Endosomes containing ERα are present in normal and tumor human breast tissues. (a) Top: Confocal images of a normal human breast tissue (reduction mammoplasty; sample N211) stained for ERα (HC-20 clone), β1-integrin, and DAPI. In the inset, arrows indicate the presence of ERα⁺ endosomes. Similar results were obtained in the four different specimens analyzed. Bottom: Confocal images of a human breast tumor (Luminal A subtype adenocarcinoma; sample T171) stained for ERα (HC-20 clone), β1-integrin, and DAPI. Yellow arrows indicate ERα⁺ endosomes. In the inset, arrows indicate the presence of ERα⁺ endosomes. Similar results were obtained in the three different specimens analyzed. (b) Magnification from the inset shown in panel a (top). Diameters of ER⁺ vesicles are shown on the right. (c) Table showing mean and SD of Pearson’s correlation index calculated for the overall colocalization between ERα and β1-integrin. Differences between groups were analyzed by two-tailed Student’s t test (per replicate: n_normal = six fields; n_tumor = seven fields). (d) OncoPrint from http://www.cbioportal.org (Cerami et al., 2012; Gao et al., 2013) showing the alterations found in ERα (ESR1) and β1-integrin (ITGB1) genes in different patients obtained from the search in four different datasets: British Columbia, Nature 2014 (Eirew et al., 2015); TCGA, Nature 2012 (Cancer Genome Atlas Network, 2012); TCGA, Cell 2015 (Ciriello et al., 2015); and Nature 2012 and Nature Communications 2016 (Pereira et al., 2016). (e) Kaplan–Meier plot of the overall survival of patients with alterations in ESR1 or ITGB1 genes using thelargest and newest dataset available in http://www.cbioportal.org (Breast Cancer-METABRIC; Cerami et al., 2012; Gao et al., 2013). Significance level after the log-rank test is shown in the plot. *, P < 0.05; **, P < 0.01; ***, P < 0.001. Bars, 20 µm (unless otherwise indicated). The results shown in this study are in whole or part based on data generated by the TCGA Research Network: http://cancergenome.nih.gov/.