title: [Lethal effect of adenovirus-mediated HSV-TK gene in combination with hydroxycamptothecin on human bladder cancer in vitro].
abstract: OBJECTIVE
  To evaluate the lethal effect of adenovirus-mediated HSV-TK-ganciclovir (GCV) gene therapy in combination with hydroxycamptothecin (HCPT) on hunman bladder carcinoma cell line T-24 cells.
  
  
  METHODS
  Human bladder carcinoma cell line T-24 was transfected with adenovirus expression vector containing TK gene and green fluorescent protein (GFP) gene, and the transfection efficiency was observed and TK expression detected by PCR. After successful cell transfection indicated by GFP expression, GCV and hydroxycamptothecin are respectively added into the cell culture with normal T-24 cells serving as the blank control group. The growth inhibition rate of hunman bladder carcinoma cells in response to HCPT treatment for 72 h and the cell survival rate of 24 h, 48 h and 72 h after transfection with different protocols were observed by MTT assay. The apoptosis of the cells treated with GCV (0.5 mg/ml)+HCPT (10 mg/L) for 4 h was observed by flow cytometry.
  
  
  RESULTS
  The cell inhibition rate increased gradually with increment of HCPT concentration, from 14% at HCPT concentration of 0.01 mg/L to 60% at 50 mg/L, but for a concentration above 100 mg/L, the inhibition rate did not exhibit further increase (P=0.216). GCV alone and GCV in combination with HCPT both resulted in significantly decreased survival rate of human bladder carcinoma cells (P=0.00), and the killing efficiency of the cells by GCV+HCPT protocol increased obviously with increment of HCPT concentration and prolongation of the action time. The cells treated with 0.5 mg/ml GCV alone for 72 h retained a cell survival rate of 34.6%, which was lowered to only 8.07% with combined treatment with GCV (0.5mg/ml) and HCPT (10 mg/L). Typical apoptotic peak before M1 phase of the cells appeared 4 h after treatment with GCV+10 mg/ml HCPT, which resulted in a apoptosis rate of 52.93%.
  
  
  CONCLUSIONS
  HSV-TK/GCV in combination with HCPT can enhance the lethal effect of suicide gene therapy against human bladder carcinoma cells and effectively induce apoptosis of the cells.
authors.affiliation: Department of Urologic Surgery, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. huanghe3000@netease.com
authors.forename: He
authors.initials: H
authors.lastname: Huang
authors.forename: Wan-long
authors.initials: WL
authors.lastname: Tan
authors.forename: Wen-hui
authors.initials: WH
authors.lastname: Zhu
authors.forename: Zhong-kun
authors.initials: ZK
authors.lastname: Liang
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis; D002166:Camptothecin / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D015774:Ganciclovir / Q000494:pharmacology*; D041722:Genes, Transgenic, Suicide; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D013937:Thymidine Kinase / Q000494:pharmacology*; D014162:Transfection; D001749:Urinary Bladder Neoplasms / Q000628:therapy*
chemical_list: C527042:hydroxycamptothecinum; D013937:Thymidine Kinase; D015774:Ganciclovir; D002166:Camptothecin