A review of the genetic and long-term effects of G-CSF injections in healthy donors: a reassuring lack of evidence for the development of haematological malignancies.

In 2007 the WMDA responded to the publication of two manuscripts suggesting a causal link between G-CSF and myeloid malignancies in healthy donors by convening an international symposium to examine this issue. At the time, registries reviewed the long-term follow-up of their healthy donors, which suggested no excess of leukaemia in PBSC donors compared with BM donors. Although the evidence for an increased risk of malignancy in healthy donors was felt to be weak, it could not be excluded. The WMDA, therefore, issued a statement, to be included in all donor consent forms, stating that it was unknown whether G-CSF increased or decreased the risk of later developing cancer. In 2012, with 5 years of additional donor follow-up and the results of several genetic studies now available, the clinical working group of the WMDA again reviewed the data. On the basis of an assessment of a continuing lack of evidence for an increased risk of malignancy in donors receiving G-CSF, the WMDA has re-issued a more reassuring statement. The revised statement was circulated to all WMDA member registries in late 2012 to replace the existing statement in consent forms, which now conclusively states that, 'Studies following large numbers of unrelated donors have shown that the risk of developing cancer within several years after the use of G-CSF is not increased compared with donors not receiving G-CSF'. Herein we review the evidence on which this statement is based.

WMDA guidelines for subsequent donations following initial BM or PBSCs.

Unrelated donor SCT activity is increasing, and in 5-10% of cases a subsequent donation of stem cells or donor lymphocytes may be requested. Second donations of stem cells are not associated with an increased chance of donor complications, but the yield of CD34+ cells may be lower in some donors. It is acceptable practice for any registry to request subsequent donations and it is recommended that donors should be counselled about this possibility before their first donation. Guidance is provided on the requirements for further medical assessment, the procedures used to agree requests, frequency and timing of donation and timing and duration of donor follow up.

Unrelated hematopoietic stem cell donors as research subjects.

Requests for participation of unrelated stem cell donors in research transplant protocols are becoming more frequent. World Marrow Donor Association calls on donor registries to participate in research activities. Here, we discuss various implications of research participation and make some recommendations as how to make this possible.

Donor characteristics as risk factors in recipients after transplantation of bone marrow from unrelated donors: the effect of donor age.

The National Marrow Donor Program (NMDP) maintains a registry of approximately 4 million volunteer unrelated donors for patients in need of a stem cell transplant. When several comparably HLA-matched volunteers are identified for a patient, various criteria are used to select a donor. A retrospective analysis of 6978 bone marrow transplantations facilitated by the NMDP from 1987 to 1999 was conducted to study the effects of various donor characteristics on recipient outcome. The evaluation addressed possible effects of donor age, cytomegalovirus serologic status, ABO compatibility, race, sex, and parity on overall and disease-free survival, acute and chronic graft-versus-host disease (GVHD), engraftment, and relapse. Age was the only donor trait significantly associated with overall and disease-free survival. Five-year overall survival rates for recipients were 33%, 29%, and 25%, respectively, with donors aged 18 to 30 years, 31 to 45 years, and more than 45 years (P =.0002). A similar effect was observed among HLA-mismatched cases (28%, 22%, and 19%, respectively). A race mismatch between recipient and donor did not affect outcome. The cumulative incidences of grade III or IV acute GVHD were 30%, 34%, and 34%, respectively, with donors aged 18 to 30 years, 31 to 45 years, and more than 45 years (P =.005). The corresponding incidences of chronic GVHD at 2 years were 44%, 48%, and 49% (P = 0.02). Recipients with female donors who had undergone multiple pregnancies had a higher rate of chronic GVHD than recipients with male donors (54% versus 44%; P <.0001). The use of younger donors may lower the incidence of GVHD and improve survival after bone marrow transplantation. Age should be considered when selecting among comparably HLA-matched volunteer donors.

Non-HLA barriers to unrelated donor stem cell transplantation.

A prospective survey involving 544 searches of the US National Marrow Donor Program (NMDP) Registry was conducted to identify reasons why many patients who have apparent HLA-matched donors do not proceed to transplant. Coordinators at NMDP transplant centers, patients and referring physicians were surveyed shortly after the initial search, and follow-up surveys were sent to the coordinators as the search was ongoing. The death of the patient, worsening of the patient's medical condition and length of the search process were the most commonly cited barriers to transplantation. Other times a decision was made not to transplant through the NMDP due to the use of a donor from another source, a preference for chemotherapy or immunotherapy, hesitancy on the part of the transplant physician or patient, or because the patient did not require a transplant. Responses differed between U.S. and international cases. An unrelated donor outside the NMDP was the most common reason cited by international coordinators (46%), whereas the death of the patient was the most common reason among US coordinators (13%). The death of the patient was the second most common reason cited by international coordinators at 9%. Financial problems were listed by 41% of US coordinators as a potential barrier at the time of initial search, but only 5% indicated this as an actual barrier on a follow-up survey. Finances were cited as the most important reason 3% of the time overall, and 6% for African Americans and Asian/Pacific Islanders.

Non-conservative substitutions distinguish previously uncharacterized HLA-A molecules.

The extent of class I HLA polymorphism is not yet realized, and to provide a glimpse of the HLA-A polymorphism which remains undetected, we have analyzed approximately 3,700 National Marrow Donor Program (NMDP) Donor/Recipient Pair Retrospective Study Samples with HLA-A DNA sequence-based typing (SBT). Seventeen new HLA-A alleles were detected, with a total of 19 nucleotide substitutions distinguishing these new alleles from their closest HLA-A relatives. Nearly all of the new alleles differ by single nucleotide substitutions; a majority of these substitutions can be explained by gene conversion events but 6 alleles likely originated by point mutation. Fifteen of the 19 nucleotide substitutions translate into amino acid differences in the molecule. Structurally, the inferred amino acid alterations were non-conservative in terms of chemical property, and most substitutions were positioned in 1 or more of the specificity pockets which determine peptide binding. Although these new alleles were identified in a primarily Caucasian sample population, 9 of the 17 new HLA-A alleles were found in samples of non-Caucasoid origin. A new allele detection rate of 1 in approximately 200 individuals in our data set would, therefore, be higher in a non-Caucasoid sample population. In summary, the single nucleotide substitutions that distinguish undetected HLA-A alleles translate into functionally distinct HLA-A molecules. Further studies of the role of HLA-A in transplantation, in disease association, and in evolution must therefore accommodate the discovery of new alleles differing by single nucleotides.

The National Marrow Donor Program. Meeting the needs of the medically underserved.

BACKGROUND: The National Marrow Donor Program operates the world's largest registry of volunteer unrelated stem cell donors. In recent years, the program has focused on building a large and diverse donor file. After initial recruitment, however, months or years may elapse before a potential donor is contacted on behalf of a searching patient. Here, the author begins to explore factors that influence donor availability at the confirmatory typing stage of the search process. METHODS: Over a 1-year period from March 1, 1999 through February 29, 2000, the author evaluated donor unavailability rates at the confirmatory typing stage of the search process. Unavailability rates by donor racial/ethnic group and by donor center were evaluated. To determine the consistency within individual donor centers, the author compared donor unavailability during the first 6 months of the observation period with unavailability during the second 6 months. RESULTS: Donor unavailability at confirmatory typing was higher among donors registered with domestic (U.S.) donor centers. The self-identified racial or ethnic group of the donor also affected the likelihood the donor will be available when requested. Between individual donor centers, there were large differences in the overall donor unavailability. Rates of donor unavailability tended to remain consistent at individual centers over time. CONCLUSIONS: This study suggests that procedures used at individual donor centers may dramatically impact donor unavailability. Future initiatives should undertake to identify best practice models for donor recruitment, retention, and subsequent contacts.

The National Marrow Donor Program: improving access to hematopoietic stem cell transplantation.

The National Marrow Donor Program has established a large and ethnically diverse registry of volunteer, unrelated stem cell donors. More than 9,000 patients world-wide have received transplants from NMDP donors. In recent years, substantial resources have been invested to increase that portion of the NMDP donor file with complete HLA-A, -B and -DR typing. While this effort has improved the likelihood of patients identifying a suitable matched unrelated donor, it has also focussed more attention on issues surrounding donor availability. Several initiatives underway or planned are intended to improve donor availability and increase the overall success of the unrelated donor stem cell transplantation.

G-CSF-mobilized donor leukocyte infusions as immunotherapy in acute leukemia relapsing after allogeneic marrow transplantation.

Eight patients who relapsed with acute leukemia within a year after allogeneic BMT were treated with G-CSF-mobilized donor leukocyte infusions (mDLI) to induce GvHD as a form of immunotherapy. Prior to mDLI, 7 who had systemic relapse received one (2 AML, 1 ALL, 1 CML myeloid blast crisis) or two (2 AML, 1 ALL) rounds of conventional dose induction chemotherapy, and 1 patient with isolated central nervous system (CNS) lymphoid blast crisis CML received intrathecal chemotherapy followed by craniospinal irradiation. G-CSF (10 microg/kg/day) was given to original HLA-matched sibling donors for 4-5 days before leukapheresis of at least 6.0 x 10(8) mononuclear cells per kilogram of recipient weight. No GvHD prophylaxis was used when mDLI was given in 6 patients at the nadir of hematologic counts and in 2 who were in hematologic remission. There was no regimen-related mortality, as pancytopenic patients had rapid recovery of neutrophil counts (6-18 days after mDLI). All patients developed moderate to severe GvHD (5 grade III/IV, 3 grade II) at a median of 30 days (range 22-59) after mDLI. Two patients died of complications from refractory GvHD while in remission. The other 6 had short remissions lasting 2.2-9.4 months until leukemic relapse as their GvHD was reversed by corticosteroids with or without cyclosporine. Patients who relapse with acute leukemia within a year after BMT still have a poor prognosis. The success of GvHD as a form of immunotherapy in these patients may depend on the ability to control it to a state that is both safe and continually exerting an antileukemia effect.

Sequence-based typing provides a new look at HLA-C diversity.

Although extensive HLA-A and HLA-B polymorphism is evident, the true diversity of HLA-C has remained hidden due to poor resolution of HLA-C Ags. To better understand the polymorphic nature of HLA-C molecules, 1823 samples from the National Marrow Donor Program research repository in North America have been typed by DNA sequencing and interpreted in terms of HLA-C diversification. Results show that HLA-Cw*0701 was the most common allele with a frequency of 16%, whereas 28% of the alleles typed as Cw12-18 (serologic blanks). The frequency of homozygotes was 9.8% as compared with previous studies of 18% for sequence-specific primers and 50% for serology. Most startling was the frequency at which new alleles were detected; 19 new HLA-C alleles were detected, representing a rate of approximately 1 in 100 samples typed. These new HLA-C alleles result from 29 nucleotide substitutions of which 4 are silent, such that coding substitutions concentrated about the Ag-binding groove predominate. Polymorphism at the HLA-C locus therefore resembles that at the HLA-A and HLA-B loci more than previously believed, indicating that antigenic stress is driving HLA-C evolution. However, sequence conservation in the alpha-helix of the first domain and a clustering of unique amino acids around the B pocket indicate that HLA-C alleles respond to antigenic pressures differently than HLA-A and HLA-B. Finally, because the samples characterized were predominantly from Caucasians, we hypothesize that HLA-C polymorphism will equal or exceed that of the HLA-A and -B loci as DNA sequence-based typing is extended to include more non-Caucasian individuals.

Long-term results of autologous marrow transplantation for relapsed or refractory male or female germ cell tumors.

Twenty-one patients with relapsed or refractory germ cell tumors were treated with high-dose chemotherapy and marrow transplantation (HDC/BMT) from 1982-1993. Primary sites of disease were testis (17), ovary (three), and pineal gland (one). Pathology included dysgerminoma (one), choriocarcinoma with adenocarcinoma (one), seminoma (four), and nonseminoma or mixed germ cell tumor (15). Nineteen had at least two prior chemotherapy regimens and eight had cisplatin-refractory disease defined as progression within 4 weeks of a cycle of cisplatin-based chemotherapy. HDC regimens were mostly combinations of cyclophosphamide with etoposide and cisplatin or carboplatin. There were only two treatment-related deaths (aspergillosis and interstitial pneumonitis). Times to engraftment of granulocytes (21+/-8.3 days) and platelets (32+/-20.2 days) were reasonable with only the last nine patients receiving growth factors. At a minimum of 4 years follow-up, eight patients have died of disease, six of whom were cisplatin-refractory prior to transplant. Eleven patients (52% overall) are alive and continuously free of disease after 4-10 years including one of three with refractory ovarian germ cell tumor. HDC/BMT provides significant long-term disease-free survival as salvage therapy for both male and female relapsed germ cell tumor patients who are not refractory to cisplatin.

Unrelated marrow donor registries.

Although large registries of volunteer, unrelated marrow donors are now operating around the world, potential recipients are frequently unable to identify suitable donors. These searching patients may benefit from 1) improved international coordination and cooperation; 2) a better understanding of the HLA system and the requirements for HLA matching; 3) improved methods for acquisition, storage, and manipulation of HLA-typing data; and 4) the development of alternative hematopoietic stem cell sources.

Cytomegalovirus surveillance and prevention in allogeneic bone marrow transplantation: examination of a preemptive plan of ganciclovir therapy.

Forty-two cytomegalovirus (CMV)-seropositive allogeneic marrow transplant patients or recipients of CMV-seropositive marrow allografts were entered into a surveillance program to detect and treat CMV infection during the first 120 days posttransplant. CMV infection was detected at a mean time of day 50 in 21/37 (58%) patients who had surveillance cultures. Twelve of 42 (28%) received preemptive ganciclovir treatment for virus isolated from blood (9 patients) or from bronchoalveolar lavage fluid (3 patients), and all had no CMV-associated sequelae. CMV disease was diagnosed in 5 patients (4 with pneumonia), 1 with gastroenteritis) who did not have positive cultures until the onset of their disease. CMV-related mortality was 4/42 (10%). Patients who earlier manifested lung injury or diffuse alveolar hemorrhage (DAH) were significantly predisposed to subsequent CMV pneumonia (P = 0.0013, Fisher's exact test) at a median onset of day 42. Restricted prophylactic use of ganciclovir in such patients may be indicated. Fifty percent of all patients never required ganciclovir during the surveillance period. When compared to a universal prophylaxis program of ganciclovir for the prevention of CMV disease, the use of ganciclovir in a preemptive strategy could avoid unnecessary therapy for a substantial number of patients and earn significant cost-savings.

High-dose carboplatin, etoposide and cyclophosphamide with autologous bone marrow transplantation for the treatment of advanced malignancies: a phase I study.

Carboplatin is a platinum-derivative widely used in conditioning regimens with ABMT, particularly in combination with cyclophosphamide and etoposide, drugs which co-express synergism in vitro. The objective of this study was to determine the maximum tolerated dose (MTD) of this combination. Thirty-four patients with refractory lymphoid or solid tumors were treated in a dose-escalation study with continuous infusion carboplatin (1.2-2 g/m2) on days -7 to -4, etoposide (1.2-2.4 g/m2) on days -7 to -5 and cyclophosphamide (120 mg/kg) given in two dose schedules: (1) day -3, -2; (2) day -9, -8. Autologous bone marrow or peripheral blood stem cells were infused on day 0. Mucositis/enterocolitis was dose limiting. In addition, severe cardiac dysfunction occurred in schedule 1 but not in schedule 2. Renal dysfunction occurred in the setting of fungemia, respiratory failure and congestive heart failure, and did not correlate with carboplatin dose. Hepatic and pulmonary dysfunction were minimal. The MTD was etoposide 2.1 g/m2 and carboplatin 2.0 g/m2, in combination with cyclophosphamide (120 mg/kg) on schedule 2. Responses were seen in 16 of 19 patients with measurable disease. Seven patients are disease-free survivors 50-60+ months post-ABMT. This study defines the MTD of carboplatin when combined with etoposide and cyclophosphamide in patients with adequate renal function and suggests significant anti-tumor activity.

Long-term survival after autologous bone marrow transplantation for metastatic breast carcinoma.

A phase II study of doxorubicin (Adriamycin)-based induction chemotherapy followed by cyclophosphamide/BCNU (CyBCNU) intensification and autologous bone marrow transplantation (ABMT) was conducted in 20 consecutive women with hormone-resistant metastatic breast cancer referred to our center. Of these 20 women, aged 24 to 56 (median age, 41), 9 had complete remission and 11 had partial remission after induction chemotherapy. Predominant sites of metastases included liver (5), lung (4), bone/bone marrow (5), and soft tissue (6). The dose of cyclophosphamide was 160 mg/kg and the dose of BCNU, 600 mg/m2, followed by infusion of a mean 2.30 x 10(8) nucleated marrow cells per kilogram of body weight. All patients achieved durable engraftment. Three patients remain disease-free at 62+, 67+, and 73+ months; two of these were in complete remission before ABMT. Actual relapse-free survival at 5 years is 15% and median survival from ABMT is 17 months. Induction chemotherapy followed by CyBCNU intensification in metastatic breast cancer can achieve prolonged relapse-free survival in 15% of patients, some of whom may be cured.

Autologous bone marrow transplantation for metastatic breast cancer.

UNLABELLED: Metastatic breast cancer accounts for 18% of cancer deaths among women in the U.S. Conventional combination chemotherapy produces responses in 50% to 80% of women with metastatic breast cancer, but is never curative. A major medical center administered high-dose chemotherapy and autologous bone marrow transplantation to 68 women with metastatic breast cancer between 1983 and 1993. Forty-nine of these women had estrogen receptor-negative tumors, a poor prognostic sign. Eighteen women with estrogen receptor-positive tumors had failed prior hormonal manipulation or had metastatic breast cancer at initial diagnosis. Prior to transplantation, 37 women were in first complete or partial remission, 8 were in their second complete or partial remission, 4 had stable disease, 14 had progressive disease, and 5 were in untreated relapse. Bone marrow transplantation preparatory regimens included high doses of single agents or combination chemotherapy. Among women not in remission before transplantation, 71% entered a partial or complete remission following transplantation. Overall, 29 women (43%) were in complete remission after marrow transplantation. Twelve women (18% overall) remain free of disease between 2 and 73+ months post-ABMT. Those in first complete or partial remission prior to transplant (37 patients) had a higher response rate (86%) and higher complete responses (62%), and 10 (27%) are free of disease. There were nine treatment-related deaths (13%). Forty-seven patients (69%) have died from breast cancer following autologous transplantation. Relapses occurred primarily at sites of previous disease. All relapses have occurred within 22 months of ABMT. CONCLUSION: Autologous bone marrow transplantation for metastatic breast cancer in first complete or partial remission has produced a 27% disease-free survival. This therapy should be considered for selected patients with metastatic breast cancer.

Autologous bone marrow transplantation for non-Hodgkin lymphoma.

UNLABELLED: Non-Hodgkin lymphomas (NHLs) are a group of malignant disorders that can be cured with chemotherapy and/or radiotherapy in 30% to 50% of cases. For those who fail initial therapy, cure is rarely achieved with standard dose chemotherapy; therefore higher doses of chemotherapy have been used with autologous bone marrow support. This major medical center has performed 74 autologous bone marrow transplants (ABMT) for patients with non-Hodgkin lymphoma who had failed initial therapy between 1984 and 1993. Preparatory regimens included high doses of chemotherapy with or without radiotherapy. There were 14 patients with low grade, 41 with intermediate grade, and 18 with high grade histologies. Among patients with low grade histologies, 90% responded and 50% are relapse-free between 1 and 33 months post-ABMT. Among patients with intermediate and high grade histologies, 25% are relapse-free between 2 and 80 months post-ABMT. CONCLUSION: Autologous bone marrow transplantation is effective in patients with relapsed non-Hodgkin lymphoma and should be considered an important therapeutic option.