Inhibition of SIRT1 sensitizes leukemic cells to genotoxic stress-mediated p53 activation and apoptosis. (A) Western blot of Molm-14 cells treated for 4 hours, as indicated, probed with antibodies directed against acetylated p53, p53, and Actin. (B) RQ-PCR analysis of p21 (top) and BAX and Puma (bottom) in Molm-14 cells treated as indicated. Target gene expression was normalized to GAPDH, and fold change is shown relative to untreated control (p21 and BAX) or irradiation only (Puma; because of no detectable expression in untreated cells). Data represent mean values ± SD of 3 independent experiments. (C) Analysis of apoptosis in BA/F3-FLT3-ITD cells infected with lentivirus expressing 2 different shRNA clones directed against p53 or scrambled shRNA after treatment with TV-6 and PKC412 (left). Immunoblot analysis demonstrating p53 knockdown efficacy (right). (D) Analysis of apoptosis after treatment with TV-6 (left) or SIRT1-knockdown (right) and genotoxic agents, as indicated. The percentage of sub-G1 cells corresponding to apoptotic cells was determined by flow cytometry. Data represent the mean values ± SD of 3 independent experiments. *P < .05, **P < .01, ***P < .001; unpaired Student t test.