Fig. 4.

Increased production of reactive oxygen species (ROS) in 17-DMAG treated cells. (a) HT29 cells were treated with 17-DMAG (1 µM) and/or NAC (20 mM) for 16 h. Cells were trypsinized and labeled with C-400 or C-368 (10 µg/ml) for 15 min at 37°C. FACS analysis was performed to measure intracellular ROS production as indicated by C-400 dye fluorescenc using the C-368 signal as a reference measurement for drug uptake. Relative ROS production is normalized to non-treated cells. (b) HT29 cells were treated with 17-DMAG (1 µM), radicicol (3 µM), and menadione (50 µM) for 16 h as indicated. Cell lysates were subjected to western blotting analysis for the expression of BRAF(V600E), phosphorylated MEK1/2 (p-MEK) and total MEK1/2 (pan-MEK).