Respiratory Syncytial Virus Fusion Glycoprotein Expressed in Insect Cells Form Protein Nanoparticles That Induce Protective Immunity in Cotton Rats
et al. (2012) Respiratory Syncytial Virus Fusion Glycoprotein Expressed in Insect Cells Form Protein
Nanoparticles That Induce Protective Immunity in Cotton Rats. PLoS ONE 7(11): e50852. doi:10.1371/journal.pone.0050852
Respiratory Syncytial Virus Fusion Glycoprotein Expressed in Insect Cells Form Protein Nanoparticles That Induce Protective Immunity in Cotton Rats
Gale Smith 0
Rama Raghunandan 0
Yingyun Wu 0
Ye Liu 0
Michael Massare 0
Margret Nathan 0
Bin Zhou 0
Hanxin Lu 0
Sarathi Boddapati 0
Jingning Li 0
David Flyer 0
Gregory Glenn 0
Steven M. Varga, University of Iowa, United States of America
0 1 Discovery , Novavax Incorporated, Rockville, Maryland , United States of America, 2 Process Development , Novavax Incorporated, Rockville, Maryland , United States of America, 3 Formulation Development , Novavax Incorporated, Rockville, Maryland , United States of America, 4 Analytical Development , Novavax Incorporated, Rockville, Maryland , United States of America, 5 Clinical Development , Novavax Incorporated, Rockville, Maryland , United States of America
Respiratory Syncytial Virus (RSV) is an important viral agent causing severe respiratory tract disease in infants and children as well as in the elderly and immunocompromised individuals. The lack of a safe and effective RSV vaccine represents a major unmet medical need. RSV fusion (F) surface glycoprotein was modified and cloned into a baculovirus vector for efficient expression in Sf9 insect cells. Recombinant RSV F was glycosylated and cleaved into covalently linked F2 and F1 polypeptides that formed homotrimers. RSV F extracted and purified from insect cell membranes assembled into 40 nm protein nanoparticles composed of multiple RSV F oligomers arranged in the form of rosettes. The immunogenicity and protective efficacy of purified RSV F nanoparticles was compared to live and formalin inactivated RSV in cotton rats. Immunized animals induced neutralizing serum antibodies, inhibited virus replication in the lungs, and had no signs of disease enhancement in the respiratory track of challenged animals. RSV F nanoparticles also induced IgG competitive for binding of palivizumab neutralizing monoclonal antibody to RSV F antigenic site II. Antibodies to this epitope are known to protect against RSV when passively administered in high risk infants. Together these data provide a rational for continued development a recombinant RSV F nanoparticle vaccine candidate.
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Competing Interests: The authors have reviewed the PLOS ONE guidelines for authors and declare: Each of the authors are employed by Novavax, Inc.,
Rockville, MD, a public biotechnology company, and GS and MM hold patents owned by Novavax. This does not alter the authors adherence to all the PLOS ONE
policies on sharing data and materials.
Respiratory syncytial virus (RSV) is the most common cause
of acute lower respiratory infection in infants and young
children, and a major disease burden in the elderly. Despite the
fact that the RSV virus was characterized half a century ago,
there is currently no vaccine for RSV and development has
been hampered by vaccine-mediated disease enhancement in
children administered a formalin inactivated RSV in the 1960s
[1,2]. Challenges in antigen production, purity, stability, and
potency of RSV vaccine candidates have also been impediments
to development [35].
The RSV fusion glycoprotein (F) mediates viral entry into
cells and cell to cell fusion, is a target of neutralizing antibodies,
and highly conserved between RSV A and B strains [6,7]. RSV
F is produced as a precursor (F0) that is cleaved at Arg109 and
Arg136 by cellular furin to three fragments, a shorter F2
polypeptide at the N-terminus covalently linked by two
disulfides to a longer F1 polypeptide with an 18 amino acid
fusion domain at the N-terminus and a hydrophobic membrane
spanning region near the C-terminus; the intervening 27 amino
acid fragment is released. Neutralizing monoclonal antibodies
palivizumab and motavizumab bind to RSV F antigenic site II
(Asn258 - Val278) [8] and have been shown to protect against
both lower and upper respiratory RSV disease in high risk and
term infants [9,10] The structures of the RSV F epitope
polypeptides that bind these neutralizing antibodies are larger
than the linear peptide and palivizumab binds with nanomolar
and motavizumab picomolar affinity to RSV F [1113].
Modeling predicts that the full extent of the binding of
palivizumab and motavizumab requires amino acids from one
or two RSV F protomers, respectively. Therefore preserving
RSV F tertiary and quaternary structures may be important in
the development of an RSV F vaccine to preserve the native
conformation of this important neutralizing region.
In this report an oligomeric form of a modified full length
RSV F was efficiently produced in Sf9 insect cells using
a baculovirus vector. Recombinant RSV F extracted from
cellular membranes and purified, assembled into nanoparticles
with morphology c (...truncated)