The present study investigates the effect of IL-12 administration on the generation of lymphoid cells that exhibit cytotoxicity against tumor cells expressing Fas Ag. Systemic injection of rIL-12 into BALB/c or (B6C3)F1 mice bearing syngeneic CSA1M or OV-HM tumor induced complete tumor regression. CSA1M tumor cells expressed Fas Ag, and exposure of these cells to IFN-gamma enhanced Fas expression. In contrast, Fas Ag was hardly detected on OV-HM cells even after IFN-gamma exposure. Only CSA1M cells were lysed by anti-Fas mAb or cells expressing Fas ligand (FasL), indicating that Fas on CSA1M cells is functional in mediating cell death. An increase in the frequency of lymphoid cells characterized as CD3+ CD4- CD8- B220+ was observed in spleens from both CSA1M and OV-HM tumor-bearing mice after IL-12 treatment. A splenic population enriched in cells with these unique phenotypes exhibited considerable degrees of cytotoxicity against Fas+ CSA1M, but not against Fas- OV-HM tumor cells. The lysis of CSA1M cells was almost completely blocked by addition of Fas-Fc, a fusion protein between the extracellular domain of mouse Fas and the Cgamma1 domain of human Ig. Regressing CSA1M and OV-HM tumor masses after IL-12 treatment exhibited a massive lymphoid cell infiltration and expressed significant levels of FasL mRNA, suggesting the infiltration of FasL-expressing cells to tumor sites. These results indicate that IL-12 induces the expansion of lymphoid cells that exhibit FasL-mediated cytolytic activity and accumulate into regressing tumor masses.