2ME2 increases NF-κB activity in LNCaP but not in DU145 or PC3 cells. A. Western blot analysis showing that treatment of LNCaP but not in DU145 or PC3 cells with 5 μM 2ME2 (M) for 24 h increases phospho (P)-IκBα and P-p65 compared to control (C). Total levels of IκBα and p65 do not change, except for a decrease of IκBα in LNCaP. Coomassie blue stain of total protein is loading control. B. Western blot analysis showing that 2ME2 (M; 5 μM) and Doc (D; 1 nM) decrease IκBα levels at 48 and 72 h in LNCaP and LN-AI compared to control (C) cells. In DU145 cells, M and D did not decrease IκBα levels. C. Luciferase reporter assay showing that 2ME2 increases NF-κB activity (fold above TK promoter) in LNCaP cells at 24 h and remains high at 72 h compared to control (C) cells. In PC3 cells, the basal NF-κB activity is 7.3-fold higher compared to LNCaP cells and 2ME2 reduces activity after 24 h. After 72 h, NF-κB activity is similar to control cells. n = 6, three independent experiments; *, P < 0.02.