Construction of an mRNA vaccine encoding hemagglutinin of influenza A H1N1 virus and investigation on booster immunization strategy

Abstract

Objective·To construct an mRNA vaccine encoding hemagglutinin (HA) of influenza A H1N1 virus, and explore the protective effects of different booster vaccination strategies.Methods·Firefly luciferase (Fluc) was used as the reporter gene to construct Fluc mRNA vaccine enveloped in lipid nanoparticles (LNP). The in vivo expression of Fluc mRNA-LNP after intramuscular injection was determined by live imaging assay in mice. Furthermore, M15-HA mRNA-LNP derived from H1N1 subtype (A/Michigan/45/2015) was constructed. Mice were immunized with 20, 10, 5, or 1 µg doses of M15-HA mRNA-LNP twice (with an interval of 3 weeks) through intramuscular injection. Serum antibody titers were measured by enzyme-linked immunosorbent assay (ELISA) at 2 weeks and 4 weeks after the second immunization, and functional antibody levels were detected by hemagglutination inhibition test. The third booster vaccination was performed 40 d after the second immunization in 1 µg dose group with 1 µg M15-HA mRNA-LNP or 10 µg HA subunit vaccine. The levels of specific antibody and functional antibody were detected by ELISA and hemagglutination inhibition test, respectively 2 weeks and 4 weeks later.Results·Live imaging assay showed that luciferase activity could be detected in mice 1 d after injection of Fluc mRNA-LNP. At 2 weeks and 4 weeks after the second immunization of M15-HA mRNA-LNP, HA-specific antibodies were significantly higher than those before the immunization in all vaccination groups at different doses (P=0.000). The hemagglutination inhibition test showed that the levels of functional antibodies in the 20 μg dose and 10 μg dose groups were significantly higher than those in the PBS control group (P<0.05). After 1 µg dose group mice were immunized with HA protein or M15-HA mRNA-LNP, higher levels of HA-specific antibody and functional antibody were induced and maintained for a long time. There was no significant difference between the two different booster immunization strategies.Conclusion·M15-HA mRNA-LNP vaccine is constructed with immunogenicity and antibody neutralization activity. Low-dose mRNA priming vaccination followed by both homologous mRNA vaccine and heterologous protein subunit vaccine booster vaccination can induce stronger immune recall responses.

Keywords

• influenza vaccine
• influenza a virus
• hemagglutinin
• mrna vaccine
• booster immunization