Experimental Studies on PNP Suicide Gene Therapy of Hepatoma

To investigate the killing effect of PNP/MeP-dR suicide gene system on hepatoma cells, pcDNA3. O/PNP, an eukaryotic expression vector harboring E. coli PNP gene, was transfected into human hepatoma HepG2 cells by liposome-mediated method. A HepG2 cell line with stable PNP gene expression, HepG2/PNP,...

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Published in:	Journal of Huazhong University of Science and Technology. Medical sciences Vol. 25; no. 2; pp. 178 - 181
Main Author:	蔡晓坤周俊立林菊生孙雪梅薛秀兰李超
Format:	Journal Article
Language:	English
Published:	China Institute for Liver Disease of Tongji Hospital, Tongji School of Medicine, Huazhong University of Science and Technology, Wuhan 430030, China%Department of Microbiology, Tongji School of Medicine, Huazhong University of Science and Technology, Wuhan 430030, China 2005
Subjects:	Bystander Effect - physiology Carcinoma, Hepatocellular - therapy Cell Line, Tumor Escherichia coli Escherichia coli - enzymology Escherichia coli - genetics Genetic Therapy - methods Humans Liver Neoplasms - therapy PNP Prodrugs - chemistry Prodrugs - therapeutic use Purine-Nucleoside Phosphorylase - genetics Purine-Nucleoside Phosphorylase - metabolism Purines - chemistry Purines - therapeutic use 基因治疗实验研究肝细胞瘤自杀基因 gene therapy hepatoma PNP/MeP-dR suicide gene system
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Summary:	To investigate the killing effect of PNP/MeP-dR suicide gene system on hepatoma cells, pcDNA3. O/PNP, an eukaryotic expression vector harboring E. coli PNP gene, was transfected into human hepatoma HepG2 cells by liposome-mediated method. A HepG2 cell line with stable PNP gene expression, HepG2/PNP, was established with presence of G418 selection. The cell growth curves were determined with trypan blue staining. The sensitivity of HepG2/PNP to MePdR and bystander effects were assayed by MTT and FCM methods. The enzymatic activity of the product of PNP gene was determined by HPLC method. The cytotoxic effects of MeP-dR on HepG2/PNP cells were obvious (IC50 =4.5 μmol/L) and all HepG2/PNP cells were killed 4 days after the treatment with 100μmol/L MeP-dR. In mixed cultures containing increasing percentages of HepG2/PNP cells, total population killing was demonstrated when HepG2/PNP cells accounted for as few as 5 % of all HepG2 cells 8 days after the treatment with 100μmol MeP-dR. Highpressure liquid chromatography (HPLC) demonstrated that the PNP enzyme could convert MePdR into 6-MP. PNP/MeP-dR suicide gene system had an advantage over traditional suicide gene systems for hepatoma gene therapy. Our e results suggest that high-level bystander effects of this system result in significant anti-tumor responses to hepatoma gene therapy, especially in vivo.
Bibliography:	R735.7 42-1679/R ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	1672-0733 1993-1352
DOI:	10.1007/bf02873570