Pharmacogenetic implications in clopidogrel therapy : a pharmacist-Ied management approach

Abstract

Pharmacists are in a position to take a leading role in the clinical implementation of pharmacogenetics as regards genotyping and interpretation of results as well as proposing recommendations to personalise therapy. The cytochrome P (CYP) 450 2C19 enzyme is the principal enzyme involved in clopidogre1 metabolism and is encoded by a highly polymorphic gene. Presence of the CYP2C 19 loss-of-function (LoF) *2 variant allele is associated with reduced CYP2C19-mediated activity which impairs clopidogrel bioactivation and increases the risk of adverse cardiac events after percutaneous coronary intervention (PCI). The principal aim of this research was to apply pharmacist-led CYP2C19 genotyping to support consultant cardiologists in the individualisation of antip1ate1et therapy prescribed in patients undergoing PCI. The study protocol was approved by the University Research Ethics Committee. A patient data collection form was developed and psychometrically evaluated. Two hundred and fifty-two patients undergoing PCI with stent deployment for stable angina or acute coronary syndrome (ACS) were recruited by non-probability sampling from the Cardiac Catheterisation Suite at Mater Dei Hospital (MDH). Patients included were ≥ 18 years and prescribed dual antiplatelet therapy with aspirin and clopidogrel. Exclusion criteria were patients with coagulation disorders, platelet disorders, chronic liver disease and/or history of stroke/transient ischaemic attack. Infonned written consent was obtained from each patient at the time of recruitment. 5mL of peripheral blood was collected into a purple-top ethylenediaminetetraacetic acid (EDT A) tube and genomic DNA (gDNA) extraction was performed with the QIAamp@ DNA Mini QIAcube kit (Qiagen). CYP2Cl9 genotyping for the *2 (rs4244285) and *17 (rsI2248560) variant alleles was undertaken using the laboratory-based TaqMan@ SNP

Drug Metabolism Assays (Life Technologies) on the 7500 real-time PCR system (Applied Biosystems). Genotypes identified were categorised into four metaboliser phenotypes according to the 'Clinical Pharmacogenetics Implementation Consortium guidelines for CYP2C19 genotype and clopidogre1 therapy' namely; 'extensive' metabo1isers (EMs; *1/*1), 'ultra-rapid' metabo1isers (UMs; *11*17, *17/*17), 'intermediate' metabolisers (IMs; * 1/*2, *2/* 17) or 'poor' metabolisers (PMs; *2/*2). The patients were followed-up 6 to 12 months post-PCI for stent thrombosis (ST) and in-stent restenosis (ISR). Thirty-four out of the 252 patients were genotyped for the LoF *2 allele with two alternative genotyping assays namely, the Spartan™ RX system (Spartan Bioscience), a rapid point-of-care (POC) assay using gDNA obtained from a buccal swab, and the laboratory-based GenID® reverse-dot blot (RDB) hybridisation assay (Autoimmun Diagnostika GmbH) using gDNA obtained from whole blood. Comparison between the three assays was carried out. Prevalence of the *2 and * 17 variant alleles in Maltese patients was compared to 12 populations bordering the Mediterranean Sea from Southern Spain to Tunisia. Eighty-two out of the total 252 patients had a history of previous PCI with stent deployment and were divided into patients who presented with angiography-confirmed ISR at time of recruitment and those who did not. Retrospective analysis was undertaken to investigate any association between various predictors including CYP2C19 *2 allele carrier status and ISR and ST. The financial impact of CYP2C19*2 genotyping to personalise antip1ate1et therapy to limit ISR and ST was estimated for the three assays using the retrospective data. Baseline characteristics of the 252 patients were: Gender (75% male), age (mean 65 years, range 26-89 years), ethnicity (99% Caucasian), weight (mean 81 kg, range 47- 134 kg), positive family history of IHD (65%), active smoking (32%), hypertension (70%), dyslipidaemia (68%) and diabetes mellitus (46%). The majority of patients (55%) were undergoing PCI for stable angina and 45% were undergoing PCI after being admitted with ACS. Polyphannacy was common and most patients (64%) were prescribed between 6 and 10 chronic medications. Distribution of CYP2C19 genotypes was divided into EMs (n=129, 51 %), UMs (n=58, 23%) and IMs (n=65, 26%). No patients in the study population were PMs. Total allele frequency of *2 and *17 was 13% and 15% respectively. Percentage agreement in genotype results between the TaqMan® and GenID® RDB assays was 100% while percentage agreement in genotype results between the Spartan ™ RX assay and both the TaqMan® and GenID® RDB assays was 97%. This difference resulted since 1 patient who was genotyped as a carrier of two *2 alleles with the Spartan ™ RX assay was genotyped as a carrier of one *2 allele with the other two assays. When the *2 allele frequency in Maltese patients was compared to the 12 populations bordering the Mediterranean Sea, prevalence in the Maltese population was in accordance (p>0.05) with all these populations. Retrospective analysis of the 82 patients with prior PCI identified 29 (35%) patients who presented with ISR at the time of recruitment and were undergoing repeat PCI. In 13 of these 29 patients the stent was deployed ≤ 1 year before recruitment into the study and the patients were still on maintenance c1opidogrel therapy at the time of repeat PCI. Univariate analysis showed a statistically significant association (p<0.05) between coronary ISR and CYP2C 19 *2 allele carrier status. Ten (12%) of the 82 patients

presented with ST at time of recruitment and were undergoing repeat PCI. In 3 of these 10 patients the stent was deployed ≤ 1 year before recruitment and the patients were still on maintenance clopidogre1 therapy at the time of repeat PCI. Univariate analysis showed a statistically significant association (p<O.05) between *2 allele carrier status and ST ≤ 1 year. The financial analysis indicated cost savings using all three CYP2C19*2 genotyping assays. The findings have important clinical implications for clopidogre1 use in Malta. According to evidence-based guidelines for CYP2C19 genotype and clopidogre1 therapy, an alternative P2Y 12-receptor inhibitor such as prasugre1 is recommended in carriers of the CYP2C 19 LoF *2 allele (26% in this study). The statistically significant association observed between CYP2C19 *2 allele carrier status and ST and ISR despite clopidogre1 therapy, indicates that carriers of the *2 alelle may benefit from treatment with prasugre1 to reduce risk of post-PCI complications. Compared to the 1aboratorybased assays, the POC Spartan™ RX assay provides very rapid results, is accurate and reliable, non-invasive and operator-friendly. Implementation of pharmacist-led antip1ate1et therapy individualisation guided by CYP2C19*2 genotyping in patients undergoing PCI has the potential to limit the incidence of ST and ISR, improve patients' quality of life and reduce healthcare costs . Keywords: clopidogre1, percutaneous coronary intervention, CYP2C 19 10ss-of-function *2 allele, stent thrombosis, coronary in-stent restenosis, laboratory-based CYP2C19 genotyping, rapid point-of-care CYP2C19 genotyping, pharmacist-1ed approach, individualised antiplatelet therapy, cost-effectiveness