Towards Effective and Safe Thrombolysis and Thromboprophylaxis: Preclinical Testing of a Novel Antibody-Targeted Recombinant Plasminogen Activator Directed Against Activated Platelets
Rationale: Fibrinolysis is a valuable alternative for the treatment of myocardial infarction when percutaneous coronary intervention is not available in a timely fashion. For acute ischemic stroke, fibrinolysis is the only treatment option with a very narrow therapeutic window. Clinically approved thrombolytics have significant drawbacks, including bleeding complications. Thus their use is highly restricted leaving many patients untreated.
Objective: We developed a novel targeted fibrinolytic drug that is directed against activated platelets.
Methods and Results: We fused single-chain urokinase plasminogen activator (scuPA) to a small recombinant antibody (scFvSCE5), which targets the activated form of the platelet-integrin GPIIb/IIIa. Antibody binding and scuPA activity of this recombinant fusion protein were on par with the parent molecules. Prophylactic in vivo administration of scFvSCE5-scuPA (75U/g BW) prevented carotid artery occlusion after ferric chloride injury in a plasminogen-dependent process compared to saline (p<0.001) and blood flow recovery was similar to high dose non-targeted urokinase (500U/g BW). Tail bleeding time was significantly prolonged with this high dose of non-targeted urokinase, but not with equally effective targeted scFvSCE5-scuPA at 75U/g BW. Real-time in vivo molecular ultrasound imaging demonstrates significant therapeutic reduction of thrombus size after administration of 75U/g BW scFvSCE5-scuPA as compared to same dose of a mutated, non-targeting scFv-scuPA or vehicle. The ability of scFvSCE5-scuPA to lyse thrombi was lost in plasminogen-deficient mice, but could be restored by intravenous injection of plasminogen.
Conclusions: Targeting of scuPA to activated GPIIb/IIIa allows effective thrombolysis and the potential novel use as a fibrinolytic agent for thromboprophylaxis without bleeding complications.
- Received August 30, 2013.
- Revision received January 22, 2014.
- Accepted January 28, 2014.