Rejection of Primary Lymphoma Cells Derived from c-myc-Transgenic Mice after Haploidentical Transplantation: Treatment of the Tumor May Confer to Resistance Against GvL-Effect.

Abstract

The long term prognosis of the High Grade B-cell-lymphoma in adults is still very poor, especially in the case of resistance to chemotherapy. To date allogeneic stem cell transplantation as a last treatment option results in poor outcomes due to high transplant related mortality or disease progression. This dilemma raises the question of whether transplant related mortality could be reduced by earlier transplantation, and whether such tumors can be recognized by an allogeneic graft providing a graft versus lymphoma effect. The immunology behind allogeneic rejection of high grade lymphomas is poorly understood due to a dearth of good animal models. Using a transgenic mouse model in which the human c-myc oncogene is driven by the human immunoglobuline lambda locus mimicking a t(8;22) translocation of Burkitt's lymphoma, we established primary lymphoma cell lines. C57BL/6c-myc tg mice were crossbred with DBA mice and lymph nodes from tumor bearing F1-mice were explanted and seeded on irradiated human MRC5 fibroblast feeder cells. Approximately 70% of all explanted lymphomas established long term cultures which could be maintained free of feeder cells after 4 to 6 weeks. FACS analysis of lymphoma cells displayed strong CD19 expression, reduced expression of MHC Class I and II and, in several cases, low expression of costimulatory molecules such as CD80, CD86 and CD54. The phenotype was maintained over time with a follow up of up to 4 months. As few as 100 lymphoma cells gave rise to orthotopic tumors or systemic disease within 40 days after s.c. or i.v. injection, respectively. On the contrary, when 50 Mio. F1-lymphoma cells were injected into haploidentical C57BL6 parental animals, no tumor growth could be observed after at least 100 days of follow up. Despite fast growth and low expression of MHC Class I and costimulatory molecules, lymphoma cells were easily rejected in a haploidentical setting. Ex vivo chemotherapy treatment of lymphoma cells resulted in resistance after 2 courses of 100nM Adriamycine. Increased doses up to 500nM Adriamycine further accelerated resistance and growth speed of the cells. Flow cytometric analysis indicated a reduced fraction of apoptotic cells in culture suggesting selection for cells bearing alteration in the apoptotic pathways. In further experiments altered cells will be tested in haploidentical settings to confirm clinical experience in our model.