Abstract

Factor Xa (fXa) inhibition has resulted in the emergence of a new class of antithrombotics. Pharmacodynamic monitoring of these agents has proven problematic. The present study was designed to

  • determine the target concentration of an oral fXa inhibitor required for clinical trials using both thrombin generation assays and three in vivo models and

  • determine whether clotting assays such as activated partial thromboplastin time (aPTT) and prothrombin time (PT) would be suitable for monitoring human dosing.

PRT54021 (PRT021) is a potent inhibitor of human fXa (Ki=117pM). PRT021 and fondaparinux, an indirect fXa inhibitor, both significantly inhibited TAT and F1.2 generation in human whole blood. Compared to a therapeutic level of fondaparinux (200nM), PRT021 (200nM) was more potent in suppressing both markers. Multiple doses of PRT021 were evaluated in three animal models. The first model, which measured clot accretion on cotton threads placed in rabbit abdominal vena cava, compared inhibition of thrombus mass by PRT021 to that of supratherapeutic doses of enoxaparin (a LMW heparin). The second model compared the ability of PRT021 to maintain vessel patency under arterial flow conditions in FeCl3 induced thrombosis in rat carotid artery to that achieved by enoxaparin or clopidogrel (an antiplatelet agent). The third model investigated inhibition of 111In labeled platelet deposition on dacron grafts and expansion chambers placed in femoral arteriovenous shunts in baboons. PRT021 and enoxaparin were administered as IV infusions and clopidogrel was dosed orally for three days. Ex vivo PT and aPTT were measured in all models. The models encompass stringent criteria of arterial and venous thrombosis and PRT021 produced dose-responsive antithrombotic activity in each of the three models. The efficacy of PRT021 compared favorably to supratherapeutic levels of enoxaparin and clopidogrel. Unlike in the rodent models, efficacy in primates was attained at a much lower dose with minimal prolongation of PT. Species specificity was also demonstrated by in vitro extensions of PT and aPTT in rat, rabbit, baboon and human plasma. A 2X change of PT was attained at concentrations of 8.9, 1.6, 1 and 0.4μM respectively. The data indicate that doses of PRT021 that inhibit thrombin generation in human blood and that provide anticoagulation similar to baboon dosed at 0.49mg/kg may be sufficient to prevent venous thrombosis in humans. Comparative modeling of extents of change in PT to levels of antithrombotic efficacy also leads us to predict that human therapeutic activity for PRT021 may be attained without concurrent changes in ex vivo clotting parameters. The targeted concentration is currently being tested in Phase II trials for its ability to prevent venous thromboembolism in orthopedic surgery patients.

Model of ThrombosisAgent, DoseAntithrombotic ActivityaPTT fold changePT fold change
Rabbit vena cava PRT021,3mg/kg 76% inhibition 2.22 2.34 
Rabbit vena cava Enoxaparin, 1.6mg/kg 96% inhibition 2.06 2.01 
Rat carotid PRT021,19.1mg/kg 90% patency 1.69 2.20 
Rat carotid Enoxaparin, 7.6mg/kg 70% patency 3.49 1.19 
Rat carotid Clopidogrel, 3mg/kg/day 80% patency 1.03 1.01 
Baboon arteriovenous PRT021,0.49mg/kg 90% inhibition (venous), 32% inhibition (arterial) 1.29 1.17 
Model of ThrombosisAgent, DoseAntithrombotic ActivityaPTT fold changePT fold change
Rabbit vena cava PRT021,3mg/kg 76% inhibition 2.22 2.34 
Rabbit vena cava Enoxaparin, 1.6mg/kg 96% inhibition 2.06 2.01 
Rat carotid PRT021,19.1mg/kg 90% patency 1.69 2.20 
Rat carotid Enoxaparin, 7.6mg/kg 70% patency 3.49 1.19 
Rat carotid Clopidogrel, 3mg/kg/day 80% patency 1.03 1.01 
Baboon arteriovenous PRT021,0.49mg/kg 90% inhibition (venous), 32% inhibition (arterial) 1.29 1.17 

Disclosures: Employees of Portola Pharmaceuticals.; Stock options in Portola Pharmaceuticals.; Stock options in Genentech, Scios.; Portola Pharmaceuticals provided research funding to OHSU.

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