一种用于细胞内定量瑞德西韦活性三磷酸代谢物GS-443902的新型高效液相色谱-串联质谱法。
A Novel HPLC-MS/MS Method for the Intracellular Quantification of the Active Triphosphate Metabolite of Remdesivir: GS-443902.
作者信息
Palermiti Alice, De Nicolò Amedeo, Antonucci Miriam, Soloperto Sara, Billi Martina, Manca Alessandra, Cusato Jessica, Menegatti Giorgia, Lamorde Mohammed, Calcagno Andrea, Waitt Catriona, D'Avolio Antonio
机构信息
Laboratory of Clinical Pharmacology and Pharmacogenetics, Department of Medical Sciences, University of Turin, Amedeo di Savoia Hospital, 10149 Turin, Italy.
ASL Città di Torino, Amedeo di Savoia Hospital, 10149 Turin, Italy.
出版信息
J Xenobiot. 2025 Jul 3;15(4):107. doi: 10.3390/jox15040107.
BACKGROUND
Remdesivir (RDV) is a broad-spectrum antiviral prodrug, which is rapidly metabolized in vivo within cells to the pharmacologically active triphosphate metabolite, GS-443902. On the other hand, the dephosphorylated metabolite GS-441524 is the main form detected in plasma. RDV acts against RNA viruses, and it was the first antiviral drug to receive EMA and FDA approval for treating COVID-19. Nevertheless, its intracellular pharmacokinetics in real life are poorly explored, particularly due to technical challenges.
METHODS
The aim of this study was to validate an HPLC-MS/MS method for the direct quantification of GS-443902 in peripheral blood mononuclear cells (PBMCs) with a chromatographic separation of 15 min.
RESULTS
The method was validated following EMA and FDA guidelines in terms of sensitivity, specificity, accuracy, precision, matrix effect, recovery, carryover, and stability, and then applied to PBMC isolates from a small cohort of patients with severe COVID-19 who received RDV.
CONCLUSIONS
This work represents the first method for the direct quantification of GS-443902 in PBMCs, with possible future application to intracellular pharmacokinetic studies in different scenarios, such as new oral prodrugs or drug-drug interaction studies.
背景
瑞德西韦（RDV）是一种广谱抗病毒前药，在体内细胞内迅速代谢为具有药理活性的三磷酸代谢物GS-443902。另一方面，去磷酸化代谢物GS-441524是血浆中检测到的主要形式。RDV对RNA病毒起作用，它是首个获得欧洲药品管理局（EMA）和美国食品药品监督管理局（FDA）批准用于治疗新冠肺炎的抗病毒药物。然而，其在现实生活中的细胞内药代动力学尚未得到充分研究，尤其是由于技术挑战。
方法
本研究的目的是验证一种高效液相色谱-串联质谱（HPLC-MS/MS）方法，用于在外周血单核细胞（PBMC）中直接定量GS-443902，色谱分离时间为15分钟。
结果
该方法按照EMA和FDA指南在灵敏度、特异性、准确性、精密度、基质效应、回收率、残留和稳定性方面进行了验证，然后应用于一小群接受RDV治疗的重症新冠肺炎患者的PBMC分离物。
结论
这项工作代表了第一种在外周血单核细胞中直接定量GS-443902的方法，未来可能应用于不同场景下的细胞内药代动力学研究，如新的口服前药或药物相互作用研究。
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