Design of nanoparticulate group 2 influenza hemagglutinin stem antigens that activate unmutated ancestor B cell receptors of broadly neutralizing antibody lineages

Abstract

30Influenza vaccines targeting the highly-conserved stem of the hemagglutinin (HA) surface 31 glycoprotein have the potential to protect against pandemic and drifted seasonal influenza viruses 32 not covered by current vaccines. While HA stem-based immunogens derived from group 1 33 influenza A have been shown to induce intra-group heterosubtypic protection, HA stem-specific 34 antibody lineages originating from group 2 may be more likely to possess broad cross-group 35 reactivity. We report the structure-guided development of mammalian cell-expressed candidate 36 vaccine immunogens based on influenza A group 2 H3 and H7 HA stem trimers displayed on 37 self-assembling ferritin nanoparticles using an iterative, multipronged approach involving helix 38 stabilization, loop optimization, disulfide bond addition, and side chain repacking. These 39 immunogens were thermostable, formed uniform and symmetric nanoparticles, were recognized 40 by cross-group-reactive broadly neutralizing antibodies (bNAbs) with nanomolar affinity, and 41 elicited protective, homosubtypic antibodies in mice. Importantly, several immunogens were 42 able to activate B cells expressing inferred unmutated common ancestor (UCA) versions of 43 cross-group-reactive human bNAbs from two multi-donor classes, suggesting they could initiate 44 elicitation of these bNAbs in humans. 45 46 47 48 common ancestor (UCA) versions of human antibody lineages associated with cross-group 59 reactive, broadly neutralizing antibodies (bNAbs). Immunization of mice led to protection 60 against a lethal homosubtypic influenza challenge. These candidate vaccines are now being 61 manufactured for clinical evaluation. 62 63 Influenza continues to be a significant global health burden, typically resulting in about 65 500,000 deaths worldwide annually (1), even though the technology for conventional egg-grown, 66 whole inactivated influenza virus vaccines was developed more than 70 years ago. Constant 67 antigenic drift of the influenza virus hemagglutinin (HA) coupled with immunodominant strain-68 specific antibody responses directed to the variable HA head domain results in conventional 69 vaccine effectiveness ranging from 10-60% (2) and the need for seasonal updates of virus strains 70 included in licensed vaccines. Furthermore, current vaccine approaches provide minimal 71 protection against pandemic influenza strains (3, 4). Improved influenza vaccines would not be 72 produced in eggs, would be designed to induce broad immunity against future drifted and 73 pandemic strains without seasonal reformulation, and elicit durable immune responses avoiding 74 the need for annual vaccination (4). One approach for achieving broadly cross-reactive 75 influenza-specific antibodies is to target highly conserved neutralization-sensitive epitopes in the 76 stem region of the influenza A HA surface glycoprotein (5). Monoclonal antibodies have been 77 identified that bind conserved HA stem epitopes and possess broad neutralizing activity across 78 diverse HA subtypes within an infl...

Nucleoside-modified mRNA-based influenza vaccines circumvent problems associated with H3N2 vaccine strain egg-adaptation

“…ELISA plates were coated overnight at 4°C with streptavidin. The next day, plates were washed with PBS-T incubated for an hour with biotinylated headless H3 (15). Plates were washed with PBS-T and then blocked for an hour with TBS supplemented with 0.05% Tween and 1% BSA.…”

Generation of antigen-specific memory CD4 T cells by heterologous immunization enhances the magnitude of the germinal center response upon influenza infection

“…Tfh cells are mainly distinguished by expression of the B cell follicle homing receptor CXCR5 (22)(23)(24)(25) and the transcriptional repressor Bcl6, which is required for Tfh cell differentiation (26)(27)(28). While natural influenza infection and a number of novel vaccine strategies have been shown to induce increased protection and broadly neutralizing antibodies (10,15,(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39), the mechanisms or involvement of CD4+ T cells in GC reaction to generate those broadly neutralizing antibodies were not described.…”