In vivo comparative analysis of live influenza vaccine candidates obtained by reverse genetics and classical reassortment

Difficulties identified in recent years while preparing reassortant live influenza vaccine (LAIV) strains related to the specific biological properties of currently circulating epidemic influenza viruses, necessitate the search for alternative development pathways that guarantee the rapid and stable production of LAIV candidates. It was shown previously that genetically engineered reassortant LAIV candidates (LAIV-RG) exhibited reduced cold adaptation in developing chicken embryos compared to the A/Leningrad/134/17/57 (H2N2) master donor virus and vaccine candidates obtained by classical reassortment (LAIV-NR). Concerns that reduced cold adaptation could negatively impact vaccine quality led to implement the objective: to conduct a comparative study of the humoral immune response in laboratory animals to immunization with RG- or NR-LAIV candidates. Materials and methods. In vivo studies involved pairs of LAIV-RG and LAIV-NR strains based on seasonal influenza A(H3N2) viruses and master donor virus for live influenza vaccine. Results. In mouse studies, LAIV-RG candidates demonstrated similar humoral immune response performance to classical reassortants. A slight reduction in the cold-adapted phenotype of LAIV-RG strains did not result in a decrease in their attenuation level. They are as safe for laboratory animals as classical reassortants suggesting that LAIV quality remains unchanged, regardless of the methods used to produce vaccine reassortants. Interestingly, the adaptive Thr-Ile amino acid substitution at position 203 of HA1, identified in three LAIV-NR A(H3N2) candidates belonging to clades 3C.2a1 and 3C.2a2, had a positive effect on increasing the thermal stability of hemagglutinin and their immunogenicity in guinea pigs. Conclusion. Classical and genetically engineered reassortment methods are equally applicable for producing attenuated LAIV candidates, and the quality of vaccine preparations may be improved by targeted introduction of well-characterized favorable mutations into the genome of LAIV-RG strains, such as the mutation in HA1 with the amino acid substitution Thr-203-Ile, which contributes to increased stability and immunogenicity of vaccine strains based on the influenza A(H3N2) viruses of clades 3C.2a1 and 3C.2a2. This should not conflict with regulatory requirements for LAIV.