fig5

Figure 5. GLS1 knockdown increased panobinostat activity. (A) Gene expression of GLS1 was analyzed using RT-PCR, and protein expression was examined through immunoblotting using antibodies against GLS1 and β-actin. Statistical significance is indicated as ^****P < 0.0001 compared to control shRNA-transfected cells. (B) Viability of shRNA-transfected U266 cells was assessed. (C-E) shRNA-transfected U266 cells were incubated with panobinostat. (C) Cell viability, (D) caspase 3/7 activity, and (E) cytotoxicity were evaluated. Statistical significance is denoted as ^****P < 0.0001 compared with untreated cells. (F) Intracellular ATP levels in shRNA-transfected U266 cells. Statistical significance is denoted as ^****P < 0.0001 compared with mock-transfected cells. (G) PBMC from leukemia and normal plasma samples were treated with panobinostat and/or CB-839 for 72 h, and cell viability was assessed. ^****P < 0.0001 compared to control. ns: not significant. The experiments were conducted twice. GLS: Glutaminase; RT-PCR: reverse transcription-polymerase chain reaction; shRNA: short hairpin RNA; ATP: adenosine triphosphate; PBMC: peripheral blood mononuclear cell.