Abstract 1102: Post Gwas Functional Genomics Analysis To Identify Non-coding Single Nucleotide Polymorphisms Controlling The Risk Of Spontaneous Coronary Artery Dissection

Abstract

Background: Spontaneous Coronary Artery Dissection (SCAD) is a non-atherosclerotic coronary artery disease and the leading cause of acute myocardial infarction in young females. SCAD results from the combination of genetic susceptibility and acquired triggering factors including inflammation and sex hormones. Three loci have been revealed by two large-scale genome-wide association studies (GWAS) in SCAD. However, GWAS only reveals the statistical associations between the genetic loci and diseases, and the biological mechanisms remain unknown. Methods: GWAS-reported tag single nucleotide polymorphisms (SNPs) and the linked haplotype SNPs were collected. Synthesized oligonucleotides representing the SNP alleles were subjected to electrophoretic mobility shift assays (EMSA) with nuclear proteins extracted from vascular endothelial cells (ECs), smooth muscle cells (SMCs) and fibroblasts. Hi-C analysis was performed to predict the potential target gene(s) based on chromatin interactions. The functionality of the SNP-associated target genes was examined in cultured vascular cells by RNA silencing. Results: The risk allele of tag SNPs in three genetic loci (chr1q21.2, 6p24.11 and 12q13.1) were associated with 1.5 to 1.8-fold higher risk of SCAD. Intronic, non-tag SNP rs7549396 (G/A) in chr1q21.2 locus was identified as the functional SNP in EMSA, given its consistent allele-imbalanced gel shifting pattern in ECs, SMCs and fibroblasts. Hi-C analysis in human aortic tissue identified a 490kb long-range chromatin interaction between SNP rs7549396 and the promoter of Annexin A9 (ANXA9), an extracellular matrix (ECM) and an inflammation-related protein potentially associated with the pathogenesis of SCAD. The knockdown of ANXA9 in vascular cells showed significant changes in inflammatory makers (IL-1β, vCAM-1 and CCL-2) and ECM degradation enzyme (MMP-1). Conclusions: Leveraging our post-GWAS functional genomics approach, we identified functional non-coding SNPs in disease-associated genetic locus and predicted a novel potential pathogenic gene ANXA9 for SCAD. Further studies on the functions of this SNP and its associated target gene ANXA9 will provide genetics and mechanistic explanation for this mysterious disease.