Association between cytochrome P450 2C19 polymorphism and clinical outcomes in clopidogrel-treated Uygur population with acute coronary syndrome: a retrospective study

Acute coronary syndromes (ACS) are triggered by fissuring or rupture of an atheromatous plaque in the coronary arterial wall. This stimulates a thrombotic response causing variable obstruction to flow in the coronary arterial lumen with downstream ischaemic myocardial injury [1]. ACS is a life-threatening disease that is becoming the leading cause of morbidity and mortality in developing countries. Clopidogrel is the most widely prescribed purinergic receptor (P2Y12) inhibitor. It is an antiplatelet drug that, when administered with aspirin, has been shown to reduce the risk for cardiovascular events after ACS or a percutaneous coronary intervention (PCI) [2]. Clopidogrel is a prodrug requiring cytochrome P450 (CYP) for biotransformation into its active thiol metabolite. Initial clopidogrel pharmacogenetic studies examined genetic variations in CYP enzymes and associated these genetic variants with active metabolite levels. The most important of these is cytochrome P450, family 2, subfamily C, polypeptide 19 (CYP2C19), which metabolize clopidogrel to its active form [3].

The CYP2C19 gene maps to the long arm of chromosome 10 (10q24) and it encodes a 490-amino-acid protein predominantly expressed in the liver and, to a lesser extent, in the small intestine. CYP2C19*2 results from a guanine (G) to adenine (A) transition at position 681 in exon5 (rs4244285), producing an aberrant splice site and it represents the most frequent CYP2C19 defect [4], with the most common allele frequencies of 29–35% in Asians. Another nonfunctional allele CYP2C19*3, in which a guanine (G) to adenine (A) transition at position 636 in exon4 (rs4986893), results in a premature stop codon, and therefore nonfunctional protein [5, 6]. CYP2C19*3 is also relatively common in Asian populations (up to 10% are carriers) [7]. Patients with loss-of-function (LOF) allele variants (CYP2C19*2 and CYP2C19*3) are at risk for thromboembolic events [8]. The CYP2C19*17 allele was previously reported to be associated with high CYP2C19 activity. CYP2C19*17 is a -806 C > T single nucleotide polymorphism (with a cytosine (C) to thymine (T) transition) that causes specific nuclear protein binding to the 5'-flanking region. This binding results in increased gene transcription and high enzyme activities [9]. The CYP2C19 gain-of-function allele (*17) is associated with increased catalytic activity [10]. In addition, polymorphisms of CYP2C19 are known to vary considerably according to ethnicity [11, 12]. Approximately 25 genetic variants in the exonic region of the CYP2C19 have been identified [4]. Other CYP2C19 gene variants that lead to loss of function are relatively rare, typically below 1% [7]. Therefore, in our study, we determined the effect of genetic polymorphisms in CYP2C19*2, *3, and *17 on variation in response to clinical outcomes in Uygur ACS patients.

In 2012, the morbidity of coronary artery disease in the Uygur population of Xinjiang was 24.2%, much higher than the national average (7.2%) [13]. Polymorphisms of the CYP2C19 gene and clinical factors are strong predictors of cardiovascular outcomes for patients with acute coronary syndrome treated with oral clopidogrel [14, 15]. Nevertheless, very little is known about the influence of such factors in Uygur populations. Therefore, we determined the long-term impact of CYP2C19 polymorphisms on the risk for major adverse cardiac events (MACE) and bleeding in Uygur patients with ACS treated with clopidogrel.

The Uygur population has a high prevalence of CHD. A major contributor is a high-fat diet and lifestyle associated with this community. Patients enrolled in this study showed high prevalences of hypertension, dyslipidemia, diabetes mellitus, and smoking. In Asian populations, obesity is defined as BMI > 26 kg/m² [16]. The average BMI of the Uygur patients in this study was 28.3 ± 3.8 kg/m². According to many recent estimates, advanced age is an independent risk factor of CHD, primarily attributed to compromised plasticity of vessels [17]. The mean age of our patients was 58.2 ± 9.0 years (range: 31–78 years), and 25.4% of these patients were > 65 years. Crimi et al. reported that cigarette smoking reduced platelet reactivity independently of clopidogrel treatment in patients with ACS [18]. Smoking is an important risk factor primarily because cigarette chemicals cause coronary inflammation [19], and 40.2% of our patients with a smoking history developed ACS. Furthermore, most of our subjects had at least one cardiovascular risk factor. Demographic characteristics of the study patients were according to the cardiovascular risk factors [20].

Clopidogrel is a therapy for ACS and emergent or elective PCI. Clopidogrel has no biological activity; it is metabolized into 15% active metabolite and 85% inactive metabolite by the P450 system. The polymorphic isoenzyme CYP2C19 plays an essential role in genetic diversity [21]. There were 101 patients (28.8%) who suffered MACE during the 1-year follow-up in the current study. By comparison, we found that the MACE group had a significantly higher CYP2C19*2 AA genotype frequency and higher GA genotype frequency than the no-MACE group (P = 0.001). These findings suggest that CYP2C19 is an independent predictor of the risk of MACE in Uygur patients, following several studies of Chinese patients undergoing PCI [15, 22]. It is well-established that patients carrying CYP2C19 (LOF) alleles have a reduced capacity for clopidogrel bioactivation, impaired platelet inhibition, and a significantly higher risk of MACE when treated with clopidogrel compared with patients without a LOF allele [23]. This finding was demonstrated in the present study; the MACE group had a significantly higher CYP2C19*2 A allele frequency than the no-MACE group (P < 0.001).

Based on their abilities to metabolize CYP2C19 substrates, individuals may be classified as EMs, IMs, PMs, or UMs [24‐26]. CYP2C19 IMs and PMs have significantly lower plasma concentrations of the active metabolite and diminished inhibition of platelet aggregation than EMs [27]. The prevalence of IMs and PMs is significantly higher in Asians, with about 57% of LOF allele carriers [28]. In the current study, about 35% of Uygur ACS patients carried an LOF allele (110 CYP2C19*2 carriers and 23 CYP2C19*3 carriers). Both IMs and PMs were significantly higher in the no-MACE group than in the MACE group (P < 0.05). This finding accords with a meta-analysis of 36,076 participants that found that IMs and PMs in Asian populations undergoing PCI and prescribed clopidogrel had a higher risk of MACE [29].

According to the characteristics and the distribution of CYP2C19 polymorphisms, we analyzed risk factors for MACE and bleeding using multivariate logistic regression analysis. In multiple logistic regression analysis, only CYP2C19*2 carriers were significantly associated with MACE (OR: 2.51, 95% CI: 1.534–4.09). Shuldiner et al. reported that, in a healthy Amish population, the CYP2C19*2 allele was the primary genetic locus associated with diminished platelet inhibitory response to clopidogrel, and that the CYP2C19*2 allele accounted for 12% of the inter-patient variability in platelet response to clopidogrel [27]. Besides, we compared clinical backgrounds between MACE and no-MACE group of CYP2C19*2 carriers. The clinical backgrounds include sex, age, BMI, alcohol intake, smoking, blood pressure, blood lipids level, transaminase, creatinine, uric acid, blood glucose, clinical presentation, final treatment, comorbidities, and concomitant medication. There were no significant differences in clinical backgrounds between MACE and no-MACE groups of CYP2C19*2 carriers (P > 0.05). This suggests that logistic regression analysis did not have the influence of other clinical backgrounds.

Another study showed that CYP2C19 UMs treated with clopidogrel exhibited increased active metabolite formation, inhibition of platelet aggregation, higher bleeding risk, and lower MACE risk [30]. CYP2C19*17 was linked to a superior response to clopidogrel but an increased risk of bleeding [31]. In the present study, bleeding occurred in 18 patients (5.1%) during the 1-year follow-up; this rate is consistent with that of Korean ACS patients after taking clopidogrel (5.3%) [32].

Although the UMs and CYP2C19*17 carriers did not contribute to the risk of bleeding events in our analysis, we believe that the results are less persuasive because of the limited sample size (only 18 patients bled). On the other hand, the frequency of UMs in the no-MACE group was significantly higher than the MACE group in Uygur patients (P = 0.004). This finding suggests that the frequency of the CYP2C19*17T allele in the bleeding group (25.0%) was higher than that in the non-bleeding group (14.7%), although not significantly (P > 0.05). Our previous report found that allele frequencies of the minor alleles of CYP2C19 *2, *3, and *17 differed significantly between Uygur and Han groups [12]. To our knowledge, this is the first study reporting the impact of the cytochrome P450 2C19 polymorphism of MACE and bleeding to clopidogrel in Uygur patients with ACS.

Our study has the following limitations. First, only 351 Uygur ACS patients were assessed; thus, our findings regarding the association between the CYP2C19 polymorphism and clinical outcomes of clopidogrel treatment should be validated in studies with larger samples. Second, because of the limitation of detection technology, we could not establish the value of CYP2C19 genotyping combined with on-treatment platelet reactivity (platelet function test). Third, we obtained some MACE and bleeding information via telephone and therefore could not precisely document symptoms. Finally, gender is a nonmodifiable risk factor of ACS [33]; the varying prevalence of ACS results might result from the unbalanced gender distribution.