title: Influenza A virus attenuation by codon deoptimization of the NS gene for vaccine development.
abstract: Influenza viral infection represents a serious public health problem that causes contagious respiratory disease, which is most effectively prevented through vaccination to reduce transmission and future infection. The nonstructural (NS) gene of influenza A virus encodes an mRNA transcript that is alternatively spliced to express two viral proteins, the nonstructural protein 1 (NS1) and the nuclear export protein (NEP). The importance of the NS gene of influenza A virus for viral replication and virulence has been well described and represents an attractive target to generate live attenuated influenza viruses with vaccine potential. Considering that most amino acids can be synthesized from several synonymous codons, this study employed the use of misrepresented mammalian codons (codon deoptimization) for the de novo synthesis of a viral NS RNA segment based on influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus. We generated three different recombinant influenza PR8 viruses containing codon-deoptimized synonymous mutations in coding regions comprising the entire NS gene or the mRNA corresponding to the individual viral protein NS1 or NEP, without modifying the respective splicing and packaging signals of the viral segment. The fitness of these synthetic viruses was attenuated in vivo, while they retained immunogenicity, conferring both homologous and heterologous protection against influenza A virus challenges. These results indicate that influenza viruses can be effectively attenuated by synonymous codon deoptimization of the NS gene and open the possibility of their use as a safe vaccine to prevent infections with these important human pathogens.
  
  
  OBJECTIVE
  Vaccination serves as the best therapeutic option to protect humans against influenza viral infections. However, the efficacy of current influenza vaccines is suboptimal, and novel approaches are necessary for the prevention of disease cause by this important human respiratory pathogen. The nonstructural (NS) gene of influenza virus encodes both the multifunctional nonstructural protein 1 (NS1), essential for innate immune evasion, and the nuclear export protein (NEP), required for the nuclear export of viral ribonucleoproteins and for timing of the virus life cycle. Here, we have generated a recombinant influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus containing a codon-deoptimized NS segment that is attenuated in vivo yet retains immunogenicity and protection efficacy against homologous and heterologous influenza virus challenges. These results open the exciting possibility of using this NS codon deoptimization methodology alone or in combination with other approaches for the future development of vaccine candidates to prevent influenza viral infections.
authors: Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA.
authors: Aitor
authors: A
authors: Nogales
authors: Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA.
authors: Steven F
authors: SF
authors: Baker
authors: Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA.
authors: Emilio
authors: E
authors: Ortiz-Riaño
authors: Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA.
authors: Stephen
authors: S
authors: Dewhurst
authors: Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA New York Influenza Center of Excellence, University of Rochester, Rochester, New York, USA Center for Biodefense and Immune Modeling, University of Rochester, Rochester, New York, USA David H. Smith Center for Vaccine Biology and Immunology, University of Rochester, Rochester, New York, USA.
authors: David J
authors: DJ
authors: Topham
authors: Department of Microbiology and Immunology, University of Rochester, Rochester, New York, USA luis_martinez@urmc.rochester.edu.
authors: Luis
authors: L
authors: Martínez-Sobrido
mesh_terms: D000818:Animals; D001483:Base Sequence; D003062:Codon; D005260:Female; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics* / Q000276:immunology; D007252:Influenza Vaccines / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology*; D007251:Influenza, Human / Q000517:prevention & control / Q000821:virology*; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D015202:Protein Engineering; D017361:Viral Nonstructural Proteins / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology*
chemical_list: D003062:Codon; D007252:Influenza Vaccines; D017361:Viral Nonstructural Proteins