Immunological characteristics of immunogenic variants induced in the MCA-F murine fibrosarcoma using 1-methyl-3-nitro-1-nitrosoguanidine, 5-aza-2'-deoxycytidine, or ultraviolet radiation.

The purpose of this study was to compare the frequency of generation and in vivo cross-reactivity of highly immunogenic (Imm+) clones induced in a single parental murine fibrosarcoma cell line MCA-F by 4 weekly treatments with either UV-B radiation, 1-methyl-3-nitro-1-nitrosoguanidine, or 5-aza-2'-deoxycytidine. These agents are believed to induce Imm+ variants by different mechanisms. The frequency of Imm+ variant generation was similar for the three different protocols, suggesting that the frequency of Imm+ generation was related more closely to the cell line than the inducing agent used. The strength of the immunogenic phenotype, however, was better correlated to the agent used, since 1-methyl-3-nitro-1-nitrosoguanidine yielded clones with the strongest immunogenicities. Three of four UV-B-induced Imm+ clones grew preferentially in chronically UV-irradiated syngeneic mice, a phenotype associated with UV-induced skin tumors. Cross-reactivity was tested with two Imm+ clones from each treatment group in a modified immunoprotection assay that selectively engendered antivariant, but not antiparental, immunity. Under these conditions each clone, except one, protected against itself. The clones displayed a complex pattern of cross-protection. Intervariant cross-protection was sensitive to the challenge dose, suggesting possible differences in the strengths of the cross-reacting immunities. Conversely, parental cross-protection was observed only with high immunizing multiplicities of Imm+ cells. The clones expressed the Imm+ phenotype in both C3H/HeN and C3H/HeJ mice, suggesting that expression of mammary tumor virus antigens did not account for the strong antitumor immune response. We also investigated whether the level of major histocompatibility complex class 1 or class 2 expression and immunogenic phenotype were correlated. Flow cytofluorography using haplotype-specific anti-Kk and anti-Dk monoclonal antibodies did not reveal a consistent difference in the constitutive or gamma-interferon-induced class 1 expression by Imm+ clones. However, we did observe a significant increase in the constitutive expression of IAk by most of the Imm+ variant clones. Together, these data demonstrate that in this system Imm+ variants engendered by a variety of mechanisms can express a range of cross-reactive tumor rejection neoantigens, independent of parental tumor antigens or major histocompatibility complex antigen expression.