Genetically engineered live attenuated influenza A virus vaccine candidates

Abstract

We have generated new influenza A virus live attenuated vaccine candidates by site-directed mutagenesis and reverse genetics. By mutating specific amino acids in the PB2 polymerase subunit, two temperature-sensitive (ts) attenuated viruses were obtained. Both candidates have 38؇C shutoff temperatures in MDCK cells, are attenuated in the respiratory tracts of mice and ferrets, and have very low reactogenicity in ferrets. Infection of mice or ferrets with either mutant conferred significant protection from challenge with the homologous wild-type virus. Three tests for genetic stability were used to assess the propensity for reversion to virulence: 14 days of replication in nude mice, growth at 37؇C in tissue culture, and serial passage in ferrets. One candidate, which contains mutations intended to reduce the ability of PB2 to bind to cap structures, was stable in all three assays, whereas the second candidate, which contains mutations found only in other ts strains of influenza virus, lost its ts phenotype in the last two assays. This approach has therefore enabled the creation of live attenuated influenza A virus vaccine candidates suitable for human testing.

Hemagglutinin (HA) Proteins from H1 and H3 Serotypes of Influenza A Viruses Require Different Antigen Designs for the Induction of Optimal Protective Antibody Responses as Studied by Codon-Optimized HA DNA Vaccines

“…Both high-level HI and neutralizing activities were easily elicited by codon-optimized DNA vaccines. The titers of protective antibodies achieved in this report were not only significantly improved over that from early versions of flu DNA vaccines (2,9,18,22,50) but also comparable to or higher than those of positive anti-HA sera reported in previous animal studies with various flu vaccines, including recombinant viral vector-based flu HA vaccines (16,30,39) and inactivated and live flu vaccines (23,38,42).…”

Avian influenza virus: prospects for prevention and control by vaccination

“…However, with the advancement in biotechnology, it may be possible to consider new vaccine approaches using genetically engineered live virus. For example, several experiments showed the possibility of attenuating influenza virus by introducing changes in the PB2 gene, exchanging the promoter region of the NA gene with a different promoter, and generating influenza viruses that have truncated NS1 genes (Muster et al, 1991;Murphy et al, 1997;Parkin et al, 1997;Talon et al, 2000). Our study with TK/OR/71-del, which has truncated NS1 protein, showed attenuation in pathogenicity and a decreased ability to replicate in chickens (Cauthen et al, 1999).…”

Study of the biological properties of attenuated variants of the virulent A/WSN/33 strain of influenza virus, obtained by the site-specific mutagenesis of PB2-gene

“…Опыты, проведенные некоторыми исследователями [12,18], показали высокую эффективность включения сайт-специфических мутаций в РВ2-ген вирулентного штамма для получения аттенуированных вариантов вируса гриппа А. Применение этой технологии в отношении РВ2-гена позволило включать в этот ген несколько аттенуирующих мутаций, что дало возможность контролировать степень аттенуации вирулентного штамма. Данные, полученные этими авторами, свидетельствовали также о высокой генетической стабильности некоторых из этих вариантов, полученных с помощью данной технологии.…”