Cellular analysis of compounds against eEF-2K activity. MDA-MB-231 tumorigenic breast cells were treated with 75 μM compound 6, 9, or 12 for 1 h in low-glucose media ± 2-DOG, along with the appropriate controls, as described under the experimental section. One set of samples was stimulated with 25 mM 2-DOG (30 min) to activate the AMPK-eEF-2K axis. Cell lysates were then analyzed by Western blotting using antibodies specific for phospho-eEF-2 (Thr-56), total eEF-2, total eEF-2K and pan-Actin as described under the experimental section.