Introduction
Aspirin use is recommended by the United States Preventive Services Task Force to prevent cardiovascular disease (CVD) in selected, high risk patients.[1] It is well known that aspirin inhibits platelet cyclooxygenase 1 (COX1)[2]; however, aspirin has additional effects on platelets, which are not fully explained by its effects on COX-1. For example, 81mg/day aspirin is sufficient to fully suppress platelet COX1[3] yet higher doses cause additional reductions in platelet function as assessed by non-COX1 dependent platelet function assays such as adenosine diphosphate (ADP) and collagen induced platelet aggregation.[4] In addition, despite consistent platelet COX-1 inhibition, non-COX1 dependent measures of platelet function wane over time[5] suggesting some adaptation of platelets to chronic aspirin exposure. Residual platelet function measured using non-COX1 dependent platelet function assays are risk markers for future cardiovascular events during aspirin therapy.[6, 7] Last after decades of uncertainty around the appropriate aspirin dose for cardiovascular risk reduction[8] only recently has there been randomized trial data comparing the effectiveness and safety of low- vs. high-dose aspirin. This trial did not identify major differences in efficacy or safety between aspirin doses; however, nearly 50% of patients crossed over to the alternative dose with 3 weeks of randomization. [9] Therefore, gaps in our knowledge around one of the most prescribed medicines worldwide remain.
Although anucleate, it is well known that platelets are a rich source of messenger RNA (mRNA). Platelets are invested with mRNA from megakaryocytes during thrombopoiesis in a regulated manner[10], can uptake transcripts from other vascular cells[11], and are also a major source of transcripts found in plasma extracellular vesicles [12]. Genetic and gene expression studies have confirmed that variable platelet mRNA levels lead to variable levels of platelet function.[13-15] We have previously shown that a blood-based, “aspirin response signature” of transcripts primarily of platelet origin correlate with platelet function on aspirin and predict future cardiovascular events in patients taking aspirin.[16] These data support the concept that the biologic basis for variable platelet function lies, at least in part, in variable platelet gene expression. We[17] and others[18] have shown that aspirin (or its stable metabolite salicylate) can alter expression of selected genes in megakaryocytes as well as in circulating platelets in patients exposed to aspirin. However, these prior studies were limited in scope as they did not systematically characterize aspirin’s effects on platelet gene expression.
Based on these knowledge gaps, we pursued the hypothesis that variability in the platelet function response to aspirin is associated with aspirin induced changes in the platelet transcriptome. We tested this hypothesis using an open label, randomized crossover study of low- vs. high-dose aspirin in human volunteers with longitudinal COX-1 direct and indirect assays of platelet function, drug concentrations, and RNA profiling at each time point.