Penicillin and streptomycin

Human neuron cell line SY5Y (ATCC CRL-2266) were obtained from American Type Culture Collection, USA. HEK293 cells (BCRC90016) were obtained from Bioresource Collection and Research Center, Taiwan. SY5Y cells and HEK293 cells were maintained in DMEM supplemented with 10% FBS (Invitrogen, Waltham, MA, USA), 1% penicillin and streptomycin (Biowest, Loire Valley, France), and 1% L-glutamine (Invitrogen) in a humidified incubator under an atmosphere of 5% CO₂ at 37 °C. Human neural stem cells (NSCs) were induced from human iPSCs. In brief, human iPSCs were first cultured as embryoid bodies (EBs) in EB medium on Matrigel (BD Biosciences; Franklin Lakes, NJ, USA)‐coated dishes and supplemented with recombinant Noggin protein (250 ng/ml, R&D). On day 10, medium was replaced with EB medium supplemented with Sonic Hedgehog (SHH, 20 ng/ml, R&D) and fibroblast growth factor 8 (100 ng/ml, R&D). Upon the appearance of rosette‐like structures (day 14), medium was changed to medium supplemented with BDNF, ascorbic acid, SHH and FGF8. On day 22, FGF8 was withdrawn, and cells were maintained in the medium supplemented with BDNF, ascorbic acid and SHH. On day 29, cells were seeded on poly‐L‐ornithine/laminin‐coated dishes in complete StemPro NSC Medium and were then expanded up to 10 passages.

The normal human mammary epithelial cells (MCF-10A), human breast cancer cells (MCF-7), human ovarian cancer cells (A278), and human ovarian fibroblasts (HOF) healthy cell lines were purchased from American Type Culture Collection (ATCC). MCF-7 was grown in the Dulbecco’s Modified Eagle’s Medium-DMEM (BioWest) containing 1% mixture of penicillin and streptomycin (BioWest), 1% of L-glutamine (BioWest) and 10% fetal bovine serum (FBS, Gibco). To cultivate MCF-10A the Dulbecco's Modified Eagle Medium, nutrient mixture F-12 was used. The medium was supplemented with 5% horse serum (Sigma-Aldrich), 10 ng·mL⁻¹ epithelial growth factor (Sigma-Aldrich), 5 μg·mL⁻¹ hydrocortisone (Sigma-Aldrich) and 10 μg·mL⁻¹ human insulin (Sigma-Aldrich). All cell cultures were mycoplasma free. To cultivate the A2780 cell lines the RPMI 1640 (BioWest) containing 1% mixture of penicillin and streptomycin (BioWest), 10% fetal bovine serum (FBS, Gibco) and 1% of L-glutamine (BioWest) was used. The HOF cells were cultivated by fibroblast medium that consisted of 500 ml of basal medium, 10 ml of fetal bovine serum (FBS), 5 ml of fibroblasts growth supplement (FGS), and 5 ml of antibiotic solution (P/S).

Origin of the four human melanoma cell lines used in this study (MeWo, Mel-2a, SK-Mel-19 and Mel-HO) was described previously [25 (link)]. All cell lines have significant tyrosinase mRNA expression [25 (link), 26 (link)]. Human embryonal kidney (HEK-293) cells were used for adenovirus amplification and quantification. All cell lines were cultivated in Dulbecco’s modified Eagle’s medium (DMEM, 4.5 g/l glucose, Biowest, Nuaillé, France), 10% fetal calf serum (c.c.pro, Oberdorla, Germany) and 1% of each penicillin and streptomycin (Biowest, Nuaillé, France) at 37 °C and 5% CO₂. Soluble TRAIL (Adipogen, San Diego, CA, USA) was added to the cell culture in a concentration of 50 ng/ml.

HCT-116, HT-29 (colon cancer) and CCD-841 (non-cancerous) cell lines were obtained from ATCC (VA, USA). Cells were cultured in DMEM medium (HyClone, Logan, Utah, USA) supplemented with 10% FBS and 1% penicillin and streptomycin (Biowest, France). Cells were incubated in incubator at 37 °C with 5% CO₂. Negative control for all assays was represented by 0.02% DMSO (vehicle) in untreated medium.

After informed consent from each individual, a skin punch biopsy from the left upper arm was performed for the index patient (IV.2), her affected brother (IV.3), and her healthy cousin (IV.7).
Primary human skin fibroblasts from patients and controls were prepared from the skin biopsies as described previously,²¹ (link) and cultivated in T75 flasks with minimal essential medium (MEM; Biowest, Renningen, Germany) supplemented with 20% fetal bovine serum (Biowest), 1.3% L-glutamine (Biowest), and 0.8% penicillin/streptomycin at 37°C in a humidified incubator with 5% CO₂. HEK293T cells were cultured in DMEM (Biowest) supplemented with 10% fetal bovine serum (Biowest), 1% L-glutamine (Biowest), and 1% penicillin and streptomycin (Biowest) at 37°C in a humidified incubator with 5% CO₂.

Mouse astrocyte cell line ALT (BCRC60581) and mouse cerebral endothelial cells bEnd.3 (BCRC60515) were obtained from Bioresource Collection and Research Center, Taiwan. ALT and bEnd.3 cells were maintained in DMEM supplemented with 10% FBS (Invitrogen, Waltham, MA, USA), 1% penicillin and streptomycin (Biowest, Loire Valley, France), and 1% l‐glutamine (Invitrogen) in a humidified incubator under an atmosphere of 5% CO₂ at 37 °C.