Stem cell transplantation in multiple myeloma : clinical aspects and experimental gene transfer studies

Abstract: Multiple myeloma (MM) is a hematological malignant disease considered essentially incurable despite its sensitivity to cytotoxic chemotherapeutic drugs and radiation. High-dose chemo/radiotherapy with allogeneic- or autologous hematopoietic stem cell transplantation (tx) can induce a disease-free state and may prolong survival and offer an improved quality of life to MM patients. The aim of this thesis was to improve the results of high-dose treatment with hematopoietic stem cell tx in MM. 523 patients reported to the Myeloma Registry of the European Group for Blood and Marrow Transplantation (EBMT) were subjected to retrospective studies: 334 patients who underwent autologous stem cell transplantation (ASCT) were analyzed with respect to parameters of prognostic importance for the outcome after tx. 189 patients who underwent allogeneic bone marrow transplantation (BMT) with an HLA-matched sibling donor were compared to an equal number of patients treated with ASCT after case-matching for gender and number of lines of previously administered treatment, and were evaluated with respect to the outcome after tx. A program utilizing two sequential cycles of high-dose treatment and autografting was applied on 23 patients at Huddinge Hospital. and was prospectively evaluated with respect to feasibility and efficacy, and selected patients in complete remission (CR) were analyzed for minimal residual disease (MRD) using tbe PCR-based immunoglobulin-gene "fingerprinting" method. In the experimental studies, the effect of including basic fibroblast growth factor (bFGF) in the prestimulation medium in addition to the three-cytokine combination of stem cell factor, interleukin-3 and interleukin-6 on the rate of retroviral-mediated transfer (RMGT) of the neomycin resistance (neoR) gene in human hematopoietic cells in vitro was assessed, and the feasibility of RMGT in antibody-separated human MM cells in vitro was studied. Finally, a clinical protocol for gene marking of CD34+ cells in the autograft, with the aim to detect a possible graft-origin of recurring MM cells at relapse after ASCT, was prepared and activated. For the 334 ASCT-patients analyzed, the median overall survival (OS) after tx was 38 months and actuarial OS at 90 months was 25%. Factors significantly predictive for a better OS or progression-free survival (PFS) following ASCT were tx as part of front-line treatment before disease progression, responsiveness to chemotherapy, male gender, post-tx alpha-interferon maintenance treatment, a low beta-2-microglobuln value at diagnosis, and to achieve CR after tx, and tbe first four of these parameters were also independently predictive for a better outcome in a multivariate analysis. In addition, for patients in first chemotherapy-responsive phase of disease, stage I-II at diagnosis and treatment in a double-ASCT program were associated with a signficantly better outcome. In the comparison between BMT and ASCT, OS was significantly better after ASCT (median OS 34 months, vs 18 forBMT), which was evident during the first 3 years post-tx, but later the survival curves merged and long-term OS was similar. The survival difference was only observed in males, while in females the survival was similar. Transplant-related mortality (TRM) was significantly higher for BMT, which was the primary reason for the poorer outcome, even though relapse rate was higher after ASCT. For the double-ASCT program, the CR-rate was 69%, actuarial OS at 49 months was 85%, and 61% of the patients were in continuous CR. There were two MM-unrelated deaths. The original clonal band could initially not be detected by PCR in any of the six patients analyzed, but later a band appeared in two patients who subsequently relapsed from CR. RMGT into human hematopoietic cells was significantly enhanced by the addition of bFGF to the three-factor medium. The transduction rate in terms of G418-resistant CFU-GM was estimated to 38% and 27% with and without bFGF, respectively, and the optimal concentration of bFGF was in the range between 20 and 2tl0 ng/ml. The feasibility of RMGT in human MM cells was demonstrated, with G418-resistance ranging between 1.5% and 50% in liquid cultures. The presence of the neoR-gene in the transduced cytohne-stimulated hematopoietic cells as well as in the transduced human MM cells was proven by PCR-analysis. In conclusion, the outcome after ASCT in MM patients is influenced by several factors of prognostic importance. Survival is significantly better after ASCT as compared to BMT, but the relapse rate is lower after BMT, and the main reason for the difference in survival is the higher rate of TRM associated with BMT. A program with two sequential ASCT-procedures is feasible, and em induce long-term CR at a substantial rate, where MRD in some CR-patients could not be detected by PCR. A foreign selectable marker gene can be introduced by RMGT into MM cells in vitro, and this forms the basis of a clinical study with the intention to mark cells in the autograft prior to reinfusion, which may enable the determination of the origin of clonal MM cells at relapse following ASCT.