Mda mb 231

Human MDA-MB-231, MCF7, and HeLa cell lines were obtained from the European Collection of Authenticated Cell Cultures and BT-20 and MCF 10A cell lines from the American Type Culture Collection. Cells were maintained in culture for less than 20 passages after receipt under standard conditions (37°C, 5% CO₂) and were regularly tested for mycoplasma contamination. All cell lines except MCF 10A cells were maintained in Dulbecco’s Modified Eagle Medium from Gibco, supplemented with 10% fetal bovine serum and antibiotics. MCF 10A cells were cultured using the MEGM^™ Mammary Epithelial Cell Growth Medium BulletKit^™ (Lonza). Cells were treated with the following drugs: veliparib (ABT-888; Enzo Life Sciences), olaparib (AZD2281; MedChemExpress), a PARG inhibitor (PDD 00017273; Tocris, bio-techne), H₂O₂ (Sigma-Aldrich), rapamycin (Sigma-Aldrich), triptolide (Sigma-Aldrich), lactacystin (Sigma-Aldrich).

The human triple-negative breast adenocarcinoma (MDA-MB-231) cell line was purchased from the European Collection of Authenticated Cell Cultures, cultured in L15 medium supplemented with 15% fetal calf serum (FCS) (Sigma-Aldrich, St. Louis, MO, USA), and maintained at 37°C without CO₂ equilibration. The myeloma (Karpas 707H) cell line was kindly gifted by Dr. Karpus from Cambridge University. Human placental choriocarcinoma (JEG-3 and BeWo) and embryonic kidney (HEK-293) cells were purchased from the American Type Collection Center (ATCC) (CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045) and stored at the Tokai University School of Medicine. JEG-3 cells were cultured in low-glucose DMEM supplemented with 10% FCS and maintained at 37°C under 5% CO₂. BeWo cells were cultured in Ham’s F12 medium supplemented with 10% FCS and maintained at 37°C under 5% CO₂. HEK-293 cells were cultured in high-glucose DMEM supplemented with 10% FCS and maintained at 37°C in 5% CO₂. Karpas 707H cells were cultured in RPMI-1640 medium (Nissui Co. Ltd., Tokyo, Japan) supplemented with 10% FCS and maintained at 37°C under 5% CO₂.

The human cervical carcinoma HeLa, human
ovarian carcinoma A2780, and human breast adenocarcinomas MDA-MB-231
and MCF-7 were purchased from the European collection of authenticated
cell cultures ECACC (Salisbury, UK), human monocytic cell line Thp-1
was purchased from the American Type Culture Collections (ATCC), human
colon carcinoma HCT-116 was kindly provided by Dr. M. Brazdova, Institute
of Biophysics (Brno, CZ), and cisplatin-resistant human ovarian carcinoma
A2780cisR (a cisplatin-resistant variant of A2780 cells) was kindly
provided by Professor B. Keppler, University of Vienna (Austria).
A2780 and A2780cisR cells were grown in RPMI-1640 medium (Biosera,
Boussens, France) supplemented with gentamycin (50 μg mL^–1, Serva, Heidelberg, Germany) and 10% heat-inactivated
FBS (Biosera). The other cells were grown in DMEM medium (high glucose
4.5 g L^–1, PAA) supplemented with gentamycin (50
μg mL^–1, Serva) and 10% heat-inactivated FBS
(Biosera). The cells were cultured in a humidified incubator at 310
K in a 5% CO₂ atmosphere and subcultured 2–3 times
a week with a desired plating density.

The cancer cell lines utilized for our investigations, including human ovarian carcinoma (A2780), human cervix carcinoma (Hela), human breast adenocarcinomas (MCF-7 and MDA-MB-231), and mouse embryonic fibroblast cell line (NIH/3T3) were purchased from the European Collection of Authenticated Cell Cultures (ECACC, Salisbury, UK). Another cervical cell line (SiHa) was acquired from the American Tissue Culture Collection (Manassas, VA, USA). For optimal growth, all cell lines were maintained in minimal essential medium (MEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS), 1% MEM non-essential amino acid solution (MEM-NEAA), and 1% penicillin/streptomycin mixture, at 37 °C, in a humidified incubator, containing 5 % carbon dioxide (CO₂). All the utilized media and supplements were purchased from Lonza Group Ltd. (Basel, Switzerland). Unless otherwise specified, the chemicals and kits used for the experiments were purchased from Merck Life Science Ltd. (Budapest, Hungary).

Human breast cell line MDA-MB-231 was obtained from the European Collection of Authenticated Cell Cultures (ECACC) and cultured at 37°C in 5% CO₂ humidity according to standard mammalian tissue culture protocols in Dulbecco’s Modified Eagle’s Medium + GlutaMax (Gibco, Waltham, MA) containing 10% fetal bovine serum (FBS) (Gibco, Waltham, MA) and 100 units/ml penicillin/100 μg/ml streptomycin (Gibco, Waltham, MA).
Athymic nude female mice (AthymicNude-Foxn1nu, ENVIGO, Indianapolis, IN) of 5 weeks of age were used to develop the breast cancer xenograft model.

The human PD-L1-expressing CHO-K1 cells overexpressing TCR activator (CHO/TCRAct/PD-L1) cells and Jurkat ECs T cells overexpressing PD-1 and carrying a luciferase reporter gene under the control of the Nuclear Factor of Activated T cells Response Element (NFAT-RE) were obtained from Promega and cultured in an RPMI-1640 medium (Biowest, Nuaillé, France) supplemented with 10% fetal bovine serum (FBS, Biowest, Nuaillé, France) and 200 mM L-glutamine (Biowest, Nuaillé, France). G418 (250 µg/mL, InvivoGen, San Diego, CA, USA) and Hygromycin B Gold (50 µg/mL, InvivoGen, San Diego, CA, USA) were also added to the culture medium as selection antibiotics. CAKI 2, LNCap, PC-3, and MDA-MB-231 cells were purchased from the European Collection of Authenticated Cell Cultures (ECACC) and cultured according to the provided protocol.

Human breast cancer cells, MCF-7, T47D, MDA-MB 231 and Hs578T were purchased from the European Collection of Authenticated Cell Cultures (ECACC) and maintained as previously published [23 (link)]. UBR60-bcl2 cell line was a generous gift from Professor Paul Harkin (Queen’s University of Belfast, Ireland). The UBR60-bcl2 cell line expresses BRCA1 under the control of tetracycline-regulated promoter and has previously been described elsewhere [25 (link)].