AEB071 targets β-catenin and its downstream transcriptional targets in B-CLL. (A) CD19⁺ cells from CLL patients (N = 3) were treated with AEB071 (2 or 5 μM) and stimulated with PMA (250 ng/mL) for 24 hours. GSK3-β phosphorylation at Ser9 and protein expression of β-Catenin, c-Myc, and Cyclin D1 was assessed in cytoplasmic (CE) and nuclear extracts (NE) by immunoblot. (B-E) CD19⁺ cells from CLL patients (N = 7) were treated with AEB071 (2 or 5 μM) and stimulated with PMA (250 ng/mL) for 24 hours, wherein surface expression of CD44 (B) was evaluated in viable cells by flow cytometry and transcript levels of Cyclin D1 (C), c-Myc (D), and CD44 (E) was estimated by real-time PCR. Dark lines represent averages. (F) CD19⁺ cells from CLL patients (N = 3) were treated with increasing concentrations of AEB071 and stimulated with PMA (250 ng/mL) for 24 hours. GSK3-β phosphorylation at Ser9 and protein expression of β-Catenin and c-Myc was assessed in whole-cell lysates by immunoblot. Results from 2 patients are presented. (G) CD19⁺ cells from CLL patients (N = 10) were incubated with increasing concentrations of AEB071 for 24 hours. Viability was determined by MTS assay and calculated relative to untreated control. Dark lines represent averages.