Molecular basis for protection and cross-protection by human antibodies targeting the parainfluenza virus hemagglutinin-neuraminidase protein

Human parainfluenza viruses (PIVs) are a leading cause of respiratory illness, particularly in vulnerable populations, where infection can lead to severe disease. Despite their clinical impact, there are currently no licensed vaccines or effective antiviral treatments available. PIVs have two large surface proteins, the fusion and hemagglutinin-neuraminidase (HN) proteins, both of which are targets of neutralizing antibodies. In this study, we identified and characterized two human monoclonal antibodies (mAbs), 5217-2 and 5217-9, which bind recombinant PIV3 HN protein, recognize PIV3-infected cells, and neutralize in vitro . We determined the antigenic sites of the PIV3 HN-specific mAbs via biolayer interferometry and found mAb 5217-9 targets a previously defined neutralizing antigenic site, while mAb 5217-2 binds a distinct antigenic site, expanding the known antigenic landscape. To further understand the newly defined epitope, we determined a cryo-electron microscopy (cryo-EM) structure of mAb 5217-2 in complex with PIV3 HN, revealing an epitope adjacent to the PIV3 HN protein receptor binding site. We also determined the structure of the previously discovered anti-PIV3 HN mAb PIV3HN-09, previously shown to be partially protective in vivo . In a hamster challenge model of PIV3, mAb 5217-2 significantly reduced lung viral titers, demonstrating its protective capacity. Furthermore, as the mAb PIV3HN-05 was previously shown to cross-neutralize PIV1, we evaluated its protective efficacy in a PIV1 challenge model, demonstrating reduced lung viral titers. Overall, these findings provide new insights into PIV3 HN epitopes to support structure-based vaccine design and demonstrate protective mAbs for both PIV3 and PIV1.