Zr 75 1

The human adenocarcinoma cell lines MCF-7, T47D, ZR-75-1, and MDA-MB-231, and an endometrial cancer cell line Ishikawa (Heraklio) 02 ER- (Ishikawa ERneg), were obtained from the European Collection of Cell Cultures (ECACC, Salisbury, UK) and the Cama-1 (HTB-21), SK-BR-3 (HTB-30), HCC1937 (CRL-2336), and MDA-MB-468 (HTB-132) cell lines from the American Type Culture Collection (ATCC, Rockville, MD, USA). The HCC 3153 cell line was kindly provided by Adi F. Gazdar (Hamon Center for Therapeutic Oncology Research and Department of Pathology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX, USA). Cryopreservation of cell cultures ranged from passages 1 to 10. Cells were used during up to 20 passages. Cells were grown routinely in Dulbecco’s modified Eagle’s medium (Biochrom, Berlin, Germany), supplemented with 10% FBS (PAA, Pasching, Austria).
For cytospin preparation, trypsinized cells were centrifuged (700 g, 10 min, 4 °C) and resuspended in phosphate-buffered saline (PBS; Biochrom, Berlin, Germany). Then, 1 million cells were spread on each cytospin and centrifuged (45 g, 5 min, room temperature). Cytospins were allowed to dry overnight at room temperature and then stored at −80 °C. They were prepared with either 1 million adenocarcinoma cells or mixed with PBMCs from healthy volunteer donors, as indicated in the legends.

Oestrogen receptor(ER)-positive (MCF-7, ZR-75-1 and T47D), triple-negative (MDA-MB-231, HCC 1806), and ER-negative/HER2-positive (SK-BR-3) human breast carcinoma cell lines and MCF-10A, a non-malignant immortalised mammary cell line, were purchased from European Collection of Cell Cultures (Wiltshire, UK). HMEC, primary human mammary epithelial cells were purchased from Life Technologies (Carlsbad, CA). HEK-293FT cells were purchased from Invitrogen (Carlsbad, CA) as a part of BLOCK-iT Lentiviral RNAi Expression System. HEK293-T cells were obtained from DSMZ (Germany), as reported previously.^{7 (link)} See Supplementary Methods for culturing conditions.

ZR-75-1, T47D, MCF7 and SKBR3 cell lines were purchased from the European Collection of Cell Cultures (Wiltshire, UK). All cell lines were maintained through continuous passaging, and were confirmed to be free of contamination by Mycoplasma spp. Cells were maintained in DMEM (ZR-75-1 cell line) or RPMI-1640 (other cell lines) media (Sigma-Aldrich, St. Louis, MO, USA), with 10% FBS (Sigma-Aldrich), 100 U/mL penicillin, 100 μg/mL streptomycin (Gibco, Rockville, MD, USA) and 1.46 mg/mL L-glutamine (Gibco). Additionally, the growth medium for ZR-75-1 was supplemented with 1 nM β-estradiol (Sigma-Aldrich), and did not contain sodium pyruvate, which mediates elimination of H₂O₂ from the culture medium [21 (link)]. Tissue culture experimental techniques are further described in the Supplementary Methods (Additional file 1).