48 months (range, 14 to 118). Of the 135 patients who received the treatment as second-line therapy, 94 were disease-free during follow-up; 22 of 49 patients who received treatment as third-line or later therapy were disease-free. Of 40 patients with cancer that was refractory to standard-dose platinum, 18 were disease-free. A total of 98 of 144 patients who had platinum-sensitive disease were disease-free, and 26 of 35 patients with seminoma and 90 of 149 patients with non-seminomatous germ-cell tumours were disease-free. It is clear from these data that testicular tumours are potentially curable by means of HDCT and haematopoietic stem-cell rescue, even when this regimen is used as third-line or later therapy or in patients with platinum-refractory disease. The role of HDCT as first-line treatment in patients with metastatic germ cell tumour (GCT) and poor-prognostic clinical features was investigated by Motzer et al. (6). They randomised 219 previously untreated patients with intermediate- or poor-risk GCT to either four cycles of standard bleomycin, etoposide and cisplatin (BEP alone), or two cycles of BEP followed by two cycles of HDCT containing carboplatin and autologous transplant. The 1-year durable complete response rate was 52% after BEP-HDCT and 48% after BEP alone (p=0.53), leading to the conclusion that the routine inclusion of HDCT in first-line treatment for GCT patients with metastases and a poor predicted outcome to chemotherapy did not improve treatment outcome. 466
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2.2. Breast cancer According to the 2006 EBMT indications, autologous transplantation for breast cancer in the adjuvant and responding metastatic setting is in a developmental phase. The role of adjuvant HDCT with autologous transplantation for primary breast cancer at high risk of recurrence (at least 4 involved axillary lymph nodes) remains ill-defined. Data from individual trials have limited power to show overall or subgroup benefit for this indication. For this reason, the MD Anderson Cancer Center and the EBMT STWP set up a single database of individual patient data from 15 known randomised trials of HDCT vs. standard-dose chemotherapy (SDCT). Disease-free survival (DFS), breast cancer-specific survival (BCSS), and overall survival (OS) were the endpoints of the analysis. Subgroup analyses were by age, hormone receptors (HmR) and menopausal (MP) status. Median follow-up for 6,210 patients (3,118 HDC, 3,092 SDC) was 6 years (range, 0–15.3). Data were presented at the 2007 SABCS meeting, and will be presented at the 2008 EBMT meeting. After adjusting for age, trial and MP status, HDCT was found to prolong DFS, but not BCSS or OS. After adjusting for HmR in the subset for which that information was available, HDC was found to prolong DFS and had modest but significant benefits on BCSS and OS (HR 0.89; 95%CI 0.81–0.98; p=0.016) compared to SDC. In conclusion, HDCT as used in these 15 randomised studies prolongs DFS when used as adjuvant therapy in breast cancer. HDCT has at most a modest benefit on BCSS and OS compared to SDC. Whether HDCT has benefit in the context of contemporary taxane-based regimens and targeted therapies is unknown and may be resolved by future clinical trials. 2.3. Allogeneic transplant Allografting is considered a developmental therapy for renal, breast and ovarian cancer, and not recommended for all other solid tumours. Allogeneic HSCT for solid tumours have been registered in the EBMT database, since 1995. The numbers of allogeneic transplants increased to 194 in 2002, concurrently with the publication of favourable reports and the implementation of an EBMT cooperative phase I study of allografting in solid tumours. Subsequently, a steady decrease in the annual numbers of transplants has occurred. Table 1 shows the numbers of allogeneic transplantations registered at EBMT in the period 2005–2007. Reasons for the recent decrease have been: • the introduction in clinical trials of molecularly targeted agents, especially for renal cancer, • the lack of well designed phase II studies, • the high rate of transplant-related mortality due to accrual of rapidly progressing, high tumour burden patients.
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Table 1: Allogeneic transplants for solid tumours, 2005-2007 (data reported to the EBMT) Tumour type
Number
Osteosarcoma
5
PNET
1
Ewing sarcoma
6
Medulloblastoma
5
Neuroblastoma
44
Ewing sarcoma/PNET, skeletal
1
Germ cell tumours
4
Ovarian carcinoma
8
Melanoma
4
Langerhans cell histiocytosis
4
Small cell lung cancer (SCLC)
1
Soft tissue sarcoma
2
Rhabdomyiosarcoma
22
Synovial sarcoma
3
Breast carcinoma
22
Undifferentiated carcinoma
2
Thymoma
1
Gastric carcinoma
1
Metastatic, unknown origin
1
Wilms tumour
3
Renal cell carcinoma
35
Other
17
TOTAL
192
A considerable amount of clinical data has been accumulated in several tumours, particularly in RCC (335 transplants), breast cancer (143 transplants), neuroblastoma (70 transplants), Ewing’s sarcoma (39), ovarian carcinoma (40), colon carcinoma (39), and other entities. The identification of the major mechanisms of neoplastic transformation has introduced into the clinic a series of innovative drugs that inhibit the molecular targets relevant to tumour phenotype. Many of these compounds have already been 468
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approved for diseases that are targets of allogeneic transplantation (e.g., renal cancer, breast cancer, colorectal cancer). Major advances have been made in advanced renal cancer, in which the inhibition of the EGF/RAS/RAF/MAP kinase pathway has resulted in the doubling of progression-free survival in previously treated patients (7), and blockade of AKT/PI3K/mTOR pathway has resulted in a survival advantage in patients with poor prognostic factors (8). Inhibition of VEGF or VEGF-initiated signalling by bevacizumab or by sunitinib has also demonstrated considerable anti-tumour activity (9, 10). Targeted therapy has distinct advantages over a complex treatment such as allogeneic transplantation: It can be conveniently administered on an outpatient basis, it can be withheld if side effects occur, it does not require the supportive care needed for allogeneic transplantation. Indeed, patient referral for allogeneic transplantation has considerably decreased over the last two years, following the introduction in clinical trials of these drugs. However, targeted therapy is not devoid of side effects, it is not active in a considerable fraction of patients, and, most importantly, complete responses occur rarely, if not at all. In spite of its relevant associated mortality, allogeneic transplantation has the potential of inducing complete responses, some of which are long-lasting.
3. Concluding remarks Relapsed germ cell tumours remain an indication for HDCT with autologous transplantation; poor-risk disease does not seem to benefit from HDCT upfront. Adjuvant breast cancer should be the focus of further randomised investigation, considering that all published studies have not included new drugs (e.g., taxanes, HER2/neu inhibitors) in the treatment program. At this stage, allogeneic transplantation should only be considered in patients without conventional therapeutic options. Further research is needed to translate the graft versus tumour effect into meaningful clinical benefit. Experimental and preliminary clinical data indicate that targeted therapies may powerfully complement the immune effect of allogeneic transplantation, rather than replace it.
References 1. Berry DA, Ueno NT, Johnson MM, et al. High-Dose Chemotherapy with Autologous StemCell Support versus Standard-Dose Chemotherapy for High-Risk Breast Cancer: MetaAnalysis of Individual Patient Data from 15 Randomized Adjuvant Trials. http://www.sabcs.org/ 2. Mobus V, Wandt H, Frickhofen N, et al. Phase III trial of high-dose sequential chemotherapy with peripheral blood stem cell support compared with standard dose chemotherapy for first-line treatment of advanced ovarian cancer: intergroup trial of the AGO-Ovar/AIO and EBMT. J Clin Oncol 2007; 25: 4187-4193.
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3. Leyvraz S. et al., J Natl Cancer Inst, in press, 2008. 4. Ljungman P, Urbano-Ispizua A, Cavazzana-Calvo M, et al. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Definitions and current practice in Europe. Bone Marrow Transplant 2006; 1–11. 5. Einhorn LH, Williams SD, Chamness A, et al. High-dose chemotherapy and stem-cell rescue for metastatic germ-cell tumors. N Engl J Med 2007; 357: 340-348. 6. Motzer RJ, Nichols GJ, Margolin KA, et al. Phase III randomized trial of conventionaldose chemotherapy with or without high-dose chemotherapy and autologous hematopoietic stem-cell rescue as first-line treatment for patients with poor-prognosis metastatic germ cell tumors. J Clin Oncol 2007; 25: 247-256. 7. Escudier B, Eisen T, Stadler WM, et al. Sorafenib in advanced clear-cell renal-cell carcinoma. N Engl J Med 2007; 356: 125-134. 8. Hudes G. Carducci M, Tomczak P, et al. Temsirolimus, interferon alfa, or both for advanced renal-cell carcinoma. N Engl J Med. 2007; 356: 2271-2281. 9. Yang JC, Haworth L, Sherry RM, et al. A randomized trial of bevacizumab, an antivascular endothelial growth factor antibody, for metastatic renal cancer. N Engl J Med 2003; 349: 427-434. 10.Motzer RJ, Hutson TE, Tomczak P, et al. Sunitinib versus interferon alfa in metastatic renalcell carcinoma. N Engl J Med 2007; 356: 115-124.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Autografting is indicated as adjuvant therapy in breast cancer: a) Only with positive hormone receptors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Only in patients below 65-yrs old . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) In patients with BRCA-1/2 positive tumours . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) None of the above . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2. High-dose chemotherapy with autologous transplantation in ovarian cancer: a) Has been utilised only in phase II trials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Has undergone phase III trials of comparison with standard-dose chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Has proven useful in suboptimally debulked patients . . . . . . . . . . . . . . . . . . . . . . d) Is the treatment of choice for optimally debulked patients . . . . . . . . . . . . . . . .
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3. Relapsed non-seminomatous germ-cell tumours may benefit from high-dose chemotherapy: a) Only in first relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Only as consolidation of remission . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) After first or subsequent relapses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Only when the disease is platinum-refractory . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. Transplant-related mortality of allografting in solid tumours is: a) Below 1% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Approximately 10% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Between 2 and 4%. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Above 20% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5. Allogeneic transplantation for solid tumours has been hampered by: a) Competition with targeted agents. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) High transplant-related mortality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The lack of well-designed phase II studies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All of the above . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 32
HSCT for autoimmune diseases in adults
R. Saccardi, D. Farge
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CHAPTER 32 • Autoimmune diseases in adults
1. Introduction Autoimmune diseases (ADs) are a heterogeneous group of diseases, affecting 10–12% of the population. Consensus indications for the use of HSCT to treat severe ADs were published in 1997 (1). Patients should be considered for HSCT when matching the following criteria: 1. Diagnosed with an AD severe enough to have an increased risk of mortality or advanced and irreversible disability; 2. The ADs must be unresponsive to conventional treatments; 3. HSCT should be undertaken before irreversible organ damage so that significant clinical benefit can be achieved. The introduction of new biotherapies has modified the therapeutic panorama since 2002, resulting in a drop of activity in HSCT for inflammatory arthritis. Today, more than 1300 patients worldwide have received an HSCT for an AD alone. Autologous HSCT phase II trials showed that in patients with a favourable outcome, a resetting of a disregulated autoaggressive immune system may occur, rather than simple ablation of auto-reactive cells. Results of allogeneic HSCT are as yet unclear, due to small numbers, and heterogeneous patient groups and treatment regimens. Peripheral blood stem cells (PBSC) have mainly been used, with a the most frequent mobilisation regimen including the combination of cyclophosphamide (Cy) and GCSF. As ADs are extremely heterogeneous, a comparison of protocols and outcome depends on careful stratification of diagnosis and phases of diseases. So far in 2007, 841 HSCT procedures have been registered in the European Group for Blood and Marrow Transplantation (EBMT) database (Table 1), the rest being registered in the US International Bone Marrow Transplantation Registry (IBMTR) and in Asia. In the EBMT database, the most commonly transplanted diseases are multiple sclerosis (MS), scleroderma (SSc), rheumatoid arthritis (RA), juvenile arthritis (JIA) and systemic lupus erythematosus (SLE), coming from over 100 transplant centres in more than 20 countries (Table 2). Prevalence of female sex
Table 1: Overview of data on HSCT for ADs reported to the EBMT database (August 2007) Patients
841
Transplant procedures
863
Centres/countries
171/25
Autografts/allografts
801/40
Age at transplant (yrs)
35
Male/female
313/526
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Table 2: Distribution of diagnoses in the EBMT database Multiple sclerosis
297
Connective tissue disorders - SSc - SLE - PM/DM - Sjogren - Other/unknown
261 147 84 12 3 15
Arthritis - Rheumatoid arthritis - Juvenile chronic arthritis: • Systemic JIA • Other JIA • Polyarticular JIA - Psoriatic arthritis - Other
154 82 38 19 9 3 3
Inflammatory bowel disease - Crohn's disease - Ulcerative colitis
11 9 2
Haematological - ITP - AIHA - Evan’s - Pure red cell - Pure white cell - Other
58 21 12 9 7 2 7
Vasculitis - Behcet’s - Wegener’s - Microscopic polyarteritis nod - Takayasu - Churg-Strauss - Other
29 6 7 3 2 2 9
Other neurological - Myasthenia gravis - Other
16 3 13
Other/missing
15
PM/DM: polymyositis/dermatomyositis; see text for other abbreviations
and young age reflects the natural distribution of the diseases. Long-lasting responses were obtained in all disease categories with an overall adjusted transplantrelated mortality (TRM) being 7±3% at three years, directly related to the type of AID disease (SSc and SLE at higher risk), the year of transplant with a learning curve and the intensity of conditioning (more intensive conditioning had a higher risk of TRM but lower probability of disease progression) (2). In the following paragraphs clinical results and indications for the major ADs will be overviewed. Less common diseases have been reviewed elsewhere (3).
2. Multiple sclerosis MS is the most frequent diagnosis for which HSCT has been used. In 2006, a retrospective analysis of 183 cases from the EBMT database, of which 99 were secondary progressive forms (SP), 19 primary progressive and 41 relapsing forms (RR), was published (4). Overall, 63% of patients did not progress in their disability after a median follow-up of 42 months. TRM was 5%, mostly concentrated prior to the year 2000. More aggressive regimens, including busulfan or a combination of graft manipulation and serotherapy, resulted in a higher toxicity without any advantage in terms of relapse prevention or disability progression. The most widely-used regimen was the association of BEAM and anti-thymocyte globulin, with an 474
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unmanipulated graft, and among the 53 patients who received this regimen, no TRM was reported. Ideally, rapidly progressing patients should be transplanted in the RR phase, before an advanced disability or a SP form has developed. An ongoing update of this analysis, carried out among patients treated with the BEAM/ATG protocol, confirms the better outcome of RR over SP forms. The ASTIMS trial comparing prospectively HSCT vs. mitoxantrone as control arm is currently ongoing, with 15 patients included so far out of the 30 required to complete the protocol.
3. Systemic sclerosis Phase I/II pilot studies showed that HSCT was feasible in carefully selected patients with diffuse SSc allowing an improvement of 25% or more in the skin score in 70% of the first 65 treated patients. However, the early TRM was higher than the 3% reported in non ADs patients and varied according to conditioning regimen, using either Cy alone (200 mg/kg total) or 8 Gy radiation plus Cy 120 mg/kg body weight. The initial TRM was 8.6% in the EBMT extended report, which could be reduced to 5.2% when stricter exclusion criteria were applied (mean pulmonary arterial pressure >50 mmHg, severe cardiac involvement or pulmonary fibrosis and uncontrolled systemic hypertension) (5). More importantly, autologous HSCT induced a major regression of SSc dermal fibrosis, confirmed by histological analysis, which had never been previously reported with any other treatment in SSc. Prolonged followup of patients confirmed sustained improved functional status, fall in skin score and stabilisation of lung function, whereas death from disease progression was strikingly lower compared to the 5-year mortality rate estimated at 30% in such severe SSc patients (6). These results were the basis for the ongoing ASTIS trial comparing HSCT (Cy, ATG and CD34 selected graft) versus monthly intravenous pulse Cy 750 mg/m2 for 12 months, with 103 patients included so far out of the 120 planned.
4. Systemic lupus erythematosus With the aim of suppressing autoimmune disease in one concentrated effort rather than drawn out over a long period, several early European phase I/II trials showed that severe SLE patients refractory to standard therapy responded to HSCT. The first EBMT/EULAR retrospective study of 55 patients, most with either renal and/or CNS involvement, showed that 66% achieved a “remission” with a 12% TRM (due to the selection of patients refractory to all previous therapies) (7). Of those achieving remission after HSCT, 32% subsequently relapsed to some degree, but were then mostly easily controlled on standard agents which were previously ineffective. The safety of HSCT is likely to be improved by better patient selection according to the consensus criteria and better choice of conditioning regimen. In a US single centre
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study, 50 patients refractory to standard therapies with either organ- or lifethreatening visceral involvement also showed a durable remission after HSCT with a 4% TRM (8). Based on these experiences, the ASTIL phase IIb trial was designed to compare the efficacy of autologous HSCT versus rituximab (anti-CD20) as induction therapy followed by maintenance therapy by mycophenolate mofetil in both arms in the treatment of severe SLE refractory to standard therapy.
5. Arthritis 5.1. Rheumatoid arthritis Analysis of the first 78 EBMT patients showed significant improvement in 67%, whereas most had failed conventional disease modifying anti-rheumatic drugs (DMARDs) before HSCT. The conditioning regimen in the majority was Cy 200 mg/m2 alone after either G-CSF or Cy/G-CSF mobilisation. Only one TRM from sepsis was reported five months after higher intensity conditioning (busulfan/Cy). Some degree of relapse was seen in 73% of patients, but was relatively easy to control with drugs which had been ineffective pre-transplant. In the past ten years, TNF blockers have emerged as safe and highly effective treatments for resistant RA, although 25% of treatment failures remain. The use of autologous HSCT as a salvage therapy is now limited to these rare patients who will also be candidates for other new biotherapies. 5.2. Phase I/II juvenile idiopathic arthritis The use of anti-TNF and anti-IL-6 receptor treatments for DMARD-resistant JIA has proven of great value, but some children still remain resistant with severe morbidity, impaired quality of life and increased mortality rate. In the EBMT database 66 children, most of whom had with Still’s disease, were treated by autologous HSCT using stem cells obtained from the bone marrow and a conditioning protocol of Cy 200 mg/kg body weight, TBI 4Gy and ATG. Impressive results showed a prolonged drug-free follow-up of 6–60 months. In a report of 34 patients (9), 18 entered complete and 6 partial drug-free remissions, for whom the corticosteroid dose could be reduced or stopped with subsequent restoration of growth. Four patients died, 3 from the macrophage activation syndrome, a well-known complication of systemic JIA, thought to be related to intercurrent infection or uncontrolled systemic activity of the disease at the time of transplantation. Since then protocols have been modified, to control systemic activity before HSCT with methyl-prednisolone, and no further such deaths have occurred. The results of phase I/II trials in JIA using Cy alone versus Cy and TBI suggested no advantage of the TBI. 476
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6. Crohn’s disease Several cases with satisfactory results have been reported, and guidelines for considering HSCT in Crohn’s disease have been published. In Europe, the ongoing phase IIb ASTIC trial (Table 3) is evaluating the potential clinical benefit of haematopoietic stem cell mobilisation followed by immediate versus delayed HSCT in patients with relapsing Crohn’s disease and clear intolerance or toxicity to a conventional treatment protocol.
Table 3: EBMT ongoing prospective trials for ADs Disease
Trial Design
Acronym
Status
Website
SSc
Autologous HSCT vs. monthly Cy
ASTIS
Started 2002 103/120
www.astistrial.com
MS
Autologous HSCT vs. monthly mitoxantrone
ASTIMS
Started 2004 15/30
www.astims.org
Crohn’s
Mobilisation followed by early vs. late autologous HSCT
ASTIC
Launch Q1 2007
www.astic.eu
SLE
Autologous HSCT vs. anti-CD20 MoAb
ASTIL
Planning Q1 2008
pending
Numbers in status box indicate patients enrolled/patients planned
7. Haematological cytopenias In this group of patients, a relatively higher proportion of allogeneic transplantation was reported, showing encouraging results in terms of relapse-free outcome. Most of the patients who received an autologous HSCT were treated with a Cy-based conditioning regimen, showing a sustained response in 33% of the cases reported to the EBMT database (10).
8. Conclusions The main indications for HSCT among ADs are MS and SSc, in which a significant subset of patients still shows an unsatisfactory response to both conventional and new immunomodulating treatments. Initial experience has been used to design several ongoing prospective, phase IIb-III randomised trials both in Europe and United States, to compare HSCT with conventional, approved treatment. Inclusion of patients in such institutional trials is encouraged.
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Acknowledgments The Authors are gratefully indebted to Virginie Chesnel for her helpful assistance in the data management.
References 1. Tyndall A, Gratwohl. A Blood and marrow stem cell transplants in auto-immune disease: A consensus report written on behalf of the European League against Rheumatism (EULAR) and the European Group for Blood and Marrow Transplantation (EBMT). Bone Marrow Transplant 1997; 19: 643-645. 2. Gratwohl A, Passweg J, Bocelli-Tyndall C, et al. Autologous haematopoietic stem cell transplantation for autoimmune diseases. Bone Marrow Transplant 2005; 35: 869-895. 3. Kapoor S, Wilson AG, Sharrack B, et Al. Haemopoietic stem cell transplantation – An evolving treatment for severe autoimmune and inflammatory diseases in rheumatology, neurology and gastroenterology. Hematology 2007; 12: 179-191. 4. Saccardi R, Kozac T, Bocelli-Tyndall C, et al. Autologous stem cell transplantation for progressive multiple sclerosis: Update of the European Group for Blood and Marrow Transplantation autoimmune diseases working party database. Mult Scler 2006; 12: 814-823. 5. Farge D, Passweg J, van Laar JM, et al. Autologous stem cell transplantation in the treatment of systemic sclerosis: Report from the EBMT/EULAR Registry. Ann Rheum Dis 2004; 63: 974-981. 6. Vonk MC, Marjanovic Z, van den Hoogen FH, et al. Long-term follow-up results after autologous haematopoietic stem cell transplantation for severe systemic sclerosis. Ann Rheum Dis, Epub 2007 May 25: 98-104. 7. Jayne D, Passweg J, Marmont A, Farge D, et al. Autologous stem cell transplantation for systemic lupus erythematosus. Lupus 2004; 13: 168-176. 8. Burt RK, Traynor A, Statkute L, et al. Nonmyeloablative hematopoietic stem cell transplantation for systemic lupus erythematosus. JAMA 2006; 295: 559-560. 9. Wulffraat NM, Sanders EA, Kamphuis SS, et al. Prolonged remission without treatment after autologous stem cell transplantation for refractory childhood systemic lupus erythematosus. Arthritis Rheum 2001; 44: 728-731. 10.Passweg JR, Rabusin M, Musso M, et al. Haematopoetic stem cell transplantation for refractory autoimmune cytopenia. Br J Haematol 2004; 125: 749-755.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. How many patients have received HSCT for an autoimmune disease in the past ten years? a) 100–500 patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 478
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b) 500–1000 patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 1000–3000 patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) More than 3000 patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which are the main indications for HSCT among ADs? a) Autoimmune cytopenia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Rheumatoid arthritis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Systemic lupus erythematosus and vasculitis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Systemic sclerosis and multiple sclerosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which is the current transplant related mortality in autologous SCT for AD? a) < 1% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 3–10% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 10–20% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 20–30% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which of the following criteria is not an indication for HSCT? a) Patients diagnosed with an AD severe enough to have an increased risk of mortality or advanced and irreversible disability . . . . . . . . . . . . . . . . . . . b) ADs not responsive to conventional treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Young patients at diagnosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Advanced disease without irreversible severe organ damage . . . . . . . . . . . . . . 5. Which the prevalent source of HSCs for transplantation in ADs? a) PBSCs mobilised by cyclophosphamide and G-CSF . . . . . . . . . . . . . . . . . . . . . . . . . . b) PBSCs mobilised by G-CSF alone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Bone marrow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Cord blood . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 33
HSCT for aplastic anaemia in adults
J. Passweg , A. Bacigalupo, A. Locasciulli for the EBMT Aplastic Anaemia Working Party
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CHAPTER 33 • AA in adults
1. Introduction Acquired aplastic anaemia (AA) is a life-threatening disorder characterised by bicytopenia or pancytopenia in the peripheral blood and an aplastic/hypoplastic bone marrow. AA is considered severe if marrow production is inadequate in at least 2 cell lines and as very severe if the neutrophil count is <0.2 x 109/L, because of the high risk of infection. The patho-physiology of AA is heterogeneous. Features include a reduced pool of haematopoietic progenitor cells, accelerated apoptosis in haematopoietic progenitor cells, immunemediated suppression of haematopoiesis, clonal abnormalities and telomere shortening in progenitor cells (1, 2). Differential diagnosis includes congenital bone marrow failure syndromes, some of which may manifest late, i.e. in early adulthood, and hypoplastic MDS. It is often wise to repeat the diagnostic marrow studies to exclude aplastic crisis due to a drug or an infection. Clonal aberrations on cytogenetic studies and dysplastic erythropoietic islands do not exclude SAA, but marrow fibrosis, the presence of blast cells, and dysplastic myelo- or megakaryopoiesis are arguments in favour of a hypoplastic MDS. Once the diagnosis is established treatment is by bone marrow transplantation (BMT) or immunosuppressive therapy (IS). In HSCT for AA the use of bone marrow rather than PB is recommended.
2. Treatment Treatment goals are to improve peripheral blood counts to achieve transfusion independence and to avoid infection risks. The outcome of SAA patients has improved considerably over time both after BMT and IS. The 5-year survival for patients reported to the Registry of the EBMT SAA Working Party is 70–80% both after immunosuppressive treatment and after HLA-identical sibling BMT (Figure 1). The choice of the primary treatment is based on availability of an HLA-identical sibling, patient age (Figure 2) and disease severity. The EBMT compared outcome after BMT and IS using a stratified proportional hazard model to estimate effects of age and neutrophil count on failure-free survival in both treatment groups (3). Table 1 presents the estimated differences in 5-year failure-free survival between the 2 initial treatment options as a function of age and neutrophil count. Negative values indicate a survival disadvantage of BMT compared with IS, while positive values indicate a survival advantage for BMT. This allows the identification of 3 groups of patients: 1. Patients in whom BMT is superior, including children regardless of neutrophil count and adults up to the age of 40 with low neutrophil counts (£ 0.3 x 109/L); 2. Patients in whom IS is superior, comprising adults above age 40; 3. Patients in whom no differences were found (3). The difference in projected survival between patients treated with BMT and IS is
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Table 1: Differences between BMT and IS in 5-year failure free survival (%) after initial treatment (3) Neutrophil count (x 109/L)
Age (Y)
0 0.1 0.2 0.3 0.4 0.5
10
20
30
40
50
24* 19 14 10 6 3
20 14 9 5 1 -2
14 8 3 -1 -4 -7
6 1 -4 -7 -10 -12
-2 -7 -11 -14 -16 -17**
Positive values: Advantage for BMT (*24% 5-yr failure free survival difference in favour of BMT) Negative values: Advantage for immunosuppression (**17% difference in favour of immunosuppression)
Figure 1: Survival after HLA-identical stem cell transplantation (1a) or immunosuppression (1b) as first line treatment stratified by year of treatment Survival Functions 1.0
1.0 0.8 Cum Survival
>2000
>2000
1996-2000 0.8 1990-1995 1986-1989 0.6 1974-1985
0.6
1996-2000 1990-1995 1986-1989 1974-1985
0.4
0.4
0.2
0.2
0.0
0
1
2
3
4
5 6 Years
7
8
9
10
0.0
0
1
2
3
4
5 6 Years
7
8
9
10
not linear but increases with time in favour of BMT. This may be explained by more early deaths with BMT, mainly due to GvHD and more late events (clonal complications and relapse) in patients treated with IS. This view has been recently challenged by studies indicating that children even with very severe disease may have excellent outcome when treated with IS, even better than patients without very severe disease (4). This may be due to improved supportive care with fewer infectious deaths possibly supporting patients long enough to benefit from treatment. 482
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Figure 2: Patients receiving BMT by age, 0–10 years, 10–20, 20–30, 30–40, 40–50 and older than 50 with youngest patients having best survival and oldest patients having poorest survival 1.0 <10 10–20 20–30 30–40 40–50 >50
Cum Survival
0.8 0.6 0.4 0.2 0.0
0
1
2
3
4
5 6 Years
7
8
9
10
3. Results: Stem cell transplantation 3.1. Conditioning for HLA-identical sibling transplantation The standard preparative regimen in HLA-identical sibling transplantation for nonsensitised patients is cyclophosphamide at a dose of 200 mg/kg b.w. (4x50 mg/kg on days -5 to -2) with or without anti-thymocyte globulin (ATG) (5). In a non-randomised trial a combination of cyclophosphamide + ATG resulted in lower incidence of chronic GvHD and improved survival compared with historical controls who received cyclophosphamide alone (6, 7). However, a prospective randomised trial in 131 patients did not detect a significant benefit from the addition of ATG to cyclophosphamide as a preparative regimen for patients with severe AA (8). Irradiation-based conditioning is generally not recommended for sibling donor transplantation. While irradiation-based regimens have been effective in reducing rejection, they have accomplished this goal at the price of increased transplantrelated complications (9–11). There are however interesting data in unrelated donor transplantation with the addition of 2Gy of TBI to reduce the graft failure rate (12). 3.2. Stem cell source The use of peripheral blood stem cells (PBSC) as a stem cell source alternative to BM for allogeneic transplantation is increasing for all indications. A number of studies have shown that PBSC is associated with higher risks of chronic GvHD and in patients with advanced malignancy in an increased graft versus leukaemia effect.
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In a joint EBMT/IBMTR retrospective analysis results of 151 HLA-identical sibling PB HSCT were compared with results of 722 HLA-identical sibling BMT for AA. Other than faster haematopoietic recovery, this study shows a higher incidence of chronic GvHD and lower survival for HSCT with PB, especially in young patients (13). Currently half the transplants reported to the EBMT for SAA use peripheral blood as a stem cell source in spite of the recommendation to use marrow as a stem cell source. 3.3. Post-transplant immunosuppression in HLA-identical sibling transplantation The introduction of cyclosporin A (CsA) in the 1980s resulted in decreased transplantrelated mortality, significantly reduced rejection rates and improved survival (14). In a prospective randomised trial comparing CsA + methotrexate (MTX) with CsA alone, the 1-y TRM rates for patients given CsA/MTX or CsA alone were 3 and 15%, respectively (15). The 5-year probability of survival was 94% in the CsA/MTX group and 78% for those in the CsA alone group. Even though these differences appear large if attributed to the addition of MTX alone, the current recommendation is to use CsA + MTX as post-transplant immunosuppression. 3.4. Alternative donor transplantation In the past, survival after alternative donor transplantation has been poor (16). Therefore it was only considered as salvage therapy for patients failing to respond to one or more courses of IS. The optimum conditioning therapy and post-transplant immunosuppression for alternative donor BMT in AA has yet to be established. Approaches currently under investigation are addition of fludarabine or radiotherapy and ATG to the standard combinations. Promising data are reported from a GITMO/EBMT AA WP trial with a survival probability of 82% in AA patients conditioned with a combination of low-dose cyclophosphamide, ATG, fludarabine and CsA + MTX as GvHD prophylaxis (17). It is also important to consider risk factors for the alternative donor BMT in AA: a fully matched donor for both Class I and Class II by high-resolution typing and early transplantation provides better results. A recent study by the EBMT using observational data shows improvement in outcome with long-term survival for patients receiving unrelated donor transplants from 32% (+ 8%) before 1998 to 57% (+ 8%), p<0.0001 in those transplanted after 1998. This improvement was associated with less graft failure, less acute and chronic GvHD and fewer infectious deaths, probably due to better donor selection and possibly progress in antimicrobial treatment (18). There is very limited information on the use of haploidentical donors and the series using cord blood as a stem cell source are too small at this time for meaningful comparisons. 484
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4. Results: Immunosuppressive treatment A combination of ATG, CsA and corticosteroids represents the current standard as first-line immunosuppressive therapy both for severe and non-severe AA (19–21). Response rates with this regimen are in the order of 65 to 75% (at 4–6 months). There is little agreement on the duration of immunosuppressive treatment and some experts recommend withdrawing CsA after 6 months whereas others treat for a longer period. Relapse in responding patients is not uncommon (30–40%), but can be retreated effectively in the majority of patients. Patients not responding to 2 cycles of ATG are unlikely to respond to additional treatment. Late complications include PNH and MDS. MDS is seen in 5–10% of patients after IS; whether incidence is linked to growth factor administration is unknown (22). Data supporting such an association in observational studies need confirmation by prospective trials. Pilot studies of G-CSF in combination with ATG + CsA have reported encouraging results with respect to early mortality, trilineage response and survival. A randomised trial confirmed improved neutrophil recovery but no advantage in terms of overall response and survival by addition of IS (23). A large EBMT multicentre randomised trial comparing ATG + CsA versus ATG + CsA + G-CSF has recently been completed and the results are awaited. The most commonly used ATG preparation in Europe, equine ATG (Lymphoglobulin), has recently been withdrawn from the market. Whether the rabbit product by the same manufacturer (Thymoglobulin), using a similar manufacturing procedure (stimulation with human thymocytes) is equivalent remains to be determined. A recent study by a Chinese group comparing different types of ATG with or without CsA and growth factors showed that not all ATG preparations are equivalent (24). There is little experience with other immunosuppressive agents. Studies are ongoing using Campath in patients refractory to ATG + CsA. The EBMT collects data not only on transplant patients but also on nontransplant treatments. This has allowed for multiple studies comparing outcomes with different treatment strategies (Table 1). Long-term survival with immunosuppressive treatment has continuously improved over the last years (Figure 1). This improvement is most likely due to improved supportive care, as the main drugs used to treat the disease (ATG and CsA) have not changed over the years.
5. Discussion and future perspectives Treatment of SAA has greatly improved the outcome of these patients over the past 30 years. Unresolved issues are: chronic GvHD in transplant recipients, transplantation for older patients not responding to immunosuppressive treatment, transplantation using alternative donors, the use of growth factors with immunosuppressive treatment and patients refractory to immunosuppressive treatment without an adequate donor.
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References 1. 2. 3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
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Schrezenmeier H, Bacigalupo A. Aplastic Anemia - Pathophysiology and Treatment. Cambridge: Cambridge University Press; 2000. Young N, Calado RT, Scheinberg P. Current concepts in the pathophysiology and treatment of aplastic anemia. Blood 2006; 108: 2509-2519. Bacigalupo A, Brand R, Oneto R, et al. Treatment of acquired severe aplastic anemia: Bone marrow transplantation compared with immunosuppressive therapy – The European Group for Blood and Marrow Transplantation experience. Semin Hematol 2000; 37: 69-80. Führer M, Rampf U, Baumann I, et al. Immunosuppressive therapy for aplastic anemia in children: A more severe disease predicts better survival. Blood 2005; 106: 21022104. Schrezenmeier H. Guidelines for treating aplastic anemia: Consensus document of a group of international experts. In: Schrezenmeier H, Bacigalupo A, eds. Aplastic Anemia: Patho-physiology and Treatment. Cambridge: Cambridge University Press; 2000: 308-315. Storb R, Etzioni R, Anasetti C, et al. Cyclophosphamide combined with antithymocyte globulin in preparation for allogeneic marrow transplants in patients with aplastic anemia. Blood 1994; 84: 941-949. Storb R, Leisenring W, Anasetti C, et al. Long-term follow-up of allogeneic marrow transplants in patients with aplastic anemia conditioned by cyclophosphamide combined with antithymocyte globulin (letter). Blood 1997; 89: 3890-3891. Champlin RE, Perez WS, Passweg JR, et al. Bone marrow transplantation for severe aplastic anemia: A randomized controlled study of conditioning regimens Blood 2007; 109: 45824585. McCann SR, Bacigalupo A, Gluckman E, et al. Graft rejection and second bone marrow transplants for acquired aplastic anaemia: A report from the Aplastic Anaemia Working Party of the European Bone Marrow Transplant Group. Bone Marrow Transplant 1994; 13: 233-237. Gluckman E, Horowitz MM, Champlin RE, et al. Bone marrow transplantation for severe aplastic anemia: Influence of conditioning and graft-versus-host disease prophylaxis regimens on outcome. Blood 1992; 79: 269-275. Deeg HJ, Socie G, Schoch G, et al. Malignancies after marrow transplantation for aplastic anemia and Fanconi anemia: A joint Seattle and Paris analysis of results in 700 patients. Blood 1996; 87: 386-392. Deeg HJ, O’Donnell M, Tolar J, et al. Optimization of Conditioning for Marrow Transplantation from Unrelated Donors for Patients with Aplastic Anemia After Failure of Immunosuppressive Therapy. Blood 2006; 108: 1485-1491. Schrezenmeier H, Passweg JR, Marsh JCW, et al. Worse outcome and more chronic GVHD with peripheral blood progenitor cells than bone marrow in HLA-matched sibling donor transplants for young patients with severe acquired aplastic anemia. Blood 2007; 110: 1397-1400.
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14. Passweg JR, Socie G, Hinterberger W, et al. Bone marrow transplantation for severe aplastic anemia: Has outcome improved? Blood 1997; 90: 858-864. 15. Locatelli F, Bruno B, Zecca M, et al. Cyclosporin A and short-term methotrexate versus cyclosporin A as graft versus host disease prophylaxis in patients with severe aplastic anemia given allogeneic bone marrow transplantation from an HLA-identical sibling: Results of a GITMO/EBMT randomized trial. Blood 2000; 96: 1690-1697. 16. Hows J, Stone JV, Camitta BM. Alternative donor transplantation for severe acquired aplastic anemia. In: Schrezenmeier H, Bacigalupo A, eds. Aplastic Anemia: Pathophysiology and Treatment. Cambridge: Cambridge University Press; 2000: 258-274. 17. Bacigalupo A, Locatelli F, Lanino E, et al. for the Severe Aplastic Anemia Working Party of the European Group for Blood and Marrow Transplantation (SAA WP-EBMT). Fludarabine, cyclophosphamide and anti-thymocyte globulin for alternative donor transplants in acquired severe aplastic anemia: A report from the EBMT-SAA Working Party. Bone Marrow Transplant 2005; 36: 947–950. 18. Viollier R, Socie G, Tichelli A, et al. for the Working Party on Severe Aplastic Anemia (WPSAA) of the European Group for Blood and Marrow Transplantation (EBMT). Recent improvement in outcome of unrelated donor transplantation for aplastic anemia. Bone Marrow Transplant 2007; epub ahead of print. 19. Marsh J, Schrezenmeier H, Marin P, et al. Prospective randomized multicenter study comparing cyclosporin alone versus the combination of antithymocyte globulin and cyclosporin for treatment of patients with nonsevere aplastic anemia: A report from the European Blood and Marrow Transplant (EBMT) Severe Aplastic Anaemia Working Party. Blood 1999; 93: 2191-2195. 20. Frickhofen N, Heimpel H, Kaltwasser JP, Schrezenmeier H. Antithymocyte globulin with or without cyclosporin A: 11-year follow-up of a randomized trial comparing treatments of aplastic anemia. Blood 2003; 101: 1236-1242. 21. Bacigalupo A, Bruno B, Saracco P, et al. Antilymphocyte globulin, cyclosporine, prednisolone, and granulocyte colony-stimulating factor for severe aplastic anemia: An update of the GITMO/EBMT study on 100 patients. European Group for Blood and Marrow Transplantation (EBMT) Working Party on Severe Aplastic Anemia and the Gruppo Italiano Trapianti di Midollo Osseo (GITMO). Blood 2000; 95: 1931-1934. 22. Socie G, Mary JY, Schrezenmeier H, et al. Granulocyte-stimulating factor and severe aplastic anemia: A survey by the European Group for Blood and Marrow Transplantation (EBMT). Blood 2007; 109: 2794-2796. 23. Gluckman E, Rokicka-Milewska R, Hann I, et al. Results and follow-up of a phase III randomized study of recombinant human-granulocyte stimulating factor as support for im-munosuppressive therapy in patients with severe aplastic anaemia. Br J Haematol 2002; 119: 1075-1082. 24. Yizhou Zheng, Yongze Liu, Yulin Chu. Immunosuppressive therapy for acquired se-vere aplastic anemia (SAA): A prospective comparison of four different regimens. Experimental Hematology 2006; 34: 826–831.
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Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The best standard immunosuppressive treatment for severe aplastic anaemia is: a) Anti-lymphocyte globulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Anti-thymocyte globulin + mycophenolate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Anti-thymocyte globulin + cyclosporin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) High dose cyclophosphamide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which characteristic is frequently found in aplastic anaemia? a) Splenomegaly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Marrow fibrosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) A positive PNH test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Increased marrow blasts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. The main factor(s) affecting mortality when it comes to decide between transplant or immunosuppression as a first line treatment is/are: a) Donor CMV status and platelet transfusion needs . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Patient age and neutrophil count . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Donor sex and a positive PNH test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Availability of an unrelated donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which of the following statements is true in aplastic anaemia? a) Peripheral blood is the best stem cell source because of faster neutrophil engraftment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Marrow is the best stem cell source because of it causes less chronic GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Peripheral blood is the best stem cell source because it is easier to harvest from the donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Marrow is the best stem cell source because stromal cells accelerate engraftment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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5. To transplant a patient with aplastic anaemia from a sibling donor the best established conditioning regimen is: a) i.v. busulfan and cyclophosphamide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 12 Gy of total body irradiation and cyclophosphamide . . . . . . . . . . . . . . . . . . . . . c) Cyclophosphamide and anti-thymocyte globulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Melphalan and fludarabine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 34
HSCT for tissue regeneration in adults
F. Prosper
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1. Introduction The development of pluripotent human embryonic stem cells (1) and the isolation of different types of stem cells derived from adult tissues (somatic and germinal) have raised the possibility of utilising these cells for non-haematological diseases. Among the different tissue-specific stem cells, some adult-derived stem cells, such as haematopoietic cells, have been endowed with a remarkable plasticity, suggesting that they may be able to contribute to regeneration of multiple tissues. An intense controversy has emerged as to whether observations supporting the plasticity of adult stem cells represent true pluripotentiality or may be explained by other mechanisms such as cell fusion, reprogramming or just a misinterpretation of certain experiments or even an excess of enthusiasm. Although from a biological perspective these are legitimate issues, the most topical question currently is whether these stem cells can be exploited clinically for non-haematological diseases in a similar way to the haematological disorders. The importance of understanding the mechanism underlying the potential benefit of stem cells for tissue regeneration remains critical.
2. Bone marrow derived stem cells The bone marrow is a remarkable reservoir of adult stem cells. Beside HSC and the side population (SP) cells, other non-haematopoietic populations of stem cells have been isolated from the bone marrow including mesenchymal stem cells (MSC) (also known as marrow stromal cells), multipotent progenitor cells (MAPC), multipotent stem cells and pre-MSC, MIAMI cells (Marrow-Isolated Adult Multilineage Inducible cells), VSEL cells (Very Small Embryonic Like stem cells) or endothelial progenitor cells (EPCs). The main characteristics of these different populations of BM-derived cells are shown in Table 1. Whether all these populations truly represent completely different types of cells or whether their differences are due to the techniques of isolation and characterisation is unclear. The potential of these cells is also under intense debate. Only HSC and MSC have been exploited in clinical trials of regenerative therapy for non-haematological diseases, while other cell populations have been studied in different experimental settings.
3. Cell therapy with stem cells from the bone marrow Multipotent adult stem cells obtained from human BM are just starting to be explored in numerous diseases and for different tissues. This Chapter will discuss those areas in which stem cell therapy has advanced the furthest. Overall, evidence for efficacy is limited to a few applications including epidermal layers, transplantation of limbo-corneal stem cells or chondrocytes for cartilage lesions. In most diseases only preclinical studies and initial phase I/II studies have been completed so that a large effort is still needed before cell therapy becomes a reality.
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Table 1: Bone marrow derived stem cells Name
Species
Tissue
Phenotype
MAPC
Human Mouse, rat, swine
BM, Brain Muscle
MIAMI
Human
VSEL
Mouse Human
ESC genes
Meso
Endo
Ecto
CD44 low, MHC-I low, Oct4+ CD45 -, Thy1-, cKit+ Nanog-
+
+
+
BM
CD44 Pos; MHC-I Neg, Oct4+ CD45 Neg Nanog?
+
+
+
BM UCB
CD45-, Lin-, Sca1 +, CXCR4 +, SSEA1+
Oct4+ Nanog+
+
+
+
Pre-MSC Mouse
BM
SSEA1+, CD45-, Ter119- , Sca1 Dim
Oct4+ Nanog+
+
+
+
MASC
Human
BM, Liver, Heart
CD45-
Oct4+ Nanog+
+
+
+
MSC
Human Mouse, rat, swine
BM, Fat UCB
CD44+, MHC-I+, CD45-, Thy1-, cKit+
Oct4Nanog-
+
+?
+?
3.1. Stem cells for cardiovascular repair The idea of repairing the diseased heart using different types of stem cells has gained widespread attention recently, leading not only to a number of preclinical studies but also to early phase I/II and even randomised clinical trials (2). In contrast to ESC, there is no convincing evidence that cells from postnatal tissues other than the heart can generate functional cardiomyocytes in vivo. Nevertheless, a number of cell populations, including total BMMNC, enriched haematopoietic stem cells, EPCs, BM, MSC and adipose MSC and skeletal myoblasts have been grafted in models of chronic as well as acute myocardial infarction (MI) (3). Improved function has been seen in a number of studies without evidence that the cells contributed to cardiac muscle itself. As a consequence of these preclinical studies, clinical trials in patients with acute and chronic myocardial ischaemia using mostly BM cells (4) (freshly isolated total BM, mononuclear cells, cultured cells to increase putative endothelial progenitor cells, CD34 or AC133 selected cells) have been developed. The largest experience has been gained in patients with acute MI using BM-MNC and while some randomised trials suggest a benefit others find no effect (5). The differences have been attributed to the heterogeneity of the types of cells used in each trial, the type of patients and/or the endpoints. BM-MNC include an heterogeneous pool of cells such as stem cells (at very low frequency), progenitor cells and mature cells. Transplantation of either pure selected 492
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populations or unselected populations each have a rationale and with the current level of knowledge both approaches are justified. Although there is much to learn in this field, some lessons are emerging. The classic idea that delivery of the appropriate cells would repair a damaged heart via active myocardial regeneration resulting from trans-differentiation of the administered cells has been superseded by the recognition of alternative mechanisms of action: - exogenous cells may stimulate proliferation of endogenous cardiac precursors or stem cells through neovascularisation or paracrine signalling actions facilitating the ability of the heart to heal itself; - exogenous cells may lead to cardiac repair via fusion of donor cells with host cardiomyocytes as has been demonstrated in animal models; - the effect of cells could also be mediated by altering the mechanical properties of the scar thereby preventing deterioration in cardiac function (5). Peripheral vascular disease (PVD) is another potential target for regenerative medicine. Approximately 15% of adults over the age of 55 have detectable haemodynamic impairments with intermittent claudication and critical limb ischaemia being the two major clinical presentations. Experimental studies indicate that infusion of progenitor cells from different sources such as peripheral blood or BM may improve recovery after limb ischaemia. Based on these experimental findings, clinical phase I trials were initiated in 2001 (6). These initial pilot trials indicated that infusion bone marrow-derived or circulating blood-derived progenitor cells improves the blood supply to the legs in patients with ischaemia resulting in longer pain-free walking distances (7). These initial studies need to be confirmed on a larger scale and with clear endpoints. 3.2. Orthopaedic applications of stem cells On the basis of the in vitro observation that MSCs can differentiate into osteocytes and chondrocytes, BM-derived MSCs were seeded onto extracellular matrices such as hydroxyapatite and then implanted in vivo into NOD/SCID mice or in animals with segmental bone defects and new bone formation was observed (8). BM-derived MSC have been transplanted into mice with osteogenesis imperfecta, a genetic disorder of mesenchymal tissues. In addition BMT had been used in children with this disease. The cells engrafted with an increase in the number of osteoblasts, formation of new lamellar bone and an increase in the total body mineral content were observed. Clinically there was a reduction in the frequency of fractures and enhancement of the body growth rate (9). Particularly promising for orthopaedic applications, especially for bone formation, is the use of natural or synthetic biomaterials as carriers for MSC delivery. Newer materials such as biodegradable polymers poly-L-lactide (PLA) and poly-L-lactide-
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co-glycolide (PLGA) can facilitate adhesion, proliferation and differentiation of cells. Clinical studies have been designed in which scaffolds were loaded with in vitro expanded autologous BM-derived MSCs and successfully implanted in patients with large bone defects. More recently, an extended mandible discontinuity was successfully repaired through heterotopic bone induction with biomaterials, autologous BM cells and growth factors (10). 3.3. Stem cells for diabetes Replacement of insulin-producing cells in patients with diabetes has become a major goal in regenerative therapy. Promising strategies have included either the use of cells with the potential to generate insulin-producing b-cells by expansion of existing‚ b-cells, differentiation of embryonic stem (ES) cells to b-cells, conversion of either pancreatic or non-pancreatic adult stem/progenitor cells into b-cells or the use pharmacological agents that seeks to regenerate b-cells in the pancreas, either by replication of existing b-cells (regeneration) or by the generation of new b-cells from other cell types (neogenesis). Numerous studies have been undertaken to establish whether insulin-producing b-cells can be developed from stemprogenitor cells from different origins. The initial claims that bone marrow stem cells could differentiate into other lineages, including insulin-producing cells are at best questionable at this time (11). It is possible however that BM cells may contribute to the endothelium of damaged pancreas, leading to the production of cytokines and growth factors that stimulate b-cell neogenesis, b-cell proliferation, or increased survival of the residual b-cells (12). 3.4. CNS disorders Diseases of the CNS, although a very attractive target for cell therapy, represent a significant challenge for cell-based strategies of repair, due to the complexity of the interactions between different types of cells and the required need for integration of the different structures. During prenatal development of the mammalian CNS, the neural stem cells (NSCs) and their progenitors expand and give rise to the functional neurons and glial cells that constitute the growing brain. A number of studies during the last 15 years have clearly demonstrated that in the adult mammalian CNS, a small number of NSCs are able to self-renew and generate different neural cell lineages including neurons, astrocytes and oligodendrocytes under specific microenvironmental stimuli (13). As we have described in other diseases, recent studies have suggested that terminal neural differentiation can also be seen with non-CNS-derived multipotent somatic stem cells, such as BM-derived stem cells. Most of these studies are based on the potential of non-neural stem cells to acquire certain “neural” markers in vitro after 494
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treatment with certain cytokines or factors that induced neural differentiation. However, studies have only recently started to address whether putative neural progeny from MSCs or other stem cells have functional characteristics consistent with neurons. There is no substantial evidence for in vivo differentiation and functional integration of any of these BM-derived stem cells. Regardless of the lack of true differentiation capacity, a number of studies in experimental models suggest that BM-derived stem cells like MSC or EPC can contribute to improve the functional capacity in patients with multiple sclerosis, stroke or spinal cord injuries (14).
Acknowledgments Supported in part by grants from the Government of Navarra, Ministerio de Sanidad, European Framework Project 6 and FEDER funds and the University of Navarra.
References 1. Thomson JA, Itskovitz-Eldor J, Shapiro SS, et al. Embryonic stem cell lines derived from human blastocysts. Science 1998; 282: 1145-1147. 2. Laflamme MA, Murry CE. Regenerating the heart. Nat Biotechnol 2005; 23: 845-856. 3. Murry CE, Field LJ, Menasche P. Cell-based cardiac repair: Reflections at the 10-year point. Circulation 2005; 112: 3174-3183. 4. Wollert KC, Drexler H. Clinical applications of stem cells for the heart. Circ Res 2005; 96: 151-163. 5. Dimmeler S, Burchfield J, Zeiher AM. Cell-Based Therapy of Myocardial Infarction. Arterioscler Thromb Vasc Biol 2007. 6. Tateishi-Yuyama E, Matsubara H, Murohara T, et al. Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells: A pilot study and a randomised controlled trial. Lancet 2002; 360: 427-435. 7. Lenk K, Adams V, Lurz P, et al. Therapeutical potential of blood-derived progenitor cells in patients with peripheral arterial occlusive disease and critical limb ischaemia. Eur Heart J 2005; 26: 1903-1909. 8. Petite H, Viateau V, Bensaid W, et al. Tissue-engineered bone regeneration. Nat Biotechnol 2000; 18: 959-963. 9. Horwitz EM, Prockop DJ, Fitzpatrick LA, et al. Transplantability and therapeutic effects of bone marrow-derived mesenchymal cells in children with osteogenesis imperfecta. Nat Med 1999; 5: 309-313. 10.Warnke PH, Springer IN, Wiltfang J, et al. Growth and transplantation of a custom vascularised bone graft in a man. Lancet 2004; 364: 766-770. 11.Ianus A, Holz GG, Theise ND, Hussain MA. In vivo derivation of glucose-competent pancreatic endocrine cells from bone marrow without evidence of cell fusion. J Clin Invest 2003; 111: 843-850. 12.Bonner-Weir S, Weir GC. New sources of pancreatic beta-cells. Nat Biotechnol 2005; 23:
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857-861. 13.Quinones-Hinojosa A, Sanai N, Soriano-Navarro M, et al. Cellular composition and cytoarchitecture of the adult human subventricular zone: A niche of neural stem cells. J Comp Neurol 2006; 494: 415-434. 14.Lindvall O, Kokaia Z, Martinez-Serrano A. Stem cell therapy for human neurodegenerative disorders-how to make it work. Nat Med 2004; 10 Suppl: S42-50.
Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. BM contains different populations of stem cells. One of the following is not a BM derived population of stem cells: a) Mesenchymal stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Haematopoietic stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Endothelial progenitor cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Myoblasts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Transplantation of BM derived stem cells in patients with acute MI: a) Is a well established and effective therapy. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Has demonstrated that HSC differentiate into functional cardiomyocytes in vivo . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Phase III clinical trials suggest that it may be associated with a functional benefit . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) All of the above . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Mesenchymal stem cells have been used in patients with one of the following diseases: a) Parkinson’s Disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Peripheral vascular disease . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Stroke . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Osteogenesis imperfecta . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Pluripotentiality is a characteristic of: a) Haematopoietic stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Side population . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 496
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c) Endothelial progenitor cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Embryonic stem cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which of the following markers are expressed by pluripotent stem cells: a) Oct4 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) CD34 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Sox2 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Both a and c . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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HSCT for acute myeloid leukaemia in children
G. Dini, M. Miano
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CHAPTER 35 • AML in children
1. Introduction The prognosis of acute myeloid leukaemia (AML) in children has significantly improved over the past two decades: with intensive chemotherapy 80–90% of children achieve complete remission (CR) and 30–70% are cured if they receive postinduction chemotherapy (1). Matched related donor (MRD) transplantation in first CR results in 45–64% long-term survival and represents an attractive option for children with high risk (HR) AML (2). However, since approximately 60% of children lack an MRD, the pros and cons of alternative approaches must be carefully weighed on a case-by-case basis.
2. Indications Recently, the European Group for Blood and Marrow Transplantation (EBMT) published indications for HSCT in all diseases, including AML. These are shown in Table 1 (3).
Table 1: Indications for HSCT for children with acute myeloid leukaemia Disease
Status
Sibling donor
Well matched unrelated / 1 Ag related
Mm unrelated / >1 Ag related
Auto-HSCT
AML
CR1 low-risk
GNR
GNR
GNR
GNR
CR1 high-risk
S
CO
GNR
S
CR1 very high-risk
S
S
CO
GNR
CR2
S
S
S
S
>CR2
CO
D
D
GNR
S: standard of care; generally indicated in suitable patients. CO: clinical option; can be carried out after careful assessment of risks and benefits. D: developmental; further trials are needed. GNR: generally not recommended. NA: not applicable. CR1, 2: first, second complete remission. Ag: antigen. Mm: mismatched. This classification does not cover patients for whom a syngeneic donor is available
3. Conditioning regimens Total body irradiation (TBI) was found to have no favourable impact on event free survival (EFS) in children undergoing HSCT for 1st CR AML. Thus, given its deleterious effect of provoking late sequelae, radiotherapy should no longer be administered to this subset of patients. Most teams currently use non-TBI-containing regimens in children transplanted for AML. The most commonly used regimens include busulfan and cyclophosphamide, almost always supplemented with melphalan. Some teams still use fractionated TBI-containing regimens, especially in high-risk
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cases (4). No studies comparing these two approaches in the treatment of paediatric AML have so far been published. However, the International-BFM-Study Group is comparing the two key preparative regimens (i.e., Bu+Cy+Mel vs. fTBI+Cy) as optional therapy in their ongoing trial on relapsed AML.
4. Role and outcome of autologous transplant Autologous HSCT has been widely used as consolidation treatment after induction therapy in children with HR AML in first or second CR lacking an MRD. However, the results of paediatric studies comparing autologous HSCT to chemotherapy are conflicting. Further randomised clinical trials are needed to address the pivotal clinical question of whether auto-HSCT is better than chemotherapy or allograft as consolidation treatment for childhood AML in first CR. A recent EBMT retrospective study involving 387 children given autologous HSCT for 1st CR AML showed that 5-year transplant related mortality (TRM), relapse rate (RR) and EFS were 3, 39 and 60%, respectively (5). A lower probability of survival was observed when; a) more than two induction courses were needed to reach CR, b) the median interval between diagnosis and achievement of CR was greater than 38 days, and c) the number of infused cells was above 2.85 x 108/kg. A trend towards a favourable effect of purging on the probability of EFS was observed. Children receiving BAVC as the conditioning regimen showed a higher relapse rate 4.1. Open questions and future directions Use of peripheral blood progenitor cells Peripheral blood progenitor cells (PBPCs) are not often used for autologous HSCT in children with AML. This is mainly due to the difficulty in collecting adequate numbers of circulating haematopoietic progenitor cells, not to mention the poorly defined effect that is observed in this subset of children when PBPCs are used to accelerate the recovery of haematopoiesis. Role of haematopoietic growth factors The beneficial impact of haematopoietic growth factors on myeloid recovery in children undergoing autologous HSCT for AML has not been proven, and given the cost of these cytokines, their use in this subset of patients should be avoided. In vitro purging In vitro purging is associated with reduced RR (p=0.04) (5). Delayed platelet engraftment is one of the drawbacks that may be observed. These data (5) suggest that in vitro purging should be used before carrying out autologous transplantation in childhood AML.
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5. Role and outcome of HLA-identical sibling transplant Several randomised trials have shown the statistically significant superiority of MRD HSCT as compared to all other options. This, in turn, is correlated with longer periods of “quality of life time” (2). In a recent, single-centre, retrospective study involving 55 children receiving MRD HSCT for 1st CR AML, the 5-year probability of survival was 74%, whereas the 5-year probability of relapse was 26% (6). None of the patients who developed acute graft versus host disease (GvHD) relapsed, confirming a graft versus leukaemia (GvL) effect following allogeneic HSCT for AML. Better EFS among the older children receiving a higher dose of TBI was observed in this study. This issue was recently confirmed by a review of the literature comparing various TBI regimens (7).
6. Role and outcome of Unrelated Donor (UD) transplant There is an absolute indication for UD HSCT in infant AML and in children with FAB M7 AML, who stand a very poor chance of being cured by chemotherapy or by autologous HSCT. FAB M0 or M6 represent more controversial indications. Timing in the identification of a suitable donor constitutes a limiting factor for this subset of patients. As compared to unrelated cord blood transplantation (UCBT), UD-HSCT has shown a similar incidence of grade III-IV acute GvHD and a higher incidence of chronic GvHD, while 2 year overall survival is similar (8). A recent study from the USA has shown that the 5-year EFS of children given an 8/8 allele matched UD-HSCT is similar to results obtained with a 1 or 2 antigen mismatched UCBT. TRM is slightly higher and relapse rate is lower after a 2 antigen mismatched UCBT (9).
7. Role and outcome of cord blood transplant Eurocord recently reported that the EFS of children with AML in 1st CR and in 2nd CR is 57 and 47%, respectively, while the RR is 10% in CR1 and 23% in CR2. The main prognostic factors are disease stage and number of infused cells (10).
8. Role and outcome of haploidentical transplants The results reported by the Perugia Group regarding patients with AML undergoing haploidentical HSCT showed that NK cells have an impressive effect on alloreactivity. In fact, no relapses occurred among the patients transplanted from haploidentical donors with KIR mismatched in the GvL direction, suggesting that the haploidentical option may play a role in the early phase of treatment for very high-risk AML patients. In vitro studies have confirmed that alloreactive NK clones exert a potent cytotoxic
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activity against the leukaemic cells taken from patients with CML and AML (11). More recently, several paediatric teams began to investigate the use of haploidentical HSCT in children with no other allogeneic donor options, or with an urgent need to proceed to transplant. Preliminary results confirm that the outcome of children in remission is similar to what can be achieved by using other donor stem cell sources (12).
9. Nature and role of Donor Lymphocyte Infusion A multi-centre study in children with AML showed that despite early donor lymphocyte infusion (DLI), relapse was still significantly more frequent in patients with increasing mixed chimerism (MC) than it was in patients with complete chimerism (CC), low-level MC (i.e. low level of host cells) or decreasing MC. Patients with increasing MC who received early DLI showed a significantly higher probability of EFS than patients with increasing MC who did not undergo immunological intervention. These results demonstrate that paediatric AML patients with increasing MC are at highest risk of relapse, and that early DLI can prevent relapse in these patients (13).
10. A treatment algorithm A suitably matched UD (8/8 or >/10 allele matched) can usually be located within 4 to 6 weeks for children with the most common haplotypes who lack an MRD. Alternative treatment options should be offered to children with rare HLA types, and a decision should be made as to whether to reduce the matching requirements or to select another type of therapy, such as UCBT or haploidentical HSCT. A matched or 1 antigen mismatched CB Unit containing more than 3 x 107 mononuclear cells should be considered equivalent to an 8/8 allele matched UD. The decision should be made based on the urgency of the HSCT. Haploidentical HSCT should be offered if no donors and no CB units with the above mentioned characteristics are available. Some teams consider haploidentical HSCT the second option when an acceptable unrelated donor is not available.
Acknowledgments G. Dini thanks V. Perricone for revising the manuscript.
References 1. Creutzig U, Ritter J, Schellong G for the AML-BFM Study Group. Identification of two risk groups in childhood acute myelogenous leukemia after therapy intensification in study
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AML-BFM-83 compared with study AML-BFM-78. Blood 1990; 75: 1932-1940. 2. Woods W G, Neudorf S, Gold S, et al. A comparison of allogeneic bone marrow transplantation, autologous bone marrow transplantation, and aggressive chemotherapy in children with acute myeloid leukaemia in remission: A report from the Children’s Cancer Group. Blood 2001; 97: 56-62. 3. Ljungman P, Urbano-Ispizua A, Cavazzana-Calvo M, et al. for the EBMT. Allogeneic and autologous transplantation for haematological diseases, solid tumours and immune disorders: Definitions and current practice in Europe. Bone Marrow Transplant 2006; 37: 439-449. 4. Vettenranta K on behalf of the Pediatric Diseases Working Party of the EBMT. Current European practice in pediatric myeloablative conditioning. Bone Marrow Transplant 2007; in press. 5. Locatelli F, Labopin M, Ortega J, et al. Factors influencing outcome and incidence of longterm complications in children who underwent autologous stem cell transplantation for acute myeloid leukaemia. Blood 2003; 15: 1611-1619. 6. Willemze AJ, Geskus RB, Noordijk, et al. HLA-identical haematopoietic stem cell transplantation for acute leukaemia in children: Less relapse with higher biologically effective dose of TBI. Bone Marrow Transplant 2007; 39: 1-9. 7. Kal HB, Loes vK-H, Heijenbrok-Kal MH, et al. Biologically effective dose in total-body irradiation and hematopoietic stem cell transplantation. Strahlenther Onkol 2006; 182: 672-679. 8. Hwang WY, Samuel M, Tan D, et al. A meta-analysis of unrelated donor umbilical cord Blood transplantation versus unrelated donor bone marrow transplantation in adult and pediatric patients. Biol Blood Marrow Transplant 2007; 13: 444-453. 9. Eapen M, Rubinstein P, Zhang MJ, et al. Outcomes of transplantation of unrelated donor umbilical cord blood and bone marrow in children with acute leukaemia: A comparison study. Lancet 2007; 369: 1947-1954. 10.Gluckman E and Rocha V on behalf of Eurocord and Paediatric WP of EBMT Indications and results of cord blood transplant in children with leukaemia. Bone Marrow Transplant 2007; in press. 11.Aversa F, Tabilio A, Velardi A, et al. Transplantation of high-risk acute leukemia with Tcell-depleted stem cells from related donor with one fully mismatched HLA haplotype. N Engl J Med 1998; 339: 1186-1193. 12.Marks DI, Khattry N, Cummins M, et al. Haploidentical stem cell transplantation for children with acute leukaemia. Br J Haematol 2006; 134: 196–201. 13.Bader P, Kreyenberg H, Hoelle W, et al. Increasing mixed chimerism defines a high-risk group of childhood acute myelogenous leukemia patients after allogeneic stem cell transplantation where pre-emptive immunotherapy may be effective. Bone Marrow Transplant 2004; 33: 815-821.
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Multiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The standard treatment for patients with high-risk AML in CR1 is: a) Allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Radiotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. The main risk factor for EFS in patients receiving allogeneic HSCT for AML is: a) Age . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) LDH levels before HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) FAB Classification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) WBC count . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. The standard treatment for patients suffering from low risk AML in CR1 is: a) Allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Radiotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. For patients with AML in CR2, which of the following criteria selects those who should receive allogeneic HSCT? a) Age . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) LDH levels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) All patients represent an indication. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) WBC number . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which of the following is true for allogeneic HSCT for patients suffering from AML in CR2? a) Generally indicated in suitable patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Not recommended . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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c) A developmental option to be demonstrated with further trials . . . . . . . . . . . d) Indicated after relapse following autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 36
HSCT for acute lymphoblastic leukaemia in children
T. Klingebiel, P. Bader
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CHAPTER 36 • ALL in children
1. Introduction ALL in childhood is a disease which is currently curable by first line treatment in about 80% of cases. The BFM Study Group has reported an EFS at 8 years of 75.9% for patients in the ALL BFM 90 study (1). The same is true for other large study groups, for example the St. Jude Group (2). Even after relapse, e.g. late bone marrow and isolated extramedullary relapse, long term EFS of 35% and 44% can be achieved with conventional chemotherapy alone (3). Therefore, indications for stem cell transplantation in childhood ALL must be defined in close collaboration with study groups conducting chemotherapy trials. An excellent overview on the role of alloand auto-HSCT is given in the systematic review of Theresa Hahn (4).
2. The role of allo-HSCT in ALL in first remission (CR1) The role of allo-HSCT particularly in first remission of childhood ALL has been a matter of intense debate. There is an ongoing cooperative trial of the BFM, I-BFM and EBMT Paediatric Diseases Working Party study groups to define exactly the role of HSCT in CR1. The trial is based on four digit high-resolution HLA typing for all non-sibling donors, standardised GvHD prophylaxis and a uniform conditioning regimen. This is a prospective trial comparing HSCT and chemotherapy. There is a broad agreement that at present a benefit for HSCT in first remission has been demonstrated only for high-risk ALL patients. Data on 326 children and young adults from study groups in several countries have been analysed (5). 267 patients achieved a complete remission after induction chemotherapy and were stratified into three subgroups; for these 267 patients the estimates of disease-free survival at 5 years were 49 ± 5% for the subgroup with the best prognosis, 30 ± 5% for patients with an intermediate prognosis and 20 ± 5% for the subgroup with the worst prognosis. An analysis of the role of post-remission therapy demonstrated that transplantation of bone marrow from HLA identical related donors (n=38) offered significantly greater benefit than intensive chemotherapy alone in terms of protecting patients from relapse or other adverse events (relative risk 0.3) whereas other types of transplantation (matched unrelated donors, mismatched related donors and autologous transplantation) had no benefit compared to chemotherapy alone. This finding was consistent in all three risk groups. In a more recent study in very high-risk ALL (defined as failure to achieve complete remission after the first four-drug induction phase; t(9;22) or t(4;11) clonal abnormalities; poor response to prednisone in combination with T-immunophenotype and/or white blood cell count >100 x 109/L) a comparison was made between children treated with chemotherapy alone or HSCT from an HLA matched sibling donor (6). 357 children entered the study of whom 280 were treated by chemotherapy and 77 by sibling donor transplantation.
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5-year-DFS was 40.6% in children treated with chemotherapy alone and 56.7% in children receiving stem cell transplantation.
3. The role of allo-HSCT in second and subsequent remission (≥CR2) There is evidence for the superiority of allo-HSCT from HLA-identical siblings in patients with bone marrow and combined relapses. However the benefit of unrelated allo-HSCT is restricted to certain subgroups. Barrett (7) analysed data of 376 HLA identical sibling transplants in comparison to 540 children treated with chemotherapy alone by the American Pediatric Oncology Group in second remission after a bone marrow relapse. The mean probability of a relapse at 5 years was significantly lower among transplant recipients (45 ± 4%) compared to the chemotherapy recipients (80 ± 3%, p<0,001); correspondingly the 5 year probability of leukaemia-free survival was higher after transplantation (40 ± 3%) than after chemotherapy (17 ± 3%, p<0,001). Uderzo (8) compared the results of allo-HSCT with those obtained with chemotherapy in second complete remission for the Italian Bone Marrow Transplantation Group and the Italian Paediatric Haematology Oncology Association. Of 287 eligible patients 230 were treated with chemotherapy and 57 underwent allo-HSCT. After transplant the DFS was significantly longer than after chemotherapy (relative risk 0.54); in this analysis, in contrast to the Barrett analysis, no benefit was found for patients with a late relapse. Harrison (9) presented the results of the MRC UKALL R1 trial analysed by HLA matched donor availability. In this analysis patients with a first remission less than 2 years having a donor had a significant advantage over patients having no donor (p = 0.05). For the BFM relapse (ALL-REZ BFM) study group Borgmann (10) performed a matched-pair analysis comparing unrelated donor transplantation with continuation of chemotherapy after a first relapse. 81 pairs were identified that could be matched for site of relapse and immunophenotype and as closely as possible for duration of first remission, age, date of diagnosis and peripheral blast cell count at relapse. For patients with intermediate prognosis no benefit for unrelated donor transplantation was found, whereas for patients with a poor prognosis benefit was evident (probability of EFS at 4 years 0.44 ± 0.7 for patients after unrelated HSCT vs. 0.00 for patients with chemotherapy alone). Taken together, for patients with a high risk of relapse related or unrelated alloHSCT is clearly of benefit.
4. Haploidentical transplantation (haplo-HSCT) For patients lacking an HLA identical sibling donor in the family and where an HLA identical unrelated donor cannot be identified in time, stem cell transplantation from 508
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a mismatched family member (haploidentical stem cell transplantation) is an alternative. It has been shown in children with ALL transplanted with large numbers of purified CD34+ stem cells and low numbers of CD3+ cells (below 2.5 x 103/kg) that an engraftment rate of more than 95% can be achieved and a long term leukaemia free survival of 44 ± 11% is possible. There was no statistical difference in survival for patients with 1, 2 or 3 antigen-mismatched donors or for patients in first, second or third remission. However as well as the relapses (10 patients of 27) there were 7 cases of transplant related mortality (11). With a new technique for CD3/CD19 depletion a more reliable engraftment and a more rapid T-cell reconstitution can be seen. It is expected that using this approach the rates of TRM can be reduced (12).
5. The role of outcome of cord blood transplantation Since the first report on the use of cord blood for stem cell transplantation many children with ALL have been treated in first and second CR. In a large registry analysis (13) 503 children with ALL transplanted with umbilical cord blood were compared with 282 bone marrow recipients. All patients had been treated in the USA. Cord blood was either fully matched (n = 35), mismatched for one HLA-antigen (n = 201) or for 2 antigens (n = 267); donor marrow was matched at the allele level for HLA A, B, C and DRB (n = 116) or mismatched (n = 166). The five year probabilities of leukaemia free survival were 38% after HLA-matched bone marrow transplants, 37% after mismatched bone marrow transplants, 60% after HLA-matched cord blood transplants, 36% after one antigen mismatched cord-blood transplant with low cell dose, 45% after one antigen mismatched cord-blood with high cell dose and 33% after two antigen-mismatched cord blood. Therefore, it seems that umbilical cord blood in experienced centres plays a role for the treatment of patients with ALL.
6. Related versus unrelated allo-HSCT In spite of a large number of studies, the outcome of related vs. unrelated donor allogeneic stem cell transplantation has not yet been adequately studied. There are no reported studies that compare outcome of HLA identical sibling donors with unrelated donors matched on allele level. Here the results of the BFM/IBFM/EMBT studies are pending. However there is an analysis from Eapen (14) in which the results of unrelated donor transplantation with bone marrow (n = 85) or cord blood (n = 81) were compared with those of HLA matched sibling transplants (n = 101) for acute leukaemia (ALL and AML) in children. Treatment related mortality was 6% for matched sibling transplantation, 15% for unrelated donor and 31% for cord blood
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HSCT respectively. The reason for the higher mortality rates is presumed to be secondary to slower myeloid recovery and higher rates of death from infections. Risks of relapse, overall and leukaemia free survival rate were closely associated with disease state at transplantation. However, leukaemia recurrence was lowest after unrelated donor HSCT in first clinical remission. After adjustment for disease status overall survival and leukaemia-free survival were similar after matched sibling and unrelated donor bone marrow or cord blood transplantation. It can be expected that using the same eligibility criteria results for related and unrelated stem cell transplantation for adequately matched donors will be identical.
7. Comparison of conditioning regimen It is accepted that TBI containing regimens in ALL have better outcomes than nonTBI containing regimen (15).
8. The role of additional cellular leukaemia therapy post transplant There has been a long-term debate as to whether ALL is a subject for immunotherapy. If additional immunotherapy is offered to patients who have already relapsed after transplantation only a very few patients will respond. It seems to be important to offer such treatment pre-emptively to avoid frank relapse. It was shown in a prospective study in 163 children with ALL in different remission states transplanted from sibling, unrelated and haploidentical donors that pre-emptive immunotherapy has an important effect on chimerism and thereby prevents relapse (16). There were 101 patients with complete chimerism, whereas increasing mixed chimerism was found in 46 and decreasing in 16 patients. Relapse was significantly more frequent in patients with increasing mixed-chimerism (26/46), whereas in patients with complete or low level chimerism only 8/101 relapsed and there were no relapses in patients with decreasing mixed-chimerism. The probability of three year EFS was 54% for all patients, 66% for patients with complete or low level mixed-chimerism, 66% for patients with decreasing mixed-chimerism and 23% for patients with increasing mixed chimerism. Of the 46 patients with increasing mixed chimerism 31 received immunotherapy and this group had a significant higher three year EFS (37%) than the 15 patients who did not receive any immunotherapy (EFS 0%). It can therefore be concluded that immunotherapy either by withdrawal of cyclosporin A or additional donor lymphocyte infusion alters the fate of patients with increasing mixed-chimerism.
9. Role of minimal residual disease monitoring after stem cell transplantation It is well known that relapse is the most frequent complication after allo-HSCT in 510
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childhood leukaemia. It is now possible to measure minimal residual disease (MRD) by real time quantitative PCR frequently and nearly in real time after HSCT. However, results of large prospective trials within IBFM and EBMT Paediatric Working Party are pending.
10. Conclusions The benefit of allo-HSCT in ALL patients in CR1 for high-risk subgroups has been established. In CR2 related HLA identical transplantation has proven as the gold standard of care. There is a clear indication for unrelated donor transplantation in all high-risk subgroups, whereas its role for low-risk groups needs to be defined within current clinical trials. Cord blood and haploidentical donors widen the spectrum of allo-HSCT in children. In experienced centres the same results can be achieved as in unrelated donor transplants. Pre-emptive immunotherapy post HSCT may be effective. The role of MRD analysis post transplant is still under investigation.
References 1. Schrappe M, Reiter A, Zimmermann M. Long-term results of four consecutive trials in childhood ALL performed by the ALL-BFM study group from 1981 to 1995. BerlinFrankfurt-Munster. Leukemia 2000; 14: 2205-2222. 2. Pui CH, Sandlund JT, Pei D. Improved outcome for children with acute lymphoblastic leukemia: Results of Total Therapy Study XIIIB at St. Jude Children’s Research Hospital. Blood 2004; 104: 2690-2696. 3. Einsiedel HG, von Stackelberg A, Hartmann R. Long-term outcome in children with relapsed ALL by risk-stratified salvage therapy: Results of trial acute lymphoblastic leukemia-relapse study of the Berlin-Frankfurt-Munster Group 87. J Clin Oncol 2005; 23: 7942-7950. 4. Hahn T, Wall D, Camitta B. The role of cytotoxic therapy with hematopoietic stem cell transplantation in the therapy of acute lymphoblastic leukemia in children: An evidencebased review. Biol Blood Marrow Transplant 2005; 11: 823-861. 5. Arico M, Valsecchi MG, Camitta B. Outcome of treatment in children with Philadelphia chromosome-positive acute lymphoblastic leukemia. N Engl J Med 2000; 342: 998-1006. 6. Balduzzi A, Valsecchi MG, Uderzo C. Chemotherapy versus allogeneic transplantation for very-high-risk childhood acute lymphoblastic leukaemia in first complete remission: Comparison by genetic randomisation in an international prospective study. Lancet 2005; 366: 635-642. 7. Barrett AJ, Horowitz MM, Pollock BH. Bone marrow transplants from HLA-identical siblings as compared with chemotherapy for children with acute lymphoblastic leukemia in a second remission. N Engl J Med 1994; 331: 1253-1258. 8. Uderzo C, Valsecchi MG, Bacigalupo A. Treatment of childhood acute lymphoblastic
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leukemia in second remission with allogeneic bone marrow transplantation and chemotherapy: Ten-year experience of the Italian Bone Marrow Transplantation Group and the Italian Pediatric Hematology Oncology Association. J Clin Oncol 1995; 13: 352-358. 9. Harrison G, Richards S, Lawson S. Comparison of allogeneic transplant versus chemotherapy for relapsed childhood acute lymphoblastic leukaemia in the MRC UKALL R1 trial. MRC Childhood Leukaemia Working Party. Ann Oncol 2000; 11: 999-1006. 10.Borgmann A, von Stackelberg A, Hartmann R. Unrelated donor stem cell transplantation compared with chemotherapy for children with acute lymphoblastic leukemia in a second remission: A matched-pair analysis. Blood 2003; 101: 3835-3839. 11.Klingebiel T, Handgretinger R, Lang P. Haploidentical transplantation for acute lymphoblastic leukemia in childhood. Blood Rev 2004; 18: 181-192. 12.Bader P, Willasch A, Niethammer D. Haploidentical stem cell transplantation in childhood. Current Cancer Therapy Reviews 2007; 3: 37-44. 13.Eapen M, Rubinstein P, Zhang MJ. Outcomes of transplantation of unrelated donor umbilical cord blood and bone marrow in children with acute leukaemia: A comparison study. Lancet 2007; 369: 1947-1954. 14.Eapen M, Rubinstein P, Zhang MJ. Comparable long-term survival after unrelated and HLAmatched sibling donor hematopoietic stem cell transplantations for acute leukemia in children younger than 18 months. J Clin Oncol 2006; 24: 145-151. 15.Eapen M, Raetz E, Zhang MJ. Outcomes after HLA-matched sibling transplantation or chemotherapy in children with B-precursor acute lymphoblastic leukemia in a second remission: A collaborative study of the Children’s Oncology Group and the Center for International Blood and Marrow Transplant Research. Blood 2006; 107: 4961-4967. 16.Bader P, Kreyenberg H, Hoelle W. Increasing mixed chimerism is an important prognostic factor for unfavorable outcome in children with acute lymphoblastic leukemia after allogeneic stem-cell transplantation: Possible role for pre-emptive immunotherapy? J Clin Oncol 2004; 22: 1696-1705.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Which one of the following statements about transplantation in ALL is correct? a) A benefit for allogeneic stem cell transplantation in ALL-patients has been demonstrated for all patients in first remission . . . . . . . . . . . . . . . . . . b) It has been proven that transplantation from all donors protects children with high-risk and very high-risk leukaemia from relapse . . . . . . . . c) It has been proven that transplantation from identical related donors protects patients with high risk ALL from relapse . . . . . . . . . . . . . . . . . . 512
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d) It has been proven that autologous stem cell transplantation protects patients with high risk ALL in first remission from relapse . . . . . . 2. Which one of the following statements about transplantation in ALL is correct? a) There is evidence that unrelated stem cell transplantation protects children with CR2 in all risk groups from relapse . . . . . . . . . . . . . . . . . b) There is evidence that patients with high risk ALL in CR2 (short first remission, T-cell-phenotype etc.) benefit from unrelated transplantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) There is evidence that chemotherapy is of advantage in CR2 in early relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) There is no controversy about the role of allo-stem cell transplantation from patients with late relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which one of the following statements about haploidentical transplantation in ALL is correct? a) The number of CD34+-cells does not affect outcome . . . . . . . . . . . . . . . . . . . . . . . b) The number of CD3+-cells should be below one million per kg. . . . . . . . . . . . . c) Patients receiving haploidentical transplantation for ALL in remission can have a long term leukaemia free survival of more than 40% . . . . . . . . . . d) TRM is not higher in haploidentical transplantation than in other forms of transplantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Which one of the following statements about transplantation in ALL is correct? a) Umbilical cord blood does not play any role for treatment of patients ALL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Even with one or two antigens mismatched cord blood results are comparable with allele matched bone marrow . . . . . . . . . . . . . . . . . . . . . . . . . . c) Treatment related mortality is not higher for cord blood transplantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) The risk of relapse is not associated with disease state at transplantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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5. Which one of the following statements about transplantation in ALL is correct? a) There is no evidence that TBI is better than other conditioning regimen in ALL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) There is evidence that DLI after stem cell transplantation can convert chimerism and prevent relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Immunotherapy (e.g. reduction of CSA) is useless in ALL . . . . . . . . . . . . . . . . . . d) There is clear proven data from large trials that MRD post transplantation has an influence on outcome . . . . . . . . . . . . . . . . . . . . . . . . .
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NOTES
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CHAPTER 37
HSCT for myelodysplastic syndromes in children
F. Locatelli, C.M. Niemeyer
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CHAPTER 37 • MDS in children
1. Introduction The myelodysplastic syndromes (MDS) are a heterogeneous group of clonal disorders, accounting for less than 5% of all haematological malignancies of childhood (1). Childhood MDS include both variants shared with the adult population (i.e. RAEB, RAEB-t) and other disorders more typical of the paediatric age, such as juvenile myelomonocytic leukaemia (JMML) (1, 2). This latter disorder predominates in infants, the median age at diagnosis being 2 years (2). About 9% of patients with JMML are diagnosed before the age of 4 months, whereas less than 10% of patients are 6 years or older. In JMML, there is a male predominance with a male/female ratio of 2:1. Hypersensitivity to GM-CSF and pathological activation of the RAS-RAF-MAP (mitogen-activated protein) kinase signalling pathway play an important role in the pathophysiology of JMML. Allogeneic HSCT is the only curative approach for children with JMML, resulting in long-term survival in a significant proportion of patients receiving an allograft (2–5). Childhood MDS other than JMML often occur in the context of congenital bone marrow failure syndromes, this fact representing a peculiarity of myelodysplasia occurring in the paediatric age group (1). HSCT is routinely offered also to all children with RAEB and RAEB-t, to paediatric patients with MDS secondary to chemo-radiotherapy, and to those with refractory cytopenia (RC) associated with cytogenetic anomalies or transfusion dependence (6).
2. Indications, results and risk factors 2.1. Juvenile myelomonocytic leukaemia A number of different studies have reported that children with JMML can be definitively cured by an allograft (3–5). In the most recent study, which included the largest number of patients with JMML given allogeneic HSCT from either a histocompatible relative or from an HLA-matched/1-antigen mismatched donor, the probability of LFS was in the order of 50% (4). In multivariate analysis, age greater than 4 years and female sex predicted poorer outcome (4). Available data indicate that, in the more recent years, using an unrelated donor offers minimal or possibly no significant disadvantage as compared to employing an HLA-identical sibling (4). While one study reported a negative impact of monosomy 7 (7), the most frequent cytogenetic anomaly in JMML, on the probability of OS after HSCT, other larger analyses documented that neither monosomy 7 nor other cytogenetic abnormalities confer a poorer prognosis (3–5). Leukaemia recurrence represents the main cause of treatment failure in children with JMML treated by HSCT, relapse rate being as high as 50% (4).
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Preparative regimens without TBI are particularly attractive for children with JMML since radiation-induced late effects, such as severe growth retardation, cataracts, hypothyroidism and neuropsychologic sequels may be especially deleterious for very young children. Moreover, in a retrospective analysis of the EWOG-MDS, busulfanbased myeloablative therapy offered a greater anti-leukaemic efficacy than TBI (3). The recommended preparative regimen of the EBMT/EWOG-MDS groups for children with JMML includes busulfan, cyclophosphamide and melphalan. Splenectomy before HSCT, as well as spleen size at time of the allograft, did not appear to have an impact on post-transplantation outcome of children with JMML. Available data are not in favour of an indiscriminate use of splenectomy before transplantation, the potential advantages having to be weighed against the risks related to the procedure or to post-splenectomy infections (3, 5). The indication of performing splenectomy has to be carefully evaluated for each individual child, the presence of massive splenomegaly with evidence of hypersplenism and/or refractoriness to platelet transfusions being an argument for considering this procedure in order to promote engraftment, to hasten haematological recovery and to lower the risk of haemorrhagic complications. Available data indicate that unrelated cord blood transplant (UCBT) is a suitable option for children with JMML lacking an HLA-compatible relative and that the search for an unrelated CB unit should be initiated at the same time as that for an unrelated BM donor (5). CB offers the advantage of prompter availability of stem cells and HSCT can be successful even in the presence of donor HLA disparities. For children with JMML experiencing leukaemia relapse after allogeneic HSCT, DLI was proved to be largely ineffective (8), while a second allograft, from either the same or a different donor, together with reduction of the intensity of GvHD prophylaxis aimed at optimizing the GvL effect, is able to rescue about one third of the patients (9). 2.2. Other types of MDS Data on outcome of HSCT in children with advanced MDS other than JMML are scanty, the reported disease-free survival (DFS) being in the order of 60% when the donor is an HLA identical sibling (10). The need for pre-HSCT remission induction chemotherapy remains a debated question in paediatric patients with RAEB and RAEB-t. In fact, it remains controversial whether cytoreductive therapy prior to HSCT for more advanced forms of MDS improves survival. A study published by the Nordic Pediatric Haematology group, comparing the outcome of children with de novo MDS (including JMML) and children with de novo AML, documented that patients belonging to the former group had a lower rate of complete remission and a higher risk of death for treatment-related complications (11). In an EWOG-MDS analysis 518
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on children with MDS other than JMML, the outcome of patients given intensive chemotherapy prior to the allograft was found to be absolutely comparable to that of children who were transplanted directly (12). Patients with RC must be considered for an early allograft from either a related or an unrelated donor if they have cytogenetic abnormalities, in particular monosomy 7. In fact, a study of the EWOG-MDS analysing children with RC has clearly demonstrated that the probability of progression to more advanced MDS (i.e. RAEB and RAEB-t), as well as to frank AML, is significantly higher in patients with monosomy 7 than in those with a normal karyotype (6). Moreover, this study has also shown that patients who had not progressed to advanced MDS prior to HSCT had a significantly better probability of survival than patients who experienced disease progression (76 versus 36%, respectively, p=0.03) (6). In the presence of a normal karyotype, a substantial proportion of children with RC may experience a long, stable course of their disease. In view of the low TRM observed in patients transplanted from an HLA-compatible sibling, HSCT may be recommended if a suitable HLA-matched relative is available. A “watch and wait” approach with careful observation may be reasonable for patients without an HLA-identical sibling in the absence of transfusion requirements, severe cytopenia or infections. As the risk of disease recurrence after the allograft in patients with RC is low, there is a great interest in testing the safety and efficacy of reduced intensity regimens in this setting. In a recent EWOG-MDS report, patients with RC and normal karyotype transplanted from an unrelated donor following a fludarabine-based reducedintensity regimen had a favourable post-transplant outcome, which was comparable to that obtained in patients transplanted following a myeloablative conditioning regimen (13). The outcome of children with MDS secondary to previous cytotoxic or radiant treatment remains still poor, for both a high risk of disease recurrence and TRM. Only one study proposed by the Children Cancer Group, which enrolled a limited number of children, has addressed the issue of autologous HSCT in childhood MDS. In this trial, children lacking an HLA-compatible sibling received intensively timed induction therapy, which was followed by 4-hydroperoxycyclophosphamide-purged autologous marrow transplantation (14). Further larger studies are needed before recommending autologous transplantation in children with MDS other than JMML lacking a suitable compatible donor.
3. Conclusions and future perspectives The available data indicate that HSCT is curative for the majority of children with MDS, the outcome of patients transplanted from either an HLA-identical sibling or
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an unrelated volunteer being comparable in more recent years. Strategies to reduce the risk of leukaemia recurrence in children with JMML, RAEB, RAEB-t and MDS secondary to previous treatment, as well as to abate TRM in children with RC, could further optimise the results of HSCT in childhood MDS.
References 1. Hasle H, Niemeyer CM, Chessells JM, et al. A pediatric approach to the WHO classification of myelodysplastic and myeloproliferative diseases. Leukemia 2003; 17: 277-282. 2. Niemeyer CM, Aricò M, Basso G, et al. Chronic myelomonocytic leukemia in childhood: A report of 110 cases. Blood 1997; 89: 3534-3543. 3. Locatelli F, Niemeyer C, Angelucci E, et al. Allogeneic bone marrow transplantation for chronic myelomonocytic leukemia in childhood: A report from the European Working Group on Myelodysplastic Syndrome in Childhood. J Clin Oncol 1997; 15: 566-573. 4. Smith FO, King R, Nelson G, et al. National Marrow Donor Program. Unrelated donor bone marrow transplantation for children with juvenile myelomonocytic leukaemia. Br J Haematol 2002; 116: 716-724. 5. Locatelli F, Nollke P, Zecca M, et al. European Working Group on Childhood MDS; European Blood and Marrow Transplantation Group. Hematopoietic stem cell transplantation (HSCT) in children with juvenile myelomonocytic leukemia (JMML): Results of the EWOGMDS/EBMT trial. Blood 2005; 105: 410-419. 6. Kardos G, Baumann I, Passmore SJ, et al. Refractory anemia in childhood: A retrospective analysis of 67 patients with particular reference to monosomy 7. Blood 2003; 102: 1997-2003. 7. Manabe A, Okamura J, Yumura-Yagi K, et al. on behalf of the MDS Committee of the Japanese Society of Pediatric Hematology. Allogeneic hematopoietic stem cell transplantation for 27 children with juvenile myelomonocytic leukemia diagnosed based on the criteria of the International JMML Working Group. Leukemia 2002; 16: 645649. 8. Yoshimi A, Bader P, Matthes-Martin S, et al. on behalf of European Working Group of MDS in Childhood (EWOG-MDS). Donor leukocyte infusion after hematopoietic stem cell transplantation in patients with juvenile myelomonocytic leukemia. Leukemia 2005; 19: 971-977. 9. Yoshimi A, Mohamed M, Bierings M, et al. on behalf of the European Working Group of MDS in Childhood (EWOG-MDS). Second allogeneic hematopoietic stem cell transplantation (HSCT) results in outcome similar to that of first HSCT for patients with juvenile myelomonocytic leukemia. Leukemia 2007; 21: 556-560. 10.Locatelli F, Pession A, Bonetti F, et al. Busulfan, cyclophosphamide and melphalan as conditioning regimen for bone marrow transplantation in children with myelodysplastic syndromes. Leukemia 1994; 8: 844-849. 11.Hasle H, Kerndrup G, Yssing M, et al. Intensive chemotherapy in childhood myelodysplastic syndrome. A comparison with results in acute myeloid leukemia. Leukemia 1996; 10: 12691273. 520
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12.Niemeyer C, Duffner U, Bender-Gotze C, et al. AML-type intensive chemotherapy prior to stem cell transplantation (SCT) does not improve survival in children and adolescents with primary myelodysplastic syndromes (MDS). Blood 2000; 96 (suppl. 1): 521a. 13.Strahm B, Locatelli F, Bader P, et al. on behalf of the EWOG-MDS Study Group Reduced intensity conditioning in unrelated donor transplantation for refractory cytopenia in childhood. Bone Marrow Transplant 2007, 40: 329-333. 14.Woods WG, Kobrinsky N, Buckley J, et al. Intensively timed induction therapy followed by autologous or allogeneic bone marrow transplantation for children with acute myeloid leukemia or myelodysplastic syndrome: A Children Cancer Group pilot study. J Clin Oncol 1993; 11: 1448-1457.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. What is the percentage of children with JMML estimated to be cured with allogeneic HSCT? a) 25% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 75% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 90% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 50% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which is the most important prognostic factor predicting poor outcome for children with JMML given allogeneic HSCT? a) An age greater than 4 years at diagnosis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A monocyte count above 3 x 109/L . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) A leukocyte count above 30 x 109/L . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Monosomy 7 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Which is the best treatment for children with JMML relapsing after allogeneic HSCT? a) Single donor leukocyte infusion . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Repeated donor leukocyte infusions. . . . . . . . . . . . . . . . . . . . . . . . . c) Second allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) AML-like chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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4. The risk of disease recurrence in children with refractory cytopenia is: a) 30% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) No more than 10% even with reduced intensity regimens . . . . . . . . . . c) No more than 10% with myeloablative regimens, but much higher in patients given reduced intensity regimens . . . . . . . . . . . . . . . . . . d) 50% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. In children with advanced MDS (i.e. RAEB and RAEB-t): a) Pre-HSCT remission induction chemotherapy should always be employed . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Pre-HSCT remission induction chemotherapy should never be employed . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Pre-HSCT remission induction chemotherapy remains a debated question . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Pre-HSCT remission induction chemotherapy should always be employed above 10% bone marrow blasts. . . . . . . . . . . . . . . . . . .
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CHAPTER 38
HSCT for chronic myeloproliferative disorders in children
F. Locatelli, P. Merli
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CHAPTER 38 • Chronic MPD in children
1. Introduction Among chronic myeloproliferative disorders (MPD), CML certainly represents the most common variant, accounting for approximately 3–5% of all leukaemias in childhood (1). The estimated incidence of Philadelphia (Ph+) CML in paediatric patients has been reported to be less than 1 in 100,000 and it is less common under the age of 2 as compared with other age groups (2). In children, the disease is characterised by the same molecular, cytogenetic, clinical and morphological features reported in adults with classical CML. As in adults, so far, allogeneic HSCT is considered to be the only proven curative treatment for children with Ph+ CML (3–6). However, it must be underlined that not all candidates for transplantation have a suitable donor, either related or unrelated, and, despite its curative potential, HSCT carries the risk of death associated with the procedure, as well as of leukaemia recurrence. Moreover, the natural history of Ph+ CML has been recently profoundly modified by the introduction of the specific Bcr/Abl tyrosine protein kinase inhibitors, the most frequently employed being imatinib mesylate, which target the enzymatic activity of the Bcr-Abl protein, occupying the ATP-binding pocket of the molecule. The tyrosine kinase inhibitors have a high probability of inducing the achievement of both major and complete cytogenetic response and a high rate of freedom from progression to AP or BC (7). So far, however, we do not have convincing evidence that the Ph+ clone can be either completely eradicated or rendered silent for many decades by prolonged treatment with tyrosine kinase inhibitors. Other MPD, such as polycythaemia vera (PV) and myelofibrosis with myeloid metaplasia (MMM), are extremely rare in children and no sound data are available for analysing the role of HSCT in children affected by these disorders.
2. Indications, results and risk factors In some of the most important studies addressing the role of HSCT in patients with Ph+ CML, children have been included in adult series, but they represented a small proportion of the patient group and their outcome was not considered separately (3–5). Recently, the Chronic Leukemia Registry of the European Group for Blood and Marrow Transplantation (EBMT) has evaluated the outcome of 314 children with CML transplanted between 1985 and 2001 from either a related donor or an unrelated one, selected using high-resolution molecular typing of HLA class II antigens only (6). In this study, 3-year OS and LFS were 66 and 55%, respectively. In multivariate analysis for both OS and LFS, outcome was superior in patients given the allograft in CP1 versus advanced phase, although it is noteworthy that more than one third of patients transplanted in AP or BC are alive and disease free 3 years after
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transplantation (6). Inferior LFS was also found in children transplanted more than 6 months from diagnosis, this finding confirming previously published studies, which documented a worse outcome for adult patients transplanted more than 12 months after diagnosis as compared to those given HSCT earlier. The TRM in the cohort of patients analysed by the EBMT group was significantly higher for children transplanted from an unrelated volunteer donor, these patients having a 35% chance of fatal transplantation-related complications as compared to 20% for recipients of sibling allografts (6). The higher incidence of transplant-related death observed in patients transplanted from an unrelated volunteer is mainly due to a greater incidence of severe GvHD in these recipients as compared to those transplanted from an HLA-compatible sibling. A more precise characterisation of HLA alleles using high-resolution typing for both Class I and Class II molecules may permit a more accurate selection of unrelated donors, thus reducing the incidence of immunemediated complications and fatal events after the allograft (8, 9). Thus, for patients transplanted in more recent years, the outcome of patients given HSCT from an unrelated volunteer is comparable to that of patients transplanted from a compatible sibling. Treatment of CML relapse after an allograft has significantly benefited from adoptive immunotherapy with DLI. In fact, in patients with CML experiencing haematological relapse in CP after HSCT, complete remission can be obtained with this treatment in approximately 70% of cases (10). Most of these remissions are sustained over time, this proving the capacity of DLI to eradicate clonogenic leukaemia cells or to control their re-growth. Patients suffering from either cytogenetic or molecular relapse have an even greater chance of benefiting from DLI than those experiencing haematological relapse, especially if in advanced phase.
3. Conclusions and future perspectives The available data indicate that HSCT is curative for the majority of children with CML, although in the past TRM unfavourably affected the outcome of patients transplanted from an unrelated volunteer donor. Long-term survival is also influenced by the stage of the disease at time of transplantation, significantly better outcome having been observed in patients transplanted in CP1. This observation, together with the finding that LFS is significantly better for children transplanted within 6 months of diagnosis, suggests that it is important to proceed to HSCT as soon as an HLA-identical donor has been identified. It is possible that in the future the choice of transplanting children with CML will have to be balanced against the results achieved with tyrosine kinase inhibitors. However, considering the long life expectancy of children, if these agents do not prove to be able to offer, either alone 526
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or in combination with other treatment, a sustained “molecular cure” or indefinitely prolonged CP of the disease, HSCT, possibly in the first year after diagnosis, remains the treatment of choice of childhood Ph+ CML, provided that a well-matched donor is available. Strict monitoring and early detection of minimal residual disease through serial quantitative evaluation of the chimaeric Bcr/Abl mRNA transcript by means of PCR can be extremely useful for ensuring the best chance of favourable outcome in patients with Ph+ CML given HSCT.
References 1. Grier HE, Civin CI. Myeloid Leukemias, myelodysplasia and myeloproliferative disease in children. In: Nathan DG, Orkin SH, editors. Hematology of Infancy and Childhood. Philadelphia: Saunders, WB, 1998: 1300-1308. 2. Hall GW. Cytogenetic and molecular genetic aspects of childhood myeloproliferative/myelodysplastic disorders. Acta Haematol 2002; 108: 171-179. 3. Goldman JM, Apperley JF, Jones L, et al. Bone marrow transplantation for patients with chronic myeloid leukemia. N Engl J Med 1986; 314: 202-207. 4. Hansen JA, Gooley TA, Martin PJ, et al. Bone marrow transplants from unrelated donors for patients with chronic myeloid leukemia. N Engl J Med 1998; 338: 962-968. 5. Gratwohl A, Hermans J, Goldman JM, et al. Risk assessment for patients with chronic myeloid leukaemia before allogeneic blood or marrow transplantation. Chronic Leukemia Working Party of the European Group for Blood and Marrow Transplantation. Lancet 1998; 352: 1087-1092. 6. Cwynarski K, Roberts IAG, Iacobelli S, et al. for the Paediatric and Chronic Leukaemia Working Parties of the EBMT. Stem cell transplantation for chronic myeloid leukemia in children. Blood 2003; 102: 1224-1231. 7. O’Brien SG, Guilhot F, Larson RA. Imatinib compared with interferon and low-dose cytarabine for newly diagnosed chronic-phase myeloid leukaemia. N Engl J Med 2003; 348: 994-1004. 8. Flomenberg N, Baxter-Lowe LA, Confer D, et al. Impact of HLA class I and class II highresolution matching on outcomes of unrelated donor bone marrow transplantation: HLA-C mismatching is associated with a strong adverse effect on transplantation outcome, Blood 2004; 104: 1923-1930. 9. Lee SJ, Klein J, Haagenson M, et al. High resolution donor-recipient HLA matching contributes to the success of unrelated donor marrow transplantation. Blood 2007, in press. 10.Locatelli F. The role of repeat transplantation of haemopoietic stem cells and adoptive immunotherapy in treatment of leukaemia relapsing following allogeneic transplantation. Br J Haematol 1998; 102: 633-638.
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Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. What is the estimated incidence of Philadelphia (Ph+) CML in paediatric patients? a) Less than 1 in 100,000. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Less than 1 in 1,000,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 1 in 50,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 1 in 10,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Which is the most important prognostic factor predicting poor outcome for children with Ph+ CML given allogeneic HSCT? a) To have been splenectomised before transplantation . . . . . . . . . . . . . . . . . . . . . . b) To be transplanted in accelerated phase, blast crisis or second chronic phase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) To have been treated with interferon before transplantation . . . . . . . . . . . . . . d) To be younger than 10 years at time of the allograft . . . . . . . . . . . . . . . . . . . . . . 3. Which is the best first-line treatment for children with Ph+ CML relapsing after allogeneic HSCT? a) Single or repeated donor leukocyte infusions . . . . . . . . . . . . . . . . . . b) Inhibitors of tyrosine kinase. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Second allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Initial treatment with inhibitors of tyrosine kinase followed by single or repeated donor leukocyte infusions . . . . . . . . . . . . . . . . 4. Which is the most important cause of treatment failure in children with 1st chronic phase Ph+ CML? a) Relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Treatment-related mortality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Equally both relapse and treatment-related mortality . . . . . . . . . . . . . d) Secondary malignancies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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5. Which is the chance of definitive cure with allogeneic HSCT in children with myelofibrosis and polycythaemia vera? a) 90% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 70% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 50% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) No sound data are available for analysing the role of HSCT in children affected by these disorders. . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 39
HSCT for lymphomas in children
M. Miano, G. Dini
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CHAPTER 39 • Lymphomas in children
1. Introduction The prognosis for children suffering from Hodgkin’s disease (HD) and non Hodgkin’s lymphoma (NHL) is good, even in patients with advanced disease (1–3). Patients who fail to respond to first line chemotherapy and radiotherapy, or who present with recurrent disease can achieve long-term, disease free survival (DFS) after autologous haematopoietic stem cell transplantation (HSCT) (4). This procedure has historically been preferred to allogeneic HSCT due to its greater availability and the absence of immunologic complications. Nevertheless, allogeneic HSCT has been attempted for the treatment of both diseases on a number of occasions, but data concerning paediatric settings are scarce. During the last few years, the high rate of transplantrelated mortality (TRM), which is also due to the advanced disease status of the patients undergoing allogeneic HSCT, has limited the use of this procedure in these patients. New approaches have been developed and adopted to reduce TRM, including the use of reduced-intensity conditioning regimens, which rely on the potential graft-versus-lymphoma (GvL) effect rather than on the conditioning regimen itself.
2. Indications Recently, the European Group for Blood and Marrow Transplantation (EBMT) published the indications for HSCT in all diseases, including lymphomas. These are shown in Table 1.
3. Non-Hodgkin’s lymphomas 3.1. Role and outcome of autologous and allogeneic HSCT There are few published reports on exclusively paediatric populations suffering from NHL undergoing HSCT. In a study of 46 paediatric patients (5), no differences were shown in terms of event free survival (EFS) (57%) between the groups of patients receiving autologous (32 pts) or allogeneic (14 pts) HSCT. Three out of 32 (9%) and 3/14 children (21%) undergoing autologous and allogeneic HSCT, respectively, died of transplant related toxicity. Currently, the true impact of allogeneic HSCT in children with NHL remains to be clarified. One study of 10 children with poor prognosis NHL treated by allogeneic HSCT showed an overall survival (OS) probability of 56% at a median follow-up of 29 months (6). An EBMT retrospective study reported data on 136 children receiving allogeneic transplantation for NHL (7). At a median follow-up of 1.2 years, 51% of patients were alive. OS was influenced by status at HSCT. Thus, OS was 74% for patients transplanted in CR, and 56% for those with sensitive relapse while only
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Table 1: Indications for HSCT in children with lymphoma Disease
Status
Sibling donor
Well matched unrelated / 1 Ag related
Mm unrelated / >1 Ag related
Auto-HSCT
NHL
CR1 low-risk
GNR
GNR
GNR
GNR
CR1 high-risk
CO
CO
GNR
CO
CR2
S
S
CO
CO
CR1
GNR
GNR
GNR
GNR
Relapse, CR2
CO
D
GNR
S
HD
S: standard of care; generally indicated in suitable patients. CO: clinical option; can be carried out after careful assessment of risks and benefits. D: developmental; further trials are needed. GNR: generally not recommended. CR1, 2: first, second complete remission. Ag: antigen. Mm: mismatched
27% with refractory disease survived. Relapse occurred in 21 of the 70 (30%) large B-cell lymphoma, but in only 4 of 28 (14%) anaplastic large cell lymphoma (ALCL) patients. ALCL represents a special category among NH lymphomas. A very low treatment failure rate (3 year EFS 75%, and TRM 15%) was observed in a series of 20 children suffering from relapsed ALCL that had been treated by allogeneic HSCT. The effectiveness of allogeneic HSCT, even in patients with chemotherapy-resistant or progressive disease suggests a role of graft vs. ALCL effect (8). 3.1.1. Risk factors The only predictive factors for EFS are the type of lymphoma and the disease status at transplant, with significantly worse EFS being observed in patients with less favourable disease status.
4. Hodgkin’s disease 4.1. Role and outcome of autologous HSCT Children suffering from HD who are treated with chemotherapy alone or associated with radiotherapy currently have a 5-year DFS rate of over 85%. Patients who are resistant to first line therapy or who have recurrent disease need to undergo salvage treatment. In the adult setting, high-dose chemotherapy with autologous stem cell transplantation represents the standard treatment for this high-risk population. This approach is also frequently used in the paediatric population. A report of 53 children and adolescents undergoing autologous HSCT showed actuarial EFS of 31% and an OS of 43% (9).
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The results of another retrospective study on 51 children showed no statistically significant differences in OS in patients who were treated with autologous HSCT or with conventional salvage therapy. Nonetheless, patients with resistant disease showed a significant survival advantage with HSCT (10). Overall, these results justify the use of autologous HSCT as a primary therapeutic option for patients with resistant or relapsing disease. 4.1.1. Risk factors Pre-transplantation LDH levels represent an important prognostic factor for 5-year EFS: 42 vs. 0% for patients with normal or high levels, respectively. Pre-transplant positive positron emission tomography proved to be an indicator of poor prognosis in a large group of patients that included children (11). 4.2. Role and outcome of allogeneic HSCT Few data are currently available regarding the outcome of allogeneic HSCT in children with Hodgkin’s disease. All available data refer to adult populations. Increasing evidence supporting the concept of a GvL effect, as well as the gradual improvement in both TRM rates and in the non-myeloablative conditioning regimens support the idea that the role of allogeneic HSCT should be re-evaluated.
5. Conclusions Autologous HSCT represents a valid treatment option for patients suffering from lymphomas after relapse, or for those with resistant disease. The true impact of allogeneic transplantation has not yet been clarified. The gradual improvement in TRM together with the possibility of administering a reduced intensity conditioning regimen should lead to a re-evaluation of the use of allogeneic HSCT for high-risk children. Novel imaging and molecular techniques should be further investigated to better identify high-risk groups of patients who need more intensive treatment.
Acknowledgments M. Miano and G. Dini thank V. Perricone for revising the manuscript.
References 1. Poetter R. Paediatric Hodgkin disease. Eur J Cancer 1999; 35: 1466-1474. 2. Kurtzberg J, Graham ML. Non Hodgkin’s Lymphoma: Biologic classification and implication for therapy. Ped Clin North Am 1991; 38: 443-456.
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3. Sandlund JT, Downing JR, Crist WM. Non Hodgkin’s lymphoma in childhood. N Engl J Med 1996; 334: 1238-1248. 4. Baker KS, Gordon BG, Gross TG, et al. Autologous Haematopoietic Stem Cell Transplantation for relapsed or refractory Hodgkin’s disease in children and adolescents. J Clin Oncol 1994; 12: 2160-2166. 5. Bureo E, Ortega JJ, Munoz A, et al. Bone marrow transplantation in 46 pediatric patients with non-Hodgkin’s lymphoma. Bone Marrow Transplant 1995; 15: 353-359. 6. O’Leary M, Ramsey NKC, Nesbit ME, et al. Bone marrow transplantation for non-Hodgkin’s lymphoma in children and young adults. Am J Med 1983; 74: 497-501. 7. Claviez A, Tiemann M, Harousseau J, et al. Allogeneic hematopoietic stem cell transplantation in children with non-Hodgkin’s lymphoma: An EBMT study on 136 patients. Bone Marrow Transplant 2003; 31: S6. 8. Woessmann W, Peters C, Lenhard M, et al. Allogeneic Stem Cell Transplantation in relapsed or refractory anaplastic large cell lymphoma of children and adolescents – a BerlinFrankfurt-Munster group report. Br J Haematol 2006; 133: 176-182. 9. Baker KS, Gordon BG, Gross TG, et al. Autologous stem cell transplantation for relapsed or refractory Hodgkin’s disease in children and adolescents. J Clin Oncol 1999; 3: 825831. 10.Stoneham S, Ashley S, Pinkerton R, et al. Outcome after Autologous Hematopoietic Stem Cell Transplantation in relapsed or refractory childhood Hodgkin’s Disease. J Pediatr Hematol Oncol 2004; 26: 740-745. 11.Jabbour E, Hosing C, Ayers G. Pre-transplant positive positron emission tomography/gallium scans predict poor outcome in patients with recurrent/refractory Hodgkin Lymphoma. Cancer 2007; 109: 2481-2489.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The standard treatment for patients suffering from HL with responding relapsed disease is: a) Allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Radiotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. The main risk factor for EFS in patients receiving allogeneic HSCT for HL is: a) LDH levels before HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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b) Disease status . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Histology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) WBC count . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. The standard treatment for patients suffering from low risk NHL in 1st CR is: a) Allogeneic HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Chemotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Radiotherapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. The main risk factor for patients receiving autologous HSCT for Hodgkin’s lymphoma is: a) LDH levels before HSCT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Disease status . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Histology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) WBC count . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Which of the following is true of allogeneic HSCT for patients suffering from relapsed NHL: a) Generally indicated in suitable patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A clinical option that can be carried out after careful assessment of risks and benefits . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Is not recommended . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Indicated after relapse following autologous HSCT . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 40
HSCT for children with severe combined immunodeficiencies (SCID)
M. Cavazzana-Calvo, W. Friedrich, A. Fischer
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1. Introduction Severe combined immunodeficiency (SCID) disorders result from genetically determined blocks in the T-lymphocyte differentiation programme. Overall, the incidence is estimated to be 1 in 75,000 births. There is considerable genetic heterogeneity as eleven different conditions all resulting in a SCID have been fully characterised (Table 1).
Table 1: SCID classification Mechanisms
Mutated gene
Inheritance
Affected cells
Premature cell death
ADA
AR
T, B, NK
Defective cytokine-dependent survival signalling
gc JAK3 IL7RA
X-L AR AR
T, NK T, NK T
Defective V(D)J rearrangement
RAG1 or RAG2 Artemis
AR AR
T, B T, B
Defective pre-TCR and TCR signalling
CD3 d, z, e CD45
AR AR
T T
AR: autosomal recessive; X-L: X-linked
Four main mechanisms have been described: a) Premature cell death caused by the accumulation of purine metabolites, as seen in adenosine deaminase (ADA) deficiency. b) Defective cytokine-dependent survival signaling in T-cell precursors (and sometimes NK cell precursors). This mechanism accounts for more than 50% of cases of SCID. Deficiency in expression or function of the gamma common (gc) cytokine receptor subunit shared by the receptors for IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 causes the X-linked form of SCID (SCID-X1), characterised by the complete absence of both T and NK lymphocytes (1). Deficiency in JAK3, which is normally associated with the cytoplasmic region of gc, results in an identical phenotype. c) Defective V(D)J rearrangements of the TCR and B-cell receptor genes. In our experience, this group accounts for 30% of SCID cases. Deficiency in either RAG1 or RAG2 (the lymphoid-specific recombination initiating elements) or artemis (a factor involved in the non-homologous end-joining repair pathway) leads to defective V(D)J rearrangements and thereby thymocyte and pre-B-cell death. d) Defective pre-TCR and TCR signalling. Pure T-cell deficiencies are caused by defects in either a CD3 subunit (such as CD3 d, e, or z) (2, 3) or the CD45 tyrosine phosphatase, key proteins involved in pre-TCR and/or TCR signalling at the positive selection stage.
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Some researchers include other T-cell immunodeficiencies in the SCID group, such as ZAP-70 deficiency, CD3g deficiency, human leukocyte antigen (HLA) class II expression deficiency, purine nucleoside phosphorylase deficiency, ligase IV or Cernunnos deficiencies, and Omenn’s syndrome. However, since these conditions are characterised by the presence of mature (though functionally defective) T cells, they raise very distinct issues as far as therapy is concerned (see below) and so are not considered in this review.
2. Clinical manifestations The clinical presentation of the different SCID conditions is fairly uniform and is characterised by the early onset of infections (usually in the respiratory tract and the gut). Common opportunistic organisms such as Pneumocystis carinii and Aspergillus and intracellular organisms such as Cytomegalovirus can cause recurrent infections and a failure to thrive. Live vaccines, such as Bacille Calmette-Guérin (BCG), can also cause life-threatening infections. The persistence and recurrence of infections in SCID patients rapidly leads to growth impairment and malnutrition. Non-infectious clinical manifestations consist mainly of graft-versus host disease (GvHD) caused by the patients’ inability to reject allogeneic cells. The two possible sources of allogeneic cells are maternal lymphocytes and transfusion. The severity of these clinical manifestations makes SCID a medical emergency that, in the absence of treatment, leads to death within the first year of life.
3. Diagnosis Diagnosis of SCID is easy when there is a family history of early death from infections and the symptoms described above. In most cases, clinical examination together with very simple tests can confirm a suspicion, i.e. lack of palpable lymph nodes, especially in the inguinal area, with absence of visible tonsils, absence of a thymic shadow on the chest X-ray film and lymphocytopenia. The latter is of great value in young children since normal absolute lymphocyte counts are high (around 6 x 109/L).
4. Allo-HSCT for SCID patients SCID is a paediatric emergency that needs to be treated as soon as possible once the diagnosis is confirmed. The treatment of choice is an allo-HSCT, which will provide the missing progenitor of T cells and allows a survival rate of more than 90% when carried out shortly after birth (4–6). Unfortunately, an early diagnosis is not always made and the survival rate is very variable, depending on a number of prognostic factors such as the clinical state at the time of diagnosis, in particular 538
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the presence of a lung infection. In the presence of an HLA genoidentical donor (20% of SCID patients), HSCT can be performed without any conditioning regimen and its course is characterised by the remarkable rarity of acute and chronic GvHD in the absence of any prophylaxis and by the rapid development of T-cell function post-transplant. HLA genotypically identical donor includes also one antigen (A or B) mismatched donor; in this last case, cyclosporin A will be used in the posttransplant period to prevent the occurrence of acute GvHD. The overall survival rate of SCID patients after HLA-genoidentical sibling transplant is good, and with recent advances in the management of severe infection management, now reaches over 90% (4). In the absence of a HLA genoidentical sibling, HSCT can be performed with: - a phenotypically identical family donor (in the case of consanguinity) or a phenoidentical cord blood - a matched unrelated donor (MUD) - HLA partially identical (haplo-identical) family donor. It should be noted that the time required to the search for a MUD is compatible only in the presence of a patient free of severe infections, in good clinical condition, waiting in a protective environment. In all these cases, the use of a conditioning regimen is recommended. This usually consists of busulfan (day –10 to day –7) (oral: 4 mg/kg/day x 4 or IV 3.2 mg/kg/day x 4) and cyclophosphamide (day –5 to day –2) (50 mg/kg/day x 4). Antithymoglobulin (ATG) will be added if a MUD is available; in this case, the GvHD prophylaxis consists in cyclosporin A. If an HLA partially identical family donor is used, the HSC harvest will be ex vivo manipulated to eliminate the mature T- and B-cells contaminating the graft; usually this is obtained by means a CD34+ selection. Cyclosporin A is not indicated except in the case of primary GvHD from maternalfoetal transfusion or Omenn syndrome. In the latter setting therapy/prophylaxis of GvHD is usually needed and should be continued for three months. Ideally, HSCT graft should contain 5 x 106 CD34/kg of body weight and less than 1 x 104 T-cells/kg, thus strongly limiting the risk of acute severe GvHD. The presence of a severe infection necessitates modification or complete omission of the conditioning regimen. A full conditioning regimen increases the chance of obtaining HSC engraftment and sustained T and B-cell reconstitution but because these regimens have considerable toxicity, research is needed to find less toxic myeloablative regimens. Patients are placed in a protective environment and usually receive prophylactic antimicrobial medication to eliminate the intestinal microflora, and IV Ig therapy weekly for three months after HSCT and then every three weeks up to the detection of B-cell function. Around 40% of all transplanted SCID forms require long-life Ig replacement because of absence of B-cell engraftment.
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The survival rate of patients treated by an haploidentical T-cell-depleted HSCT is less good than that reported for an HLA genoidentical donor, and ranges from 50 to 78% (4, 5). Again, results have improved over time (from 35% survival in patients transplanted before 1985 in Europe to 75% in those treated from 1996 to 1999) (4). Better prevention of GvHD and treatment of infections probably account for the improved survival rates. Three parameters play a role in determining overall survival rate: GvHD, graft rejection and kinetics of T-cell development, the latter being the major parameter affecting the different outcome after HLA partially identical HSCT. In the light of these partially unsatisfying clinical results, gene therapy protocols have been initiated in Europe to try to improve the clinical results for SCID patients lacking a genoidentical sibling donor.
5. Gene therapy trials for SCID The first clinical trial for SCID-X1 was initiated in 1999. The protocol has been described elsewhere (7) and is quite simple in its principle. Patients with no HLAidentical siblings were eligible, and no myeloablative treatment was given. Among the 10 treated patients with typical SCID-X1, T-cell development occurred in nine (7–9). A correlation between quality of T-cell reconstitution and number of transduced CD34+ cells infused was detected (Figure 1).
Figure 1: T-lymphocyte development after gene therapy for SCID-X1 9000
P4
T-lymphocytes/µl
7000
> 3x106 CD34+ gc+/kg
5000 P8
P7
3000 P6 P10
1000 P9
0 0
10
20
P2
P5
P1
1x106 CD34+ gc+/kg 30
40
50
60
70
80
90
100
Months after gene therapy
P1 Æ10: each curve represents T-cell counts for a given patient. In P6 and 10, who received 1 x 106 g-chain (gc)+CD34+ cells/kg T-cell development was suboptimal (below the line) as compared to the other patients who received > 3 x 106 gc+CD34+ cells/kg
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Suboptimal T-cell development occurred in two patients who received 1 x 106 CD34+ gc+/kg, whereas the T-cell pool fully developed in those who received >3 x 106 CD34+ gc+/kg, providing a minimal threshold of total number of cells to inject. These results have since been confirmed in 10 additional patients treated by Thrasher et al. (10) in London with a similar protocol. Until now, correction of immunodeficiency, which also includes in part B-cell immunity, is good enough so that patients cope normally with infections, including those caused by VZV, and live normally without therapy. Efficacy of gene therapy also has now been reported in the treatment of 10 patients with ADA deficiency using similar methodology (11). Usage of a low-dose myeloablative therapy (busulfan, 4 mg/kg) may have increased the observed rate of myeloid cell transduction, thereby potentially improving long-term efficacy. These results open the door to an extension of gene therapy to other SCID conditions, as the selective growth advantage concept should apply, albeit with a variable intensity in different forms of SCID. There are some limitations to gene therapy of SCID. Failure to correct immunodeficiency occurred in a child with SCID-X1 who had an enlarged spleen caused by a disseminated BCG infection (8). It appeared from analysis of the spleen upon removal that transduced cells were probably trapped in the spleen, thus impairing T-cell differentiation in the thymus. Therefore, such (rare) patients might require splenectomy prior to be eligible for gene therapy. Gene therapy in older SCID-X1 patients, either patients with an incomplete phenotype or a patient in whom HSCT at least partially failed, did not succeed. This failure is likely the consequence of the premature loss of thymic function in SCID patients in the absence of functional T-cell precursor cells. Four patients from our SCID-X1 gene therapy trial developed clonal T-cell proliferation, as described in detail elsewhere (9). In two of these clonal proliferations, the primary cause was insertion of the provirus within the LMO-2 locus, leading to aberrant expression of LMO-2 in mature T cells and thereby uncontrolled proliferation. It was found that there is a higher risk of this serious adverse effect occurring in the context of SCID-X1, as also suggested in a murine model of leukaemogenesis. Further application of gene therapy in SCID nevertheless should try to use potentially safer vectors, including self-inactivated long terminal repeats and perhaps insulators and a rescue “suicide” gene. Note While writing this Chapter, one case of leukaemia-like lymphoproliferative disease occurred in the English trial.
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References 1. Notarangelo LD, Giliani S, Mazza C, et al. Of genes and phenotypes: The immunological and molecular spectrum of combined immune deficiency. Defects of the gamma(c)-JAK3 signaling pathway as a model. Immunol Rev 2000; 178: 39-48. 2. de Saint Basile G, Geissmann F, Flori E, et al. Severe combined immunodeficiency caused by deficiency in either the delta or the epsilon subunit of CD3. J Clin Invest 2004; 114: 1512-1517. 3. Rieux-Laucat F, Hivroz C, Lim A, et al. Inherited and somatic CD3zeta mutations in a patient with T-cell deficiency. N Engl J Med 2006; 354: 1913-1921. 4. Antoine C, Muller S, Cant A. Long-term survival and transplantation of haematopoietic stem cells for immunodeficiencies: Report of the European experience 1968-1999. Lancet 2003; 361: 553-560. 5. Buckley RH, Schiff SE, Schiff RI. Hematopoietic stem-cell transplantation for the treatment of severe combine immunodeficiency. N Engl J Med 1999; 340: 508-516. 6. Bertrand Y, Landais P, Friedrich W. Influence of severe combine immunodeficiency phenotype on the outcome of HLA non-identical, T cell depleted bone marrow transplantation: a retrospective European survey from the European group for bone marrow transplantation and the European society for immunodeficiency. J Pediatr 1999; 134: 740-748. 7. Cavazzana-Calvo M, Hacein-Bey S, de Saint-Basile G, et al. Gene Therapy of human severe combined immunodeficiency (SCID)-X1 disease. Science 2000; 288: 669-672. 8. Hacein-Bey S, Le Deist F, Carlier F, et al. Correction of human X-linked severe combined immunodeficiency (SCID-X1) by ex-vivo gene therapy. N Engl J Med 2002; 346: 1185-1193. 9. Hacein-Bey-Abina S, von Kalle C, Schmidt M, et al. LMO2-associated T-cell proliferation in two patients after gene therapy for SCID-X1. Science 2003; 302; 415-419. 10.Gaspar HB, Parsley KL, Howe S, et al. Gene therapy of X-linked severe combined immunodeficiency by use of a pseudotyped gammaretroviral vector. Lancet 2004; 364: 2181-2187. 11.Aiuti A, Slavin S, Aker M, et al. Correction of ADA-SCID by stem cell gene therapy combined with nonmyeloablative conditioning. Science 2002; 296: 2410-2413.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The common characteristic of the different forms of SCID is: a) Absence of circulating white blood cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Absence of T- and B-cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Absence of T- and NK cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Absence of mature T-cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) Leukopenia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 542
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2. SCID forms are primary immunodeficiencies and the short-term prognosis without treatment is: a) A few days . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) A few weeks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Some months . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 1 year . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) A few years . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. The search for a MUD is indicated in the presence of: a) Severe lung infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Generalised BCG infection. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) CMV infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Adenovirus infection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) No opportunistic infections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4. The outcome of an haploidentical HSCT is heavily influenced by: a) The frequency of rejection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) The occurrence of GvHD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) The occurrence of VOD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) A long lasting immunodeficiency . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) The frequency of pulmonary acute hypertension . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5. The major risk factor linked to gene therapy is: a) Failure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Partial immunological reconstitution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Age of patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Insertional mutagenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e) Splenomegaly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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CHAPTER 41
HSCT for inborn errors of metabolism
J.J. Boelens, R.F. Wynn, M. Bierings
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CHAPTER 41 • HSCT for IEM
1. Introduction For the past two decades, HSCT has been used as an effective therapy for selected inborn errors of metabolism (IEM). This group of conditions consists of several lysosomal storage diseases (LSDs) and peroxisomal disorders, osteopetrosis and a variety of rare miscellaneous disorders (e.g. purine nucleoside phosphorylase- and adenosine deaminase deficiency, mevalonic aciduria). Frantantoni and Neufeld laid the foundation for our understanding of transferable lysosomal enzymes by demonstrating cross-correction of metabolic defects in cocultures of fibroblasts from Hurler and Hunter syndrome patients. These results provided the rationale for Hobbs to trial HSCT in a Hurler patient in the early eighties (1). Dramatic improvement in the clinical phenotype of this first patient resulted in over 1000 HSCTs for various IEM (>20 diseases/disorders) worldwide. Unfortunately, for unknown reasons not all LSDs benefit from HSCT, and therefore careful evaluation of the effect of HSCT is important to make clear guidelines on indications for HSCT. In addition, success has been limited by high rates of graftfailure (15–75%), transplant-related-mortality and absence of instant availability of unrelated donors for rapidly progressive diseases (2, 3). Risk-factor analyses and improved transplantation techniques including the rapid availability of umbilical cord blood (UCB) have resulted in less graft-failure and less transplantrelated mortality. In osteopetrosis HSCT can correct the disease by providing haematopoietic stem cells as well as osteoclasts. Success is dependent on timing of the procedure, and possibly also depends on stem cell source as well as on the underlying genetic defect. In this Chapter, indications for HSCT are discussed and current guidelines of HSCT for IEM are summarised.
2. Lysosomal storage diseases and peroxisomal disorders 2.1. Indications Despite the fact that HSCT for IEM have been performed for more than 25 years, series of considerable size are only present for Hurler’s syndrome, infantile Krabbe Disease and X-ALD. In other disorders the effects and outcome of HSCT are difficult to assess because of the limited number of cases, wide range of clinical heterogeneity and the absence of a good functioning registry for proper long-term follow-up. If damage to the central nervous system (CNS) is present, this is irreversible and therefore a contra-indication for HSCT in all candidate diseases (4, 5). The indications and contra-indications are listed in Table 1. This guideline however should be interpreted in the context of progress in transplantation.
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2.2. Results Quality of life in successfully transplanted patients for certain indications, like Hurler’s syndrome (HS), seems encouraging. However the relatively high rates of graft-failure (15–75%) and transplant-related toxicity limits its success (2, 6). Since HS is the most frequent indication for HSCT within the LSD-group of IEM, HSCT for HS serves as a model for a risk factor analysis for graft failure. In multivariate analysis (n=146 HS-patients, transplanted between 1994–2004), T-cell depletion and reduced intensity conditioning were found to be serious risk-factors for graft failure (6). Busulfan-targeting (Therapeutic-Drug-Monitoring) on the other hand protected against graft failure. No difference in graft failure was noted between the different cell sources used (BM, PBSC or UCB). However, significantly (p=0.037) more patients receiving cord-blood achieved full-donor chimerism. This is in line with other studies (4, 7) using UCB in IEM and was once again observed in a recent ‘EUROCORDEBMT Working Party Inborn Errors’ survey (n=42): the EFS after 2001, was 84% using UCB, similar to that found in the same period using BM or PBSC. All UCB recipients however (except two with a donor chimerism of 90 and 94%) had full-donor chimerism and all had normal enzyme levels, in comparison to 60% in the BM/PBSC matched controls from the same period (publication pending). Although no large studies have examined the effect of lower enzyme levels after HSCT
Table 1: Guideline for indications (Peters et al. (2); Boelens et al. (3)) Indication
Disease Mucopolysaccharidoses (MPS) MPS I - Hurler - Hurler-Scheie MPS VI; Maroteaux-Lamy - Severe phenotypes MPS VII; Sly Other MPS Leukodystrophy X-linked adrenoleukodystrophy - Cerebral Metachromatic leukodystrophy - Juvenile subtype - “Late subtype” Globoid leukodystrophy - Early infantile subtype (Krabbe’s disease) - Late onset type
Yes No ** No ** Yes No
Yes In development*** Yes Yes Yes continue
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Disease
Indication
Other inborn errors of metabolism Fucosidosis a-mannosidosis Aspartylglucosaminuria Farber’s lipogranulomatosis Gangliosidosis - GM1 - GM2 Gaucher - Type I - Type III Mucolipidosis I Neuronal ceroid lipofuscinosis (NCL) - NCL 1 - NCL 2 Niemann-Pick - Type B - Type A and C Osteopetrosis
In development*** Yes In development*** In development*** In development*** No No ** No/ Yes * In development*** No No Yes In development***
Exclude neuronopathic osteopetrosis (e.g. in OSTM1) and carbonic anhydrase type II deficiency. Be cautious in case of mild or transient phenotype: discuss with experts
- Malignant infantile subtype - Wolmans disease Adenosine-deaminase-deficiency Purine-nucleoside-phosphatase-deficiency Mevalonic aciduria
Yes * Yes Yes Yes In development***
ERT: enzyme replacement therapy, MPS: Mucopolysaccharidosis. * No/Yes: depending on clinical phenotype, ** No: SCT is not indicated because of ERT, *** In development: not a clear indication, but is being evaluated in clinical and pre-clinical studies or single cases suggest efficacy. This guideline is applicable for patients who do not have any central nervous system (CNS) involvement or those who are only slightly affected. Advanced disease is always a contra-indication for HSCT. Since intravenous enzyme replacement therapy (ERT) does not cross the blood-brain-barrier, ERT is not a treatment option where there is CNS involvement. Donor derived monocytes do cross the blood brain barrier. This table is mainly based on patients receiving bone marrow or peripheral blood stem cells. Emerging stem cell sources, such as umbilical cord blood, as well as improvement of transplantation techniques, may in future extend the list
on long-term outcome parameters, there are recent suggestions that it does influence the neuro-cognitive outcome. Since cord-blood appeared to increase the likelihood of sustained engraftment resulting in full-donor chimerism and normal enzyme levels, cord-blood should be considered as a preferential stem cell source. Because the enzyme level is reduced in heterozygous family donors – and further reduced again if there is mixed donor chimerism after the transplant when using BM/PBSC, carrier donors will produce lower enzyme levels making them a less preferable donor, despite HLAHAEMATOPOIETIC STEM CELL TRANSPLANTATION
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Table 2: Donor hierarchy and conditioning a. Hierarchy of preferred stem cell source: 1. SIB/MFD (not carriers) 2. UD (10/10 allel match)* or unrelated cord blood (UCB: 6/6) 3. UCB (5/6 Ag match) 4. UCB (4/6 Ag match) or mismatched-UD (non-T-depleted) 5. UCB (3/6 Ag match) or HAPLO Notes: - UD (10/10) may be bypassed depending on institutional preference or because of time - For UD: either BM or PBSC (>10 x 106/kg) could be chosen - Cell dose for UCB: 5-6/6 match: >3.0 x 107 NC/kg and/or 2 x 105 CD34+/kg, 4/6-match >5 x 107 NC/kg and/or >3 x 105 CD34+/kg. Matching according to intermediate resolution criteria (low resolution on A and B, high resolution on DR) - Unrelated donors are regarded as non-carriers of the mutation - In case of osteopetrosis: currently no preference for unrelated marrow, cord blood or haploidentical transplant can be advised. Minimum cord blood stem cell dose as defined above should be used.
b. Pretransplant immunosuppression: - SIB: - UD/UCB:
no either Campath-1H 3 x 0.3 mg (day –9 to –7) or ATG-SangStat 4 x 2.5 mg/kg (day –4 to –1)
c. Conditioning: - SIB/UCB/UD: Busulfan 480 mg/m2 total dose (IV: day –9 to –6) with therapeutic drug monitoring AUC 20000 mg x h/L (cumulative per day) Cyclophosphamide 200 mg/kg total dose (day –5 to –2) -HAPLO: Bu/Cy + fludarabine 160 mg/m2 (4 x 40 mg) Note: For osteopetrosis the WP-IE of the EBMT is currently writing a protocol. The conditioning regimen will probably be busulfan (IV), fludarabine, +/- thiotepa based.
d. GvHD-prophylaxis: - SIB: CsA (+ MTX: 10 mg/m2; day +3, +6 and +11) - UD (BM): • with Campath-1H: CsA • with ATG: CsA + MTX (10 mg/m2; day +3, +6 and +11) - UD (PBSC) UD/Mismatched-UD (BM): CsA + MMF (30 mg/kg: stop day +28 if no GvHD) - UCB: CsA + prednisolone 1mg/kg (until day +28, taper in 2wks) In all cases aim for a CsA-trough level: 200 mg/L Tapering CsA-prophylaxis: - SIB/UD: CsA until day +50. Then taper 20% per week - UCB: CsA until + 6 months. Then taper over 3 months SIB: HLA matched sibling; MFD: matched family donor; HAPLO: haploidentical donor; UD: unrelated donor
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CHAPTER 41 • HSCT for IEM
matching (see guidelines EBMT: www.EBMT.org: Table 2). A study on the long-term outcome and the influence of mixed-chimerism/enzyme levels within the European series began in summer 2007. The observation of less mixed-chimerism in patients receiving UCB is intriguing. Possible factors contributing to this higher level of donor engraftment include better graft-versus-marrow (GvM) effect because of the greater mismatch, the greater proliferative potential of cord blood stem cells and the presence of mesenchymal cells in the cord blood that favourably influence engraftment. 2.3. Risk factors/morbidity In the early reports on HSCT of IEM (mainly HS) high rates of GvHD and idiopathic pneumonia syndrome (IPS) were reported. Within the EMBT study the incidence of GvHD and pulmonary complications was considerably lower, 16 and 8%, respectively. The incidence of chronic GvHD was only 5–15%. Since graft failure was common until recently second transplants were performed quite often. Survival after second transplant in the EBMT-series in Hurler’s syndrome (n=33) was over 80%. 2.4. Enzyme replacement therapy and SCT Enzyme replacement therapy (ERT) has become available for various indications (e.g. Gaucher, MPS-1, MPS-2, MPS-6). However for diseases with CNS involvement, like Hurler’s syndrome, HSCT remains the treatment of choice. In a study analysing the effects of ERT on EFS and transplant-related morbidity, patients receiving HSCT + ERT were compared with those receiving HSCT alone. Neither a positive nor a negative effect on EFS after receiving ERT prior to HSCT was noted, as compared to a historic cohort. However, in those patients in a poor clinical condition (e.g. cardiomyopathy, severe respiratory problems) prior to HSCT, ERT can be considered to improve the general clinical condition making them eligible for HSCT (8). 2.5. The future More detailed long term follow up outcome should be measured in this group of patients to define better guidelines regarding selection of patients eligible for transplantation as well as better guidelines for transplantation techniques. For Hurler’s syndrome such a study has been initiated recently. More than 150 successfully transplanted patients will be analysed for predefined long-term outcome parameters within the upcoming years. Neonatal screening might also influence the outcome. Early detection and transplantation might further influence the effect of HSCT, since HSCT in presymptomatic infants is safer than in symptomatic patients with organ damage.
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3. Osteopetrosis 3.1. Indications and results A study has recently been published proving the principle that TCIRG1 based osteopetrosis in mice can be corrected by introducing the gene in haematopoietic stem cells (9). Clinically, the largest series of HSCT for osteopetrosis reported to date is described by Driessen et al. (n=122: Transplants between 1980–2001) (10). The majority of patients (n=40) were transplanted with an HLA-identical sibling donor, resulting in a 5 yr DFS of 73%. For unrelated donor and haplo-identical transplants the DFS was less impressive, 40% and 24%, respectively. However, haploidenticaltransplants performed in experienced centres show a clear improvement over time. UCB appears to be a good alternative stem cell source: a recent EUROCORD-study on the outcome of UCB-transplantation (n=25) showed an “alive and engrafted”rate of about 50% (Bierings et al., EBMT abstract 2007). For indications and contra-indications in different geno- and phenotypes of the disease see Table 1. Functional outcome and the impact of various genetic subtypes have not been assessed successfully so far. 3.2. Risk factors and morbidity Transplant toxicity in HSCT for osteopetrosis is mainly caused by graft-failure, pulmonary complications and veno-occlusive disease (VOD). VOD can successfully be prevented by defibrotide (11). Pulmonary arterial hypertension is mainly seen in the first 3 months after HSCT, while outcome of treatment for pulmonary hypertension is generally poor. 3.3. The future Careful evaluation of long-term follow-up is of utmost importance. An analysis of the impact of different genotypes on outcome is currently ongoing. A new IV busulfan – fludarabine ± thiotepa regimen is being discussed within the EBMT-WP-IE.
4. Miscellaneous IEM In addition to the IEM discussed above, some miscellaneous IEM will profit from HSCT. These indications include adenosine deaminase (ADA) and purine-nucleosidephosphorylase deficiencies (PNP) resulting in a (severe) combined immunodeficiency due to storage of toxic metabolites in lymphocytes, leading to profound lymphopenia. For ADA-SCID more treatment options are available: PEG-ADA and gene-therapy. ADA and PNP are discussed in more detail in Chapter 40. Recently, a successful transplantation has been performed in mevalonic aciduria refractory to steroid and 550
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therapeutic monoclonal antibody. After successful sibling transplantation, HSCT resulted in almost normalisation of the excretion of mevalonic acid and in a long-term complete remission of the disease without any immunosuppressive medication.
5. Conclusion HSCT is an effective procedure for selected IEM. The procedure has become much safer with EFS rate of more than 80% in a-symptomatic/slightly affected LSD patients in recent years. Clear guidelines for indications, donor selections and conditioning regimens have been made recently. Umbilical cord blood has become the cell source of preference for several indications. Guidelines should be interpreted in the context of transplantation progress. Improvement of transplantation techniques and alternative therapies may change the recommended indications and contraindications for IEM.
References 1. Hobbs JR, Hugh-Jones K, Barrett AJ, et al. Reversal of clinical features of Hurler’s disease and biochemical improvement after treatment by bone-marrow transplantation. Lancet 1981; 2: 709-712. 2. Peters C, Steward CG. Hematopoietic cell transplantation for inherited metabolic diseases: an overview of outcomes and practice guidelines. Bone Marrow Transplantation 2003; 31: 229-239. 3. Boelens JJ. Trends in haematopoietic cell transplantation for inborn errors of metabolism. J Inherit Metab Dis 2006; 29: 413-420. 4. Escolar ML, Poe MD, Provenzale JM, et al. Transplantation of umbilical-cord blood in babies with infantile Krabbe’s disease. New Engl J Med 2005; 352: 2069-2081. 5. Peters C, Charnas LR, Tan Y, et al. Cerebral X-linked adrenoleukodystrophy: The international hematopoietic cell transplantation experience from 1982 to 1999. Blood 2004; 104: 881-888. 6. Boelens JJ, Wynn RF, O’meara A, et al. Outcomes of hematopoietic stem cell transplantation for Hurler’s syndrome in Europe: A risk factor analysis for graft failure. Bone Marrow Transplant 2007; 40:225-233. 7. Martin P, Carter S, Kernan N, et al. Results of the Cord Blood Transplantation Study (COBLT): Outcomes of Unrelated Donor Umbilical Cord Blood Transplantation in Pediatric Patients with Lysosomal and Peroxisomal Storage Diseases. Biology of Blood and Marrow Transplantation 2006; 12: 184-194. 8. Cox-Brinkman J, Boelens JJ, Wraith JE, et al. Haematopoietic cell transplantation (HCT) in combination with enzyme replacement therapy (ERT) in patients with Hurler syndrome. Bone Marrow Transplant 2006; 38: 17-21. 9. Johansson MK, de Vries TJ, Schoenmaker T, et al. Hematopoietic stem cell-targeted neonatal gene therapy reverses lethally progressive osteopetrosis in oc/oc mice. Blood 2007; 109:
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5178-5185. 10.Driessen GJ, Gerritsen EJ, Fischer A, et al. Long-term outcome of haematopoietic stem cell transplantation in autosomal recessive osteopetrosis: an EBMT report. Bone Marrow Transplant 2003; 32: 657-663. 11.Corbacioglu S, Honig M, Lahr G, et al. Stem cell transplantation in children with infantile osteopetrosis is associated with a high incidence of VOD, which could be prevented with defibrotide. Bone Marrow Transplant 2006; 38: 547-553.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. There are no treatment options for patients with an IEM: a) Correct statement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) HSCT is an effective treatment for all IEM . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) HSCT is a treatment option for a selected group of IEM. . . . . . . . . . . . . . . . . . . . d) HSCT is only effective in Hurler syndrome patients . . . . . . . . . . . . . . . . . . . . . . . . .
2. Which of the following statements about enzyme replacement therapy (ERT) is correct? a) ERT is a safer and more effective treatment option for IEM than HSCT . . . b) ERT is only effective in IEM without neurological involvement, since ERT does not cross the “blood brain barrier” . . . . . . . . . . . . . . . . . . . . . . . . . c) ERT is necessary pre-transplant to prevent transplantation related mortality and graft failure. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) ERT + HSCT is the most effective treatment for IEM for all indications . . .
3. Which of the following statements about graft failure in HSCT for IEM is correct? a) Historically, high graft failure rates were reported mainly associated with T-cell depleted grafts and “reduced intensity conditioning” . . . . . . . . . b) Graft failure has never been a problem in HSCT for IEM . . . . . . . . . . . . . . . . . . . . c) Not graft failure but TRM is the major problem in HSCT for IEM . . . . . . . . . . . d) Less graft failure is reported using unrelated cord blood as stem cell source . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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4. The period between diagnosis and HSCT is not important, the outcome depends on the degree of donor matching: a) No, the shorter the period between diagnosis and HSCT, the better the outcome . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Correct, you should not use mismatched grafts in IEM . . . . . . . . . . . . . . . . . . . . . c) Correct, do not use unrelated cord blood . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Correct, even neonatal screening will not improve outcome . . . . . . . . . . . . . . . 5. For every genotype of osteopetrosis HSCT is a potentially curative treatment option: a) Correct statement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) HSCT is not a treatment option for any genotype. . . . . . . . . . . . . . . . . . . . . . . . . . . c) HSCT is only a treatment option for selected genotypes . . . . . . . . . . . . . . . . . . . d) Not HSCT but ERT is currently the state of the art treatment for all genotypes of osteopetrosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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HSCT for hereditary bone marrow failure syndromes
E. Gluckman, J.E. Wagner
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CHAPTER 42 • Hereditary BMF syndromes
1. Introduction Aplastic anaemia is a rare disease in children. It is most commonly idiopathic and less often due to a hereditary disorder. However, hereditary bone marrow failure (BMF) syndromes should be considered in both children and adults before the institution of any therapeutic treatment plan. While new genetic tests are being developed, these are not widely available. Genomic instability in the presence of clastogenic agents is the hallmark of Fanconi anaemia (FA). In contrast, marked telomere dysregulation is characteristic of dyskeratosis congenita (DKC). Mutations affecting ribosome assembly and function are associated with DKC, Shwachman-Diamond syndrome, and Diamond-Blackfan anaemia (DBA) (1). The principal characteristics of these diseases are detailed in Table 1.
2. Fanconi anaemia (FA) FA is a rare autosomal recessive disease characterised by congenital abnormalities, progressive BMF, chromosome breakage, and cancer susceptibility. At least 13 genes have been involved in the disease; abnormalities in any of these gene products that normally interact disrupt the FA/BRCA biochemical pathway (2) (see Table 1). FA patients often have skeletal, thumb or limb abnormalities and abnormal skin pigmentation (café au lait spots). Other commonly involved organ systems include cardiac, renal system and auditory. Low birth weight and growth retardation are frequent (see Table 2). The haematological consequences of FA often develop in the first decade of life but absence of malformations and reversion due to somatic mosaicism can result in delayed or failed diagnosis in a small proportion of patients. Death, however, often results from the complications of BMF or occurrence of malignancy. The most frequent malignancy is AML with clonal cytogenetic abnormalities in the bone marrow; older patients are at high risk of squamous cell carcinomas of the oesophagus, head and neck and urogenital tract. 2.1. Disease-specific pre-HSCT work-up FA being a heterogeneous disease, clinical diagnosis is not always sufficient to assess the correct diagnosis in children or young adults with AA. Other constitutional AA may have similar congenital abnormalities and FA patients may have no abnormalities.
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Table 1: Principal characteristics of hereditary BMF syndromes Disease
Mutations
Diagnostic tests
Clinical expression
Fanconi anaemia
FANC A FANC B FANCC FANC D1/BRCA2 FANC D2 FANC E FANC F FANCG/XRCC9 FANCI FANCJ/BACH1/BRIP1 FANCL/PHF9/POG FANCM/Hef FANCN/PALB2
Mitomycin (MMC) or diexpoxybutane (DEB)induced chromosomal breakage in blood lymphocytes
Aplastic anaemia
Dyskeratosis congenita
Seckel syndrome
ShwachmanDiamond
Malformations: thumbs, cafe au lait spots, microcephaly, short stature, kidney, heart, etc.
Flow cytometry evidence of cycle arrest on exposure Malignancies: acute to MMC leukaemia (AML), squamous FANCD2 monoubiquination cell carcinoma head and neck, anogenital, liver for 90% of patients with abnormalities in the FA/BRCA2 pathway upstream of FANC D2
DCK1 TERC TERT
Telomere length
Aplastic anaemia
SCKL1 SCKL2 SCKL3
None
SBDS
Decreased serum trypsinogen and/or pancreatic isoamylase
Aplastic anaemia
Malformations Pigmentation, nail dystrophy, oral leukoplakia Malignancies: AML, solid tumours: oesophagus, gastro intestinal tract, bladder Aplastic anaemia Malformations: bird-head, dwarfism, mental retardation
Pancreatic exocrine deficiency
Kostmann
Neutrophil elastase ELA2
None
Chronic neutropenia, leukaemia
DiamondBlackfan
RSP19
Elevated adenosine deaminase (ADA)
Erythroblastopenia Malformations: thumb, renal cardiac
Congenital amegakaryocytic thrombocytopenia
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Amegakaryocytosis, aplastic anaemia
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CHAPTER 42 • Hereditary BMF syndromes
Table 2: Evaluating new onset cytopenia in children 6. BM aspiration with cytogenetics, BM biopsy if necessary 7. Chromosome breaks with DEB or MMC 8. Alfa foeto-protein 9. HLA typing 10. FANCD2
1. Family history 2. Malformations 3. Date of onset 4. Liver function tests 5. Blood counts
Diagnosis is suspected with:
• Blood
Diagnosis is confirmed with:
•
Other tests
New tests not for routine use
•
counts: pancytopenia with macrocytic anaemia Raised alfa-foetoprotein and haemoglobin F
PB lymphocyte cytogenetics with clastogenic agents: nitrogen mustard, DEB or MMC showing increased chromosome breaks with tri- and quadri-radial figures • Study of the cell cycle showing a G2/M arrest increased by incubation with clastogenic agents •
BM cytogenetic abnormalities for diagnosis of leukaemia or myelodysplastic abnormalities with abnormalities in chromosomes 1, 3, 7, 5, 8 and 11 being the most common
Ubiquitination of FANCD2: this test is specific and sensitive, if negative skin fibroblasts may be positive for FA and this confirms the existence of mosaicism • Identification of the complementation group with retroviral or lentiviral vectors • Sequencing and identification of the mutation. This test is useful for preimplantation diagnosis and perhaps for assessing the prognostic •
2.2. Results of allo-HSCT in FA HSCT is currently the only treatment that definitively restores normal haematopoiesis. FA anaemia cells are hypersensitive to DNA cross-linking agents. Cellular exposure to genotoxic agents including cyclophosphamide (Cy), busulfan (Bu) or irradiation increases chromosome breaks and tissue damage. Graft-versus-host disease (GvHD) induces severe tissue damage and absence of repair (3). Standard conditioning with
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high dose Cy and total body irradiation (TBI) as for patients with SAA must not be used. 2.2.1. HLA identical sibling transplant With the use of reduced intensity conditioning regimens, most reports over the past decade have demonstrated good results. In our series of 117 FA patients, conditioned with low dose CY and total lymphoid irradiation (TLI), 5-year survival was 85%. In general, most series have reported younger patient age, higher pre-transplant platelet counts, absence of previous treatment with androgens, normal pre-transplant liver function tests and limited malformations as factors associated with better survival after transplantation (4). Unfortunately, a rising risk of cancer has been observed in long-term survivors, particularly cancers of the head and neck (5). In an analysis of 700 patients with FA (n=79) or AA (n=621) treated with allogeneic HSCT in Seattle or Paris, the KaplanMeier estimate for developing any malignancy by 20 years after transplantation was 14% (6). Among patients with FA, a single hazard peak for solid tumours occurred between 8 and 9 years after HSCT. The Kaplan-Meier estimate of developing any malignancy by 20 years after HSCT was 42% (95%CI, 10–74%), with all being solid tumours. In the multivariate analysis of the 700 patients with marrow failure (FA and non-FA), the diagnosis of FA (relative risk [RR] 11.2, p=0.0001) and treatment with azathioprine (RR 11.7, p=0.0001) were independent predictors of secondary malignancy. Absence of irradiation in the conditioning regimen did not abolish the risk of secondary tumours. As solid tumours occur in FA patients without prior exposure to chemotherapy and radiation, it is clear that cancer is at least in part related to the specific genetic defect present and environment, as shown by different phenotypic expression of the disease in homozygous twins. In an attempt to reduce the potential impact of irradiation and GvHD on the risk of late effects, including cancer, newer regimens have replaced TLI with fludarabine in combination with low dose CY as well as used T-cell depletion to reduce the risk of GvHD. While limited in number and follow-up, early results are encouraging (7). It is too early to tell if there is any impact upon cancer risk at this time. Nonetheless, the good results of HLA-identical sibling HSCT raise several questions regarding the optimal timing of BMT and the best conditioning regimen. Concerning the former, there is a general agreement that HLA-identical sibling HSCT should be performed as first-line therapy, without first using androgens or corticosteroids, which have considerable side effects. When blood counts fulfil the criteria of severe AA (Hb <8 g/dL, neutrophils <0.5 x 109/L, or platelets <20 x 109/L), transfusions and infections are more likely, making this a suitable time to perform HSCT. During the 558
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CHAPTER 42 • Hereditary BMF syndromes
waiting period, it is important to regularly perform bone marrow aspiration and cytogenetic analysis for detection of clonal abnormalities or leukaemic transformation. Results show that transplants performed late (after a long period of aplasia or during leukaemic transformation) are associated with markedly poorer results. Most patients treated for acute leukaemia do not tolerate standard dose chemotherapy and have a very poor prognosis, although some long-term survivors have been reported after HSCT. In terms of the best conditioning regimen, the aims are to 1) avoid rejection in a population of patients who have received multiple transfusions, 2) limit early and late toxicities and 3) minimise risk of GvHD. Several combinations have been used in a limited number of patients. Fludarabine containing regimens, in combination with low dose CY or Bu and ATG, appear to be well tolerated. GvHD must be prevented, as it is more likely to be severe in FA patients because of the underlying DNA repair defect and because lichen planus lesions associated with chronic GvHD may be a precursor of squamous cell carcinoma. Cyclosporin A (CsA) and mycophenolate mofetil (MMF) are being used more frequently as methods of GvHD prophylaxis with some groups incorporating T-cell depletion of sibling donor marrow. 2.2.2. Results of unrelated donor adult volunteer HSCT The EBMT working party on AA has analysed the outcome of alternative donor HSCT in 67 FA patients. The median 2-year survival was 28±8%. Causes of death included infection, haemorrhage, acute GvHD, chronic GvHD, hepatic veno-occlusive disease (VOD), and multi organ failure (MOF) (8). The CIBMTR analysed data from 98 patients transplanted with unrelated donor marrow (excluding those with peripheral blood or umbilical cord blood [CB] grafts) between 1990 and 2003 (9). Probabilities of neutrophil (89 vs. 69%, p=0.02) and platelet recovery (74 vs. 23%, p<0.001) were higher after fludarabine (Flu) than non-Flu containing regimens. Risks of acute (RR 2.95, p=0.003) and chronic GvHD (RR 3.30, p=0.03) were significantly higher in recipients of non T-cell depleted than T-cell depleted grafts. Day-100 mortality rate was significantly higher after non-Flu than Flu containing regimens (65 vs. 24%, respectively p<0.001). Corresponding 3-year adjusted overall survival rates were 13% vs. 52% (p<0.001) with best survival in those treated a Flu-based regimen (Figure 1). In addition, mortality was higher in recipients who were older (>10 years), CMV seropositive and who had received >20 blood product transfusions pre-transplant. Based on these results significant changes in practice were suggested: use of a Flu-containing conditioning regimen in the context of T-cell depleted marrow allografts and earlier referral with transplantation prior to excessive transfusions.
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Figure 1: Impact of fludarabine on survival in patients with FA treated with unrelated donor HSCT 100
Adjusted Probability, %
80 FLU (n=46)
60
40
20 Non FLU (n=52) 0 Months: 0 FLU: Non FLU:
12
24
36
(26) (9)
(25) (9)
(15) (7)
2.2.3. Results of related and unrelated donor CB transplantation (CBT) In the circumstance where an HLA identical sibling donor is available, CBT and BMT give similar results in terms of survival. However, reports comparing the two demonstrate a reduction in the frequency and severity of acute and chronic GvHD related to the relative immaturity of neonatal T-cells. However, the majority of patients do not have an HLA matched sibling donor. Eurocord analysed the results of unrelated CBT in 93 FA patients (10). The incidence of neutrophil recovery at 60 days was 60±5%. In addition to high cell dose, Flucontaining regimens (as in marrow recipients) (9) were associated with better neutrophil engraftment. The incidence of acute and chronic GvHD was 32.5% and 16%, respectively. Overall survival was 40±5%. In multivariate analysis factors associated with favourable outcome were use of Flu, high number of cells infused and negative recipient CMV serology. To date, there has been no formal comparison of outcomes in recipients of unrelated CB and marrow. However, results demonstrate that Flu is associated with better survival regardless of stem cell source in patients with FA. This suggests that Flu, a potent immune suppressive agent, enhances engraftment without paying the price 560
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of extra-medullary toxicity. In the future, studies may help us determine the place of CB. For now, CB is clearly indicated in those FA patients for whom an HLA-A, B, C, DRB1 allele-matched unrelated volunteer donor cannot be identified. Preimplantation genetic diagnosis (PGD) to select an embryo produced by in vitro fertilisation that is both unaffected by a heritable genetic disease and HLA identical to the affected recipient has been performed. Clearly, this approach is controversial (11) with marked differences in its acceptance by different countries worldwide. Globally, the strategy has been most often used for couples at high risk of having children with thalassaemia. However, the first successful use of PGD for a specific HLA type was for a child with FA. With this approach, the couple can avoid the risk of having additional affected children (and the consequent consideration of abortion) and also have a healthy child that will be an HLA identical match with the existing child needing HSCT. In these cases, it is typical for the CB to be collected at birth, eliminating risk to the newborn child. In the U.S., the use of the technology is expanding. To date, 5 transplants have been successfully performed in patients with FA. 2.2.4. Post-HSCT monitoring in FA Patients with FA require particular attention because of their sensitivity to toxic agents, various organ dysfunctions due to congenital malformations and increased risk of developing malignancies. This should include at least yearly endocrinological and growth follow-up, bone marrow cytogenetic and oral follow-up. Patients with oral lichen planus should be biopsied regularly and lesions removed.
3. Other congenital cytopenias 3.1. Dyskeratosis congenita Dyskeratosis congenita (DKC), also known as Zinsser-Engman-Cole syndrome, is a rare, progressive bone marrow failure syndrome characterised by the triad of reticulated skin hyperpigmentation, nail dystrophy, and oral leukoplakia. Evidence exists for telomerase dysfunction, ribosome deficiency, and protein synthesis dysfunction in this disorder. Early mortality is often associated with bone marrow failure, infections, fatal pulmonary complications, or malignancy. Results of HSCT are disappointing because of severe late effects including diffuse vasculitis and lung fibrosis (9). Conditioning with a Flu-containing non-myeloablative regimen may give better shortterm results but may not delay fatal outcome due to multi-organ failure related to the underlying genetic defect.
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3.2. Seckel syndrome Seckel syndrome is a rare autosomal recessive disorder with growth retardation, microcephaly with mental retardation and a characteristic bird-headed facial appearance. Two loci have been identified. Very few transplants have been reported in the literature. In one case late pulmonary fibrosis occurring 2 years after transplant was the cause of death despite an early favourable outcome. 3.3. Shwachman-Diamond syndrome Shwachman-Diamond syndrome is an autosomal recessive disorder with clinical features that include pancreatic exocrine insufficiency, skeletal abnormalities and pancytopenia (10). AML transformation has been observed. Very few patients have been treated by allogeneic HSCT. 3.4. Diamond–Blackfan anaemia DBA is characterised by chronic constitutional aregenerative anaemia with absent or decreased erythroid precursors in the BM. Both autosomal dominant and recessive inheritance are described. Most patients present with anaemia in the neonatal period or in infancy. Approximately 30% patients have a variety of physical anomalies including thumb, upper limb, craniofacial, heart and urogenital malformations. Usually, the patients are treated with transfusions and steroids and at least 50% patients respond. Allogeneic HSCT is an option in steroid resistant patients. In a report from the DBA Registry, 354 patients were registered and 20 were transplanted. The 5-year OS for HLA identical sibling transplants was 87.5%. Results were poor with alternative donors. CIBMTR reported 61 patients transplanted from 1984 to 2000. Most patients (67%) were transplanted with HLA identical donors. The 3 year probability of overall survival was 64%. Results were better in HLA identical sibling transplants (11). 3.5. Kostmann syndrome Kostmann syndrome is an inherited disorder with severe neutropenia and early onset of severe bacterial infections. More than 90% of the patients respond to r-HuG-CSF but approximately 10% will develop MDS/AML, regardless of their treatment or response. Allo-HSCT is the treatment of choice in patients refractory to G-CSF or with acute leukaemia (12). In the French chronic neutropenia registry including 101 patients, 9 patients were transplanted, 7 with an unrelated donor and 2 with an HLA identical sibling donor. Four patients had acute leukaemia, 4 were refractory to GCSF and 1 had BMF. The OS at 5 years was 61% indicating that HSCT should be considered in these patients even if there is no HLA identical sibling. 562
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3.6. Inherited thrombocytopenia 3.6.1. Congenital amegakaryocytic thrombocytopenia Affected infants are identified within days or weeks of birth. Transmission is autosomal recessive. Despite optimal supportive care, AA develops leading to death in the absence of HSCT, which is the only chance of cure in this disease. 3.6.2. Thrombocytopenia with absent radii TAR syndrome includes shortened or absent forearms due to the absence of development of the bilateral radii, associated with severe thrombocytopenia at birth. Skeletal anomalies are also seen in other bones but do not affect the hands and fingers. Usually the degree of thrombocytopenia is greatest at birth requiring transfusions, however thrombocytopenia becomes less severe during the first year of life and most patients will not require platelet transfusion after infancy. HSCT is not recommended. 3.7. Other rare inherited BMF syndromes Nijmegen breakage syndrome (NBS) is a rare autosomal recessive condition of chromosomal instability that is clinically characterised by microcephaly, a distinct facial appearance, short stature, immunodeficiency, radiation sensitivity, and a strong predisposition to lymphoid malignancy. Mutations in the NBS1 gene located in band 8q21 are responsible for NBS. Pearson syndrome is currently recognised as a rare, multisystemic, mitochondrial cytopathy. Its features are refractory sideroblastic anaemia, pancytopenia, defective oxidative phosphorylation, exocrine pancreatic insufficiency, and variable hepatic, renal, and endocrine failure. Death often occurs in infancy or early childhood due to infection or metabolic crisis. Patients may recover from the refractory anaemia. Older survivors have Kearns-Sayre syndrome (KSS), which is a mitochondropathy characterised by progressive external ophthalmoplegia and weakness of skeletal muscle. DNA ligase IV deficiencies due to mutation of LIG4 gene is a rare disease; one case of successful transplant has been described.
References 1. Shinamura A. Inherited bone marrow failure syndrome: molecular features. American Society Education program book. 2006; pp. 65-71. 2. Tanigushi T, D’Andrea AD. Molecular pathogenesis of Fanconi anemia: Recent progress. 2006; 107: 4223-4233. 3. Guardiola P, Socié G, Li X, et al. Acute graft versus host disease in patients with Fanconi
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anemia or acquired aplastic anemia undergoing bone marrow transplantation from HLA identical sibling donors: Risk factors and influence on outcome. Blood 2004; 103: 73-77. 4. Socié G, Devergie A, Girinski T, et al. Transplantation for Fanconi’s anemia: Long-term followup of fifty patients transplanted from a sibling donor after low-dose Cyclophosphamide and thoraco-abdominal irradiation for conditioning. Br J Haematol 1998; 103: 249–255. 5. Rosenberg PS, Greene MH, Alter BP. Cancer incidence in persons with Fanconi anemia Blood 2003; 101: 822-826. 6. Guardiola P, Pasquini R, Dokal I, et al. Outcome of 69 allogeneic stem cell transplants for Fanconi anemia using HLA- matched unrelated donors: A study of the European Group for Blood and Marrow Transplantation. Blood 2000; 95: 422-429. 7. Wagner JE, Eapen M, Mac Millan ML, et al. Unrelated donor bone marrow transplantation fro the treatment of Fanconi anemia. Blood 2007; 109: 2256-2262. 8. Gluckman E, Rocha V, Ionescu I, et al. on behalf of Eurocord Netcord and EBMT. Results of unrelated cord blood transplant in Fanconi anemia patients: Risk factor analysis of engraftment and survival. BBMT 2007; 13: 1073-1082. 9. Rocha V, Devergie A, Socie G, et al. Unusual complications after bone marrow transplantation for dyskeratosis congenita. Br J Haematol 1998; 103: 243-248. 10.Dror Y, Freedman MH. Shwachman-Diamond syndrome. Br J Haematol 2002; 118: 701-713. 11.Roy V, Perez WS, Eapen M, et al. Non malignant marrow disorders working committee of the international bone marrow transplant registry. Bone marrow transplantation for Diamond-Blackfan anemia. Biol Blood Marrow Transplant 2005; 11: 600-608. 12.Zeitler C, Welte K, Barak Y, et al. Stem cell transplantation in patients with severe congenital neutropenia without evidence of leukemic transformation. Blood 2000; 95: 1195-1198.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. Fanconi anaemia is diagnosed by the finding of which one of the following: a) Presence of malformations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Aplastic anaemia. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Bone marrow biopsy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Cytogenetics in PBL with clastogenic agents. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Presence of bone marrow failure with malformations, leukoplakia and nail dystrophy is more likely to be which of the following? a) Fanconi anaemia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Dyskeratosis congenita . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 564
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c) Shwachman syndrome . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Diamond-Blackfan syndrome . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. The best first line therapeutic option in Kostmann disease is: a) Supportive care . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Antibiotics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) G-CSF. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Haematopoietic stem cell transplantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Prior to initiating treatment of an adult with aplastic anaemia, which tests should be done? a) DEB or MMC to rule out FA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Bone marrow aspirate with cytogenetic evaluation . . . . . . . . . . . . . . . . . . . . . . . . . c) HLA type family members and patient . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Perform molecular testing to rule out Shwachman-Diamond syndrome and DBA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. In Fanconi anaemia conditioning for HSCT must be reduced because: a) Aplastic anaemia is not severe . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Hypersensitivity to clastogenic agents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Age of the patients . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Absence of leukaemia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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*
CHAPTER 43
HSCT in the haemoglobinopathies
I.A.G. Roberts, J. de la Fuente
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CHAPTER 43 • Haemoglobinopathies
1. Introduction Although advances in iron chelation therapy, transfusion and supportive care have dramatically improved life expectancy in thalassaemia major and sickle cell disease (SCD), patients continue to suffer disabling symptoms, particularly in adulthood, and most die prematurely from complications of the disorders and/or their treatment. HSCT is the only proven cure, effecting both resolution of symptoms and freedom from life-long, emotionally- and physically-demanding treatment (1, 2). The decision to undergo HSCT is often difficult since these disorders are not usually immediately life-threatening.
2. Indications for HSCT in thalassaemia (Table 1) All transfusion-dependent children with HLA-identical family members are eligible for HSCT, including the rare cases of a-thalassaemia major. For patients with HbE/b-thalassaemia or thalassaemia intermedia, treatment must be individually tailored, as these diseases are heterogeneous: severely affected patients are transfusion-dependent and can benefit from HSCT. Improving medical treatment and inferior outcome with unrelated donors and in adults means that HSCT for these patients should only be considered in carefully selected patients and the procedure carried out in centres experienced in management of haemoglobinopathies. We advise delaying HSCT until both patient and donor are ≥2 years.
Table 1: Indications for HSCT in thalassaemias 1. Definite indication for HSCT: - Transfusion-dependent a- or b-thalassaemia major; transfusion-dependent HbE/bthalassaemia - Age £16 years - HLA-identical family donor 2. Candidates who may be considered for HSCT in special circumstances: - Transfusion-dependent thalassaemia major in adults aged 17–35 years - Thalassaemia relapsing after previous HSCT - Transfusion-dependent S-b0 thalassaemia - Thalassaemia intermedia
3. Indications for HSCT in SCD (Table 2) Most clinicians offer HSCT only to patients with specific complications of SCD which predict for a poor prognosis. Many also offer HSCT to families wishing to return to countries without reliable access to good medical care. Pre-HSCT assessment must
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include neurological assessment (MRI/MRA, transcranial Doppler studies, neurocognition) and may reveal unanticipated severe cerebral vasculopathy (“moya moya”), which increases TRM and morbidity; such cases must be discussed with experts in SCD before proceeding to HSCT.
Table 2: Indications for BMT in sickle cell disease (SCD) 1. Definite indication for HSCT: - One or more of the following clinical complications: • SCD-related neurological deficit, stroke or subarachnoid haemorrhage • Recurrent acute sickle chest syndrome (>2 episodes) which has failed to respond to a trial of hydroxycarbamide of at least 6 months or where hydroxycarbamide is contraindicated • Recurrent, severe debilitating pain due to vaso-occlusive crises which has failed to respond to a trial of hydroxycarbamide of at least 6 months or where hydroxycarbamide is contraindicated - Age £16 years - HLA-matched family donor 2. Candidates who may be considered for HSCT in special circumstances: - Problems relating to future medical care e.g. unavailability of adequately screened blood products - SCD relapsing after previous HSCT - Transfusion-dependent S-b0 thalassaemia - Adults aged 17–35 years (as part of clinical trial) Modified from the British Paediatric Haematology Forum Criteria (Amrolia et al., 2003a)
4. Conditioning regimens for haemoglobinopathies (Table 3) The most commonly employed myeloablative regimens use oral busulfan (14–16 mg/kg) and intravenous cyclophosphamide (200 mg/kg); thiotepa, fludarabine or melphalan may be added to reduce graft rejection but their role is unproven (1, 2). Some groups, including ours, add ATG or alemtuzumab to pre-HSCT conditioning to reduce graft rejection with encouraging results, although it is important not to employ the high doses used to prevent GvHD (3). Hypertransfusion (maintaining Hb >13 g/dL) for 6 weeks prior to and after HSCT also reduces graft rejection. Reduced cyclophosphamide (120 mg/kg) is recommended for Pesaro Class 3 patients (see “Prognostic factors”) to avoid high TRM but must be combined with other measures to prevent graft rejection (e.g. ATG, red cell hypertransfusion, fludarabine). Most groups use cyclosporin/methotrexate as GvHD prophylaxis; many now use low-dose methotrexate (2 doses of 10 mg/m2) as graft rejection is less (3, 4). Busulfan dose-adjustment via blood levels has no significant impact on OS or EFS. Methotrexate is generally omitted in cord blood transplants (CBT) due to increased risk of graft rejection (5). 568
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CHAPTER 43 • Haemoglobinopathies
Table 3: Conditioning regimens for haemoglobinopathies Drug
Total dose
Schedule of administration
Alternatives
Busulfan
14–16 mg/kg
1 mg/kg orally per dose; usually given day -9 to -6
iv busulfan treosulfan
Cyclophosphamide
200 mg/kg
50 mg/kg iv per dose usually given day -5 to -2 Give Mesna (twice cyclophosphamide dose)/ Use lower dose (120–160 mg/kg) in Pesaro Class 3
Alemtuzumab
0.3 mg/kg
0.1 mg/kg iv per dose usually days -8 to -6
ATG fludarabine melphalan thiotepa
5. Results of HSCT for thalassaemias 5.1. Role of HSCT in thalassaemia The aim of HSCT is to improve long-term survival and/or quality of life (QoL). There are no controlled survival or QoL trials of HSCT versus medical treatment in thalassaemia. However, recent data show that patients who comply with iron chelation, and have no cardiac or liver damage, survive into their fifth decade (6). Since TRM is 2–5% and the risk of graft failure is ≤5%, such patients have a predicted survival of around 90–95% to age 40 with either therapeutic approach and the decision to proceed to HSCT should be based on QoL, i.e. the perceived benefit of freedom from life-long transfusions, chelation and their long-term complications. By contrast, for patients with poor compliance, few survive to age 40 and for them HSCT offers not only improved QoL, but also increased long-term survival (6). However, it is often very difficult to predict which children will accumulate iron in their organs, sometimes despite adequate chelation, and the results of transplantation before significant iron accumulation and organ damage has occurred can assure long term cure. 5.2. Prognostic factors (Tables 4 and 5) 5.2.1. Pesaro class Analysis of the first large series of patients undergoing HSCT for thalassaemia identified 3 independent prognostic factors for outcome in children (Table 4): - Hepatomegaly
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- Liver biopsy evidence of portal fibrosis - Irregular compliance with chelation. These factors form the basis of the Pesaro classification into good, intermediate and poor risk (Class 1, 2 and 3 respectively). Table 5 shows how Pesaro class predicted OS, thalassaemia-free survival and graft rejection (1). Even modest organ damage pre-HSCT increased TRM. Pesaro classification does not predict outcome in all centres- perhaps due to low TRM in many recent series. Nevertheless, it is useful for identifying high-risk patients who may not benefit from HSCT or who may need modified conditioning (4).
Table 4: Pesaro Classification for predicting outcome of HSCT for thalassaemia major Risk factors
Class 1
Class 2*
Class 3
Hepatomegaly (>2 cm below costal margin)
No
Yes/no
Yes
Irregular chelation#
No
Yes/no
Yes
Portal fibrosis on liver biopsy
No
Yes/no
Yes
* One or two of any of the 3 risk factors; # Desferrioxamine started >18 months after regular transfusions commenced or desferrioxamine administered <8 hours/night on at least 5 nights per week
Table 5: Outcome of BMT for thalassaemia by Pesaro Risk Classification* Class 1
Class 2
Class 3#
Survival (%)
93
87
93
Thalassaemia-free survival (%)
90
84
85
Transplant-related mortality (%)
6
13
6
Graft rejection (%)
7
4
8
Data are mostly taken from Schrier and Angelucci, 2005 (1). * See Table 4; # Data from Sodani et al., 2004 (4) using modified conditioning regimen
5.2.2. Other factors In children <17 years, age is not an independent predictor of outcome. However, adults consistently show inferior OS and EFS to children (64% OS; 62% EFS), even using reduced-intensity conditioning (RIC) (1, 7). Hepatitis B/C infection affects neither TRM nor EFS. Thalassaemia trait in the donor does not affect outcome. 570
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5.3. Outcome of HSCT for thalassaemia using HLA-matched family donors (Table 5) Most groups currently report OS of 90–95% and thalassaemia-free survival of 80–90% in children (1, 3). Acute GvHD and infections are the commonest causes of TRM. Severe chronic GvHD after sibling SCT is uncommon (2–5% of patients). Graft rejection (~10%) is more common and usually occurs within 6 months. Therefore, monthly donor/host chimerism (peripheral blood) should be monitored for the first 3–6 months; falling donor chimerism usually responds to manipulation of immune suppression. Anecdotal experience suggests DLI rarely reverses established rejection but may be successful if used early. Failure of primary engraftment with aplasia is rare and has a poor outcome even with second HSCT and autologous HSCT may be safer; therefore cryopreservation of autologous marrow prior to allogeneic HSCT may be advisable. Most HSCT for thalassaemia use marrow from HLA-identical siblings; where there is consanguinity, other close relatives may be HLA-identical and outcome is not significantly different. Families without sibling donors often extend their families, with/without prenatal or pre-implantation genetic diagnosis. PBSC are not generally chosen for thalassaemia HSCT since most donors are young children (reluctance to use G-CSF or central venous catheters) and the largest series showed more severe GvHD in PBSC transplants (8). 5.4. Outcome and role of cord blood transplantation (CBT) for thalassaemia CBT data from HLA-identical siblings are limited and show excellent OS (100%) but significant graft rejection (21%) which can be reduced by omitting methotrexate and including thiotepa in conditioning (4). Since graft rejection is significant, it is reasonable to use cord blood only where the total nucleated cell dose is >3 x 107/kg and to wait until the donor is ≥2 years of age so “back up” donor marrow is available. 5.5. Outcome and role of unrelated donor HSCT for thalassaemia Experience of unrelated donor SCT for thalassaemia is limited to small numbers with a variety of conditioning regimens, age groups and donor types and relatively short follow up (9). Recent data confirm lower OS (79%) and thalassaemia-free survival (66%) after unrelated compared to HLA-identical family donors. At present the role of unrelated donor HSCT in thalassaemia remains to be established and is best investigated through well-designed clinical trials.
6. Results of HSCT for SCD 6.1. Role of HSCT in SCD SCD is very heterogeneous. Although all patients have impaired QoL because of
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unpredictable pain and progressive organ damage, specific clinical features (e.g. recurrent chest syndrome, stroke and frequent severe painful crises) best identify SCD patients with a poor prognosis (2). For them HSCT offers both improved QoL and, since 50–60% of adults with SCD fail to reach their 50s, better prospects for long-term survival. 6.2. Prognostic and risk factors The Pesaro classification does not apply to SCD and there is no equivalent prognostic score for SCD. However, careful evaluation of patients enrolled in transfusion programmes for CNS disease is recommended. Donor sickle cell trait does not affect outcome. 6.3. Outcome of HSCT for SCD using HLA-matched family donors (Table 6) Approximately 250 patients have been transplanted worldwide, virtually all <16 years and most for complicated SCD, particularly SCD-related stroke (Table 6) (2, 10–12). OS is ~90%; SCD-free survival 82–86%; TRM 7–8% and graft rejection 8%. All patients with stable engraftment no longer had clinical manifestations of SCD. Commonest causes of TRM are GvHD and infections. The risk of neurological complications is increased in the peri-HSCT period (1/3 of patients), particularly seizures and intracranial haemorrhage. However, these neurological problems are reduced significantly by prophylactic anticonvulsants before HSCT continuing for at least 6 months; maintaining platelets >50 x 109/L; and rigorous control of cyclosporin, magnesium and blood pressure. Graft failure occurs in 10–18% almost always with autologous reconstitution/relapse of SCD. Mixed haematopoietic chimerism is usually stable but falling donor haematopoiesis has been successfully treated by DLI. Chimerism should be monitored monthly for the first 6 months. Table 6: Outcome of BMT for sickle cell disease Walters et al. 2000 (n=50)
Bernaudin et al. Vermylen et al. 1997 (n=26) 1998 (n=50)
Panepinto et al. 2007 (n=67)
Overall survival
94% (6 yr)
92% (8 yr)
93% (11 yr)
97% (5 yr)
Event free survival
84% (6 yr)
75% (8 yr)
82% (11 yr)
85% (5 yr)
Graft rejection*
10%
18%
10%
13%
Acute GvHD ≥ Grade 2
8%
23%
20%
10%
8%
6%
4%
Chronic GvHD - extensive 4%
* All had autologous reconstitution with relapse of SCD 572
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6.4. Outcome and role of CBT for SCD CBT data from HLA-identical sibling donors are limited and show excellent OS (100%) (4). Graft rejection occurs but numbers are too small for analysis of risk factors. Similar principles about the role of CBT apply in SCD as those in thalassaemia. Methotrexate should be omitted; thiotepa may be added to conditioning and cord blood should be used only where the total nucleated cell dose is >3 x 107/kg. It is recommended to wait until the donor is ≥2 years old so “back up” donor marrow is available. 6.5. Outcome and role of unrelated donor HSCT for SCD Unrelated donor HSCT for SCD is not routinely recommended and experience is limited to occasional case reports.
7. Long-term effects of SCT for haemoglobinopathies: special features Iron overload improves slowly post-HSCT but can be accelerated by regular phlebotomy or chelation beginning 9–12 months after HSCT until the total iron burden approaches normal (liver iron <5 mg/g DW; serum ferritin <700 ng/mL) (1). There are few data on fertility post-HSCT for thalassaemia and SCD although occasional pregnancies and paternities have been reported. Advances in cryopreservation of testicular and ovarian tissue show promise even for pre-pubertal children; available options should be discussed with families prior to HSCT.
8. Conclusions HSCT remains the only cure for thalassaemia and SCD. Cure appears life-long and associated with acceptable long-term risks. Families and physicians must weigh up the risks, including TRM and graft failure, against expected survival and QoL with medical treatment. For thalassaemia outcome of HSCT is best in patients <16 years old who comply well with chelation and have no liver dysfunction. They can expect long-term survival of 95% and thalassaemia-free survival of 90%. Patients with poor risk features have a reduced chance of cure (56–82%) and higher TRM (up to 20%) but still have a long-term survival advantage over conventional medical management. SCD is more heterogeneous; the patients predicted to benefit most from HSCT are those with CNS disease or recurrent chest syndrome despite hydroxycarbamide. Longterm EFS after HSCT for SCD in childhood is 82–86%, almost identical to recent US data on survival in SCD with medical treatment. Future prospects include effective RIC regimens, which currently have unacceptably high rejection rates in haemoglobinopathies, and gene therapy (2).
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References 1. Schrier S, Angelucci E. New strategies in the treatment of the thalassemias. Ann Rev Med 2005; 56: 157-171. 2. Walters MC. Stem cell therapy for sickle cell disease: Transplantation and gene therapy. Hematology (Am Soc Educ Program 2005) 2005; 66-73. 3. Lawson S, Roberts IAG, Amrolia P, et al. Bone marrow transplantation for b-thalassaemia major: The UK experience in two paediatric centres. Br J Haematol 2003; 120: 289-295. 4. Sodani P, Gaziev D, Polchi P, et al. New approach for bone marrow transplantation in patients with class 3 thalassemia aged younger than 17 years. Blood 2004; 104: 1201-1203. 5. Locatelli F, Rocha V, Reed W, et al. Related Umbilical Cord Blood Transplant in Patients With Thalassemia and Sickle Cell Disease. Blood 2003; 101: 2137-2143. 6. Borgna-Pignatti C, Rugolotto S, di Stefano P, et al. Survival and complications in patients with thalassemia major treated with transfusion and deferoxamine. Haematologica 2004; 89: 1187-1193. 7. Gaziev J, Sodani P, Polchi P, et al. Bone marrow transplantation in adults with thalassemia: Treatment and long-term follow up. Ann NY Acad Sci 2005; 1054: 196-205. 8. Mohyeddin Bonab M, Alimoghaddam K, Vatandoust S, et al. Are HLA antigens a risk factor for acute GVHD in thalassemic patients receiving HLA-identical stem cell transplantation? Transplant Proc 2004; 36: 3190-3193. 9. La Nasa G, Argiolu F, Giardini C, et al. Unrelated Donor Bone Marrow Transplantation for Thalassemia: The Effect of Extended Haplotypes. Blood 2002; 99: 4350-4356. 10.Vermylen C. Hematopoietic stem cell transplantation in sickle cell disease. Blood Rev 2003; 17: 163-166. 11.Panepinto J, Walters M, Carreras J, et al., on behalf of the Non-Malignant Marrow Disorders Working Committee, Center for International Blood and Marrow Transplant Research. Matched-related donor transplantation for sickle cell disease: Report from the Center for International Blood and Marrow Transplant Research. Br J Haematol 2007; 137: 479-485. 12.Bernaudin F, Socie G, Kuentz M, et al., on behalf of the SFGM-TC. Long-term results of related myeloabalative stem-cell transplantation to cure sickle cell disease. Blood 2007; 110: 2749-2756.
Mutiple Choice Questionnaire To find the correct answer, go to http://www.esh.org/ebmt-handbook2008answers.htm 1. The following are standard indications for HSCT in children with sickle cell disease: a) Recurrent splenic sequestration if there is an HLA-matched family donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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b) Stroke if there is an HLA-matched family donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Stroke if there any HLA-matched donor. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Severe growth and pubertal delay if there is an HLA-matched family donor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. The following are prognostic factors in the Pesaro classification for outcome of HSCT for thalassaemia: a) Hepatomegaly, portal fibrosis and cardiac dysfunction . . . . . . . . . . . . . . . . . . . . . b) Hepatosplenomegaly, portal fibrosis and serum ferritin >2000 ng/mL . . . . c) Hepatosplenomegaly, hepatic cirrhosis and cardiac dysfunction . . . . . . . . . . d) Hepatomegaly, portal fibrosis and a history of irregular chelation . . . . . . . . 3. Overall survival in children after HSCT for thalassaemia from an HLA-identical family donor is: a) 90-95% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) 80-85% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) 70-75% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) 75-80% . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Most conditioning regimens for haemoglobinopathy use a combination of: a) Cyclophosphamide and TBI . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Cyclophosphamide and busulfan. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Cyclophosphamide and ATG . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Cyclophosphamide and fludarabine. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. The following are common long-term complications of HSCT for haemoglobinopathies: a) Growth delay . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b) Secondary malignancy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c) Relapse . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d) Infertility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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ABBREVIATIONS
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(-) = negative (+) = positive A AA = aplastic anaemia ABMTR = Autologous Bone Marrow Transplant Registry ADA = adenosine deaminase ADs = autoimmune diseases AE = adverse event Ag = antigen aGvHD = acute graft versus host disease AI = alloimmunisation AL = acute leukaemia AL amyloid = primary (or immunoglobulin light-chain) amyloidosis ALCL = anaplastic large cell lymphoma ALL = acute lymphoblastic leukaemia allo-HSCT = allogeneic haematopoietic stem cell transplant ALWP = Acute Leukaemia Working Party AmB = amphotericin B AML = acute myeloid leukaemia ANC = absolute neutrophil count AP = accelerated phase APCs = antigen presenting cells APL = acute promyelocytic leukaemia APTT = activated partial thromboplastin time ARA-C = cytosine arabinoside ASR = annual safety report ATD = adult therapeutic dose ATG = antithymocyte globulin ATRA = all-trans retinoic acid auto-HSCT = autologous haematopoietic stem cell transplant AVN = avascular necrosis of bone B BAL = broncho-alveolar lavage BC = blast crisis BCNU = 1,3-bis(2-chloroethyl)-1-nitrosourea BCSS = breast cancer-specific survival BFM = Berlin-Frankfurt-Munster study group
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BM = bone marrow BMF = bone marrow failure BMDW = Bone Marrow Donor Worldwide BMT = bone marrow transplantation BNLI = British National Lymphoma Investigation BO = bronchiolitis obliterans BOOP = bronchiolitis obliterans with organising pneumonia Bu = busulfan BUN = blood urea nitrogen C CA = Competent Authority and National legislation CARD15 = caspase recruitment domain family member 15 CB = cord blood CBT = cord blood transplantation CD = cluster designation CFU-GM = colony forming unit granulocyte macrophage CGPs = cytokine gene polymorphisms cGvHD = chronic graft versus host disease CI = cranial irradiation CLL = chronic lymphocytic leukaemia CLWP = Chronic Leukaemia Working Party CLS = capillary leak syndrome CML = chronic myeloid leukaemia CMV = cytomegalovirus CNS = central nervous system CP = chronic phase C-PI = coordinating principle investigator CR = complete remission CR1 = first complete remission CsA/CSA = cyclosporin A CSF = colony stimulating factor CSI = craniospinal irradiation CT = chemotherapy CTA = Clinical Trial Authorisation CT/RT = chemo-radiotherapy CTL = cytotoxic T-lymphocytes CTLp = cytotoxic T-lymphocyte precursor CVC = central venous catheter 578
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Cy = cyclophosphamide D DAH = diffuse alveolar haemorrhage DBA = Diamond-Blackfan anaemia DC = dendritic cell DFS = disease-free survival DIC = disseminated intravascular coagulation DKC = dyskeratosis congenita DLBCL = diffuse large B-cell lymphoma DLI = donor lymphocyte infusion DLT = dose limiting toxicity DMARDs = disease modifying antirheumatic drugs DMSO = dimethylsulfoxide E EBMT = European Group for Blood and Marrow Transplantation EBV = Epstein-Barr virus EC = Ethics Committee ECIL = European Conference on Infections in Leukaemia EES = extra-osseous Ewing’s sarcoma EFS = event-free survival EPCs = endothelial progenitor cells EPO = erythropoietin ERα = estrogen receptor alpha ERT = enzyme replacement therapy ES = embryonic stem cells ES = engraftment syndrome ET = essential thrombocythaemia EU = European Union EULAR = European League against Rheumatism EWOG-MDS = European working group on myelodysplastic syndromes F FA = Fanconi anaemia FAHCT = Foundation for Accreditation of Hematopoietic Cell Therapy FFP = fresh frozen plasma FHCRC = Fred Hutchinson Cancer Research Centre FISH = fluorescent in situ hybridisation
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FL = follicular lymphoma FLC = free immunoglobulin light-chain FLIPI = follicular lymphoma international prognostic index Fluda = fludarabine FNHTR = febrile non-haemolytic transfusion reactions G G-CSF = granulocyte colony stimulating factor GCT = germ cell tumour GH = growth hormone GHD = growth hormone deficiency GI = gastrointestinal GITMO = Gruppo Italiano Trapianto di Midollo Osseo GM-CSF = granulocyte macrophage colony stimulating factor GMP = good manufacturing practice GnRH = gonadotrophin releasing hormone GOELAMS = Groupe Ouest Est d'étude des Leucémies et Autres Maladies du Sang GSH = glutathione GT = granulocyte transfusions GvHD = graft versus host disease GvL= graft versus leukaemia H HBV = hepatitis B virus HC = haemorrhagic cystitis HCMV = human cytomegalovirus HCV = hepatitis C virus HD = Hodgkin’s disease HDCT = high-dose chemotherapy HDT = high-dose therapy HEPA = high efficiency particle extraction HHV6 = human herpes virus 6 HIV = human immunodeficiency virus HLA = human leukocyte antigen HmR = hormone receptors HR = high-risk HS = Hurler’s syndrome HSC = haematopoietic stem cell HSCT = haematopoietic stem cell transplantation 580
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HSV = herpes simplex virus HSV–TK = herpes simplex virus 1–thymidine kinase HTLV = human T-leukaemia virus HUS = haemolytic uraemic syndrome HVGP = hepatic venous gradient pressure I IBMTR = International Bone Marrow Transplant Registry IDWP = Infectious Diseases Working Party IEM = inborn errors of metabolism IFN = interferon Ig = immunoglobulin IGF = insulin-like growth factor IGFBP-3 = insulin-like growth factor binding protein 3 IL = interleukin IL-1Ra = interleukin 1 receptor antagonist IMP = investigational medicinal product IP = interstitial pneumonia IPS = interstitial pneumonia syndrome IPSS = International Prognostic Scoring System IR = immune reconstitution ITIM = immunoreceptor tyrosine-based inhibition motifs IV = intravenous IVIg = intravenous immunoglobulin J JACIE = Joint Accreditation Committee EBMT-ISCT Europe JIA = juvenile arthritis JMML = juvenile myelomonocytic leukaemia K KGF = keratinocyte growth factor KIR = killer-cell inhibitory receptor L LAP = leukaemia-associated phenotype LC = immunoglobulin light-chain LCT = long chain triglycerides LDH = lactate dehydrogenase
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LEWP = Late Effects Working Party LFS = leukaemia-free survival LGL = large granular lymphocytes LH = luteinising hormone LSCs = leukaemic stem cells LSDs = lysosomal storage diseases LSK = Lin-sca-1+c-kit+ cells LTC-ICs = long term culture-initiating cells LVEF = left ventricular ejection fraction M MA = marketing authorisation MAH = marketing authorisation holder MAPCs = multipotent adult progenitor cells MBL = mannose binding lectin MC = mixed chimerism MCT = medium chain triglycerides MDS = myelodysplastic syndrome MEL = melphalan MethylPDN = methyl prednisolone MHA = microangiopathic haemolytic anaemia MHC = major histocompatibility complex MHag = minor histocompatibility antigens MMF = mycophenolate mofetil MM = multiple myeloma MMM = myelofibrosis with myeloid metaplasia MNC = mononuclear cells MoAb = monoclonal antibody MODS = multiple-organ dysfunction syndrome MOF = multi-organ failure MP = menopausal MPD = myeloproliferative disorders MPO = myeloperoxidase MRD = matched related donor MRD = minimal residual disease MRA = magnetic resonance angiography MRI = magnetic resonance imaging MS = multiple sclerosis MSC = mesenchymal stem cell 582
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MTD = maximum tolerated dose MTHFR = methylenetetrahydrofolate reductase MTX = methotrexate MUD = matched unrelated donor MVA = modified vaccinia Ankara N NBS = Nijmegen breakage syndrome NHL = non-Hodgkin’s lymphoma NK = natural killer NMDP = National Marrow Donor Program NOD2 = nucleotide-binding oligomerisation domain containing 2 NOD/SCID = non-obese diabetic/severe combined immunodeficiency (mice) NRM = non-relapse mortality O OAS = optimal additive solution OS = overall survival P PAI-1 = plasminogen activator inhibitor-1 PB = peripheral blood PBSC = peripheral blood stem cells PBSCT = peripheral blood stem cell transplantation PC = platelet concentrate PCR = polymerase chain reaction PCV = packed cell volume PDN = prednisone / prednisolone PFS = progression-free survival PFT = pulmonary function test Ph(+) = Philadelphia positive PGD = preimplantation genetic diagnosis PI = principal investigator pNET = primitive neuroectodermal tumour PNH = paroxysmal nocturnal haemoglobinuria PNP = purine-nucleoside-phosphorylase p.o. = per os PR = partial remission PRB = plasma reduced blood
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PRCA = pure red cell aplasia PRCT = prospective randomised controlled trial PRD = primary refractory disease PRRs = pattern recognition receptors PS = performance status PT = prothrombin time PTCL = peripheral T-cell lymphoma PTLD = post-transplant lymphoproliferative disorder PV = polycythaemia vera PVD = peripheral vascular disease Q QoL = quality of life R RA = rheumatoid arthritis RAEB = refractory anaemia with excess of blasts RAEB-t = refractory anaemia with excess of blasts in transformation RC = refractory cytopenia REM = remission RI = relapse incidence RIA = radioimmune assay RIC = reduced intensity conditioning RMS = rhabdomyosarcoma RR = relapse risk RSV = respiratory syncytial virus RT = radiotherapy/irradiation S SAA = severe aplastic anaemia SC = stem cell SCC = squamous cell carcinoma SCD = sickle cell disease SCF = stem cell factor SCID = severe combined immunodeficiency SCID-X1 = X-linked form of severe combined immunodeficiency SD = standard deviation SDCT = standard-dose chemotherapy SEC = sinusoidal endothelial cell 584
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SHRT = sex hormone replacement therapy SIB = sibling SLE = systemic lupus erythematosus SMC = stable mixed chimerism SNPs = single nucleotide polymorphisms SOS = sinusoidal obstruction syndrome SP = side population SRV = survival SSc = scleroderma (systemic sclerosis) SSL = small lymphocytic lymphoma STRs = short tandem repeats T TA = transfusion-associated TBI = total body irradiation TCD = T-cell depleted TCR = T-cell receptor TGF-beta = tumour growth factor beta Th-1 = T helper TKI = tyrosine kinase inhibitor TNF = tumour necrosis factor TLRs = toll like receptors TMA = thrombotic microangiopathy TMC = transient mixed chimerism TPN = total parenteral nutrition TREC = T-cell receptor rearrangement excision DNA circles Treg = regulatory T-cells TRM = transplant related mortality TSH = thyroid stimulating hormone TT = thiothepa TTP = thrombotic thrombocytopenic purpura U UCBT = umbilical cord blood transplantation UD = unrelated donor UD CBT = unrelated donor cord blood transplantation V VDR = vitamin D receptor VNTRs = variable number tandem repeats
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VOD = veno-occlusive disease VP/VP16 = etoposide vs. = versus VZV = varicella zoster virus W WBC = white blood cell WHO = World Health Organisation WMDA = World Marrow Donor Association WP = working party Y yrs = years
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NOTES
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