Pharmacologic inhibition of GSK-3 induces apoptosis in CLL cells. (A) Malignant B cells from 3 patients with CLL were treated with 25 μmol/L concentrations of the 3 distinct GSK-3 inhibitors AR-A014418 (AR), SB216763 (SB), TDZD8 (TD), or diluent (DMSO); 24 hours after treatment, the cell pellet was collected and protein was obtained. Cell lysates were separated by SDS-PAGE, transferred to PVDF membrane, and immunoblotted with the indicated antibodies. (B) MEC1 CLL cells were treated with DMSO or AR-A014418 at indicated concentrations for 24 hours and protein expression was analyzed as described in (A). ARA10 = 10 μmol/L AR-A014418; ARA25 = 25 μmol/L AR-A014418. (C) MEC1 cells were treated for 24 hours with DMSO or AR-A014418 at indicated concentrations, then assayed for apoptosis using Annexin-V-FITC staining as determined by flow cytometry. Columns, mean; bars, standard deviation (SD). (D) Malignant B cells from 10 patients with CLL were treated for 48 hours with diluent (DMSO) or AR-A014418 at indicated concentrations, then assayed for apoptosis using Annexin-V-FITC staining as determined by flow cytometry (mean ± SD; n = 10). Columns, mean; bars, SD.