title: A Diverse Panel of Hepatitis C Virus Glycoproteins for Use in Vaccine Research Reveals Extremes of Monoclonal Antibody Neutralization Resistance.
abstract: Despite significant advances in the treatment of hepatitis C virus (HCV) infection, the need to develop preventative vaccines remains. Identification of the best vaccine candidates and evaluation of their performance in preclinical and clinical development will require appropriate neutralization assays utilizing diverse HCV isolates. We aimed to generate and characterize a panel of HCV E1E2 glycoproteins suitable for subsequent use in vaccine and therapeutic antibody testing. Full-length E1E2 clones were PCR amplified from patient-derived serum samples, cloned into an expression vector, and used to generate viral pseudoparticles (HCVpp). In addition, some of these clones were used to generate cell culture infectious (HCVcc) clones. The infectivity and neutralization sensitivity of these viruses were then determined. Bioinformatic and HCVpp infectivity screening of approximately 900 E1E2 clones resulted in the assembly of a panel of 78 functional E1E2 proteins representing distinct HCV genotypes and different stages of infection. These HCV glycoproteins differed markedly in their sensitivity to neutralizing antibodies. We used this panel to predict antibody efficacy against circulating HCV strains, highlighting the likely reason why some monoclonal antibodies failed in previous clinical trials. This study provides the first objective categorization of cross-genotype patient-derived HCV E1E2 clones according to their sensitivity to antibody neutralization. It has shown that HCV isolates have clearly distinguishable neutralization-sensitive, -resistant, or -intermediate phenotypes, which are independent of genotype. The panel provides a systematic means for characterization of the neutralizing response elicited by candidate vaccines and for defining the therapeutic potential of monoclonal antibodies.
  
  
  OBJECTIVE
  Hepatitis C virus (HCV) has a global burden of more than 170 million people, many of whom cannot attain the new, expensive, direct-acting antiviral therapies. A safe and effective vaccine that generates both T cell responses and neutralizing antibodies is required to eradicate the disease. Regions within the HCV surface glycoproteins E1 and E2 are essential for virus entry and are targets for neutralizing antibodies. Screening of vaccine candidates requires suitable panels of glycoproteins that represent the breadth of neutralization resistance. Use of a standard reference panel for vaccine studies will ensure comparability of data sets, as has become routine for HIV-1. Here, we describe a large panel of patient-derived HCV glycoproteins with an assessment of their neutralization sensitivity to defined monoclonal antibodies, which has enabled us to predict their likely efficacy in the wider HCV-infected population. The panel could also be important for future selection of additional therapeutic antibodies and for vaccine design.
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Urbanowicz
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom.
authors.forename: C Patrick
authors.initials: CP
authors.lastname: McClure
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom.
authors.forename: Richard J P
authors.initials: RJ
authors.lastname: Brown
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom.
authors.forename: Theocharis
authors.initials: T
authors.lastname: Tsoleridis
authors.affiliation: Karolinska Institutet, Department of Clinical Neurosciences, Center for Molecular Medicine, Karolinska University Hospital Solna, Stockholm, Sweden.
authors.forename: Mats A A
authors.initials: MA
authors.lastname: Persson
authors.affiliation: Unité de Virologie Structurale, Département de Virologie, Institut Pasteur, Paris, France CNRS UMR 3569, Paris, France.
authors.forename: Thomas
authors.initials: T
authors.lastname: Krey
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom.
authors.forename: William L
authors.initials: WL
authors.lastname: Irving
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom jonathan.ball@nottingham.ac.uk.
authors.forename: Jonathan K
authors.initials: JK
authors.lastname: Ball
authors.affiliation: School of Life Sciences, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom NIHR Nottingham Digestive Diseases Biomedical Research Unit, The University of Nottingham, Nottingham University Hospitals NHS Trust, Nottingham, United Kingdom.
authors.forename: Alexander W
authors.initials: AW
authors.lastname: Tarr
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000627:therapeutic use; D057134:Antibodies, Neutralizing / Q000276:immunology*; D002460:Cell Line; D057809:HEK293 Cells; D016174:Hepacivirus / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D006526:Hepatitis C / Q000276:immunology / Q000821:virology; D018937:Hepatitis C Antibodies / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D014759:Viral Envelope Proteins / Q000276:immunology*; D014761:Viral Hepatitis Vaccines / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; C087035:E1 protein, Hepatitis C virus; D018937:Hepatitis C Antibodies; D014759:Viral Envelope Proteins; D014761:Viral Hepatitis Vaccines; C091015:glycoprotein E2, Hepatitis C virus