title: A chimeric platelet-targeted urokinase prodrug  selectively blocks new thrombus formation.
abstract: The use of fibrinolytic agents to prevent new thrombus formation is limited by an increased risk of bleeding due to lysis of hemostatic clots that prevent hemorrhage in damaged blood vessels. We sought to develop an agent that provides thromboprophylaxis without carrying a significant risk of causing systemic fibrinolysis or disrupting hemostatic clots. We previously showed that platelet (PLT) α granule-delivered urokinase plasminogen activator (uPA) is highly effective in preventing thrombosis, while being associated with little systemic fibrinolysis or bleeding. Here, we generated a chimeric prodrug composed of a single-chain version of the variable region of an anti-αIIbβ3 mAb fused to a thrombin-activatable, low-molecular-weight pro-uPA (PLT/uPA-T). PLT/uPA-T recognizes human αIIbβ3 on both quiescent and activated platelets and is enzymatically activated specifically by thrombin. We found that this prodrug binds tightly to human platelets even after gel filtration, has a prolonged half-life in mice transgenic for human αIIb compared with that of uPA-T, and prevents clot formation in a microfluidic system. Importantly, in two murine injury models, PLT/uPA-T did not lyse preexisting clots, even when administration was delayed by as little as 10 minutes, while it concurrently prevented the development of nascent thrombi. Thus, PLT/uPA-T represents the prototype of a platelet-targeted thromboprophylactic agent that selectively targets nascent over preexisting thrombi.
authors.forename: Rudy E
authors.initials: RE
authors.lastname: Fuentes
authors.forename: Sergei
authors.initials: S
authors.lastname: Zaitsev
authors.forename: Hyun Sook
authors.initials: HS
authors.lastname: Ahn
authors.forename: Vincent
authors.initials: V
authors.lastname: Hayes
authors.forename: M  Anna
authors.initials: MA
authors.lastname: Kowalska
authors.forename: Michele P
authors.initials: MP
authors.lastname: Lambert
authors.forename: Yuhuan
authors.initials: Y
authors.lastname: Wang
authors.forename: Donald L
authors.initials: DL
authors.lastname: Siegel
authors.forename: Daniel W
authors.initials: DW
authors.lastname: Bougie
authors.forename: Richard H
authors.initials: RH
authors.lastname: Aster
authors.forename: Daniel D
authors.initials: DD
authors.lastname: Myers
authors.forename: Victoria
authors.initials: V
authors.lastname: Stepanova
authors.forename: Douglas B
authors.initials: DB
authors.lastname: Cines
authors.forename: Vladimir R
authors.initials: VR
authors.lastname: Muzykantov
authors.forename: Mortimer
authors.initials: M
authors.lastname: Poncz
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D016503:Drug Delivery Systems / Q000379:methods*; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D018345:Mice, Knockout; D016513:Mice, SCID; D011355:Prodrugs / Q000493:pharmacokinetics / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000493:pharmacokinetics / Q000494:pharmacology; D013927:Thrombosis / Q000097:blood / Q000188:drug therapy*; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000493:pharmacokinetics / Q000494:pharmacology*
chemical_list: D011355:Prodrugs; D011993:Recombinant Fusion Proteins; D014568:Urokinase-Type Plasminogen Activator