The Study Of Tumor-specific TK Gene Therapy Induced By HTERT Promoter For Human Nasopharyngeal Carcinoma Cells

PURPOSE: To construct human Telomerase Reverse TranscriptasePromoter/TK specific tumor-specific expressing system, and investigateits directional effect of killing and wounding the Nasopharyngealcarcinoma tumor cells.METHODS: The hTERT promoter and TK gene were cloned, andwhich was connected to the PGL3 carrier after enzyme digestion, toconstruct the hTERTp/PGL3,TK/PGL3,hTERTp/TK/PGL3 recombinantcarriers, and then to construct the CMV/TK/PGL3 recombinant carrierswith the similar method and regard CMV as the positive control. ThehTERTp/PGL3,TK/PGL3,hTERTp/TK/PGL3 and CMV/TK/PGL3 weretransfected into the nasopharyngeal carcinoma HNE1 cells and thenormal fibroblast transiently, with the application of bangosome throughRT-PCR. After 24 hours, it was added in GCV, to examine the appearanceof the cell, and detect the death of nasopharyngeal carcinoma cell. After48 hours, the cell was harvested to detect the expression of telomeraseand its subunit hTERT and TK gene in the transfection cell, and detect thecytoactive with MTT.RESULTS: There was a successful construction in identifyhTERTp/PGL3,TK/PGL3,hTERTp/TK/PGL3 and CMV/TK/PGL3recombinant plasmid by double digestion with restriction endonucleases.The luciferase activity test manifested that the activity of CMV/PGL3 issuperior to hTERTp/PGL3, the effect of killing NPC HNE1 cell ofhTERTp/TK/PGL3 is higher than the hTERTp/PGL3, TK/PGL3 controlgroup, but lower than the positive group. The expressions of TK genewere higher than the hTERTp/PGL3, TK/PGL3 control groups, and theexpression of telomerase and its subunit hTERTp were lower than thepositive control groups.CONCLUSION: hTERTp/TK, a tumor-specific expressing system,has a special killing effect to nasopharyngeal carcinoma cell in vitro.