Use of interfering RNA to investigate the role of endogenous gastrin in the survival of gastrointestinal cancer cells

Grabowska, Anna; Hughes, Jaime; Watson, Sue‐Ann

doi:10.1038/sj.bjc.6603588

Cited by 19 publications

(13 citation statements)

References 41 publications

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“…A universal negative control siRNA was used as a negative control (Eurogentec, Southampton, Hampshire, UK). Transfection was carried out using siPORT Amine siRNA transfection reagent (Ambion, Warrington, UK) following the manufacturer's instructions as described previously 23. After transfection, cells were cultured in a normal cell culture environment for 48 h before further experimentation.…”

Section: Methodsmentioning

confidence: 99%

Helicobacter pylori potentiates epithelial:mesenchymal transition in gastric cancer: links to soluble HB-EGF, gastrin and matrix metalloproteinase-7

Yin

Grabowska

Clarke

et al. 2010

Gut

106

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Background and aimsHelicobacter pylori (H pylori) infection is a major risk factor in the development of distal gastric adenocarcinoma. Development of the invasive phenotype is associated with the phenomenon of epithelial:mesenchymal transition (EMT). Soluble heparin-binding epidermal growth factor (HB-EGF) has been implicated in this process. A study was undertaken to investigate the possibility that matrix metalloproteinase (MMP)-7 is upregulated in H pylori infection as a result of hypergastrinaemia, which may enhance shedding of HB-EGF and contribute towards EMT in gastric adenocarcinoma cell lines.MethodsThree gastric epithelial cell lines (AGS, MGLVA1 and ST16) were co-cultured with the pathogenic H pylori strain 60190 and non-pathogenic strain Tx30a in an in vitro infection model. Gene expression was quantified by real-time PCR, HB-EGF shedding by ELISA and protein expression by immunofluorescence or immunohistochemistry. The INS-GAS mouse, a transgenic mouse model of gastric carcinogenesis which overexpresses amidated gastrin, was used to investigate the in vivo relationship between HB-EGF, MMP-7, gastrin and EMT.ResultsThe pathogenic strain of H pylori significantly upregulated EMT-associated genes Snail, Slug and vimentin in all three gastric cell lines to a greater degree than the non-pathogenic strain. Pathogenic H pylori also upregulated HB-EGF shedding, a factor implicated in EMT, which was partially dependent on both gastrin and MMP-7 expression. Gastrin and MMP-7 siRNAs and MMP-7 neutralising antibody significantly reduced upregulation of HB-EGF shedding in H pylori infected gastric cell lines and reduced EMT gene expression. The effect of H pylori on EMT was also reversed by gastrin siRNA. Neutralisation of gastrin in the INS-GAS mouse model reduced expression of MMP-7, HB-EGF and key EMT proteins.ConclusionThe upregulation of MMP-7 by pathogenic H pylori is partially dependent on gastrin and may have a role in the development of gastric cancer, potentially through EMT, by indirectly increasing levels of soluble HB-EGF.

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Section: Methodsmentioning

confidence: 99%

Helicobacter pylori potentiates epithelial:mesenchymal transition in gastric cancer: links to soluble HB-EGF, gastrin and matrix metalloproteinase-7

Yin

Grabowska

Clarke

et al. 2010

Gut

106

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“…Progastrin growth-promoting effects are mediated through the activation of several key signal transduction pathways (21,22). In addition, several works including ours showed that depletion of endogenous progastrin produced by colon cancer cells leads to an inhibition of tumor growth (23,24). Moreover, high concentrations of progastrin are found in colon tumors and in blood of 80% of patients with colorectal cancer.…”

Section: Introductionmentioning

confidence: 99%

A New Biomarker That Predicts Colonic Neoplasia Outcome in Patients with Hyperplastic Colonic Polyps

Castro

Palasse

et al. 2012

Cancer Prevention Research

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The most frequently occurring lesions in the colon are the hyperplastic polyps. Hyperplastic polyps have long been considered as lesions with no malignant potential and colonoscopy for these patients is not recommended. However, recent works suggest that hyperplastic polyps may represent precursor lesions of some sporadic colorectal cancers. Until now, no biomarker allows to identify the subset of hyperplastic polyps that may have a malignant potential. Because the hormone precursor progastrin has been involved in colon carcinogenesis, we investigated whether its expression in hyperplastic polyps predicts the occurrence of colonic neoplasm after resection of hyperplastic polyps. We retrospectively analyzed progastrin expression in hyperplastic polyps from 74 patients without history of colorectal pathology. In our study, 41% of patients presenting an initial hyperplastic polyp subsequently developed adenomatous polyps, recognized as precursor lesions for colorectal adenocarcinomas. Progastrin was overexpressed in the hyperplastic polyps in 40% of the patients. We showed a significant association between progastrin overexpression and shortened neoplasm-free survival (P ¼ 0.001). Patients with high overexpression of progastrin had a 5-year neoplasm-free survival rate of 38% as compared with 100% for the patients with low progastrin expression. In addition, we established a predictive test on the basis of progastrin staining and patients' age that predicts occurrence of neoplasm after developing a first hyperplastic polyp with a sensitivity of 100% [95% confidence interval (CI), 79%-100%] and a specificity of 74% (51%-90%). We show that progastrin expression evaluation in hyperplastic polyps is an efficient prognostic tool to determine patients with higher risk of metachronous neoplasms who could benefit from an adapted follow-up. Cancer Prev Res; 5(4);

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“…When combined with gemcitabine, a newer orally active gastrin receptor antagonist (Z-360) has shown modest efficacy against orthotopic pancreatic tumors in mice 30. Transient knockdown of gastrin gene expression by transfecting pancreatic cancer cells with antisense gastrin constructs 31 or with gastrin siRNA 32 decreased gastrin levels and tumor cell proliferation in vitro .…”

Section: Introductionmentioning

confidence: 99%

Growth of Human Pancreatic Cancer Is Inhibited by Down-Regulation of Gastrin Gene Expression

Matters¹,

Harms²,

McGovern³

et al. 2009

Pancreas

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Objectives This study evaluated the effects of gastrin mRNA down-regulation on growth of human pancreatic cancer. Methods Gastrin expression was examined in human pancreatic cancer cell lines by RT-PCR and peptide expression was assessed by immunocytochemistry. Gastrin was down-regulated using either stable transfection of an antisense gastrin cDNA or one of three shRNA (short hairpin RNA) constructs. Tumor formation was evaluated following either subcutaneous or orthotopic injections into nude mice. The effect of nanoliposomes loaded with gastrin siRNA was tested in mice bearing pancreatic tumors. Results Stable transfection of gastrin antisense or shRNAs into BxPC-3 cells resulted in clones with >90% reduction in gastrin mRNA. Tumor growth rate and incidence of metastases in both wild type and transfected pancreatic cancer cells was directly proportional to the degrees of gastrin mRNA expression. Immunofluoresence analysis confirmed that gastrin peptide levels were decreased in antisense and shRNA tumors. Gastrin knockdown clones had lower Ki-67 and increased cleaved caspase-3 staining, consistent with known effects of gastrin on proliferation and apoptosis. Tumors in mice treated with gastrin siRNA were smaller than controls. Conclusions These results suggest that RNAi targeting of gastrin could serve as an effective treatment for pancreatic cancer.

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Use of interfering RNA to investigate the role of endogenous gastrin in the survival of gastrointestinal cancer cells

Cited by 19 publications

References 41 publications

Helicobacter pylori potentiates epithelial:mesenchymal transition in gastric cancer: links to soluble HB-EGF, gastrin and matrix metalloproteinase-7

Helicobacter pylori potentiates epithelial:mesenchymal transition in gastric cancer: links to soluble HB-EGF, gastrin and matrix metalloproteinase-7

A New Biomarker That Predicts Colonic Neoplasia Outcome in Patients with Hyperplastic Colonic Polyps

Growth of Human Pancreatic Cancer Is Inhibited by Down-Regulation of Gastrin Gene Expression

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