Consensus H3N2 HA immunogens incorporating key antigenic mutations elicit broad-spectrum neutralizing responses.

Seasonal influenza A/H3N2 poses a significant public health burden due to its rapid antigenic evolution, necessitating frequent vaccine updates. This study systematically analysed the antigenic evolution of 14 H3N2 vaccine strains (2000-2022) using pseudovirus-based neutralization assays and antigenic cartography. Vaccine strains exhibited cross-neutralization patterns, with higher reactivity among strains from adjacent years, forming six antigenic clusters. Key amino acid mutations (e.g. 144K, 189N) were identified as critical drivers of antigenic variation, with clustering-specific effects on neutralization sensitivity. Based on these findings, a consensus HA immunogen (consensus up-HA) was designed by combining mutations at key sites. This immunogen demonstrated broad-spectrum neutralization against pseudoviruses representing circulating strains from 2011 to 2024, including recent variants. These results highlight the potential of targeted HA modifications to develop broadly cross-reactive H3N2 vaccine candidate, offering broader and more durable neutralizing activity against evolving H3N2 strains.