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. Author manuscript; available in PMC: 2019 Aug 16.
Published in final edited form as: Cell Chem Biol. 2018 Jun 14;25(8):996–1005.e4. doi: 10.1016/j.chembiol.2018.05.008

Figure 3.

Figure 3

Cells treated with PAC-1 and vemurafenib or osimertinib have sustained inhibition of MAPK signaling. (A) A375 and SK-MEL-5 melanoma cells were treated with PAC-1 (5 μM), vemurafenib (10 μM), trametinib (30 nM), or the indicated combinations for 48 h. Inhibition of both ERK1/2 and MEK1/2 phosphorylation was only observed in cells treated with PAC-1 + vemurafenib. (B) Timecourse of phospho-MEK1/2 and phospho-ERK1/2 inhibition in A375 cells treated with DMSO, vemurafenib, vemurafenib + PAC-1 or vemurafenib + trametinib for 6, 24, or 48 hours. (C) After 48 hours of treatment with PAC-1 (5 μM) + osimertinib (4 nM), sustained inhibition of both MEK1/2 and ERK1/2 phosphorylation were observed in H1975 and PC-9 GR cells. Treatment with PAC-1 + gefitinib (4 nM) for a similar time period did not lead to similar observations. Sustained inhibition of MEK1/2 phosphorylation was also not observed in cells treated with trametinib (30 nM) + osimertinib. (D) Timecourse of phospho-MEK1/2 and phospho-ERK1/2 inhibition in PC-9 GR cells treated with DMSO, osimertinib, osimertinib + PAC-1 or osimertinib + trametinib for 6, 24, or 48 hours. See also Figure S4.