Effect of BAX499 aptamer on tissue factor pathway inhibitor function and thrombin generation in models of hemophilia

Introduction: In hemophilia, thrombin generation is significantly suppressed due to decreased factor (F) X activation. Clinical studies and experiments with transgenic mice have suggested that the severity of hemophilia is substantially reduced by tissue factor pathway inhibitor (TFPI) deficiency. Methods: We evaluated the effect of TFPI antagonist aptamer BAX499 (formerly ARC19499) on TFPI function in purified systems and on thrombin generation and clot formation in plasma and blood. Results: BAX499 effectively neutralized TFPI inhibition of FXa and FXa dependent inhibition of TF/FVIIa by TFPI. BAX499 did not inhibit FXa or TF/FVIIa when used up to 500 nM. In the synthetic coagulation proteome with TFPI at its mean physiologic concentration, BAX499 at 1 - 10 nM increased thrombin generation triggered with 5 pM relipidated TF in a concentration-dependent manner. In severe hemophilia A or B models using the synthetic coagulation proteome, the addition of BAX499 at 5 nM increased thrombin generation to the levels observed in normal control. Thrombin generation measured in induced hemophilia B plasma required similar to 100 nM BAX499 to restore thrombin levels to those seen in untreated plasma. In induced hemophilia B whole blood, BAX499 repaired the clotting time but failed to appreciably impact the propagation phase of thrombin generation. Conclusion: These data suggest that inhibition of TFPI by BAX499 may have potential for hemophilia treatment but requires further study in blood-based hemophilia systems. (C) 2012 Elsevier Ltd. All rights reserved.