Apixaban, a direct factor Xa inhibitor, inhibits tissue-factor induced human platelet aggregation in vitro: Comparison with direct inhibitors of factor VIIa, XIa and thrombin

Pancras C. Wong; Xiaosui Jiang

doi:10.1160/TH10-02-0097

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2010; 104(02): 302-310
DOI: 10.1160/TH10-02-0097

Platelets and Blood Cells

Schattauer GmbH

Apixaban, a direct factor Xa inhibitor, inhibits tissue-factor induced human platelet aggregation in vitro: Comparison with direct inhibitors of factor VIIa, XIa and thrombin

Pancras C. Wong

¹Thrombosis Research, Bristol-Myers Squibb Company, Pennington, New Jersey, USA

,

Xiaosui Jiang

¹Thrombosis Research, Bristol-Myers Squibb Company, Pennington, New Jersey, USA

› Author Affiliations

Abstract

Summary

Apixaban is an oral, direct and highly selective factor Xa (FXa) inhibitor in late-stage clinical development. This study evaluated the in vitro effect of apixaban on human platelet aggregation induced by thrombin derived via the extrinsic pathway. Direct inhibitors of FXa (rivaroxaban), FVIIa (BMS-593214), thrombin (dabigatran, argatroban) and FXIa (BMS-262084) were included for comparison. Citrated human plateletsrich plasma (PRP) was treated with 50 μg/ml corn trypsin inhibitor (to block the contact factor pathway) and 3 mM H-Gly-Pro-Arg- Pro-OH-AcOH (to prevent fibrin polymerisation). Human tissue factor (TF) (Innovin®; dilution 1:1,000 to 1:1,500) plus 7.5 mM CaCl₂ was added to PRP pre-incubated with vehicle or increasing concentrations of inhibitors. The TF-induced platelet aggregation was measured by optical aggregometry. TF produced 85 ± 3% aggregation of human platelets in the vehicle-treated group (n=10). Apixaban and other factor inhibitors, except the FXIa inhibitor, inhibited TF-induced platelet aggregation with IC₅₀ (nM) values as follows: 4 ± 1 (apixaban), 8 ± 2 (rivaroxaban), 13 ± 1 (BMS-593214), 46 ± 1 (dabigatran) and 79 ± 1 (argatroban). BMS-262084 (IC₅₀ = 2.8 nM vs. human FXIa) had no effect on TF-induced platelet aggregation at 10 μM. These inhibitors at 10 μM had no effect on platelet aggregation induced by ADP and collagen, as expected from their mechanism of action. This study demonstrates that inhibition of thrombin generation by blocking upstream proteases (FVIIa and FXa) in the blood coagulation cascade is as effective as direct thrombin inhibition in preventing TF-induced platelet aggregation. Under these experimental conditions, a FXIa inhibitor did not prevent TF-induced platelet aggregation.

Keywords

Apixaban - dabigatran - direct factor Xa inhibitor - platelet aggregation - rivaroxaban - tissue factor

Full Text

References

References
1 Pinto DJ, Orwat MJ, Koch S. et al. Discovery of 1-(4-methoxyphenyl)-7-oxo-6-(4-(2-oxopiperidin-1-yl)phenyl)-4,5,6,7-tetrahydro- 1H-pyrazolo[3,4-c>pyridine-3-carboxamide (apixaban, BMS-562247), a highly potent, selective, efficacious, and orally bioavailable inhibitor of blood coagulation factor Xa. J Med Chem 2007; 50: 5339-5356.
2 Wong PC, Crain EJ, Xin B. et al. Apixaban, an oral, direct and highly selective factor Xa inhibitor: in vitro, antithrombotic and antihemostatic studies. J Thromb Haemost 2008; 06: 820-829.
3 Wong PC, Watson CA, Crain EJ. Arterial antithrombotic and bleeding time effects of apixaban, a direct factor Xa inhibitor, in combination with antiplatelet therapy in rabbits. J Thromb Haemost 2008; 06: 1736-41.
4 Wong PC, Crain EJ, Watson CA. et al. Favorable therapeutic index of the direct factor Xa inhibitors, apixaban and rivaroxaban, compared to the thrombin inhibitor dabigatran in rabbits. J Thromb Haemost 2009; 07: 1313-1320.
5 Luettgen JM, Wang Z, Seiffert DA. et al. Inhibition of measured thrombin generation in human plasma by apixaban: a predictive mathematical model based on experimentally determined rate constants. J Thromb Haemost 2007; 05 (Suppl. 02) PT633.
6 Jiang X, Crain EJ, Luettgen JM. et al. Apixaban, an oral direct factor Xa inhibitor, inhibits human clot-bound factor Xa activity in vitro. Thromb Haemost 2009; 101: 780-782.
7 Frost C, Yu Z, Moore K. et al. Apixaban, an oral direct factor Xa inhibitor: multiple-dose safety, pharmacokinetics, and pharmacodynamics in healthy subjects. J Thromb Haemost 2007; 05 (Suppl. 02) PM664.
8 Lassen MR, Davidson BL, Gallus A. et al. The efficacy and safety of apixaban, an oral, direct factor Xa inhibitor, as thromboprophylaxis in patients following total knee replacement. J Thromb Haemost 2007; 05: 2368-2375.
9 Lassen MR, Raskob GE, Gallus A. et al. Apixaban or enoxaparin for thromboprophylaxis after knee replacement. N Engl J Med 2009; 361: 594-604.
10 Lassen MR, Raskob GE, Gallus A. et al. ADVANCE-2 investigators. Apixaban versus enoxaparin for thromboprophylaxis after knee replacement (ADVANCE-2): a randomised double-blind trial. Lancet 2010; 375: 807-815.
11 Botticelli Investigators, Writing Committee Büller H, Deitchman D, Prins M. et al. Efficacy and safety of the oral direct factor Xa inhibitor apixaban for symptomatic deep vein thrombosis. The Botticelli DVT dose-ranging study. J Thromb Haemost 2008; 06: 1313-1318.
12 APPRAISE Steering Committee and Investigators Alexander JH, Becker RC, Bhatt DL. et al. Apixaban, an oral, direct, selective factor Xa inhibitor, in combination with antiplatelet therapy after acute coronary syndrome. Results of the apixaban for prevention of acute ischemic and safety events (APPRAISE) trial. Circulation 2009; 119: 2877-2885.
13 Coughlin SR. Protease-activated receptors in hemostasis, thrombosis and vascular biology. J Thromb Haemost 2005; 03: 1800-1814.
14 Mann KG, Brummel K, Butenas S. What is all that thrombin for?. J Thromb Haemost 2003; 01: 1504-1514.
15 Mackman N, Tilley RE, Key NS. Role of the extrinsic pathway of blood coagulation in hemostasis and thrombosis. Arterioscler Thromb Vasc Biol 2007; 27: 1687-1693.
16 De Lucca I, Zhou J, Saiah E. et al. A selective FVIIa inhibitor with efficacy in an arterial thrombosis model. Abstracts of Papers, 230th ACS National Meeting, Washington, DC, United States, Aug. 28-Sept. 1. 2005
17 Wong P, Luettgen JM, Rendina AR. et al. BMS-593214, an active site-directed factor VIIa inhibitor: Enzyme kinetics, antithrombotic and antihemostatic studies. Thromb Haemost 2010; 104: 261-269.
18 Schumacher WA, Seiler SE, Steinbacher TE. et al. Antithrombotic and hemostatic effects of a small molecule factor XIa inhibitor in rats. Eur J Pharmacol 2007; 570: 167-174.
19 Wong PC, Schumacher WA, Watson CA. et al. Nonpeptide factor XIa (FXIa) Inhibitor. In vitro kinetic evaluation and in vivo studies of BMS-262084 on arterial thrombosis and hemostasis in rabbits. Circulation 2006; 114: @@II_104.
20 Yeh RW, Jang IK. Argatroban: update. Am Heart J 2006; 151: 1131-1138.
21 Perzborn E, Strassburger J, Wilmen A. et al. In vitro and in vivo studies of the novel antithrombotic agent BAY 59–7939--an oral, direct factor Xa inhibitor. J Thromb Haemost 2005; 03: 514-521.
22 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; 05: 1237-1242.
23 Eriksson I B, Quinlan DJ, Weitz I J. Comparative pharmacodynamics and pharmacokinetics of oral direct thrombin and factor Xa inhibitors in development. Clin Pharmacokinet 2009; 48: 1-22.
24 Mega I JL, Braunwald E, Mohanavelu I S. ATLAS ACS-TIMI 46 study group. Rivaroxaban versus placebo in patients with acute coronary syndromes (ATLAS ACS-TIMI 46): a randomised, double-blind, phase II trial. Lancet 2009; 374: 29-38.
25 Stiles I S. Oral anticoagulants REDEEMed? Daily dabigatran „safe“ with dual antiplatelets after MI. http://www.theheart.org/article/1026791.do Accessed 24 Jan 2010
26 Sutton JC, Bolton SA, Hartl KS. et al. Synthesis and SAR of 4-carboxy-2-azetidinone mechanism-based tryptase inhibitors. Bioorg Med Chem Lett 2002; 12: 3229-3233.
27 Hauel NH, Nar H, Priepke H. et al. Structure-based design of novel potent nonpeptide thrombin inhibitors. J Med Chem 2002; 45: 1757-1766.
28 Roehrig S, Straub A, Pohlmann J. et al. Discovery of the novel antithrombotic agent 5-chloro-N-({(5S)-2-oxo-3– [4-(3-oxomorpholin-4-yl)phenyl>-1,3-oxazolidin-5-yl}methyl)thiophene- 2-carboxamide (BAY 59–7939): an oral, direct factor Xa inhibitor. J Med Chem 2005; 48: 5900-5908.
29 Cawthern KM, van 't Veer C, Lock JB. et al. @@@Blood coagulation in hemophilia A and hemophilia C. Blood 1998; 91: 4581-4592.
30 He R, Xiong S, He X. et al. The role of factor XI in a dilute thromboplastin assay of extrinsic coagulation pathway. Thromb Haemost 2001; 85: 1055-1059.
31 Gailani D, Renné T. Intrinsic pathway of coagulation and arterial thrombosis. Arterioscler Thromb Vasc Biol 2007; 27: 2507-2513.
32 Blat Y, Seiffert D. A renaissance for the contact system in blood coagulation?. Thromb Haemost 2008; 99: 457-460.
33 von dem Borne PA, Meijers JC, Bouma BN. Feedback activation of factor XI by thrombin in plasma results in additional formation of thrombin that protects fibrin clots from fibrinolysis. Blood 1995; 86: 3035-3042.
34 Keularts IM, Zivelin A, Seligsohn U. et al. The role of factor XI in thrombin generation induced by low concentrations of tissue factor. Thromb Haemost 2001; 85: 1060-1065.
35 Perzborn E, Lange U. Rivaroxaban – an oral, direct factor Xa inhibitor – inhibits tissue factor-mediated platelet aggregation. J Thromb Haemost 2007; 05 (Suppl. 02) PW642.
36 van Ryn J, Kink-Eiband M, Hauel N. et al. Effects of dabigatran, a direct thrombin inhibitor, as compared to the direct factor Xa inhibitors, rivaroxaban and apixaban, on tissue factor-induced human platelet aggregation in platelet rich plasma. Blood (ASH Annual Meeting Abstracts) 2007 @@110 Abstract 1884
37 Swan SK, Hursting MJ. The pharmacokinetics and pharmacodynamics of argatroban: effects of age, gender, and hepatic or renal dysfunction. Pharmacotherapy 2000; 20: 318-329.
38 Eriksson I B, Quinlan DJ, Weitz I J. Comparative pharmacodynamics and pharmacokinetics of oral direct thrombin and factor Xa inhibitors in development. Clin Pharmacokinet 2009; 48: 1-22.
39 Stangier J. Clinical pharmacokinetics and pharmacodynamics of the oral direct thrombin inhibitor dabigatran etexilate. Clin Pharmacokinet 2008; 47: 285-295.
40 Mueck W, Borris LC, Dahl OE. et al. Population pharmacokinetics and pharmacodynamics of once- and twice-daily rivaroxaban for the prevention of venous thromboembolism in patients undergoing total hip replacement. Thromb Haemost 2008; 100: 453-461.