Factor Xa FXa has emerged as a particularly

Factor Xa (FXa) has emerged as a particularly promising target for the development of effective anticoagulant drugs because FXa acts at the convergence point of the extrinsic and intrinsic coagulation pathways. It efficiently catalyzes the conversion of prothrombin to thrombin with one molecule of FXa resulting in the generation of more than 1000 thrombin N6022 receptor (Krishnaswamy, 2013, Majumder et al., 2013, Mani and Lindhoff-Last, 2014). An oral, directly acting FXa inhibitor that does not require routine coagulation monitoring would offer significant advantages over current therapies. In this study, we investigated zifaxaban inhibition of FXa activity in vitro as well as its in vivo antithrombotic efficacy in rat venous thrombosis, arteriovenous-shunt thrombosis and carotid thrombosis models. In addition, the bleeding risks of zifaxaban and rivaroxaban were compared in a rat model of venous thrombosis by the tail-off method. Furthermore, the PT prolongation by zifaxaban was investigated in human, rat, and rabbit models. The aim of the study was to determine the clinical pharmacodynamics of zifaxaban, to provide experimental evidence for a further clinical study of the drug for the prevention and treatment of arteriovenous thromboembolic diseases.
Materials and methods
Results
Discussion FXa is a critical factor in the blood coagulation cascade and has become a target for treating the thrombotic diseases that are a leading cause of morbidity and mortality worldwide (Misumida et al., 2015, Wolberg et al., 2015). Inhibition of FXa activity can reduce thrombin generation, there by inhibiting blood clotting and the formation of intravascular thrombi and emboli. The existing oral FXa inhibitors include rivaroxaban, apixaban, edoxaban and betrixaban. Zifaxaban is a novel FXa inhibitor developed by the Tianjin Institute of Pharmaceutical Research, and it has a potency that is similar to that of rivaroxaban. In the present study, we investigated the pharmacology of zifaxaban, through a series of in vitro and in vivo assays. We demonstrated that zifaxaban is a highly potent and selective FXa inhibitor. Moreover, zifaxaban significantly inhibited clotting in various rat models of thrombosis and showed a trend of less bleeding than rivaroxaban. Previous studies have shown that there are species differences in FXa inhibition in humans, rabbits, rats and dogs with the FXa inhibitors. The current study extended these findings, and showed that zifaxaban was more potent in prolonging APTT and PT in human and rabbit plasma, compared to rat plasma. Zifaxaban was more potent in prolonging PT and APTT, compared with TT, in vitro and in vivo. We studied zifaxaban in vivo in rats, administering the agent by gavage, and then evaluated its antithrombotic efficacy in different experimental models of thrombosis. These included a venous thrombosis model, in which an occlusive thrombus is formed. Venous thrombosis was mimicked through the induction of stasis and hypercoagulability (Albadawi et al., 2017, Diaz et al., 2012). In an arteriovenous bypass thrombosis model, thrombosis was induced on silk thread in an arteriovenous shunt. This was performed as previously reported, with some minor modifications (Yu et al., 2014). Finally, in a carotid thrombosis model, thrombosis was produced in segment of carotid artery that was subjected to electrolytic injury, mimicking arterial thrombosis (Wong et al., 2000). Zifaxaban exhibited strong antithrombotic activity in these rat models, and had comparable effects to those reported in previous studies of rivaroxaban. Zifaxaban was equally potent for prevention of both arterial and venous thrombosis. The wet weight and dry weights of the thrombi were significantly reduced following a dose-effect relationship for zifaxaban. Moreover, the bleeding time of tail-off, as well as the PT and APTT were significantly prolonged after the administration of zifaxaban in rats. Moreover, the activity of FXa was significantly reduced in a dose-dependent manner after the administration of zifaxaban. The results indicate that the antithrombotic effects of zifaxaban are consistent with selective inhibition of FXa. In vitro studies showed that zifaxaban was highly selective for FXa and did not directly change platelet aggregation. Ex vivo studies showed that zifaxaban inhibited FXa activity. However, by inhibiting FXa, zifaxaban should inhibit thrombin generation in vivo, thus reducing thrombin-induced platelet activation and producing an indirect antiplatelet effect.