A combination of recombinant single-chain urokinase-type plasminogen activator and streptokinase has a synergistic non-fibrin specific action on in vitro clot lysis

Abstract

Thrombolytic effects of streptokinase (SK), a recombinant single-chain urokinase-type plasminogen activator (rscu-PA), and their combinations were measured in an in vitro system composed of 125I-fibrin labelled human plasma clot immersed in a citrated human plasma at 37°C. Clot lysis was monitored by release of 125I-fibrin degradation products and an activation of the fibrinolytic system in the surrounding plasma was measured by fibrinogen, plasminogen and α2-antiplasmin levels. SK and rscu-PA induced dose- and time-dependent clot lysis: 50% lysis in 4h was obtained with 75 nM SK and 38nM rscu-PA, respectively. At these concentrations, no significant activation of the fibrinolytic system in plasma was observed with rscu-PA, whereas SK produced fibrinogen breakdown. As was judged by a mathematical analysis of dose-response curves by the isobole method, simultaneous and sequential equimolar combinations of SK and rscu-PA caused a significant non-fibrin specific synergistic thrombolytic effect. The synergy of thrombolytic action of these combinations persisted for 24h and was more pronounced at the subsequent addition of SK followed by rscu-PA (in 1h). 4h after the addition of 25 nM of SK or rscu-PA, they had induced 6% and 25% lysis respectively, while simultaneous or sequential addition of both plasminogen activators at the same concentrations induced 55% and 80% clot lysis. The maximal achievable 5-fold increase in the thrombolytic action of sequential combinations was observed in the molar ratio of plasminogen activators 1:1, as compared with the total effect of the agents at the same concentrations as used in simultaneous combinations. The synergistic thrombolytic effect of the sequential equimolar combinations was more fibrin-specific than that of the simultaneous combinations. It is concluded that the simultaneous and sequential equimolar combinations of SK and rscu-PA possessed synergistic thrombolytic action on clot lysis in vitro. The sequential combination of equimolar concentrations of these plasminogen activators induced a moderate activation of the fibrinolytic system while the simultaneous combination produced a significant exhaustion of the fibrinolytic system.