Haemagglutinin substitutions N125D, D127E, D222G and R223Q improve replicative fitness and vaccine effectiveness of an A/H1N1pdm09 live attenuated influenza vaccine virus by enhancing α-2,6 receptor binding

• Published in: PLoS pathogens 2022-05, Vol.18 (5), p.e1010585-e1010585
• Main Authors: Dempsey, Rachael, Tamburrino, Giulia, Schewe, Katarzyna E, Crowe, Jonathan, Nuccitelli, Annalisa, Dibben, Oliver
• Format: Article
• Language: English
• Subjects: Animals; Antibodies, Viral; Antigens; Avidity; Binding sites; Biology and Life Sciences; Competition; Eggs; Epidemics; Epithelial cells; Epithelium; Ferrets; Fever; Fitness; HA protein; Hemagglutinins; Humans; Immunogenicity; Infections; Influenza; Influenza Vaccines; Influenza, Human; Medicine and Health Sciences; Mustela; Mutagenesis; Mutation; Pandemics; Proteins; Receptors; Replication; Research and Analysis Methods; Vaccine Efficacy; Vaccines; Vaccines, Attenuated; Viruses
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1010585

During 2013-14 and 2015-16, A/H1N1pdm09 live attenuated influenza vaccine (LAIV) viruses replicated inefficiently in primary human nasal epithelial cells (hNEC). This led to reduced vaccine effectiveness (VE) in quadrivalent formulations, mediated by inter-strain competition. By mutating the haemagglutinin (HA) protein, we aimed to enhance hNEC replication of a novel A/H1N1pdm09 vaccine strain to overcome competition and improve VE. Combinations of N125D, D127E, D222G and R223Q substitutions were introduced to the HA protein of A/Slovenia/2903/2015 (A/SLOV15). A/SLOV15 S13, containing all four HA substitutions, produced approximately 1000-fold more virus than parental V1 during hNEC infection. Immunogenicity in ferrets was increased by approximately 10-fold, without compromising yield in eggs or antigenic match to wild-type (wt) reference strains. Despite S13 and V1 being antigenically similar, only S13 protected ferrets from wt virus shedding and fever post-challenge. Crucially, these data suggested that enhanced fitness allowed S13 to overcome inter-strain competition in quadrivalent LAIV (QLAIV). This improved efficacy was later validated by real-world VE data. S13 displayed increased binding avidity to a mammalian-like α-2,6 receptor analogue (6-SLN), relative to V1, while maintaining avian-like 3-SLN avidity. In silico modelling of the HA receptor binding site revealed additional interactions in the S13:6-SLN binding network and a mild increase in 6-SLN binding energy, indicating a possible mechanism for increased α-2,6 receptor-binding avidity. These data confirm that rational HA mutagenesis can be used to optimise hNEC replication and VE for A/H1N1pdm09 LAIV viruses.