Study on killing efficacy of yeast cytosine deaminase /5 - fluorocytosine gene therapy system on K562B cell line
- VernacularTitle:酵母菌胞嘧啶脱氨酶/5-氟胞嘧啶基因治疗系统对K562B细胞杀伤效应的初步研究
- Author:
Yusheng ZHANG
;
Jianmin WANG
;
Yongping ZHAI
- Publication Type:Journal Article
- Keywords:
cytosine deaminase,yeast;
fluorocytosine;
efficacy,killing;
vector,retroviral
- From:
Academic Journal of Second Military Medical University
1982;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To clone the yeast cytosine deaminase(YCD) gene and to elucidate the killing efficacy of yeast cytosine deaminase / 5 - fluorocytosine gene therapy system on gene- transfered cells. Methods:YCD gene was cloned and YCD expression retroviral vector was constructed,the vector was transfered into packaging cell line and high- titer virus was abtained. Then the tum origenic leukemia cell line K5 6 2 B was infected,selected and evaluated. Finally,the killing efficacy of 5 - FC on gene- transfered cell clone observed,and the 5 - FC to 5 - FU conversion of YCD- K5 6 2 B cell lysate was estim ated. Results:YCD gene was cloned and the sequence was confirm ed. A YCD expression retroviral vector,MSCV- YCD- IRES- EGFP was constructed. Transfering it into packaging cell line? NX- A ,high- titer (3.5? 10 6 CFU / ml) virus was obtained. The virus were used to infect tumorigenic leukem ia cell line,K5 6 2 B,and the infecting rate was quite high (30 % ) . A gene- transfered cell clone,YCD- K5 6 2 B,was selected,and YCD gene m RNA expression was detected in it.YCD- K5 6 2 B cell lysate demonstrated CD enzyme activity,and it could deaminase 5 - FC to 5 - FU .MTT assay showed 5 - FC could kill YCD- K5 6 2 B cell in 96 h even at lower concentration(15?mol/ L ) . Conclusion:YCD/ 5 - FC suicide gene therapy system has a significantkilling efficacy on gene- transfered cells. [
