Heterogeneity of estrogen receptor expression in circulating tumor cells from metastatic breast cancer patients

PLoS One. 2013 Sep 18;8(9):e75038. doi: 10.1371/journal.pone.0075038. eCollection 2013.


Background: Endocrine treatment is the most preferable systemic treatment in metastatic breast cancer patients that have had an estrogen receptor (ER) positive primary tumor or metastatic lesions, however, approximately 20% of these patients do not benefit from the therapy and demonstrate further metastatic progress. One reason for failure of endocrine therapy might be the heterogeneity of ER expression in tumor cells spreading from the primary tumor to distant sites which is reflected in detectable circulating tumor cells (CTCs).

Methods: A sensitive and specific staining protocol for ER, keratin 8/18/19, CD45 was established. Peripheral blood from 35 metastatic breast cancer patients with ER-positive primary tumors was tested for the presence of CTCs. Keratin 8/18/19 and DAPI positive but CD45 negative cells were classified as CTCs and evaluated for ER staining. Subsequently, eight individual CTCs from four index patients (2 CTCs per patient) were isolated and underwent whole genome amplification and ESR1 gene mutation analysis.

Results: CTCs were detected in blood of 16 from 35 analyzed patients (46%), with a median of 3 CTCs/7.5 ml. In total, ER-negative CTCs were detected in 11/16 (69%) of the CTC positive cases, including blood samples with only ER-negative CTCs (19%) and samples with both ER-positive and ER-negative CTCs (50%). No correlation was found between the intensity and/or percentage of ER staining in the primary tumor with the number and ER status of CTCs of the same patient. ESR1 gene mutations were not found.

Conclusion: CTCs frequently lack ER expression in metastatic breast cancer patients with ER-positive primary tumors and show a considerable intra-patient heterogeneity, which may reflect a mechanism to escape endocrine therapy. Provided single cell analysis did not support a role of ESR1 mutations in this process.

Publication types

  • Clinical Trial
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Breast Neoplasms / blood*
  • Breast Neoplasms / genetics*
  • Breast Neoplasms / pathology
  • Estrogen Receptor alpha / blood*
  • Estrogen Receptor alpha / genetics*
  • Female
  • Gene Amplification
  • Gene Expression Regulation, Neoplastic*
  • Humans
  • Keratins / blood
  • Keratins / genetics
  • Leukocyte Common Antigens / blood
  • Leukocyte Common Antigens / genetics
  • Neoplastic Cells, Circulating / metabolism*
  • Neoplastic Cells, Circulating / pathology


  • ESR1 protein, human
  • Estrogen Receptor alpha
  • Keratins
  • Leukocyte Common Antigens
  • PTPRC protein, human

Grant support

This research was supported by the Roggenbuck-Stiftung. KP was supported by ERC advanced investigator grant: ERC-2010-AdG_2010317, grant agreement number 269081, DISSECT; and BMBF grant 01ES0724, Metfinder. SJ was supported by the Research Promotion Fund of the Medical Faculty UKE (FFM). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.