title: Deliberate attenuation of chikungunya virus by adaptation to heparan sulfate-dependent infectivity: a model for rational arboviral vaccine design.
abstract: Mosquito-borne chikungunya virus (CHIKV) is a positive-sense, single-stranded RNA virus from the genus Alphavirus, family Togaviridae, which causes fever, rash and severe persistent polyarthralgia in humans. Since there are currently no FDA licensed vaccines or antiviral therapies for CHIKV, the development of vaccine candidates is of critical importance. Historically, live-attenuated vaccines (LAVs) for protection against arthropod-borne viruses have been created by blind cell culture passage leading to attenuation of disease, while maintaining immunogenicity. Attenuation may occur via multiple mechanisms. However, all examined arbovirus LAVs have in common the acquisition of positively charged amino acid substitutions in cell-surface attachment proteins that render virus infection partially dependent upon heparan sulfate (HS), a ubiquitously expressed sulfated polysaccharide, and appear to attenuate by retarding dissemination of virus particles in vivo. We previously reported that, like other wild-type Old World alphaviruses, CHIKV strain, La Réunion, (CHIKV-LR), does not depend upon HS for infectivity. To deliberately identify CHIKV attachment protein mutations that could be combined with other attenuating processes in a LAV candidate, we passaged CHIKV-LR on evolutionarily divergent cell-types. A panel of single amino acid substitutions was identified in the E2 glycoprotein of passaged virus populations that were predicted to increase electrostatic potential. Each of these substitutions was made in the CHIKV-LR cDNA clone and comparisons of the mutant viruses revealed surface exposure of the mutated residue on the spike and sensitivity to competition with the HS analog, heparin, to be primary correlates of attenuation in vivo. Furthermore, we have identified a mutation at E2 position 79 as a promising candidate for inclusion in a CHIKV LAV.
authors.affiliation: Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America ; Department of Microbiology & Molecular Genetics, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Christina L
authors.initials: CL
authors.lastname: Gardner
authors.affiliation: CEITEC, Masaryk University, Brno, Czech Republic.
authors.forename: Jozef
authors.initials: J
authors.lastname: Hritz
authors.affiliation: Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America ; Department of Microbiology & Molecular Genetics, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Chengqun
authors.initials: C
authors.lastname: Sun
authors.affiliation: Department of Diagnostic Medicine & Pathobiology, Biosecurity Research Institute, Kansas State University, Manhattan, Kansas, United States of America.
authors.forename: Dana L
authors.initials: DL
authors.lastname: Vanlandingham
authors.affiliation: Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America ; Department of Computational and Systems Biology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Timothy Y
authors.initials: TY
authors.lastname: Song
authors.affiliation: Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America ; Department of Computational and Systems Biology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Elodie
authors.initials: E
authors.lastname: Ghedin
authors.affiliation: Department of Diagnostic Medicine & Pathobiology, Biosecurity Research Institute, Kansas State University, Manhattan, Kansas, United States of America.
authors.forename: Stephen
authors.initials: S
authors.lastname: Higgs
authors.affiliation: Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America ; Department of Microbiology & Molecular Genetics, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: William B
authors.initials: WB
authors.lastname: Klimstra
authors.affiliation: Center for Vaccine Research, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America ; Department of Microbiology & Molecular Genetics, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
authors.forename: Kate D
authors.initials: KD
authors.lastname: Ryman
mesh_terms: D000220:Adaptation, Biological / Q000187:drug effects / Q000235:genetics; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology; D002646:Chikungunya virus / Q000187:drug effects / Q000235:genetics / Q000276:immunology / Q000472:pathogenicity; D016207:Cytokines / Q000378:metabolism; D006497:Heparan Sulfate / Q000494:pharmacology*; D051379:Mice; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D055672:Static Electricity; D014613:Vaccines, Attenuated / Q000235:genetics* / Q000276:immunology; D014759:Viral Envelope Proteins / Q000235:genetics; D014765:Viral Vaccines / Q000235:genetics* / Q000276:immunology; D014774:Virulence / Q000235:genetics
chemical_list: D057134:Antibodies, Neutralizing; D016207:Cytokines; D006497:Heparan Sulfate; D014613:Vaccines, Attenuated; D014759:Viral Envelope Proteins; D014765:Viral Vaccines