Novel Discovery and Functional Characterization of Key Genetic Variants that Influence the Pharmacogenomics of Anti-Platelet Therapy

Abstract

Coronary artery disease (CAD) is among the leading causes of death in the United States and worldwide. The most commonly prescribed treatment for the secondary prevention of cardiovascular events in at-risk patients is concurrent administration of clopidogrel and aspirin (DAPT). However, significant inter-individual variability in response has been documented, resulting in sub-optimal therapy in some patients and an increased risk of recurrent events. While genetic evaluations of ex vivo platelet reactivity have identified notable clopidogrel response modifiers such as CYP2C19*2, heritability estimates suggest most of the genetic determinants of variable DAPT response remain unidentified. We conducted the first genome-wide analysis of clopidogrel active metabolite concentration, a direct measure of clopidogrel pharmacokinetics, and discovered novel loci on chromosomes 3p25 and 17q11that significantly influence clopidogrel metabolism, as well as several other loci with suggestive evidence of association with clopidogrel response. While extension to on-clopidogrel platelet aggregation and ischemic events in an independent population was inconclusive, this research highlights the utility of clopidogrel active metabolite concentration as a novel endophenotype for clopidogrel efficacy. In addition to investigating novel antiplatelet response modifiers, we conducted clinical and functional characterization of rs12041331, a well-studied intronic variant in Platelet Endothelial Aggregation Receptor 1 (PEAR1) previously correlated with platelet aggregation and cardiovascular event risk. We used a fixed sequence, crossover design to prospectively evaluate platelet reactivity pre- and post- 3 different therapeutic aspirin doses in 67 healthy subjects recruited by rs12041331 genotype. rs12041331 was associated with platelet aggregation before and after aspirin exposure in a pathway- and agonist- specific manner, although higher aspirin dose did not impact platelet reactivity in a genotype-dependent manner. PEAR1 is also suggested to be a modifier of endothelial function. We therefore functionally characterized the impact of PEAR1 knockdown and rs12041331 on human umbilical vein endothelial cell (HUVEC) behavior in vitro and found PEAR1 to have a significant role in endothelial migration, extracellular matrix adhesion, and proliferation. While further follow-up is necessary, these experiments demonstrate the importance of incorporating multiple measures to evaluate genetic determinants of antiplatelet efficacy, and the results herein have promise to contribute to improved cardiovascular care in the future.