Molnupiravir Metabolite--N4 -hydroxycytidine Causes Cytotoxicity and DNA Damage in Mammalian Cells in vitro
Keywords:
N4 -hydroxycytidine, Molnupiravir, Cytotoxicity, DNA amage, Comet assayAbstract
N4-hydroxycytidine (NHC) is the active metabolite of molnupiravir—a new drug for COVID-19 treatment. NHC exerts antiviral activity by incorporating into SAR-CoV-2 RNA leading to false base-paring and lethal mutations to the virus. However, the risk of non-specific mutagenesis to host cells has been a concerned. The goal of this study is to detect cytotoxic activity and DNA damage induced by NHC in rapid-growing cells including human keratinocyte (HaCaT), and human adenocarcinomic alveolar basal epithelial (A549) cells in vitro by using sulforhodamine B (SRB) colorimetric and comet assays. NHC induced cytotoxicity in a concentration-dependent manner (0.1-30µM) in HaCaT and A549 cells. Half-maximal inhibitory concentration (IC50) values of NHC were lower in HaCaT compared to A549 cells after 3, 5, 10 days of exposure (4.40±0.09 vs 23.21±3.42, 5.82±0.91 vs 16.35±2.04, and 5.41±0.88 vs 13.83±2.05 µM, respectively), suggesting that the cytotoxic effect of NHC is more potent in HaCaT cells than in A549 cells. Significant increase in DNA damage parameters were observed in comet assay for HaCaT and A549 cells after exposure to NHC. NHC-induced DNA damage in HaCaT cells was concentration-dependent (1-10µM), and time-dependent (3-10 days). NHC-induced DNA damage in A549 cells was concentration-dependent (1-10µM), but not time-dependent (3-10days). Within the limitations of this in vitro study, we conclude that NHC could induce cytotoxic and DNA damage in mammalian cells at therapeutic and supratherapeutic concentrations. We propose caution in the use and supervision of molnupiravir, especially in patients with impaired xenobiotic clearance.
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