达比加群抗凝机理的光谱研究

doi:10.3964/j.issn.1000-0593(2024)03-0699-08

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摘要：达比加群酯（DE）是一种新型口服抗凝药，用于预防非瓣膜性房颤患者的卒中和全身性血管栓塞。达比加群酯本身没有药理活性，其活性成分是在血浆和肝脏中经酯酶催化水解生成的达比加群（DAB）。尽管已经被用于临床治疗，活性成分达比加群的极性基团苯甲脒和羧基在其与凝血酶（Thr）结合时的作用机理仍不清晰，并且目前还未找到针对达比加群的特定逆转剂。采用稳态和瞬态荧光光谱、分子对接模拟等方法研究了DAB与凝血酶在模拟生理条件下的相互作用。通过研究极性基团酯化前后DAB与凝血酶作用时的时间分辨荧光光谱，揭示了DAB对凝血酶的荧光猝灭包含动态猝灭和静态猝灭的双重机制，指出DAB苯甲脒基团与凝血酶之间的静电导向效应是DAB与凝血酶快速有效形成复合物的关键动力学因素。运用分子对接模拟方法研究了DAB与凝血酶相互作用时的分子构象，探究DAB极性基团的酯化对两者结合能的影响。通过比较DAB双极性基团酯化后的达比加群酯（DE）、DAB羧酸基团酯化后的达比加群乙酯（DAE）、以及DAB苯甲醚基团酯化后的达比加群己酯（DAH）与凝血酶相互作用的荧光光谱和分子对接模拟结果，进一步验证了极性基团在DAB与凝血酶结合时的重要作用。获得了DAB和DE与牛血清白蛋白（BSA）在生理pH条件下相互作用的稳态和瞬态荧光光谱的对照结果，提出DAB的极性基团在其与凝血酶的选择性结合中发挥重要作用。研究结果为提高达比加群药物生理活性以及逆转剂的研究提供了理论和实验依据。

关键词：达比加群；抗凝机理；荧光光谱；静电导向效应

Abstract：Dabigatran etexilate (DE) is a new oral anticoagulant drug used to prevent non valvular atrial fibrillation stroke and vascular embolism. DE itself has no pharmacological activity. Its active component is Dabigatran (DAB), the product of DE by catalyzed hydrolysis in plasma and liver. However, the interaction mechanism between the polar groups of DAB and thrombin is still unclear, especially in physiological conditions, although the drug has already been used for clinical treatment. No specific reversal agent has been found for dabigatran.This work used steady-state and time-resolved fluorescence spectroscopy methods to study the interaction between DAB and thrombin in pH 7.4 phosphate buffer. A combination of dynamic and static fluorescence quenching of thrombin when in contact with DAB wasobserved, suggesting that the electrostatic effect between thrombin and the benzamidine group of DAB was the key factor of a fast and effective formation of the DAB-thrombin complex. The molecular conformation of DAB when in interaction with thrombin was studied by molecular docking simulation and the effect of DAB polar groups on the binding energy was investigated. By comparing the fluorescence spectra and molecular docking simulation results of the interaction between thrombin and dabigatran ester (DE, esterification of both DAB polar groups), dabigatran ethyl ester (DAE, esterification of DAB carboxylic group), and dabigatran hexyl ester (DAH, esterification of DAB benzamidine group), the role of polar groups in the combination of dabigatran and thrombin was further verified. Steady-state and transient fluorescence spectra of DAB and DE interacting with bovine serum albumin (BSA) under physiological pH conditions were also obtained,confirming that the polar groups of DAB play an important role in the selective binding of DAB with thrombin. The results provide theoretical and experimental bases for improving the drug efficiency and finding reversal agents.

Key words：Dabigatran; Anticoagulant Mechanism; Fluorescence spectroscopy; Electrostatic guidance

收稿日期: 2022-11-16 修订日期: 2023-07-24

中图分类号:

O643.12

基金资助: 国家自然科学基金项目（22173012，21773011）资助

通讯作者: 顾洪斌，祖莉莉 E-mail: hongbingu@hotmail.com; zull@bnu.edu.cn

作者简介: 龚梦洁，女，1997年生，北京师范大学化学学院硕士研究生 e-mail: Monica_G@mail.bnu.edu.cn

引用本文:

龚梦洁，海滢，吕凯文，顾洪斌，祖莉莉. 达比加群抗凝机理的光谱研究[J]. 光谱学与光谱分析, 2024, 44(03): 699-706.
GONG Meng-jie, HAI Ying, LÜ Kai-wen, GU Hong-bin, ZU Li-li. Spectral Study on Anticoagulation Mechanism of Dabigatran. SPECTROSCOPY AND SPECTRAL ANALYSIS, 2024, 44(03): 699-706.

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[1] Salem J, Sabouret P, Funck-Brentano C, et al. Fundamental & Clinical Pharmacology, 2015, 29(1): 10.
[2] Shnayder N A, Petrova M M, Shesternya P A, et al. Biomedicines, 2021, 9(5): 451.
[3] Diener H C, Sacco R L, Easton J D, et al. New England Journal of Medicine, 2019, 380(20): 1906.
[4] Ruff C T, Giugliano R P, Braunwald E, et al. The Lancet, 2014, 383(9921): 955.
[5] Hauel N H, Nar H, Priepke H, et al. Journal of Medicinal Chemistry, 2002, 45: 1757.
[6] Laizure S C, Parker R B, Herring V L, et al. Drug Metabolism and Disposition, 2014, 42(2): 201.
[7] Levy J H, Douketis J, Weitz J I, Nature Reviews Cardiology, 2018, 15: 273.
[8] Yang H, Liu Q, Gao X, et al. European Journal of Medicinal Chemistry, 2017, 126: 799.
[9] Naik R S, Pawar S K, Tandel R D, et al. Journal of Molecular Liquids, 2018, 251: 119.
[10] Michaelis S, Marais A, Schrey A K, et al. Journal of Medicinal Chemistry, 2012, 55: 3934.
[11] Remko M, Broer R, Remková A, RSC Advances, 2014, 4: 8072.
[12] Fuglestad B, Gasper P M, Tonelli M, et al. Biophysical Journal, 2012, 103: 79.
[13] Meng X, Zhang H, Mezei M, et al. Current Computer-Aided Drug Design, 2011, 7(2): 146.
[14] Wang Z, Sun H, Yao X, et al. Physical Chemistry Chemical Physics, 2016, 18(18): 12964.
[15] Burley S K, Bhikadiya C, Bi C, et al. Nucleic Acids Research, 2023, 51(D1): D488.
[16] Frisch M J, Trucks G W, Schlegel H B, et al. Gaussian 16 Rev. A. 03, Gaussian, Inc., Wallingford, CT, 2016.
[17] Morris G M, Huey R, Lindstrom W, et al. Journal of Computational Chemistry, 2009, 30(16): 2785.
[18] Trott O, Olson A J. Journal of Computational Chemistry, 2010, 31(2): 455.
[19] Baerga-Ortiz A, Rezaie A R, Komives E A. Journal of Molecular Biology, 2000, 296: 651.
[20] Borisevich N, Loznikova S, Sukhodola A, et al. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2013, 104: 158.
[21] Huntington J A. Thrombosis and Haemostasis, 2014, 111: 583.
[22] Coburn C A. Expert Opinion on Therapeutic Patents, 2001, 11: 721.
[23] Schwienhorst A. Cellular and Molecular Life Sciences, 2006, 63: 2773.
[24] Simone G De, Pasquadibisceglie A, Masi A di, et al. Journal of Molecular Recognition, 2021, 34(3): e2877.