Development and in vitro Characterization of the IgA1 and IgG1 Isotypes of the Broadly Neutralizing Antibody FM08 Against Influenza A Virus

Rationale. According to the World Health Organization, seasonal influenza causes approximately 3 to 5 million cases of severe illness requiring hospitalization annually, with 290,000 to 650,000 of these cases resulting in death. Recombinant antibodies represent one of the most modern and promising classes of drugs for effective influenza therapy.

Objective. To evaluate the properties, specificity, and virus-neutralizing activity of the recombinant antibody FM08, targeting the stem domain of influenza virus hemagglutinin, in the IgA1 and IgG1 formats.

Methods. Expression plasmids were constructed using molecular genetic techniques. Experimental samples of FM08 antibody of IgA1 and IgG1 isotypes were produced by transient expression in HEK293 cell culture under serum-free conditions and subsequently purified. Biological activity was assessed comparatively using electrophoretic separation, ELISA, and a microneutralization assay. The study utilized influenza A virus strains of subtypes H1N1pdm, H2N2, H3N2, H5N1, as well as influenza B virus strains (from both genetic lineages).

Results. The genetic constructs were shown to enable the accumulation of recombinant FM08 antibodies of IgA1 and IgG1 isotypes in eukaryotic cell cultures. Both FM08 antibody isotypes exhibited nearly identical binding patterns and, starting from a concentration of 16 ng/µl, interacted with influenza A virus strains from both phylogenetic groups, with the exception of the H1N1 1934 strain. The 50% inhibitory concentration (IC₅₀) for binding was 1.49 µg/ml. FM08 antibodies of both IgA1 and IgG1 isotypes neutralized influenza A/California/07/09 (H1N1pdm09) virus with IC₅₀ values of 1–4 µg/ml and influenza A/Cambodia/e0826360/2020 (H3N2) virus with IC₅₀ values of 20 µg/ml.

Conclusion. The study of the in vitro biological activity of the obtained experimental FM08 antibody samples of IgA1 and IgG1 isotypes demonstrated their high potential as an antiviral agent.