Structure-Guided Improvement of a Dual HPIV3/RSV Fusion Inhibitor

Human parainfluenza virus 3 (HPIV3) and respiratory syncytial virus (RSV) are leading causes of lower respiratory tract infections. There are currently no vaccines or antiviral therapeutics to treat existing HPIV3 or RSV infections. We recently reported a peptide (VIQKI), derived from the C-terminal heptad repeat (HRC) domain of the HPIV3 fusion (F) glycoprotein that inhibits infection by both HPIV3 and RSV. The dual inhibitory activity of VIQKI is due to its unique ability to bind to the N-terminal heptad repeat (HRN) domains of both HPIV3 and RSV F, thereby preventing the native HRN-HRC interactions required for viral entry. Here we describe the structure-guided design of dual inhibitors of HPIV3 and RSV fusion with improved efficacy. We show that VIQKI derivatives possessing one (I456F) or two (I454F/I456F) phenylalanine substitutions near the N-terminus exhibit more stable assemblies with RSV HRN domain and enhanced antiviral efficacy against both HPIV3 and RSV infection. Co-crystal structures of the new Phe-substituted inhibitors co-assembled with HPIV3 or RSV HRN domains reveal that the I456F substitution makes intimate hydrophobic contact with the core trimers of both HPIV3 and RSV F.