Serine hydrolases activate/inactivate trisubstituted nitrosoureas in dependence on intra- and extracellular enzyme location: an SCE study in Chinese hamster V79-E cells.

Abstract

Trisubstituted nitrosoureas are very stable in aqueous systems. But they are potent genotoxins in Chinese hamster V79-E cells, if no exogenous metabolizing system is added, and the mechanism of their genotoxic and carcinogenic activity has been largely unknown. This investigation shows that the sister-chromatid-exchange (SCE)-inducing capacity of 1,3-dimethyl-3-phenyl-1-nitrosourea (DMPNU) is eliminated by adding diisopropylflurophosphate (DFP) or porcine liver carboxylesterase to the incubation system. These effects are caused by two different mechanisms: (i) DFP inhibits endogenous amidases existing in V79-E cells, thus preventing the intracellular decomposition, which means an activation; and (ii) exogenous carboxylesterase cleaves DMPNU extracellularly, and the genotoxic decomposition product is obviously too short-lived to reach a critical intracellular target. A second trisubstituted nitrosourea, 3,3-diethyl-1-methyl-1-nitrosourea (DEMNU), which is mainly activated by monooxygenases, but in the absence of an exogenous metabolizing system also induces SCEs in V79-E cells, was studied in the same way. It was found that the 'direct' genotoxicity of DEMNU may be inhibited by DFP as well, but carboxylesterase decomposes this trialklynitrosourea with a much lower efficiency than DMPNU suggesting a low substrate affinity. The SCE-inducing capacity of both compounds is strongly influenced by the presence of calf serum in the culture medium. The nature of the serum factor is still unknown. Pathways for the amidase catalysis of DMPNU and for the activation of DEMNU by monooxygenases and amidases are proposed and discussed with respect to the topical or systemic carcinogenicity of these agents.