The Effects Of Wild Type P53 And MDM2 Genes On The Growth Of Pancreatic Carcinoma Cells

There is increasing evidences that the carcinogenesis of cancers including pancreatic carcinoma is related to the accumulation of genetic abnormalities. Frequency of K-ras gene mutation in pancreatic cancer is very high,and in recent studies,p53 gene mutation is also found to be high in pancreatic cancer,which suggests that p53 gene mutation may be one of the genetic defects that plays a role in the pathogenesis of pancreatic cancer. Conventional methods of treatment for pancreatic carcinoma including surgery,radiotherapy and chemotherapy offer little hope of cure and 5-year survival rate is less than 5%. For exploiting new approaches of pancreatic cancer treatment,we studied the gene therapy of pancreatic cancer by transducing exogenic wild type p53 gene into the pancreatic cancer cells.Single-strand conformation polymorphism (SSCP) and DNA sequencing of the polymerase chain reaction (PCR) products techniques were used to determine p53 gene mutations in 5 pancreatic cancer cell lines established by our laboratory . Two of the 5 pancreatic cancer cell lines (PC-2,PC-4) have p53 mutations. PC-2 cell line was used for the gene transfer studies.(1). A recombinant retroviral vector expressing wild-type p53 (pBabe-swtp53) was constructed and packaged by packaging cell lines PA317 cells using calcium phosphate coprecipitation method. The supernatant of the virus producing cell was used to transfect the pancreatic carcinoma cell lines,PC-2,which contains p53 gene mutation at codon 240. Transduction with the retroviral vector resulted in integration and expression of wtp53 gene in PC-2 cells,verified by PCR and Northern blot analysis. The transfonnant cell line (PC-2/pBabe-swtp53) showed significant inhibition of cell growth, decrease of ~3H-TdR incorporation rate,soft-agar cology-formation and tumorigenesis in athymic nude mice,as compared with the control cell lines PC-2/pBabe-puro and PC-2 cell lines. These results suggest that retroviral wild-type p53 expression vector can be used to induce potential gene therapeutic effects in pancreatic cancer cell lines.(2). The Ad5CMVwtp53, a recombinant replication-deficient adenoviral vector containing a human CMV promoter,a human wild-type p53 and SV40 polyadenylation signal was amplified in 293 packaging cells. The PC-2 cell line was transfected by Ad5CMVwtp53 and the control adenoviral vector Ad5PXJ. The cells transfected with Ad5CMVwtp53...