Outflanking immunodominance to target subdominant broadly neutralizing epitopes.

A major obstacle to vaccination against antigenically variable viruses is skewing of antibody responses to variable immunodominant epitopes. For influenza virus hemagglutinin (HA), the immunodominance of the variable head impairs responses to the highly conserved stem. Here, we show that head immunodominance depends on the physical attachment of head to stem. Stem immunogenicity is enhanced by immunizing with stem-only constructs or by increasing local HA concentration in the draining lymph node. Surprisingly, coimmunization of full-length HA and stem alters stem-antibody class switching. Our findings delineate strategies for overcoming immunodominance, with important implications for human vaccination.

Protective efficacy of influenza group 2 hemagglutinin stem-fragment immunogen vaccines.

The stem of the influenza A virus hemagglutinin (HA) is highly conserved and represents an attractive target for a universal influenza vaccine. The 18 HA subtypes of influenza A are phylogenetically divided into two groups, and while protection with group 1 HA stem vaccines has been demonstrated in animal models, studies on group 2 stem vaccines are limited. Thus, we engineered group 2 HA stem-immunogen (SI) vaccines targeting the epitope for the broadly neutralizing monoclonal antibody CR9114 and evaluated vaccine efficacy in mice and ferrets. Immunization induced antibodies that bound to recombinant HA protein and viral particles, and competed with CR9114 for binding to the HA stem. Mice vaccinated with H3 and H7-SI were protected from lethal homologous challenge with X-79 (H3N2) or A/Anhui/1/2013 (H7N9), and displayed moderate heterologous protection. In ferrets, H7-SI vaccination did not significantly reduce weight loss or nasal wash titers after robust 107 TCID50 H7N9 virus challenge. Epitope mapping revealed ferrets developed lower titers of antibodies that bound a narrow range of HA stem epitopes compared to mice, and this likely explains the lower efficacy in ferrets. Collectively, these findings indicate that while group 2 SI vaccines show promise, their immunogenicity and efficacy are reduced in larger outbred species, and will have to be enhanced for successful translation to a universal vaccine.

Influenza hemagglutinin stem-fragment immunogen elicits broadly neutralizing antibodies and confers heterologous protection.

Influenza hemagglutinin (HA) is the primary target of the humoral response during infection/vaccination. Current influenza vaccines typically fail to elicit/boost broadly neutralizing antibodies (bnAbs), thereby limiting their efficacy. Although several bnAbs bind to the conserved stem domain of HA, focusing the immune response to this conserved stem in the presence of the immunodominant, variable head domain of HA is challenging. We report the design of a thermotolerant, disulfide-free, and trimeric HA stem-fragment immunogen which mimics the native, prefusion conformation of HA and binds conformation specific bnAbs with high affinity. The immunogen elicited bnAbs that neutralized highly divergent group 1 (H1 and H5 subtypes) and 2 (H3 subtype) influenza virus strains in vitro. Stem immunogens designed from unmatched, highly drifted influenza strains conferred robust protection against a lethal heterologous A/Puerto Rico/8/34 virus challenge in vivo. Soluble, bacterial expression of such designed immunogens allows for rapid scale-up during pandemic outbreaks.