Evidence in Escherichia coli that N3-methyladenine lesions and cytotoxicity induced by a minor groove binding methyl sulfonate ester can be modulated in vivo by netropsin

Abstract

The use of DNA equilibrium binding molecules to transfer alkyl groups to specific positions on DNA is an approach to generating cytotoxic DNA damage while avoiding the formation of promutagenic lesions that increase the risk for the development of secondary cancer. We have previously reported that in vitro a neutral DNA equilibrium binding agent based on an N-methylpyrrolecarboxamide dipeptide (lex) and modified with an O-methyl sulfonate ester functionality (Me-lex) selectively affords N3-methyladenine lesions in >90% yield relative to the formation of other adducts. While in vitro interactions between the lex dipeptide and DNA have been thoroughly studied, in vivo interactions are more difficult to elucidate. We report herein the relationship between the in vivo formation of N3-methyladenine and toxicity in wild-type and base excision repair defective mutant Escherichia coli. In addition, it is demonstrated that both N3-methyladenine adduction and cytotoxicity can be inhibited in vivo with netropsin, a potent competitive inhibitor of binding of lex to DNA. The results show a clear relationship between the levels of N3-methyladenine and toxicity in an alkA/tag glycosylase mutant that cannot remove the adduct from its genome. For methyl methanesulfonate, which does not sequence selectively methylate DNA, a relationship between the formation of N3-methyladenine and toxicity is also observed. However, netropsin affects neither the level of N3-methyladenine nor the toxicity of methyl methanesulfonate in E. coli.