Table 2

Induction of Apoptosis by CP-654577

BT474 cells or MCF7 cells in growth medium were treated as indicated for 48 h and apoptosis was estimated by depolarization of the mitochondrial membrane (δΨm) as described in Materials and Methods. After incubation with CP-654577 for 48 h, BT474 and MCF7 cells were stained with JC-1. Healthy and apoptotic cells were visualized simultaneously by flow cytometry using the FL2 (propidium iodide) channel to detect JC-1 red aggregates and the FL1 (fluorescein) channel to detect JC-1 green monomers. Red−/green+ cells were scored as apoptotic and were easily distinguished from nonapoptotic (red+/green+) cells. The ionophore valinomycin (100 nM) was used as a positive control and caused loss of JC-1 red fluorescence in both BT474 and MCF7 cells. Determinations were performed in duplicate and the results are representative of two independent experiments.

CellTreatment% apoptotic cells
BT474Vehicle (DMSO)5
BT474CP-654577 100 nm8
BT474CP-654577 300 nm18
BT474CP-654577 1000 nm31
BT474Valinomycin 100 nm66
MCF7Vehicle (DMSO)9
MCF7CP-654577 1000 nm11
MCF7Valinomycin 100 nm91