Molecular mechanisms of fibrinolysis and their application to fibrin-specific thrombolytic therapy

Abstract

The fibrinolytic system comprises a proenzyme, plasminogen, which can be converted to the active enzyme, plasmin, which degrades fibrin. Plasminogen activation is mediated by plasminogen activators, which are classified as either tissue-type plasminogen activators (t-PA) or urokinase-type plasminogen activators (u-PA). Inhibition of the fibrinolytic system may occur at the level of the activators or at the level of generated plasmin.

Plasmin has a low substrate specificity, and when circulating freely in the blood it degrades several proteins including fibrinogen, factor V, and factor VIII. Plasma does, however, contain a fast-acting plasmin inhibitor, α₂-antiplasmin, which inhibits free plasmin extremely rapidly but which reacts much slower with plasmin bound to fibrin. A “systemic fibrinolytic state” may, however, occur by extensive activation of plasminogen and depletion of α₂-antiplasmin. Clot-specific thrombolysis therefore requires plasminogen activation restricted to the vicinity of the fibrin.

Two physiological plasminogen activators, t-PA and single-chain u-PA (scu-PA) induce clot-specific thrombolysis, via entirely different mechanisms, however. t-PA is relatively inactive in the absence of fibrin, but fibrin strikingly enhances the activation rate of plasminogen by t-PA. This is explained by an increased affinity of fibrin-bound t-PA for plasminogen and not by alteration of the catalytic rate constant of the enzyme. The high affinity of t-PA for plasminogen in the presence of fibrin thus allows efficient activation on the fibrin clot, while no significant plasminogen activation by t-PA occurs in plasma. scu-PA has a high affinity for plasminogen (K_m = 0.3 μM) but a low catalytic rate constant (k_cat = 0.02 sec⁻¹). However, scu-PA does not activate plasminogen in plasma in the absence of a fibrin clot, owing to the presence of (a) competitive inhibitor(s). Fibrin-specific thrombolysis appears to be due to the fact that fibrin reverses the competitive inhibition.

The thrombolytic efficacy and fibrin specificity of natural and recombinant t-PA has been demonstrated in animal models of pulmonary embolism, venous thrombosis, and coronary artery thrombosis. In all these studies intravenous infusion of t-PA at sufficiently high rates caused efficient thromblysis in the absence of systemic fibrinolytic activation.

The efficacy and relative fibrinogen-sparing effect of t-PA was recently confirmed in three multicenter clinical trials in patients with acute myocardial infarction. Intravenous infusion of 0.5–1 mg of t-PA per kg body weight over 1–3 hr resulted in coronary reperfusion in approximately 70% of patients. It raised the plasma level about 1,000-fold but was associated with an average decrease of the plasma fibrinogen level by 30%.

Specific thrombolysis by scu-PA has also been demonstrated in animal models of pulmonary embolism, venous thrombosis, and coronary thrombosis, Again, intravenous infusion of scu-PA at sufficiently high rates caused thrombolysis in the absence of systemic fibrinolytic activation. We have treated six patients with acute myocardial infarction with scu-PA and have obtained coronary reperfusion during intravenous infusion of 40 mg scu-PA over 60 min in four of the patients and during subsequent intracoronary infusion in one additional patient. A decrease of fibrinogen to 25% of the preinfusion value was observed in one patient.