Rat Liver Cytochrome P450-mediated Metabolic Activation of Methoxsalen and Structurally Related Compounds and its Relation to Enzyme Inhibition

Abstract

Abstract The metabolic activation and enzyme inhibition characteristics of methoxsalen were investigated in rat liver microsomes obtained from untreated animals and those treated with a number of prototypic inducers of cytochrome P450. Glutathione depletion assays have been carried out which show reactive metabolite generation to be markedly increased following phenobarbitone and β-naphthoflavone induction. Moreover, isoniazid induction led to levels of glutathione depletion significantly higher than those seen with other forms of induction, suggesting an important role for the cytochrome P4502E1 isozyme in the metabolic activation process. Methoxsalen was shown to be an extremely potent inhibitor of 7-ethoxycoumarin-O-de-ethylase activity, with inhibition constants of the order of μm with microsomes obtained from untreated, phenobarbitone- and β-aphthoflavone-induced animals. In contrast, constants obtained with microsomes obtained from isoniazid-induced animals were found to be markedly higher. Comparisons of the inhibition of 7-ethoxy and 7-pentoxyresorufin-O-dealkylase activities by methoxsalen and a number of structurally-related compounds have shown that a complete tricyclic ring system and an unsaturated 4′,5′-bond are structural prerequisites in the formation of reactive metabolites which inhibit cytochrome P450. These data implicate the furan ring system as the source of these metabolites and rule out the involvement of the pyrone ring system in the inhibition process.