GX15-070 and bortezomib combination enhances Bak-dependent apoptotic signaling in MCL cell lines. (A) Jeko cells were treated with 0.5 μM GX15-070 and/or 10 nM bortezomib for 5 hours. Mcl-1 immunoprecipitation was performed as described in “Patients, materials, and methods,” analyzing Mcl-1–bound and –unbound fractions by Western blotting for Mcl-1, Bak, and Noxa proteins. Western blot images are representative results from 3 independent experiments. (B) Jeko cells were treated with 0.5 μM GX15-070 and/or 10 nM bortezomib for 18 hours. Bax/Bak conformational changes, caspase-3 activation, loss of ΔΨ_m, and PS exposure were analyzed as described in “Patients, materials, and methods.” The percentage inside each chart refers to the population in black. These experiments have been performed twice with similar results, and therefore 1 representative experiment is shown. (C) NOXA siRNA and nonsilencing siRNA were introduced in Jeko cells by electroporation as described in “Patients, materials, and methods.” Total RNA was isolated 6 hours after transfection. NOXA mRNA levels were determined by quantitative RT-PCR (Taqman technology) using GUS as a housekeeping gene. The results showed are the mean ± SD of 2 different experiments. (D) Jeko cells transfected with nonsilencing siRNA (ns-siRNA) and with NOXA siRNA (NOXA-siRNA) were treated with 0.5 μM GX15-070 and/or 10 nM bortezomib for 18 hours. Loss of ΔΨ_m and Bak conformational change were analyzed as described in “Patients, materials, and methods.” The percentage inside each chart refers to the population in black.