Abstract
Influenza A virus (IAV) remains a significant global health threat due to its rapid mutation rate, seasonal epidemics, and pandemic potential. Continuous viral evolution and emerging resistance to existing antivirals necessitate the evaluation of alternative or repurposed antiviral strategies. Tenofovir alafenamide (TAF), a phosphonamidite prodrug of tenofovir primarily used in antiretroviral therapy, has demonstrated intracellular stability and a favorable safety profile. In this study, we investigated the time- and concentration-dependent antiviral activity and cytotoxicity of TAF in Influenza A–infected cell culture over a 120-h period. Antiviral efficacy was quantified as percentage inhibition of viral replication, while cytotoxicity was assessed using the MTT assay to determine cell viability. TAF exhibited progressive, exposure-dependent suppression of Influenza A replication, with inhibition exceeding 80% at higher concentrations after prolonged incubation. Importantly, host cell viability remained ≥70% under corresponding conditions, and the CC₅₀ was not reached within the tested concentration range. The estimated EC₅₀ values indicated significant antiviral activity with a favourable selectivity window. These findings suggest that TAF exerts time-dependent inhibitory effects on Influenza A virus while preserving cellular viability, supporting further investigation of nucleotide-analogue–based strategies for RNA virus replication control.