Conjunctive thrombolysis under the action of tissue-type plasminogen activator and conjugate urokinase-fibrinogen after double sequential bolus administration in the model of vein thrombosis in dogs

Abstract

Summary Objective : The aim of the study was to define thrombolytic efficacy of the combination of tissue plasminogen activator (t-PA) and urokinase-fibrinogen (uPA-Fbg) covalent conjugate using a simplified scheme of administration in vivo. Design : Plasminogen (Pmg) activation by t-PA and uPA-Fbg separately and combined was evaluated in vitro, both in the presence of fibrin-monomer (FM) and without it. We have conducted a comparative study of thrombolysis efficacy on the model of venous thrombosis using t-PA preparations and uPA-Fbg conjunction applying various schemes of administration. Materials : Recombinant t-PA and covalent conjugate uPA-Fbg (obtained earlier) which was exhibiting pronounced prolonged thrombolytic action were used in the study. Interventions : Thrombolytic efficacy of preparations was assayed on the model of 125 I-labelled venous thrombosis in dogs. Bolus, bolus-infusion and sequential double bolus administration of thrombolytics were used to compare their effects. Main outcome measures : Pmg activation by t-PA and uPA-Fbg preparations (in the presence of FM and without it) was monitored by hydrolysis of S-2390 chromogenic substrate in vitro. Thrombolytic efficacy of the preparations was evaluated by radioactive level and content of hemostatic proteins (fibrinogen (Fbg), Pmg and alpha-2-antiplasmin) in blood samples taken from dogs. Radioactivity increase in blood showed thrombolysis dynamics, while the change in hemostatic protein content showed disturbed action of thrombolytics on the fibrinolysis system. Results : The in vitro model system showed additivity of t-PA and uPA-Fbg action, both in the presence and without FM. Sequential double bolus administration of t-PA and (then after 15 min) uPA-Fbg conjugate into dogs with venous thrombosis revealed a fast and stable thrombolytic action of the conjuction mentioned (2.5 mg t-PA and 250 000 IU of uPA-Fbg), as well as its noticeable, but moderate (not exceeding 40% decrease from the initial level) influence on the indexes of systemic fibrinolysis. Thrombolytic action of t-PA in conjunction with uPA-Fbg is considerable and surpasses effects of the same doses of its components administered separately. It was significantly similar to the conjunction action after the bolus-infusion-bolus scheme of administration and the action after bolus-infusion administration of 10 mg t-PA (at later stages of monitoring). Data demonstrated eligibility of t-PA and uPA-Fbg thrombolytic composition for effective research application with a simplified scheme of administration. Conclusion : Conjunctive application of t-PA and uPA-Fbg conjugate exhibiting complementary action and various pharmacokinetic profiles is promising for efficient and safe thrombolysis with a simplified scheme of the preparation administration. Thrombolytic compositions (mixtures) on the basis of these, or similar short- and long-term acting plasminogen activators, are likely to become one of the main and widespread means of treatment of thromboembolism disease in the 21st century.