Antibodies targeting a conserved cryptic epitope at the influenza hemagglutinin head-stem interface via distinct binding modes

Influenza A virus continues to pose pandemic threats, as demonstrated by the recent global spread of highly pathogenic H5N1 strains. Consequently, the development and characterization of vaccines targeting zoonotic influenza subtypes have become a major focus of the field. In this study, we characterize two broadly reactive antibodies previously isolated from H5N1 vaccines. Although they have distinct immunoglobulin germline gene usage and binding modes, cryo-EM analysis shows that both antibodies share a highly conserved cryptic epitope at the interface of the hemagglutinin (HA) head and stem domains. The antigenicity of this epitope is influenced by natural amino acid variation at residue 293 of HA subunit 1, and its access is facilitated by acid-induced conformational change of HA. Nevertheless, both antibodies are non-neutralizing and possess only marginal protective efficacy against seasonal and H5N1 viruses in vivo, thus representing suboptimal vaccine responses. Overall, our findings expand the interactions of cross-reactive antibodies with an under-characterized HA epitope and provide insights for next-generation influenza vaccine design.