title: Gene therapy of pancreatic cancer targeting the K-Ras oncogene.
abstract: Ras mutations are present in ∼95% of pancreatic cancer (PC) cases leading to increased proliferation and apoptosis resistance. The aim of this study is to selectively kill Ras-transformed cells by overexpressing the pro-apoptotic protein, p53 upregulated modulator of apoptosis (PUMA) under a Ras-responsive promoter. Colo357, Panc1 and MiaPaca, PC cell lines harboring K-Ras mutations, normal rat IEC18 enterocytes, and their K-Ras transformed R1 counterparts, were tested. We constructed adenoviral vectors containing the PUMA gene downstream to: (1) Four or five repetitive Ras-responsive elements (Ad-PY4/PY5-PUMA) and (2) a negative control (Ad-SV40-PUMA). Cell viability was estimated by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and apoptosis was evaluated by FACS. In vivo potency of the adenoviruses was evaluated in athymic nude mice. Infection with Ad-PY4/PY5-PUMA markedly inhibited cell growth (∼40-50%), and apoptosis was detected in all cells with high Ras activity, whereas IEC18 cells remained unaffected. The control vector, Ad-SV40-PUMA, did not induce any cell death. Selective and high expression of PUMA was detected in Ad-PY4-PUMA-infected cells. In vivo, Ad-PY4-PUMA inhibited by ∼35% the growth of established tumors compared with the Ad-SV40-PUMA. Selective overexpression of PUMA efficiently inhibits the growth of Ras-transformed cells while sparing the normal ones. This treatment modality may become a useful, effective and safe approach to selectively target Ras-mutated tumor cells.
authors.affiliation: The Integrated Cancer Prevention Center, Tel Aviv Sourasky Medical Center, Tel Aviv, Israel.
authors.forename: V
authors.initials: V
authors.lastname: Lisiansky
authors.forename: I
authors.initials: I
authors.lastname: Naumov
authors.forename: S
authors.initials: S
authors.lastname: Shapira
authors.forename: D
authors.initials: D
authors.lastname: Kazanov
authors.forename: A
authors.initials: A
authors.lastname: Starr
authors.forename: N
authors.initials: N
authors.lastname: Arber
authors.forename: S
authors.initials: S
authors.lastname: Kraus
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis / Q000235:genetics; D048708:Cell Growth Processes / Q000235:genetics; D045744:Cell Line, Tumor; D005260:Female; D011905:Genes, ras; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D010190:Pancreatic Neoplasms / Q000235:genetics* / Q000473:pathology / Q000628:therapy*; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics; D016283:Proto-Oncogene Proteins p21(ras); D051381:Rats; D020218:Response Elements; D023041:Xenograft Model Antitumor Assays; D018631:ras Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D051017:Apoptosis Regulatory Proteins; C438378:BBC3 protein, human; C117307:KRAS protein, human; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins