Microarray analysis of gene expression in gastric cancer cells from ascitic fluids before and after capecitabine and oxaliplatin (CapeOX).

60 Background: Resistance to anti-cancer drugs is a critical issue in treatment of cancer. Alteration of gene expression profile accompanied with acquisition of drug-resistance is considered to be related with resistance mechanisms. Methods: 1. Ascitic fluids were collected before the beginning of CapeOX, during the treatment was effective, and after the disease was progressed. They were cultured for ten days and passed to new flasks, and were cultured in additional two weeks to remove normal cells including white blood cells and mesothelial cells. 2. AGS cells were cultured in medium with either fluorouracil or oxaliplatin, and fluorouracil- or oxaliplatin-resistant cells were established. AGS cells were also treated with a demethylating agent decitabine. 3. Expression of 25,147 genes was analyzed with microarray, and was compared among ascitic fluid cells before and after CapeOX, drug-resistant AGS cells, and decitabine-treated AGS cells. Results: Genes with alteration in expression due to CapeOX-resistance were classified into genes commonly changed with fluorouracil-resistant cells, those with oxaliplatin-resistant cells, and others. Among 873 genes with decreased expression after CapeOX, some genes including PCDH20 and DEFB4A have been reported as potential tumor suppressor genes. Expression of PCHD20 in AGS increased after treatment with decitabine. Conclusions: Drug-resistance in gastric cancer may be related with expression of genes including tumor suppressors regulated by epigenetic mechanisms. Repetitive collection of gastric cancer cells in ascitic fluids before and after chemotherapy is useful to understand drug-resistance mechanisms.

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Alterations in gene expression and DNA methylation profiles in gastric cancer cells obtained from ascitic fluids collected before and after chemotherapy

Molecular and Clinical Oncology ◽

10.3892/mco.2019.1858 ◽

2019 ◽

Author(s):

Osamu Maeda ◽

Ayumu Matsuoka ◽

Kazuhiro Furukawa ◽

Ryoji Miyahara ◽

Yoshiki Hirooka ◽

...

Keyword(s):

Gene Expression ◽

Gastric Cancer ◽

Dna Methylation ◽

Cancer Cells ◽

Gastric Cancer Cells ◽

Before And After

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TSPAN9 suppresses the chemosensitivity of gastric cancer to 5-fluorouracil by promoting autophagy

Cancer Cell International ◽

10.1186/s12935-019-1089-2 ◽

2020 ◽

Vol 20 (1) ◽

Cited By ~ 4

Author(s):

Yaoyue Qi ◽

Weiwei Qi ◽

Shihai Liu ◽

Libin Sun ◽

Aiping Ding ◽

...

Keyword(s):

Gastric Cancer ◽

Drug Resistance ◽

Cancer Cells ◽

Transmembrane Protein ◽

Pi3k Pathway ◽

Drug Resistant ◽

Gastric Cancer Cells ◽

Autophagy Pathway ◽

Resistant Cells

Abstract Background The issue of drug resistance in gastric cancer has attracted global attention. TSPAN9, a 4-transmembrane protein that plays an important role in tumor progression and signal transduction, has been found to be closely related to tumor invasion, metastasis, and autophagy. Methods Immunoblotting was used to evaluate TSPAN9 expression in parental and drug-resistant gastric cancer cells. Functional assays, such as the CCK-8 assay, were used to detect the proliferation of gastric cancer cells and the response of TSPAN9 to 5-fluorouracil (5-FU). Western blotting was used to analyze the expression of constituents of the PI3K/AKT/mTOR-mediated autophagy pathway induced by TSPAN9. Coimmunoprecipitation was performed to assess the specific mechanism by which TSPAN9 affects the PI3K pathway. Results We demonstrated that TSPAN9 is overexpressed in 5-FU-resistant cells compared to parental cells. 5-FU-mediated inhibition of cell proliferation can be significantly restored by increasing TSPAN9 expression, and inhibiting this expression in drug-resistant cells can restore the sensitivity of the cells to 5-FU. In addition, TSPAN9 also significantly promoted autophagy in gastric cancer cells in vitro. Further studies indicated that TSPAN9 downregulates the expression of PI3K and proteins associated with PI3K-mediated autophagy. In addition, TSPAN9 interacts with PI3K and inhibits its catalytic activity. Conclusion The current study reveals the important role of TSPAN9 in drug resistance to 5-FU in gastric cancer. It also provides a new target to clinically address drug-resistant gastric cancer and will contribute to the treatment strategy of this disease.

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Effects of siRNA-mediated silencing of Bmi-1 gene expression on proliferation of gastric cancer cells

European Journal of Inflammation ◽

10.1177/2058739219845534 ◽

2019 ◽

Vol 17 ◽

pp. 205873921984553

Author(s):

Ying Guo ◽

Li Zhang ◽

Guangyu Zhou ◽

Qingjie Ma ◽

Shi Gao ◽

...

Keyword(s):

Gene Expression ◽

Gastric Cancer ◽

Flow Cytometry ◽

Cancer Cells ◽

Cancer Cell ◽

Gastric Cancer Cell ◽

Negative Control ◽

Control Group ◽

Gastric Cancer Cells ◽

Ags Cells

This study was designed to investigate the effects of siRNA-mediated silencing of Bmi-1 gene expression on proliferation of AGS gastric cancer cell. siRNA Bmi-1 was transfected into human AGS gastric cancer cells by liposome (as siRNA Bmi-1 group) with negative control (as control group); the expressions of Bmi-1 and apoptosis-related genes like P21, Bax, and Bcl-2 in AGS cells were determined by Western blot method; the apoptosis of AGS cells was detected by flow cytometry double staining and Hoechst staining; and cell cycle was measured by flow cytometry. Compared with the control group, the expression of Bmi-1 in the siRNA Bmi-1 group was significantly decreased ( P < 0.05), the apoptosis rate was increased ( P < 0.05), and cell cycles were arrested at G1 phase (P < 0.05); the expression level of P21 and Bax in cells was significantly up-regulated while that of Bcl-2 down-regulated ( P < 0.05). The down regulation of Bmi-1 can inhibit the proliferation of AGS gastric cancer cell and promote its apoptosis, which takes such effects mainly by up-regulating P21 as well as Bax and down-regulating Bcl-2.

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The CAGE–MiR-181b-5p–S1PR1 Axis Regulates Anticancer Drug Resistance and Autophagy in Gastric Cancer Cells

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.666387 ◽

2021 ◽

Vol 9 ◽

Author(s):

Minjeong Yeon ◽

Youngmi Kim ◽

Deepak Pathak ◽

Eunju Kwon ◽

Dong Young Kim ◽

...

Keyword(s):

Gastric Cancer ◽

Drug Resistance ◽

Cancer Cells ◽

Anticancer Drugs ◽

Anticancer Drug ◽

Autophagic Flux ◽

Gastric Cancer Cells ◽

Ags Cells ◽

Promoter Sequences ◽

Anticancer Drug Resistance

Cancer-associated gene (CAGE), a cancer/testis antigen, has been known to promote anticancer drug resistance. Since the underlying mechanisms of CAGE-promoted anticancer drug resistance are poorly understood, we established Anticancer drug-resistant gastric cancer cells (AGSR) to better elucidate possible mechanisms. AGSR showed an increased expression level of CAGE and autophagic flux compared with anticancer drug-sensitive parental gastric cancer cells (AGS cells). AGSR cells showed higher invasion potential, growth rate, tumor spheroid formation, and angiogenic potential than AGS cells. CAGE exerted effects on the response to anticancer drugs and autophagic flux. CAGE was shown to bind to Beclin1, a mediator of autophagy. Overexpression of CAGE increased autophagic flux and invasion potential but inhibited the cleavage of PARP in response to anticancer drugs in CAGE CRISPR–Cas9 cell lines. TargetScan analysis was utilized to predict the binding of miR-302b-5p to the promoter sequences of CAGE, and the results show that miR-302b-5p directly regulated CAGE expression as illustrated by luciferase activity. MiR-302b-5p regulated autophagic flux and the response to anticancer drugs. CAGE was shown to bind the promoter sequences of miR-302b-5p. The culture medium of AGSR cells increased CAGE expression and autophagic flux in AGS cells. ImmunoEM showed CAGE was present in the exosomes of AGSR cells; exosomes of AGSR cells and human recombinant CAGE protein increased CAGE expression, autophagic flux, and resistance to anticancer drugs in AGS cells. MicroRNA array revealed miR-181b-5p as a potential negative regulator of CAGE. MiR-181b-5p inhibitor increased the expression of CAGE and autophagic flux in addition to preventing anticancer drugs from cleaving poly(ADP-ribose) polymerase (PARP) in AGS cells. TargetScan analysis predicted sphingosine 1-phosphate receptor 1 (SIPR1) as a potential target for miR-181b-5p. CAGE showed binding to the promoter sequences of S1PR1. The downregulation or inhibition of S1PR1 led to decreased autophagic flux but enhanced the sensitivity to anticancer drugs in AGSR cells. This study presents a novel role of the CAGE–miR-181b-5p–S1PR1 axis in anticancer drug resistance and autophagy.

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Metabolomic analysis of dynamic response and drug resistance of gastric cancer cells to 5-fluorouracil.

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.15_suppl.e22053 ◽

2013 ◽

Vol 31 (15_suppl) ◽

pp. e22053-e22053

Author(s):

Shinsuke Sasada ◽

Yoshihiro Miyata ◽

Yasuhiro Tsutani ◽

Jun Hihara ◽

Morihito Okada

Keyword(s):

Gastric Cancer ◽

Cancer Research ◽

Drug Resistance ◽

Mrna Expression ◽

Cancer Cells ◽

Human Gastric Cancer ◽

Metabolomic Analysis ◽

Gastric Cancer Cells ◽

Short Term Treatment ◽

Resistant Cells

e22053 Background: Metabolomics has developed as an important new tool in cancer research. It is expected to lead to the discovery of biomarker candidates for cancer diagnosis and treatment. The current study aimed to perform a comprehensive metabolomic analysis of the intracellular dynamic responses of human gastric cancer cells to 5-fluorouracil (5-FU), referencing the mechanism of drug action and drug resistance. Methods: Small metabolites in gastric cancer cells and 5-FU–resistant cells were measured by liquid chromatography-mass spectrometry. Candidates for drug targets were selected according to the presence or absence of resistance, before and after 5-FU treatment. In addition, the gene expression of each candidate was assessed by reverse transcription-polymerase chain reaction. Results: The number of metabolites in cancer cells dramatically changed during short-term treatment with 5-FU. Particularly, proline was reduced to one-third of its original level and glutamate was increased by a factor of 3 after 3 h of treatment. The metabolic production of glutamate from proline proceeds by proline dehydrogenase (PRODH), producing superoxide. After 5-FU treatment, PRODH mRNA expression was upregulated 2-fold and production of superoxide was increased by a factor of 3. In 5-FU–resistant cells, proline and glutamate levels were less affected than in non-resistant cells, and PRODH mRNA expression and superoxide generation were not increased following treatment. Conclusions: The authors identified a candidate biomarker, PRODH, for drug effects using the metabolomic approach, a result that was confirmed by conventional methods. In the future, metabolomics will play an important role in the field of cancer research.

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OSI-027 alleviates oxaliplatin chemoresistance in gastric cancer cells by suppressing P-gp induction

Current Molecular Medicine ◽

10.2174/1566524020666201120113538 ◽

2020 ◽

Vol 20 ◽

Author(s):

En Xu ◽

Hao Zhu ◽

Feng Wang ◽

Ji Miao ◽

Shangce Du ◽

...

Keyword(s):

Gastric Cancer ◽

Cell Cycle ◽

Cancer Cells ◽

Mammalian Target Of Rapamycin ◽

Cell Counting ◽

Gastric Cancer Cells ◽

Ags Cells ◽

Dual Inhibitor ◽

Induced Apoptosis

: Gastric cancer is one of the most common malignancies worldwide and the third leading cause of cancer-related death. In the present study, we investigated the potential activity of OSI-027, a potent and selective mammalian target of rapamycin complex 1/2 (mTOR1/2) dual inhibitor, alone or in combination with oxaliplatin against gastric cancer cells in vitro. Cell counting kit-8 assays and EdU staining were performed to examine the proliferation of cancer cells. Cell cycle and apoptosis were detected by flow cytometry. Western blot was used to detect the elements of the mTOR pathway and Pgp in gastric cancer cell lines. OSI-027 inhibited the proliferation of MKN-45 and AGS cells by arresting the cell cycle in the G0/G1 phase. At the molecular level, OSI-027 simultaneously blocked mTORC1 and mTORC2 activation, and resulted in the downregulation of phosphor-Akt, phpspho-p70S6k, phosphor-4EBP1, cyclin D1, and cyclin-dependent kinase4 (CDK4). Additionally, OSI-027 also downregulated P-gp, which enhanced oxaliplatin-induced apoptosis and suppressed multidrug resistance. Moreover, OSI-027 exhibited synergistic cytotoxic effects with oxaliplatin in vitro, while a P-gp siRNA knockdown significantly inhibited the synergistic effect. In summary, our results suggest that dual mTORC1/mTORC2 inhibitors (e.g., OSI-027) should be further investigated as a potential valuable treatment for gastric cancer.

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