title: Expression, Polyubiquitination, and Therapeutic Potential of Recombinant E6E7 from HPV16 Antigens Fused to Ubiquitin.
abstract: Ubiquitin-proteasome system plays an essential role in the immune response due to its involvement in the antigen generation and presentation to CD8+T cells. Hereby, ubiquitin fused to antigens has been explored as an immunotherapeutic strategy that requires the activation of cytotoxic T lymphocytes. Here we propose to apply this ubiquitin fusion approach to a recombinant vaccine against human papillomavirus 16-infected cells. E6E7 multi-epitope antigen was fused genetically at its N- or C-terminal end to ubiquitin and expressed in Escherichia coli as inclusion bodies. The antigens were solubilized using urea and purified by nickel affinity chromatography in denatured condition. Fusion of ubiquitin to E6E7 resulted in marked polyubiquitination in vitro mainly when fused to the E6E7 N-terminal. When tested in a therapeutic scenario, the fusion of ubiquitin to E6E7 reinforced the anti-tumor protection and increased the E6/E7-specific cellular immune responses. Present results encourage the investigation of the adjuvant potential of the ubiquitin fusion to recombinant vaccines requiring CD8+T cells.
authors.affiliation: Laboratório de Biotecnologia Molecular I, Centro de Biotecnologia, Instituto Butantan, Av. Vital Brasil 1500, São Paulo, SP, Brazil.
authors.forename: Liliane M Fernandes
authors.initials: LM
authors.lastname: de Oliveira
authors.affiliation: Laboratório de Biotecnologia Molecular I, Centro de Biotecnologia, Instituto Butantan, Av. Vital Brasil 1500, São Paulo, SP, Brazil.
authors.forename: Mirian G
authors.initials: MG
authors.lastname: Morale
authors.affiliation: Laboratório de Biotecnologia Molecular I, Centro de Biotecnologia, Instituto Butantan, Av. Vital Brasil 1500, São Paulo, SP, Brazil.
authors.forename: Agtha A M
authors.initials: AA
authors.lastname: Chaves
authors.affiliation: Laboratório de Bioquímica e Biofísica, Instituto Butantan, Av. Vital Brasil 1500, São Paulo, SP, Brazil.
authors.forename: Marilene
authors.initials: M
authors.lastname: Demasi
authors.affiliation: Laboratório de Biotecnologia Molecular I, Centro de Biotecnologia, Instituto Butantan, Av. Vital Brasil 1500, São Paulo, SP, Brazil. paulo.ho@butantan.gov.br.
authors.forename: Paulo L
authors.initials: PL
authors.lastname: Ho
mesh_terms: D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D005786:Gene Expression Regulation; D052162:Human papillomavirus 16 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009856:Oncogene Proteins, Viral / Q000235:genetics* / Q000378:metabolism; D050725:Papillomavirus E7 Proteins / Q000235:genetics* / Q000378:metabolism; D030361:Papillomavirus Infections / Q000188:drug therapy* / Q000276:immunology; D053918:Papillomavirus Vaccines / Q000008:administration & dosage* / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D025801:Ubiquitin / Q000235:genetics* / Q000378:metabolism
chemical_list: C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D053918:Papillomavirus Vaccines; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D025801:Ubiquitin; D007371:Interferon-gamma
keywords: HPV; Polyubiquitination; Recombinant antigen; Ubiquitin fusion; Vaccine