Characterization of variant and parental-cross-protective immunity to immunogenic variants of a murine fibrosarcoma using the local adoptive transfer assay

Abstract

The purpose of this study was to characterize the lymphocyte populations responsible for rejection of immunogenic (Imm⁺) tumor variants, and the crossprotective immunity engendered by Imm⁺ variants against the weakly immunogenic parental tumor. Immunogenic clones of the weakly immunogenic methylcholanthreneinduced fibrosarcoma MCA-F have been generated using 1-methyl-3-nitro-1-nitrosoguanidine, 5-aza-2′-deoxycytidine, or ultraviolet radiation (UV-B; 280–320 nm). These clones grow progressively in immunosuppressed adult-thymectomized irradiated mice, but are rejected by immunocompetent syngeneic hosts. The parental MCA-F tumor grows progressively in both groups. Mice that have rejected a challenge of 1 × 10⁵ Imm⁺ cells show an anamnestic immune response against both the Imm⁺ clone and the parental MCA-F tumor. Using the local adoptive transfer assay and depletion of T-cell subsets with antibody plus complement, we show that immunity induced by the Imm⁺ variants against the parent MCA-F was mediated by the Thy1.2⁺, L3T4a⁺ population without an apparent contribution by Lyt2.1⁺ cells. Although antivariant immunity was also dependent upon Thy1.2⁺ cells, depletion of either the L3T4a⁺ or the Lyt2.1⁺ cells failed to abolish immunity against the variant. A role for Lyt2.1⁺ T lymphocytes in antivariant immunity, but not antiparent immunity, was supported by the results of cytotoxic T lymphocyte (CTL) assays. Following immunization with high numbers (1 × 10⁵ to 5 × 10⁵) of viable Imm⁺ cells, antivariant, but not antiparent CTL activity was detected in mixed lymphocyte tumor cell cultures. Immunization with lower numbers (3 × 10⁴) of viable Imm⁺ or with high numbers of mitomycin-C-treated Imm⁺ engenders only antivariant immunity without parental cross-protection. Under these conditions lymphocytes mediating immunity against the variant in the local adoptive transfer assay were exclusively of the Thyl1.2⁺, L3T4a⁺ phenotype, with no contribution from the Lyt2.1⁺ cells. Identical results were obtained for Imm⁺ clones of MCA-F induced by methylnitronitrosoguanidine, 5-azadeoxycytidine, and UV-B, suggesting that the nature of the antitumor immunity engendered by Imm⁺ is not significantly affected by the agent used. Furthermore, these results demonstrate that the cross-reactivity and cellular effectors of antitumor immunity in this system are influenced by the immunizing dose of Imm⁺ cells: the predominant effectors of both antivariant and parental-cross-reactive immunity were of the CD4⁺ T cell subclass, with a CD8⁺ cytotoxic population contributing to antivariant immunity only after highdose immunization.