Accumulation of fusion protein in pulmonary vasculature facilitates local fibrinolysis. (A) Dose-response curve of pulmonary thrombolysis. Dissolution of ¹²⁵I-labeled microemboli lodged in mouse pulmonary vasculature by bolus injection of 300, 100, and 30 μg fusion protein, the equivalent amounts of lmw-scuPA (150, 50, and 15 μg) versus a mixture of lmw-scuPA and parental antibody, respectively. Thrombolytic potency was expressed as the percentage of lysis versus the dose administered. Interrupted line indicates spontaneous lysis. Error bars indicate SEM. (B) Simplified model of a proposed strategy for thromboprophylaxis using vascular immunotargeting of genetically engineered anti-PECAM scFv-uPA fusion protein. Anti-PECAM scFv-uPA circulates in a form of a prodrug, single-chain uPA, binds to PECAM-1, and remains anchored on the luminal surface of endothelium for at least several hours. In situ thrombosis or embolism induces initial local conversion of plasminogen (Pg) into plasmin (Pn) by endogenous plasminogen activators (Endo-PA). Plasmin (and perhaps other enzymes) formed in the vicinity of the clot converts the endothelium-bound scFv-scuPA into enzymatically active tcuPA (Figure 3A), which in turn amplifies local formation of plasmin, reinforcing local thrombolysis, preventing clot extension and reocclusion.