Figure 6. Specific inhibition of Akt, but not Src, significantly enhances lapatinib-mediated growth inhibition and long-term suppressive effects on colony formation.

A. SKBR3-Pool2 and BT474-HR20 cells were untreated or treated with lapatinib (0.1 μmol/L) or lapatinib combined with Akt/Src inhibitor (Lap+Akti/Lap+Srci) for 24 hr. Cells were collected and subjected to western blot analyses of P-Akt (S473), Akt, P-Src (Y416), Src. B. SKBR3-Pool2 and BT474-HR20 cells were plated onto 96-well plates. After 24 hr, the culture medium was replaced with 0.1 ml fresh medium containing 0.5% FBS or the same medium containing Akti (0.5 μmol/L), Srci (0.2 μmol/L), lapatinib (0.1 μmol/L), lapatinib+Akti, lapatinib+Srci for another 72 hr. The percentages of surviving cells from each cell line relative to controls, defined as 100% survival, were determined by reduction of MTS. Bars, SD. Data show a representative of three independent experiments. C. colony formation assays. 1 × 10³ SKBR3-Pool2 or BT474-HR20 cells were seeded onto 12-well plates. Cells were cultured with 0.1 ml fresh medium containing 0.5% FBS or the same medium containing lapatinib (0.1 μmol/L), lapatinib+Akti (0.5 μmol/L), lapatinib+Srci (0.2 μmol/L) for 2 weeks. The medium changed very 3 day. Bars, SD. Data show a representative of three independent experiments.