Figure 4.

(A and B) HER2 down-regulation affects both viability and cell death in the CDC25A-overexpressing cell line. SKBR3 and BT474 cells were treated or not with AG825 (45 µM) for 96 hours. Cell viability changes were measured by MTT assay. Results were expressed as mean percentage of cell viability ± SE of six replicates from a representative experiment that was repeated three independent times. HER2 down-regulation resulted in a lower viability of the CDC25A-overexpressing cell line (SKBR3) compared to the CDC25A basal expressing cells (BT474) (A). Cell death was evaluated by immunoblot for cleaved PARP: PARP cleavage was greater in SKBR3 compared to BT474. β-Tubulin was used as a loading control (B). (C and D) CDC25A inhibition by PM-20 induces breast cancer cell death. SKBR3 and BT474 were treated or not with PM-20 (3 µM) for 96 hours. Cell viability changes were measured by MTT assay as performed for HER2 inhibition studies: A marked reduction in SKBR3 viability was observed (C); PM-20 significantly inhibited CDC25A phosphatase activity, as evinced by increased pCDK1/2Y15 levels, and associated to increased apoptotic response, as deduced by increased PARP cleavage. β-Tubulin was used as a loading control (D).