Lipid nanoparticles as adjuvant of Norovirus VLP vaccine augment cellular and humoral immune responses in a TLR 9- and type I IFN-dependent pathway

Norovirus (NoV) virus-like particles (VLPs) adjuvanted with aluminum hydroxide (Alum) are common vaccine candidates in clinical studies. Alum adjuvants usually inefficiently assist recombinant proteins to induce cellular immune responses. Thus, novel adjuvants are required to develop NoV vaccines that could induce both efficient humoral and robust cellular immune responses. Lipid nanoparticles (LNPs) are well-known mRNA delivery vehicles. Increasing evidence suggests that LNPs may have intrinsic adjuvant activity and can be used as adjuvants for recombinant protein vaccines; however, the underlying mechanism remains poorly understood. In this study, we compared the adjuvant effect of LNPs and Alum for a bivalent GI.1/GII.4 NoV VLP vaccine. Compared with Alum, LNP-adjuvanted vaccines induced earlier production of binding, blocking and neutralizing antibodies and promoted a more balanced IgG2a/IgG1 ratio. It is crucial that LNP-adjuvanted vaccines induced stronger Th1-type cytokine-producing CD4+ T cell and CD8+ T cell responses than Alum. The adjuvant activity of LNPs depended on the ionizable lipid components. Mechanistically, LNPs activated innate immune responses in a type I IFN-dependent manner and were partially dependent on Toll-like receptor (TLR) 9, thus affecting the adaptive immune responses of the vaccine. This conclusion was supported by RNA-seq analysis and in vitro cell experiments and by the deeply blunted T cell responses in IFNαR1−/− mice immunized with LNP-adjuvanted vaccines. This study not only identified LNPs as a high quality adjuvant for NoV VLP vaccines, but also clarified the underlying mechanism of LNPs as a potent immunostimulatory component for improving protein subunit vaccines.