Fig. 4
Top lead GR-28 decreases functional activity of REST and is cytotoxic against high-REST GBM cells. A Effect of GR-28 on REST protein level in A172 cells (4 μM for 24 h) and T98G cells (10 μM for 48 h). Shown is one representative WB replicate (left) and quantification (right) from three to four independent cell treatments (mean ± SEM). Comparison vs DMSO was performed using paired one-tailed t-tests. Dashed lines indicate that adjacent blots were processed on different days. Individual data values are provided in Additional File 10H-I. B Effect of GR-28 on mRNA level of REST-target genes in A172 cells (4 μM for 18 h, left) and T98G cells (10 μM for 36 h, right). Shown are fold changes (FC) vs DMSO derived from three to four independent biological replicates. Gene expression was measured using ddCt method and normalized by ACTB expression. Comparison vs DMSO was performed using paired one-tailed t-tests. Dashed line indicates FC = 1.5. Individual data values are provided in Additional File 10J-K. C Survival rates (72 h) of high-REST GBMs (A172 and T98G) and control cells (HepG2) under single drug treatment with GR-28. Shown are viability rates (mean ± SEM) normalized to that of solvent-control wells derived from three to four independent experiments, n = 9–18. D Sensitivity (72 h) of high-REST GBMs (A172 and T98G) and control cells (HepG2) to GR-28. Shown are LD50s (lethal doses 50) with 95% confidence intervals calculated from three to four independent biological replicates using “drc” R package. TrC = Triacsin C (sensitization at 0.625 and 2.5 μM in T98G and A172, respectively). E Protective effect of REST-OE under GR-28 treatment (24 h) in A172 cells (two independent replicates combined, n = 6). Group comparison was performed using multiple t-tests. Validation of transient REST overexpression is shown on the right side. *p < 0.05, ns not significant