Publication: Selective disruption of heparin and antithrombin-mediated regulation of human factor IX
Issued Date
2015-01-01
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ISSN
15387836
15387933
15387933
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2-s2.0-84930178037
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Mahidol University
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SCOPUS
Bibliographic Citation
Journal of Thrombosis and Haemostasis. Vol.13, No.6 (2015), 1053-1063
Suggested Citation
P. R. Westmark, P. Tanratana, J. P. Sheehan Selective disruption of heparin and antithrombin-mediated regulation of human factor IX. Journal of Thrombosis and Haemostasis. Vol.13, No.6 (2015), 1053-1063. doi:10.1111/jth.12960 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/36698
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Title
Selective disruption of heparin and antithrombin-mediated regulation of human factor IX
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Abstract
© 2015 International Society on Thrombosis and Haemostasis. Interaction with antithrombin and heparin regulates distribution, activity, and clearance of factor IXa (FIXa). Hemophilia B prophylaxis targets plasma FIX levels > 1% but neglects extravascular FIX, which colocalizes with antithrombin-heparan sulfate. Objective: Combined mutagenesis of FIX was undertaken to selectively disrupt heparin- and antithrombin-mediated regulation of the protease. Methods: Human FIX alanine substitutions in the heparin (K126A and K132A) and antithrombin (R150A) exosites were characterized with regard to coagulant activity, plasma thrombin generation, antithrombin inhibition, and plasma half-life. Results: Single or combined (K126A/R150A or K132A/R150A) exosite mutations variably reduced coagulant activity relative to wild-type (WT) for FIX (27-91%) and FIXa (25-91%). Double mutation in the heparin exosite (K126A/K132A and K126A/K132A/R150A) markedly reduced coagulant activity (7-21%) and plasma TG. In contrast to coagulant activity, FIX K126A (1.8-fold), R150 (1.6-fold), and K132A/R150A (1.3-fold) supported increased tissue factor-initiated plasma TG, while FIX K132A and K126A/R150A were similar to WT. FIXa K126A/R150A and K132A/R150A (1.5-fold) demonstrated significantly increased FIXa-initiated TG, while FIXa K132A, R150A, and K126A (0.8-0.9-fold) were similar to WT. Dual mutations in the heparin exosite or combined mutations in both exosites synergistically reduced the inhibition rate for antithrombin-heparin. The half-life of FIXa WT in FIX-deficient plasma was remarkably lengthy (40.9 ±1.4 min) and further prolonged for FIXa R150A, K126A/R150A, and K132A/R150A (> 2 h). Conclusion: Selective disruption of exosite-mediated regulation by heparin and antithrombin can be achieved with preserved or enhanced thrombin generation capacity. These proteins should demonstrate enhanced therapeutic efficacy for hemophilia B.