Sunitinib malate

GSCs were grown in Neurobasal medium, and supplemented with B27 1:50 (Invitrogen, Carlsbad, CA), epidermal growth factor (Sigma; 20 ng/ml) and basic fibroblast growth factor (20 ng/ml), as previously described (34 (link)). U87MG, U373, LN18 and T98G (ATCC) cells were cultured in DMEM containing 10% FBS and antibiotics. U87MG DK and EGFRvIII cells were generously provided by Dr. Matthew J. Lazzara (University of Pennsylvania). Staurosporine (Cell Signaling) was used induce cell death at 1–2 µM for four hrs. To create chemotherapeutic microenvironmental stress, we combined serum starvation with Temozolomide (TMZ; 250 µm) for 72 hrs. To activate MAPK signaling pathway, human epidermal growth factor (hEGF) was used at a concentration of 100ng/mL for 30 min. A dual mTOR kinase inhibitor, AZD8055 (Selleck Chemicals), was used at a final concentration of 500 nM. The RTK inhibitors sunitinib malate and AG-1478 hydrochloride (Tocris Bioscience) were used at a final concentration of 10 µM for 24 hrs. RTK inhibitor SU6668 (Tocris Bioscience) was used at a final concentration of 10 µM in T4121 cells and 50 µM in U373 cells for 24 hrs. For inhibiting methylation, 5 azacytidine was used at 10 µM final concentration for 6 days and dosing medium was changed every two days. EZH2 inhibitor (3-deazaneplanocin A; DZNep) was used for 24 hrs at a final concentration of 10 µM.