pervanadate and Breast-Neoplasms

pervanadate has been researched along with Breast-Neoplasms* in 7 studies

Other Studies

7 other study(ies) available for pervanadate and Breast-Neoplasms

ArticleYear
Spontaneous and pronase-induced HER2 truncation increases the trastuzumab binding capacity of breast cancer tissues and cell lines.
    The Journal of pathology, 2013, Volume: 229, Issue:3

    A subgroup of HER2-overexpressing breast tumours co-expresses p95(HER2), a truncated HER2 receptor that retains a functional HER2 kinase domain but lacks the extracellular domain, thus impairing trastuzumab binding. We evaluated p95(HER2) expression in 99 frozen breast carcinoma samples by western blot analysis. The HER2-positive cell line BT474 treated with pervanadate or pronase was used as a positive control for p95(HER2) expression. Immunohistochemistry was performed on parallel formalin-fixed, paraffin-embedded sections of the same case series using antibodies directed against either the intra- or extra-cellular binding domain of HER2. In particular, biotinylated trastuzumab (BiotHER) was used to evaluate the binding capacity of the humanized antibody. To avoid a subjective evaluation of the score values and the percentage of immunostained cells, the slides were scanned and automatically analysed. The number of cases with HER2 overexpression (score 3+) and HER2 gene amplification was higher in the p185(HER2)-positive/p95(HER2)-positive samples than in the p185(HER2)-positive/p95(HER2)-negative group. Automated analysis confirmed a significantly higher percentage of 3+ scored cells in p95(HER2)-positive cases. Conversely, the percentage of 2+ scored cells was higher inp95(HER2)-negative cases. The status of the HER2 extracellular domain was then studied using flow cytometry on BT474 cells after pronase enzymatic digestion using trastuzumab and pertuzumab, while the presence of HER2-HER3 dimers was studied using a proximity-ligation assay. In vitro experiments showed that short-term pronase digestion of BT474 cells produced two HER2 fragments (of 95 and 150 kDa, detectable in tissue specimens as well), increased the binding affinity of trastuzumab, reduced the rate of HER2-HER3 dimers, and did not interfere with pertuzumab-binding capacity. In conclusion, the presence of p95(HER2 as detected by western blot analysis does not compromise the immunohistochemical detection of HER2. Our data suggest that a reduction of the receptor steric hindrance as induced by enzymatic shedding may facilitate the binding capacity of trastuzumab.

    Topics: Antibodies, Monoclonal, Humanized; Antineoplastic Agents; Blotting, Western; Breast Neoplasms; Cell Line, Tumor; DNA, Neoplasm; Enzyme Inhibitors; Female; Flow Cytometry; Humans; In Situ Hybridization, Fluorescence; Pronase; Protein Binding; Receptor, ErbB-2; Trastuzumab; Vanadates

2013
PTP1B suppresses prolactin activation of Stat5 in breast cancer cells.
    The American journal of pathology, 2010, Volume: 177, Issue:6

    Basal levels of nuclear localized, tyrosine phosphorylated Stat5 are present in healthy human breast epithelia. In contrast, Stat5 phosphorylation is frequently lost during breast cancer progression, a finding that correlates with loss of histological differentiation and poor patient prognosis. Identifying the mechanisms underlying loss of Stat5 phosphorylation could provide novel targets for breast cancer therapy. Pervanadate, a general tyrosine phosphatase inhibitor, revealed marked phosphatase regulation of Stat5 activity in breast cancer cells. Lentiviral-mediated shRNA allowed specific examination of the regulatory role of five tyrosine phosphatases (PTP1B, TC-PTP, SHP1, SHP2, and VHR), previously implicated in Stat5 regulation in various systems. Enhanced and sustained prolactin-induced Stat5 tyrosine phosphorylation was observed in T47D and MCF7 breast cancer cells selectively in response to PTP1B depletion. Conversely, PTP1B overexpression suppressed prolactin-induced Stat5 tyrosine phosphorylation. Furthermore, PTP1B knockdown increased Stat5 reporter gene activity. Mechanistically, PTP1B suppression of Stat5 phosphorylation was mediated, at least in part, through inhibitory dephosphorylation of the Stat5 tyrosine kinase, Jak2. PTP1B knockdown enhanced sensitivity of T47D cells to prolactin phosphorylation of Stat5 by reducing the EC(50) from 7.2 nmol/L to 2.5 nmol/L. Immunohistochemical analyses of two independent clinical breast cancer materials revealed significant negative correlations between levels of active Stat5 and PTP1B, but not TC-PTP. Collectively, our data implicate PTP1B as an important negative regulator of Stat5 phosphorylation in invasive breast cancer.

    Topics: Breast Neoplasms; Carcinoma; Down-Regulation; Drug Synergism; Dual Specificity Phosphatase 3; Enzyme Inhibitors; Female; Humans; Phosphorylation; Prolactin; Protein Tyrosine Phosphatase, Non-Receptor Type 1; Protein Tyrosine Phosphatase, Non-Receptor Type 11; Protein Tyrosine Phosphatase, Non-Receptor Type 2; Protein Tyrosine Phosphatase, Non-Receptor Type 6; STAT5 Transcription Factor; Tumor Cells, Cultured; Vanadates

2010
Tyrosine kinase p56lck regulates cell motility and nuclear factor kappaB-mediated secretion of urokinase type plasminogen activator through tyrosine phosphorylation of IkappaBalpha following hypoxia/reoxygenation.
    The Journal of biological chemistry, 2003, Dec-26, Volume: 278, Issue:52

    Nuclear factor kappaB (NFkappaB) plays major role in regulating cellular responses as a result of environmental injuries. The molecular mechanism(s) by which hypoxia/reoxygenation (H/R) regulates p56lck-dependent activation of NFkappaB through tyrosine phosphorylation of IkappaBalpha and modulates the expression of downstream genes that are involved in cell migration in human breast cancer cells are not well defined. In this paper, we investigated the involvement of protein-tyrosine kinase p56lck in the redox-regulated activation of NFkappaB following H/R in highly invasive (MDA-MB-231) and low invasive (MCF-7) breast cancer cells. We demonstrated that H/R induces tyrosine phosphorylation of p56lck, nuclear translocation of NFkappaB, NFkappaB-DNA binding, and transactivation of NFkappaB through tyrosine phosphorylation of IkappaBalpha. Transfection of these cells with wild type Lck but not with mutant Lck F394 followed by H/R induces the tyrosine phosphorylation of inhibitor of nuclear factor kappaB (IkappaBalpha) and transcriptional activation of NFkappaB, and these are inhibited by Lck inhibitors. In vitro kinase assay demonstrated that immunoprecipitated p56lck but not Lyn or Fyn directly phosphorylate IkappaBalpha in presence of H/R. Pervanadate, H2O2, and H/R induce the interaction between Lck and tyrosine-phosphorylated IkappaBalpha, and this interaction is inhibited by Src homology 2 domain inhibitory peptide, suggesting that tyrosine-phosphorylated IkappaBalpha interacts with Src homology 2 domain of Lck. Luciferase reporter gene assay indicated that Lck induces NFkappaB-dependent urokinase type plasminogen activator (uPA) promoter activity in presence of H/R. Furthermore, H/R stimulates the cell motility through secretion of uPA. To our knowledge, this is the first report that p56lck in presence of H/R regulates NFkappaB activation, uPA secretion, and cell motility through tyrosine phosphorylation of IkappaBalpha and further demonstrates an important redox-regulated pathway for NFkappaB activation following H/R injury that is independent of IkappaB kinase/IkappaBalpha-mediated signaling pathways.

    Topics: Active Transport, Cell Nucleus; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Cell Nucleus; DNA, Complementary; Enzyme Inhibitors; Genes, Reporter; Humans; Hydrogen Peroxide; Hypoxia; I-kappa B Proteins; Luciferases; Lymphocyte Specific Protein Tyrosine Kinase p56(lck); Microscopy, Fluorescence; Models, Biological; Mutation; NF-kappa B; NF-KappaB Inhibitor alpha; Oxidation-Reduction; Oxygen; Phosphorylation; Plasmids; Precipitin Tests; Promoter Regions, Genetic; Protein Binding; Serine; Signal Transduction; Time Factors; Transfection; Tyrosine; Urokinase-Type Plasminogen Activator; Vanadates

2003
Regulation of the EphA2 kinase by the low molecular weight tyrosine phosphatase induces transformation.
    The Journal of biological chemistry, 2002, Oct-18, Volume: 277, Issue:42

    Intracellular signaling by protein tyrosine phosphorylation is generally understood to govern many aspects of cellular behavior. The biological consequences of this signaling pathway are important because the levels of protein tyrosine phosphorylation are frequently elevated in cancer cells. In the classic paradigm, tyrosine kinases promote tumor cell growth, survival, and invasiveness, whereas tyrosine phosphatases negatively regulate these same behaviors. Here, we identify one particular tyrosine phosphatase, low molecular weight tyrosine phosphatase (LMW-PTP), which is frequently overexpressed in transformed cells. We also show that overexpression of LMW-PTP is sufficient to confer transformation upon non-transformed epithelial cells. Notably, we show that the EphA2 receptor tyrosine kinase is a prominent substrate for LMW-PTP and that the oncogenic activities of LMW-PTP result from altered EphA2 expression and function. These results suggest a role for LMW-PTP in transformation progression and link its oncogenic potential to EphA2.

    Topics: Breast Neoplasms; Cell Line, Transformed; Cells, Cultured; Chelating Agents; Dose-Response Relationship, Drug; Egtazic Acid; Electrophoresis, Polyacrylamide Gel; Enzyme Inhibitors; Epithelial Cells; Gene Expression Regulation, Enzymologic; Humans; Immunoblotting; Isoenzymes; Oligonucleotides, Antisense; Phosphorylation; Phosphotyrosine; Precipitin Tests; Protein Tyrosine Phosphatases; Proto-Oncogene Proteins; Receptor, EphA2; Signal Transduction; Time Factors; Transfection; Tumor Cells, Cultured; Tyrosine; Vanadates

2002
Role of Src kinases in the ADAM-mediated release of L1 adhesion molecule from human tumor cells.
    The Journal of biological chemistry, 2000, May-19, Volume: 275, Issue:20

    The ectodomain of certain transmembrane molecules can be released by proteolysis, and the solubilized antigens often exert important biological functions. We demonstrated before that the L1 adhesion molecule is shed from the cell surface. Here we show that L1 release in AR breast carcinoma cells is mediated by a member of the disintegrin metalloproteinase (ADAM) family of proteinases. Up-regulation of L1 shedding by phorbol ester or pervanadate involved distinct mechanisms. Pervanadate induced shedding and rounding-up of cells from the substrate, which was blocked by the Src kinase inhibitor PP2. Tyr phosphorylation of the L1 cytoplasmic tail and the Src kinase Fyn was observed following pervanadate treatment. Up-regulation of L1 release and activation of Fyn occurred also when cells were detached by EDTA suggesting that the regulation of L1 shedding by this pathway was linked to cell morphology and adhesion. The phorbol 12-myristate 13-acetate-induced shedding was inhibited by the protein kinase C inhibitor bisindolylmaleimide I and by PD98059, a specific inhibitor of the mitogen-activated protein kinase pathway. Soluble L1 binds to the proteoglycan neurocan and in bound form could support integrin-mediated cell adhesion and migration. We propose that the release of cell-associated adhesion molecules such as L1 may be relevant to promote cell migration.

    Topics: Animals; Antigens, Surface; Breast Neoplasms; Cell Size; CHO Cells; Cricetinae; Disintegrins; Enzyme Inhibitors; Female; Flavonoids; Humans; Leukocyte L1 Antigen Complex; Membrane Glycoproteins; Metalloendopeptidases; Mice; Neural Cell Adhesion Molecules; Neuroblastoma; Phosphorylation; Protease Inhibitors; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-fyn; Recombinant Proteins; Signal Transduction; src-Family Kinases; Tetradecanoylphorbol Acetate; Transfection; Tumor Cells, Cultured; Vanadates

2000
Cleavage of the HER2 ectodomain is a pervanadate-activable process that is inhibited by the tissue inhibitor of metalloproteases-1 in breast cancer cells.
    Cancer research, 1999, Mar-15, Volume: 59, Issue:6

    HER2/neu, a Mr 185,000 tyrosine kinase receptor that is overexpressed in breast cancer, undergoes proteolytic cleavage of its extracellular domain (ECD). In contrast with other membrane-bound proteins, including growth factor receptors, that are cleaved by a common machinery system, we show that HER2 cleavage is a slow process and is not activated by protein kinase C. Pervanadate, a general inhibitor of protein-tyrosine phosphatases, induces a rapid and potent shedding of HER2 ECD. The shedding of HER2 ECD is inhibited by the broad-spectrum metalloprotease inhibitors EDTA, TAPI-2, and batimastat. The tissue inhibitor of metalloproteases-1; an inhibitor of matrix metalloproteases that does not inhibit cleavage by the general protein kinase C-dependent shedding machinery, also inhibited HER2 ECD shedding, whereas tissue inhibitor of metalloproteases-2 did not. These data suggest that HER2 cleavage is a process regulated by an as-yet-unidentified distinct protease.

    Topics: Breast Neoplasms; Endopeptidases; Humans; Hydroxamic Acids; Metalloendopeptidases; Phenylalanine; Phosphorylation; Protease Inhibitors; Receptor, ErbB-2; Thiophenes; Tissue Inhibitor of Metalloproteinase-1; Tumor Cells, Cultured; Tyrosine; Vanadates

1999
The receptor-like protein-tyrosine phosphatase DEP-1 is constitutively associated with a 64-kDa protein serine/threonine kinase.
    The Journal of biological chemistry, 1997, May-02, Volume: 272, Issue:18

    Protein-tyrosine phosphatases (PTPs) are involved in the regulation of diverse cellular processes and may function as positive effectors as well as negative regulators of intracellular signaling. Recent data demonstrate that malignant transformation of cells is frequently associated with changes in PTP expression or activity. Our analysis of PTP expression in mammary carcinoma cell lines resulted in the molecular cloning of a receptor-like PTP, also known as DEP-1. DEP-1 was found to be expressed at varying levels in mammary carcinoma cell lines and A431 cells. In all tumor cell lines analyzed, DEP-1 was constitutively phosphorylated on tyrosine residues. Phosphorylation of DEP-1 increased significantly after treatment of cells with the PTP inhibitor pervanadate. In A431 cells, tyrosine phosphorylation of DEP-1 was also observed after stimulation with epidermal growth factor, however, only after prolonged exposure of the cells to the ligand, suggesting an indirect mechanism of phosphorylation. In addition, DEP-1 coprecipitated with several tyrosine-phosphorylated proteins from pervanadate-treated cells. In vitro binding experiments using a glutathione S-transferase fusion protein containing the catalytically inactive PTP domain of DEP-1 (Gst-DEP-1-C/S) identify these proteins as potential substrates of DEP-1. In addition, we found a 64-kDa serine/threonine kinase to be constitutively associated with DEP-1 in all tumor cell lines tested. The 64-kDa kinase forms a stable complex with DEP-1 and phosphorylates DEP-1 and DEP-1-interacting proteins in vitro. These data suggest a possible mechanism of DEP-1 regulation in tumor cell lines involving serine/threonine and/or tyrosine phosphorylation.

    Topics: Adenosine Triphosphate; Amino Acid Sequence; Breast Neoplasms; Cell Line; Cloning, Molecular; Conserved Sequence; Female; Humans; Kinetics; Molecular Weight; Phosphoproteins; Phosphorylation; Protein Serine-Threonine Kinases; Protein Tyrosine Phosphatases; Receptor-Like Protein Tyrosine Phosphatases, Class 3; Recombinant Proteins; Transfection; Tumor Cells, Cultured; Vanadates

1997
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