PF798 ANTITHROMBOTIC EFFECT OF SULFATED NON‐ANTICOAGULANT HEPARIN WITHOUT COMPROMIZING HEMOSTASIS
Authors: Mousa S. , Godugu K. , Darwish N. , Mousa S. 
Published: June 2019

Background:

The pathophysiology of thrombosis comprises a complex interplay of factors associated with vascular endothelial activation, platelet activation and adhesion. Blood coagulation process initiated by plasma protease factor VIIa in complex with the membrane protein tissue factor. Although heparin can be administered therapeutically to prevent and treat thrombosis, the effects of unfractionated heparin are more unpredictable, Low molecular weight heparin (LMWH) was recommended for prophylaxis and treatment of deep vein thrombosis as it significantly shortened both duration of thromboembolic crisis and hospitalization by ∼40%. Safety concerns associated with the narrow therapeutic index (bleeding risks) of LMWH are a major barrier to dose escalation/optimization of treatments. Here, we used the novel sulfated non‐anticoagulant LMWH, referred to as S‐NACH, which has a similar activity of LMWH but devoid of systemic anti‐factor Xa and IIa activity. Also, the sulfated form enhanced S‐NACH binding to the vascular endothelial cells and potentiate the action of tissue factor pathway inhibitors (TFPI‐1 and 2).

Aims:

To determine if S‐NACH exerts antithrombotic activity without compromizing hemostasis (bleeding) side effects that are associated with the clinical use of LMWHs. In the present study we investigated the effects of S‐NACH on clot kinetics in comparison to LMWH using thrombelstography (TAG) and examined its in vitro and in vivo effects on vascular endothelial release capacity of TFPI.

Methods:

We conducted both in vitro and in vivo experiments using our compound (S‐NACH) to test its efficacy on coagulation properties in comparison to LMWH (enoxaparin). We compared the effects on the coagulation time, time to clot initiation and thrombus strength level in vitro, with and without HUVEC endothelial cell line coated into cups used for the TEG analysis. TFPI‐1 and TFPI‐2 levels were also estimated in both in vitro and in vivo models at different doses of S‐NACH. In addition, the effect of S‐NACH on factors Xa and IIa was assessed in mice and rabbits.

Results:

TEG assay have shown that, increasing concentrations of S‐NACH (0.1 – 10 ug) did not affect the clotting time (R) and maximum amplitude of the clot formation (MA), while LMWH (enoxaparin) was associated with marked increased clotting time (R) and decreased maximum amplitude of the clot formation (MA) in comparison to control. Using HUVEC cell line coated cups; both S‐NACH and LMWH were associated with increased clotting time (R) and decreased maximum amplitude of the clot formation (MA) (Table 1). This is due to the ability of HUVEC cell to release of TFPI‐1 and TFPI‐2. In the in vivo study, administration of enoxaparin inhibited both factors Xa and IIa, while administration of S‐NACH did not affect the activities of these coagulation factors. However, S‐NACH caused 3 folds greater tissue factor pathway inhibitor (TFPI) release from human endothelial cells and in mice versus enoxaparin (Figure 1).

Summary/Conclusion:

S‐NACH is devoid of anti‐thrombin binding and inhibition of systemic anti‐thrombin‐dependent coagulation factors such as factor Xa and factor IIa but has an optimal releasing capacity of vascular endothelial TFPI that is associated with antithrombotic activities without any effect on hemostasis.

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