title: In vivo assessment of NS1-truncated influenza virus with a novel SLSYSINWRH motif as a self-adjuvanting live attenuated vaccine.
abstract: Mutants of influenza virus that encode C-terminally truncated NS1 proteins (NS1-truncated mutants) characteristically induce high interferon responses. The dual activity of interferon in blocking virus replication and enhancing the development of adaptive immune responses makes these mutants promising as self-adjuvanting live-attenuated influenza vaccine (LAIV) candidates. Yet, among the NS1-truncated mutants, the length of NS1 is not directly correlated with the interferon-inducing efficiency, the level of attenuation, or effectiveness as LAIV. Using quantitative in vitro biologically active particle subpopulation analysis as a tool to identify potential LAIV candidates from a pool of NS1-truncated mutants, we previously predicted that a NS1-truncated mutant pc2, which was less effective as a LAIV in chickens, would be sufficiently effective as a LAIV in mammalian hosts. In this study, we confirmed that pc2 protected mice and pigs against heterologous virus challenge in terms of preventing clinical signs and reducing virus shedding. pc2 expresses a unique SLSYSINWRH motif at the C-terminus of its truncated NS1. Deletion of the SLSYSINWRH motif led to ~821-fold reduction in the peak yield of type I interferon induced in murine cells. Furthermore, replacement of the SLSYSINWRH motif with the wildtype MVKMDQAIMD sequence did not restore the interferon-inducing efficiency. The diminished interferon induction capacity in the absence of the SLSYSINWRH motif was similar to that observed in other mutants which are less effective LAIV candidates. Remarkably, pc2 induced 16-fold or more interferon in human lung and monkey kidney cells compared to the temperature-sensitive, cold-adapted Ann Arbor virus that is currently used as a master backbone for LAIVs such as FluMist. Although the mechanism by which the SLSYSINWRH motif regulates the vaccine properties of pc2 has not been elucidated, this motif has potential use in engineering self-adjuvanting NS1-truncated-based LAIVs.
authors: Department of Molecular and Cell Biology, University of Connecticut, Storrs, CT, United States of America; Food Animal Health Research Program, The Ohio State University, Wooster, OH, United States of America.
authors: John M
authors: JM
authors: Ngunjiri
authors: Department of Poultry Diseases, Faculty of Veterinary Medicine, Beni-Suef University, Beni-Suef, 62511, Egypt; Department of Preventive Medicine, The Ohio State University, Columbus, OH, United States of America.
authors: Ahmed
authors: A
authors: Ali
authors: Department of Veterinary Bioscience, The Ohio State University, Columbus, OH, United States of America.
authors: Prosper
authors: P
authors: Boyaka
authors: Department of Molecular and Cell Biology, University of Connecticut, Storrs, CT, United States of America.
authors: Philip I
authors: PI
authors: Marcus
authors: Food Animal Health Research Program, The Ohio State University, Wooster, OH, United States of America; Department of Preventive Medicine, The Ohio State University, Columbus, OH, United States of America.
authors: Chang-Won
authors: CW
authors: Lee
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics; D000704:Analysis of Variance; D000818:Animals; D017931:DNA Primers / Q000235:genetics; D066298:In Vitro Techniques; D007252:Influenza Vaccines / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology*; D007370:Interferon Type I / Q000276:immunology*; D051379:Mice; D016296:Mutagenesis; D009975:Orthomyxoviridae / Q000235:genetics* / Q000276:immunology; D015202:Protein Engineering / Q000379:methods; D013552:Swine; D017361:Viral Nonstructural Proteins / Q000235:genetics*; D017201:Virus Shedding / Q000187:drug effects
chemical_list: D017931:DNA Primers; C085820:INS1 protein, influenza virus; D007252:Influenza Vaccines; D007370:Interferon Type I; D017361:Viral Nonstructural Proteins