title: Production of Human papillomavirus pseudovirions in plants and their use in pseudovirion-based neutralisation assays in mammalian cells.
abstract: Human papillomaviruses (HPV) cause cervical cancer and have recently also been implicated in mouth, laryngeal and anogenital cancers. There are three commercially available prophylactic vaccines that show good efficacy; however, efforts to develop second-generation vaccines that are more affordable, stable and elicit a wider spectrum of cross-neutralising immunity are still ongoing. Testing antisera elicited by current and candidate HPV vaccines for neutralizing antibodies is done using a HPV pseudovirion (PsV)-based neutralisation assay (PBNA). PsVs are produced by transfection of mammalian cell cultures with plasmids expressing L1 and L2 capsid proteins, and a reporter gene plasmid, a highly expensive process. We investigated making HPV-16 PsVs in plants, in order to develop a cheaper alternative. The secreted embryonic alkaline phosphatase (SEAP) reporter gene and promoter were cloned into a geminivirus-derived plant expression vector, in order to produce circular dsDNA replicons. This was co-introduced into Nicotiana benthamiana plants with vectors expressing L1 and L2 via agroinfiltration, and presumptive PsVs were purified. The PsVs contained DNA, and could be successfully used for PBNA with anti-HPV antibodies. This is the first demonstration of the production of mammalian pseudovirions in plants, and the first demonstration of the potential of plants to make DNA vaccines.
authors: Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, 7701, South Africa.
authors: Renate L
authors: RL
authors: Lamprecht
authors: Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, 7701, South Africa.
authors: Paul
authors: P
authors: Kennedy
authors: Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, 7701, South Africa.
authors: Suzanne M
authors: SM
authors: Huddy
authors: Pharmaceutical Product Development, Fraunhofer IME, Aachen, 52074, Germany.
authors: Susanne
authors: S
authors: Bethke
authors: Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, 7701, South Africa.
authors: Megan
authors: M
authors: Hendrikse
authors: Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, 7701, South Africa.
authors: Inga I
authors: II
authors: Hitzeroth
authors: Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, 7701, South Africa.|Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Rondebosch, 7701, South Africa.
authors: Edward P
authors: EP
authors: Rybicki
mesh_terms: D057134:Antibodies, Neutralizing / Q000276:immunology*; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology*; D036022:Capsid Proteins / Q000276:immunology / Q000378:metabolism; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D009500:Neutralization Tests; D027383:Papillomaviridae / Q000276:immunology*; D053918:Papillomavirus Vaccines / Q000276:immunology*; D010957:Plasmids; D014026:Nicotiana / Q000235:genetics / Q000821:virology*; D014162:Transfection; D014771:Virion / Q000276:immunology* / Q000378:metabolism
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D036022:Capsid Proteins; D053918:Papillomavirus Vaccines