Rapid analysis of base-pair substitutions induced by mutagenic drugs through their oxygen radical or epoxide derivatives

Abstract

Among the drugs that induce base-pair substitution mutations in the Salmonella reversion assay are the nitric oxide (NO)-delivery drug, diethylamine NONOate (DeaNO), and the ovarian cancer chemotherapeutic drug, treosulphan (TE). The present study compared the mutation spectra generated by DeaNO and TE in the hisG46 strains, TA1535 and TA100, the hisG428 strain, TA102, and the six Ames II™ 7000 series strains. Using these strains, it was feasible to conduct rapid analysis of the type and magnitude of induced mutation without resorting to DNA amplification and sequencing. A putative hydrolysis product of TE, 1,2:3,4-diepoxybutane (DEB), and hydrogen peroxide (H 2O 2) were included in the study to allow for further comparisons between epoxide-induced damage and that induced by the hydroxyl radical. TE (0.93 μmole/pl) induced 16.8-fold-over-background reversion or a mutagenicity ratio (MR) of 16.8 in TA1535. The response was weaker in TA100 (MR of 3), and negative in strain TA102. Only two Ames II™ strains demonstrated sensitivity to TE, and they were TA7004 (CG:AT) and TA7005 (GC:AT). Like TE, DeaNO (33 μmole/pl) was mutagenic in TA1535 (MR of 24.6), TA100 (MR of 5.3), TA7004 (MR of 13.7), and TA7005 (MR of 7.7), and non-mutagenic in TA102. These results showed a preferential sensitivity to reversion of the CCC target in TA100 and TA1535, and a lack of sensitivity to reversion of the TAA target in TA102. In addition, they elucidated the selectivity of the Ames II™ strains, with AT targets showing little or no sensitivity to reversion. The TE-epoxide derivative DEB was mutagenic in TA1535 and TA7004, but in contrast to TE, DEB was mutagenic in TA102. Interestingly, TA102 was reverted by DEB and H 2O 2 but not by TE or DeaNO. This study showed that analysis of mutations is achievable using the battery of strains listed above. The fact that DNA damage can be detected by reversion at specific bases offers a tool for understanding the mechanisms through which drugs may exert their DNA and cellular damage.