Figure 7

SAHA and panobinostat inhibit proliferation and induce apoptosis in erbB2-overexpressing breast cancer cells associated with the reduction of mRNA levels and protein expression of erbB2/erbB3. (a) MDA-MB-453 or BT474 cells were plated onto 96-well plates. After 24 h incubation, cells were grown in either control medium, or the same medium containing indicated concentrations of SAHA or panobinostat for another 72 h. The percentages of surviving cells relative to controls, defined as 100% survival, were determined by reduction of MTS. Bars, S.D. The data are representative of three independent experiments. (b) MDA-MB-453 cells were treated with SAHA (300 nmol/l) or panobinostat (8 nmol/l) for 24 h. BT474 cells were treated with SAHA (1.5 μmol/l) or panobinostat (20 nmol/l) for 24 h. All cells were collected and subjected to western blot analyses with specific antibody directed against erbB2, erbB3, PARP, or β-actin (top) and a specific apoptotic ELISA (bottom). Bars, S.D. The data are representative of three independent experiments. (c) MDA-MB-453 or BT474 cells were treated as described in (b) for 16 h. All cells were collected and subjected to total RNA extraction. First-strand cDNA was synthesized using a reverse transcription kit from Applied Biosystems. The mRNA levels of erbB2 and erbB3 were measured by qRT-PCR. Bars, S.D. The data are representative of three independent experiments. *P<0.03 versus control. (d), MDA-MB-453 (453) or BT474 cells were treated as described in (c). All cells were collected and subjected to total RNA extraction, inclusive of the small RNA fraction. The expression levels of miR-125a, miR-125b, and miR-205 were measured by qRT-PCR using TaqMan miRNA assays. All results were normalized with the internal control RNU6B. Bars, S.D. The data are representative of three independent experiments