ISCOMs/MPLA‐Adjuvanted SDAD Protein Nanoparticles Induce Improved Mucosal Immune Responses and Cross‐Protection in Mice

The epidemics caused by the influenza virus are a serious threat to public health and the economy. Adding appropriate adjuvants to improve immunogenicity and finding effective mucosal vaccines to combat respiratory infection at the portal of virus entry are important strategies to boost protection. In this study, a novel type of core/shell protein nanoparticle consisting of influenza nucleoprotein (NP) as the core and NA1‐M2e or NA2‐M2e fusion proteins as the coating antigens by SDAD hetero‐bifunctional crosslinking is exploited. Immune‐stimulating complexes (ISCOMs)/monophosphoryl lipid A (MPLA) adjuvants further boost the NP/NA‐M2e SDAD protein nanoparticle‐induced immune responses when administered intramuscularly. The ISCOMs/MPLA‐adjuvanted protein nanoparticles are delivered through the intranasal route to validate the application as mucosal vaccines. ISCOMs/MPLA‐adjuvanted nanoparticles induce significantly strengthened antigen‐specific antibody responses, cytokine‐secreting splenocytes in the systemic compartment, and higher levels of antigen‐specific IgA and IgG in the local mucosa. Meanwhile, significantly expanded lung resident memory (RM) T and B cells (TRM/BRM) and alveolar macrophages population are observed in ISCOMs/MPLA‐adjuvanted nanoparticle‐immunized mice with a 100% survival rate after homogeneous and heterogeneous H3N2 viral challenges. Taken together, ISCOMs/MPLA‐adjuvanted protein nanoparticles could improve strong systemic and mucosal immune responses conferring protection in different immunization routes. In this study, novel protein nanoparticles are generated by conjugating the influenza M2e‐NA fusion protein onto influenza nucleoprotein nanoparticle cores using a hetero‐bifunctional crosslinker SDAD (NHS‐SS‐Diazirine). The resulting protein nanoparticles, when formulated with ISCOMs/monophosphoryl lipid A adjuvants, exhibit significantly improved immune responses in both systemic and local mucosal compartments. These outcomes position this formulation as a promising mucosal vaccine candidate.