MTERT Promoter-driven M4-1BBL Gene Targeting Immunotherapy Of Hepatocellular Carcinoma In Mice In Vivo Study

Object:To construct a recombinant adenoviral vector carrying mTERT promoter and m4-1BBL gene.To observe the growth of hepatocellular carcinoma(HCC) in vivo,and evaluate the effect of m4-1BBL gene under the control of Mouse Telomerase Reverse Transcriptase Gene Promoter (mTERTpromoter) on HCC.Methods:A recombinant adenoviral plasmid(pAd-mTERT-promoter- m4-1BBL)containing m4-1BBL gene under the mTERT promoter was constructed by the homologous recombination method in bacteria. We made the adenoviruses (pAd-mTERT-promoter- m4-1BBL) packaged and amplified in the QBI 293A cells.We could get recombinant adenoviruses rAd-mTERTpromoter-m4-1BBL . And the same we get recombinant adenoviruses rAd.We make recombinant adenoviruses(rAd-mTERT promoter-m4-1BBL,rAd)infecting the mouse hepatocellular carcinoma cell line hepa1-6.RT-PCR was used to detect the m4-1BBLgene and mTERT promoter gene in hepa1-6 cells. A mouse hepatocellular carcinoma model was successfully established in the C57BL/6 mice.We observed the weight,volume of the mice and the antitumor for eight weeks.We use immunohistochemical dectected the expression of the m4-1BBL gene and mTERT promoter gene in tumour.Results : We successfully constructed a recombinant adenoviruse containing mTERT-promoter gene and m4-1BBl gene.The viral titer was 1×1010pfu/ml.The replication-deficient adenovirus liquid can use for treatment.In vitro, rAd-mTERT promoter- m4-1BBL induced Hepa1-6 cells effectly,beside this result,the growth rate of HCC significantly reduced CD8+cells expressing increased and the survivals prolonged in the rAd-mTERT promoter-m4-1BBL-infected C57BL/6 mice.Conclusion:The m4-1BBL gene under the control of mTERTpromoter gene can effectly inhibit mouse HCC cells growth and promote the apoptosis of Hepa1-6 cells,and effectively Extend the survival of tumor-bearing mice period, there is a good effect of targeted immunotherapy.