A Novel HPLC-MS/MS Method for the Intracellular Quantification of the Active Triphosphate Metabolite of Remdesivir:
Alice Palermiti1, Amedeo De Nicolò1, Miriam Antonucci2
1Laboratory of Clinical Pharmacology and Pharmacogenetics, Department of Medical Sciences, University of Turin, Amedeo di Savoia Hospital, 10149 Turin, Italy.
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.
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