title: Rapid testing of clopidogrel resistance by genotyping of CYP2C19 and CYP2C9 polymorphisms using denaturing on-chip capillary electrophoresis.
abstract: Antiplatelet therapy is a cornerstone of cardiovascular treatment in patients with coronary artery disease and after myocardial infarction. Clopidogrel has become a popular antiplatelet agent due to its fast action and low frequency of adverse effects. Kinetics of clopidogrel metabolism is driven by enzymatic activity of the Cytochrome P450 system. Genotyping of CYP2C19 and CYP2C9 polymorphisms allows to identify slow metabolizers showing resistance to clopidogrel therapy. Today, a number of PCR-based techniques for single nucleotide polymorphism genotyping directed at clopidogrel resistance polymorphisms are in use. Here, we describe a new alternative genotyping approach combining the separation power of denaturing capillary electrophoresis with the analysis speed and ease of use of Bioanalyzer chipCE platform. Using an upgraded heater control, we present an optimization for allele separation of CYP2C19 I331V, CYP2C9 R144C, and CYP2C9 I359L polymorphisms employing run temperatures of up to 55°C. We demonstrate rapid and accessible approach to reproducible clopidogrel resistance with feasibility and low cost.
authors.affiliation: Center for Applied Genomics of Solid Tumors, Genomac Research Institute, Prague, Czech Republic. mminarik@email.com
authors.forename: Marek
authors.initials: M
authors.lastname: Minarik
authors.forename: Marta
authors.initials: M
authors.lastname: Kopeckova
authors.forename: Marcus
authors.initials: M
authors.lastname: Gassman
authors.forename: Pavel
authors.initials: P
authors.lastname: Osmancik
authors.forename: Lucie
authors.initials: L
authors.lastname: Benesova
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics* / Q000378:metabolism; D000077144:Clopidogrel; D065731:Cytochrome P-450 CYP2C19; D065729:Cytochrome P-450 CYP2C9; D004351:Drug Resistance; D046209:Electrophoresis, Microchip / Q000379:methods*; D006801:Humans; D010597:Pharmacogenetics; D010975:Platelet Aggregation Inhibitors / Q000493:pharmacokinetics / Q000494:pharmacology; D020641:Polymorphism, Single Nucleotide; D015203:Reproducibility of Results; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics / Q000494:pharmacology
chemical_list: D010975:Platelet Aggregation Inhibitors; D000077144:Clopidogrel; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; D065731:Cytochrome P-450 CYP2C19; D013988:Ticlopidine