Figure 2.

CK2 inhibition by CX-4945 decreases NOTCH1 and MYC levels in human T-cell acute lymphoblastic leukemia (T-ALL) cells by destabilizing NOTCH1. (A) CX-4945 treatment in both human JURKAT and ALL-SIL T-ALL cells for 8 hours leads to decreased expression of cleaved-NOTCH1 (clv-NOTCH1) and phospho-AKT at Serine 129 site (P-AKT ser129). (B) Blocking proteasome-mediated degradation by MG132 for 8 hours rescues the decreased levels of clv-NOTCH1 in JURKAT and ALL-SIL T-ALL cells upon CX-4945 treatment. (C) Pulse-chase analysis of the half-life for clv-NOTCH1 reveals a less stable NOTCH1 upon CX-4945 (5 μM) treatment, compared with those treated with DMSO (3.96±0.45 hours vs. 1.59±1.54 hours in JURKAT cells and more than 8 hours vs. 5.75±1.77 hours in ALL-SIL cells; n=3 per group). CHX: cycloheximide. Data from one out of three biological repeats are shown. (D) qRT-PCR analysis revealing significantly decreased transcript levels of MYC in both JURKAT (P=0.017) and ALL-SIL (P<0.0001) T-ALL cells treated with CX-4945, compared with those in DMSO-treated cells. Data from one out of two biological repeats are shown. (E) CX-4945 treatment for 24 hours leads to decreased protein levels of both clv-NOTCH1 and MYC in JURKAT-BCL-2 over-expressing (o/e) T-ALL cells that are resistant to apoptosis. Clv-NOTCH1 protein amounts (relative to ACTIN) are shown in the bottom of panels (A) and (C). hr: hours.