Differential Neutralization of Apixaban, Betrixaban, Edoxaban, and Rivaroxaban by Andexanet Alfa as Measured by Whole Blood Thromboelastographic Analysis.

Original Manuscript Differential Neutralization of Apixaban, Betrixaban, Edoxaban, and Rivaroxaban by Andexanet Alfa as Measured by Whole Blood Thromboelastographic Analysis Clinical and Applied Thrombosis/Hemostasis Volume 28: 1-9 © The Author(s) 2022 Article reuse guidelines: sagepub.com/journals-permissions DOI: 10.1177/10760296221138297 journals.sagepub.com/home/cat Siddharth Mehrotra1, Debra Hoppensteadt, PhD1,2 , Walter Jeske, PhD1,2, Fakiha Siddiqui, BS2,3, Omer Iqbal, MD1, Alfonso Tafur, MD4,5, Bruce Lewis, MD5, Mark Jaradeh, BS1 , Bulent Kantarcioglu, MD1, and Jawed Fareed, PhD1,2 Abstract Introduction: The available oral anti-Xa agents are routinely used for the management of thrombotic disorders. A molecularly modified recombinant coagulation FXa, also known as Andexanet Alfa (AA), that has been developed as an antidote to neutralize the bleeding effects of oral FXa inhibitors, such as Apixaban and Rivaroxaban. Materials and Methods: This study utilized thromboelastography (TEG 5000 Hemostasis System), to investigate the neutralizing effects of AA at different concentrations of oral FXa inhibitors measuring such parameters as R-Time, K-Time, Angle, and Max Amplitude (MA). Apixaban, Betrixaban, Edoxaban, and Rivaroxaban were obtained commercially in powdered form. Each of these drugs was supplemented with freshly drawn whole citrated blood at a concentration of 1 μg/mL. And subsequently mixed with AA at 50 or 100 μg/mL. Results: At a concentration of 1 μg/mL, all FXa inhibitors produced variable anticoagulant effects in the order of Edoxaban > Betrixaban > Rivaroxaban > Apixaban. AA at 100 μg/mL produced a complete neutralization of these inhibitors whereas at 50 μg/ mL relatively weaker neutralization as measured by various parameters. Conclusion: These results suggest that regardless of the variable anticoagulant effects exhibited by the FXa Inhibitors, AA at FC = 100 μg/mL fully neutralized these agents as measured by the TEG parameters. AA was shown to be more effective in neutralizing Betrixaban and least effective in Apixaban. The neutralization of various FXa inhibitors was dose and donor-dependent warranting dosage adjustment for optimal outcomes. Keywords anti-Xa agents, anticoagulant, Andexanet Alfa Date received: 25 July 2022; revised: 7 October 2022; accepted: 26 October 2022. 1 Department of Molecular Pharmacology & Neuroscience, Cardiovascular Research Institute, Loyola University Chicago, Health Sciences Division, Maywood, IL, USA 2 Department of Pathology and Laboratory Medicine, Cardiovascular Research Institute, Loyola University Chicago, Health Sciences Division, Maywood, IL, USA 3 Program in Health Sciences, UCAM - Universidad Católica San Antonio de Murcia, Murcia, Spain 4 Department of Vascular Medicine, Northshore Cardiovascular Institute, NorthShore University Health Systems, Skokie, IL, USA 5 Department of Internal Medicine, Division of Cardiology, Loyola University Medical Center, Maywood, IL, USA Corresponding Author: Jawed Fareed, Department of Molecular Pharmacology & Neuroscience, Loyola University Health System, 2160 S First Avenue Bldg 115, Rm 432 Maywood, IL 60153, USA. Email: Creative Commons Non Commercial CC BY-NC: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 License (https://creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access page (https://us.sagepub.com/en-us/nam/open-access-at-sage). 2 Introduction Three oral anti-Xa agents are currently available for clinical use; Apixaban, Edoxaban, and Rivaroxaban, with a fourth agent, Betrixaban, approved but off market. These oral direct factor Xa (FXa) inhibitors are effective anticoagulants and aid in the prevention and treatment of thrombotic events such as thromboembolism. FXa inhibitors bind to FXa and prevent the conversion of prothrombin to thrombin.1 These FXa inhibitors are also being investigated for efficacy in the treatment of other conditions such as acute coronary syndrome, cancer-associated thrombosis, and peripheral arterial disease.2,3 Unlike other commonly used anticoagulants, such as heparins and heparin derivatives, FXa inhibitors can be taken orally and do not require any routine or episodic monitoring.4 However, similar to other anticoagulant options, these anti-Xa agents can cause uncontrolled and life-threatening bleeding.5 FXa inhibitors such as Rivaroxaban and Apixaban are now widely used in various indications where vitamin K inhibitors such as warfarin were indicated. Despite the widespread use of these agents the bleeding complications are frequently reported and require the use of such antidotes as the four-factor prothrombin concentrates and AA. The efficacy of AA in neutralizing FXa inhibitors is well documented however the dosing regimen widely varies.6,7 Therefore the circulating levels of these inhibitors may vary during the administration of AA. For this reason, two concentrations of AA were selected in these studies to neutralize the fixed concentration of various FXa inhibitors. Andexanet Alfa (AA, inactivated-zhzo, coagulation factor Xa) is a recombinant factor Xa protein developed by Portola Pharmaceuticals for use as a universal antidote for the anticoagulant effects of direct or indirect factor Xa inhibitors. Andexanet alfa acts as a decoy protein that binds to FXa inhibitors, neutralizing the anticoagulant effects of FXa inhibitors by preventing the inhibitors from binding to endogenous FXa.8 In this recombinant protein, the serine active site is replaced by an alanine residue to eliminate catalytic activity and the membrane-binding glutamic acid domain is removed to prevent the assemblage of the prothrombinase complex.9 Through accelerated approval, the United States Food and Drug Administration approved the use of AA as an antidote for the reversal of Apixaban and Rivaroxaban anticoagulation in the case of life-threatening or uncontrolled bleeding.8 Recently, studies have shown AA may also be effective in neutralizing the anticoagulant effects of Betrixaban and Edoxaban, in addition to other anticoagulants, such as enoxaparin and UFH, due to its highaffinity binding to both direct and indirect FXa inhibitors.10 While AA is not used or directed as an active procoagulant, its use has been reported to result in thromboembolic events.5,11 The purpose of this study was to investigate the anticoagulant effects of these FXa Inhibitors and their relative neutralization by AA. Additionally, AA was investigated for its potential of creating a hypercoagulable state. Materials and Methods The active pharmaceutical ingredient (API) forms of various oral anti-Xa agents including Apixaban, Betrixaban, Edoxaban, and Clinical and Applied Thrombosis/Hemostasis Rivaroxaban were commercially obtained. Working solutions of the (...truncated)