title: In vitro biotransformation studies of 2-oxo-clopidogrel: multiple thiolactone ring-opening pathways further attenuate prodrug activation.
abstract: The biotransformation of clopidogrel has been under extensive investigation to address the observed high clinical variability and resistance of its antithrombotic prodrug therapy. Clopidogrel (M0) is first activated to its thiolactone intermediate, 2-oxo-clopidogrel (M2), by hepatic cytochrome P450 (P450) enzymes. Subsequent P450-catalyzed S-oxidation is followed by thioester hydrolysis, which cleaves the thiolactone ring and yields a sulfenic acid intermediate (M12); this intermediate is reduced to the final active metabolite (M13). The aim of the present study is to characterize the metabolic fates of M2 more comprehensively with focus on the thiolactone ring-opening pathways. It was found that the bioactivating S-oxidation confers on the thiolactone moiety not only one electrophilic site at the carbonyl C-atom (Site A), but also a second one at the allylic bridge C-atom (Site B). Both sites can react with H2O or other nucleophiles, like glutathione (GSH), leading to different thiolactone ring-opening pathways. In addition to the pharmacologically desired A-H2O pathway leading to M13 formation, the A-GSH pathway leads to the formation of a glutathione conjugate (GS-3), the B-H2O pathway leads to the formation of a desulfurized hydroxyl metabolite (M17), and the B-GSH pathway leads to the formation of a desulfurized glutathione conjugate (GS-2). These results demonstrate the reactive nature of the electrophilic thiolactone S-oxide intermediate (M11) and suggest that M13 formation from M2 might be accompanied by more attenuating pathways than previously reported. The research presented here may facilitate future studies exploring the clinical antithrombotic response to clopidogrel as well as the susceptibility to the adverse effect of clopidogrel and its close prodrug analogues.
authors.affiliation: MetabQuest Research Laboratory , 202 Chengfu Road, Beijing 100871, P. R. China.
authors.forename: Yaoqiu
authors.initials: Y
authors.lastname: Zhu
authors.forename: Jiang
authors.initials: J
authors.lastname: Zhou
mesh_terms: D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D004229:Dithiothreitol / Q000737:chemistry; D005978:Glutathione / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008862:Microsomes, Liver / Q000378:metabolism; D010084:Oxidation-Reduction; D010103:Oxygen Isotopes / Q000737:chemistry; D011355:Prodrugs / Q000032:analysis / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013455:Sulfur / Q000737:chemistry; D053719:Tandem Mass Spectrometry; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism; D014867:Water / Q000737:chemistry
chemical_list: D010103:Oxygen Isotopes; D011355:Prodrugs; D011994:Recombinant Proteins; D014867:Water; C540904:2-oxo-clopidogrel; D013455:Sulfur; D003577:Cytochrome P-450 Enzyme System; D005978:Glutathione; D013988:Ticlopidine; D004229:Dithiothreitol