Figure 2.

Bortezomib enhances PP2A activity, which is regulated by CIP2A, and down-regulates CIP2A and P-Akt in sensitive leukemia cells. (A) Analysis of PP2A activity in drug-treated leukemia cells. Columns, mean (n=3); bars, SD; *P<0.05. Cells were treated with DMSO or bortezomib at 50 nM or okadaic acid at 20 nM (as negative control) or forskolin ⁴⁰ μM (as a positive control) for 24 h. Cell lysates were prepared for detecting PP2A activity. (B) Analysis of PP2A activity in CIP2A-overexpressed HL-60 cells. Ectopic expression of CIP2A reduced PP2A activity in HL-60 cells. Columns, mean (n=3); bars, SD; *P<0.05. (C) (Top) Dose-dependent analysis of CIP2A, P-Akt and caspase-3. Cells were exposed to bortezomib (BTZ) for 6 h at the indicated doses. Cell lysates were prepared and assayed for these molecules by Western blotting. Representative of 3 independent experiments. CF: cleaved form (activated form). (Bottom) Ratio of CIP2A to actin levels. Immunoblots were scanned by a UVP BioSpectrum AC image system and quantitated using VisionWork LS software. Columns, mean (n=3); bars, SD. (D) Bortezomib did not significantly alter the phosphorylation of Erk 1/2 in HL-60 cells. Cells were exposed to bortezomib at the indicated doses for 6 h. Cell lysates were prepared and assayed for p-Erk 1/2 and Erk by Western blotting. Representative of 3 independent experiments.