Prophylactic thrombolysis by thrombin-activated latent prourokinase targeted to PECAM-1 in the pulmonary vasculature

Blood. 2008 Feb 15;111(4):1999-2006. doi: 10.1182/blood-2007-07-103002. Epub 2007 Nov 28.

Abstract

A recombinant prodrug, single-chain urokinase-type plasminogen activator (scuPA) fused to an anti-PECAM-1 antibody single-chain variable fragment (anti-PECAM scFv/scuPA) targets endothelium and augments thrombolysis in the pulmonary vasculature.(1) To avoid premature activation and inactivation and to limit systemic toxicity, we replaced the native plasmin activation site in scFv/low-molecular-weight (lmw)-scuPA with a thrombin activation site, generating anti-PECAM scFv/uPA-T that (1) is latent and activated by thrombin instead of plasmin; (2) binds to PECAM-1; (3) does not consume plasma fibrinogen; (4) accumulates in mouse lungs after intravenous injection; and (5) resists PA inhibitor PAI-1 until activated by thrombin. In mouse models of pulmonary thrombosis caused by thromboplastin and ischemia-reperfusion (I/R), scFv/uPA-T provided more potent thromboprophylaxis and greater lung protection than plasmin-sensitive scFv/uPA. Endothelium-targeted thromboprophylaxis triggered by a prothrombotic enzyme illustrates a novel approach to time- and site-specific regulation of proteolytic reactions that can be modulated for therapeutic benefit.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Endothelium, Vascular / drug effects
  • Endothelium, Vascular / physiology*
  • Fibrinogen / metabolism
  • Humans
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Platelet Endothelial Cell Adhesion Molecule-1 / metabolism
  • Platelet Endothelial Cell Adhesion Molecule-1 / physiology*
  • Protein Binding
  • Pulmonary Circulation / physiology*
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Proteins / metabolism
  • Recombinant Proteins / pharmacology
  • Thrombin / physiology*
  • Thrombolytic Therapy*
  • Tissue Distribution
  • Urokinase-Type Plasminogen Activator / genetics
  • Urokinase-Type Plasminogen Activator / metabolism
  • Urokinase-Type Plasminogen Activator / physiology*

Substances

  • Platelet Endothelial Cell Adhesion Molecule-1
  • Recombinant Fusion Proteins
  • Recombinant Proteins
  • Fibrinogen
  • Thrombin
  • Urokinase-Type Plasminogen Activator
  • saruplase