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Argininosuccinate synthetase 1 suppression and arginine restriction inhibit cell migration in gastric cancer cell lines.

ABSTRACT: Gastric cancer metastasis remains a major cause of cancer-related deaths. There is an urgent need to develop new therapeutic approaches targeting metastatic gastric cancer. Argininosuccinate synthetase 1 (ASS1) expression is increased in gastric cancer. We detected the protein expression of ASS1 in human gastric cancer cell lines (AGS, NCI-N87, and MKN45) and in murine gastric cancer cell lines (3I and 3IB2). We used vector-mediated short hairpin RNA (shRNA) expression to silence ASS1 expression in the MKN45 and 3IB2 cell lines, and analyzed the effects of this protein on cell migration and metastasis. We demonstrated that ASS1 silencing suppressed cell migration in the MKN45 and 3IB2 cell lines. ASS1 knockdown significantly reduced liver metastasis in mice after the intrasplenic implantation of 3IB2 cancer cell clones. To determine whether arginine restriction may represent a therapeutic approach to treat gastric cancer, the sensitivity of tumor cells to arginine depletion was determined in gastric cancer cells. Arginine depletion significantly inhibited cell migration in the gastric cancer cell line. The silencing of ASS1 expression in MKN45 and 3IB2 gastric cancer cells markedly decreased STAT3 protein expression. In conclusion, our results indicate that the ASS1 protein is required for cell migration in gastric cancer cell lines.

SUBMITTER: Shan YS

PROVIDER: S-EPMC4415574 | biostudies-literature | 2015 Apr

REPOSITORIES: biostudies-literature

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Argininosuccinate synthetase 1 suppression and arginine restriction inhibit cell migration in gastric cancer cell lines.

Shan Yan-Shen YS Hsu Hui-Ping HP Lai Ming-Derg MD Yen Meng-Chi MC Chen Wei-Ching WC Fang Jung-Hua JH Weng Tzu-Yang TY Chen Yi-Ling YL

Scientific reports 20150430

Gastric cancer metastasis remains a major cause of cancer-related deaths. There is an urgent need to develop new therapeutic approaches targeting metastatic gastric cancer. Argininosuccinate synthetase 1 (ASS1) expression is increased in gastric cancer. We detected the protein expression of ASS1 in human gastric cancer cell lines (AGS, NCI-N87, and MKN45) and in murine gastric cancer cell lines (3I and 3IB2). We used vector-mediated short hairpin RNA (shRNA) expression to silence ASS1 expression ...[more]

PMID: 25928182

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Project description:Arginine (Arg) deprivation is a promising therapeutic approach for tumors with low argininosuccinate synthetase 1 (ASS1) expression. However, its efficacy as a single agent therapy needs to be improved as resistance is frequently observed. Methods: A tissue microarray was performed to assess ASS1 expression in surgical specimens of pancreatic ductal adenocarcinoma (PDAC) and its correlation with disease prognosis. An RNA-Seq analysis examined the role of ASS1 in regulating the global gene transcriptome. A high throughput screen of FDA-approved oncology drugs identified synthetic lethality between histone deacetylase (HDAC) inhibitors and Arg deprivation in PDAC cells with low ASS1 expression. We examined HDAC inhibitor panobinostat (PAN) and Arg deprivation in a panel of human PDAC cell lines, in ASS1-high and -knockdown/knockout isogenic models, in both anchorage-dependent and -independent cultures, and in multicellular complex cultures that model the PDAC tumor microenvironment. We examined the effects of combined Arg deprivation and PAN on DNA damage and the protein levels of key DNA repair enzymes. We also evaluated the efficacy of PAN and ADI-PEG20 (an Arg-degrading agent currently in Phase 2 clinical trials) in xenograft models with ASS1-low and -high PDAC tumors. Results: Low ASS1 protein level is a negative prognostic indicator in PDAC. Arg deprivation in ASS1-deficient PDAC cells upregulated asparagine synthetase (ASNS) which redirected aspartate (Asp) from being used for de novo nucleotide biosynthesis, thus causing nucleotide insufficiency and impairing cell cycle S-phase progression. Comprehensively validated, HDAC inhibitors and Arg deprivation showed synthetic lethality in ASS1-low PDAC cells. Mechanistically, combined Arg deprivation and HDAC inhibition triggered degradation of a key DNA repair enzyme C-terminal-binding protein interacting protein (CtIP), resulting in DNA damage and apoptosis. In addition, S-phase-retained ASS1-low PDAC cells (due to Arg deprivation) were also sensitized to DNA damage, thus yielding effective cell death. Compared to single agents, the combination of PAN and ADI-PEG20 showed better efficacy in suppressing ASS1-low PDAC tumor growth in mouse xenograft models. Conclusion: The combination of PAN and ADI-PEG20 is a rational translational therapeutic strategy for treating ASS1-low PDAC tumors through synergistic induction of DNA damage.

Project description:BackgroundPancreatic cancer is a leading cause of cancer-related deaths in the world with a 5-year survival rate of less than 6%. Currently, there is no successful therapeutic strategy for advanced pancreatic cancer, and new effective strategies are urgently needed. Recently, an arginine deprivation agent, arginine deiminase, was found to inhibit the growth of some tumor cells (i.e., hepatocellular carcinoma, melanoma, and lung cancer) deficient in argininosuccinate synthetase (ASS), an enzyme used to synthesize arginine. The purpose of this study was to evaluate the therapeutic efficacy of arginine deiminase in combination with gemcitabine, the first line chemotherapeutic drug for patients with pancreatic cancer, and to identify the mechanisms associated with its anticancer effects.MethodsIn this study, we first analyzed the expression levels of ASS in pancreatic cancer cell lines and tumor tissues using immunohistochemistry and RT-PCR. We further tested the effects of the combination regimen of arginine deiminase with gemcitabine on pancreatic cancer cell lines in vitro and in vivo.ResultsClinical investigation showed that pancreatic cancers with reduced ASS expression were associated with higher survivin expression and more lymph node metastasis and local invasion. Treatment of ASS-deficient PANC-1 cells with arginine deiminase decreased their proliferation in a dose- and time-dependent manner. Furthermore, arginine deiminase potentiated the antitumor effects of gemcitabine on PANC-1 cells via multiple mechanisms including induction of cell cycle arrest in the S phase, upregulation of the expression of caspase-3 and 9, and inhibition of activation of the NF-?B survival pathway by blocking NF-?B p65 signaling via suppressing the nuclear translocation and phosphorylation (serine 536) of NF-?B p65 in vitro. Moreover, arginine deiminase can enhance antitumor activity of gemcitabine-based chemotherapy in the mouse xenograft model.ConclusionsOur results suggest that arginine deprivation by arginine deiminase, in combination with gemcitabine, may offer a novel effective treatment strategy for patients with pancreatic cancer and potentially improve the outcome of patients with pancreatic cancer.

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Argininosuccinate synthetase 1 suppression and arginine restriction inhibit cell migration in gastric cancer cell lines.

Publications

Argininosuccinate synthetase 1 suppression and arginine restriction inhibit cell migration in gastric cancer cell lines.

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