title: Packaging and Prefusion Stabilization Separately and Additively Increase the Quantity and Quality of Respiratory Syncytial Virus (RSV)-Neutralizing Antibodies Induced by an RSV Fusion Protein Expressed by a Parainfluenza Virus Vector.
abstract: Human respiratory syncytial virus (RSV) and human parainfluenza virus type 3 (HPIV3) are major pediatric respiratory pathogens that lack vaccines. A chimeric bovine/human PIV3 (rB/HPIV3) virus expressing the unmodified, wild-type (wt) RSV fusion (F) protein from an added gene was previously evaluated in seronegative children as a bivalent intranasal RSV/HPIV3 vaccine, and it was well tolerated but insufficiently immunogenic for RSV F. We recently showed that rB/HPIV3 expressing a partially stabilized prefusion form (pre-F) of RSV F efficiently induced "high-quality" RSV-neutralizing antibodies, defined as antibodies that neutralize RSV in vitro without added complement (B. Liang et al., J Virol 89:9499-9510, 2015, doi:10.1128/JVI.01373-15). In the present study, we modified RSV F by replacing its cytoplasmic tail (CT) domain or its CT and transmembrane (TM) domains (TMCT) with counterparts from BPIV3 F, with or without pre-F stabilization. This resulted in RSV F being packaged in the rB/HPIV3 particle with an efficiency similar to that of RSV particles. Enhanced packaging was substantially attenuating in hamsters (10- to 100-fold) and rhesus monkeys (100- to 1,000-fold). Nonetheless, TMCT-directed packaging substantially increased the titers of high-quality RSV-neutralizing serum antibodies in hamsters. In rhesus monkeys, a strongly additive immunogenic effect of packaging and pre-F stabilization was observed, as demonstrated by 8- and 30-fold increases of RSV-neutralizing serum antibody titers in the presence and absence of added complement, respectively, compared to pre-F stabilization alone. Analysis of vaccine-induced F-specific antibodies by binding assays indicated that packaging conferred substantial stabilization of RSV F in the pre-F conformation. This provides an improved version of this well-tolerated RSV/HPIV3 vaccine candidate, with potently improved immunogenicity, which can be returned to clinical trials.
  
  
  OBJECTIVE
  Human respiratory syncytial virus (RSV) and human parainfluenza virus type 3 (HPIV3) are major viral agents of acute pediatric bronchiolitis and pneumonia worldwide that lack vaccines. A bivalent intranasal RSV/HPIV3 vaccine candidate consisting of a chimeric bovine/human PIV3 (rB/HPIV3) strain expressing the RSV fusion (F) protein was previously shown to be well tolerated by seronegative children but was insufficiently immunogenic for RSV F. In the present study, the RSV F protein was engineered to be packaged efficiently into vaccine virus particles. This resulted in a significantly enhanced quantity and quality of RSV-neutralizing antibodies in hamsters and nonhuman primates. In nonhuman primates, this effect was strongly additive to the previously described stabilization of the prefusion conformation of the F protein. The improved immunogenicity of RSV F by packaging appeared to involve prefusion stabilization. These findings provide a potently more immunogenic version of this well-tolerated vaccine candidate and should be applicable to other vectored vaccines.
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Bo
authors: B
authors: Liang
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Joan O
authors: JO
authors: Ngwuta
authors: Experimental Primate Virology Section, Comparative Medicine Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Poolesville, Maryland, USA.
authors: Richard
authors: R
authors: Herbert
authors: Experimental Primate Virology Section, Comparative Medicine Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Poolesville, Maryland, USA.
authors: Joanna
authors: J
authors: Swerczek
authors: Rocky Mountain Laboratory, Microscopy Unit, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA.
authors: David W
authors: DW
authors: Dorward
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Emerito
authors: E
authors: Amaro-Carambot
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Natalie
authors: N
authors: Mackow
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Barbora
authors: B
authors: Kabatova
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Matthias
authors: M
authors: Lingemann
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Sonja
authors: S
authors: Surman
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Lijuan
authors: L
authors: Yang
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Man
authors: M
authors: Chen
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Syed M
authors: SM
authors: Moin
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Azad
authors: A
authors: Kumar
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Jason S
authors: JS
authors: McLellan
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Peter D
authors: PD
authors: Kwong
authors: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Barney S
authors: BS
authors: Graham
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Anne
authors: A
authors: Schaap-Nutt
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors: Peter L
authors: PL
authors: Collins
authors: RNA Viruses Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA munirs@niaid.nih.gov.
authors: Shirin
authors: S
authors: Munir
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000235:genetics* / Q000276:immunology; D000914:Antibodies, Viral / Q000235:genetics / Q000276:immunology; D002213:Capsid / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D002522:Chlorocebus aethiops; D006224:Cricetinae; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D008253:Macaca mulatta; D029101:Parainfluenza Virus 3, Bovine / Q000235:genetics* / Q000276:immunology; D010224:Parainfluenza Virus 3, Human / Q000235:genetics* / Q000276:immunology; D018357:Respiratory Syncytial Virus Infections / Q000276:immunology / Q000821:virology; D022261:Respiratory Syncytial Virus Vaccines / Q000235:genetics / Q000276:immunology; D018113:Respiratory Syncytial Virus, Human / Q000235:genetics* / Q000276:immunology*; D010253:Respirovirus Infections / Q000276:immunology / Q000821:virology; D014709:Vero Cells; D014760:Viral Fusion Proteins / Q000235:genetics* / Q000276:immunology; D014779:Virus Replication / Q000235:genetics
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D022261:Respiratory Syncytial Virus Vaccines; D014760:Viral Fusion Proteins