Inhibition of von Willebrand factor‐mediated platelet activation and thrombosis by the anti‐von Willebrand factor A1‐domain aptamer ARC1779

Abstract

Summary. Background: von Willebrand factor (VWF) has a role in both hemostasis and thrombosis. Platelets adhere to damaged arteries by interactions between the VWF A1-domain and glycoprotein Ib receptors under conditions of high shear. This initial platelet binding event stimulates platelet activation, recruitment, and activation of the clotting cascade, promoting thrombus formation. Objective: To characterize the inhibitory activity of a VWF inhibitory aptamer. Methods: Using in vitro selection, aptamer stabilization, and conjugation to a 20-kDa poly(ethylene glycol), we generated a nuclease-resistant aptamer, ARC1779, that binds to the VWF A1-domain with high affinity (K D 2 nM). The aptamer was assessed for inhibition of VWF-induced platelet aggregation. In vitro inhibition of platelet adhesion was assessed on collagen-coated slides and injured pig aortic segments. In vivo activity was assessed in a cynomolgus monkey carotid electrical injury thrombosis model. Results and Conclusion: ARC1779 inhibited botrocetin-induced platelet aggregation (IC 90 300 nM) and shear force-induced platelet aggregation (IC 95 400 nM). It reduced adhesion of platelets to collagen-coated matrices and formation of platelet thrombi on denuded porcine arteries. ARC1779 also inhibited the formation of occlusive thrombi in cynomolgus monkeys. We have discovered a novel anti-VWF aptamer that could have therapeutic use as an anti-VWF agent in the setting of VWFmediated thrombosis.

A novel interaction between FlnA and Syk regulates platelet ITAM-mediated receptor signaling and function

“…Stable platelet adhesion and aggregate formation under arterial shear condition, events which normally require activation of the IIb3 integrin downstream of GPVI ligation by collagen and that of GPIb by VWF, were severely impaired in the absence of FlnA (Ruggeri and Mendolicchio, 2007;Diener et al, 2009). Syk participates in the signaling cascade induced by GPIb ligation, particularly in the calcium rise and activation of PI-3 kinase which follow ligation, and is intimately involved in downstream signaling to IIb3 Yanabu et al, 1997;Falati et al, 1999;Ozaki et al, 2005).…”

“…Binding to a collagencoated surface was measured in whole blood after labeling of platelets and normalization of platelet counts. Perfusion was performed under arterial shear condition (shear rate 1,500/s), which mediates binding of plasma VWF to surface-bound collagen (Ruggeri and Mendolicchio, 2007;Diener et al, 2009). Fluorescent FlnA-null platelets covered 70% less of the collagen-coated surface than did WT platelets (Fig.…”

Active platelet‐binding conformation of plasma von Willebrand factor in young women with acute myocardial infarction

“…It remains to be established by means of prospective and/or intervention studies whether this abnormality of VWF is causally related to coronary thrombosis or is only an epiphenomenon of acute disease. Intervention may become possible, for instance, by using an aptamer, which by blocking the interaction between VWF and its platelet receptor glycoprotein Iba, inhibited thrombus formation in animal models [15] and was safe in healthy volunteers following short-term administration in a phase 1 trial [16]. Phase 2 trials are being planned in humans in clinical conditions in which VWF-dependent platelet aggregation is thought to play a role in causing thrombosis, such as TTP and carotid endarterectomy.…”

Aptamers Are New Pharmacological Substances for the Development of Anticoagulants

“…К 3'-концу олигонуклеотида для его защиты при-креплен инвертированный дезокситимидин, а к 5'-концу -20-кДа ПЭГ, как для защиты, так и для увеличения молекулярной массы аптамера (олигонуклеотид имеет мол. массу ~1 3 кДА) и замедления его выведения из кровотока [6][7][8].…”

“…При этом ARC1779 не влиял на агрегацию тромбоцитов, индуциро-ванную такими агонистами, как АДФ и арахи-доновая кислота. В модели тромбоза на обезь-янах ARC1779 предотвращал образование окк-люзивных тромбов после повреждения каро-тидных артерий [6][7][8].…”