Suppression of Pancreatic Cancer by the Dominant NegativerasMutant, N116Y

Shichinohe, Toshiaki; Senmaru, Naoto; Furuuchi, Keiji; Ogiso, Yoshifumi; Ishikura, Hiroshi; Yoshiki, Takashi; Takahashi, Toshiyuki; Kato, Hiroyuki; Kuzumaki, Noboru

doi:10.1006/jsre.1996.0383

Cited by 40 publications

(32 citation statements)

References 7 publications

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“…Human pancreatic cancer cell lines, PCI-19, PCI-24, PCI-35 and PCI-43, were established from surgically resected, primary carcinoma tissues. 2 Among these cell lines, PCI-35 and PCI-43 are CEA-producers. 24 These pancreatic cancer cell lines were maintained in Dulbecco's modified Eagle's medium (DMEM), supplemented with 10% fetal calf serum and 2 mm l-glutamine.…”

Section: Cell Linesmentioning

confidence: 99%

“…Each PCR reaction contained 2 l of RT reaction product as template DNA, 1 × PCR buffer (Boehringer), 160 m of each deoxynucleotide, 1 unit of Taq DNA polymerase (Boehringer), and 10 pmol of each 5Ј and 3Ј primer pair specific to N116Y cDNA. 2 Amplification conditions were 94°C for 1 min, 55°C for 30 s and 72°C for 1 min for 35 cycles. Equal aliquots (10 pmol) of secondary glyceraldehydephosphate dehydrogenase (GAPDH) primer sets were added at the sixth cycle (30 cycles remaining) by the primer-dropping method.…”

Section: Adenoviral Infection In Vitromentioning

confidence: 99%

“…[1][2][3][4][5][6] In particular, liver metastasis frequently occurs (about 60%) after a surgical resection for pancreatic adenocarcinoma even if it is performed with a curative intent. 5 No effective therapeutic modality has been established to prevent liver metastasis or to treat it successfully.…”

Section: Introductionmentioning

confidence: 99%

“…17 We have pursued another strategy of suppressing intracellular Ras function using a dominant negative Hras mutant, N116Y. 2,[18][19][20][21][22] It was derived from the v-H-ras oncogene by substituting asparagine with tyrosine at codon 116, which is in the GTP-binding domain. N116Y is thought to prevent the activation of oncogenic Ras protein with which it competes for a guanine nucleotide exchange factor.…”

Section: Introductionmentioning

confidence: 99%

“…18 N116Y expression significantly inhibited colony formation in selection medium when cotransfected into human pancreatic cancer cell lines with an oncogenic c-K-ras mutated at codon 12 (PCI-10, PCI-19, PCI-24, PCI-35, PCI-43, PCI-55, PCI-64, and PCI-66). 2 Moreover, the N116Y transfectants (PCI-35-N116Y) have been rendered nontumorigenic when injected subcutaneously into nude mice. Recently, we have reported that a recombinant N116Y expressing adenoviral construct driven by the cytomegalovirus promoter (Ad CMV-N116Y) suppressed the activation of extra-cellular signalrelated kinase 2 (Erk2) after EGF stimulation in serum starved esophageal cancer cells (TE8, SGF3 and SGF7).…”

Section: Introductionmentioning

confidence: 99%

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The dominant negative H-ras mutant, N116Y, suppresses growth of metastatic human pancreatic cancer cells in the liver of nude mice

et al. 2000

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In pancreatic cancer, the mutation of c-K-ras is a critical event of tumor growth and metastasis. We have previously demonstrated a dominant negative effect of N116Y on the growth of pancreatic cancer cells. To evaluate the potential of N116Y for suppressing the metastatic growth of pancreatic tumor cells, we made a replication-deficient recombinant N116Y adenovirus driven by the carcinoembryonic antigen (CEA) promoter (Ad CEA-N116Y). We demonstrated that the expression of N116Y, growth inhibition, and apoptotic death induction were all specific to pancreatic cancer cell lines (PCI-35 and PCI-43) that were promoter positive, whereas no growth retardation was observed in human embryonic pancreas-derived cell line 1C3D3 after Ad CEA-

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Section: Cell Linesmentioning

confidence: 99%

Section: Adenoviral Infection In Vitromentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

The dominant negative H-ras mutant, N116Y, suppresses growth of metastatic human pancreatic cancer cells in the liver of nude mice

et al. 2000

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Suppression of Erk activation andin vivo growth in esophageal cancer cells by the dominant negativeRas mutant, N116Y

et al. 1998

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Our previous studies demonstrated that introduction of a dominant negative H-ras mutant, N116Y, inhibits the growth of various types of cancer cells in vitro. In this study, we tested the efficacy of N116Y in blocking the growth of esophageal cancer cells using an adenoviral vector. Infection with N116Y adenovirus, (AdCMV-N116Y), in which N116Y expression is driven by the cytomegalovirus promoter, significantly reduced the in vitro growth of all esophageal cancer cell lines studied. Esophageal cancer cells that contained wild-type K-ras and H-ras (TE8, SGF3, SGF7) were more sensitive to AdCMV-N116Y than HEC46 cells that expressed mutant K-ras protein.Most importantly, direct injection of AdCMVN116Y into TE8-or SGF3-induced tumors in nude mice suppressed their growth significantly. To examine the suppressive mechanism of N116Y, cell cycle profile and the activation of extracellular signal-regulated kinase 2 (Erk2) were examined by ow cytometry and Western blot analysis, respectively. In TE8 cells, progression into S phase was clearly blocked after infection with AdCMV-N116Y. Infection with AdCMV-N116Y did not strongly suppress the activation of Erk2 after EGF stimulation in serum-starved HEC46 cells, whereas it completely suppressed activation in TE8, SGF3 and SGF7 cells. Our observations suggest that N116Y reduces growth of human esophageal cancer cells and suppresses the activation of Erk2; they also indicate that N116Y is a potential candidate gene for human esophageal cancer gene therapy.

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Adenovirus-mediated gene therapy for bladder cancer in an orthotopic model using a dominant negative H-ras mutant

et al. 2001

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It has been suggested that abnormal Ras function is important in the carcinogenesis and progression of bladder cancer. Our aim was to investigate the efficacy of transurethral inoculation of an adenovirus expressing the dominant negative H‐ras mutant N116Y against orthotopically implanted human bladder‐cancer cells in nude mice. We used a replication‐defective adenovirus vector containing the β‐galactosidase gene (AdCMV‐LacZ) as a control and the N116Y gene (AdCMV‐N116Y) as the therapeutic vector under the transcriptional control of the cytomegalovirus promoter. We initially investigated the in vitro growth‐suppressive effects of AdCMV‐N116Y on 2 human bladder‐cancer cell lines, KU‐7 and UMUC‐2. Thereafter, we examined the inhibitory effects of AdCMV‐N116Y on the 2 orthotopically implanted cell lines in nude mice. Intravesically created, orthotopic human bladder cancers were established in female KSN athymic nude mice with 1× 107 cancer cells. Then, 2, 3 and 4 days following implantation, 1 × 109 pfu of AdCMV‐LacZ or AdCMV‐N116Y were administered transurethrally. In vitro growth assays revealed significant growth suppression (>95%) with apoptosis of target cells treated with AdCMV‐N116Y compared to AdCMV‐LacZ. Transurethral inoculation of AdCMV‐N116Y into the bladder brought about a significant reduction in size (73% to 90%) and number (47% to 78%) of orthotopically implanted human bladder tumors compared to AdCMV‐LacZ or PBS. Normal mucosa in nude mice had minor inflammation with the infiltration of mononuclear cells. Our results suggest that gene therapy via transurethral inoculation of AdCMV‐N116Y holds promise for the treatment of human bladder cancer. © 2001 Wiley‐Liss, Inc.

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Suppression of Pancreatic Cancer by the Dominant NegativerasMutant, N116Y

Cited by 40 publications

References 7 publications

The dominant negative H-ras mutant, N116Y, suppresses growth of metastatic human pancreatic cancer cells in the liver of nude mice

The dominant negative H-ras mutant, N116Y, suppresses growth of metastatic human pancreatic cancer cells in the liver of nude mice

Suppression of Erk activation andin vivo growth in esophageal cancer cells by the dominant negativeRas mutant, N116Y

Adenovirus-mediated gene therapy for bladder cancer in an orthotopic model using a dominant negative H-ras mutant

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