Figure 1

Vemurafenib treatment induces selective ErbB3 phosporylation in melanoma cells. Simultaneous detection of the phosphorylation status of RTKs (n = 49) using a human phospho-RTK array in LOX IMVI (a) and MST-L (b) melanoma cells treated or not for 24 h with 0.3 μM vemurafenib. Membranes were incubated with cell lysates (100 μg) overnight according to the manufacturer' s protocol. The array detects the tyrosine-phosphorylated RTKs simultaneously in duplicate (1, pErbB1; 2, pErbB2; 3, pErbB3; 4, pErbB4). Duplicate dots in each corner are positive controls. Array data on images were analyzed using Photoshop Quantity One Program (Bio-Rad LaboratoriesGmbH). The phosphorylation of ErbB3 is strongly increased by vemurafenib treatment. LOX IMVI (c) and MST-L (d) cells were serum starved for 24 h, treated or not with different doses of vemurafenib for 6 h or 24 h. Western blot analysis performed using the indicated antibodies shows that vemurafenib induces a strong dose-dependent and time-dependent phosphorylation of ErbB3 and AKT. For densitometric analysis pErbB3/ErbB3, pERK/ERK and pAKT/ATK values are expressed as fold change with respect to the control unstimulated cells to which value = 1 was assigned. Results are reported as mean values ± standard deviation (SD) from three independent experiments.