title: Incorporation of membrane-anchored flagellin into influenza virus-like particles enhances the breadth of immune responses.
abstract: We have designed a membrane-anchored form of the Toll-like receptor 5 ligand flagellin, the major proinflammatory determinant of enteropathogenic Salmonella, which was found to be glycosylated and expressed on cell surfaces. A chimeric influenza virus-like particle (cVLP) vaccine candidate containing A/PR8/34 (H(1)N(1)) hemagglutinin (HA), matrix protein (M1), and the modified flagellin as a molecular adjuvant was produced. The immunogenicity, including the serum antibody levels and cellular immune responses, and the protective efficacy against homologous and heterologous live virus challenge of the resulting VLPs were tested after intramuscular administration in a mouse model. The results demonstrated that flagellin-containing VLPs elicited higher specific immunoglobulin G (IgG) responses than standard HA and M1 VLPs, indicating the adjuvant effect of flagellin. Enhanced IgG2a and IgG2b but not IgG1 responses were observed with flagellin-containing VLPs, illuminating the activation of Th1 class immunity. The adjuvant effects of flagellin were also reflected by enhanced specific cellular responses revealed by the secretion of cytokines by freshly isolated splenocyte cultures when stimulated with pools of major histocompatibility complex class I or II peptides. When immunized mice were challenged with homologous live PR8 virus, complete protection was observed for both the standard and cVLP groups. However, when a heterosubtypic A/Philippines (H(3)N(2)) virus was used for challenge, all of the standard VLP group lost at least 25% of body weight, reaching the experimental endpoint. In contrast, for the cVLP group, 67% of mice survived the challenge infection. These results reveal that cVLPs designed by incorporating flagellin as a membrane-anchored adjuvant induce enhanced cross-protective heterosubtypic immune responses. They also indicate that such cVLP vaccines are a promising new approach for protection against pandemic influenza viruses.
authors.affiliation: Department of Microbiology and Immunology and Emory Vaccine Center, Emory University School of Medicine, 1510 Clifton Road, Atlanta, Georgia 30322, USA.
authors.forename: Bao-Zhong
authors.initials: BZ
authors.lastname: Wang
authors.forename: Fu-Shi
authors.initials: FS
authors.lastname: Quan
authors.forename: Sang-Moo
authors.initials: SM
authors.lastname: Kang
authors.forename: Jadranka
authors.initials: J
authors.lastname: Bozja
authors.forename: Ioanna
authors.initials: I
authors.lastname: Skountzou
authors.forename: Richard W
authors.initials: RW
authors.lastname: Compans
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis; D002460:Cell Line; D005260:Female; D005408:Flagellin / Q000235:genetics / Q000276:immunology*; D006031:Glycosylation; D006385:Hemagglutination Inhibition Tests; D007114:Immunization; D007252:Influenza Vaccines / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D018411:Spodoptera; D018417:Th1 Cells / Q000276:immunology; D018418:Th2 Cells / Q000276:immunology; D051198:Toll-Like Receptor 5 / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis; D014771:Virion / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D007252:Influenza Vaccines; D051198:Toll-Like Receptor 5; D014409:Tumor Necrosis Factor-alpha; D005408:Flagellin
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