Inhibition of purified factor Xa amidolytic activity may not be predictive of inhibition of in vivo thrombosis: implications for identification of therapeutically active inhibitors.

Objective-In this study we test the hypothesis that blood/plasma-based prothrombinase assays, rather than inhibition of purified factor Xa (fXa), are predictive of in vivo antithrombotic activity. Methods and Results-Six fXa inhibitors with equivalent nanomolar K-i were studied in thrombin generation assays using human plasma/blood and endogenous macromolecular substrate. In all assays, benzamidine inhibitors were more potent (100 to 800 nmol/L) than the aminoisoquinolines (5 to 58 mumol/L) or neutral inhibitors (3 to 10 mumol/L). A similar rank order of compound inhibition was also seen in purified prothrombinase assays as well as in a rabbit model of deep vein thrombosis. Conclusions-Assays using prothrombinase with protein substrates are better predictors of in vivo efficacy than fXa Ki using amidolytic substrates.