Covalently linking heparin to antithrombin enhances prothrombinase inhibition on activated platelets

Ivan Stevic; Howard H. W. Chan; Ankush Chander; Leslie R. Berry; Anthony K. C. Chan

doi:10.1160/TH12-10-0766

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2013; 109(06): 1016-1024
DOI: 10.1160/TH12-10-0766

Blood Coagulation, Fibrinolysis and Cellular Haemostasis

Schattauer GmbH

Covalently linking heparin to antithrombin enhances prothrombinase inhibition on activated platelets

Ivan Stevic

¹Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada

³Department of Pediatrics, McMaster University, Hamilton, Ontario, Canada

,

Howard H. W. Chan

¹Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada

²Department of Medicine, McMaster University, Hamilton, Ontario, Canada

,

Ankush Chander

¹Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada

³Department of Pediatrics, McMaster University, Hamilton, Ontario, Canada

,

Leslie R. Berry

¹Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada

³Department of Pediatrics, McMaster University, Hamilton, Ontario, Canada

,

Anthony K. C. Chan

¹Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada

³Department of Pediatrics, McMaster University, Hamilton, Ontario, Canada

› Author Affiliations

Abstract

Summary

Factor (F)Xa within the prothrombinase complex is protected from inhibition by unfractionated heparin (UFH), enoxaparin and fondaparinux. We have developed a covalent antithrombin-heparin complex (ATH) with enhanced anticoagulant activity. We have also demonstrated that ATH is superior at inhibiting coagulation factors when assembled on artificial surfaces. The objective of the present study is to determine the ability of ATH vs AT+UFH to inhibit FXa within the prothrombinase complex when the enzyme complex is assembled on the more native platelet system. Discontinuous inhibition assays were performed to determine final k ₂-values for inhibition of FXa, FXa within the platelet-prothrombinase, or FXa within prothrombinase devoid of various components. Thrombin generation and plasma clotting was also assayed in the presence of resting/activated platelets ± inhibitors. Protection of FXa was not observed for ATH, whereas a moderate 60% protection was observed for AT+UFH. ATH inhibited platelet-prothrombinase ∼4-fold faster than AT+UFH. Relative to intact prothrombinase, rates for FXa inhibition by AT+UFH in prothrombinase complexes devoid of either prothrombin (II)/activated platelets/FVa were higher. However, inhibition by AT+UFH of prothrombinase devoid of FII yielded slightly lower rates compared to free FXa inhibition. Thrombin generation and plasma clotting was enhanced with activated platelets, while inhibition was better by ATH compared to AT+UFH, thus suggesting an overall enhanced anticoagulant activity of ATH against platelet-bound prothrombinase complexes.

Keywords

Coagulation - covalent antithrombin-heparin - heparin - platelets - prothrombinase complex

Full Text

References

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