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Minutes from 13th IHWG Stem Cell Transplantation Component Meeting Donor Registries Conference, July, 1999 Paris |
| Moderator: | Dr. Effie Petersdorf (USA) | |
| Panel Members: | Dr. Carolyn Hurley (USA) Dr. Marie-Marthe Tongio (France) Dr. Jean-Denis Bignon (France) Dr. Chaim Brautbar (Israel) Dr. Carlheinz Muller (Germany) Dr. John Hansen (USA) Dr. Effie Petersdorf (USA) |
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Introduction This session was opened by a presentation of the overall goals and organization of the 13th International Histocompatibility Workshop and Conference (IHWC) by Dr. John Hansen, IHWC Chairman. The goal of the 13th IHWC is to define the full extent of genomic diversity of the MHC and to understand biological implications. The workshop is comprised of Core Components that provide basic resources for the workshop participants, and Scientific Components that are each designed to address new questions pertaining to HLA polymorphism and function. Three components have direct relevance to stem cell transplantation: Core Component E (SSOP and SBT Typing), Hematopoietic Cell Transplantation (HCT), and Donor Registries. Hematopoietic Cell Transplantation Dr. Effie Petersdorf summarized the goals and organization of the Hematopoietic Cell Transplantation Component. The overall goal is to develop resources for scientific inquiry into the genomics of transplantation. The working group will first develop an international cell and DNA repository of donor-recipient transplant pairs that are not already available through existing repositories. The working group will not duplicate cells and DNA if these materials are available in other repositories. The working group will also identify and collect samples that have, up until now, been missing from existing repositories. The working group has defined a minimum requisite of 2 mg of genomic DNA or 2 vials of 5 X 106 PBL on each donor and patient. For laboratories not wishing to transform cell lines, the IHWC will transform cell lines at the Fred Hutchinson Cancer Research Center (FHCRC). For all transplants henceforth, the working group desires to save a minimum of 2 vials of 5 X 106 PBL and 2 mg of genomic DNA. The FHCRC will transform cell lines if the contributing laboratory does not have the facilities to do so. The second specific aim of the component is to fully characterize each donor-recipient pair for alleles encoded at HLA-A, B, C, DRB, DQB1 and DPB1. This will be accomplished by collecting all HLA typing information available at the time of transplant, identification of samples requiring further allele-level typing, and use of SSOP and SBT standardized protocols developed by Core E for completion of the typing. In the third aim of the study, the relevance of complete allele matching to clinical outcome will be evaluated. For the purposes of reducing heterogeneity in the study population and to decrease inter-observer variation in the definition of clinical endpoints, the working group will focus on the analysis of CML unrelated transplants with survival as the primary endpoint. Acute and chronic GVHD are the secondary endpoints of the study. Clinical data collection will use existing report forms from the NMDP and IBMTR. If clinical data is not submitted via the NMDP or IBMTR, the working group will identify those transplants and liaison directly with the transplant centers. The fourth aim of the study will be to evaluate the clinical impact of disparity for microsatellites and for single nucleotide polymorphisms (SNPs). The overall objectives are to determine whether microsatellite markers, in defining a larger part of the MHC can provide new information on donor-recipient matching than is currently gained by typing each class I and II gene. The presence of disparity for microsatellite markers may indicate that the patient and donor have different extended haplotypes. The analysis of SNPs will represent the stepping stones for investigating the role of polymorphisms encoded outside the MHC in clinical transplantation. Finally, the project will seek to define permissible mismatches. The specific mismatches to be analyzed will depend on the overall allele and haplotype frequencies encoded in the international set of data, but will likely enable examination of common alleles encoded by A2, B44 and DR4 as a minimum. SSOP and SBT Standardized Protocols Dr. Carolyn Hurley summarized the overall goals and activity to date of the Core Component on SSOP and SBT Typing. The goal is to define standard HLA polymorphism identification strategies and reagents to be used in all components of the 13th IHWS. The specific aims are to first establish DNA testing strategies; in this regard, SSOP techniques provide robust, high volume typing, and resolution of known polymorphisms. All HLA loci will be included (HLA-A, B, C, DR, DQ, DP A and B). The Core Component will provide a strategy and reagents to workshop participants and will also provide quality control. The oligonucleotide reagents will provide two levels of resolution: allele level and "registry" level (serologic level to intermediate resolution). Reagents, strategies and protocols have already been posted on the IHWC web site. Reagents will be provided at reasonable cost by a commercial vendor. The typing strategy uses over 30 amplification groups to yield allele-level typing. Thus far, the component has completed development of DRB registry-level reagents. Strategies for HLA-A and B are currently under discussion. Over 630 reagents have been tested. A quality control process has been established and a panel of reference cells to be used for quality control testing is being collected. By October 1999 the reagents, QC, and reference cells will be available to workshop participants. The use of standardized reagents and protocols by all components of the workshop will enable a uniform set of data to be collected in a highly rigorous manner. Advantages to stem cell donor registries are three-fold: 1) the establishment and standardization of DNA-based HLA typing of volunteer donors; 2) the establishment of a network of high quality laboratories through quality control process, and 3) the interaction between participants will enable development of flexible and accurate systems for retaining and using HLA data in donor selection. Updates: France Dr. Marie-Marthe Tongio presented a summary of the retrospective transplants on behalf of French laboratories. In total, 26 grafting centers, 40 HLA laboratories and France Greffe de Moelle are involved in the 13th IHWC Hematopoietic Cell Transplantation effort. In review of the retrospective transplants performed between 1990 - 1998 inclusive, a total of 989 transplants were performed during this period, of which 812 were performed between 1993-1998. Among the 812 transplants, 173 (21%) were for CML and 22 (3%) were for JCML. A total of 287 transplant pairs have stored DNA and/or PBL available for workshop studies. The following centers are contributing data to the component: Besancon (10), Grenoble (10), Lille (30), Lyon (24), Marseille (25; 16 cord blood), Montpellier (6), Nantes (36), Paris Armees (4), Paris St. Louis (60), Poitiers (20), Rennes (20), Saint Etienne (7), and Strasbourg (35). The extension of the proposed workshop studies to include cord blood transplants and pediatric transplants was presented. Clinical outcome data for all cases contributed by French centers will be submitted by the French Society of Bone Marrow Transplantation, SFGM. The SFGM has substantial clinical data already available. Pre-transplant variables include diagnosis, therapy prior to transplantation, and history of the disease. Data pertaining to the transplant include extent of donor-recipient matching (serology, HLA, ABO), disease stage, conditioning regimen, GVHD prophylaxis, and source and number of infused stem cells. Post-transplant data include engraftment, chimerism, acute and chronic GVHD, relapse, survival, and cause of death. For retrospective transplants performed between 1990-1998, DNA and PBL are currently located in HLA laboratories. The participants propose that the HLA laboratories first provide patient identifiers and HLA typing data to France Greffe de Moelle. France Greffe de Moelle will then submit this information to SFGM for release of clinical data. France Greffe de Moelle will be the primary communicator with the IHWC. A significant number of additional testing will be feasible from the participating laboratories including MIC, TAP, LMP, KIR, HA-1, microsatellites and CD1. Dr. Jean-Denis Bignon summarized anticipated contributions from prospective transplants performed in France between now and 2001. In is anticipated that 300-350 additional transplants will have been performed by June 2001. The categories by disease are: CML (60-80), AML (30-70), ALL (80-100), SAA (15-20), FA (12-15), MDS (25-30), immune deficiencies (10-12), other (20-25). France Greffe de Moelle sent a questionnaire in April 1999 to all participants. In total, 12-15 HLA labs out of 26 grafting centers will be participating in this component. Approximately 40-60 CML transplants are anticipated from this effort. France Greffe de Moelle has requested each laboratory and grafting center to collect and store 50 micrograms of DNA on each donor and recipient, and cells for eventual EBV transformation. DNA will be stored in HLA laboratories and at France Greffe de Moelle. HLA laboratories will store cells. France Greffe de Moelle will update unrelated donor transplant files and request centers to collect and store adequate amounts of DNA and cells. France Greffe de Moelle will regularly verify that the collection of donor-recipient samples is carried out efficiently at all centers. The Scientific Committee of SFGM has agreed to provide clinical data from transplants included in the Hematopoietic Cell Transplantation Component. In France, local centers will take on the responsibility of LCL transformation. Centers will forward to the Seattle Repository 2 mg of genomic DNA and 2 vials of 5 X 106 PBL on each pair. Storage of additional samples at France Greffe de Moelle is currently under discussion. France Greffe de Moelle has sent a questionnaire to participating laboratories and centers to determine the highest resolution typing that will be attained in this effort. Based on response from 10 laboratories, the majority of alleles recognized by the WHO Nomenclature Committee as of April 1999, can be typed by the participating laboratories. Currently, the majority of centers are using SSP methodology. Many laboratories are in the process of developing SBT typing methods. In conclusion, 13 French laboratories have agreed to contribute DNA/cells on prospective transplants and to perform allele-level typing. France Greffe de Moelle could serve as a centralized repository to facilitate the handling of donor-recipient DNA/Cells to the IHWC. Updates: Germany Dr. Carlheinz Müller gave a presentation of the envisaged German contribution to the Hematopoietic Stem Cell Transplant component of the Workshop and recent developments in HLA information technology that are relevant to the workshop and which may have a significant impact on the way donor registries work and cooperate. A large multi-center study will be carried out between 1999 and 2001 in Germany retyping 800 unrelated and 100 related donor patient pairs. The major objective of this study funded by the German José Carreras Leukemia Foundation is to assess the impact of HLA matching on the major parameters of transplantation outcome. There is a special focus on the role of class I and class II subtype disparities in view of recent results from Seattle and Japan whose relevance for the German population is still unclear. All pairs will be allele level typed for DRB1 and DQB1 by SSP or SSOP and sequenced for HLA-A, B and C. The principal investigators, Dr. Shraga F. Goldmann, Ulm, and Dr. Hans Grosse-Wilde, Essen, have assured a broad cooperation among German clinicians and immunogeneticists including the German national donor and patient registries, ZKRD and DRST. They intend to provide the intermediate data of this study to the 13th IHWC. Although the clinical follow-up time will still be limited, the data will encompass a large and fairly comprehensive set of unrelated transplants performed in Germany in 1999 and 2000. A significant part of HLA class II typing performed for potential stem cell donors in recent years was performed with intermediate resolution molecular methods, mostly SSP and SSOP. Multiple allele letter coding introduced by the NMDP a few years ago has been used to communicate and enter the typing data into registry databases. The major drawback of this coding is twofold. First, typing usually results in a number of genotypes compatible with the raw laboratory data. With the multi-allele approach, a (hopefully minimal) set rectangle is drawn around an actual result set, which in most cases, then also encompasses genotypes already excluded by means of existing results. Second, and even worse, the interpretation of the laboratory results will almost surely change as soon as new alleles for the locus in question is described. In the setting of a stem cell donor registry, this will gradually impair any existing data and, in the end, render them wrong and useless. Currently, a number of promising approaches to tackle this challenge exist. Their common purpose is to "immortalize" (Dr. Martin Maiers) the data obtained in the laboratory, but their theoretical background and general requirements are quite different. They are essentially based on storing the data obtained in the laboratory or some data set derived from them without an interpretation depending on the current underlying set of alleles. These concepts are currently discussed and reviewed in an informal "Virtual Sequence Group" that involves the practical contributors Dr. Wolfgang Helmberg (Kit Pattern a.k.a. Virtual DNA Analysis), Dr. Martin Maiers (Logical Constraint Programming) and Dr. Erik Rozemuller (Virtual Sequence). For donor registries, the major benefit from using such an approach in the future is to maximize the usability of their data. All users should have a precise idea of which information a donor typing conveys independently of when it was performed. The financial relevance of this topic cannot be overestimated since the resources necessary for donor typing are by far the largest sum in the budget of the registry system of any country. Making such innovative ideas work for registries on a national and international level will require significant efforts of all parties involved and revolutionize the structure of the HLA databases and the communication. The 13th IHWC could make a major contribution in this area with regard to validating the power, establishing the feasibility and standardizing the implementation of a new way of handling HLA typing data. Updates: Israel Dr. Chaim Brautbar presented an update of the unrelated marrow donor program and cord blood registry at Hadassah Hospital, Jerusalem. Overall, Caucasian patients seeking unrelated donors in the NMDP registry have an 83% success rate of identifying a suitable donor. In contrast, Jewish patients initiating an unrelated donor search through international registries have a 45% success rate in identifying suitable donors. The explanation for the lower rate in the Jewish population may be explained by the HLA genetic background of the Jewish population. The Jewish population in Israel is classified into two major groups, Ashkenazi from Central and Eastern Europe, and non-Ashkenazi Jewish originating from Mediterranean and North African countries. The relationship between the Ashkenazi Jewish populations can be readily described by a population tree displaying genetic distances for HLA class II allele distributions among Israeli, non-Jewish Caucasians and African populations. This dendrogram demonstrates that the different Jewish groups, Israeli Arabs, European and American Caucasians, and African populations from North America and South Africa can be clustered into three major groups, with the exception of the Ethiopian Jews: all Jewish groups and the Israeli Arabs; non-Jewish Caucasians; and the two African populations. Hence, the Jewish population differs from other population groups in HLA allele and haplotype frequencies. Furthermore, it is most important for Israelis as well as Jews worldwide, to have donor registries containing as many of the alleles and haplotypes represented among Ashkenazi and non-Askenazi ethnic groups in order for all Jewish patients to have increased chances of finding suitable unrelated donors for transplantation. As of May, 1999, the registry contained a total of 13,277 bone marrow donors, and 243 cord blood (CB) units. 11,346 of the bone marrow donors have been HLA-A,B typed, 1931 have been HLA-A,B,DR typed, and 1200 have DRB1 and DQB1 allele typing. Of the 243 CB units, 34 have been HLA-A,B typed, 209 are HLA-A,B,DR typed, and 207 have DRB1, DQB1 allele typing. The age distribution of donors is as follows: 1% between 18 - 20 years, 36% between 21-30 years, 23% between 31-40 years, 27% between 41-40 years, and 13% over the age of 50 years. In 1998, there were 1,120 preliminary searches, one bone marrow donation, three PBSC donations and one second PBSC donation. As of June 15, 1999, there have been 653 preliminary searches, 3 bone marrow donations, 2 PBSC donations and 1 second PBSC donation. HLA typing of unrelated donor-recipient pairs is performed for HLA-A,B, and C using serological and low-resolution SSP techniques for patients, and low-resolution SSP methods for donors. HLA-DRB1, DRB3, DRB4, DRB5 and DQB1 alleles are typed using SSOP techniques for both patients and donors. Additional testing of donor-recipient compatibility includes MLC and an assessment of sensitization using PRA and crossmatch. To date, 72 unrelated donor transplants have been performed. For the 13th IHWC Hematopoietic Cell Transplantation Component, 22 unrelated donor-recipient CML pairs are available for analysis, 18 pairs have DNA available for further genomic testing, and 15 pairs have cells available. All 22 pairs have complete clinical data. Prospective CML transplants performed between 1999 - 2001 will have genomic DNA prepared on both donor and recipient and PBL frozen for future EBV transformation. Clinical data is submitted by Dr. Arnon Naglar of the Transplant Center. Dr. Chaim Brautbar will submit HLA and clinical data to the IHWC database. Conclusions The Hematopoietic Cell Transplantation Component represents a worldwide effort to understand the genomics of transplantation. It is essential that all future transplants have sufficient cells collected and stored in order to guarantee longitudinal studies. The worldwide community must minimize any further loss of samples and clinical data. The IHWC venue is ideal for completion of the proposed studies for several reasons. The IHWC has historically excelled in the study of large human populations distinguished by their racial and ethnic diversity. The use of standardized methods has enabled the collection of high quality data that is necessary for the correlation of genetic and phenotypic data. |