Figure 5. miR-181a/b sensitize cancer cells to Olaparib treatment. (A) Colony formation of MDA-MB-231 cells transfected with miR-181a, miR-181b, ATM (siATM) or control siRNA (CTRL) and treated with 1 µM Olaparib for 10 d. Percentage of surviving fraction are reported. Surviving fractions were calculated as ratio between plating efficiency in Olaparib-treated cells and plating efficiency in untreated cells. Plating efficiency was obtained according the following formula: # of colonies/# of plated cells. (B) MDA-MB-231 cells were transfected with a combination of miR-181a and miR-181b or control siRNA and 72 h hour later splitted and treated with 10 µM Olaparib. After 7 d, cells were harvested and half of them permeabilized and stained with propidium iodide, and the other half stained with PI/Annexin V, and analyzed by flow cytometry. The percentage of sub G₁ population (black columns) and Annexin V positive cells (gray columns) are reported. (C) SUM159PT, OVCAR-3 and PANC1 cells transfected with a combination of miR-181a and miR-181b or control siRNA, were treated with 10 µM Olaparib for 4 (SUM159PT and PANC1) or 7 (OVCAR) d. Cells were then harvested, stained with propidium iodide and subjected to FACS analysis. The percentage of SubG1 population is reported. A representative western blot showing the levels of ATM and Vinculin as loading control is reported. (D) SUM159PT cells transfected with a combination of antagomiR against miR-181a and miR-181b (ant-181a/b) or a control antagomir mutated in five residues (Ant-181mut), and treated as in (C). A representative western blot showing the levels of ATM and Vinculin as loading control is reported. Graphs show means and s.d. for at least three independent experiments. p values were calculated with two tailed t-test: *, p < 0.05; **, p < 0.01; ***, p < 0.001.