Improvement of spatial fibrin formation by the anti‐TFPI aptamer BAX499: changing clot size by targeting extrinsic pathway initiation

Summary. Background: Tissue factor pathway inhibitor (TFPI) is a major regulator of clotting initiation and a promising target for pro‐ and anticoagulation therapy. The aptamer BAX499 (formerly ARC19499) is a high‐affinity specific TFPI antagonist designed to improve hemostasis. However, it is not clear how stimulation of coagulation onset by inactivating TFPI will affect spatial and temporal clot propagation. Objective: To examine the BAX499 effect on clotting in a spatial, reaction‐diffusion experimental system in comparison with that of recombinant activated factor VII (rVIIa). Methods: Clotting in plasma activated by immobilized tissue factor (TF) was monitored by videomicroscopy. Results: BAX499 dose‐dependently improved coagulation in normal and hemophilia A plasma activated with TF at 2 pmole m^?2 by shortening lag time and increasing clot size by up to ～2‐fold. The effect was TFPI specific as confirmed by experiments in TFPI‐depleted plasma with or without TFPI supplementation. Clotting improvement was half‐maximal at 0.7 nm of BAX499 and reached a plateau at 10 nm, remaining there at concentrations up to 1000 nm . The BAX499 effect decreased with TF surface density increase. RVIIa improved clotting in hemophilia A plasma activated with TF at 2 or 20 pmole m^?2, both by shortening lag time and increasing spatial velocity of clot propagation; its effects were strongly concentration dependent. Conclusions: BAX499 significantly improves spatial coagulation by inhibiting TFPI in a spatially localized manner that is different to that observed with rVIIa.