Recombinant human C1-inhibitor prevents non-specific proteolysis by mutant pro-urokinase during optimal fibrinolysis

• Published in: Thrombosis and haemostasis Vol. 102; no. 2; p. 279
• Main Authors: Gurewich, Victor, Pannell, Ralph
• Format: Journal Article
• Language: English
• Published: Germany 01.08.2009
• Subjects: alpha-2-Antiplasmin - pharmacology; Animals; Complement C1 Inactivator Proteins - pharmacology; Dogs; Fibrinogen - metabolism; Fibrinolysin - antagonists & inhibitors; Fibrinolysis - drug effects; Fibrinolysis - physiology; Fibrinolytic Agents - administration & dosage; Fibrinolytic Agents - adverse effects; Humans; In Vitro Techniques; Kinetics; Mutagenesis, Site-Directed; Plasminogen - metabolism; Plasminogen Activator Inhibitor 1 - pharmacology; Recombinant Proteins - administration & dosage; Recombinant Proteins - adverse effects; Recombinant Proteins - antagonists & inhibitors; Recombinant Proteins - genetics; Recombinant Proteins - pharmacology; Thrombolytic Therapy - adverse effects; Thrombolytic Therapy - methods; Tissue Plasminogen Activator - administration & dosage; Tissue Plasminogen Activator - adverse effects; Tissue Plasminogen Activator - antagonists & inhibitors; Urokinase-Type Plasminogen Activator - administration & dosage; Urokinase-Type Plasminogen Activator - adverse effects; Urokinase-Type Plasminogen Activator - antagonists & inhibitors; Urokinase-Type Plasminogen Activator - genetics
• ISSN: 0340-6245
• DOI: 10.1160/TH08-09-0598

A single-site mutant of prouPA (M5) spared haemostatic fibrin during thrombolysis in dogs. Zymograms of plasma from these dogs showed an unusual inhibitor complex with C1-inhibitor (C1I). Purified C1I added to human plasma enhanced the fibrin-specificity of M5. In the present study, the effect of recombinant human C1I (recC1I) on high-dose M5 and tPA were compared using fluorescein-labeled standardised clots in a plasma milieu. The shortest time to complete clot lysis (maximum rate) was first determined. This was approximately 65% per hour for both activators. By contrast, their top fibrin-specific lysis rate (<20% fibrinogen depletion) was less than half maximum (25-30% per hour). Adding recC1I (250-750 microg/ml) did not affect fibrinolysis, but prevented fibrinogenolysis and plasminogen depletion by M5, raising its fibrin-specific lysis rate to the maximum. With tPA, the recC1I modestly attenuated fibrinogenolysis, raising its fibrin-specific rate to about half the maximum. Consistent with this, the t(1/2) inhibition by C1I was approximately 90 min for tPA compared with ~10 min for tcM5. The t(1/2) of C1I for plasmin was ~2 min. Zymograms of plasma after clot lysis indicated that recC1I prevented non-specific tcM5 generation from M5, as evidenced by suppression of tcM5:C1I complexes. In conclusion, recC1I raised the fibrin-specificity of M5 in plasma so that a maximum lysis rate could be achieved without fibrinogenolysis. The inhibition by C1I of non-specific but not fibrin-dependent plasminogen activation could not be duplicated by other serpins. The findings provide a potential means to optimize both the efficacy and safety of thrombolysis.