Over the past 20 years, emerging coronaviruses from animal reservoirs have caused severe and lethal diseases in humans – including SARS–CoV, MERS and SARS–CoV–2, which causes COVID–19. Early in the COVID–19 pandemic, researchers turned to novel and repurposed antivirals to search for effective new treatments for the disease.
Remdesivir, originally developed for the treatment of Hepatitis C and Ebola, has now been approved for treating patients with severe COVID–19. But this drug must be given to patients intravenously – requiring hospitalisation – and it is difficult to manufacture and expensive. Researchers are continuing the hunt for other antiviral medicines for treating COVID–19.
Molnupiravir (MPV), is an oral antiviral drug currently in clinical trials for the treatment of COVID–19. Taken as a prodrug, MPV is converted to its active form in the cell, ß–d–N4–hydroxycytidine (NHC). MPV was one of the first drugs selected for testing on the Accelerating COVID–19 Drug Development – Phase I/II platform (AGILE), which was launched early in the pandemic to fast–track the evaluation of potential new treatments for the disease.
Understanding how a drug is absorbed, distributed, metabolised and eliminated from the body is an important part of early-stage clinical trials. These clinical pharmacokinetics studies require analytical methods that can accurately measure the concentration of target analytes in the body.
The team extracted the analytes from samples by protein precipitation, followed by chromatographic separation. They then conducted analyte detection in negative ionisation mode, performing the analysis using stable isotopically labelled internal standards.
Their new method was validated according to the European Medicines Agency (EMA) and US Food and Drug Administration (FDA) guidelines over a linear range of 2.5–5000 ng/ml for both plasma and saliva. Across four different concentrations, precision and accuracy were 15% – and the recovery of both analytes from plasma and saliva was between 95% and 100% and 65–86% respectively.
The researchers present a sensitive, selective, accurate and robust new LC–MS/MS method that is validated to quantify MPV and NPC in human plasma and saliva.
This bespoke method, which was devised for the AGILE trial, will help provide greater clarity about the clinical pharmacokinetics of MPV and NPC under different treatment scenarios – forming the basis of selecting the most effective dose regimen.
Additionally, this assay can serve as the foundation method for other trials involving the same analytes.
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1) Amara, A.A. et al. The development and validation of a novel LC-MS/MS method for the simultaneous quantification of Molnupiravir and its metabolite ß–d–N4–hydroxycytidine in human plasma and saliva. J Pharm Biomed Anal. 2021;206:114356
Dr Alison Halliday
After completing an undergraduate degree in Biochemistry & Genetics at Sheffield University, Alison was awarded a PhD in Human Molecular Genetics at the University of Newcastle. She carried out five years as a Senior Postdoctoral Research Fellow at UCL, investigating the genes involved in childhood obesity syndrome. Moving into science communications, she spent ten years at Cancer Research UK engaging the public about the charity’s work. She now specialises in writing about research across the life sciences, medicine and health.