Breast Cancer Research and Treatment

, Volume 129, Issue 2, pp 623–628 | Cite as

Simultaneous measurement of ERα, HER2, and PhosphoERK1/2 in breast cancer cell lines by flow cytometry

  • Ulas Darda Bayraktar
  • Tae Kon Kim
  • Katherine Drews-Elger
  • Cara Benjamin
  • Dorraya El-Ashry
  • Eric Wieder
  • Krishna V. Komanduri
Brief Report

Abstract

The activation of human epidermal growth factor receptor-2 (HER2) results in the activation of the mitogen-activated protein kinase (MAPK) cascade that may lead to the resistance to anti-estrogen therapy in estrogen receptor (ERα) expressing breast cancer by means of phosphorylation of ERα in the N-terminal region by phosphorylated extracellular signal-regulated kinase 1/2 (pERK1/2) and by means of decreasing ERα expression. Immunohistochemistry is the most widely used technique for the detection of ERα and HER2 in breast cancer specimens, however, is inadequate in its ability to assess the relationship between ERα, HER2, and MAPK cascade at the single cell level. To clear this major hurdle, we devised a novel flow cytometric method to quantify the expression of ERα, HER2, and the activation of MAPK cascade simultaneously in single cells. The method was validated by concurrent Western blotting in established cell lines: MDA-231 (ERα and HER2-negative), MCF-7 (ERα-positive, HER2-negative), MCF-7 cells overexpressing ERα after long-term incubation in estrogen-free medium, and HER2 transfected MCF7 cells. Using the flow cytometry method, we confirmed the previous finding that ERα expression is down-regulated upon epidermal growth factor mediated ERK1/2 phosphorylation in EGFR/MCF-7 cells. To our knowledge, this is the first such assay to incorporate simultaneous single cell measurement for all of these pathways, which may prove useful to determine the intratumoral heterogeneity in breast tumors or the receptor status in circulating tumor cells.

Keywords

Estrogen receptor Human epidermal growth factor receptor Extracellular signal-regulated kinase Multicolor flow cytometry 

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Copyright information

© Springer Science+Business Media, LLC. 2011

Authors and Affiliations

  • Ulas Darda Bayraktar
    • 1
  • Tae Kon Kim
    • 1
  • Katherine Drews-Elger
    • 1
  • Cara Benjamin
    • 1
  • Dorraya El-Ashry
    • 1
  • Eric Wieder
    • 1
  • Krishna V. Komanduri
    • 1
  1. 1.Department of Medicine, Division of Hematology/OncologyUniversity of MiamiMiamiUSA

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