Killing of Fas ligand-resistant renal carcinoma cells by interleukin-2-andBCG-activated effector cells

Activated cytolytic effector cells like lymphokine-activated killer (LAK) a nd the recently described bacillus-Calmette-C ui rin-activated killer (BAK) cells are thought to mediate antitumor effects against metastatic renal ce ll carcinoma (RCC) and superficial bladder cancer respectively, Perforin an d Fas ligand (FasL) have been described as the major lytic principles in ce llular cytotoxicity. Using a radioactive-release assay and specific inhibit ors, we investigated the molecular mechanisms used by LAK and BAK cells in the Lysis of renal carcinoma cells. In addition, we evaluated the susceptib ility of RCC cells to FasL-mediated cytotoxicity. LAK and BAK cells effecti vely lysed the renal cancer cell line SK-RC-35 upon cell-cell contact. Both effector cell populations were shown to produce perforin and Fast as deter mined by reverse transcriptase/polymerase chain reaction RT-PCR. Using fluo rescence-activated cell sorting analyses and RT-PCR, we detected a marked F as receptor (Fas, CD95) expression on RCC cells. However, RCC cells were sh own to be resistant to killing by recombinant Fast rind lysis by BAK and LA K cells was not inhibited in the presence of anti-FasL antibody. In contras t, the cytotoxicity exerted by LAK and BAK cells was drastically reduced il l the presence of the Ca2(+)- chelating agent EGTA as well as concanamycin A, a specific inhibitor of perforin-mediated lysis. These results demonstra te that cytolysis of Fast-resistant RCC cells by activated immune cells is mediated via perforin. Our findings give Further insights into the molecula r mechanisms involved in the elimination of RCC by cytotoxic lymphocytes ac tivated with biological response modifiers.