Objective Mechanism of β-d-N₄-hydroxycytidine (HNCEIDD-1931) of Molnupiravir Mol EIDD-2801）in inhibiting the highly mutable influenza A virus (IAV), which resisted to drugs and rendered vaccination ineffective, was investigated.

Method A549 cells were infected with A/Puerto Rico/8/1934 (H1N1) and treated with NHC in varied concentrations. RNA replication, protein expression, and progeny virus particle release of the virus were determined by qRT-PCR, Western blot, and plaque assay. Molnupiravir, the precursor drug of NHC, were administered to mice. Body weight, survival rate, lung tissue viral load, inflammatory factor expression, and pathological damage on the control and treatment mice were compared, and genome mutations of the virus sequenced.

Results NHC displayed in vitro as well as in vivo a dose-dependent inhibitory effect on IAV. Administration of Mol alleviated virus-induced inflammation and lung injury on mice. The genome mutations on the virus were mainly manifested as A-G and U-C transitions.

Conclusion NHC in Mol was highly effective against IAV. The accumulated specific mutations in genome induced by NHC contributed to the antiviral effect as observed, suggesting plausible application of the nucleoside analogue for combating the difficult virus.