title: Mutations affecting the stability of the haemagglutinin molecule impair the immunogenicity of live attenuated H3N2 intranasal influenza vaccine candidates lacking NS1.
abstract: The isolation and cultivation of human influenza viruses in embryonated hen eggs or cell lines often leads to amino acid substitutions in the haemagglutinin (HA) molecule. We found that the propagation of influenza A H3N2 viruses on Vero cells may trigger the appearance of HA destabilising mutations, affecting viral resistance to low pH or high temperature treatment. Two ΔNS1 reassortants, containing the HA sequences identical to the original human H3N2 influenza virus isolates were constructed. Passages of these viruses on Vero cells led to the appearance of single mutations in the HA(1) L194P or HA(2) G75R subunits that impaired virus stability. The original HA sequences and the stable phenotypes of the primary isolates were preserved if reassortants were passaged by infection at pH 5.6 and cultivation in medium at pH 6.5. Corresponding ΔNS1 reassortants were compared for their immunogenicity in ferrets upon intranasal immunisation. Vaccine candidates containing HA mutations demonstrated significantly lower immunogenicity compared to those without mutations. Thus, the retaining of the original HA sequences of human viruses during vaccine production might be crucial for the efficacy of live attenuated influenza vaccines.
authors.affiliation: Avir Green Hills Biotechnology AG, Forsthausgasse 11, A-1200 Vienna, Austria.
authors.forename: Sabine
authors.initials: S
authors.lastname: Nakowitsch
authors.forename: Markus
authors.initials: M
authors.lastname: Wolschek
authors.forename: Alexander
authors.initials: A
authors.lastname: Morokutti
authors.forename: Tanja
authors.initials: T
authors.lastname: Ruthsatz
authors.forename: Brigitte M
authors.initials: BM
authors.lastname: Krenn
authors.forename: Boris
authors.initials: B
authors.lastname: Ferko
authors.forename: Nicole
authors.initials: N
authors.lastname: Ferstl
authors.forename: Andrea
authors.initials: A
authors.lastname: Triendl
authors.forename: Thomas
authors.initials: T
authors.lastname: Muster
authors.forename: Andrej
authors.initials: A
authors.lastname: Egorov
authors.forename: Julia
authors.initials: J
authors.lastname: Romanova
mesh_terms: D000281:Administration, Intranasal; D000818:Animals; D057134:Antibodies, Neutralizing / Q000097:blood; D000914:Antibodies, Viral / Q000097:blood; D002460:Cell Line; D002522:Chlorocebus aethiops; D005260:Female; D005289:Ferrets; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics* / Q000276:immunology; D006863:Hydrogen-Ion Concentration; D056724:Immunity, Humoral; D007114:Immunization; D053122:Influenza A Virus, H3N2 Subtype / Q000235:genetics* / Q000276:immunology; D007252:Influenza Vaccines / Q000235:genetics / Q000276:immunology; D008297:Male; D009154:Mutation; D016865:Reassortant Viruses / Q000235:genetics / Q000276:immunology; D014709:Vero Cells
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D007252:Influenza Vaccines