Quantitative forward-mutation specificity of mono-functional alkylating agents, ICR-191, and aflatoxin B1 in mouse...

We have analyzed forward-mutation specificity in S49 mouse T lymphoma cells. Our criteria of specificity were based upon relative mutabilities of a panel of 3 genetic markers: resistance to 6-thioguanine (6TGr), dibutyryl cAMP (bt2cAMPr), and ouabain (OUAr). We tested 2 monofunctional alkylating agents, ethyl methane-sulfonate (EMS) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), and 2 heterocyclic compounds, ICR-191 and aflatoxin B1 (AFb1). AFB1 was activated with rat-liver microsomal S9 plus cofactors. Expression-time lags of each genetic marker ranged from 2 days of OUAr mutations to 6 and 8 days for bt2cAMPr and 6TGr cells, with stable induced mutant fractions thereafter. The relative activity of each agent for each marker was assessed on the basis of its mutagenic efficiency at equitoxic doses. Specificity differences between the agents were determined by taking ratios of mutagenic efficiencies (RME) for the 3 possible pairs of markers. From these quantitative correlations and other data we conclude that both MNNG and EMS induce ouabain resistance (and probably bt2cAMPr and 6TGr) by similar mechanisms, almost certainly base substitutions. In contrast, ICR-191 and AFB1 are respectively less than 2 and 3% as efficient as MNNG for OUAr mutant induction relative to the activity of each agent for 6TGr mutagenesis. We infer that ICR-191 and AFB1 very rarely cause base substitutions in S49 cells, but that their activities are consistent with production of deletions, insertions or chromosomal aberrations. S49 cells demonstrate an unusually high specificity of mutagenesis at the OUAr locus compared to several other rodent cell lines. Thus, this panel of markers in S49 cells can be used as a sensitive, reliable screening system for mutagen detection and to discriminate among major classes of mutagenic mechanisms.