A, U251-MG cells were treated with JSI-124 (1 μM) for the indicated times. Cells were lysed and immunoblotted with the indicated Ab. B, U251-MG cells were treated with JSI-124 (1 μM) for the indicated times, followed by isolation of RNA, generation of cDNA and quantitative RT-PCR using Taqman primers. Data represent mean ± SD, replicates of three (*, p<0.001; ANOVA). C, U251-MG cells were treated with TNF-α (10 ng/ml) for 15 min or JSI-124 (1 μM) for 15 or 30 min. Cells were lysed and nuclear and cytosolic fractions were isolated, followed by immunoblotting with the indicated Ab. Caspase 3 and PARP Ab were used to verify purity of cytosolic and nuclear fractions, respectively. D, U251-MG cells were treated with TNF-α (10 ng/ml) or JSI-124 (1 μM) for the indicated times. Cells were lysed and immunoblotted with the indicated Ab.