In vitro metabolism of exemestane by hepatic cytochrome P450s: impact of nonsynonymous polymorphisms on formation of the active metabolite 17β‐dihydroexemestane

Exemestane (EXE) is an endocrine therapy commonly used by postmenopausal women with hormone-responsive breast cancer due to its potency in inhibiting aromatase-catalyzed estrogen synthesis. Preliminary in vitro studies sought to identify phase I EXE metabolites and hepatic cytochrome P450s (CYP450s) that participate in EXE biotransformation. Phase I metabolites were identified by incubating EXE with HEK293-overexpressed CYP450s. CYP450s 1A2, 2C8, 2C9, 2C19, 2D6, 3A4, and 3A5 produce 17β-dihydroexemestane (17β-DHE), an active major metabolite, as well as two inactive metabolites. 17β-DHE formation in pooled human liver microsomes subjected to isoform-specific CYP450 inhibition was also monitored using tandem mass spectrometry. 17β-DHE production in human liver microsomes was unaffected by isoform-specific inhibition of CYP450s 2A6, 2B6, and 2E1 but decreased 12-39% following inhibition of drug-metabolizing enzymes from CYP450 subfamilies 1A, 2C, 2D, and 3A. These results suggest that redundancy exists in the EXE metabolic pathway with multiple hepatic CYP450s catalyzing 17β-DHE formation in vitro. To further expand the knowledge of phase I EXE metabolism, the impact of CYP450 genetic variation on 17β-DHE formation was assessed via enzyme kinetic parameters. Affinity for EXE substrate and enzyme catalytic velocity were calculated for hepatic wild-type CYP450s and their common nonsynonymous variants by monitoring the reduction of EXE to 17β-DHE. Several functional polymorphisms in xenobiotic-metabolizing CYP450s 1A2, 2C8, 2C9, and 2D6 resulted in deviant enzymatic activity relative to wild-type enzyme. Thus, it is possible that functional polymorphisms in EXE-metabolizing CYP450s contribute to inter-individual variability in patient outcomes by mediating overall exposure to the drug and its active metabolite, 17β-DHE.