Dmd064949 1632..1641

Clopidogrel, a thienopyridine antiplatelet prodrug, is metabolized by oxidation to 2-oxo-clopidogrel, followed by conversion to its pharmacologically active thiol metabolite. After oral administration of clopidogrel to humans, two thiol isomers (H3 andH4) are observed in plasma,with similar concentrations, and only H4 is active in humans. In this work, themechanism of stereoselectivity in the formation and S-methylation of H3 and H4 was investigated in vitro. The two diastereomers of 2-oxo-clopidogrel were epimerized rapidly at physiologic pH. The intrinsic clearance (CLint) for H3 formation from 2-oxo-clopidogrel in human liver microsomes (HLMs) was 3.1-fold higher than that for H4 formation, indicating stereoselective metabolism. Kinetic studies using expressed enzymes demonstrated that the contributions of CYP2B6, CYP2C19, and CYP3A4 to the formation of H4 from 2-oxo-clopidogrel were 18.5%, 26.1%, and 53.5%, respectively. The CLint ratios of H3 formation to H4 formation from 2-oxo-clopidogrel by CYP2B6, CYP2C19, and CYP3A4 were 2.2, 1.0, and 1.7, respectively. In HLMs, H3 and H4 were further S-methylated, and the S-methylation was inhibited by 2,3dichloromethyl benzylamine, indicating the involvement of thiol S-methyltransferase. The CLint value for the S-methylation of H3 in HLMs was 98.1-fold higher than that for H4. The stereoselective formation of H3 from 2-oxo-clopidogrel and the stereoselective S-methylation of H3may lead to the similar exposure levels of H3 and H4 previously reported in humans. The epimerization of 2-oxoclopidogrel and the variations of thiol S-methyltransferase may affect the exposure to H4 in humans.