Endothelial surface and circulating glycoprotein von Willebrand factor (vWF) regulates platelet adhesion and is associated with thrombotic diseases, including ischemic stroke, myocardial infarction, and peripheral vascular disease. Thrombosis, as manifested in these diseases, is the leading cause of disability and death in the western world. Current parenteral antithrombotic and thrombolytic agents used to treat these conditions are limited by a short therapeutic window, irreversibility, and major risk of hemorrhage. To overcome these limitations, we developed a novel anti-vWF aptamer, called DTRI-031, that selectively binds and inhibits vWF-mediated platelet adhesion and arterial thrombosis while enabling rapid reversal of this antiplatelet activity by an antidote oligonucleotide (AO). Aptamer DTRI-031 exerts dose-dependent inhibition of platelet aggregation and thrombosis in whole blood and mice, respectively. Moreover, DTRI-031 can achieve potent vascular recanalization of platelet-rich thrombotic occlusions in murine and canine carotid arteries. Finally, DTRI-031 activity is rapidly (<5 min) and completely reversed by AO administration in a murine saphenous vein hemorrhage model, and murine toxicology studies indicate the aptamer is well tolerated. These findings suggest that targeting vWF with an antidote-controllable aptamer potentially represents an effective and safer treatment for thrombosis patients having plateletrich arterial occlusions in the brain, heart, or periphery.

BackgroundDrug-eluting stents (DES) represent the main-stream management of restenosis following treatments of occlusive cardiovascular diseases. However, DES cannot eliminate instent restenosis yet exacerbate thrombogenic risks. To achieve dual inhibition of restenotic smooth muscle cell (SMC) de-differentiation/proliferation and thrombogenic endothelial cell (EC) dysfunction, a common target in both cell types, has been long-sought after. We evaluated the potential of protein kinase RNA-like endoplasmic reticulum kinase (PERK) as such a target for low-thrombogenic anti-restenotic intervention.Methods and ResultsWe used a rat angioplasty model of restenosis and a FeCl3-induced mouse model of thrombosis. Loss-or gain-of-function was achieved by PERK inhibition (GSK2606414, siRNA) or overexpression (adenovirus). Restenosis was robustly mitigated by GSK2606414 administered either via injected (i.v.) lesion-homing platelet membrane-coated nanoclusters or a perivascular hydrogel; it was enhanced by PERK transgene. Whereas PERK inhibition blocked, its overexpression exacerbated PDGF-induced human aortic SMC de-differentiation (reduced smooth muscle α-actin or αSMA) and proliferation (BrdU incorporation). Further, PERK activity promoted STAT3 activation but inhibited SRF transcriptional (luciferase) activity; its protein co-immunoprecipitated with STAT3 and also MRTF-A, the SRF activator for αSMA transcription. Importantly, PERK inhibition also prevented TNFα-induced impairment of human EC growth and upregulation of thrombogenic tissue factor, both in vitro and ex vivo. In vivo, oral gavage of GSK2606414 preserved ~50% of the normal blood flow 60 min after FeCl3-induced vascular injury.ConclusionsPERK inhibition is dual beneficial in mitigating restenosis and thrombosis, thus implicating a potential design for anti-restenotic intervention to overcome the thrombogenicity of DES.

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