JY-1-106 sensitizes tumor cells to metabolic stress. (A) A549 cells were exposed to metabolic stress and serial doses of JY-1-106. After 72 hours, cell viability was measured using the XTT assay. The results shown are the average of triplicate assays and the experiment was repeated twice. (B) A549 cells were subjected to metabolic stress with or without the addition of JY-1-106 for 48 hours. The same numbers of viable cells were used for each time point. The intracellular ATP levels in these cells were measured fluorometrically. The differences between these two groups at each time point were statistically evaluated using a Student’s t-test (p < 0.01). (C) Western blot analysis of BH3-only proteins after metabolic stress. A549 cells were exposed to metabolic stress medium for 0, 1, 3, 6, 12 and 24 hours and analyzed by western blotting using appropriate monoclonal and polyclonal antibodies specific for PUMA and Bim proteins. (D) Determination of apoptotic cells with the Annexin V binding assay. A549 cells were labeled with Annexin V-FITC and counterstained with PI after exposure to JY-1-106 or metabolic stress medium or combination of JY-1-106 or metabolic stress medium for 24 hours. The cells were then analyzed by flow cytometry. The differences between combination and JY-1-106 or TAXOL were measured using the Student’s t-test (p < 0.01). The final results presented are the average of three independent experiments.