Thromb Haemost 2006; 96(03): 317-324
DOI: 10.1160/TH06-04-0219
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Amplified anticoagulant activity of tissue factor-targeted thrombomodulin

In-vivo validation of a tissue factor-neutralizing antibody fused to soluble thrombomodulin
Yi-Xin Wang
1   Berlex Biosciences, Richmond, California, USA
,
Chenliang Wu
1   Berlex Biosciences, Richmond, California, USA
,
Jon Vincelette
1   Berlex Biosciences, Richmond, California, USA
,
Baby Martin-McNulty
1   Berlex Biosciences, Richmond, California, USA
,
Serene Alexander
1   Berlex Biosciences, Richmond, California, USA
,
Brent Larsen
1   Berlex Biosciences, Richmond, California, USA
,
David R. Light
1   Berlex Biosciences, Richmond, California, USA
,
Kirk McLean
1   Berlex Biosciences, Richmond, California, USA
› Author Affiliations
Further Information

Publication History

Received 21 April 2006

Accepted after revision 27 July 2006

Publication Date:
30 November 2017 (online)

Summary

Tissue factor (TF) exposure isa potent pro-thrombotic trigger that initiates activation of the coagulation cascade, while thrombomodulin (TM) is a potent anticoagulant protein that limits the extent of activation. Both TF neutralizing antibodies and soluble TM (sTM) are effective anticoagulants. We have developed a novel anticoagulant fusion protein, Ab(TF)-TM, by fusing a TF-neutralizing single-chain antibody, Ab(TF), to an active fragment of TM. Ab(TF)-TM is a novel anticoagulant targeting to sites of TF exposure with a dual mechanism of action. The Ab(TF) portion of the molecule inhibits TF/factor VIIa mediated activation of FIX and FX, and the TM portion of the molecule acts as a cofactor for activation of protein C. In-vitro coagulation assays show that Ab(TF)-TM more potently inhibits TF-initiated coagulation (prothrombin time) than can its individual components, Ab(TF) (20-fold) and sTM (80-fold) alone, or in combination (10-fold). In contrast, the potency of Ab(TF)-TM in the activated partial thromboplastin and thrombin clotting time assays was similar to sTM alone. Ina rat model of disseminated intravascular coagulation (DIC), intravenous injection of a human TF-containing thromboplastin reagent (0.5 ml/kg) resulted in an immediate death in ∼60% of the animals and a clinical score of ∼2.5. Pre-injection of Ab(TF)-TM or Ab(TF) and sTM, given alone or in combination, showed dose-dependent efficacy. At a dose of 0.7 nmol/kg, Ab(TF)-TM completely prevented death and reduced clinical scores by 79%, while neither Ab(TF) nor sTM, given alone or in combination, showed significant therapeutic effects. Calculated effective doses that reduced mortality by 50% relative to that in the control group (ED50, nmol/kg) were 0.21 for Ab(TF)-TM, 3.2 for an equimolar mixture of Ab(TF) and sTM, 4.3 for sTM and 20 for Ab(TF). Thus, Ab(TF)-TM presented 10– to 100-fold enhancement of the anticoagulant potency, relative to the ED50 in Ab(TF) and sTM given either alone or in combination, in a rat DIC model.