Differentiation of antitumor-specific cytotoxic T lymphocytes from autologous tumor infiltrating lymphocytes in non-Hodgkin's lymphomas

The present study describes a new culture protocol allowing the activation and proliferation of autologous tumor infiltrating T lymphocytes (TIL), and the generation of antitumor specific CTL in non-Hodgkin's lymphoma (NHL). Cells from eight patients with indolent NHL were used. We performed 3-week co-cultures of TIL with irradiated autologous malignant B cells in the pres ence of low doses of IL-1 beta, IL-2 and IL-12. The proliferation, phenotyp e and cytotoxicity, and antitumor specificity of T cells recovered were stu died, T-cell clonality was analyzed using TCR gamma gene rearrangement ampl ification by a multiplex PCR, Under these culture conditions, TIL prolifera ted, and the CD8+ T lymphocytes that were in a minority at the beginning of the culture increased dramatically in 6 out of 8 cases. In two cases, CD4 T lymphocytes expanded. We showed that an oligoclonal selection of reactiv e T cells occurred in culture, Specific cytotoxicity developed against auto logous malignant B cells in the 6 cases where there was an expansion of CD8 + T lymphocytes. Inhibition experiments performed with mAb directed against HLA class I and II molecules, CD4, CD8 and TCR gamma delta showed that the cytotoxic effector cells were CD8+ T lymphocytes probably expressing TCR a lpha beta+. Cytokine secretion was analyzed in culture medium, and we detec ted significant levels of IFN-gamma, TNF-alpha, and IL-10 and no IL-4 (exce pt in one case). Our results demonstrate that memory T cells from lymphoma patients can be amplified and differentiated into antitumor cytotoxic cells using a combination of the cytokines IL-1 beta, IL-2, and IL-12 in associa tion with non modified tumor cells. (C) 1999 International Society for Expe rimental Hematology. Published by Elsevier Science Inc.