[Show abstract][Hide abstract] ABSTRACT: Metastasis is a major cause of cancer-related mortality in patients with gastric cancer. Ursolic acid, a pentacyclic triterpenoid compound derived from medicinal herbs, has been demonstrated to exert anticancer effects in various cancer cell systems. However, to the best of our knowledge, the inhibitory effect of ursolic acid on the invasive phenotype of gastric cancer cells has yet to be reported. Therefore, the aim of the present study was to investigate the effect of ursolic acid on the invasiveness of SNU-484 human gastric cancer cells. Ursolic acid efficiently induced apoptosis, possibly via the downregulation of B-cell lymphoma 2 (Bcl-2), the upregulation of Bcl-2-associated X protein and the proteolytic activation of caspase-3. Furthermore, the activation of p38 mitogen-activated protein kinase and c-Jun N-terminal kinase was increased by the administration of ursolic acid. In addition, ursolic acid significantly suppressed the invasive phenotype of the SNU-484 cells and significantly decreased the expression of matrix metalloproteinase (MMP)-2, indicating that MMP-2 may be responsible for the anti-invasive activity of ursolic acid. Taken together, the results of the present study demonstrate that ursolic acid induces apoptosis and inhibits the invasive phenotype of gastric cancer cells; therefore, ursolic acid may have a potential application as a chemopreventive agent to prevent the metastasis of gastric cancer or to alleviate the process of metastasis.
[Show abstract][Hide abstract] ABSTRACT: Lysyl-tRNA synthetase (KRS), a protein synthesis enzyme in the cytosol, relocates to the plasma membrane after a laminin signal and stabilizes a 67-kDa laminin receptor (67LR) that is implicated in cancer metastasis; however, its potential as an antimetastatic therapeutic target has not been explored. We found that the small compound BC-K-YH16899, which binds KRS, impinged on the interaction of KRS with 67LR and suppressed metastasis in three different mouse models. The compound inhibited the KRS-67LR interaction in two ways. First, it directly blocked the association between KRS and 67LR. Second, it suppressed the dynamic movement of the N-terminal extension of KRS and reduced membrane localization of KRS. However, it did not affect the catalytic activity of KRS. Our results suggest that specific modulation of a cancer-related KRS-67LR interaction may offer a way to control metastasis while avoiding the toxicities associated with inhibition of the normal functions of KRS.
Full-text · Article · Nov 2013 · Nature Chemical Biology
[Show abstract][Hide abstract] ABSTRACT: Genipin, a constituent of Gardenia jasminoides Ellis, is used in the treatment of hepatic disorders and inflammatory diseases in traditional medicine. Although mounting evidence suggests an anti-tumor activity of genipin in several cancer cell systems, the inhibitory effect of genipin on the growth of breast cancer cells has not been reported yet. The present study aimed to investigate the anti-proliferative activity of genipin in MDA-MB-231 human breast cancer cells. Herein, we showed that genipin efficiently induced apoptosis in MDA-MB-231 cells by the down-regulation of Bcl-2, up-regulation of Bax and proteolytic activation of caspase-3. Activation of JNK and p38 MAPK also increased by genipin. Importantly, genipin significantly inhibited invasive and migratory phenotypes of MDA-MB-231 cells. Taken together, this study demonstrates that genipin induces apoptosis and inhibits invasive/migratory abilities of highly invasive MDA-MB-231 human breast cancer cells, suggesting a potential application of genipin as a chemopreventive agent that may prevent or alleviate metastatic breast cancer.
No preview · Article · Feb 2012 · Oncology Reports
[Show abstract][Hide abstract] ABSTRACT: Hypoxia and growth factor stimulation induce hypoxia-inducible factor-1α (HIF-1α), conferring upon cancer cells the ability to adapt to microenvironments and enhance proliferation, angiogenesis and metastasis. Hemin, an iron-binding porphyrin, has been used to treat porphyria attacks, particularly in acute intermittent porphyria. Although the anti-inflammatory and antitumor effects of hemin were reported, no information is available regarding its effect on HIF-1α. Our study investigated whether hemin and other protoporphyrin compounds have the ability to inhibit HIF-1α activity, and if so, what is the molecular basis of inhibition. Hemin treatment prevented CoCl(2) -induced HIF-1α expression. HIF-1α inhibition by hemin resulted from an increase in its facilitated ubiquitination and degradation, as shown by the experimental results using cychloheximide treatment and ubiquitination assays. Consistently, hemin repressed HIF-1α-dependent gene transactivation. Intriguingly, hemin directly impeded the binding between heat shock protein 90 (HSP90) and HIF-1α, which was reversed by the addition of an excess amount of ATP required for HSP90 activity. In addition, hemin decreased the expression of client proteins of HSP90. Thus, the inhibition of HIF-1α activity by hemin might result from its interaction with HSP90. Moreover, treatment of protoporphyrin IX, ZnPP or Co(III)PP, but not Mn(III)PP, inhibited HIF-1α induction, indicating that protoporphyrin ring in association with the nature of binding metal leads to HSP90 inhibition. In an in vivo model, hemin treatment inhibited not only the formation of new vessels but also cancer cell proliferation and migration/invasion, supporting the notion that hemin may be applied to the prevention and/or treatment of angiogenesis and/or cancer metastasis.
Preview · Article · Feb 2012 · International Journal of Cancer
[Show abstract][Hide abstract] ABSTRACT: Recent evidence suggests that inflammation is involved in malignant progression of breast cancer. Sphingosine 1-phosphate (S1P), acting on the G-protein-coupled receptors, is known as a potent inflammatory mediator. In this study, the effect of the inflammatory lipid S1P on the regulation of invasive/migratory phenotypes of MCF10A human breast epithelial cells was investigated to elucidate a causal relationship between inflammation and the control of invasiveness of breast cells. We show that S1P causes induction of matrix metalloproteinase-9 (MMP-9) in vitro and in vivo, and thus enhances invasion and migration. We also show that fos plays a crucial role in the transcriptional activation of MMP-9 by S1P. In addition, activation of extracellular-signal-regulated kinases 1 and 2 (ERK1/2), p38 and alpha serine/threonine-protein kinase (Akt) are involved in the process of S1P-mediated induction of MMP-9 expression and invasion. Activation of the S1P receptor S1P₃ and G(αq) are required for S1P-induced invasive/migratory responses, suggesting that the enhancement of S1P-mediated invasiveness is triggered by the specific coupling of S1P₃ to the heterotrimeric G(αq) subunit. Activation of phospholipase C-β₄ and intracellular Ca²⁺ release are required for S1P-induced MMP-9 upregulation. Taken together, this study demonstrated that S1P regulates MMP-9 induction and invasiveness through coupling of S1P₃ and G(αq) in MCF10A cells, thus providing a molecular basis for the crucial role of S1P in promoting breast cell invasion.
[Show abstract][Hide abstract] ABSTRACT: Ras expression has been suggested to be a marker for tumor aggressiveness of breast cancer. We previously showed that H-Ras, but not N-Ras, induced invasive/migratory phenotypes in MCF10A human breast epithelial cells. The present study aimed to determine the role of granulocyte colony-stimulating factor in H-Ras-induced malignant progression of human breast epithelial cells. Here, we show that G-CSF plays a crucial role in H-Ras-induced MCF10A cell invasion and migration. The siRNA-mediated knockdown of G-CSF significantly reduced H-Ras-induced matrix metalloproteinase (MMP)-2 expression, as well as invasion/migration, suggesting the functional significance of G-CSF in the invasive phenotype of human breast cells. Importantly, the induction of G-CSF expression conferred the invasive/migratory phenotypes to MCF10A cells with up-regulation of MMP-2 and activation of Rac1, MKK3/6, p38 MAPK, Akt, and ERKs. Knockdown of Rac1 by siRNA significantly inhibited MMP-2 upregulation and invasiveness of G-CSF MCF10A cells, demonstrating that G-CSF-induced MMP-2 upregulation and invasive phenotype is mediated by Rac1. Using human breast tissues and sera from breast cancer patients, we further demonstrate that the expression level of G-CSF is strongly correlated with pathologically-diagnosed breast cancer. These data provide a molecular basis for the crucial role of G-CSF in promoting invasiveness of human breast epithelial cells.
[Show abstract][Hide abstract] ABSTRACT: The G12 subfamily of the heterotrimeric G proteins, Galpha12 and Galpha13, has been implicated as an important signaling component in various cellular processes including oncogenesis and cells invasion. Our previous report showed that the expression of an activated mutant of Galpha12 (Galpha12QL) or Galpha13 (Galpha13QL) leads to cell invasion in MCF10A human breast epithelial cells. The present study aimed to investigate the role of Galpha12 and Galpha13 in the malignant phenotypic conversion of NIH3T3 mouse fibroblast cells. Galpha12QL and Galpha13QL induced an invasive phenotype in NIH3T3 cells. In addition, the activation of Galpha12 and Galpha13 upregulated matrix metalloproteinase (MMP)-2 while MMP-9 was not affected by either Galpha12QL or Galpha13QL. Using female NOD/SCID mice injected with NIH3T3 cells stably expressing Galpha12QL, we provided in vivo confirmation of Galpha12-mediated MMP-2 upregulation. Taken together, this study elucidated the role of Galpha12/13 in regulating malignant phenotypic conversion of NIH3T3 fibroblast cells, validating the role of Galphal2/13 in tumorigenesis.
No preview · Article · Jun 2011 · Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics
[Show abstract][Hide abstract] ABSTRACT: Although mounting evidence suggests a role for G(12) proteins, G(α12) and G(α13), in tumor progression, a direct role of G(12) proteins has not been determined. This study aims to elucidate the molecular mechanism for a tumorigenic and invasive potential of G(α12) and G(α13) in MCF10A human breast epithelial cells. Here, we report, for the first time, that G(α12) and G(α13) induce upregulation of matrix metalloproteinase (MMP)-2 leading to the invasive and migratory phenotypes in MCF10A cells. We further show that p53 is an important transcription factor for induction of MMP-2 transcriptional activation by G(α12/13). G(α12/13)-induced MMP-2 upregulation, invasion, and migration are dependent on the activation of Ras, Rac1, MKK3/6, p38, and Akt. Using human breast tissue samples, we demonstrate that the expression levels of G(α12) and MMP-2 are strongly correlated with the pathogenically diagnosed cancer (P < 0.0001). Moreover, the expression of G(α12) shows a strong correlation with that of MMP-2 in human breast cancer tissues, implicating the in vivo tumorigenic potential of G(α12). Taken together, this study elucidated the role of G(12) proteins in regulating processes for MMP-2 expression and malignant phenotypic conversion of MCF10A human breast epithelial cells, providing a molecular basis for the promoting role of G(α12) and G(α13) in breast cell invasion.
No preview · Article · Nov 2010 · Breast Cancer Research and Treatment
[Show abstract][Hide abstract] ABSTRACT: Breast cancer mortality is strongly related to the invasive and metastatic potential of tumor cells. We previously showed that an active mutant of H-Ras induced invasive phenotype of MCF10A human breast epithelial cells. Membrane anchoring of Ras requires isoprenylation which involves the activity of 3-hydroxy 3-methylglutaryl (HMG)-CoA reductase. In this study, we investigated the inhibitory effect of HMG-CoA reductase inhibitors, widely used for hypercholesterolemia, on H-Ras-induced invasion of MCF10A cells. Treatment of H-Ras MCF10A cells with simvastatin and lovastatin markedly decreased isoprenylated H-Ras in membrane fraction while the unprenylated H-Ras was increased in cytosol fraction, demonstrating that these statins inhibited membrane anchoring of H-Ras in MCF10A cells. Simvastatin and lovastatin significantly inhibited H-Ras-induced invasion which was reversed by farnesyl pyrophosphate (FPP), indicating that the inhibitory effect was related to inhibition of the biosynthesis of prenylated derivatives. Statins downregulated matrix metalloproteinase (MMP)-9 and, to a lesser extent, MMP-2 in H-Ras MCF10A cells. Simvastatin and lovastatin inactivated H-Ras downstream signaling molecules, possibly by inhibiting H-Ras membrane localization and thus its function in MCF10A cells. Taken together, this study clearly demonstrated the inhibitory effect of simvastatin and lovastatin on H-Ras-induced invasion, MMP expression and signal transduction in MCF10A breast epithelial cells, providing supporting rationale for future statin trials as a therapeutic intervention to regulate breast cancer metastasis.
[Show abstract][Hide abstract] ABSTRACT: Tumor cell invasion and metastasis are often associated with matrix metalloproteinases (MMPs), among which MMP-2 and MMP-9 are of central importance. We previously showed that H-Ras, but not N-Ras, induced invasion of MCF10A human breast epithelial cells in which the enhanced expression of MMP-2 was involved. MMP-2 is produced as a latent pro-MMP-2 (72 kDa) to be activated resulting the 62 kDa active MMP-2. The present study investigated if H-Ras and/or N-Ras induces pro-MMP-2 activation of MCF10A cells when cultured in two-dimensional gel of type I collagen. Type I collagen induced activation of pro-MMP-2 only in H-Ras MCF10A cells but not in N-Ras MCF10A cells. Induction of active MMP-2 by type I collagen was suppressed by blocking integrin alpha2, indicating the involvement of integrin signaling in pro-MMP-2 activation. Membrane-type (MT)1-MMP and tissue inhibitor of metalloproteinase (TIMP)-2 were up-regulated by H-Ras but not by N-Ras in the type I collagen-coated gel, suggesting that H-Ras-specific up-regulation of MT1-MMP and TIMP-2 may lead to the activation of pro-MMP-2. Since acquisition of pro-MMP-2 activation can be associated with increased malignant progression, these results may help understanding the mechanisms for the cell surface matrix-degrading potential which will be crucial to the prognosis and therapy of breast cancer metastasis.
Preview · Article · Oct 2007 · Journal of biochemistry and molecular biology
[Show abstract][Hide abstract] ABSTRACT: We have previously shown that transforming growth factor (TGF)-beta up-regulates matrix metalloproteinase (MMP)-2 leading to the induction of oncogenic signaling in preneoplastic MCF10A human breast epithelial cells. The present study investigated the mechanism of transcriptional regulation of MMP-2 by TGF-beta in MCF10A cells. By using 5' deletion constructs of MMP-2 promoter, we demonstrated that binding sites for p53, S1, AP-1 and Sp1, and to a lesser extent CREB, GCN-His and PEA3, were potential cis-acting elements for TGF-beta-induced transcriptional activation of MMP-2 in MCF10A cells. Since activating transcription factor (ATF)2 was shown to mediate the TGF-beta-induced cellular responses, we examined the involvement of ATF2 in TGF-beta-activated MMP-2 gene transcription. TGF-beta increased DNA binding activity of AP-1 in which ATF2 was involved as evidenced by electrophoretic mobility shift assay. TGF-beta induced phosphorylation of ATF2 through p38 MAPK signaling. A dominant-negative (DN) ATF2 significantly inhibited the TGF-beta-induced up-regulation of MMP-2, but not that of MMP-9, suggesting that ATF2 may be a transcription factor responsible for transcriptional activation of MMP-2 gene by TGF-beta. Invasive and migratory phenotypes induced by TGF-beta were significantly inhibited by DN ATF2, indicating a critical role of ATF2 in TGF-beta-induced oncogenic progression of MCF10A cells. Taken together, this study demonstrates that ATF2 mediates the TGF-beta-induced MMP-2 transcriptional activation, elucidating a molecular mechanism for the malignant progression of human breast epithelial cells exerted by TGF-beta.
[Show abstract][Hide abstract] ABSTRACT: To address how transforming growth factor (TGF)-beta and oncogenic H-ras signal transduction pathways interact with each other in the malignant progression of breast epithelial cells, we investigated the role of TGF-beta signaling pathway in invasive and migrative properties of H-ras-transformed MCF10A human breast epithelial cells in this study. Here we show that TGF-beta treatment significantly enhanced invasion and migration of H-ras MCF10A cells. H-ras-mediated activation of p38 MAPK and ERK-1/2 was stimulated by TGF-beta. TGF-beta increased expression of matrix metalloproteinase (MMP)-2 through transcriptional activation while TGF-beta-stimulated MMP-9 up-regulation did not occur at transcription level. Activation of p38 MAPK pathway was required for TGF-beta-induced cell migration, invasion and MMP-2/-9 up-regulation, indicating a critical role of p38 MAPK signaling in TGF-beta-promoted tumor progression of H-ras-activated cells. ERKs signaling was also crucial for TGF-beta-enhanced invasive and migrative phenotypes but the up-regulation of MMP-2/-9 was not dependent on ERKs activity. Taken together, we show that TGF-beta promotes H-ras-mediated cell migration and invasive phenotypes in which p38 MAPK and ERKs signaling pathways are involved. Our findings revealing how H-ras and TGF-beta signal pathways interact with each other in MCF10A human breast cells may provide an insight into molecular mechanisms for contribution of TGF-beta to a malignant progression of breast cancer in collaboration with activated H-ras.
[Show abstract][Hide abstract] ABSTRACT: Transforming growth factor (TGF)-beta has been reported to exert growth inhibitory activity in normal epithelial cells whereas it induces cell proliferation and invasive phenotypes in advanced carcinomas. Our previous study showed that MCF10A, a spontaneously immortalized "normal" breast epithelial cell line, is resistant to TGF-beta-induced growth inhibition, suggesting that conversion of TGF-beta growth inhibitory signaling into an oncogenic pathway may occur at the early stage of tumor development/progression. To address this issue, we investigated the TGF-beta signaling pathway and its role in phenotypic transformation of MCF10A cells. TGF-beta treatment induced changes in the MCF10A cell morphology from cuboidal to an elongated spindle-like shape, accompanied with down-regulation of epithelial cell marker E-cadherin. TGF-beta treatment was sufficient to induce migrative and invasive phenotypes in these cells, an important phenotypic conversion during tumor progression. We also showed that TGF-beta treatment rapidly activated ERK-1/2 and p38 MAPK leading to upregulation of matrix metalloproteinase (MMP)-2 and MMP-9. Using chemical inhibitors and dominant negative mutants of MAPKs, we provide evidence that while both p38 MAPK and ERKs are required for TGF-beta-induced MCF10A cell migration and invasion, TGF-beta-induced MMP-2 and MMP-9 expression depends on p38 MAPK signaling, but is independent of ERK activity. This study demonstrates the roles of TGF-beta signaling pathways for induction of oncogenic signaling in preneoplastic human breast epithelial cells and will deepen our understanding of TGF-beta signaling in the progress of breast cancer.
No preview · Article · Dec 2004 · International Journal of Oncology