Novel Thrombolytic Drug Based on Thrombin Cleavable Microplasminogen Coupled to a Single‐Chain Antibody Specific for Activated GPIIb/IIIa

BACKGROUND: Thrombolytic therapy for acute thrombosis is limited by life‐threatening side effects such as major bleeding and neurotoxicity. New treatment options with enhanced fibrinolytic potential are therefore required. Here, we report the development of a new thrombolytic molecule that exploits key features of thrombosis. We designed a recombinant microplasminogen modified to be activated by the prothrombotic serine‐protease thrombin (HtPlg), fused to an activation‐specific anti–glycoprotein IIb/IIIa single‐chain antibody (SCE5), thereby hijacking the coagulation system to initiate thrombolysis. METHODS AND RESULTS: The resulting fusion protein named SCE5‐HtPlg shows in vitro targeting towards the highly abundant activated form of the fibrinogen receptor glycoprotein IIb/IIIa expressed on activated human platelets. Following thrombin formation, SCE5‐HtPlg is activated to contain active microplasmin. We evaluate the effectiveness of our targeted thrombolytic construct in two models of thromboembolic disease. Administration of SCE5‐HtPlg (4 μg/g body weight) resulted in effective thrombolysis 20 minutes after injection in a ferric chloride–induced model of mesenteric thrombosis (48±3% versus 92±5% for saline control, P<0.01) and also reduced emboli formation in a model of pulmonary embolism (P<0.01 versus saline). Furthermore, at these effective therapeutic doses, the SCE5‐HtPlg did not prolong bleeding time compared with saline (P=0.99). CONCLUSIONS: Our novel fusion molecule is a potent and effective treatment for thrombosis that enables in vivo thrombolysis without bleeding time prolongation. The activation of this construct by thrombin generated within the clot itself rather than by a plasminogen activator, which needs to be delivered systemically, provides a novel targeted approach to improve thrombolysis.