Biomarkers of PIM inhibition. DLBCL cells were incubated for 4 hours with ETP-39010 (10μM) or vehicle DMSO alone (represented as ETP and C, respectively). (A-B) Cellular levels of AKT, pAKT (Ser473), 4E-BP1, and p4E-BP1(Thr37/65) were detected by Western blot using antibodies for these proteins and tubulin as the loading control. Bands were quantified using ImageJ 1.36b (National Institutes of Health), and the ratios of phosphorylated to total protein are presented. The blot is representative of 2 independent experiments. (C) Cellular levels of pBAD (Ser112) and BAD were measured by flow cytometry, and the ratios of phosphorylated to total protein are presented (each column represents ratios from triplicate experiments).