Application of tetracycline inducible promoter based CRAd to cell vehicle system

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The aim of this project is to establish the cell vehicle system for the viral vector carrier. To do this, drug inducible promoter based conditionally replicative adenovirus was made at first. Tetracycline inducible promoter was chosen to switch the promoter activation easily. The tetracycline inducible promoter based replicative adenovirus, AdTRE-E1, was constructed by the replacement of wild type El promoter with tetracycline inducible promoter. AdTRE-E1 proliferated in the infected human fibroblast culture cells (WI38) in the presence of tetracycline in the culture medium. Contrary, the virus was silent in the absence of tetracycline. Therefore, addition of tetracycline in the medium functioned as a switch for viral replication initiation. In the next step, telomerase promoter based replicative adenovirus (AdTERT-E1) was used with the tetracycline inducible system for cancer treatment. WI38 cells were co-infected with AdTRE-E1 and AdTERT-E1 simultaneously, and then tetracycline was added into the culture medium. After 72 hours from infection, the viruses proliferated rapidly and destroyed the infected cells and were released into the medium. Apart of the medium containing viruses was transferred to the intact cancer cell culture flask. AdTERT-E1 infected into the cancer cells and proliferated in the cells independently without tetracycline. As a result, infected WI38 cells carried the viruses in the cells keeping the virus ability of its infectivity. The tetracycline inducible system functioned in the peritoneal space in the nude mouse. When the infected WI38 cells were implanted into the mouse intraperitoneally, viruses proliferated in the peritoneal space after feeding of tetracycline in the drinking water. Therefore, this system functioned in vivo as well as in vitro, and was possible for the cell vehicle system for virus carrier.