A comparative study of tamoxifen metabolism in female rat, mouse and human liver microsomes

Abstract

The metabolisms of tamoxifen in female rat, mouse and human liver microsomal preparations were compared. Rat, mouse and human liver microsomes were incubated with tamoxifen in the presence of NADPH and MgCl2 and the metabolites formed were analysed by on-line HPLC-electrospray ionization MS. The major metabolites formed by rat liver microsomes were 4-hydroxytamoxifen, 4'-hydroxytamoxifen, N-desmethyltamoxifen and tamoxifen N-oxide. In addition, two epoxide metabolites, 3,4-epoxytamoxifen and 3',4'-epoxytamoxifen, and their hydrolysed derivatives, 3,4-dihydrodihydroxytamoxifen and 3',4'-dihydrodihydroxytamoxifen, have been identified. The pattern of the main metabolites obtained with human liver microsomes resembles qualitatively that of rat liver microsomes. The major differences between rat and human liver microsomes were that the amount of hydroxylated metabolites were much lower in human and only traces of 3,4-epoxytamoxifen and the corresponding dihydrodihydroxy derivative were detected. No 3',4'-epoxytamoxifen was detected in human liver microsomes. The four major metabolites were also formed in much larger amounts and with faster rates of formation by mouse liver microsomes, though tamoxifen N-oxide clearly predominated in this species. Polar metabolites, 3,4-dihydroxytamoxifen and 4-hydroxytamoxifen N-oxide, which were undetectable in rat and human, were formed in significant amounts in mouse microsomes. As in human microsomes, there was only one epoxide metabolite, 3,4-epoxytamoxifen, produced by mouse liver microsomes at levels lower than that found in rat. The faster rate of metabolism and the production of polar metabolites may indicate the ability of mouse to detoxify tamoxifen by rapid elimination compared with rat and human. The production of a larger amount of potentially reactive epoxide metabolites in rat may be responsible for the liver carcinogenesis in this species.

New insights into the metabolism of tamoxifen and its role in the treatment and prevention of breast cancer

“…Although many candidates were described [75][76][77][78], the metabolite found to be responsible for the initiation of rat liver carcinogenesis is α-hydroxytamoxifen [79][80][81][82][83] (Figure 5) Alpha-hydroxytamoxifen has been resolved into R-(+) and S-(−) enantiomers. Metabolism by rat liver microsomes gave equal amounts of the two forms, but in hepatocytes the R form gave 8x the level of DNA adducts as the S form.…”

Evaluation of tamoxifen and alpha-hydroxytamoxifen 32P-post-labelled DNA adducts by the development of a novel automated on-line solid-phase extraction HPLC method

“…At least two distinct metabolic pathways lead to DNA adduct formation, the first is via α-hydroxytamoxifen (21-23) and the second is via an arene oxide or 4-hydroxylation, which leads to the formation of a quinone methide (24)(25)(26). Adduct peaks formed by 4-hydroxytamoxifen HPLC analysis of post-labelled DNA reacted in vitro with 4-hydroxytamoxifen gave one major peak that co-eluted with rat peak number 1.…”

α-Hydroxytamoxifen, a genotoxic metabolite of tamoxifen in the rat: identification and quantification in vivo and in vitro

“…The reactive intermediate generated during the oxidative metabolism of tamoxifen by microsomal P450 binds irreversibly to protein and DNA (16,18). Tamoxifen is metabolized variously via primary N-demethylation, 4-hydroxylation, α-hydroxylation and N-oxidation ( Figure 1) by hepatic microsomes and cell lines (19)(20)(21)(22)(23), and additionally via secondary O-glucuronidation in hepatocytes (24) and in vivo (25)(26)(27)(28)(29), Ndemethylation being the principal biotransformation in humans and human liver microsomes (20,23,28). Both aromatic hydroxylation and α-hydroxylation have been consistently implicated in the formation of reactive tamoxifen metabolites in vitro and in experimental animals, and they appear to represent the initial steps of distinct bioactivation pathways in vivo (16,30).…”