Effects of
CD/5-FC Suicide Gene Therapy System
on Human Malignant Glioma Cells in Vitro
LV
Sheng-Qing1,2*, YANG Hui2, HE Jia-Quan2,
WANG Bin2, YOSHIMURA Ichiro3, LIU Yun-Sheng1*
( 1 Department of Neurosurgery, Xiangya Hospital, Central South
University, Changsha 410008, China;
2 Department of Neurosurgery,
Xinqiao Hospital, Third Military Medical University, Chongqing 400037, China;
3 Department of Urology,
National Defense Medical College, 3-2 Namiki, Tokorozawa, Saitama 359-8513,
Japan )
Abstract The purpose of this paper is to
investigate the antitumor effects of cytosine deaminase/5-fluorocytosine
(CD/5-FC) suicide gene therapy system on human malignant glioma cells in
vitro. The pCMVCD plasmid was constructed through the CD gene insertion in
the multicloning site of eukaryotic expression vector pcDNA3.0, and confirmed
by restriction endonuclease digestion/gene sequencing. The construct was
subsequently transfected into the U251 human malignant glioma cells by using
LipofectAMINE2000-mediated method. Resistant clones (named U251/CD cells) were
isolated by screening with G418 presence. U251/CD cells were incubated with
5-FC in different concentrations to determine viability ratios (or cytotoxicity
assay), measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide (MTT) assay. The concentrations of 5-fluorouracil (5-FU) in the media
were measured by high-performance liquid chromatography (HPLC) detector. Our results
suggested that the untreated U251 cells were insensitive to 5-FC, with the IC
Key words cytosine deaminase; 5-fluorocytosine; gene therapy; malignant glioma
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