Mem nonessential amino acids solution 100x

The human cell lines ALL-SIL, SKW-3/KE-37, CTV-1, MEC-1, JEKO-1, REC-1, and Granta-519 were purchased from the Leibniz Institut DSMZ-German collection of microorganisms and cell cultures (Germany). Cells were cultured in RPMI 1640 (#MT10040CV, Thermo Fisher Scientific, Waltham MA, USA) with 10% or 20% fetal bovine serum (FBS) (#10270–106, Thermo Fisher Scientific), 1% penicillin-streptomycin (P/S) (#3MT30002CI, Thermo Fisher Scientific), and 1% of MEM Non-Essential Amino Acids Solution (100X) (#11140050, Thermo Fisher Scientific), 2 mM L-Glutamine (#25030-081, Thermo Fisher Scientific), and 1% HEPES Buffer 1M (#MS013D1006, Biowest, Nuaillé, France). Granta-519 was maintained in DMEM (#11960-044, Thermo Fisher Scientific) with 20% FBS, 1% P/S, and 2 mM L-Glutamine (#25030-081, Thermo Fisher Scientific). Cell lines were grown in a humidified incubator at 37 °C, and 5% CO₂ and monitored for mycoplasma contamination.

The H1_DL2 cell line used in this study is based on the H1 cell line isolated from a patient biopsy of human melanoma brain metastases as described previously [47 (link)]. The H1_DL2 cell line was developed by transducing H1 cells with two lentiviral vectors encoding Luciferase and a GFP variant of Dendra [37 (link)]. The cells were cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum (Gibco), 2% L-glutamine (Thermofisher Scientific), and MEM Non-Essential Amino Acids Solution (100X) (Thermofisher Scientific) diluted 1:25. The growth medium was exchanged twice a week.
Female NOD/SCID mice (eight weeks of age, 18–22 g of weight) were purchased from Harlan. The mice were housed in individually ventilated cages (Techniplast). In accordance with the recommendations set forth by the Federation of European Laboratory Animal Science Associations, animal housing conditions were free of specific pathogens. The mice were provided with sterile water and food ad libitum. All animal procedures were approved by the Norwegian National Animal Research Authorities, Mattilsynet, https://www.mattilsynet.no/.

Two cell lines were used, i) cervical cancer cells (Caski) and ii) fibroblast cells (HFL). Caski cells were cultured in Gibco RPMI 1640 Medium 1X, supplemented with 10% fetal bovine serum (FBS - Gibco) and 1% Penicillin-Streptomycin. For HFL1, we used 15% fetal bovine serum (FBS - Gibco), 1% Penicillin-Streptomycin (Sigma), 2 mM glutamine (Gibco), MEM-NEAA (MEM Non-Essential Amino Acids Solution (100X) - Gibco) and DMEM Medium 4.5 g/L D-glucose (Sigma)/Nutrient Mixtures F-12 HAM (Sigma) (v 1:1). Cell lines were maintained at 37 °C and 5% CO2 in a humidified incubator.

In this study, the experiments were performed on triple negative breast cancer (TNBC) cell lines, Hs578T and MDA-MB-231. The Hs578T cell line was cultured in D-MEM high glucose (D-MEM Gibco®) supplemented with 10% fetal bovine serum (FBS- Gibco®), 2 mM L-glutamine (Gibco®), 1% MEM Non-Essential Amino Acids Solution (100X, Gibco®), 0.01 mg/ml insulin and 1% Penicillin-Streptomycin (Gibco®). MDA-MB-231 cell line was cultured in RPMI-1640 (RPMI-1640 Gibco®), supplemented with 10% fetal bovine serum (FBS- Gibco®), 2 mM L-glutamine (Gibco®) and 1% Penicillin-Streptomycin (Gibco®). Cells were maintained in a humidified atmosphere at 37 °C with 95% air and 5% of CO₂ (carbon dioxide). The doxorubicin-resistant TNBC cells were established by multiple dose exposure. The drug concentration used for maintaining the drug resistance of Hs578T/Dox and MDA-MB-231/Dox was 50 nM. Cells were treated with this dose for 12, and respectively 24 passages (Figure S1).

The human promonocytic cell lines U-937 and THP-1, the murine myeloma Ag8 cells (obtained from the American Type Culture Collection, ATCC, Manassas, VA), and THP-1 L2 cells (THP-1 cells with low expression of CD13 [18 (link)]) were cultured in RPMI-1640 medium (Gibco by Life Technologies, NY, USA). HMEC-1 microvasculature endothelial cells (originally from ATCC, obtained from Dra. Dolores Correa, Instituto Nacional de Pediatría, México) were cultured in MCDB-131 medium supplemented according to the manufacturer's instructions (Invitrogen, Carlsbad, CA, USA). HEK-293 cells (ATCC CRL-1573™) or HEK-ANPEP cells (HEK-293 cells expressing human CD13-GFP, obtained as previously described [18 (link)]) were cultured in DMEM-high glucose (Invitrogen, CA, USA). J744-hCD13 mouse macrophages expressing human CD13 [18 (link)] were cultured in DMEM-high glucose (Gibco by Life Technologies, USA). All media were supplemented with 10% heat-inactivated FBS (Invitrogen, Carlsbad, CA, USA), 2 mM L-glutamine, 100 μg/ml streptomycin, 100 U/ml penicillin (Sigma-Aldrich, St. Louis MO, USA), 1 mM sodium pyruvate solution, and 1% MEM nonessential amino acids solution (100x) (Gibco by Life Technologies, NY, USA). Cultures were maintained at 37°C in a humidified atmosphere with 6% CO₂.