Profibrinolytic activity of the direct thrombin inhibitor melagatran and unfractionated heparin in platelet-poor and platelet-rich clots

Summary

Anticoagulants have been shown to stimulate fibrinolysis principally via inhibition of thrombin-mediated activation of TAFI (thrombin activatable fibrinolysis inhibitor). Their profibrinolytic effect,however,may vary according to their mechanism of action and to the clot composition. We compared the fibrinolytic activity of the direct thrombin inhibitor melagatran with that of unfractionated heparin in platelet-poor (PPP) and platelet-rich (PRP) models consisting of tissue-factor-induced clots exposed to exogenous t-PA (25 ng/ml). In the PPP clot model, both heparin (0.1–0.6 U/ml) and melagatran (20–320 ng/ml) caused a concentration- dependent shortening of lysis time. However, when drug profibrinolytic activity (lysis ratio) was expressed in function of the aPTT prolongation (aPTT ratio), melagatran was more efficient than heparin. In the PRP clot model, melagatran displayed a fibrinolytic activity fairly comparable to that observed in PPP whilst heparin caused a modest reduction of lysis time only at the highest concentrations. Assay of thrombin and TAFIa generation in defibrinated plasma showed that the presence of platelets markedly reduced the ability of heparin,but not that of melagatran, to inhibit the formation of these enzymes. Altogether these data indicate that melagatran is more efficient than heparin in promoting fibrinolysis, particularly in plateletrich clots, and may thus grant a greater antithrombotic activity by enhancing thrombus dissolution.

Keywords

Fibrinolysis - anticoagulants - thrombin - TAFI - platelets

^* Contributed equally to the work.

• References

• 1 Broze Jr. GJ. Thrombin-dependent inhibition of fibrinolysis. Curr Opin Hematol 1996; 3: 390-394.
• 2 Bajzar L. Thrombin activatable fibrinolysis inhibitor and an antifibrinolytic pathway. Arterioscler Thromb Vasc Biol 2000; 20: 2511-2518.
• 3 Leurs J, Hendriks D. Carboxypeptidase U (TAFIa): a metallocarboxypeptidase with a distinct role in haemostasis and a possible risk factor for thrombotic disease. Thromb Haemost 2005; 94: 471-487.
• 4 Bajzar L, Nesheim ME, Tracy PB. The profibrinolytic effect of activated protein C in clots formed from plasma is TAFI-dependent. Blood 1996; 88: 2093-2100.
• 5 Lisman T, de Groot PG. Rebuttal to: Effect of heparin on TAFI-dependent inhibition of fibrinolysis. J Thromb Haemost 2003; 1: 200-201.
• 6 Lisman T, Adelmeijer J, Nieuwenhuis HK. et al. Enhancement of fibrinolytic potential in vitro by anticoagulant drugs targeting activated factor X, but not by those inhibiting thrombin or tissue factor. Blood Coagul Fibrinolysis 2003; 14: 557-562.
• 7 Mann KG, Brummel K, Butenas S. What is all that thrombin for?. J Thromb Haemost 2003; 1: 1504-1514.
• 8 Bouma BN, Meijers JC. Thrombin-activatable fibrinolysis inhibitor (TAFI, plasma procarboxypeptidase B, procarboxypeptidase R, procarboxypeptidase U). J Thromb Haemost 2003; 1: 1566-1574.
• 9 Lindhout T, Baruch D, Schoen P. et al. Thrombin generation and inactivation in the presence of antithrombin III and heparin. Biochemistry 1986; 25: 5962-5969.
• 10 Taube J, McWilliam N, Luddington R. et al. Activated protein C resistance: effect of platelet activation, platelet-derived microparticles, and atherogenic lipoproteins. Blood 1999; 93: 3792-3797.
• 11 Eitzman DT, Chi L, Saggin L. et al. Heparin neutralization by platelet-rich thrombi. Role of platelet factor 4. Circulation 1994; 89: 1523-1529.
• 12 Hrebickova L, Nawarskas JJ, Anderson JR. Ximelagatran: a new oral anticoagulant. Heart Dis 2003; 5: 397-408.
• 13 Nagashima H. Studies on the different modes of action of the anticoagulant protease inhibitors DX-9065a and Argatroban. II. Effects on fibrinolysis. J Biol Chem 2002; 277: 50445-50449.
• 14 Colucci M, Pentimone A, Binetti BM. et al. Effect of heparin on TAFI-dependent inhibition of fibrinolysis: relative importance of TAFIa generated by clotbound and fluid phase thrombin. Thromb Haemost 2002; 88: 282-287.
• 15 Colucci M, Binetti BM, Tripodi A. et al. Hyperprothrombinemia associated with prothrombin G20210A mutation inhibits plasma fibrinolysis through a TAFImediated mechanism. Blood 2004; 103: 2157-2161.
• 16 Mutch NJ, Thomas L, Moore NR, Lisiak KM, Booth NA. TAFIa, PAI-1 and alpha-antiplasmin: complementary roles in regulating lysis of thrombi and plasma clots. J Thromb Haemost 2007; 5: 812-817.
• 17 Sakharov D, Plow EF, Rijken DC. On the mechanism of the antifibrinolytic activity of plasma carboxypeptidase B. J Biol Chem 1997; 272: 14477-14482.
• 18 Hemker HC, Al Dieri R, De Smedt E. et al. Thrombin generation, a function test of the haemostaticthrombotic system. Thromb Haemost 2006; 96: 553-561.
• 19 Nesheim M, Wang W, Boffa M. et al. Thrombin, thrombomodulin and TAFI in the molecular link between coagulation and fibrinolysis. Thromb Haemost 1997; 78: 386-391.
• 20 Bajzar L, Fredenburgh JC, Nesheim M. The activated protein C-mediated enhancement of tissue-type plasminogen activator-induced fibrinolysis in a cellfree system. J Biol Chem 1990; 265: 16948-16954.
• 21 Colucci M, Binetti BM, Semeraro N. Platelets abolish the profibrinolytic effect of activated protein C. Thromb Haemost 2000; 84: 922-923.
• 22 Weitz JI, Hudoba M, Massel D. et al. Clot-bound thrombin is protected from inhibition by heparin-antithrombin III but is susceptible to inactivation by antithrombin III-independent inhibitors. J Clin Invest 1990; 86: 385-391.
• 23 Ofosu FA, Cherskus AL, Hirsh J. et al. The inhibition of the anticoagulant activity of heparin by platelets, brain phospholipids and tissue factor. Br J Haematol 1984; 57: 229-238.
• 24 Bostrom SL, Dagnelid E, Hansson GF. et al. Inhibition of thrombin-induced feedback activation of factor V: a potential pathway for inhibition of thrombin generation by melagatran. Blood Coagul Fibrinolysis 2004; 15: 25-30.
• 25 Mattsson C, Bjorkman JA, Abrahamsson T. et al. Local proCPU (TAFI) activation during thrombolytic treatment in a dog model of coronary artery thrombosis can be inhibited with a direct, small molecule thrombin inhibitor (melagatran). Thromb Haemost 2002; 87: 557-562.
• 26 Crawley JTB, Zanardelli S, Chion CKNK, Lane DA. The central role of thrombin in hemostasis. J Thromb Haemost 2007; 5 (Suppl. 01) 95-101.
• 27 Wolberg AS. Thrombin generation and fibrin clot structure. Blood Rev 2007; 21: 131-142.
• 28 Sidelmann JJ, Gram J, Jespersen J. et al. Fibrin clot formation and lysis: basic mechanisms. Semin Thromb Hemost 2000; 26: 605-618.
• 29 Boos CJ, Lip GYH. Ximelagatran: an eulogy. Thromb Res 2006; 118: 301-304.
• 30 Hauel N, Nar H, Priepke H. et al. Stucture-based design of novel potent nonpeptide thrombin inhibitors. J Med Chem 2002; 45: 1757-1766.
• 31 Eriksson BI, Dahl OE, Buller HR. et al. A new oral direct thrombin inhibitor, dabigatran etexilate, compared with enoxaparin for prevention of thromboembolic events following total hip or knee replacement: the BISTRO II randomized trial. J Thromb Haemost 2005; 3: 103-111.
• 32 Wienen W, Stassen JM, Priepke H. et al. Antithrombotic and anticoagulant effects of the direct thrombin inhibitor dabigatran, and its oral prodrug, dabigatran etexilate, in a rabbit model of venous thrombosis. J Thromb Haemost 2007; 5: 1237-1242.
• 33 Wienen W, Stassen JM, Priepke H. et al. Effects of the direct thrombin inhibitor dabigatran and its orally active prodrug, dabigatran etexilate, on thrombus formation and bleeding time in rats. Thromb Haemost 2007; 98: 333-338.