elafin and Stomach-Neoplasms

The purposes of this study were to determine the expression profiles of microRNA-34a (miR-34a) in human gastric cancer cell line (SGC-7901) and cisplatin-resistant cell lines (SGC-7901/DDP), and to establish the correlation between miR-34a expression profile and the sensitivity of human gastric cancer cell to cisplatin-based pattern, thereby providing new methods and strategies for treating gastric cancer. Gastric cancer cell line (SGC-7901) and cisplatin-resistant cell line (SGC-7901/DDP) were cultivated in vitro, respectively. Quantitative real-time PCR (qRT-PCR) and Western blot were utilized to determine the expression profiles of miR-34a and survivin in both gastric cancer cell lines. With miR-34a mimic and miR-34a inhibitor transfected into SGC-7901 and SGC-7901/DDP for 48 h, post-transfection changes of miR-34a expression was determined; the effects of miR-34a ectopic expression on the viability of cisplatin-induce gastric cancer cell were assayed by the MTT method. The effects of miR-34a ectopic expression on apoptosis of cisplatin-induce gastric cancer cell were determined by Annexin V/propidium iodide (PI) double staining method and flow cytometry. The effects of miR-34a ectopic expression on the AKT and p-AKT expression of cisplatin-induce gastric cancer cells were determined by Western blot and flow cytometry with the PI3K pathway inhibitor Wortmannin. As shown by qRT-PCR and Western blot analyses, the expression of miR-34a in cisplatin-resistant cell lines decreased significantly in comparison to that of SGC-7901 cell line (p < 0.05), while significant up-regulation of survivin expression was also observed (p < 0.05). Compared with the control group, the expression of miR-34a increased significantly in SGC-7901 cells transfected with miR-34a mimic for 48 h (p < 0.01). After miR-34a inhibitor transfection, the expression of miR-34a decreased significantly (p < 0.05). The viability of cisplatin-induce gastric cancer cells increased significantly (p < 0.05) with significant decrease of apoptosis after miR-34a expression inhibition, as demonstrated by MTT and flow cytometry with miR-34a over-expression, the viability of cisplatin-induce gastric cancer cells decreased significantly (p < 0.05), with significant apoptosis increase (p < 0.05). As shown by Western blot and flow cytometry, in comparison to the control group, Wortmannin could inhibit miR-34a inhibitor and DDP induced up-regulation of p-AKT significantly (p < 0.05) and stimulated apoptos

Topics: Animals; Apoptosis; Cell Proliferation; Cisplatin; Elafin; Female; Gene Expression Regulation, Neoplastic; Humans; Mice; MicroRNAs; Proto-Oncogene Proteins c-akt; Signal Transduction; Stomach Neoplasms

G-protein coupled receptor 34 (GPR34), which belongs to the G-protein coupled receptors superfamily, is reportedly expressed highly in the spread of several solid tumors. However, its expression in gastric primary tumor and potential role in gastric cancer development and progression have not been determined.. Immunohistochemistry, real-time RT-PCR and western blot methods were used to determine GPR34 expression in human gastric cancer tissues/cell lines and matched adjacent tissues/ normal mucosal cell line. A statistical analysis was performed to establish the potential correlation between GPR34 expression and the patients' clinicopathological characteristics, tumor progression, and prognosis. Stably transfected NCI-N87 cell lines with either GPR34 over-expression or knock-down were constructed to determine the effect of GPR34 on gastric cancer cell invasion and migration, and to explain the preliminary molecular mechanism of GPR34 in gastric cancer metastasis.. GPR34 is up-regulated in primary gastric cancer tissues/cell lines compared with matched adjacent tissues/normal mucosal cell line, and when the relationship between GPR34 expression and the the clinicopathological characteristics was analyzed, it was shown that GPR34 expression is significantly correlated with tumor differentiation, infiltration depth, and lymph node status and had a significant influence on prognosis. Furthermore, GPR34-overexpression increased while GPR34-knockdown inhibited NCI-N87 cell invasion in vitro by PI3K/PDK1/AKT pathway.. Taken together, up-regulation of GPR34 expression in human gastric carcinoma may play a critical role in tumor progression and in determining patient prognosis. GPR34 may be a useful diagnostic or prognostic molecular biomarker, and a potential target for therapeutic intervention.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Blotting, Western; Disease Progression; Elafin; Female; Humans; Immunohistochemistry; Male; Middle Aged; Prognosis; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-akt; Pyruvate Dehydrogenase Acetyl-Transferring Kinase; Real-Time Polymerase Chain Reaction; Receptors, Lysophospholipid; Retrospective Studies; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Stomach Neoplasms; Up-Regulation; Young Adult

The cyclin-dependent kinase (CDK) inhibitor p27 has an important role in cell cycle regulation. Reduced expression of p27 is commonly associated with poor prognosis in many malignancies, including gastric cancer. Cytoplasmic p27 mislocalization may be an additional indicator of high-grade tumors and poor prognosis in cancer. As chronic infection by Helicobacter pylori is the most important risk factor for gastric cancer development, we evaluated the effects of H. pylori on p27 expression and localization in gastric cancer cells. Co-culture of gastric cells with H. pylori induced cytoplasmic p27 expression and reduced nuclear p27 expression in vitro. Cytoplasmic p27 expression was associated with and dependent upon phosphorylation of p27 at T157 and T198: wild-type p27 accumulated in the cytoplasm, but non-phosphorylatable mutants affecting T157 or T198 were nuclear in H. pylori-infected cells. These post-translational p27 changes were secondary to activation of cellular phosphoinositide-3 kinase (PI3K) and AKT signaling pathways, and dependent upon a functional H. pylori cag pathogenicity island. We investigated the clinical significance of cytoplasmic p27 mislocalization in 164 cases of resected gastric cancer in tissue microarrays. In 97 cases (59%), cytoplasmic p27 mislocalization was observed, and this was associated with increased mortality in multivariate analysis. These results show that H. pylori infection induces AKT/PI3K-mediated phosphorylation of p27 at T157 and T198 to cause cytoplasmic p27 mislocalization in gastric cancer, and that p27 mislocalization is an adverse prognostic feature in gastric cancer. This is the first demonstration of the translocation of a specific bacterial virulence factor that post-translationally regulates a host cell CDK inhibitor. This is of particular significance, because p27 has both tumor-suppressive and oncogenic activities, depending upon its subcellular localization. Cytoplasmic mislocalization of p27 induced by H. pylori may be an important mechanistic link between H. pylori infection and gastric carcinogenesis.

Topics: Cell Line, Tumor; Coculture Techniques; Cyclin-Dependent Kinase Inhibitor p27; Cytosol; Elafin; Female; Helicobacter Infections; Helicobacter pylori; Humans; Male; Phosphorylation; Prognosis; Stomach Neoplasms

Therapies targeting receptor tyrosine kinases have shown efficacy in molecularly defined subsets of cancers. Unfortunately, cancers invariably develop resistance, and overcoming or preventing resistance will ultimately be key to unleashing their full therapeutic potential. In this study, we examined how cancers become resistant to MET inhibitors, a class of drugs currently under clinical development. We utilized the highly sensitive gastric carcinoma cell line, SNU638, and two related MET inhibitors PHA-665752 and PF-2341066. To our surprise, we observed at least two mechanisms of resistance that arose simultaneously. Both resulted in maintenance of downstream PI3K (phosphoinositide 3-kinase)-AKT and MEK (MAP/ERK kinase)-ERK signaling in the presence of inhibitor. One mechanism, observed by modeling resistance both in vitro and in vivo, involved the acquisition of a mutation in the MET activation loop (Y1230). Structural analysis indicates that this mutation destabilizes the autoinhibitory conformation of MET and abrogates an important aromatic stacking interaction with the inhibitor. The other cause of resistance was activation of the epidermal growth factor receptor (EGFR) pathway due to increased expression of transforming growth factor α. Activation of EGFR bypassed the need for MET signaling to activate downstream signaling in these cells. This resistance could be overcome by combined EGFR and MET inhibition. Thus, therapeutic strategies that combine MET inhibitors capable of inhibiting Y1230 mutant MET in combination with anti-EGFR-based therapies may enhance clinical benefit for patients with MET-addicted cancers. Importantly, these results also underscore the notion that a single cancer can simultaneously develop resistance induced by several mechanisms and highlight the daunting challenges associated with preventing or overcoming resistance.

Topics: Animals; Cell Line, Tumor; Crizotinib; Drug Resistance, Neoplasm; Elafin; ErbB Receptors; Extracellular Signal-Regulated MAP Kinases; Humans; Indoles; MAP Kinase Kinase Kinases; Mice; Mice, Nude; Mutation; Oncogene Protein v-akt; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-met; Pyrazoles; Pyridines; Receptors, Growth Factor; Stomach Neoplasms; Sulfones; Transcription Factors; Transforming Growth Factor alpha; Xenograft Model Antitumor Assays