Fig. 6: PHI1 is a selective BRAF dimer inhibitor that displays positive co-operativity.

a Melanoma A375 and SKMEL239-C4 cell lines were treated with increasing. concentrations of PON or PHI1 for 1 h. Whole-cell lysate were assayed by western blot with the indicated antibodies to assess ERK-pathway inhibition. Representative blots from n = 3 independent experiments are shown. b Quantitation of p-ERK inhibition; normalized values (mean ± SEM, n = 3 independent experiments) of p-ERK levels obtained by densitometry with corresponding fitted curves (see “Methods” section). c SKMEL239-C4 cells without Encorafenib (−Enco) treatment and after Encorafenib (+Enco) treatment for 1 h, followed by exchange with fresh medium for another hour, were treated with increasing concentrations of c PHI1, e LY3009120, g AZ-628 and i TAK-632 for 1 h and cell lysates were assayed by western blot with the indicated antibodies to assess ERK-pathway inhibition. A representative blot from n = 3 (c, d) and n = 2 (e–j) independent experiments is shown. Normalized values and non-linear regression fits of p-ERK activity for different compounds in c, e, g, and i is shown respectively. Error bars represent mean ± SEM, n = 3 (d) and mean of two replicates from n = 2 independent experiments (f, h, j). k Table summarizing p-ERK inhibition results from all inhibitors in Encorafenib-free and Encorafenib-treated SKMEL239-C4 cells. Source data are provided as a Source Data file.