O ⁶-Methylguanine-DNA methyltransferase activity and induction of novel immunogenicity in murine tumor cells treated with methylating agents

Summary

To investigate the mechanism of the generation of immunogenic tumor variants by mutagenic drugs, murine leukemia cells exhibiting different sensitivity to killing by the alkylator 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and different ability to repairO ⁶-methylguanine in their DNA were treated in vitro with a series of methylating agents, including triazene derivatives, temozolomide, and streptozotocin. At the population level, we found that BCNU-resistant cells (L1210/BCNU) that appeared to be cross-resistant to killing by a dimethyltriazene and expressed high levels ofO ⁶-methylguanine-DNA methyltransferase activity (mer⁺ phenotype) failed to generate highly immunogenic variant sublines on repeated exposure to the methylating agents. In contrast, all cells (L1210) that were susceptible to DNA alkylation damage and deficient inO ⁶-methylguanine repair (mer⁻) developed immunogenic variant sublines. A noticeable exception was represented by streptozotocin treatment, which was equally effective in mer⁺ and mer⁻ cells. At the clonal level, a single exposure to streptozotocin or a triazene derivative resulted in a high incidence (33% and 50%, respectively) of immunogenic cell generation in mer⁻ cells only. In mer⁺ cells, streptozotocin treatment led to a 33% incidence of immunogenic clones only when the cells were concurrently exposed toO ⁶-methylguanine as a free base. The activity ofO ⁶-methylguanine-DNA methyltransferase in mer⁺ cells was greatly reduced by treatment withO ⁶-methylguanine or streptozotocin, and the combination of the two drugs led to enzyme levels similar to those observed in mer⁻ cells. Taken together, these data suggest that the mechanism ofO ⁶-alkylation may be operative in the induction of novel tumor-cell antigenicity by methylating agents.