Design, synthesis and pharmacological evaluation of phenylthiophene-2-carboxamide-based RSV fusion protein inhibitors

Respiratory syncytial virus (RSV) remains a significant cause of serious lower respiratory tract infection, highlighting the urgent need for new antivirals. A series of amide bond containing fluorinated benzimidazole derivatives was synthesized, and substitution of the N-phenylcarboxamide moiety with five-membered heteroaryl groups led to enhanced potency, with 2- and 3-halogenated thiophene rings significantly improved anti-RSV activity. Structural optimization afforded compound 21, bearing a 3-bromo-N-phenylthiophene-2-carboxamide moiety, which exhibited subnanomolar potency against RSV (IC₅₀ = 0.10 nM). Molecular docking studies suggest compound 21 forms stable hydrogen bonds and π-π stacking interactions with the pre-fusion F protein, providing a mechanistic basis for its activity. Compound 21 dose-dependently suppressed RSV F protein expression in HEp-2 cells and significantly reduced viral burden in the lungs in an RSV-infected murine model. These results identify compound 21 as a highly potent RSV fusion inhibitor and a promising lead for further antiviral development.