Open AccessA specific plasminogen activator inhibitor‐1 antagonist derived from inactivated urokinase
Author(s) -
Gong Lihu,
Proulle Valerie,
Fang Chao,
Hong Zebin,
Lin Zhonghui,
Liu Min,
Xue Guangpu,
Yuan Cai,
Lin Lin,
Furie Barbara,
Flaumenhaft Robert,
Andreasen Peter,
Furie Bruce,
Huang Mingdong
Publication year - 2016
Publication title -
journal of cellular and molecular medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.44
H-Index - 130
eISSN - 1582-4934
pISSN - 1582-1838
DOI - 10.1111/jcmm.12875
Subject(s) - fibrinolysis , urokinase , plasminogen activator inhibitor 1 , plasminogen activator , urokinase receptor , serine protease , chemistry , plasmin , microbiology and biotechnology , thrombus , pharmacology , tissue plasminogen activator , biology , protease , biochemistry , medicine , endocrinology , enzyme
Fibrinolysis is a process responsible for the dissolution of formed thrombi to re‐establish blood flow after thrombus formation. Plasminogen activator inhibitor‐1 ( PAI ‐1) inhibits urokinase‐type and tissue‐type plasminogen activator ( uPA and tPA ) and is the major negative regulator of fibrinolysis. Inhibition of PAI ‐1 activity prevents thrombosis and accelerates fibrinolysis. However, a specific antagonist of PAI ‐1 is currently unavailable for therapeutic use. We screened a panel of uPA variants with mutations at and near the active site to maximize their binding to PAI ‐1 and identified a potent PAI ‐1 antagonist, PAI trap. PAI trap is the serine protease domain of urokinase containing active‐site mutation (S195A) and four additional mutations (G37bR–R217L–C122A–N145Q). PAI trap inhibits human recombinant PAI ‐1 with high potency ( K d = 0.15 nM ) and high specificity. In vitro using human plasma, PAI trap showed significant thrombolytic activity by inhibiting endogenous PAI ‐1. In addition, PAI trap inhibits both human and murine PAI ‐1, allowing the evaluation in murine models. In vivo, using a laser‐induced thrombosis mouse model in which thrombus formation and fibrinolysis are monitored by intravital microscopy, PAI trap reduced fibrin generation and inhibited platelet accumulation following vascular injury. Therefore, this work demonstrates the feasibility to generate PAI ‐1 inhibitors using inactivated urokinase.