Global characterization of signalling networks associated with tamoxifen resistance in breast cancer

Authors

  • Brigid C. Browne,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Falko Hochgräfe,

    1. Competence Center – Functional Genomics, Junior Research Group Pathoproteomics, University of Greifswald, Germany
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  • Jianmin Wu,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
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  • Ewan K. A. Millar,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. School of Medical Sciences, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
    3. Department of Anatomical Pathology, South Eastern Area Laboratory Service, St George Hospital, Kogarah, New South Wales, Australia
    4. School of Medicine and Health Sciences, University of Western Sydney, Campbelltown, New South Wales, Australia
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  • Jane Barraclough,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Andrew Stone,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Rachael A. McCloy,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Christine S. Lee,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Caroline Roberts,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Naveid A. Ali,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Alice Boulghourjian,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
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  • Fabian Schmich,

    1. Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
    2. Swiss Institute of Bioinformatics, Basel, Switzerland
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  • Rune Linding,

    1. Cellular Signal Integration Group, Center for Biological Sequence Analysis, Department of Systems Biology, Technical University of Denmark, Lyngby, Denmark
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  • Lynn Farrow,

    1. Cardiff School of Pharmacy & Pharmaceutical Sciences, Cardiff University, UK
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  • Julia M. W. Gee,

    1. Cardiff School of Pharmacy & Pharmaceutical Sciences, Cardiff University, UK
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  • Robert I. Nicholson,

    1. Cardiff School of Pharmacy & Pharmaceutical Sciences, Cardiff University, UK
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  • Sandra A. O'Toole,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
    3. Department of Tissue Pathology and Diagnostic Oncology, Royal Prince Alfred Hospital, Camperdown, New South Wales, Australia
    4. Sydney Medical School, University of Sydney, Westmead, New South Wales, Australia
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  • Robert L. Sutherland,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
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    • Deceased
  • Elizabeth A. Musgrove,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
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  • Alison J. Butt,

    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
    Current affiliation:
    1. Research Investment, National Breast Cancer Foundation, Sydney, New South Wales, Australia
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    • These authors contributed equally to this work
  • Roger J. Daly

    Corresponding author
    1. Cancer Research Program, The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, New South Wales, Australia
    2. St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Darlinghurst, New South Wales, Australia
    3. Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Melbourne, Victoria, Australia
    • Correspondence

      R. J. Daly, Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Level 1, Building 77, Monash University, Melbourne, Victoria 3800, Australia

      Fax: +61 3 990 29500

      Tel: +61 3 990 29301

      E-mail: roger.daly@monash.edu

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    • These authors contributed equally to this work

Abstract

Acquired resistance to the anti-estrogen tamoxifen remains a significant challenge in breast cancer management. In this study, we used an integrative approach to characterize global protein expression and tyrosine phosphorylation events in tamoxifen-resistant MCF7 breast cancer cells (TamR) compared with parental controls. Quantitative mass spectrometry and computational approaches were combined to identify perturbed signalling networks, and candidate regulatory proteins were functionally interrogated by siRNA-mediated knockdown. Network analysis revealed that cellular metabolism was perturbed in TamR cells, together with pathways enriched for proteins associated with growth factor, cell–cell and cell matrix-initiated signalling. Consistent with known roles for Ras/MAPK and PI3-kinase signalling in tamoxifen resistance, tyrosine-phosphorylated MAPK1, SHC1 and PIK3R2 were elevated in TamR cells. Phosphorylation of the tyrosine kinase Yes and expression of the actin-binding protein myristoylated alanine-rich C-kinase substrate (MARCKS) were increased two- and eightfold in TamR cells respectively, and these proteins were selected for further analysis. Knockdown of either protein in TamR cells had no effect on anti-estrogen sensitivity, but significantly decreased cell motility. MARCKS expression was significantly higher in breast cancer cell lines than normal mammary epithelial cells and in ER-negative versus ER-positive breast cancer cell lines. In primary breast cancers, cytoplasmic MARCKS staining was significantly higher in basal-like and HER2 cancers than in luminal cancers, and was independently predictive of poor survival in multivariate analyses of the whole cohort (< 0.0001) and in ER-positive patients (= 0.0005). These findings provide network-level insights into the molecular alterations associated with the tamoxifen-resistant phenotype, and identify MARCKS as a potential biomarker of therapeutic responsiveness that may assist in stratification of patients for optimal therapy.

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