title: Antitumor effects of human interferon-alpha 2b secreted by recombinant bacillus Calmette-Guérin vaccine on bladder cancer cells.
abstract: OBJECTIVE
  Our objective was to construct a recombinant bacillus Calmette-Guérin vaccine (rBCG) that secretes human interferon-alpha 2b (IFNα-2b) and to study its immunogenicity and in vitro antitumor activity against human bladder cancer cell lines T24 and T5637.
  
  
  METHODS
  The signal sequence BCG Ag85B and the gene IFNα-2b were amplified from the genome of BCG and human peripheral blood, respectively, by polymerase chain reaction (PCR). The two genes were cloned in Escherichia coli-BCG shuttle-vector pMV261 to obtain a new recombinant plasmid pMV261-Ag85B-IFNα-2b. BCG was transformed with the recombinant plasmid by electroporation and designated rBCG-IFNα-2b. Mononuclear cells were isolated from human peripheral blood (PBMCs) and stimulated with rBCG-IFNα-2b or wild type BCG for 3 d, and then cultured with human bladder cancer cell lines T24 and T5637. Their cytotoxicities were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.
  
  
  RESULTS
  BCG was successfully transformed with the recombinant plasmid pMV261-Ag85B-IFNα-2b by electroporation and the recombinant BCG (rBCG-IFNα-2b) was capable of synthesizing and secreting cytokine IFNα-2b. PBMC proliferation was enhanced significantly by rBCG-IFNα-2b, and the cytotoxicity of PBMCs stimulated by rBCG-IFNα-2b to T24 and T5627 was significantly stronger in comparison to wild type BCG.
  
  
  CONCLUSIONS
  A recombinant BCG, secreting human IFNα-2b (rBCG-IFNα-2b), was constructed successfully and was superior to control wild type BCG in inducing immune responses and enhancing cytotoxicity to human bladder cancer cell lines T24 and T5637. This suggests that rBCG-IFNα-2b could be a promising agent for bladder cancer patients in terms of possible reductions in both clinical dosage and side effects of BCG immunotherapy.
authors.affiliation: Department of Urology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, China.
authors.forename: Guo-qing
authors.initials: GQ
authors.lastname: Ding
authors.forename: Yan-lan
authors.initials: YL
authors.lastname: Yu
authors.forename: Zhou-jun
authors.initials: ZJ
authors.lastname: Shen
authors.forename: Xie-lai
authors.initials: XL
authors.lastname: Zhou
authors.forename: Shan-wen
authors.initials: SW
authors.lastname: Chen
authors.forename: Guo-dong
authors.initials: GD
authors.lastname: Liao
authors.forename: Yue
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D001500:BCG Vaccine / Q000008:administration & dosage* / Q000235:genetics; D019496:Cancer Vaccines / Q000008:administration & dosage*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D006801:Humans; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000008:administration & dosage* / Q000235:genetics; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics; D016896:Treatment Outcome; D001749:Urinary Bladder Neoplasms / Q000628:therapy*; D014765:Viral Vaccines / Q000008:administration & dosage*
chemical_list: D001500:BCG Vaccine; D019496:Cancer Vaccines; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D014765:Viral Vaccines