eIF4E KD and selinexor synergistically inhibit MM cell proliferation. Dox-inducible eIF4E KD U266 cells were treated with selinexor (200 nM, unless otherwise indicated) and/or dox (500 ng/mL) for 72 hours. (A) The protein expression of eIF4E level was detected using WB with β-actin as a loading control. The densities of the bands were measured using NIH ImageJ and normalized for β-actin. The immunoblot is representative of 3 independent experiments. (B) Cell proliferation was assessed and the combination of doxycycline and selinexor was compared with selinexor alone. ∗P < .05. Quantification of the maximal inhibitory effects of selinexor is shown underneath the bar graph. (C) Example dot plots (top) and quantification of replicates (bottom) for cell apoptosis were detected via flow cytometry. (D) Immunofluorescence microscopy for eIF4E (red) and with 4′,6-diamidino-2-phenylindole for nuclear counterstaining (blue). Scale bar, 20 μM. (E) Cytoplasmic and nuclear fractions were extracted and analyzed using WB using antibodies against XPO1, eIF4E, IKZF1, and c-MYC. LaminB1 and β-actin were blotted as the markers for nuclear and cytoplasmic fractions, respectively.