Figure 5

ZAP-70 expression sensitizes Raji cells to gefitinib treatment. Raji cells transduced with GFP control vector or ZAP-70 vector (a) were treated with DMSO, and 20 or 30 μM gefitinib, or left untreated for 1 h and analyzed for Syk/ZAP-70 phosphorylation by western blot (b). (c) Co-immunoprecipitation of Syk or ZAP-70 with CD79a in Raji-ZAP-70 cells treated with DMSO or 20 μM gefitinib for 24 h and then lysed in 0.2% CHAPS buffer. (d) Lysates of untreated (-) and treated with DMSO (D) or gefitinib (G) Raji-ZAP-70 and Vector cells were analyzed by western blot for Mcl-1 degradation. (e) Raji-vector and Raji-ZAP-70 cells were treated with 20 μM gefitinib and stained with propidium iodide and analyzed by flow cytometry. Results from three independent experiments are shown (unpaired t-test). (f) Raji-vector and Raji-ZAP-70 cells were treated with DMSO, 20 μM gefitinib, 30 μM dasatinib, 20 μM ibrutinib, or 20 μM fludarabine for 24 h and then stained with Annexin V-APC and 7-AAD and analyzed by flow cytometry. Cell death of Raji-ZAP-70 cells normalized to Raji-vector cells. Results from three independent experiments are shown. *P<0.05