SBIR-STTR Award

The development of a platelet-rich thrombus on damaged endothelium or atherosclerotic plaques can severely impair the blood flow to vital organs, including the brain, heart, lungs, and kidneys. The contribution of platelets to the pathogenesis of potentially fatal ischemic and/or thromboembolic events, including stroke, myocardial infarct, and pulmonary embolism, is well documented. Therefore, the discovery of effective modulators of platelet function that can prevent thrombus formation is the focus of intensified efforts in translational hematology and cardiovascular biology research.The rationally designed small molecule chemical compound a-cyano-Beta-hydroxy-Beta-methyl-N-(2,5-dibromophenyl)-propenamide (LFM-A13) is a specific inhibitor of the TEC family protein tyrosine kinases, Bruton's tyrosine kinase (BTK) and TEC. Both BTK and TEC play an important role in platelet physiology by regulating the glycoprotein GPVI-FcRy-coupled collagen receptor signaling pathway. We have recently found that LFM-A13 inhibits (a) collagen-induced BTK/TEC stimulation, (b) BTK/TEC-dependent downstream signaling events, (c) biochemical and ultrastructural changes indicative of platelet activation, and (d) collagen-induced platelet aggregation. LFM-A13 was not toxic to mice when administered systemically at dose levels ranging from 1 mg/kg to 100 mg/kg. At nontoxic dose levels, LFM-A13 prolonged the tail bleeding times of mice and improved event-free survival in two mouse models of fatal pulmonary thromboembolism.An oral capsule formulation of LFM-A13 (LFM-A13-F) was developed and showed excellent bioavailability both in mice and dogs. We are now proposing to test the activity of this clinically applicable oral formulation in a mouse model of collagen-induced fatal thromboembolism. After establishing the single agent activity of LFM-A 13-F, we will also examine its antithrombotic effects in combination with the standard antiplatelet agents, dipyndamole or aspirin (alone or in combination). Also examined will be the effect of LFM-A13 alone or in combination with other drugs on the bleeding/clotting times in mice

We have rationally designed the leflunomide metabolite analog alpha-cyano-beta-hydroxy-beta-methyl-N-(2,5-dibromophenyl)-propenamide (LFM-A13) as a specific inhibitor of the protein tyrosine kinase, Bruton's tyrosine kinase (BTK). LFM-A13 is a chemosensitizing anti-leukemic agent with anti-thrombotic properties. Our preclinical studies established that LFM-A13 has a favorable safety and pharmacokinetics profile. LFM-A13 has been formulated for oral administration (LFM-A13-F). Under SPECIFIC AIM 1, we will study the toxicity profile of LFM-A13-F in rats and dogs and 2) synthesize and formulate large quantities of LFM-A13 under GMP conditions to be used for human clinical trials. Based on the toxicity studies done in mice, we hypothesize that LFM-A13-F will not cause acute, subacute, or chronic toxicity at the proposed dose levels. Under SPECIFIC AIM 2, large scale synthesis and evaluation of LFM-A13-F as hard gelatin capsules will be carried under Good Manufacturing Practice (GMP) to comply with FDA regulations and to ensure preparation of clinical grade LFM-A13-F for human clinical trials. We will use synthetic procedures developed during the phase I studies. In order to achieve specific aim 2 we have already identified FDA approved facilities for the synthesis and formulation of LFM-A13. The results obtained in specific aim #1 and specific aim #2 will provide the critical information for further testing of this agent in human subjects under Phase III. This research will facilitate the bench-to-bedside development of LFM-A13 as the protype of a new class of anti-leukemic agents with anti-thrombotic properties.

Thesaurus Terms:
anticoagulant, antineoplastic, chemosensitizing agent, drug design /synthesis /production, leukemia, neoplasm /cancer chemotherapy antithrombin, collagen, oral administration, pharmacokinetics, platelet activation dog, laboratory rat