Figure 5

NF-κB functions as a critical regulator for cell survival in response to lapatinib treatment. (A,B) SkBr3 (A), MDA-MB-231 (B), and their lapatinib-treated derivatives were infected with lentivirus of Luc or p65 shRNA in the presence or absence of 1 μM lapatinib for three days. Cell viability and p65 protein expression were determined by MTT and Western blot analyses, respectively. (C) MDA-MB-231 and 231/Lap cells were treated with 10 μM IKKα inhibitor in the presence or absence of 1 μM lapatinib for 24 hours. The cell viability was determined by MTT assay. (D-I) Parental and lapatinib-selected cells of SkBr3 (D-F) and MDA-MB-231 (G-I) cells were treated with various doses of proteasome inhibitors, including MG-132 (D and G), lactacystin (E and H) and proteasome inhibitor I (F and I) in the presence or absence of lapatinib for 24 hours and then subjected to MTT assay. (J) MDA-MB-231 cells and their lapatinib-treated clones were treated with or without the indicated concentrations of bortezomib for 48 hours. Two apoptotic markers, PARP and caspase 3 cleavages, were then examined by Western blot analysis with anti-PARP and anti-caspase 3 specific antibodies. *, P <0.05; **, P <0.01; ***, P <0.001. IKK, IKB kinase; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PARP, poly ADP ribose polymerase.