Heparin-tissue Factor Pathway Inhibitor Complexes: Anticoagulant and Pharmacokinetic Properties

Abstract

Exogenous addition of full-length tissue factor pathway inhibitor (FL-TFPI) to plasma caused a greater pro longation of prothrombin time (PT) than activated partial thromboplastin time (APTT). In contrast, heparin elicited a greater prolongation of APTT than PT. These results suggest that FL-TFPI and heparin exert their anticoagulant activity pri marily through inhibition of the extrinsic and intrinsic path ways, respectively. Using a dilute thromboplastin-induced clot ting assay, it was found that FL-TFPI was ~37-fold more potent than the carboxyl terminus truncated form (CT-TFPI) in pro longing the clotting time, which indicated that the positively charged carboxyl terminus of FL-TFPI was crucial for its an ticoagulant activity. Both FL-TFPI and CT-TFPI could exert synergistic anticoagulant action with heparin when TFPIs and heparin were added sequentially to plasma. However, when FL-TFPI was complexed with heparin before addition to the plasma, the effect on anticoagulant activity was dependent on the weight ratio of heparin:FL-TFPI. Addition of the hepa rin:FL-TFPI complex at weight ratios <1.25:1 gave a dPT clot ting time shorter than that of addition of FL-TFPI alone sug gesting that neutralization of the positively charged carboxyl terminus of FL-TFPI by heparin could also decrease its anti coagulant activity. Addition of heparin:FL-TFPI complex at weight ratios ≥1.25:1 gave an additive or synergistic antico agulant effect compared to the individual anticoagulant effects of heparin and FL-TFPI. In contrast, addition of preformed N-acetyl heparin:FL-TFPI and low molecular weight hepa rin :FL-TFPI complexes in above weight ratios to plasma caused only inhibition of the anticoagulant activity of FL-TFPI. Pharmacokinetic studies of FL-TFPI and heparin:FL-TFPI complex were carried out in rabbits. Both pharmacokinetic data could be fitted into a biexponential clearance model. Full- length TFPI had a very short t1/2α (1.4 min) and a relatively long t1/2β (92 min) with AUCβ (92%) dominant. Heparin :FL- TFPI, in contrast, had a prolonged t1/2α (15 min) and a similar t1/2β (116 min) with AUCα (88%) dominant. The overall clear ance was ∼2.6-fold faster for FL-TFPI than heparin:FL-TFPI complex. Constant infusion studies confirmed that it was pos sible to achieve the same steady state level of TFPI in the circulating plasma by infusing 2.6-fold less complex than in fusion of the FL-TFPI alone.