Cytoplasmic Estrogen Receptor in Breast Cancer
ABSTRACT In addition to genomic signaling, it is accepted that estrogen receptor-α (ERα) has nonnuclear signaling functions, which correlate with tamoxifen resistance in preclinical models. However, evidence for cytoplasmic ER localization in human breast tumors is less established. We sought to determine the presence and implications of nonnuclear ER in clinical specimens.
A panel of ERα-specific antibodies (SP1, MC20, F10, 60c, and 1D5) was validated by Western blot and quantitative immunofluorescent (QIF) analysis of cell lines and patient controls. Then eight retrospective cohorts collected on tissue microarrays were assessed for cytoplasmic ER. Four cohorts were from Yale (YTMA 49, 107, 130, and 128) and four others (NCI YTMA 99, South Swedish Breast Cancer Group SBII, NSABP B14, and a Vietnamese Cohort) from other sites around the world.
Four of the antibodies specifically recognized ER by Western and QIF analysis, showed linear increases in amounts of ER in cell line series with progressively increasing ER, and the antibodies were reproducible on YTMA 49 with Pearson correlations (r(2) values) ranging from 0.87 to 0.94. One antibody with striking cytoplasmic staining (MC20) failed validation. We found evidence for specific cytoplasmic staining with the other four antibodies across eight cohorts. The average incidence was 1.5%, ranging from 0 to 3.2%.
Our data show ERα is present in the cytoplasm in a number of cases using multiple antibodies while reinforcing the importance of antibody validation. In nearly 3,200 cases, cytoplasmic ER is present at very low incidence, suggesting its measurement is unlikely to be of routine clinical value.
Full-textDOI: · Available from: Lisa Ryden, Jul 16, 2014
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ABSTRACT: The translation of basic research into improved therapies for breast cancer patients requires relevant pre-clinical models that incorporate spontaneous metastasis. We have completed a functional and molecular characterisation of a new isogenic C57BL/6 mouse model of breast cancer metastasis, comparing and contrasting it with the established BALB/c 4T1 model. Metastatic EO771.LMB tumours were derived from parental poorly metastatic EO771 mammary tumours. Functional differences were evaluated using both in vitro assays and spontaneous metastasis assays in mice. Results were compared to non-metastatic 67NR and metastatic 4T1.2 tumours of the 4T1 model. Protein and transcript levels of markers of human breast cancer molecular subtypes were measured in the four tumour lines, as well as p53 tumour suppressor gene status and responses to Tamoxifen in vivo and in vitro. Array-based expression profiling of whole tumours identified genes and pathways that were deregulated in metastatic tumours. EO771.LMB cells metastasised spontaneously to lung in C57BL/6 mice and displayed increased invasive capacity compared to parental EO771. By immunohistochemical assessment, EO771 and EO771.LMB were basal-like, as was the 4T1.2 tumour, while 67NR had a luminal phenotype. Primary tumours from all lines were negative for progesterone receptor, Erb-b2/Neu, and cytokeratin 5/6, but positive for epidermal growth factor receptor. Only 67NR displayed nuclear estrogen receptor alpha positivity. EO771 and EO771.LMB expressed mutant p53 while 67NR and 4T1.2 were p53 null. Integrated molecular analysis of both the EO771/EO771.LMB and 67NR/4T1.2 pairs indicated that upregulation of MMP-3, Pthlh and S100a8 and downregulation of Cd36 may be causally involved in metastatic dissemination of breast cancer.Disease Models and Mechanisms 01/2015; DOI:10.1242/dmm.017830 · 5.54 Impact Factor
Molecular Endocrinology 09/2014; 28(9):1403-7. DOI:10.1210/me.2014-1230 · 4.20 Impact Factor
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ABSTRACT: Tamoxifen is still the most frequently used antiestrogen for the treatment of patients with premenopausal, estrogen receptor positive breast cancer. However, in 20-30% of these cases, tamoxifen therapy fails due to an existing or developing resistance. The prediction of tamoxifen resistance by appropriate biomarker analysis and the development of novel therapies for tamoxifen resistance in premenopausal breast cancer is, therefore, an important goal of ongoing research. Tamoxifen resistance is associated with altered estrogen receptor expression especially on the plasma membrane, including the alternative G-protein coupled receptor GPR-30 (GPER) and estrogen receptor splice products, such as ERα36. Tamoxifen resistant cells often use alternative pathways to promote proliferation in the absence of genomic estrogen signaling. These pathways involve the epidermal growth factor EGF, the inflammation associated transcription factor NF-κB- and the IGF-1 pathway. Tamoxifen resistant mamma carcinoma cell lines are useful models to understand tamoxifen resistance in-vitro and to search for prognostic or predictive biomarkers. Furthermore, such cell lines can be used to identify potential targets for therapy. Copyright © 2015 Elsevier GmbH. All rights reserved.Pathology - Research and Practice 01/2015; 211(3). DOI:10.1016/j.prp.2015.01.004 · 1.56 Impact Factor