PROJECT SUMMARY/ABSTRACT Occlusive arterial thrombosis leading to stroke and myocardial infarction contribute to ~13 million deaths around the world every year. Over the past two decades, recombinant tissue plasminogen activator (rTPA) has remained the only drug approved to treat acute ischemic stroke. Unfortunately, patients who receive rTPA have a significant increase in symptomatic hemorrhage with a conversion rate of 6.4%. Moreover, rTPA only achieves an average of ~30% recanalization in arterial thrombi, which are commonly platelet rich and are notoriously resistant to rTPA. Evidence for this was seen in a recent study in patients who presented with large vessel occlusion (LVO) stroke and received rTPA, demonstrated vessel recanalization only 10% of the time. The limitations of rTPA therapy result in only ~5% of patients actually receive the drug. Therefore, a critical need exists to develop anti-thrombotic agents that: 1) prevent occlusive thrombus formation, 2) recanalize acute arterial occlusions and 3) prevent restenosis, all with a superior safety profile than that of rTPA. Von Willebrand Factor (VWF) is an optimal target for anti-thrombotic therapy. Under high shear seen in thrombosis, VWF binds to glycoprotein Ib (GPIb) of the platelet receptor complex GPIb-IX-V (GPIb-IX-V) as well as to GPIIb-IIIa, resulting in platelet activation and aggregation. VWF also self-associates, extending into the vessel lumen where it serves not only as a scaffold for platelets but also red blood cells. These central processes ultimately result in arterial thrombosis as seen in ischemic stroke. Aptamers are an innovative class of drug molecules that consist of oligonucleotides that specifically and efficiently bind to and inhibit target proteins. Basking Biosciences Inc has developed an RNA aptamer that inhibits VWF, named DTRI-031. It has also designed a reversal oligonucleotide, named DTRI-025 that rapidly neutralizes DTRI-031 within minutes. DTRI-031 prevents thrombus formation, lyses fully formed arterial occlusions and reduces the radiographic burden of ischemia better than rTPA in small and large animal models of arterial thrombosis and stroke. It is currently in the final series of IND enabling studies. DTRI-025 rapidly reverses DTRI-031 activity in vitro and in vivo in small animal models. The overall goal of this proposal is to define the optimum dose of DTRI-025 to neutralize DTRI-031 in a large animal model and to develop a bioassay to perform GLP safety pharmacology and toxicity studies with GMP manufactured DTRI-025. The Aims of this proposal will allow Basking Biosciences to submit a Pre-IND application to FDA and ultimately lead to First In Human (FIH) trials in conjunction with DTRI-031.