Clarifying the importance of CYP2C19 and PON1 in the mechanism of clopidogrel bioactivation and in vivo antiplatelet response

Gong, Inna Y.; Crown, Natalie; Suen, Colin; Schwarz, Ute I.; Dresser, George K.; Knauer, Michael J.; Sugiyama, Daisuke; DeGorter, Marianne K.; Woolsey, Sarah; Tirona, Rommel G.; Kim, Richard B.

doi:10.1093/eurheartj/ehs042

Cited by 71 publications

(68 citation statements)

References 33 publications

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“…In contrast, in our study, the formation of H4 from thiolactone 2 by CYP2C8 was negligible (data not shown). Gong et al (2012) reported that CYP2C19 and CYP3A4 were involved in the formation of H4 from thiolactone 2, which was consistent with our study. CYP2B6 and CYP3A4 are subjected to induction and inhibition, which may affect the metabolism and efficacy of clopidogrel.…”

Section: Discussionsupporting

confidence: 93%

Human Liver Cytochrome P450 Enzymes and Microsomal Thiol Methyltransferase Are Involved in the Stereoselective Formation and Methylation of the Pharmacologically Active Metabolite of Clopidogrel

et al. 2015

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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 and H4) are observed in plasma, with similar concentrations, and only H4 is active in humans. In this work, the mechanism 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 (CL int ) 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 CL int 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 2,3-dichloromethyl benzylamine, indicating the involvement of thiol S-methyltransferase. The CL int 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 H3 may 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.

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Section: Discussionsupporting

confidence: 93%

Human Liver Cytochrome P450 Enzymes and Microsomal Thiol Methyltransferase Are Involved in the Stereoselective Formation and Methylation of the Pharmacologically Active Metabolite of Clopidogrel

et al. 2015

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“…This was followed by a series of additional healthy volunteer studies that confirmed the impact of CYP2C19 genotype on the antiplatelet effect and provided evidence for loss of function variant carriers having significantly lower plasma concentrations of the clopidogrel active metabolite (Brandt et al, 2007;Kim et al, 2008;Umemura et al, 2008). Cardiovascular Pharmacogenetics and Personalized Medicine Numerous studies in patients with cardiovascular disease further documented that CYP2C19 loss of function carriers have reduced concentrations of active metabolite and less antiplatelet effect (Mega et al, 2009;Varenhorst et al, 2009;Collet et al, 2011;Hulot et al, 2011;Gong et al, 2012;Price et al, 2012). As a result, there is little to no discrepancy in the literature regarding the impact of the CYP2C19 loss of function alleles on clopidogrel active metabolite concentrations or on-treatment platelet reactivity.…”

Section: B Cyp2c19 Genetic Polymorphismsmentioning

confidence: 92%

Pharmacogenetics and Cardiovascular Disease—Implications for Personalized Medicine

Johnson¹,

Cavallari²

2013

Pharmacol Rev

115

104

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“…At last, ] i ) was determined as described previously. [27][28][29][30][31] Briefly, washed platelets were resuspended at concentrations of 10 8 /mL in modified Tyrode buffer (136.5 mM NaCl, 2.68 mM KCl, 11.9 mM NaHCO 3 , 0.42 mM NaH 2 PO 4 , 1 mM MgCl 2 , 5 mM N-(2-hydroxyethyl) piperazine-N′-2-ethanesulfonic acid (Hepes), 1.11 mM dextrose, 0.35% BSA, pH 7.4) with 10 U/mL heparin, 0.2 U/mL Apyrase and 1 µM prostaglandin I 2 (PGI 2 ), and then incubated for 8 min. After centrifugation, about 10 8 platelets were incubated in 300 µL of modified Tyrode buffer with 2.5 mM probenecid, 0.02% pluronic, 10 µM Fluo-3, 0.5 µM PGI 2 and 0.02 U/mL apyrase for 40 min at 37°C and washed, while the test drug was added in the last 10 min.…”

Section: Methodsmentioning

confidence: 99%

Delineation of Platelet Activation Pathway of Scutellarein Revealed Its Intracellular Target as Protein Kinase C

Tian

Chang

et al. 2016

Biological & Pharmaceutical Bulletin

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Erigeron breviscapus has been widely used in traditional Chinese medicine (TCM) and its total flavonoid component is commonly used to treat ischemic stroke, coronary heart disease, diabetes and hypertension. Scutellarin is the major ingredient of E. breviscapus and scutellarein is one of the main bioactive metabolites of scutellarin in vivo, but the latter's pharmacological activities have not been fully characterized. Provided evidence that could inhibit platelet aggregation, the effect of scutellarein on rat washed platelets and its underlying mechanisms were evaluated in our research. Scutellarein inhibited platelet adhesion and aggregation induced by multiple G protein coupled receptor agonists such as thrombin, U 46619 and ADP, in a concentration-dependent manner. Furthermore, the mild effect of scutellarein on intracellular Ca 2 mobilization and cyclic AMP (cAMP) level was observed. On the other hand, the role of scutellarein as potential protein kinase C (PKC) inhibitor was confirmed by PKC activity analysis and molecular docking. The phorbol myristate acetate-induced platelets aggregation assay with or without ADP implied that the scutellarein takes PKC(s) as its primary target(s), and acts on it in a reversible way. Finally, scutellarein as a promising agent exhibited a high inhibition effect on ADP-induced platelet aggregation among its analogues. This study clarifies the PKC-related signaling pathway involved in antiplatelet action of scutellarein, and may be beneficial for the treatment of cardiovascular diseases.

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Clarifying the importance of CYP2C19 and PON1 in the mechanism of clopidogrel bioactivation and in vivo antiplatelet response

Abstract: Our results demonstrate that PON1 does not mediate clopidogrel active metabolite formation or antiplatelet action, while CYP2C19 activity and genotype remains a predictor of clopidogrel pharmacokinetics and antiplatelet response.

Cited by 71 publications

References 33 publications

Human Liver Cytochrome P450 Enzymes and Microsomal Thiol Methyltransferase Are Involved in the Stereoselective Formation and Methylation of the Pharmacologically Active Metabolite of Clopidogrel

Human Liver Cytochrome P450 Enzymes and Microsomal Thiol Methyltransferase Are Involved in the Stereoselective Formation and Methylation of the Pharmacologically Active Metabolite of Clopidogrel

Pharmacogenetics and Cardiovascular Disease—Implications for Personalized Medicine

Delineation of Platelet Activation Pathway of Scutellarein Revealed Its Intracellular Target as Protein Kinase C

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