A SOLUBLE TISSUE FACTOR MUTANT IS A SELECTIVE ANTICOAGULANT AND ANTITHROMBOTIC AGENT

One approach to developing safer and more efficacious agents for the t reatment of thrombotic disease involves the design and testing of inhi bitors that block specific steps in the coagulation cascade. We descri be here the development of a mutant of human tissue factor (TF) as a s pecific antagonist of the extrinsic pathway of blood coagulation and t he testing of this mutant in a rabbit model of arterial thrombosis, Al anine substitutions of Lys residues 165 and 166 in human TF have been shown previously to diminish the cofactor function of TF in support of factor X (FX) activation catalyzed by factor VIIa (FVIIa), The K165A: K166A mutations have been incorporated into soluble TF (sTF; residues 1-219) to generate the molecule ''hTFAA,'' hTFAA binds FVIIa with kine tics and affinity equivalent to wild-type sTF, but the hTFAA FVIIa com plex shows a 34-fold reduction in catalytic efficiency for FX activati on relative to the activity measured for sTF FVIIa, hTFAA inhibits the activation of FX catalyzed by the complex formed between FVIIa and re lipidated TF(1-243), hTFAA prolongs prothrombin time (PT) determined w ith human plasma and relipidated TF(1-243) or membrane bound TF, and h as no effect on activated partial thromboplastin time, but is 70-fold less potent as an inhibitor of PT with rabbit plasma. The rabbit homol ogue of this mutant (''rTFAA'') was produced and shown to have greater potency with rabbit plasma, Both hTFAA and rTFAA display an antithrom botic effect in a rabbit model of arterial thrombosis with rTFAA givin g full efficacy at a lower dose than hTFAA, Compared to heparin doses of equal antithrombotic potential, hTFAA and rTFAA cause less bleeding as judged by measurements of the cuticle bleeding time. These results indicate that TF FVIIa is a good target for the development of new an ticoagulant drugs for the treatment of thrombotic disease. (C) 1997 by The American Society of Hematology.