CRISPLD1 rs12115090 polymorphisms alters antiplatelet potency of clopidogrel in coronary artery disease patients in Chinese Han

Highlights

• CYP2C19*2 is associated with HTPR and the PRI is significantly increased in the *2 A allele carriers.

• The PRI is significantly decreased in the patients with CRISPLD1 rs12115090 C allele.

• CRISPLD1 rs12115090 is associated with decreased risk of HTRP in Han Chinese CAD patients.

Abstract

Background

Dual antiplatelet therapy (DAPT) with aspirin and clopidogrel is a recommended treatment for coronary artery disease (CAD) patients undergoing percutaneous coronary intervention (PCI) to reduce the rate of ischemic events and stent thrombosis. However, high on-treatment platelet reactivity (HTPR) during clopidogrel therapy for some patients may lead to outcome failure and occurrence of cardiovascular events. Amounts of studies have proved that genetic factors may contribute to HTPR. In our study, we explored the predictive value of 10 single nucleotide polymorphisms (SNPs) in 8 genes indicated by exome sequencing with clopidogrel efficacy.

Methods

Two hundred and forty-one Han Chinese CAD patients (mean age: 61 ± 10 years) receiving dual antiplatelet therapy were recruited, among which 118 patients administered with 300 mg loading dose (LD) clopidogrel for 12–24 h and 123 subjects administered with 75 mg/day maintain dose (MD) clopidogrel for at least 5 days before discharge. The platelet reaction index (PRI) was determined to reflect clopidogrel response in the patients. Venous blood samples were drawn from all participants to extract genomic DNA. MassARRAY, Sanger sequencing and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were used to determine the genotypes of 10 SNPs.

Results

Allelic tests showed significant differences in genotype distribution between HTPR and normal on-treatment platelet reactivity (NTPR) patients for 3 SNPs including CYP2C19 rs4244285 (CYP2C19*2) (co-dominant model: p = 0.003, dominant model: p = 0.004, recessive model: p = 0.012), CRISPLD1 rs12115090 (co-dominant model: p = 0.011, dominant model: p = 0.004), and LTA4H rs11108379 (dominant model: p = 0.041). After adjusting for covariates including clinical characteristics of patients, concomitant medications and complications, we confirmed that carriers of the CYP2C19*2 showed significantly increased risk of HTPR (*2/*2 vs *1/*1: OR = 12.266, 95% CI: 1.336–112.592, p = 0.027; *1/*2 + *2/*2 vs *1/*1: OR = 2.202, 95% CI: 1.083–4.480, p = 0.029). Contrarily, carriers of the CRISPLD1 rs12115090 C allele showed significantly reduced risk of HTPR (CC vs AA: OR = 0.242, 95% CI: 0.078–0.752, p = 0.014; CA + CC vs AA: OR = 0.457, 95% CI: 0.232–0.904, p = 0.024) in Chinese CAD patients. In addition, carriers of the CYP2C19*2 allele showed significantly increased PRI (*1/*2 vs *1/*1: p = 0.008, 2/*2 vs 1/*1: p < 0.001, *2/*2 vs 1/*2: p = 0.011), while patients carrying the rs12115090 C allele showed significantly decreased PRI than the wild-type AA homozygotes (CA vs AA: p = 0.046, CA + CC vs AA: p = 0.023).

Conclusion

CYP2C19*2 reduced the antiplatelet potency of clopidogrel and increased the risk of HTPR, while CRISPLD1 rs12115090 A>C polymorphism increased the antiplatelet potency of clopidogrel. Genetic tests, especially for CYP2C19*2 are recommended in Han Chinese CAD patients before using of clopidogrel.

Introduction

Cardiovascular diseases including acute coronary syndrome (ACS) lead to the first death worldwide. Approximately 1.2 million patients are diagnosed and hospitalized due to cardiovascular events every year in USA and the patients with coronary artery disease (CAD) account for more than half (Roger et al., 2012). Platelet activation and aggregation play crucial roles in the occurrence of cardiovascular events after percutaneous coronary interventions (PCI) (Lange and Hillis, 2004). Clopidogrel is a second-generation thienopyridine antiplatelet drug, which irreversibly inhibits P2Y12 receptor to exert antiplatelet effect. Dual antiplatelet therapy (DAPT), especially aspirin and clopidogrel became the recommended treatment of CAD for its reduction of >80% cardiovascular events and improvement in the long-term clinical outcomes (Angiolillo et al., 2007; O'Gara et al., 2013; Levine et al., 2016; Resor et al., 2016; Yu et al., 2017). However, nearly 5%–44% patients still suffered from therapeutic failure manifested by ischemic events due to clopidogrel resistance or high on-treatment platelet reactivity (HTPR) of clopidogrel (Gurbel and Tantry, 2007; Stone et al., 2013; Amin et al., 2017).

Clopidogrel, which is an inactive pro-drug, needs to be absorbed from the intestine and converts into an active metabolite (H4) in the liver by a two-step metabolism. H4 competes with ADP for P2Y12 receptors and inhibits activation of downstream signaling pathway, and further exerts antiplatelet activity. A recent study has shown that about 50% of clopidogrel is absorbed from the intestine (Simon et al., 2009) and only 10%–15% of the absorbed clopidogrel is metabolized into an active thiol metabolite by several cytochrome P450 (CYP450) micro-enzymes such as CYP2C19, CYP3A4, CYP2C9, and CYP1A2 (Bonello et al., 2010; Kazui et al., 2010; Zhu et al., 2013; Zhang et al., 2015; Zhang et al., 2017). The CYP450 genetic polymorphisms may be one of the most important factors for the amount of clopidogrel metabolism and bioactivation. CYP2C19 plays a substantial role in the bioactivation of clopidogrel. The loss-of-function (LOF) variants in CYP2C19, including CYP2C19*2 (rs4244285) and CYP2C19*3 (rs4986893), were widely reported to decrease the activity of CYP2C19 and inhibit clopidogrel bioactivation. The CYP2C19 LOF alleles are observed to decrease the antiplatelet effect of clopidogrel and increase the risk of major adverse clinical events (MACE) during clopidogrel therapy (Trenk et al., 2008; Shuldiner et al., 2009; Jang et al., 2012). On the contrary, CYP2C19*17 (rs12248560) leading to CYP2C19 gain of function (GOF) by increasing the gene transcription is associated with increased risk of bleeding during clopidogrel therapy (Sibbing et al., 2010). However, the CYP2C19 polymorphisms can only explain 5%–12% of individual differences in the efficacy of clopidogrel (Shuldiner et al., 2009; Karazniewicz-Lada et al., 2012). Other genetic polymorphisms, ages, smoking, co-medications and combined disease also account for its variability (Simon et al., 2009; Frelinger et al., 2013), although the underlying mechanisms related to these intrinsic and extrinsic factors are not yet fully understood.

With the development of high throughput genotyping techniques including genome-wide association study (GWAS) and exome sequencing, more genetic polymorphisms are observed to affect clopidogrel response. By using exome sequencing with 4 pairs of matched CYP2C19*1/*1 wild-type patients (European descent) with extreme phenotype for clopidogrel response indicated by P2Y12 reaction units (PRU) < 50 or PRU > 235, Scott et al. (2016) observed 18 SNPs associated with clopidogrel efficacy. However, influence of these SNPs on clopidogrel response in Chinese population is still unknown. In this study, we selected 10 SNPs in 8 genes (CYP2C19, B4GALT2, CRISPLD1, STAB2, LTA4H, PSMD13, MED23, CDK18) according to the findings from Scott's report (Scott et al., 2016) and based on allele frequencies of the SNPs in Chinese population from the 1000 Genomes database as well, and investigate the association between these SNPs with clopidogrel response in Han Chinese CAD patients.