Mechanistic analysis of an ERK2-interacting compound that inhibits mutant-BRAF expressing melanoma cells by inducing oxidative stress
Conlon, Ivie L
Kane, Maureen A
JournalJournal of Pharmacology and Experimental Therapeutics
PublisherAmerican Society for Pharmacology and Experimental Therapy
MetadataShow full item record
AbstractConstitutively active extracellular signal-regulated kinase (ERK1/2) signaling promotes cancer cell proliferation and survival. We previously described a class of compounds containing a 1,1-dioxido-2,5-dihydrothiophen-3-yl 4-benzenesulfonate scaffold that targeted ERK2 substrate docking sites and selectively inhibited ERK1/2-dependent functions, including activator protein-1 (AP-1)-mediated transcription, and growth of cancer cells containing active ERK1/2 due to mutations in Ras G-proteins or BRAF kinase. The current study identified chemical features required for biological activity and global effects on gene and protein levels in A375 melanoma cells containing mutant BRAF (V600E). STD-NMR and mass spectrometry analyses revealed interactions between a lead compound (SF-3-030) and ERK2 including the formation of a covalent adduct on cysteine 252 located near the docking site for ERK, FXF (DEF) motif for substrate recruitment. Cells treated with SF-3-030 showed rapid changes in immediate early gene (IEG) levels including DEF motif containing ERK1/2 substrates in the Fos family. Analysis of transcriptome and proteome changes showed that the SF-3-030 effects overlapped with ATP-competitive or catalytic site inhibitors of MEK1/2 or ERK1/2. Like other ERK1/2 pathway inhibitors, SF-3-030 induced reactive oxygen species (ROS) and genes associated with oxidative stress, including nuclear factor erythroid 2-related factor 2 (NRF2). Whereas the addition of the ROS inhibitor N-acetyl cysteine reversed SF-3-030 induced ROS and inhibition of A375 cell proliferation, the addition of NRF2 inhibitors have little effect on cell proliferation. These studies provide mechanistic information on a novel chemical scaffold that selectively regulates ERK1/2-targeted transcription factors and inhibits the proliferation of A375 melanoma cells through a ROS-dependent mechanism. Significance Statement Constitutive activation of the extracellular signal-regulated kinase (ERK1/2) pathway drives the proliferation and survival of many cancer cell types. Given the diversity of cellular functions regulated by ERK1/2, the current studies have examined the mechanism of a novel chemical scaffold that targets ERK2 near a substrate binding site and inhibits select ERK functions. Using transcriptomic and proteomic analyses, we provide a mechanistic basis for how this class of compounds inhibits melanoma cells containing mutated BRAF and active ERK1/2.
Rights/Terms© 2020 The Authors. This is an open access article under the terms of the Creative Commons Attribution Non-Commercial License, which permits use, distribution, and reproduction in any medium, provided that the original work is properly cited and is not used for commercial purposes.
covalent drug binding
reactive oxygen species (ROS)
Identifier to cite or link to this itemhttp://hdl.handle.net/10713/14301
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