U 87 mg

SKRC-17 (kind gift from J. Vissers, Nijmegen), RCC-53 (derived from a patient with stage IV disease (pT2N1MxG2-3)), sunitinib-resistant SKRC-17 and RCC-53 [18 (link)], DU145, cabacitaxel-resistant DU-145 [19 (link),20 (link)], U87-MG (from European Collection of Authenticated Cell Cultures), temozolomide-resistant U87-MG, and MFE-319 (established from the primary adenosquamous endometrium carcinoma, grading G1–G2) [22 (link)] were cultured in RPMI 1640 medium, and EN (primary endometrial carcinoma, stage IIIC) [21 (link)] was cultured in DMEM GlutaMAX medium, both supplemented with 10% fetal calf serum (FCS “Gold Plus”, Bio & Sell GmbH, Feucht, Germany), 2 mM L-glutamine, 1 mM sodium pyruvate, and 1% minimal essential medium (Invitrogen, Life Technologies GmbH, Darmstadt, Germany) at 37 °C in humidified incubator with 5% CO₂. CSCs were cultured in DMEM/F12 culture medium, containing 2% B-27 (Invitrogen, Life Technologies, Darmstadt, Germany) 10 ng/mL human recombinant basic fibroblast growth factor (bFGF, Sigma Aldrich Chemie GmbH, Taufkirchen, Germany), and 10 ng/mL epidermal growth factor (EGF, Sigma Aldrich).

The human glioblastoma cell lines LN229 (ATCC-CRL-2611), U251 (ECACC 89081403; formerly known as U373MG), U87MG (ECACC 89081402), and T98G (ECACC No. 92090213) were obtained from the European Collection of Authenticated Cell Cultures (ECACC, Salisbury, UK) or the American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured as described previously [24 (link)]. Human primary GBM cultures (n = 2) were produced by dissociation and cultured according to established techniques as described before [24 (link)]. Human primary GBM stem-like cell cultures (n = 8) as well as GBM cell line-derived stem-like cells were established and intensively characterized by the formation of neurospheres, the ability to survive and proliferate under stem cell conditions, and the ability to differentiate into more mature cells as described before [3 (link),25 (link),26 (link),27 (link)]. The purity of the GBM cells was ascertained by immunostaining with cell type-specific markers and by the absence of contamination with mycoplasms. GBM cell line identity was verified by short tandem repeat profiling at the Department of Forensic Medicine (Kiel, Germany) using the Powerplex HS Genotyping Kit (Promega, Madison, WI, USA) and the 3500 Genetic Analyzer (Thermo Fisher Scientific, Waltham, MA, USA) as previously described [3 (link)].

The following cell lines were used in the research: MOLT-4 (T-cell acute lymphoblastic leukemia), U87-MG (human glioblastoma), MDA-MB-231 (human breast adenocarcinoma), A431 (epidermoid carcinoma), MCF-7 (breast cancer) and normal human fibroblasts. The MOLT-4 and U87-MG cells were purchased from the European Collection of Authenticated Cell Cultures (ECACC, Salisbury, UK), the MDA-MB-231 cells were purchased from Cell Biolabs (San Diego, California, USA), the A431 and MCF-7 cell lines were purchased from American Type Culture Collection (ATCC, Manassas, Virginia, USA), the human fibroblasts were purchased from Celther Polska (Lodz, Poland). All the cell lines were maintained at 37 °C in an atmosphere of 5% CO₂ in the medium appropriate for the cell type. The MCF-7 and A431 cells were cultured in Dulbecco's modified Eagle's medium (Sigma-Aldrich, St. Louis, MO) supplemented with 10% fetal bovine serum (FBS) (Sigma-Aldrich, St. Louis, MO), MOLT-4 in RPMI 1640 medium (Sigma-Aldrich, St. Louis, MO) with 10% FBS, U87-MG in MEM medium containing 10% FBS, MDA-MB-231 cells in DMEM medium with 10% FBS supplemented with l-glutamine and non-essential amino acids. All media were enriched in antibiotics – 100 U per L penicillin G and 100 U per L streptomycin.