Figure 4.

The combination of ABT-199 with S63845 significantly inhibited sphere-forming capacity of the Melanoma Initiating Cells. Melanoma cells were subjected to the primary sphere assay (a–c). Spheres were treated with the indicated compounds either alone, or in combination, for 48 h, and the number of surviving spheres were counted and quantified (a). (b) Shows example images by phase contrast microscopy. (c) Dot plot of normalized primary sphere (expressed as percentage) for combination treatment in BRAF-V600E MUT and WT lines. Secondary sphere assay was conducted with surviving cells from each treatment conditions from the primary sphere assay (d–f). (d) Quantification of the number of secondary spheres; (e) the images of a representative secondary sphere, and (f) dot plot of the relative number of secondary spheres in the combination wells for BRAF-V600E MUT and WT lines. In all melanoma lines, the combination treatment significantly reduced the primary and secondary sphere formation compared with all other treatments (DMSO or single drug). For visual clarity, we have not marked the significance in panel (a) and (d). * indicates p < 0.05. Error bars represent +/− SEM. For clarity and readability of panels a and b, we listed p values in Table S5. Scale bar = 100 μm.