Construction Of Adenoviral Vector Encoding GLI1siRNA And Its Effect In The Treatment Of Pancreatic Cancer

The purpose of the present study is to explore the therapeutic effect of adenovirus encoding GLI1siRNA on human pancreatic cancer through combining RNA interference technique and gene therapy. First,RT-PCR was used to detect expression of GLI1 and PTCH1mRNA in five pancreatic cancer cell lines and the RT-PCR products were sequenced and demonstrated to have the same sequences as those of GLI1 and PTCH1 in GenBank.Then an expression cassette encoding GLI1siRNA was inserted into pAdTrack and the resulting recombinant pAdTrack-U6-GLI1siRNA was used to electrotransform BJ5183-AdEasy-1 to form pAd-U6-GLI1siRNA,which was packaged in 293 cells to yield Ad-U6-GLI1siRNA.In vitro experiment,quantitative real-time PCR and Western blot were performed to measure the expression of GLL1 mRNA and protein in cells treated with Ad-U6-GLI1siRNA.MTT and flow cytometry were performed to measure cellular proliferation,cell cycle and apoptosis.Finally, two pancreatic tumor xenograft models were established and treated with Ad-U6-GLI1siRNA for further study of its anti-tumor effect in vivo.Our study show that all five pancreatic cancer cell lines expresses GLI1 and PTCH1 mRNA and Ad-U6-GLI1siRNA effectively inhibits the expression of GLI1 mRNA and protein in pancreatic cancer cell lines. Ad-U6-GLI1siRNA efficiently suppresses the growth of pancreatic cancer in vitro and in vivo without remarkable adeverse effect,showing a wide indication and a promising future in its application.