Figure 4.

Concentration-dependent effects of c-Met inhibition. (a) Treatment of melanoma cells with PHA665752 results in a concentration-dependent inhibition of c-Met phosphorylation. Cells were treated hours for four hours with PHA665752 at a concentration range of 0 to 1000 nM in the absence or presence of HGF 100 nM. (b) Similar concentrations inhibit phosphorylation of Akt, with the NRAS-mutated lines demonstrating the greatest sensitivity. Cells were treated with PHA665752 for 4 hours at a concentration 0, 50 or 100 nM in the presence or absence of HGF 100 nM. Cell lysates were subsequently examined for effects on Akt phosphorylation by western blotting. (c) Growth inhibitory effects of PHA665752 also appear to be influenced by genotype as demonstrated in the charted IC₅₀ values. Cells were plated in triplicate and treated with PHA665752 at a concentration range of 0 to 5 uM for 72 hours. Cell viability was determined by MTT assay and IC₅₀ values were subsequently calculated. The mean IC₅₀ for NRAS mutants was significantly lower than those of the other two genotypes, with p-values of < 0.001.