title: CYP-independent inhibition of platelet aggregation in rabbits by a mixed disulfide conjugate of clopidogrel.
abstract: Dual antiplatelet therapy with clopidogrel and aspirin has been the standard of care in the United States for patients with acute coronary syndromes (ACS) and/or undergoing percutaneous coronary interventions (PCI). However, the effectiveness of clopidogrel varies significantly among different sub-populations due to inter-individual variability. In this study we examined the antiplatelet potential of a novel mixed disulfide conjugate of clopidogrel with the aim to overcome the inter-individual variability. In the metabolic studies using human liver microsomes and cDNA-expressed P450s, we confirmed that multiple P450s are involved in the bioactivation of 2-oxoclopidogrel to H4, one of the diastereomers of the pharmacologically active metabolite (AM) possessing antiplatelet activity. Results from kinetic studies demonstrated that 2C19 is the most active in converting 2-oxoclopidogrel to H4 with a catalytic efficiency of 0.027 µM⁻¹min⁻¹ in the reconstituted system. On the basis of this finding, we were able to biosynthesise the conjugate of clopidogrel with 3-nitropyridine-2-thiol, referred to as clopNPT, and examined its antiplatelet activity in male New Zealand white rabbits. After administration as intravenous bolus at 2 mg/kg, the clopNPT conjugate was rapidly converted to the AM leading to the inhibition of platelet aggregation (IPA). Analyses of the blood samples drawn at various time points showed that intravenous administration of clopNPT led to ~70% IPA within 1 hour and the IPA persisted for more than 3 hours. Since the antiplatelet activity of clopNPT does not require bioactivation by P450s, the mixed disulfide conjugate of clopidogrel has the potential to overcome the inter-individual variability in clopidogrel therapy.
authors.affiliation: Dr. Haoming Zhang, Department of Pharmacology, The University of Michigan Medical School, 1150 West Medical Center Drive, 2220A MSRB III, Ann Arbor, MI 48109-5632, USA, Tel.: +1 734 764 6182, Fax: +1 734 763 4450, E-mail: haom@umich.edu.
authors.forename: H
authors.initials: H
authors.lastname: Zhang
authors.forename: D A
authors.initials: DA
authors.lastname: Lauver
authors.forename: P F
authors.initials: PF
authors.lastname: Hollenberg
mesh_terms: D000818:Animals; D001711:Biotransformation; D000077144:Clopidogrel; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004220:Disulfides / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007275:Injections, Intravenous; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D010974:Platelet Aggregation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D011355:Prodrugs / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D013988:Ticlopidine / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology
chemical_list: D004220:Disulfides; D010975:Platelet Aggregation Inhibitors; D011355:Prodrugs; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D000077144:Clopidogrel; D013988:Ticlopidine
keywords: Clopidogrel; active metabolite; antiplatelet; inter-individual variability; mixed disulfide conjugate