Fig. 1

HCQ blocked autophagy and increased the sensitivity of the PI3K inhibitor BKM120 in tumor cells. GFP-LC3 puncta (A) and RFP-GFP-LC3 puncta (B) in SKOV-3, MKN-1 and HBC-5 tumor cells treated with 1 μM BKM120 or/and 20 μM HCQ (the same concentrations in subsequent studies unless indicated) for 24 h. The number of GFP-LC3 puncta per cell was quantified from 50 randomly selected cells (n = 3). C Western blot analysis of LC3B and p62 in SKOV-3, MKN-1 and HBC-5 cells treated with BKM120 and/or HCQ for 48 h. D Cell viability was measured by PrestoBlue after treatment with BKM120 (0.5 μM, 1 μM and 2 μM) alone or in combination with 20 μM HCQ for 72 h. E Colony-forming abilities of cells treated with HCQ and/or BKM120 were determined. Colony quantification is graphed in (F). G Cells were treated with BKM120 and/or HCQ for 48 h and then subjected to flow cytometric analysis of apoptosis with Annexin V-FITC/PI staining. H FACS quantification of the total apoptotic cell population, including Annexin V+/propidium iodide−early apoptotic cells and Annexin V+/propidium iodide+late apoptotic cells. I Western blot analysis of Caspase-3 and PARP in SKOV-3, MKN-1 and HBC-5 cells treated with HCQ and/or BKM120. Scale bars, 10 μm. Graphs are normalized to 100% per treatment and are shown as the mean ± SD from three independent experiments; *p < 0.05; **p < 0.01; ***p < 0.001