Small Interfering RNA Therapy Targeting the Long Noncoding RNA SMILR for Therapeutic Intervention in Coronary Artery Bypass Graft Failure.

Abstract

Coronary artery bypass graft (CABG) surgery remains the gold standard of care to prevent myocardial ischemia in patients with advanced atherosclerosis; however, poor long-term graft patency remains a considerable and long-standing problem. Excessive vascular smooth muscle cell (SMC) proliferation in the grafted tissue is recognized as central to late CABG failure. We previously identified SMILR, a human-specific SMC-enriched long noncoding RNA that drives SMC proliferation, suggesting that targeting SMILR expression could be a novel way to prevent neointima formation, and thus CABG failure. Here, we sought to identify a lead siRNA for clinical development. We describe the design and synthesis of a library of 76 chemically enhanced SMILR-targeting siRNA. From this library, we identify a lead siRNA, BHF7, which demonstrates potent and reproducible silencing of SMILR expression, and which robustly blocks vascular smooth muscle cell proliferation, both in vitro and in the ex vivo human saphenous vein model. We further demonstrate using RNA-sequencing that BHF7 down-regulates the expression of genes associated with proliferation and does not induce the expression of interferon or apoptosis genes, suggesting it has a favorable safety profile, both on- and off-target. Finally, we performed TUNEL staining on BHF7-treated tissues and measured the levels of cleaved caspase-3 by enzyme-linked immunosorbent assay after BHF7 treatment. This demonstrated that BHF7 does not induce a cytotoxic response either in vitro or ex vivo. Collectively, these data represent a preclinical package into the function and specificity of BHF7 which warrants further investigation into the possibility of utilizing BHF7 as a novel, ex vivo RNA therapeutic for the prevention of CABG failure in humans.