›› 2009, Vol. 29 ›› Issue (7): 802-.

• Original article (Basic research) • Previous Articles     Next Articles

Construction and application of conditionally replicative adenovirus for selective cytotoxicity in CEA positive colorectal cancer cells

WANG Meng-yun, WEI Fang, WANG Hui-ping, JI Xun-da, CHEN Xia-fang, LI Hui-ming, FENG Ye, WANG Yu-ei, HUANG Qian   

  1. Experimental Center, The First People's Hospital, Shanghai Jiaotong University, Shanghai 200080, China
  • Online:2009-07-25 Published:2009-09-16
  • Supported by:

    National Basic Research Program of China, 973 program, 2004CB518804; National Natural Science Foundation of China, 30325043, 30428015

Abstract:

Objective To construct a new conditionally replicative adenovirus (CRAds) targeting carcinoembryonic antigen (CEA) positive colorectal cancer cells. Methods The DNA fragment of the CEA gene promoter was amplified through PCR and cloned into the vector carrying fusion reporter gene EGFP-Luc to construct expression plasmid pCEA-EGFP-Luc. The constructed plasmid pCEA-EGFP-Luc was transfected into CEA positive and negative cells by liposome. The activity of CEA gene promoter was evaluated by detecting the expression of EGFP and luciferase activity. The conditionally replicative adenovirus Ad.CEA-E1A/CMV-TK carrying suicide gene HSVtk was constructed, in which the E1A gene was controlled under CEA promoter. CEA positive(Lovo and SW620)and negative tumor cells(HeLa)were infected with Ad.CEA-E1A/CMV-TK. The selective cytotoxicity of Ad.CEA-E1A/CMV-TK and the synergistic effect of the virus with GCV in CEA positive tumor cells were evaluated by the expression of E1A, cytopathic effect and cell survival rate. Results CEA promoter possesses a good specificity as well as high activity. The expression of E1A only presented in CEA positive tumor cells. After infection with Ad.CEA-E1A/CMV-TK, the cell survival rates of Lovo and SW620 were (36.72±2.49)% and (39.82±4.76)%, respectively, significantly lower than that of Hela[(87.44±2.76)%](P<0.01). When combined with GCV, Ad.CEA-E1A/CMV-TK had better oncolytic effect on Lovo and SW620 cells, with cell survival rates of (17.26±3.65)% and (23.93±5.40)%, respectively, significantly lower than those without GCV\[(36.72±2.49)% and (39.82±4.76)%, respectively](P<0.01). Conclusion Ad.CEA-E1A/CMV-TK under the control of CEA promoter has selective cytotoxic effect on CEA positive colorectal cancer cells, and the effect can be enhanced when combined with GCV.

Key words: colorectal cancer, carcinoembryonic antigen, promoter, recombinant adenovirus, E1A gene, HSVtk suicide gene