title: The paraoxonase-1 pathway is not a major bioactivation pathway of clopidogrel in vitro.
abstract: OBJECTIVE
  Clopidogrel is a prodrug bioactivated by cytochrome P450s (CYPs). More recently, paraoxonase-1 (PON1) has been proposed as a major contributor to clopidogrel metabolism. The purpose of this study was to assess the relative contribution of CYPs and PON1 to clopidogrel metabolism in vitro.
  
  
  METHODS
  Clopidogrel metabolism was studied in human serum, recombinant PON1 enzyme (rePON1), pooled human liver microsomes (HLMs), HLMs with the CYP2C19*1/*1 genotype and HLMs with the CYP2C19*2/*2 genotype. Inhibition studies were also performed using specific CYP inhibitors and antibodies. Clopidogrel and its metabolites were measured using LC/MS/MS method.
  
  
  RESULTS
  PON1 activity was highest in the human serum and there was no difference in PON1 activity between any of the HLM groups. The production of clopidogrel's active metabolite (clopidogrel-AM) from 2-oxo-clopidogrel in pooled HLMs was approximately 500 times that in serum. When 2-oxo-clopidogrel was incubated with rePON1, clopidogrel-AM was not detected. Clopidogrel-AM production from 2-oxo-clopidogrel was lower in CYP2C19*2/*2 HLMs compared with CYP2C19*1/*1 HLMs, while PON1 activity in HLMs with both genotypes was similar. Moreover, incubation with inhibitors of CYP3A, CYP2B6 and CYP2C19 significantly reduced clopidogrel bioactivation while a PON1 inhibitor, EDTA, had only a weak inhibitory effect.
  
  
  CONCLUSIONS
  This in vitro study shows that the contribution of PON1 to clopidogrel metabolism is limited at clinically relevant concentrations. Moreover, CYP2C19, CYP2B6 and CYP3A play important roles in the bioactivation of clopidogrel.
authors.affiliation: Clinical Pharmacology and Toxicology Service, Geneva University Hospitals and Faculty of Medicine, Geneva, Switzerland.
authors.forename: V
authors.initials: V
authors.lastname: Ancrenaz
authors.forename: J
authors.initials: J
authors.lastname: Desmeules
authors.forename: R
authors.initials: R
authors.lastname: James
authors.forename: P
authors.initials: P
authors.lastname: Fontana
authors.forename: J-L
authors.initials: JL
authors.lastname: Reny
authors.forename: P
authors.initials: P
authors.lastname: Dayer
authors.forename: Y
authors.initials: Y
authors.lastname: Daali
mesh_terms: D000911:Antibodies, Monoclonal; D043303:Aryldialkylphosphatase / Q000235:genetics / Q000378:metabolism*; D002853:Chromatography, Liquid; D000077144:Clopidogrel; D003577:Cytochrome P-450 Enzyme System; D015971:Gene Expression Regulation, Enzymologic; D005838:Genotype; D006801:Humans; D008861:Microsomes; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D015394:Molecular Structure; D053719:Tandem Mass Spectrometry; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D003577:Cytochrome P-450 Enzyme System; D000077144:Clopidogrel; D043303:Aryldialkylphosphatase; C468406:PON1 protein, human; D013988:Ticlopidine