Fig. 1: GDC-0941 (Pictilisib) and Rapamycin affect PI3K-mediated signalling in glioblastoma cell lines

a The effect of the indicated concentrations of the two pharmacological inhibitors, GDC-0941 from Selleckchem (Munich, Germany) (GDC) left and Rapamycin from Sigma-Aldrich (Hamburg, Germany) (Rapa) right, on the viability of two glioblastoma cell lines, U87 (upper panel) and A172 (lower panel). Viability was assessed by MTT assay, while cell lines were obtained from ATCC (Manassas, VA, USA) and cultured as previously described^15,40. b Specific DNA fragmentation was used as surrogate readout for apoptosis as assessed by flow cytometric analysis of propidium iodide-stained nuclei, as previously described⁴⁰, for up to 144 h treatment with either 0.6 μM GDC-0941 (left) or 5 nM Rapamycin (right). c Relative changes in cell numbers (control set at 1 for each individual time point), for up to 144 h treatment with either GDC-0941 (left) or Rapamycin (right). Cells were counted using CASY1 DT (Innovatis, Reutlingen, Germany), as previously described²⁶. d Specific DNA fragmentation used as surrogate readout for apoptosis as assessed by flow cytometric analysis of propidium iodide-stained nuclei, for a 144 h treatment with either 0.6 μM GDC-0941 (left) or 5 nM Rapamycin (right) and 100 μM temozolomide from Sigma-Aldrich (TMZ) and a combination of either inhibitor with TMZ (Combi). Red line indicates additive effect, i.e. below red line: antagonism, above red line: synergism. e Relative changes in cell numbers (solvent control set at 1), for a 144 h treatment with either 0.6 μM GDC-0941 (left) or 5 nM Rapamycin (right) and 100 μM temozolomide (TMZ) and a combination of either inhibitor with TMZ (Combi). Red line indicates additive effect, i.e. below red line: synergism, above red line: antagonism. f U87 (left) and A172 (right) cells were incubated with 0.6 μM GDC-0941, 5 nM Rapamycin or solvent (control). Scratches were introduced 1 h after treatment initiation and from then semi-directional cell migration was examined after 16 h by fixation of the cells and subsequent staining, as previously described¹⁵. Scale bars equal 500 μm. Experiments shown in a were performed at least three times in sextet, shown are mean and +SD, while b–e show results of at least three independent experiments performed in triplicate, shown are mean and +SD. An unpaired two-tailed t test was used for statistical evaluation (*p < 0.05, **p  < 0.01, ***p < 0.001). In f, one of two representative independent experiments performed in triplicate is shown