Effects of PON1 Gene Promoter DNA Methylation and Genetic Variations on the Clinical Outcomes of Dual Antiplatelet Therapy for Patients Undergoing Percutaneous Coronary Intervention.

ABSTRACT

INTRODUCTION AND OBJECTIVE: The relationship between either paraoxonase 1 (PON1) gene promoter DNA methylation or genetic variations and bleeding or major adverse cardiac events after dual antiplatelet therapy has been incompletely characterized. We aimed to systematically investigate the role of genetic variations and DNA methylation of the PON1 CpG island promoter on the clinical outcomes of dual antiplatelet therapy for patients with coronary artery disease (CAD) who underwent percutaneous coronary intervention (PCI). METHODS: This study included 653 patients with CAD undergoing PCI and receiving dual antiplatelet therapy. Genomic DNAs were isolated from whole blood and were genotyped for the three single nucleotide polymorphisms (SNPs) of the PON1 gene. The DNA methylation levels in the PON1 promoter region were determined by bisulfite sequencing or pyrosequencing at five CpG sites (positions -142, -161, -163, -170, and -184 from the transcription start site). Clopidogrel and its metabolites in plasma were examined using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), and platelet function analysis was performed using the VerifyNow assay. RESULTS: Statistically significant associations between methylation levels at five PON1 CpG sites and bleeding were observed -184 [odds ratio (OR) 0.98, 95% confidence interval (CI) 0.96-1.00, p = 0.028]; -170 (OR 0.99, 95% CI 0.97-1.00, p = 0.048); -163 (OR 0.98, 95% CI 0.96-1.00, p = 0.029); -161 (OR 0.98, 95% CI 0.97-1.00, p = 0.026); and -142 (OR 0.98, 95% CI 0.97-1.00, p = 0.042) at a false discovery rate of <5%. Statistical analysis also revealed that aspirin reaction units (ARUs) were significantly associated with PON1 methylation level at CpG site -163 (p = 0.0342). The ARUs of patients with the PON1 126 CC genotype was 527 ± 94, which was higher than the ARUs (473 ± 89) of patients with the 126 CG genotype (p = 0.0163). Multivariate logistic regression analysis indicated that the PON1 methylation level at CpG site -161 (OR 0.95, 95% CI 0.92-0.98, p = 0.002) and the use of angiotensin-converting enzyme inhibitors (OR 0.48, 95% CI 0.26-0.89, p = 0.021) were associated with a decreased risk of bleeding events. CONCLUSIONS: Hypomethylation of CpGs in the PON1 promoter may be a weak, albeit statistically significant, risk factor of bleeding after dual antiplatelet therapy. Further large-scale studies are needed to verify our results.