Phenotypic and genotypic characterization of influenza virus mutants selected with the sialidase fusion protein DAS181

Abstract

Influenza viruses (IFVs) frequently achieve resistance to antiviral drugs, necessitating the development of compounds with novel mechanisms of action. DAS181 (Fludase^®), a sialidase fusion protein, may have a reduced potential for generating drug resistance due to its novel host-targeting mechanism of action. IFV strains B/Maryland/1/59 and A/Victoria/3/75 (H3N2) were subjected to >30 passages under increasing selective pressure with DAS181. The DAS181-selected IFV isolates were characterized in vitro and in mice. Despite extensive passaging, DAS181-selected viruses exhibited a very low level of resistance to DAS181, which ranged between 3- and 18-fold increase in EC₅₀. DAS181-selected viruses displayed an attenuated phenotype in vitro, as exhibited by slower growth, smaller plaque size and increased particle to pfu ratios relative to wild-type virus. Further, the DAS181 resistance phenotype was unstable and was substantially reversed over time upon DAS181 withdrawal. In mice, the DAS181-selected viruses exhibited no greater virulence than their wild-type counterparts. Genotypic and phenotypic analysis of DAS181-selected viruses revealed mutations in the haemagglutinin (HA) and neuraminidase (NA) molecules and also changes in HA and NA function. Results indicate that resistance to DAS181 is minimal and unstable. The DAS181-selected IFV isolates exhibit reduced fitness invitro, likely due to altered HA and NA functions.