The Feasibility Study On Gene Therapy Of Pancreatic Cacinoma With Ad-PUMA

Background:The extremely poor prognosis of patients with pancreatic carcinoma indicates the need for novel therapeutic approaches.Gene therapy is the new hope of pancreatic carcinoma treatments for the effect of typical therapies is not satisfactory at present.PUMA(p53 up-regulated modulator of apoptosis) is a potent proapoptotic molecule that is rapidly induced in cells following DNA damage and is required for p53-induced apoptosis.We evaluated the therapeutic potential of PUMA adenovirus against pancreatic carcinoma cell lines.Infection with Ad-PUMA(PUMA Adenovirus) resulted in the more powerful cytotoxicity in these cell lines compared with Ad-p53. Furthermore,we assessed the efficacy of a combined treatment with Ad-PUMA and anticancer drug(5-fluorouracil,cisplatin,gemcitabine hydrochloride,respectively) for these ceils and found PUMA significantly increased the chemosensitivity of pancreatic carcinoma cells,which may result from more abundant apoptosis induction.Interestingly, Ad-PUMA was found to be more efficient than Ad-p53 in inhibiting cell growth and enhancing the chemosensitivity of pancreatic carcinoma cell lines irrespective of the p53 expression level.These results suggest that Ad-PUMA is a potent cytotoxic agent and could be a promising alternative in the cancer gene therapy in combination with chemotherapeutic agents.Objective:To investigate the expression of PUMA and p53 protein and in cell lines derived from pancreatic ductal adenocarcinoma cancer.To study the effect of overexpression of PUMA on upstream p53 and downstream gene caspase-3 and parp in human pancreatic carcinoma cell line and effect of Ad-PUMA combined with chemotherapy on human pancreatic carcinoma cells.Methods:An adenovirus expressing PUMA(Ad-PUMA),alone or in combination with chemotherapeutic agents,was used to treat pancreatic carcinoma cells.The growth inhibitory and apoptotic effects of PUMA in vitro and in vivo were examined.The mechanisms of PUMA-mediated growth suppression and apoptosis were investigated through analysis of caspase activation and cleavage of PARE.The efficacy of PUMA and p53 in suppressing the growth of pancreatic carcinoma cells was also compared.Results:We have analyzed the biological activity of PUMA upon pancreatic ductal adenocarcinoma cancer-derived cell lines.We report that PUMA is variously expressed in cell lines derived from pancreatic ductal adenocarcinoma cancer and adenoviral-mediated expression of PUMA(Ad-PUMA) in these cells results in apoptosis via caspase activation and no cell-cycle change.And PUMA is more potent than p53 in growth suppression.RT-PCR and Western Blot revealed that the expression of exogenous Ad-PUMA was elevated 6 hours after infection.Furthermore,we assessed the efficacy of a combined treatment with Ad-PUMA and anticancer drugs(5-fluorouracil, cisplatin,gemcitabine hydrochloride,respectively) for Mia PaCa-2 and PANCI cell line and found that PUMA significantly increased the chemosensitivity of pancreatic carcinoma cell lines to chemotherapeutic agents,which may be resulted from abundant apoptosis induction.In Mia PaCa-2 xenograft models,Ad-PUMA alone and combined with chemotherapeutic significantly inhibited the growth of tumor.Conclusions:PUMA is more potent than p53 in growth suppression and chemosensitization in pancreatic carcinoma cells.PUMA-mediated growth suppression and apoptosis may be accomplished by activating procaspase-3 and cleaving PARP. Collectively,our results suggest that PUMA may play nagative roles in cancer cell growth and may be a promising molecule used for the cancer gene therapy in combination with chemotherapeutic agents.