title: [High expression of HPV16L2N120E7E6 fusion protein in E. coli and its inhibitory effect on tumor growth in mice].
abstract: OBJECTIVE
  To investigate the high expression of HPV16L2N120E7E6 fusion protein by prokaryotic expression system, and evaluate its immunogenicity and antitumor efficacy in vaccinated mice.
  
  
  METHODS
  The HPV16L2N120E7E6 fusion gene, its codons were optimized to increase the expression of the protein, was constructed by overlap extension PCR and inserted into prokaryotic expression vector pET9a. Then the fusion protein was expressed by inducing with IPTG in E. coli strain BL21 (DE3) harboring with plasmid pETL2N120E7E6, and further detected by SDS-PAGE and Western-blot. Finally, the humoral and cellular immune responses were measured by ELISA and ELISPOT, respectively, in vaccinated mice with the purified HPV16L2N120E7E6 fusion protein, and the antitumor efficacy was assessed in mice using the TC-1 tumor challenge model.
  
  
  RESULTS
  The codon-optimized HPV16L2N120E7E6 fusion gene was highly expressed in E. coli strain BL21 (DE3) harboring with plasmid pETL2N120E7E6, and the amount of fusion protein was nearly 48.6% of the total bacterial protein. The purified fusion protein could induce high titer of specific antibody against L2, E7 and E6 in vaccinated mice. When accompanied with the adjuvant CpG, the fusion protein was able to elicit strong and moderate cellular immune responses in vaccinated mice against peptide HPV16E7(49-57) and peptide pools of HPV16E6, respectively. Furthermore, the tumor therapeutic experiment showed that HPV16L2N120E7E6 + CpG could prevent the tumor formation in 80.0% (8/10) vaccinated mice.
  
  
  CONCLUSIONS
  The data of this study suggest that HPV16L2N120E7E6 fusion protein could be a promising candidate vaccine for treatment of chronic HPV16 infection and post-operative adjuvant therapy for cervical cancer.
authors.affiliation: National Institute for Viral Disease Control and Prevention, Beijing, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhao
authors.forename: Meng
authors.initials: M
authors.lastname: Gao
authors.forename: Jian
authors.initials: J
authors.lastname: Gao
authors.forename: Jiao
authors.initials: J
authors.lastname: Ren
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
authors.forename: Hou-wen
authors.initials: HW
authors.lastname: Tian
authors.forename: Wen-jie
authors.initials: WJ
authors.lastname: Tan
authors.forename: Li
authors.initials: L
authors.lastname: Ruan
mesh_terms: D000276:Adjuvants, Immunologic / Q000494:pharmacology; D000818:Animals; D019496:Cancer Vaccines / Q000276:immunology* / Q000627:therapeutic use; D036022:Capsid Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003062:Codon; D004926:Escherichia coli / Q000276:immunology / Q000378:metabolism; D005260:Female; D006801:Humans; D007114:Immunization / Q000379:methods; D007167:Immunotherapy / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D009368:Neoplasm Transplantation; D009838:Oligodeoxyribonucleotides / Q000276:immunology; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D050725:Papillomavirus E7 Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D053918:Papillomavirus Vaccines / Q000276:immunology / Q000627:therapeutic use; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D000276:Adjuvants, Immunologic; D019496:Cancer Vaccines; D036022:Capsid Proteins; D003062:Codon; C423449:CpG ODN 1826; C068651:E6 protein, Human papillomavirus type 16; C063316:L2 protein, Human papillomavirus type 16; D009838:Oligodeoxyribonucleotides; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D053918:Papillomavirus Vaccines; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C059731:oncogene protein E7, Human papillomavirus type 16