Inhibition of Arterial Thrombus Formation in Two Canine Models: Comparison of Ancrod, a Fibrinogen-Depleting Agent, the Thrombin-Inhibitor r-Hirudin, and the Glycoprotein IIb/IIIa-Receptor Antagonist Ro 43-8857

Summary

Inhibition of arterial thrombus formation by ancrod, a fibrinogen depleting agent isolated from a snake venom, r-hirudin, an inhibitor of thrombin-mediated fibrinogen cleavage, or the glycoprotein (GP)IIb/IIIa-receptor antagonist Ro 43-8857 interfering with fibrinogen binding to platelets, was evaluated in two canine models. As a marker of platelet-dependent thrombus formation, cyclic blood flow reductions (CFR) were induced in the left coronary artery (LAD) of mongrel dogs by mechanical injury of the endothelium combined with critical stenosis. In the second model CFRs were induced by thrombolysis of a copper coil-induced thrombus in the carotid artery. Blood flow rate during the reocclusion phase was used as an additional parameter of efficacy.

The frequency of CFRs used as indicator of platelet aggregation and adhesion was significantly diminished by all treatments in both the carotid artery- and the LAD-model. In the LAD-model, following ancrod treatment, CFRs were correlated with plasma fibrinogen concentrations. Carotid artery blood flow after reperfusion, used as indicator of occlusive thrombus formation, rapidly declined to zero in the control group but remained at a high level after treatment with ancrod or r-hirudin. Ro 43-8857 at the selected dose improved blood flow rate only to a minor degree but prolonged the bleeding time from a mean value of 87.2 ± 10.9 s (n = 24) to values >300 s in 50% of the animals.

Our results indicate that CFRs as indicator of platelet aggregation and adhesion are inhibited by either treatments. Blood flow as indicator of occlusive thrombus formation, however, is effectively improved by ancrod and r-hirudin only. Inhibition of fibrinogen binding to platelet GPIIb/IIIa receptors alone was found to be a less potent antithrombotic principle in this model.

• References

• 1 Chesebro JH, Badimon L, Fuster V. New approaches to treatment of myocardial infarction. Am J Cardiol 1990; 65: 12C-19C
• 2 Yasuda T, Gold H, Fallon JT, Leinbach RC, Guerrero JL, Scudder LE, Kanka M, Shealy D, Ross MJ, Collen D, Coller BS. Monoclonal antibody against the platelet glycoprotein (GP) Ilb/IIIa receptor prevents coronary artery reocclusion after reperfusion with recombinant tissue-type plasminogen activator in dogs. J Clin Invest 1988; 81: 1284-1291
• 3 Yasuda T, Gold HK, Yaoita H, Leinbach RC, Guerrero JL, Jang I-K, Holt R, Fallon JT, Collen D. Comparative effects of aspirin, a synthetic thrombin inhibitor and a monoclonal antiplatelet glycoprotein Ilb/IIIa antibody on coronary artery reperfusion, reocclusion and bleeding with recombinant tissue-type plasminogen activator in a canine preparation. J Am Coll Cardiol 1990; 16: 714-722
• 4 Folts JD, Crowell EB, Rowe GG. Platelet aggregation in partially obstructed vessels and its elimination with aspirin. Circulation 1976; 54: 365-370
• 5 Coller BS, Folts JD, Smith SR, Scudder LE, Jordan R. Abolition of in vivo platelet thrombus formation in primates with monoclonal antibodies to the platelet GPIIb/IIIa receptor. Correlation with bleeding time, platelet aggregation,and blockade of GPIIb/IIIa receptors Circulation 1989; 80: 176674
• 6 Mirshahi M, Soria J, Soria C, Faivre R, Lu H, Courtney M, Roitsch C, Tripier D, Caen J. Evaluation of the inhibition by heparin and hirudin of coagulation activation during rt-PA-induced thrombolysis. Blood 1989; 74: 1025-1030
• 7 Weitz JI, Hudoba M, Massel D, Maraganore J, Hirsh J. Clot-bound thrombin is protected from inhibition by heparin-antithrombin III but is susceptible to inactivation by antithrombin IlI-independent inhibitors. J Clin Invest 1990; 86: 385-391
• 8 Esnouf MP, Tunnah GW. The isolation and properties of the thrombin-like activity from Ancistrodon rhodostoma venom. Brit J Haemat 1968; 15: 589-602
• 9 Bell WR, Bolton G, Pitney WR. The effect of arvin on blood coagulation factors. Brit J Haemat 1968; 15: 589-602
• 10 Edgar W, Prentice CR M. The proteolytic action of ancrod on human fibrinogen and its polypeptide chains. Thromb Res 1973; 2: 85-96
• 11 Brown CH, Bell WR, Shreiner DP, Jackson DP. Effect of arvin on blood platelets In vitro and in vivo studies. J Lab Clin Med 1972; 79: 758-769
• 12 Kordenat RK, Kezdi P, Stanley EL. A new catheter technique for producing experimental coronary thrombosis and selective coronary visualization. Amer Heart J 1972; 83: 360-364
• 13 Brassard JA, Meyers KM. Evaluation of the buccal bleeding time and platelet glass bead retention as assays of hemostasis in the dog: The effects of acetylsalicylic acid, warfarin and von Willebrand factor deficiency. Thromb Haemost 1991; 65: 191-195
• 14 Clauss A. Rapid physiological coagulation method for the determination of fibrinogen. Acta Haematol 1957; 17: 237-239
• 15 Alig L, Edenhofer A, Hadvary P, Hiirzeler M, Knopp D, Muller M, Steiner B, Trzeciak A, Weller T. Low molecular weight, non-peptide fibrinogen receptor antagonists. J Med Chem 1992; 35: 4393-4407
• 16 Folts J. An in vivo model of experimental arterial stenosis, intimal damage, and periodic thrombosis. Circulation 1991; 83 Suppl (Suppl. 04) IV-3-IV-14
• 17 Cerek B, Lew AS, Hod H, Yano J, Lewis B, Reddy KN N, Ganz W. Ancrod enhances the thrombolytic effect of streptokinase and urokinase. Thromb Res 1987; 47: 417-426
• 18 Tomaru T, Uchida Y, Nakamura F, Sonoki H, Tsukamoto M, Sugimoto T. Enhancement of arterial thrombolysis with native tissue-type plasminogen activator by pretreatment with heparin or batroxobin: An angioscopic study. Am Heart J 1989; 117: 275-281
• 19 Apprill PG, Ashton J, Guerrero J, Glas-Greenwalt P, Buja LM, Willerson JT. Ancrod decreases the frequency of cyclic flow variations and causes thrombolysis following acute coronary thrombosis. Am Heart J 1987; 113: 898-906
• 20 Rapold HJ, Gold HK, Wu Z, Napier M, Bunting S, Collen D. Effects of G4120, a ARG-GLY-ASP containing synthetic platelet glycoprotein Ilb/IIIa receptor antagonist, on arterial and venous thrombolysis with recombinant tissue-type plasminogen activator in dogs. Fibrinolysis 1993; 7: 248-56
• 21 Rote WE, Wems SW, Davis JH, Feigen LP, Kilgore KS, Lucchesi BR. Platelet GPIIb/IIIa receptor inhibition by SC-49992 prevents thrombosis and rethrombosis in the canine carotid artery. Cardiovasc Res 1993; 27: 500-507
• 22 Haskel EJ, Prager NA, Sobel BE, Abendschein DR. Relative efficacy of antithrombin compared with antiplatelet agents in accelerating coronary thrombolysis and preventing early reocclusion. Circulation 1991; 83: 1048-1056
• 23 Cook NS, Bruttger O, Pally C, Hagenbach A. The effects of two synthetic glycoprotein Ilb/IIIa antagonists, Ro 43-8857 and L-700,462, on platelet aggregation and bleeding in guinea-pigs and dogs: Evidence that Ro 438857 is orally active. Thromb Haemost 1993; 70: 838-847
• 24 Rote WE, Mu DX, Bates ER, Nedelman MA, Lucchesi BR. Prevention of rethrombosis after coronary thrombolysis in a chronic canine model. I. Adjunctive therapy with monoclonal antibody 7E3 F(ab’)₂fragment J Cardiovasc Pharmacol 1994; 23: 194-202
• 25 Rote WE, Mu DX, Bates ER, Nedelman MA, Lucchesi BR. Prevention of rethrombosis after coronary thrombolysis in a chronic canine model. II. Adjunctive therapy with r-hirudin J Cardiovasc Pharmacol 1994; 23: 203-211
• 26 Heras M, Chesebro JH, Webster MW I, Mruk JS, Grill DE, Penny WJ, Bowie EJ W, Badimon L, Fuster V. Hirudin, heparin, and placebo during deep arterial injury in the pig. The in vivo role of thrombin in platelet- mediated thrombosis Circulation 1990; 82: 1476-1484
• 27 Yao SK, McNatt J, Anderson HV, Eidt J, Cui K, Golino P, Glas-Greenwalt P, Maraganore J, Buja LM, Willerson JT. Thrombin inhibition enhances tissue-type plasminogen activator-induced thrombolysis and delays reocclusion. Am J Physiol 1992; 262: H374-H379
• 28 Rigel DF, Olson RW, Lappe RW. Comparison of hirudin and heparin as adjuncts to streptokinase thrombolysis in a canine model of coronary thrombosis. Circulation Res 1993; 72: 1091-1102
• 29 Prager NA, Torr-Brown SR, Sobel BE, Abendschein DR. Maintenance of patency after thrombolysis in stenotic coronary arteries requires combined inhibition of thrombin and platelets. J ACC 1993; 22: 296-301
• 30 Nicolini FA, Lee P, Rios G, Kottke-Marchant K, Topol EJ. Combination of platelet fibrinogen receptor antagonist and direct thrombin inhibitor at low doses markedly improves thrombolysis. Circulation 1994; 89: 1802-1809
• 31 Kirchengast M, Münter K, Rübsamen K. The Ca²⁺antagonist verapamil potentiates the effect of the ACE-inhibitor trandolapril on cyclic coronary blood flow reductions. J Vase Med Biol 1994; 5: 144-151
• 32 The Ancrod Stroke Study Investigators. Ancrod for the treatment of acute ischemic brain infarction. Stroke 1994; 25: 1755-1759
• 33 Garabedian HK, Gold HK, Leinbach RC, Yasuda T, Johns JA, Thornton D, Collen D. Laboratory monitoring of hemostasis during thrombolytic therapy with recombinant human tissue-type plasminogen activator. Thromb Res 1988; 50: 121-133