Registry Of Clopidogrel Metabolism Of Gene Polymorphisms And Antiplatelet Function In Acute Coronary Syndromes

Dual antiplatelet treatment with clopidogrel and aspirin is currently the standard choice for the prevention of thrombotic events in patients with acute coronary syndromes (ACS) or those undergoing percutaneous coronary intervention (PCI) with stent implantation. Although some patients took enough dose of aspirin and clopidogrel in clinical, they still had adverse cardiovascular events, such as sudden death, nonfatal myocardial infarction, stent thrombosis and stroke. Some studies have demonstrated wide inter-individual variability in the platelet inhibitory response to clopidogrel.Clopidogrel, an inactive prodrug, requires intestinal absorption by the P-glycoprotein multidrug resistance-1(MDR1) efflux transporter that is encoded by the ATP binding cassette, sub-family B, member1(ABCB1) gene. After absorption, clopidogrel requires several biotransformation steps, mainly by the hepatic cytochrome P450(CYP) system to generate its active thiol metabolite, which targets and irreversibly blocks the platelet adenosine diphosphate (ADP) P2Y12receptor. The highly polymorphic hepatic CYP system plays a key role in the conversion of clopidogrel into its active metabolite and may be a reasonable explanation for the increased risk of developing adverse clinical events. Several studies have demonstrated that carriers of the CYP2C19*2and*3loss-of-function alleles generate a smaller amount of the active metabolite of clopidogrel, resulting in an attenuated antiplatelet effect. There are considerable ethnic differences in the distribution and type of the CYP2C19loss-of-function (LOF) alleles. The carriage prevalence of the CYP2C19LOF variant is35-45%and25-35%among Caucasians and Africans, respectively, whereas it ranges from55to70%among Asians. Recently, Q192R within the gene encoding the paraoxonase-1(PON1) enzyme has recently been reported to be linked to clopidogrel bioactivation, and clinical outcomes of clopidogrel-treated patients.Platelet reactivity assessment with bedside monitoring assays was widely used in clinical, and VerifyNow P2Y12assay system with its fast and accurate detection of platelet function has become into the mainstream. However, VerifyNow P2Y12assay system’s data was relatively small in China. Pharmacodynamics studies have demonstrated wide inter-individual variability in the platelet inhibitory response to clopidogrel. And high residual platelet reactivity (HRPR) is well established as an independent predictor of increased cardiovascular events.Some studies had confirmed the consistent link between CYP2C19LOF allele carriage and adverse cardiovascular events among Caucasian patients, however, there are no substantial clinical data linking the CYP2C19LOF, ABCB1C3435T, and PON1Q192R alleles to an increased incidence of cardiovascular events in Chinese populations. Therefore, the aim of this study was to evaluate the relationship with the CYP2C19*2,*3,*17, ABCB1, and PON1variants on clopidogrel pharmacodynamics and short-term clinical outcomes in ACS patients of China.Above all, the present study was designed with following3subtopics.Part I Clopidogrel Metabolism Related Gene Polymorphisms in Acute Coronary Syndrome Patients and clinical characteristicsObjective:To detect the single nucleotide polymorphisms of CYP2C19*2,*3,*17, ABCB1C3435T and PON1Q192R related to clopidogrel metabolism in ACS patients by genotype analysis.Methods:Patients admitted to Fuwai Hospital emergence department (ED) from May2012to April2013with ACS were enrolled. We collected the general clinical data of patients, and record the basic information, patient related history and hospitalization treatment All patients had informed consents for blood samples and genetic analysis. The detection of polymorphisms was performed by TaqMan real-time PCR method. The alleles genotyped were CYP2C19*2,*3,*17, ABCB1C3435T and PON1Q192R. Minor allele frequency (MAF) was calculated. Patients were classified as one of the5groups by clopidogrel metabolizer phenotypes as Extensive [without any "loss-of-function"(LOF) allele*2,*3], Intermediate (with only one LOF allele), Poor (with two or more LOF alleles), Ultra [with one or two "gain-of-function"(GOF) allele*17GOF alleles] or Unknown (with one LOF allele and one GOF allele). The ABCB1C3435T and PON1Q192R genes are grouped according to genotype, divided into high expression (CC), the intermediate expression (CT) and low expression (TT).Results:A total of299Chinese ACS patients were enrolled with mean age of59.1years and233(74.6%) males. Current smoking patients were150(50.2%). Risk factors: hypertension180(60.2%), history of diabetes100(33.4%), hypercholesterolemia250(83.6%) and history of stroke28(9.4%).229(76.6%) patients had PCI and10(3.3%) had CABG operation during hospitalization. The main drug treatment during hospitalization:proportion of aspirin, beta blockers, clopidogrel, ACEI, ARB, CCB and nitric drugs were:99%,95.6%,85.6%,61.2%,11%,13.4%and83.9%. The diagnosis was ST-segment elevation myocardial infarction (STEMI) in208(69.6%), non-ST-segment elevation myocardial infarction (NSTEMI) in63(21.1%) and unstable angina (UA) in28(9.4%). The minor allele frequency (MAF) for each genotype of CYP2C19*2,*3,*17were31.9%,5.2%and0.8%, CYP2C19LOF allele’s included genotypes and accounted for the proportion of people:*1/*2(40.8%),*1/*3(6.7%),*2/*2(10.4%),*2/*3(1.7%) and*3/*3(0.7%); and CYP2C19GOF allele’s included only one patients:*1/*17(0.3); the Unknown phenotypes:*2/*17(0.7%) and*3/*17(0.7%). Patient clopidogrel metabolizer groups were defined as Extensive in38.1%, Intermediate in47.5%, Poor in12.8%, Ultra in0.3%and Unknown in1.4%, respectively. The MAF for ABCB1C3435T and PON1Q192R were38.3%and37%. The ABCB1C3435T high expression type was (CC)37.1%, intermediate expression type (CT) of49.2%and a low expression type (TT) of13.7%. PON1gene divided into high expression (CC), intermediate expression type (CT) and low expression type (TT) each accounted for38.8%,48.5%and12.7%of all the patients. Conclusion:Our study demonstrated a high distribution of the loss of function (LOF) allele of CYP2C19in China ACS patients, but we compared to other populations, the gene types of ABCB1and PON1were carrying similar proportion. The ABCB1and PON1genotype associated with poor metabolism of clopidogrel, which the proportion of our population carried lower. Part Ⅱ The effect of Clopidogrel Metabolism Related GenePolymorphisms on the pharmacodynamics and adverse clinical events of clopidogrel in ACS patientsObjective:The objective of this study was to evaluate the effect of the CYP2C19, ABCB1, and PON1Q192R variants on clopidogrel pharmacodynamics and clinical outcomes in acute coronary syndrome patients of China.Methods:This was a prospective observational study performed at a single-center from May2012to April2013. Two hundred and ninety-nine ACS patients presenting to the ED were enrolled and the patients had not taken clopidogrel within one month before enrollment. All patients received oral loading doses of300mg clopidogrel on the first day and75mg once daily for five consecutive days. The main outcome measure was the P2Y12reaction unit (PRU) which was measured using the VerifyNow P2Y12assay (Accumetrics, San Diego, CA, USA) at6hours after first loading dose and at five days after one hour oral clopidogrel. The secondary outcome measure was primary composite clinical endpoints which included death, nonfatal myocardial infarction, refractory angina, stent thrombosis and stroke. The follow-up period was30days.Results:We grouped according to whether they contain LOF and how many LOF genes they contain, CYP2C19alleles can be divided into strong and intermediary-poor metabolism, as well as a strong metabolism, intermediary metabolism and poor metabolism groups. Baseline demographics, clinical presentations, and treatments were mostly balanced between the CYP2C19LOF carrier groups. A lower response to clopidogrel (216.9PRU vs.192.2PRU, P<0.001) and composite endpoints increased with the CYP2C19loss-of-function alleles (6.7%vs.1.7%, P=0.043). However,there were not significant differences in clopidogrel pharmacodynamics and clinical outcomes across the ABCB1and PON1genotype groups; there was no obvious difference in minor bleeding events among the CYP2C19, ABCB1, and PON1genotype groups.Conclusion:The CYP2C19loss-of-function alleles had a gene effect on the pharmacodynamics and composite endpoints of clopidogrel in our study population. Neither the ABCB1nor the PON1genotype significantly influenced the antiplatelet effect and clinical outcomes of clopidogrel in these patients. Part Ⅲ Incidence and Outcome of Clopidogrel Treatment Platelet Reactivity in ACS Patients by VerifyNow P2Y12Objective:Among the ACS patients who receive clopidogrel therapy, the low response to clopidogrel has been one of the important factors affecting prognosis. The aim of this part was to assess the incidence, predictors, and clinical consequences by the results of VerifyNow P2Y12in a large population of ACS patients from ED.Methods:Overall,279ACS patients received point-of-care testing of platelet inhibition at five days after one hour oral clopidogrel. Poor responders of clopidogrel was diagnosed on the basis of P2Y12reaction units≥230, which divided into the poor responders and the normal responders.Results:There were100patients who were classified as poor responders (35.8%). In the poor responders groups, they are more females (39.2%vs.16.4%, P<0.001), older(63.8y vs.56.2y, P<0.001), less smoker (39.2%vs.55.9%, P<0.001), lower RBC (4.4×1012/L vs.4.7×1012/L, P<0.001), HGB (136.1g/L vs.147.8g/L, P<0.001) and HCT (39.1%vs.41.9%, P<0.001), but higher CREA (84.9μmol/L vs.79.9μmol/L, P<0.001). At30 days, the clinical endpoint events occurred no difference between the two groups. After adjustment for confounders, males (OR=0.37,95%CI0.16-0.97, P=0.041), CREA (OR=1.02,95%CI1.004-1.04, P=0.017), PPI drugs (OR=2.47,95%CI1.32-4.61, P=0.005) and CYP2C19LOF (OR=1.89,95%CI1.03-3.48, P=0.04) did emerge as independent predictor of poor responders.Conclusion:1/3of patients with acute coronary syndromes patients showed HRPR on bedside monitoring. They were more in CYP2C19LOF groups. CREA, PPI drugs and CYP2C19LOF were independently risk factors associated with poor responders to clopidogrel, while women were independent risk factors.