The expression of adenoviral mediated gfp reporter gene in tumor cells induced and regulated by irradiation

In order to establish a radiation inducible gene expression system for cancer gene therapy, the promoter sequence of radiation-inducible Egr-1 gene was amplified front genomic DNA of BALB/c mouse with PCR method, and linked to gfP reporter gene. Then the pEgr-gfp expression cassette was subcloned into an adenoviral shuttle plasmid to generate recombinant adenovirus of AdEgr-GFP by using a novel, high efficient method of homologous recombination in bacteria. After infection with AdEgr-GFP, MM45T. Li tumor cells were exposed to different doses of gamma -irradiation from. 0 Gy to 15 Gy in vitro. The percentage of GFP expression positive cells increased greatly, in a dose-dependent manner as detected by FAGS and Western blot analysis. For in vivo study, AdEgr-GFP were injected intratumorally, and tumor site received different doses of local gamma -irradiation 45 h after injection, and after 8 h the tumor samples were biopsed for investigating the GFP expression. Tumor tissue image analysis revealed that gamma -irradiation could markedly increase GFP expression in a dose-dependent manner as compared with chat of non-irradiated control group. Our results indicate that the irradiation can effectively control adenoviral-mediated GFP expression in tumor cells via Egr-1 promoter, and these data laid basis for further gene radiotherapy study.