Improving outcomes in transplantation

Improving Outcomes in Transplantation Wolfram Brugger Follicular lymphoma and mantle cell lymphoma are incurable with standard chemotherapy regimens. One approach to improve outcome in patients with these diseases is high-dose therapy and autologous stem cell transplantation. Rituximab, an anti-CD20 monoclonal antibody, is specific for the B-cell surface antigen and can be used in autologous stem cell transplantation to eliminate lymphoma cells before the harvest (in vivo purging) or to prevent regrowth of malignant cells following transplant (post-transplant therapy). Preliminary data from an ongoing multicenter study evaluating the safety and efficacy of rituximab as a post-transplant consolidation therapy in patients with follicular lymphoma and mantle cell lymphoma are presented. After high-dose therapy and autologous stem cell transplantation together with rituximab treatment, 92% of patients are in complete remission at the 18month study follow-up, suggesting that rituximab is a valuable and potentially curative treatment for patients with follicular lymphoma and mantle cell lymphoma. Six months after treatment, all evaluable patients became polymerase chain reaction-negative for the bcl-1 and bcl-2 chromosomal rearrangements and remained so during follow-up, indicating that rituximab is able to eliminate minimal residual disease and bring about high rates of durable remissions in these patients. Semin Oncol 29 (suppl 6):23-26. Copyright 2002, Elsevier Science (USA). All rights reserved.

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OLLICULAR lymphoma (FL) and mantle cell lymphoma (MCL) are incurable with current treatments, leading to the study of new therapeutic approaches. Follicular lymphoma comprises approximately 20% of all non-Hodgkin’s lymphomas (NHLs) reported in Europe and the United States and exhibits an indolent clinical course.1 Most patients with FL are over the age of 50 and have widespread disease at diagnosis. Although conventional chemotherapy offers the possibility of remission, patients consistently relapse, with progressively shorter remission periods. No current therapy offers a cure. Mantle cell lymphoma represents approximately 8% of all NHLs, and has an indolent presentation but an aggressive clinical course.2 The disease is more than twice as common in males as in females and has a median age of onset of 58 years.3 Mantle cell lymphoma is characterized by the t(11;14) chromosomal translocation leading to overexpression of cyclin D1 and inhibition of cell-cycle control.4,5 Mantle cell lymphoma has a particularly poor prognosis with conventional therapy and has an overall survival of only 3 to 5 years, leading to Seminars in Oncology, Vol 29, No 2, Suppl 6 (April), 2002: pp 23-26

an urgent need for the development of new therapeutic strategies. The failure of conventional chemotherapy in the treatment of FL and MCL has led to the increasing use of high-dose therapy (HDT) with autologous stem cell transplantation (ASCT). Recent studies have shown that HDT with stem-cell transplantation may offer the chance of longer survival, particularly in the younger patient.6 Bone marrow had traditionally been an accepted source for repopulation of the hemopoietic system following HDT, but since the early 1990s, peripheral blood stem cells, collected by apheresis, have largely replaced bone marrow as a source of stem cells for autologous transplantation. Hemopoietic repopulation using peripheral blood stem cells appears to produce a shorter interval to the recovery of neutrophils and platelets than can be achieved by bone marrow transplantation.7 Although ASCT and high-dose chemotherapy can provide the possibility of longer survival rates,6 the rate of relapse remains unsatisfactorily high.8 Relapse is caused by regrowth of residual malignant cells, arising either as a result of contamination of the reinfused stem cells, or from cells remaining in the patient following high-dose chemotherapy. This has led to the implementation of treatments to improve the outcome of ASCT by eliminating the source of contamination. IN VITRO STEM CELL PURGING

To prevent contamination of the reinfused stem cells, several investigators have attempted to purge the harvested stem cells of malignant cells before reinfusion into the patient. Such in vitro purging involves treatment of the stem cell harvest with antibodies or agents to selectively kill or remove the malignant cells.9 Alternative methods involving positive selection of CD34⫹ stem cells have also been attempted.10 Although in vitro purging

From the Eberhard-Karls Universita¨t, Ha¨matologie und Onkologie, Medizinische Klinik II, Tu¨bingen, Germany. Address reprint requests to Wolfram Brugger, MD, EberhardKarls Universita¨t, Medizinische Klinik II, Abtlg. Ha¨matologie und Onkologie, Otfried-Mu¨ller Str. 10, 72076 Tu¨bingen, Germany. Copyright 2002, Elsevier Science (USA). All rights reserved. 0093-7754/02/2902-0605$35.00/0 doi:10.1053/sonc.2002.32751 23

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has shown that it is possible to reduce the level of tumor cell contamination, the procedures are time consuming and costly and, in most cases, lymphoma cells remain detectable. IN VIVO PURGING WITH RITUXIMAB

Rituximab is a human-mouse chimeric monoclonal antibody that is specific for the CD20 B-cell surface antigen. CD20 is a cell-surface phosphoprotein that is strongly and exclusively expressed at all stages of B-cell development. Studies have shown that 93% of B-cell lymphomas express the CD20 antigen.11 Rituximab has multiple mechanisms of action, which are distinct from those of chemotherapeutic agents. Rituximab, when bound to cells, initiates the classical complement activation pathway, thereby mediating direct lysis of CD20⫹ cells.12 Antibody-dependent cell-mediated cytotoxicity is initiated as natural killer cells and macrophages are mobilized by rituximab, causing a depletion of B cells from the lymph nodes, bone marrow, and peripheral blood, beginning from the first dose.12 Rituximab also can directly inhibit tumor cell proliferation by the induction of apoptosis.13 Rituximab does not deplete bone marrow stem cells, because uncommitted hemopoietic progenitor cells do not express CD20.14 As a monotherapy, rituximab has shown good efficacy in patients with indolent and aggressive NHL, and can induce remission in patients with MCL.15-17 In combination with cyclophosphamide/ adriamycin/vincristine/prednisone (CHOP) therapy, efficacy can be further improved and overall response rates of up to 95% have been achieved in patients with indolent NHL.18 During in vivo purging, rituximab is administered either before and/or during the stem cell mobilization process. Malignant cells are thus prevented from contaminating the stem cell harvest.

Killing malignant cells before harvest using activation of the patient’s own immune mechanisms is potentially much more efficient than trying to eradicate them later by in vitro methods. Importantly, several studies have shown that in vivo purging with rituximab does not reduce the yield of stem cells, and the time to engraftment and hemopoietic recovery is not compromised.19,20 RITUXIMAB: POST-TRANSPLANT MAINTENANCE THERAPY

While in vivo purging may be able to eliminate malignant cells from the harvest, it is possible that the HDT (along with rituximab) will not eliminate all tumor cells remaining in the patient. Killing residual malignant cells before they have had the chance to re-establish themselves and proliferate may reduce the probability of relapse. The administration of rituximab as a post-transplant consolidation or maintenance therapy is currently being evaluated in clinical trials. Studies have shown that administration of rituximab following ASCT causes a rapid depletion of CD20⫹ cells without any increase in infection.21-23 An ongoing, multicenter phase II study, set up to determine the safety and efficacy of rituximab for FL and MCL following HDT and ASCT, is generating encouraging data.23 Thirty patients were enrolled on to the trial and data from 28 patients (19 with FL, 9 with MCL) are currently available for efficacy and safety. The treatment schedule (summarized in Fig 1) consisted of VACOP-B chemotherapy (etoposide, doxorubicin, cyclophosphamide, vincristine, prednisone, and bleomycin) for 6 weeks, followed by mobilization of stem cells using etoposide/ifosfamide/cisplatin/epirubicin (VIP-E). CD34⫹ stem cells were then harvested from the patients’ peripheral blood.

Fig 1. Summary of the treatment schedule used during the rituximab plus CD34ⴙPBPCT transplantation trial. TBI, total body irradiation; Cy, cyclophosphamide; PBPCT, peripheral blood progenitor cell transplantation.

IMPROVING OUTCOMES IN TRANSPLANTATION

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Table 1. Response Rates Over Time Following HighDose Chemotherapy and Rituximab Treatment

Before TBI/Cy After TBI/Cy After rituximab 6 mos 12 mos 18 mos

No. of Patients

Clinical Response (%)

Partial Response (%)

28 28 28 24 16 12

9 43 55 50 88 92

91 57 45 50 12 8

Abbreviations: TBI, total body irradiation; Cy, cyclophosphamide.

Patients received a second round of VIP-E therapy. CD34⫹ stem cells, positively selected using magnetic cell sorting (eg, with the CliniMACS system; Bergisch-Gladbach, Germany), were transplanted back into the patient after conditioning with total body irradiation and cyclophosphamide. Approximately 8 weeks post-transplantation, patients were given four once-weekly infusions of rituximab. The clinical response rate has been impressive and has shown improvements over time (Table 1). Although only 12 patients have reached 18 months follow-up so far, the percentage of patients in complete remission has continuously increased, and at 18 months post-transplantation 92% were in complete remission. The t(14;18) chromosomal translocation is present in over 90% of patients with FL and leads to overexpression of the bcl-2 gene.24 The t(11;14) translocation (a characteristic of MCL) leads to overexpression of the bcl-1 gene.5 The presence of

Fig 2. Molecular remission in molecularly evaluable patients over the course of treatment and follow-up. * P ⴝ .042; † P ⴝ .0019; ‡ P < .001.

bcl-1 and bcl-2 can be detected by the highly sensitive polymerase chain reaction and may thus be used as molecular markers of minimal residual disease. Clearance from the bone marrow of cells carrying the bcl-2 rearrangement has been associated with a 48-fold lower risk of relapse and the clearance of bcl-2 positive cells can be viewed as a possible indicator of treatment efficacy.25,26 In parallel with the clinical observations, 25 patients from our study have been evaluable for molecular remission. Immediately following transplantation, 55% of evaluable patients were polymerase chain reaction-negative for bcl-1 (MCL) or bcl-2 (FL). This rose to 76% immediately following four infusions of rituximab, and at 6 months post-rituximab infusion, all evaluable patients were polymerase chain reaction-negative (Fig 2). These results suggest that clearance of residual lymphoma cells may continuously occur over a prolonged period following rituximab therapy. At a mean follow-up of 20 months (range, 5 to 39 months), all evaluable patients are still in complete molecular remission. In terms of the safety of rituximab given after HDT, the number of serious adverse events has been low throughout this study with only seven observed. The most common and serious adverse event was pneumonia, which resolved in all patients. High-dose therapy and ASCT resulted in one death (before rituximab treatment), producing an overall treatment mortality of only 3.6%. CONCLUSIONS

Indications from ongoing trials suggest that rituximab will also prove to be a valuable treatment

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for patients receiving ASCT. Initial results indicate that rituximab consolidation therapy, following high-dose chemotherapy and ASCT, is potentially curative for patients with follicular and MCL. Polymerase chain reaction analysis of bcl-1 and bcl-2 chromosomal rearrangements has shown that rituximab is able to eliminate minimal residual disease and is able to bring about high rates of durable remissions. Continued assessment of patients already on the trial, together with accrual of additional patients, will be necessary for corroboration of these results. REFERENCES 1. The Non-Hodgkin’s Lymphoma Classification Project: A clinical evaluation of the International Lymphoma Study Group classification of non-Hodgkin’s lymphoma. Blood 89: 3909-3918, 1997 2. Hiddemann W, Unterhalt M, Herrmann R, et al: Mantlecell lymphomas have more widespread disease and a slower response to chemotherapy compared with follicle-center lymphomas: Results of a prospective comparative analysis of the German Low-Grade Lymphoma Study Group. J Clin Oncol 16:1922-1930, 1988 3. Press O, Grogan T, Fisher R: Evaluation and management of mantle cell lymphoma. Adv Leuk Lymph 6:3, 1996 4. Tsujimoto Y, Yunis J, Onorato-Showe L, et al: Molecular cloning of the chromosomal breakpoint of B-cell lymphomas and leukemias with the t(11; 14) chromosome translocation. Science 224:1403-1406, 1984 5. Campo E, Raffeld M, Jaffe ES: Mantle-cell lymphoma. Semin Hematol 36:115-127, 1999 6. Rohatiner AZ, Johnson PW, Price CG, et al: Myeloablative therapy with autologous bone marrow transplantation as consolidation therapy for recurrent follicular lymphoma. J Clin Oncol 12:1177-1184, 1994 7. Champlin RE, Schmitz N, Horowitz MM, et al: Blood stem cells compared with bone marrow as a source of hematopoietic cells for allogeneic transplantation. IBMTR Histocompatibility and Stem Cell Sources Working Committee and the European Group for Blood and Marrow Transplantation (EBMT). Blood 95:3702-3709, 2000 8. Bierman PJ, Armitage JO: Non-Hodgkin’s lymphoma. Curr Opin Hematol 3:266-272, 1996 9. Tarella C, Corradini P, Astolfi M, et al: Negative immunomagnetic ex vivo purging combined with high-dose chemotherapy with peripheral blood progenitor cell autograft in follicular lymphoma patients: Evidence for long-term clinical and molecular remissions. Leukemia 13:1456-1462, 1999 10. Paulus U, Dreger P, Viehmann K, et al: Purging peripheral blood progenitor cell grafts from lymphoma cells: Quantitative comparison of immunomagnetic CD34⫹ selection systems. Stem Cells 15:297-304, 1997 11. Anderson KC, Bates MP, Slaughenhoupt BL, et al: Expression of human B cell-associated antigens on leukemias and lymphomas: A model of human B cell differentiation. Blood 63:1424-1433, 1984 12. Reff ME, Carner K, Chambers KS, et al: Depletion of B

cells in vivo by a chimeric mouse human monoclonal antibody to CD20. Blood 83:435-445, 1994 13. Shan D, Ledbetter JA, Press OW: Signaling events involved in anti-CD20-induced apoptosis of malignant human B cells. Cancer Immunol Immunother 48: 673-683, 2000 14. Nadler LM, Ritz J, Hardy R, et al: A unique surface antigen identifying lymphoid malignancies of B cell origin. J Clin Invest 67:134-140, 1981 15. McLaughlin P, Grillo-Lopez, AJ, Link BK, et al: Rituximab chimeric anti-CD20 monoclonal antibody therapy for relapsed indolent lymphoma: Half of patients respond to a four-dose treatment program. J Clin Oncol 16:2825-2833, 1998 16. Coiffier B, Haioun C, Ketterer N, et al: Rituximab (anti-CD20 monoclonal antibody) for the treatment of patients with relapsing or refractory aggressive lymphoma: A multicenter phase II study. Blood 92:1927-1932, 1998 17. Foran JM, Cunningham D, Coiffier B, et al: Treatment of mantle-cell lymphoma with rituximab (chimeric monoclonal anti-CD20 antibody): Analysis of factors associated with response. Ann Oncol 11:117-121, 2000 (suppl 1) 18. Czuczman MS, Grillo-Lopez A J, McLaughlin PCA, et al: Treatment of patients with low-grade B-cell lymphoma with the combination of chimeric anti-CD20 monoclonal antibody and CHOP chemotherapy. J Clin Oncol 17:268-276, 1999 19. Salles G, Moullet I, Charlot C, et al: In vivo purging with rituximab before autologous peripheral blood progenitor cell (PBPC) transplantation in lymphoma patients (pts). Blood 94:141a, 1999 (suppl 1) (abstr) 20. Haioun C, Delfau-Larue MH, Beaujean F: Efficiency of in vivo purging with rituximab followed by high-dose therapy (HDT) with autologous peripheral blood stem cell transplantation (PBSCT) in B-cell non-Hodgkin’s lymphomas (NHL). A single institution study. Blood 96:184a, 2000 (suppl 1) (abstr) 21. Horwitz SM, Breslin S, Negrin RS: Adjuvant rituximab after autologous peripheral blood stem cell transplant (APBSCT) results in delayed immune reconstitution without increase in infectious complications. Blood 96:384a, 2000 (suppl 1) (abstr) 22. Buckstein R, Imrie K, Spaner D, et al: High frequency of molecular remissions associated with Rituxan or IFN immunotherapy following ASCT for follicular NHL. Blood 96:791a, 2000 (suppl 1) (abstr) 23. Brugger W, Hirsch J, Repp R, et al: Treatment of follicular and mantle cell non-Hodgkin’s lymphoma with antiCD20 antibody rituximab after high-dose chemotherapy with autologous CD34⫹ enriched peripheral blood stem cell transplantation. Blood 96:482a-483a, 2000 (suppl 1) 24. Buchonnet G, Lenain P, Ruminy P, et al: Characterisation of BCL2-JH rearrangements in follicular lymphoma: PCR detection of 3’ BCL2 breakpoints and evidence of a new cluster. Leukemia 14:1563-1569, 2000 25. Lopez-Guillermo A, Cabanillas F, McDonnell TI, et al: Correlation of bcl-2 rearrangement with clinical characteristics and outcome in indolent follicular lymphoma. Blood 93:30813087, 1999 26. 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 81:3449-3457, 1993