title: Severe acute respiratory syndrome coronavirus spike protein expressed by attenuated vaccinia virus protectively immunizes mice.
abstract: The spike protein (S), a membrane component of severe acute respiratory syndrome coronavirus (SARS-CoV) is anticipated to be an important component of candidate vaccines. We constructed recombinant forms of the highly attenuated modified vaccinia virus Ankara (MVA) containing the gene encoding full-length SARS-CoV S with and without a C-terminal epitope tag called MVA/S-HA and MVA/S, respectively. Cells infected with MVA/Sor MVA/S-HA synthesized a 200-kDa protein, which was recognized by antibody raised against a synthetic peptide of SARS-CoV S or the epitope tag in Western blot analyses. Further studies indicated that S was N-glycosylated and migrated in SDS polyacrylamide gels with an apparent mass of approximately 160 kDa after treatment with peptide N-glycosidase F. The acquisition of resistance to endoglycosidase H indicated trafficking of S to the medial Golgi compartment, and confocal microscopy showed that S was transported to the cell surface. Intranasal or intramuscular inoculations of BALB/c mice with MVA/S produced serum antibodies that recognized the SARS S in ELISA and neutralized SARS-CoV in vitro. Moreover, MVA/S administered by either route elicited protective immunity, as shown by reduced titers of SARS-CoV in the upper and lower respiratory tracts of mice after challenge. Passive transfer of serum from mice immunized with MVA/S to naïve mice also reduced the replication of SARS-CoV in the respiratory tract after challenge, demonstrating a role for antibody to S in protection. The attenuated nature of MVA and the ability of MVA/S to induce neutralizing antibody that protects mice support further development of this candidate vaccine.
authors.affiliation: Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Himani
authors.initials: H
authors.lastname: Bisht
authors.forename: Anjeanette
authors.initials: A
authors.lastname: Roberts
authors.forename: Leatrice
authors.initials: L
authors.lastname: Vogel
authors.forename: Alexander
authors.initials: A
authors.lastname: Bukreyev
authors.forename: Peter L
authors.initials: PL
authors.lastname: Collins
authors.forename: Brian R
authors.initials: BR
authors.lastname: Murphy
authors.forename: Kanta
authors.initials: K
authors.lastname: Subbarao
authors.forename: Bernard
authors.initials: B
authors.lastname: Moss
mesh_terms: D000818:Animals; D002460:Cell Line; D006367:HeLa Cells; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D051379:Mice; D008807:Mice, Inbred BALB C; D018613:Microscopy, Confocal; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D045473:Severe acute respiratory syndrome-related coronavirus / Q000502:physiology*; D064370:Spike Glycoprotein, Coronavirus; D014616:Vaccinia virus / Q000235:genetics*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology / Q000502:physiology*
chemical_list: D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins