Induction of enhanced stem-directed neutralizing antibodies by HA2-16 ferritin nanoparticles with H3 influenza virus boost†

Current seasonal influenza vaccines offer limited protection against influenza viruses due to genetic drift. The urgent need for a universal influenza vaccine to combat highly mutated strains is evident. This study utilized the conserved HA2 subunit of hemagglutinin (HA) and a short linear epitope of HA2 (HA2-16) from the H3 influenza virus to conjugate with ferritin, resulting in the construction of recombinant immunogens termed HA2-F and HA2-16-F, respectively. In vitro characterization confirmed the self-assembly of prokaryotically expressed HA2-F and HA2-16-F into nanoparticles (NPs). To simulate natural virus infection in the vaccinated population, intranasal infection with the whole H3N2 virus was administered as a final boost. Enhanced binding activity to A/Hong Kong/4801/2014 (H3N2) and A/17/California/2009/38 (H1N1) virus was detected in the HA2-16 group induced by the A/Wisconsin/67/2005 (H3N2) virus boost (Titer >10⁴). Furthermore, higher titers of neutralizing antibodies were elicited by HA2-16-F NP (ID₅₀: 50.4–631.0) compared to those by HA2-F NP (ID₅₀: 20.3–178.2). These results demonstrated that the H3N2 virus boost focused the antibody response on the HA2-16 epitope. Additionally, our immunization strategy was found to reduce serum ferritin reactive antibodies. In summary, HA2-16 not only holds promise as a vaccine candidate but also exhibits significant potential for influenza vaccine production, particularly in enhancing the levels of induced stem-directed antibodies. This study contributes to the development of recombinant immunogens for improved influenza vaccine efficacy.