Publications by authors named "Laia Caja"

27 Publications

  • Page 1 of 1

The noncoding MIR100HG RNA enhances the autocrine function of transforming growth factor β signaling.

Oncogene 2021 May 4;40(21):3748-3765. Epub 2021 May 4.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Box 582, Biomedical Center, Uppsala University, Uppsala, Sweden.

Activation of the transforming growth factor β (TGFβ) pathway modulates the expression of genes involved in cell growth arrest, motility, and embryogenesis. An expression screen for long noncoding RNAs indicated that TGFβ induced mir-100-let-7a-2-mir-125b-1 cluster host gene (MIR100HG) expression in diverse cancer types, thus confirming an earlier demonstration of TGFβ-mediated transcriptional induction of MIR100HG in pancreatic adenocarcinoma. MIR100HG depletion attenuated TGFβ signaling, expression of TGFβ-target genes, and TGFβ-mediated cell cycle arrest. Moreover, MIR100HG silencing inhibited both normal and cancer cell motility and enhanced the cytotoxicity of cytostatic drugs. MIR100HG overexpression had an inverse impact on TGFβ signaling responses. Screening for downstream effectors of MIR100HG identified the ligand TGFβ1. MIR100HG and TGFB1 mRNA formed ribonucleoprotein complexes with the RNA-binding protein HuR, promoting TGFβ1 cytokine secretion. In addition, TGFβ regulated let-7a-2-3p, miR-125b-5p, and miR-125b-1-3p expression, all encoded by MIR100HG intron-3. Certain intron-3 miRNAs may be involved in TGFβ/SMAD-mediated responses (let-7a-2-3p) and others (miR-100, miR-125b) in resistance to cytotoxic drugs mediated by MIR100HG. In support of a model whereby TGFβ induces MIR100HG, which then enhances TGFβ1 secretion, analysis of human carcinomas showed that MIR100HG expression correlated with expression of TGFB1 and its downstream extracellular target TGFBI. Thus, MIR100HG controls the magnitude of TGFβ signaling via TGFβ1 autoinduction and secretion in carcinomas.
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http://dx.doi.org/10.1038/s41388-021-01803-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8154591PMC
May 2021

The TGFB2-AS1 lncRNA Regulates TGF-β Signaling by Modulating Corepressor Activity.

Cell Rep 2019 Sep;28(12):3182-3198.e11

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, and Ludwig Cancer Research Box 582, Biomedical Center, Uppsala University, 751 23 Uppsala, Sweden. Electronic address:

Molecular processes involving lncRNAs regulate cell function. By applying transcriptomics, we identify lncRNAs whose expression is regulated by transforming growth factor β (TGF-β). Upon silencing individual lncRNAs, we identify several that regulate TGF-β signaling. Among these lncRNAs, TGFB2-antisense RNA1 (TGFB2-AS1) is induced by TGF-β through Smad and protein kinase pathways and resides in the nucleus. Depleting TGFB2-AS1 enhances TGF-β/Smad-mediated transcription and expression of hallmark TGF-β-target genes. Increased dose of TGFB2-AS1 reduces expression of these genes, attenuates TGF-β-induced cell growth arrest, and alters BMP and Wnt pathway gene profiles. Mechanistically, TGFB2-AS1, mainly via its 3' terminal region, binds to the EED adaptor of the Polycomb repressor complex 2 (PRC2), promoting repressive histone H3K27me modifications at TGF-β-target gene promoters. Silencing EED or inhibiting PRC2 methylation activity partially rescues TGFB2-AS1-mediated gene repression. Thus, the TGF-β-induced TGFB2-AS1 lncRNA exerts inhibitory functions on TGF-β/BMP signaling output, supporting auto-regulatory negative feedback that balances TGF-β/BMP-mediated responses.
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http://dx.doi.org/10.1016/j.celrep.2019.08.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6859500PMC
September 2019

Downregulation of Epidermal Growth Factor Receptor in hepatocellular carcinoma facilitates Transforming Growth Factor-β-induced epithelial to amoeboid transition.

Cancer Lett 2019 Nov 26;464:15-24. Epub 2019 Aug 26.

TGF-β and Cancer Group, Oncobell Program, Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain; Oncology Program, CIBEREHD, National Biomedical Research Institute on Liver and Gastrointestinal Diseases, Instituto de Salud Carlos III, Madrid, Spain; Department of Physiological Sciences, Faculty of Medicine and Health Sciences, University of Barcelona, Spain. Electronic address:

The Epidermal Growth Factor Receptor (EGFR) and the Transforming Growth Factor-beta (TGF-β) are key regulators of hepatocarcinogenesis. Targeting EGFR was proposed as a promising therapy; however, poor success was obtained in human hepatocellular carcinoma (HCC) clinical trials. Here, we describe how EGFR is frequently downregulated in HCC patients while TGF-β is upregulated. Using 2D/3D cellular models, we show that after EGFR loss, TGF-β is more efficient in its pro-migratory and invasive effects, inducing epithelial to amoeboid transition. EGFR knock-down promotes loss of cell-cell and cell-to-matrix adhesion, favouring TGF-β-induced actomyosin contractility and acquisition of an amoeboid migratory phenotype. Moreover, TGF-β upregulates RHOC and CDC42 after EGFR silencing, promoting Myosin II in amoeboid cells. Importantly, low EGFR combined with high TGFB1 or RHOC/CDC42 levels confer poor patient prognosis. In conclusion, this work reveals a new tumour suppressor function for EGFR counteracting TGF-β-mediated epithelial to amoeboid transitions in HCC, supporting a rational for targeting the TGF-β pathway in patients with low EGFR expression. Our work also highlights the relevance of epithelial to amoeboid transition in human tumours and the need to better target this process in the clinic.
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http://dx.doi.org/10.1016/j.canlet.2019.08.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6853171PMC
November 2019

TANK-binding kinase 1 is a mediator of platelet-induced EMT in mammary carcinoma cells.

FASEB J 2019 07 26;33(7):7822-7832. Epub 2019 Mar 26.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden.

Platelets can promote several stages of the metastatic process and thus contribute to malignant progression. As an example, platelets promote invasive properties of tumor cells by induction of epithelial to mesenchymal transition (EMT). In this study, we show that tumor necrosis factor receptor-associated factor (TRAF) family member-associated NF-κB activator (TANK)-binding kinase 1 (TBK1) is a previously unknown mediator of platelet-induced EMT in mammary carcinoma cells. Coculture of 2 mammary carcinoma cell lines, Ep5 from mice and MCF10A(MII) from humans, with isolated platelets induced morphologic as well as molecular changes characteristic of EMT, which was paralleled with activation of TBK1. TBK1 depletion using small interfering RNA impaired platelet-induced EMT in both Ep5 and MCF10A(MII) cells. Furthermore, platelet-induced activation of the NF-κB subunit p65 was suppressed after TBK1 knockdown, demonstrating that TBK1 mediates platelet-induced NF-κB signaling and EMT. Using an metastasis assay, we found that depletion of TBK1 from mammary carcinoma cells during preconditioning with platelets subsequently suppressed the formation of lung metastases in mice. Altogether, these results suggest that TBK1 contributes to tumor invasiveness and may be a driver of metastatic spread in breast cancer.-Zhang, Y., Unnithan, R. V. M., Hamidi, A., Caja, L., Saupe, F., Moustakas, A., Cedervall, J., Olsson, A.-K. TANK-binding kinase 1 is a mediator of platelet-induced EMT in mammary carcinoma cells.
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http://dx.doi.org/10.1096/fj.201801936RRRDOI Listing
July 2019

TGF-β and the Tissue Microenvironment: Relevance in Fibrosis and Cancer.

Int J Mol Sci 2018 Apr 26;19(5). Epub 2018 Apr 26.

TGF-β and Cancer Group, Oncobell Program, Bellvitge Biomedical Research Institute (IDIBELL), Gran Via de l'Hospitalet, 199, 08908 Barcelona, Spain.

Transforming growth factor-β (TGF-β) is a cytokine essential for the induction of the fibrotic response and for the activation of the cancer stroma. Strong evidence suggests that a strong cross-talk exists among TGF-β and the tissue extracellular matrix components. TGF-β is stored in the matrix as part of a large latent complex bound to the latent TGF-β binding protein (LTBP) and matrix binding of latent TGF-β complexes, which is required for an adequate TGF-β function. Once TGF-β is activated, it regulates extracellular matrix remodelling and promotes a fibroblast to myofibroblast transition, which is essential in fibrotic processes. This cytokine also acts on other cell types present in the fibrotic and tumour microenvironment, such as epithelial, endothelial cells or macrophages and it contributes to the cancer-associated fibroblast (CAF) phenotype. Furthermore, TGF-β exerts anti-tumour activity by inhibiting the host tumour immunosurveillance. Aim of this review is to update how TGF-β and the tissue microenvironment cooperate to promote the pleiotropic actions that regulate cell responses of different cell types, essential for the development of fibrosis and tumour progression. We discuss recent evidences suggesting the use of TGF-β chemical inhibitors as a new line of defence against fibrotic disorders or cancer.
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http://dx.doi.org/10.3390/ijms19051294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5983604PMC
April 2018

Snail regulates BMP and TGFβ pathways to control the differentiation status of glioma-initiating cells.

Oncogene 2018 05 16;37(19):2515-2531. Epub 2018 Feb 16.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Box 582, Biomedical Center, Uppsala University, SE-75123, Uppsala, Sweden.

Glioblastoma multiforme is a brain malignancy characterized by high heterogeneity, invasiveness, and resistance to current therapies, attributes related to the occurrence of glioma stem cells (GSCs). Transforming growth factor β (TGFβ) promotes self-renewal and bone morphogenetic protein (BMP) induces differentiation of GSCs. BMP7 induces the transcription factor Snail to promote astrocytic differentiation in GSCs and suppress tumor growth in vivo. We demonstrate that Snail represses stemness in GSCs. Snail interacts with SMAD signaling mediators, generates a positive feedback loop of BMP signaling and transcriptionally represses the TGFB1 gene, decreasing TGFβ1 signaling activity. Exogenous TGFβ1 counteracts Snail function in vitro, and in vivo promotes proliferation and re-expression of Nestin, confirming the importance of TGFB1 gene repression by Snail. In conclusion, novel insight highlights mechanisms whereby Snail differentially regulates the activity of the opposing BMP and TGFβ pathways, thus promoting an astrocytic fate switch and repressing stemness in GSCs.
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http://dx.doi.org/10.1038/s41388-018-0136-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5945579PMC
May 2018

Snail mediates crosstalk between TGFβ and LXRα in hepatocellular carcinoma.

Cell Death Differ 2018 05 11;25(5):885-903. Epub 2017 Dec 11.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Box 582, Biomedical Center, Uppsala University, SE-75123, Uppsala, Sweden.

Understanding the complexity of changes in differentiation and cell survival in hepatocellular carcinoma (HCC) is essential for the design of new diagnostic tools and therapeutic modalities. In this context, we have analyzed the crosstalk between transforming growth factor β (TGFβ) and liver X receptor α (LXRα) pathways. TGFβ is known to promote cytostatic and pro-apoptotic responses in HCC, and to facilitate mesenchymal differentiation. We here demonstrate that stimulation of the nuclear LXRα receptor system by physiological and clinically useful agonists controls the HCC response to TGFβ. Specifically, LXRα activation antagonizes the mesenchymal, reactive oxygen species and pro-apoptotic responses to TGFβ and the mesenchymal transcription factor Snail mediates this crosstalk. In contrast, LXRα activation and TGFβ cooperate in enforcing cytostasis in HCC, which preserves their epithelial features. LXRα influences Snail expression transcriptionally, acting on the Snail promoter. These findings propose that clinically used LXR agonists may find further application to the treatment of aggressive, mesenchymal HCCs, whose progression is chronically dependent on autocrine or paracrine TGFβ.
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http://dx.doi.org/10.1038/s41418-017-0021-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5943406PMC
May 2018

Transforming growth factor β as regulator of cancer stemness and metastasis.

Br J Cancer 2016 09 18;115(7):761-9. Epub 2016 Aug 18.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Uppsala University, Box 582, SE 751 23 Uppsala, Sweden.

Key elements of cancer progression towards metastasis are the biological actions of cancer stem cells and stromal cells in the tumour microenvironment. Cross-communication between tumour and stromal cells is mediated by secreted cytokines, one of which, the transforming growth factor β (TGFβ), regulates essentially every cell within the malignant tissue. In this article, we focus on the actions of TGFβ on cancer stem cells, cancer-associated fibroblasts and immune cells that assist the overall process of metastatic dissemination. We aim at illustrating intricate connections made by various cells in the tumour tissue and which depend on the action of TGFβ.
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http://dx.doi.org/10.1038/bjc.2016.255DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5046208PMC
September 2016

Transforming growth factor β and bone morphogenetic protein actions in brain tumors.

FEBS Lett 2015 Jun 6;589(14):1588-97. Epub 2015 May 6.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Uppsala University, Box 582, SE-751 23 Uppsala, Sweden; Ludwig Cancer Research, Science for Life Laboratory, Uppsala University, Box 595, SE-751 24 Uppsala, Sweden. Electronic address:

Members of the transforming growth factor β (TGF-β) family are implicated in the biology of several cancers. Here we focus on malignancies of the brain and examine the TGFβ and the bone morphogenetic protein (BMP) signaling branches of the family. These pathways exhibit context-dependent actions during tumorigenesis, acting either as tumor suppressors or as pro-tumorigenic agents. In the brain, the TGF-βs associate with oncogenic development and progression to the more malignant state. Inversely, the BMPs suppress tumorigenic potential by acting as agents that induce tumor cell differentiation. The latter has been best demonstrated in grade IV astrocytomas, otherwise known as glioblastoma multiforme. We discuss how the actions of TGF-βs and BMPs on cancer stem cells may explain their effects on tumor progression, and try to highlight intricate mechanisms that may link tumor cell differentiation to invasion. The focus on TGF-β and BMP and their actions in brain malignancies provides a rich territory for mechanistic understanding of tumor heterogeneity and suggests ways for improved therapeutic intervention, currently being addressed by clinical trials.
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http://dx.doi.org/10.1016/j.febslet.2015.04.058DOI Listing
June 2015

Vascular smooth muscle cell phenotypic changes in patients with Marfan syndrome.

Arterioscler Thromb Vasc Biol 2015 Apr 15;35(4):960-72. Epub 2015 Jan 15.

From the Department of Cell Biology, Immunology and Neurosciences (E.C.-M., T.M., C.S.-P, J.S., D.G, Y.M., V.H., E.S., G.E.), Departments of Physiological Sciences I (J.J.U., D.N.) and Physiological Sciences II (I.F.), Department of Pathology and Experimental Therapeutics (E.C.), University of Barcelona School of Medicine, Barcelona, Spain; Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain (M.C., G.E.); Institut de Nanociència i Nanotecnologia (IN2UB), Barcelona, Spain (G.E.); Institut de Bioenginyeria de Catalunya (IBEC), Barcelona, Spain and CIBER de Enfermedades Respiratorias (CIBERES) (D.N.); Vascular Biology Laboratory, Heart Institute (InCor), University of São Paulo School of Medicine, São Paulo, Brazil (T.M.); Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain (E.C.-M., J.L.-L. L.C., E.B., I.F.); Centro de Biología Molecular Severo Ochoa (CSIC-UAM), Madrid, Spain (O.B., F.R.-P.); Hospital de Bellvitge-IDIBELL, L'Hospitalet de Llobregat, Barcelona, Spain (C.G.-C., E.C., D.T.); Cardiovascular Surgery Department, Hospital Clínic i Provincial, Barcelona, Spain (M.C.); Cardiac Surgery Department, Marfan Syndrome Unit, Hospital Universitario 12 de Octubre, Madrid, Spain (A.F.); and William S. Smilow Center for Marfan Syndrome Research, Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, Baltimore, MD (H.C.D.).

Objective: Marfan's syndrome is characterized by the formation of ascending aortic aneurysms resulting from altered assembly of extracellular matrix microfibrils and chronic tissue growth factor (TGF)-β signaling. TGF-β is a potent regulator of the vascular smooth muscle cell (VSMC) phenotype. We hypothesized that as a result of the chronic TGF-β signaling, VSMC would alter their basal differentiation phenotype, which could facilitate the formation of aneurysms. This study explores whether Marfan's syndrome entails phenotypic alterations of VSMC and possible mechanisms at the subcellular level.

Approach And Results: Immunohistochemical and Western blotting analyses of dilated aortas from Marfan patients showed overexpression of contractile protein markers (α-smooth muscle actin, smoothelin, smooth muscle protein 22 alpha, and calponin-1) and collagen I in comparison with healthy aortas. VSMC explanted from Marfan aortic aneurysms showed increased in vitro expression of these phenotypic markers and also of myocardin, a transcription factor essential for VSMC-specific differentiation. These alterations were generally reduced after pharmacological inhibition of the TGF-β pathway. Marfan VSMC in culture showed more robust actin stress fibers and enhanced RhoA-GTP levels, which was accompanied by increased focal adhesion components and higher nuclear localization of myosin-related transcription factor A. Marfan VSMC and extracellular matrix measured by atomic force microscopy were both stiffer than their respective controls.

Conclusions: In Marfan VSMC, both in tissue and in culture, there are variable TGF-β-dependent phenotypic changes affecting contractile proteins and collagen I, leading to greater cellular and extracellular matrix stiffness. Altogether, these alterations may contribute to the known aortic rigidity that precedes or accompanies Marfan's syndrome aneurysm formation.
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http://dx.doi.org/10.1161/ATVBAHA.114.304412DOI Listing
April 2015

A trifluorinated thiazoline scaffold leading to pro-apoptotic agents targeting prohibitins.

Angew Chem Int Ed Engl 2014 Sep 5;53(38):10150-4. Epub 2014 Sep 5.

Departament de Ciències Fisiològiques II, University of Barcelona, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), L'Hospitalet de Llobregat (Spain).

A new class of small molecules, with an unprecedented trifluorothiazoline scaffold, were synthesized and their pro-apoptotic activity was evaluated. With an EC50 in the low micromolar range, these compounds proved to be potent inducers of apoptosis in a broad spectrum of tumor cell lines, regardless of the functional status of p53. Fast structure-activity relationship studies allowed the preparation of the strongest apoptosis-inducing candidate. Using a high performance affinity purification approach, we identified prohibitins 1 and 2, key proteins involved in the maintenance of cell viability, as the targets for these compounds.
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http://dx.doi.org/10.1002/anie.201405758DOI Listing
September 2014

Overactivation of the TGF-β pathway confers a mesenchymal-like phenotype and CXCR4-dependent migratory properties to liver tumor cells.

Hepatology 2013 Dec 11;58(6):2032-44. Epub 2013 Oct 11.

Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.

Unlabelled: Transforming growth factor-beta (TGF-β) is an important regulatory suppressor factor in hepatocytes. However, liver tumor cells develop mechanisms to overcome its suppressor effects and respond to this cytokine by inducing other processes, such as the epithelial-mesenchymal transition (EMT), which contributes to tumor progression and dissemination. Recent studies have placed chemokines and their receptors at the center not only of physiological cell migration but also of pathological processes, such as metastasis in cancer. In particular, CXCR4 and its ligand, stromal cell-derived factor 1α (SDF-1α) / chemokine (C-X-C motif) ligand 12 (CXCL12) have been revealed as regulatory molecules involved in the spreading and progression of a variety of tumors. Here we show that autocrine stimulation of TGF-β in human liver tumor cells correlates with a mesenchymal-like phenotype, resistance to TGF-β-induced suppressor effects, and high expression of CXCR4, which is required for TGF-β-induced cell migration. Silencing of the TGF-β receptor1 (TGFBR1), or its specific inhibition, recovered the epithelial phenotype and attenuated CXCR4 expression, inhibiting cell migratory capacity. In an experimental mouse model of hepatocarcinogenesis (diethylnitrosamine-induced), tumors showed increased activation of the TGF-β pathway and enhanced CXCR4 levels. In human hepatocellular carcinoma tumors, high levels of CXCR4 always correlated with activation of the TGF-β pathway, a less differentiated phenotype, and a cirrhotic background. CXCR4 concentrated at the tumor border and perivascular areas, suggesting its potential involvement in tumor cell dissemination.

Conclusion: A crosstalk exists among the TGF-β and CXCR4 pathways in liver tumors, reflecting a novel molecular mechanism that explains the protumorigenic effects of TGF-β and opens new perspectives for tumor therapy.
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http://dx.doi.org/10.1002/hep.26597DOI Listing
December 2013

The Epstein-Barr virus nuclear antigen-1 reprograms transcription by mimicry of high mobility group A proteins.

Nucleic Acids Res 2013 Mar 28;41(5):2950-62. Epub 2013 Jan 28.

Department of Cell and Molecular Biology, Karolinska Institutet, 17177 Stockholm, Sweden.

Viral proteins reprogram their host cells by hijacking regulatory components of protein networks. Here we describe a novel property of the Epstein-Barr virus (EBV) nuclear antigen-1 (EBNA1) that may underlie the capacity of the virus to promote a global remodeling of chromatin architecture and cellular transcription. We found that the expression of EBNA1 in transfected human and mouse cells is associated with decreased prevalence of heterochromatin foci, enhanced accessibility of cellular DNA to micrococcal nuclease digestion and decreased average length of nucleosome repeats, suggesting de-protection of the nucleosome linker regions. This is a direct effect of EBNA1 because targeting the viral protein to heterochromatin promotes large-scale chromatin decondensation with slow kinetics and independent of the recruitment of adenosine triphosphate-dependent chromatin remodelers. The remodeling function is mediated by a bipartite Gly-Arg rich domain of EBNA1 that resembles the AT-hook of High Mobility Group A (HMGA) architectural transcription factors. Similar to HMGAs, EBNA1 is highly mobile in interphase nuclei and promotes the mobility of linker histone H1, which counteracts chromatin condensation and alters the transcription of numerous cellular genes. Thus, by regulating chromatin compaction, EBNA1 may reset cellular transcription during infection and prime the infected cells for malignant transformation.
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http://dx.doi.org/10.1093/nar/gkt032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3597695PMC
March 2013

Context-dependent action of transforming growth factor β family members on normal and cancer stem cells.

Curr Pharm Des 2012 ;18(27):4072-86

Ludwig Institute for Cancer Research, Uppsala University, Sweden.

The transforming growth factor β (TGFβ) family embraces many growth factors including the Activins and bone morphogenetic proteins (BMPs). The pathways mediated by these growth factors are implicated in many fundamental biological processes such as early embryonic development, organ morphogenesis and adult tissue homeostasis and in a large number of pathologies including cancer. The action of these pathways is often contextual, which means that different cell types present different physiological responses to these ligands or that the response of one cell type to a certain ligand differs depending on the presence of other signaling proteins that stimulate the target cell together with TGFβ/BMP. The latter usually reflects developmental stage or progression to a specific pathological stage. Not only diverse growth factors and cytokines can influence the response of tissues to TGFβ/BMP, but a single cell type may also show drastically different physiological outcomes to TGFβ or Activin signaling as compared to BMP signaling. This review describes differential physiological outcomes of TGFβ and BMP signaling in normal embryonic or adult stem cells and eventually in cancer stem cells and the process of epithelial-mesenchymal transition. We also summarize evidence on the mechanistic antagonism between TGFβ and BMP signaling as established in vascular differentiation and the progression of tissue fibrosis and cancer. The article ends by discussing possible advantages that the acquired knowledge of these signaling mechanisms offers to new regimes of cancer therapy and the ever-lasting problem of drug resistance elicited by tumor initiating cells.
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http://dx.doi.org/10.2174/138161212802430459DOI Listing
March 2013

Protein-tyrosine phosphatase 1B (PTP1B) deficiency confers resistance to transforming growth factor-β (TGF-β)-induced suppressor effects in hepatocytes.

J Biol Chem 2012 May 16;287(19):15263-74. Epub 2012 Mar 16.

Biological Clues of the Invasive and Metastatic Phenotype Group, Bellvitge Biomedical Research Institut (IDIBELL), L'Hospitalet de Llobregat, 08908 Barcelona, Spain.

Transforming growth factor-β (TGF-β) plays a dual role in hepatocytes, mediating both tumor suppressor and promoter effects. The suppressor effects of the cytokine can be negatively regulated by activation of survival signals, mostly dependent on tyrosine kinase activity. The aim of our work was to study the role of the protein-tyrosine phosphatase 1B (PTP1B) on the cellular responses to TGF-β, using for this purpose immortalized neonatal hepatocytes isolated from both PTP1B(+/+) and PTP1B(-/-) mice. We have found that PTP1B deficiency conferred resistance to TGF-β suppressor effects, such as apoptosis and growth inhibition, correlating with lower Smad2/Smad3 activation. Both responses were recovered in the presence of the general tyrosine kinase inhibitor genistein. PTP1B(-/-) cells showed elevated NF-κB activation in response to TGF-β. Knockdown of the NF-κB p65 subunit increased cell response in terms of Smads phosphorylation and apoptosis. Interestingly, these effects were accompanied by inhibition of Smad7 up-regulation. In addition, lack of PTP1B promoted an altered NADPH oxidase (NOX) expression pattern in response to TGF-β, strongly increasing the NOX1/NOX4 ratio, which was reverted by genistein and p65 knockdown. Importantly, NOX1 knockdown inhibited nuclear translocation of p65, promoted Smad phosphorylation, and decreased Smad7 levels. In summary, our results suggest that PTP1B deficiency confers resistance to TGF-β through Smad inhibition, an effect that is mediated by NOX1-dependent NF-κB activation, which in turn, increases the level of the Smad inhibitor Smad7 and participates in a positive feedback loop on NOX1 up-regulation.
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http://dx.doi.org/10.1074/jbc.M111.303958DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3346142PMC
May 2012

Regulation of transcription factor Twist expression by the DNA architectural protein high mobility group A2 during epithelial-to-mesenchymal transition.

J Biol Chem 2012 Mar 11;287(10):7134-45. Epub 2012 Jan 11.

Ludwig Institute for Cancer Research, Biomedical Center, Uppsala University, Uppsala SE-751 24, Sweden.

Deciphering molecular mechanisms that control epithelial-to-mesenchymal transition (EMT) contributes to our understanding of how tumor cells become invasive and competent for intravasation. We have established that transforming growth factor β activates Smad proteins, which induce expression of the embryonic factor high mobility group A2 (HMGA2), which causes mesenchymal transition. HMGA2 associates with Smad complexes and induces expression of an established regulator of EMT, the zinc finger transcription factor Snail. We now show that HMGA2 can also induce expression of a second regulator of EMT, the basic helix-loop-helix transcription factor Twist. Silencing of endogenous Twist demonstrated that this protein acts in a partially redundant manner together with Snail. Double silencing of Snail and Twist reverts mesenchymal HMGA2-expressing cells to a more epithelial phenotype when compared with single silencing of Snail or Twist. Furthermore, HMGA2 can directly associate with A:T-rich sequences and promote transcription from the Twist promoter. The new evidence proposes a model whereby HMGA2 directly induces multiple transcriptional regulators of the EMT program and, thus, is a potential biomarker for carcinomas displaying EMT during progression to more advanced stages of malignancy.
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http://dx.doi.org/10.1074/jbc.M111.291385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3293571PMC
March 2012

Lack of amino acids in mouse hepatocytes in culture induces the selection of preneoplastic cells.

Cell Signal 2012 Jan 22;24(1):325-32. Epub 2011 Sep 22.

Biological Clues of the Invasive and Metastatic Phenotype Group, Bellvitge Biomedical Research Institut (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.

Protein malnutrition occurs when there is insufficient protein to meet metabolic demands. Previous works have indicated that cycles of protein fasting/refeeding enhance the incidence of early lesions during chemical carcinogenesis in rat liver. The general objective of this work was to study the effect of aminoacids (Aa) deprivation on the proliferation and survival of hepatocytes, to understand its possible involvement in the generation of pre-neoplastic stages in the liver. Lack of Aa in the culture medium of an immortalized mice hepatocyte cell line induced loss in cell viability, correlating with apoptosis. However, a subpopulation of cells was able to survive, which showed a more proliferative phenotype and resistance to apoptotic stimuli. Escaping to Aa deprivation-induced death is coincident with an activated mTOR signaling and higher levels of phospho-AKT and phospho-ERKs, which correlated with increased activation of EGFR/SRC pathway and overexpression of EGFR ligands, such as TGF-α and HB-EGF. Lack of Aa induced a rapid increase in reactive oxygen species (ROS) production. However, cells that survived showed an enhancement in the levels of reduced glutathione and a higher expression of γ-GCS, the regulatory enzyme of glutathione synthesis, which can be interpreted as an adaptation of the cells to counteract the oxidative stress. In conclusion, results presented in this paper indicate that it is possible to isolate a subpopulation of hepatocytes that are able to grow in the absence of Aa, showing higher capacity to proliferate and survive, reminiscent of a preneoplastic phenotype.
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http://dx.doi.org/10.1016/j.cellsig.2011.09.018DOI Listing
January 2012

The tyrphostin AG1478 inhibits proliferation and induces death of liver tumor cells through EGF receptor-dependent and independent mechanisms.

Biochem Pharmacol 2011 Dec 12;82(11):1583-92. Epub 2011 Aug 12.

Biological Clues of the Invasive and Metastatic Phenotype Group, Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.

Hepatocellular carcinoma (HCC) is one of the most common causes of cancer-related death. Different signaling pathways are de-regulated in this pathogenesis, among them the epidermal growth factor receptor one (EGFR/Erb1). Here we show that blockage of this pathway by the tyrphostin 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG1478) in different liver tumor cell lines promotes both inhibition of cell proliferation and induction of cell death, which are coincident with arrest in the G1 phase of the cell cycle, caspase-3 activation and DNA fragmentation. AG1478 up-regulates the expression of the pro-apoptotic member of the BCL-2 family BIM and down-regulates the expression of the anti-apoptotic BCL-XL and MCL1. Furthermore, it also decreases the levels of the caspase inhibitors HIAP2 and XIAP. The treatment of HCC cells with AG1478 enhanced the apoptosis induced by other pro-apoptotic stimuli, such as the physiological cytokine, TGF-β, highly expressed in liver tumors, or the chemotherapeutic drug doxorubicin. The effects observed by AG1478 were broader than the ones seen by silencing of the EGFR with siRNA, which indicates that this drug might act on other targets different from the EGFR. In this same line of evidence, AG1478 retained some cytotoxic effects in cells where EGFR has been targeted knock-down with shRNA. Interestingly, AG1478 preferentially acts on liver tumor cells, being untransformed cells much less responsive to its cytotoxic effects. In conclusion, AG1478 could be a potential therapeutic drug to be used in HCC.
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http://dx.doi.org/10.1016/j.bcp.2011.08.011DOI Listing
December 2011

Sorafenib sensitizes hepatocellular carcinoma cells to physiological apoptotic stimuli.

J Cell Physiol 2012 Apr;227(4):1319-25

Biological Clues of the Invasive and Metastatic Phenotype Group, Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.

Sorafenib increases survival rate of patients with advanced hepatocellular carcinoma (HCC). The mechanism underlying this effect is not completely understood. In this work we have analyzed the effects of sorafenib on autocrine proliferation and survival of different human HCC cell lines. Our results indicate that sorafenib in vitro counteracts autocrine growth of different tumor cells (Hep3B, HepG2, PLC-PRF-5, SK-Hep1). Arrest in S/G2/M cell cycle phases were observed coincident with cyclin D1 down-regulation. However, sorafenib's main anti-tumor activity seems to occur through cell death induction which correlated with caspase activation, increase in the percentage of hypodiploid cells, activation of BAX and BAK and cytochrome c release from mitochondria to cytosol. In addition, we observed a rise in mRNA and protein levels of the pro-apoptotic "BH3-domain only" PUMA and BIM, as well as decreased protein levels of the anti-apoptotic MCL1 and survivin. PUMA targeting knock-down, by using specific siRNAs, inhibited sorafenib-induced apoptotic features. Moreover, we obtained evidence suggesting that sorafenib also sensitizes HCC cells to the apoptotic activity of transforming growth factor-β (TGF-β) through the intrinsic pathway and to tumor necrosis factor-α (TNF) through the extrinsic pathway. Interestingly, sensitization to sorafenib-induced apoptosis is characteristic of liver tumor cells, since untransformed hepatocytes did not respond to sorafenib inducing apoptosis, either alone or in combination with TGF-β or TNF. Indeed, sorafenib effectiveness in delaying HCC late progression might be partly related to a selectively sensitization of HCC cells to apoptosis by disrupting autocrine signals that protect them from adverse conditions and pro-apoptotic physiological cytokines.
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http://dx.doi.org/10.1002/jcp.22843DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3227741PMC
April 2012

Dissecting the effect of targeting the epidermal growth factor receptor on TGF-β-induced-apoptosis in human hepatocellular carcinoma cells.

J Hepatol 2011 Aug 13;55(2):351-8. Epub 2010 Dec 13.

Laboratori d'Oncologia Molecular, Universitat de Barcelona, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.

Background & Aims: Transforming growth factor-beta (TGF-β) induces apoptosis in hepatocytes, a process that is inhibited by the epidermal growth factor receptor (EGFR) pathway. The aim of this work was to ablate EGFR in hepatocellular carcinoma (HCC) cells to understand its role in impairing TGF-β-induced cell death.

Methods: Response to TGF-β in terms of apoptosis was analyzed in different HCC cell lines and the effect of canceling EGFR expression was evaluated.

Results: TGF-β induces apoptosis in some HCC cells (such as Hep3B, PLC/PRF/5, Huh7, or SNU449), but it also mediates survival signals, coincident with the up-regulation of EGFR ligands. Inhibition of the EGFR, either by targeted knock-down with specific siRNA or by pharmacological inhibition, significantly enhances apoptotic response. TGF-β treatment in EGFR targeted knock-down cells correlates with higher levels of the NADPH oxidase NOX4 and changes in the expression profile of BCL-2 and IAP families. However, other HCC cells, such as HepG2, which show over activation of the Ras/ERKs pathway, SK-Hep1, with an endothelial phenotype, or SNU398, where the TGF-β-Smad signaling is altered, show apoptosis resistance that is not restored through EGFR blockade.

Conclusions: The inhibition of EGFR in HCC may enhance TGF-β-induced pro-apoptotic signaling. However, this effect may only concern those tumors with an epithelial phenotype which do not bear alterations in TGF-β signaling nor exhibit an over-activation of the survival pathways downstream of the EGFR.
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http://dx.doi.org/10.1016/j.jhep.2010.10.041DOI Listing
August 2011

The transforming growth factor-beta (TGF-β) mediates acquisition of a mesenchymal stem cell-like phenotype in human liver cells.

J Cell Physiol 2011 May;226(5):1214-23

Biological Clues of Invasive and Metastatic Phenotype Group, Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Spain.

Transforming growth factor-beta (TGF-β) mediates several and sometime opposite effects in epithelial cells, inducing growth inhibition, and apoptosis but also promoting an epithelial to mesenchymal transition (EMT) process, which enhances cell migration and invasion. TGF-β plays relevant roles in different liver pathologies; however, very few is known about its specific signaling and cellular effects in human primary hepatocytes. Here we show that TGF-β inhibits proliferation and induces pro-apoptotic genes (such as BMF or BIM) in primary cultures of human fetal hepatocytes (HFH), but also up-regulates anti-apoptotic genes, such as BCL-XL and XIAP. Inhibition of the epidermal growth factor receptor (EGFR), using gefitinib, abrogates the increase in the expression of the anti-apoptotic genes and significantly enhances cell death. Simultaneously, TGF-β is able to induce an EMT process in HFH, coincident with Snail up-regulation and a decrease in E-cadherin levels, cells showing mesenchymal proteins and reorganization of the actin cytoskeleton in stress fibers. Interestingly, these cells show loss of expression of specific hepatic genes and increased expression of stem cell markers. Chronic treatment with TGF-β allows selection of a population of mesenchymal cells with a de-differentiated phenotype, reminiscent of progenitor-like cells. Process is reversible and the mesenchymal stem-like cells re-differentiate to hepatocytes under controlled experimental conditions. In summary, we show for the first time that human hepatocytes may respond to TGF-β inducing different signals, some of them might contribute to tumor suppression (growth inhibition and apoptosis), but others should mediate liver tumor progression and invasion (EMT and acquisition of a stem-like phenotype).
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http://dx.doi.org/10.1002/jcp.22439DOI Listing
May 2011

Overactivation of the MEK/ERK pathway in liver tumor cells confers resistance to TGF-{beta}-induced cell death through impairing up-regulation of the NADPH oxidase NOX4.

Cancer Res 2009 Oct 22;69(19):7595-602. Epub 2009 Sep 22.

Laboratori de Oncologia Molecular and Departament de Ciències Fisiològiques II, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Universitat de Barcelona, L'Hospitalet de Llobregat, Barcelona, Spain.

Transforming growth factor-beta (TGF-beta) induces apoptosis in hepatocytes, being considered a liver tumor suppressor. However, many human hepatocellular carcinoma (HCC) cells escape from its proapoptotic effects, gaining response to this cytokine in terms of malignancy. We have recently reported that the apoptosis induced by TGF-beta in hepatocytes requires up-regulation of the NADPH oxidase NOX4, which mediates reactive oxygen species (ROS) production. TGF-beta-induced NOX4 expression is inhibited by antiapoptotic signals, such as the phosphatydilinositol-3-phosphate kinase or the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathways. The aim of the present work was to analyze whether resistance to TGF-beta-induced apoptosis in HCC cells is related to the impairment of NOX4 up-regulation due to overactivation of survival signals. Results indicate that inhibition of the MAPK/ERK kinase (MEK)/ERK pathway in HepG2 cells, which are refractory to the proapoptotic effects of TGF-beta, sensitizes them to cell death through a mitochondrial-dependent mechanism, coincident with increased levels of BIM and BMF, decreased levels of BCL-XL and MCL1, and BAX/BAK activation. Regulation of BMF, BCL-XL, and MCL1 occurs at the mRNA level, whereas BIM regulation occurs post-transcriptionally. ROS production and glutathione depletion are only observed in cells treated with TGF-beta and PD98059, which correlates with NOX4 up-regulation. Targeting knockdown of NOX4 impairs ROS increase and all the mitochondrial-dependent apoptotic features by a mechanism that is upstream from the regulation of BIM, BMF, BCL-XL, and MCL1 levels. In conclusion, overactivation of the MEK/ERK pathway in liver tumor cells confers resistance to TGF-beta-induced cell death through impairing NOX4 up-regulation, which is required for an efficient mitochondrial-dependent apoptosis.
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http://dx.doi.org/10.1158/0008-5472.CAN-09-1482DOI Listing
October 2009

Role of CXCR4/SDF-1 alpha in the migratory phenotype of hepatoma cells that have undergone epithelial-mesenchymal transition in response to the transforming growth factor-beta.

Cell Signal 2009 Nov 6;21(11):1595-606. Epub 2009 Jul 6.

Laboratori d'Oncologia Molecular, Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), Spain.

Treatment of FaO rat hepatoma cells with TGF-beta selects cells that survive to its apoptotic effect and undergo epithelial-mesenchymal transitions (EMT). We have established a cell line (T beta T-FaO, from TGF-beta-treated FaO) that shows a mesenchymal, de-differentiated, phenotype in the presence of TGF-beta and is refractory to its suppressor effects. In the absence of this cytokine, cells revert to an epithelial phenotype in 3-4 weeks and recover the response to TGF-beta. T beta T-FaO show higher capacity to migrate than that observed in the parental FaO cells. We found that FaO cells express low levels of CXCR4 and do not respond to SDF-1 alpha. However, TGF-beta up-regulates CXCR4, through a NF kappaB-dependent mechanism, and T beta T-FaO cells show elevated levels of CXCR4, which is located in the presumptive migration front. A specific CXCR4 antagonist (AMD3100) attenuates the migratory capacity of T beta T-FaO cells on collagen gels. Extracellular SDF-1 alpha activates the ERKs pathway in T beta T-FaO, but not in FaO cells, increasing cell scattering and protecting cells from apoptosis induced by serum deprivation. Targeted knock-down of CXCR4 with specific siRNA blocks the T beta T-FaO response to SDF-1 alpha. Thus, the SDF-1/CXCR4 axis might play an important role in mediating cell migration and survival after a TGF-beta-induced EMT in hepatoma cells.
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http://dx.doi.org/10.1016/j.cellsig.2009.06.006DOI Listing
November 2009

The inhibition of the epidermal growth factor (EGF) pathway enhances TGF-beta-induced apoptosis in rat hepatoma cells through inducing oxidative stress coincident with a change in the expression pattern of the NADPH oxidases (NOX) isoforms.

Biochim Biophys Acta 2009 Feb 24;1793(2):253-63. Epub 2008 Sep 24.

Centre d'Oncologia Molecular (COM), IDIBELL-Institut d'Investigació Biomèdica de Bellvitge, L'Hospitalet, Barcelona, Spain.

Transforming growth factor-beta (TGF-beta) induces apoptosis in hepatocytes, through a mechanism mediated by reactive oxygen species (ROS) production. Numerous tumoral cells develop mechanisms to escape from the TGF-beta-induced tumor suppressor effects. In this work we show that in FaO rat hepatoma cells inhibition of the epidermal growth factor receptor (EGFR) with the tyrphostin AG1478 enhances TGF-beta-induced cell death, coincident with an elevated increase in ROS production and GSH depletion. These events correlate with down-regulation of genes involved in the maintenance of redox homeostasis, such as gamma-GCS and MnSOD, and elevated mitochondrial ROS. Nonetheless, not all the ROS proceed from the mitochondria. Emerging evidences indicate that ROS production by TGF-beta is also mediated by the NADPH oxidase (NOX) system. TGF-beta-treated FaO cells induce nox1 expression. However, the treatment with TGF-beta and AG1478 greatly enhanced the expression of another family member: nox4. NOX1 and NOX4 targeted knock-down by siRNA experiments suggest that they play opposite roles, because NOX1 knockdown increases caspase-3 activity and cell death, whilst NOX4 knock-down attenuates the apoptotic process. This attenuation correlates with maintenance of GSH and antioxidant enzymes levels. In summary, EGFR inhibition enhances apoptosis induced by TGF-beta in FaO rat hepatoma cells through an increased oxidative stress coincident with a change in the expression pattern of NOX enzymes.
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http://dx.doi.org/10.1016/j.bbamcr.2008.09.003DOI Listing
February 2009

Upregulation of the NADPH oxidase NOX4 by TGF-beta in hepatocytes is required for its pro-apoptotic activity.

J Hepatol 2008 Dec 19;49(6):965-76. Epub 2008 Sep 19.

Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense, 28040 Madrid, Spain.

Background/aims: The transforming growth factor-beta (TGF-beta) induces apoptosis in hepatocytes through an oxidative stress process. Here, we have analyzed the role of different NADPH oxidase isoforms in the intracellular signalling induced by TGF-beta in hepatocytes, to later explore whether this mechanism is altered in liver tumor cells.

Methods: Primary cultures of rat and human hepatocytes, HepG2 and Hep3B cells were used in in vitro studies to analyze the TGF-beta response.

Results: TGF-beta-induced apoptosis in rat hepatocytes does not require Rac-dependent NADPH oxidases. TGF-beta upregulates the Rac-independent Nox4, which correlates with its pro-apoptotic activity. Regulation of Nox4 occurs at the transcriptional level and is counteracted by intracellular survival signals. siRNA targeted knock-down of Nox4 attenuates NADPH oxidase activity, caspase activation and cell death in rat hepatocytes. NOX4 upregulation by TGF-beta is also observed in human hepatocytes, coincident with apoptosis. In human hepatocellular carcinoma (HCC) cell lines, NOX4 upregulation by TGF-beta is only observed in cells that are sensitive to its cytotoxic effect, such as Hep3B cells. siRNA targeted knock-down of NOX4 in these cells impairs TGF-beta-induced apoptosis.

Conclusions: Upregulation of NOX4 by TGF-beta is required for its pro-apoptotic activity in hepatocytes. Impairment of this TGF-beta-induced response might confer apoptosis resistance in HCC cells.
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http://dx.doi.org/10.1016/j.jhep.2008.07.021DOI Listing
December 2008

Inhibition of the EGF receptor blocks autocrine growth and increases the cytotoxic effects of doxorubicin in rat hepatoma cells: role of reactive oxygen species production and glutathione depletion.

Biochem Pharmacol 2008 May 19;75(10):1935-45. Epub 2008 Feb 19.

Fundació Institut d'Investigació Biomèdica de Bellvitge, Centre d'Oncologia Molecular, Gran Via s/n, Km 2.7, 08907 L'Hospitalet, Barcelona, Spain.

FaO rat hepatoma cells show increased levels of the epidermal growth factor receptor (EGFR) ligands, when compared with adult normal hepatocytes, and higher activity of the TNF-alpha converting enzyme (TACE/ADAM17), which is required for EGFR ligand proteolysis and activation. In this work we have analysed the consequences of inhibiting the EGFR in FaO rat hepatoma cells, focusing the attention on autocrine growth and protection from apoptosis. Results have indicated that FaO cells show overactivation of the EGFR pathway, which induces basal growth (in the absence of serum) and protection from pro-apoptotic agents, such as doxorubicin, generating drug resistance. Treatment of cells with the combination of doxorubicin and the tyrphostin 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG1478, a potent and specific inhibitor of EGFR tyrosine kinase) potently inhibits autocrine growth and induces apoptosis. The apoptotic effect correlates with high expression and activation of the pro-apoptotic Bax and decreased transcript and protein levels of the anti-apoptotic Mcl-1 and Bcl-x(L). Furthermore, the combination of AG1478 and doxorubicin induces reactive oxygen species (ROS) production and glutathione depletion in FaO cells, coincident with up-regulation of the NADPH oxidase NOX4 and down-regulation of the gamma-glutamylcysteine synthetase (gamma-GCS), a key regulatory enzyme of the glutathione synthesis. Incubation of cells with glutathione ethyl ester attenuates the apoptosis induced by the combination of doxorubicin and AG1478, which indicates that glutathione depletion is required for an efficient cell death. In conclusion, targeting EGFR combined with other conventional pro-apoptotic drugs should potentially be effective in antineoplastic therapy towards liver cancer.
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http://dx.doi.org/10.1016/j.bcp.2008.02.015DOI Listing
May 2008

Differential intracellular signalling induced by TGF-beta in rat adult hepatocytes and hepatoma cells: implications in liver carcinogenesis.

Cell Signal 2007 Apr 16;19(4):683-94. Epub 2006 Sep 16.

Centre d'Oncologia Molecular, Institut de Recerca Oncològica-Institut d'Investigació Biomèdica de Bellvitge (IDIBELL), L'Hospitalet, 08907, Barcelona, Spain.

The transforming growth factor-beta (TGF-beta) regulates hepatocyte growth, inhibiting proliferation and inducing apoptosis. Indeed, escaping from the TGF-beta suppressor actions might be a prerequisite for liver tumour progression. In this work we show that TGF-beta plays a dual role in regulating apoptosis in FaO rat hepatoma cells, since, in addition to its pro-apoptotic effect, TGF-beta also activates survival signals, such as AKT, the epidermal growth factor receptor (EGFR) being required for its activation. TGF-beta induces the expression of the EGFR ligands transforming growth factor-alpha (TGF-alpha) and heparin-binding EGF-like growth factor (HB-EGF) and induces intracellular re-localization of the EGFR. Cells that overcome the apoptotic effects of TGF-beta undergo morphological changes reminiscent of an epithelial-mesenchymal transition (EMT) process. In contrast, TGF-beta does not activate AKT in adult hepatocytes, which correlates with lack of EGFR transactivation and no response to EGFR inhibitors. Although TGF-beta induces TGF-alpha and HB-EGF in adult hepatocytes, these cells show very low expression of TACE/ADAM 17 (TNF-alpha converting enzyme), which is required for EGFR ligand proteolysis and activation. Furthermore, adult hepatocytes do not undergo EMT processes in response to TGF-beta, which might be due, at least in part, to the fact that F-actin re-organization induced by TGF-beta in FaO cells require the EGFR pathway. Finally, results indicate that EGFR transactivation does not block TGF-beta-induced cell cycle arrest in FaO cells, but must be interfering with the pro-apoptotic signalling. In conclusion, TGF-beta is a suppressor factor for adult quiescent hepatocytes, but not for hepatoma cells, where it plays a dual role, both suppressing and promoting carcinogenesis.
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http://dx.doi.org/10.1016/j.cellsig.2006.09.002DOI Listing
April 2007