Figure S1.

Characterization of 885-A, an Analog of SB590885, Relates to Figure 1

(A) Sorafenib, a pan-RAF, multi-kinase class II inhibitor.

(B) PLX4720, a selective class I BRAF inhibitor.

(C) SB590885 and its analog 885-A, selective class I BRAF inhibitors.

(D) PD184352, a MEK inhibitor (CI1040).

(E) Inhibition of ^V600EBRAF by 885-A in vitro. Insect cell purified ^V600EBRAF kinase activity was measured at increasing concentrations of 885-A using a 96-well DELFIA-based assay system (Perkin Elmer, Amersham, UK). The assays were performed in the linear range of the assay and in duplicate using a concentration response of 11 points. IC₅₀ values were determined using GraphPad Prism software (GraphPad Software, San Diego, CA) and the reported IC₅₀ values are the mean of 3 independent assays.

(F) 885-A selectively inhibits the growth of BRAF mutant melanoma cells. The growth inhibitory activity of 885-A against a panel of cell lines (7 V600 BRAF mutant lines; 10 wild-type BRAF lines) is represented. The individual IC₅₀ values for each line are represented by the symbols, and the horizontal line represents the mean IC₅₀ for these two populations.

(G) SKMel24, SKMel28, D25 and WM266.4 cells were treated with DMSO (-), PD184352 (PD; 1μM), sorafenib (SF; 10 μM), 885-A (1μM) and PLX4720 (PLX; 0.3μM) for 4 hr. Cell extracts were western blotted for phospho-ERK (ppERK) and total ERK2 (loading control).