Effects of Rivaroxaban on Platelet Activation and Platelet-Coagulation Pathway Interaction: In Vitro and In Vivo Studies

Abstract

Introduction: Activation of coagulation and platelets is closely linked, and arterial thrombosis involves coagulation activation as well as platelet activation and aggregation. In these studies, we investigated the possible synergistic effects of rivaroxaban in combination with antiplatelet agents on thrombin generation and platelet aggregation in vitro and on arterial thrombosis and hemostasis in rat models.

Materials and methods: Thrombin generation was measured by the Calibrated Automated Thrombogram method (0.5 pmol/L tissue factor) using human platelet-rich plasma (PRP) spiked with rivaroxaban (15, 30, or 60 ng/mL), ticagrelor (1.0 µg/mL), and acetylsalicylic acid (ASA; 100 µg/mL). Tissue factor-induced platelet aggregation was measured in PRP spiked with rivaroxaban (15 or 30 ng/mL), ticagrelor (1 or 3 µg/mL), or a combination of these. An arteriovenous (AV) shunt model in rats was used to determine the effects of rivaroxaban (0.01, 0.03, or 0.1 mg/kg), clopidogrel (1 mg/kg), ASA (3 mg/kg), and combinations on arterial thrombosis.

Results: Rivaroxaban inhibited thrombin generation in a concentration-dependent manner and the effect was enhanced with ticagrelor and ticagrelor plus ASA. Rivaroxaban and ticagrelor also concentration-dependently inhibited tissue factor-induced platelet aggregation, and their combination increased the inhibition synergistically. In the AV shunt model, rivaroxaban dose-dependently reduced thrombus formation. Combining subefficacious or weakly efficacious doses of rivaroxaban with ASA or ASA plus clopidogrel increased the antithrombotic effect.

Conclusion: These data indicate that the combination of rivaroxaban with single or dual antiplatelet agents works synergistically to reduce platelet activation, which may in turn lead to the delayed/reduced formation of coagulation complexes and vice versa, thereby enhancing antithrombotic potency.

Keywords: antiplatelet agents; combination drug therapy; platelet aggregation; rivaroxaban; thrombosis.

Figure 1.

Representative thrombograms in platelet-rich plasma: (A) in the presence of a control, rivaroxaban (15, 30, or 60 ng/mL), or ASA (100 µg/mL); (B) in the presence of a control, rivaroxaban (60 ng/mL), ticagrelor (1.0 µg/mL), or in the combination of rivaroxaban (60 ng/mL) plus ticagrelor (1.0 µg/mL) or rivaroxaban (60 ng/mL) plus ticagrelor (1.0 µg/mL) and ASA (100 µg/mL). ASA indicates acetylsalicylic acid.

Figure 2.

Thrombin generation in platelet-rich plasma in the presence of rivaroxaban (15, 30, or 60 ng/mL), ticagrelor (1 µg/mL), and ASA (100 µg/mL), and the combination of rivaroxaban plus ticagrelor or rivaroxaban plus ticagrelor and ASA. (A) C_max; (B) lag time; (C) T_max; and (D) mean velocity. Results are presented as mean values ± standard error of the mean (n = 11). *P < .05 versus control; **P < .01 versus control; ***P < .001 versus control. ASA indicates acetylsalicylic acid; C_max, peak thrombin generation; Riva, rivaroxaban; Tica, ticagrelor; T_max, time to peak thrombin generation.

Figure 3.

Inhibition of tissue factor-induced platelet aggregation in human platelet-rich plasma by rivaroxaban and ticagrelor. Results are presented as mean values ± standard error of the mean (ticagrelor, n = 9-13; rivaroxaban, n = 11-13). ***P < .001 versus control.

Figure 4.

Inhibition of thrombus formation after administration of (A) rivaroxaban (0.1 mg/kg [n = 10], 0.3 mg/kg [n = 9], or 1.0 mg/kg [n = 10] intravenous); (B) clopidogrel (1.0, 3.0, 10, or 30 mg/kg oral; n = 6 per dosing group); or (C) ASA (1.0, 3.0, 10, 30, or 100 mg/kg oral; n = 6 per dosing group) in an arteriovenous shunt model in rats. Results are presented as mean values ± standard error of the mean. *P < .05 versus control; **P < .01 versus control; ***P < .001 versus control. ASA indicates acetylsalicylic acid.