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Tumor Infiltrating Lymphocytes (TILs) is one of the most promising sources of autologous cytotoxic T-cells for adoptive immunotherapy, which has already shown high efficiency in the treatment of metastatic melanoma. In this study, we isolated TILs from surgical material obtained by resection of solid tumors (primary and metastatic adenocarcinomas of various localization, melanoma, glioblastoma), studied their population composition and developed protocols for the purification expanding, and activation of CD4 +, CD8 + cytotoxic antitumor lymphocytes. Cells were obtained from samples of resected tumors in 15 patients; in each case we obtain an autologous pair: the primary tumor culture and the TILs culture. We could isolate viable lymphocytes in 100% of cases. Isolated TILs were successfully expanded in our specialized medium using various combinations of IL-2 / IL-15 / IL-21 // IL-7, anti-CD3 and anti-CD28. Immunophenotyping showed that the obtained TILs are a heterogeneous mixture of CD4 +, CD8 + cells containing populations of CD3 +, CD4, CD8 cells, CD4 + CD25 + CD127 T-regulatory cells, CD45RA, CCR7 + memory T-cells, CD56 + CD16- NK cells. The initial cultures of TILs were also characterized by a high level of PD1 expression, indicating their low antitumor cytotoxicity. Using different protocols of isolation, expansion, and activation, we obtained a cell preparation containing 80% of CD8+ PD1 activated TILs in an amount sufficient for adoptive therapy (500106 or more). An in vitro study of the cytotoxicity of obtained TILs in primary cultures of homologous tumors using RTCA Icelligence showed high cytotoxicity, providing almost 100% tumor cell death. Our developed protocol for the production and activation of TILs can be recommended for the phase I-II clinical trials of adoptive immunotherapy of recurrent, highly metastatic solid tumors.

TUMOR INFLATING LYMPHOCYTES. PURIFICATION, EXPANDING AND CYTOTOXICITY ANALISYS ON PRIMARY TUMOR CULTURES