Fig 1: The combination treatment of gamitrinib and HDAC inhibitors interferes with GBM cell energy metabolism. (A) Mitochondrial stress test of U87 cells treated with gamitrinib, panobinostat or the combination of both for 24 h. OM: oligomycin, FCCP: Carbonyl cyanide-4 (trifluoromethoxy) phenylhydrazone, R/A: rotenone and antimycin A (n = 4); (B) The basal respiration and maximal respiration from experiment in (A) were calculated (n = 3, 4); (C) Shown is the OCR/ECAR ratio from mitochondrial stress test of U87 cells treated with the indicated concentrations of gamitrinib, panobinostat or the combination of both for 24 h (n = 4). OCR: oxygen consumption rate, ECAR: extracellular acidification rate (a surrogate for glycolytic activity); (D) U87 and LN229 cells were treated with gamitrinib, panobinostat or the combination of both for 24 h and the whole cell lysates were subjected to protein capillary electrophoresis. The double asterisk indicates a stronger exposure of SDHB. Vinculin is the loading control for U87 and actin is the loading control for LN229. The expression levels of V-ATP5A, I-NDUFB8 and II-SDHB were quantified by using ImageJ (shown in cursive font); (E) U87 cells were transfected with non-targeting (siNT) or TRAP1 specific siRNA (siTRAP1)(single or pool (p)) and the whole cell lysates were subjected to protein capillary electrophoresis; (F) Shown are the respective tracks of ChIP sequencing around SDHA and SDHB of parental and panobinostat treated U87 GBM cells. Highlighted is the transcriptional start site (TSS) by a red rectangle. Shown are means and SD. ANOVA was used for the statistical analysis. ***/**** p < 0.001.
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