Design, synthesis, and biological activity of potent and selective inhibitors of blood coagulation factor Xa.

Factor Xa (FXa) has materialized as a key enzyme for the intervention of the blood coagulation cascade and for the development of new antithrombotic agents. FXa is the lone enzyme responsible for the production of thrombin and therefore is an attractive target for the control of thrombus formation. We have designed and synthesized a unique series of quinoxalinone FXa inhibitors. This series resulted in 3-[4-[5-((2S,6R)-2,6-dimethylpiperidin-1-yl)pentyl]-3-oxo-3,4-dihydroquinoxolin-2-yl]benzamidine (35) with 0.83 nM activity against FXa and excellent selectivity over similar serine proteases. An X-ray crystal structure of compound 35 bound to trypsin along with molecular modeling has led to a predicted binding conformation of compound 35 in FXa. Compound 35 has also been proven to be efficacious in vivo in both the rabbit veno-venous shunt and dog electrolytic injury models. In addition, it was shown that compound 35 did not significantly increase bleeding times in a rabbit model except at the highest doses and plasma concentrations were elevated in a dose dependent manner following a bolus dose and continuous intravenous infusion.

Assessment of ex vivo pharmacodynamic markers during inhibition of thrombosis by CI-1031 (ZK-807834), a novel direct factor Xa inhibitor.

CI-1031 (ZK-807834) is a novel, synthetic factor Xa (FXa) inhibitor with a Ki of 0.11 nM against human FXa. In human plasma in vitro, CI-1031 doubled PT and aPTT at 0.23 and 0.49 microM, respectively. The in vivo antithrombotic effect of CI-1031 was evaluated in a veno-venous shunt model of thrombosis in anesthetized rabbits. After thrombus formation was verified in the first shunts, rabbits received either vehicle or CI-1031 intravenously (bolus injection of 60, 240, or 480 microg/kg followed by an infusion of 2, 8, or 16 microg/kg/min for 140 min, respectively). The second shunts were inserted after 20 min of infusion of CI-1031 or vehicle. CI-1031 dose-dependently prolonged time to occlusion (TTO) in the second shunts (35 +/- 21, 62 +/- 24, and 120 +/- 0 min for the three dose groups, respectively, vs. 10 +/- 1 min for vehicle). Thrombus mass (TM) was reduced in a dose-dependent manner by CI-1031 (42 +/- 7, 27 +/- 6, and 18 +/- 4 mg vs. 50 +/- 4 mg for vehicle). Maximal TM reduction was 70% with an IC(50) of 0.6 microg/ml. Among all the coagulation parameters tested, PT had the best correlation with plasma CI- 1031 concentration (r = 0.97). Ex vivo plasma anti-FXa activity was also well correlated with plasma concentration of CI-1031 and with PT (r = 0.96 and 0.98, respectively). These results indicate that CI-1031, which is currently undergoing clinical evaluation, is an effective antithrombotic compound with a favorable efficacy-to-bleeding ratio. In addition, CI-1031 concentration in plasma can be monitored using PT or anti-Xa assays, thereby providing reliable methods to ensure safe and accurate dose titration of CI-1031.