title: Utilizing a TLR5-Adjuvanted Cytomegalovirus as a Lentiviral Vaccine in the Nonhuman Primate Model for AIDS.
abstract: Despite tremendous progress in our understanding of human immunodeficiency virus (HIV) natural history and advances in HIV treatment, there is neither an approved vaccine nor a cure for infection. Here, we describe the development and characterization of a novel replicating vaccine vector utilizing Cytomegalovirus (CMV) and a TLR5 adjuvant. After partial truncation of the central, immunodominant hypervariable domain, flagellin (fliC) from Salmonella was cloned downstream of a codon optimized gag gene from simian immunodeficiency virus (SIV) and transiently expressed in telomerized rhesus fibroblast (TeloRF) cells in culture. Lysates generated from these transfected cells induced the tumor necrosis factor alpha (TNF-α), in a mouse macrophage cell line, in a TLR5-dependent manner. The Gag/FliC expression construct was cloned into a bacterial artificial chromosome encoding the rhesus CMV (RhCMV) genome, and infectious RhCMV was generated following transfection of TeloRF cells. This virus stably expressed an SIV Gag/FliC fusion protein through four serial passages. Lysates generated from infected cells induced TNF-α in a TLR5-dependent manner. Western blot analysis of infected cell lysates verified expression of a Gag/FliC fusion protein using a SIV p27 capsid monoclonal antibody. Lastly, rhesus macaques inoculated with this novel RhCMV virus demonstrated increased inflammatory responses at the site of inoculation seven days post-infection when compared to the parental RhCMV. These results demonstrate that an artificially constructed replicating RhCMV expressing an SIV Gag/FliC fusion protein is capable of activating TLR5 in a macrophage cell line in vitro and induction of an altered inflammatory response in vivo. Ongoing animals studies are aimed at determining vaccine efficacy, including subsequent challenge with pathogenic SIV.
authors.affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Jesse D
authors.initials: JD
authors.lastname: Deere
authors.affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: W L William
authors.initials: WL
authors.lastname: Chang
authors.affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Luis D
authors.initials: LD
authors.lastname: Castillo
authors.affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Kim A
authors.initials: KA
authors.lastname: Schmidt
authors.affiliation: Department of Veterinary Medicine and Epidemiology, School of Veterinary Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Hung T
authors.initials: HT
authors.lastname: Kieu
authors.affiliation: Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Renzette
authors.affiliation: Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.
authors.forename: Timothy
authors.initials: T
authors.lastname: Kowalik
authors.affiliation: Department of Veterinary Pathology, Microbiology and Immunology, School of Veterinary Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Stephen W
authors.initials: SW
authors.lastname: Barthold
authors.affiliation: Department of Medical Microbiology and Immunology, School of Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Barbara L
authors.initials: BL
authors.lastname: Shacklett
authors.affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Barry
authors.affiliation: Department of Veterinary Medicine and Epidemiology, School of Veterinary Medicine, University of California Davis, Davis, California, United States of America.
authors.forename: Ellen E
authors.initials: EE
authors.lastname: Sparger
mesh_terms: D016915:AIDS Vaccines / Q000276:immunology*; D000276:Adjuvants, Immunologic; D000818:Animals; D003587:Cytomegalovirus / Q000235:genetics / Q000276:immunology; D005260:Female; D005786:Gene Expression Regulation; D023061:Gene Order; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology; D005822:Genetic Vectors / Q000235:genetics / Q000276:immunology; D016086:Lentivirus / Q000276:immunology*; D008253:Macaca mulatta; D009154:Mutation; D054730:Protein Interaction Domains and Motifs; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D016097:Simian Acquired Immunodeficiency Syndrome / Q000276:immunology* / Q000517:prevention & control*; D015302:Simian Immunodeficiency Virus / Q000276:immunology*; D051198:Toll-Like Receptor 5 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics
chemical_list: D016915:AIDS Vaccines; D000276:Adjuvants, Immunologic; D015683:Gene Products, gag; D011993:Recombinant Fusion Proteins; D051198:Toll-Like Receptor 5; D014409:Tumor Necrosis Factor-alpha