Covalent interaction of 5-nitroimidazoles with DNA and protein in vitro: mechanism of reductive activation

Abstract

Human hepatic microsomal enzymes catalyzed the NADPH-dependent anaerobic reductive activation of [1-14C]metronidazole [1-(2-hydroxyethyl)-2-methyl-5-nitroimidazole] and [4,5-14C]ronidazole [(1-methyl-5-nitroimidazole-2-yl)methyl carbamate] to species that became covalently bound to proteins. Due to the low efficiency of the enzyme-catalyzed covalent binding of metronidazole, the stoichiometry of anaerobic reductive activation was studied with dithionite as the reductant. Two moles of dithionite was consumed per mole of [1-14C]metronidazole for maximal covalent binding to either DNA or immobilized sulfhydryl groups, demonstrating that four electrons are required for the reductive activation of metronidazole. These data implicate the involvement of a hydroxylamine in covalent binding. Maximal covalent binding of [4,5-14C]ronidazole to DNA also required four-electron reduction, consistent with previous studies of the covalent binding of this agent to immobilized sulfhydryl groups [Kedderis et al. (1988) Arch. Biochem. Biophys. 262, 40-48]. Studies of the covalent binding of variously radiolabeled ronidazole molecules to DNA suggested that the imidazole ring was intact while greater than 80% of the 2-carbamoyl group and the C4 proton were not present in the DNA adduct. Studies of both the chemical and human hepatic microsomal reduction of [4-3H]metronidazole demonstrated that covalent binding occurred with the stoichiometric loss of this label, implicating binding at the C4 position. These results suggest that the reductive activation of 5-nitroimidazoles generally proceeds via four-electron reduction to form hydroxylamines followed by nucleophilic attack at C4.

Activation of Benznidazole by Trypanosomal Type I Nitroreductases Results in Glyoxal Formation

“…Where oxygen is limited, the nitroso form can be produced either by a further single-electron reduction step or by the interaction of two nitro anions (26). The nitroso molecule is highly reactive and can lead to cellular damage directly or indirectly through the formation a hydroxylamine derivative (16,18). This pathway is believed to underlie the selective toxicity of nitroimidazoles against infections caused by anaerobic/microaerophilic microbes and prodrug activation in hypoxic tumors.…”

“…3). Because of their transient nature, we could not analyze the effect of these intermediates, but by analogy with the equivalent derivatives obtained from other nitroimidazoles, we can postulate that the hydroxylamine form of benznidazole could react with nucleic acids and proteins, while the nitrenium ion could conjugate with free thiols (16,45). Such interactions may account for some of this prodrug's pleiotropic effects: benznidazole is mutagenic and causes a thiol depletion (4,22).…”

Effects of Renal Impairment on the Pharmacokinetics of Morinidazole: Uptake Transporter-Mediated Renal Clearance of the Conjugated Metabolites

“…5-Nitroimidazole antimicrobial drugs act via nitroreduction of the molecules in the pathogens (1). As an analog of metronidazole, morinidazole appears to show greater antiparasitic potency against T. vaginalis and amoebic protozoa, but it exhibited relatively weaker toxicity in vitro and in preclinical animal studies compared with metronidazole (2).…”

Metabolic Activation of Benzodiazepines by CYP3A4

“…In the reductive pathways from nitro to the fully reduced amine catalyzed by P450 and/or reductase, several reactive metabolites including nitroso and N-hydroxylamine derivatives could be produced. Such reactive metabolites seem to bind covalently to nucleophilic targets of proteins and nucleic acids, leading to the cytotoxic effects (Biaglow et al, 1986;Rickert, 1987;Kedderis and Miwa, 1988;Kedderis et al, 1989). On the other hand, arylamines are metabolically activated by P450-mediated N-hydroxylation.…”