pd318088  (MedChemExpress)

Journal: Nature Communications

Article Title: MEK inhibitors activate Wnt signalling and induce stem cell plasticity in colorectal cancer

doi: 10.1038/s41467-019-09898-0

Figure Lengend Snippet: Compound screens identify MEK inhibitors as activators of Wnt in CRC. a Schematic overview of the screening procedure. Colorectal cancer cell lines stably expressing TCF/Wnt luciferase reporters were seeded onto 384 well plates in two separate sets. 24 h after seeding, compound libraries were added and cells were treated for 24 h, followed by measurement of cell viability in one set and Wnt reporter activity in the other set of plates. Two biological replicates were performed for each screen. b A large compound screen identifies PD-0325901 as an activator of Wnt signalling. Waterfall plots showing the effect of a compound library containing 2399 drugs on Wnt reporter activity in HCT116, SW480 and DLD1 cell lines. The MEK inhibitor PD-0325901 is shown as a red dot. The GSK3 inhibitor BIO serves as positive control, whereas the porcupine inhibitor LGK974 and tankyrase inhibitor IWR-1 are negative controls for Wnt reporter activity (see also Supplementary Fig. ). The mean value of two independent experiments is presented. c – d A kinase-focused compound screen confirms Wnt activation by MEK inhibitors. Four colorectal cancer cell lines stably expressing TCF/Wnt reporters were treated with a compound library containing 274 kinase inhibitors. c Heatmap of Wnt reporter activities for all EGFR, RAF, MEK and GSK3 inhibitors. Wnt reporter activity was z-normalised for all drugs and high activity is presented in red and low activity in blue. d Dot plot showing average Wnt reporter activity levels resulting from treatment with different classes of Ras pathway and GSK3 inhibitors. Wnt reporter activity is significantly increased by MEK inhibitors compared to all other kinase inhibitors, and the increase is similar to GSK3 inhibition (two-sided Student’s t -test). Mean values of two independent experiments are presented. e Chemical structure of selected MEK1/2 inhibitors trametinib (PubChem CID 11707110), U0126 (Pubchem CID 3006531), selumetinib (PubChem CID 10127622) and PD-318088 (PubChem CID 10231331)

Article Snippet: Trametinib, ICG-001, PRI-724, dabrafenib, GDC-0994, PD318088, selumetinib were purchased from Selleckchem.

Techniques: Stable Transfection, Expressing, Luciferase, Activity Assay, Drug discovery, Positive Control, Activation Assay, Inhibition

Journal: Nature Communications

Article Title: MEK inhibitors activate Wnt signalling and induce stem cell plasticity in colorectal cancer

doi: 10.1038/s41467-019-09898-0

Figure Lengend Snippet: Characteristics of Wnt activation by MEK inhibitors. a – b Wnt reporter activity and target gene expression are increased by MEK inhibitors. SW480–7TFP were treated with different concentrations of the MEK inhibitors selumetinib, PD318088 or trametinib. TCF/Wnt-luciferase and CellTiterGlo signals were determined and normalised to DMSO controls ( a ) and expression of AXIN2 was measured by qPCR ( b ). c Concentration dependent activation of Wnt signalling by MEK inhibitor trametinib. SW480–7TFP cells were treated with different concentrations of trametinib for 24 h. TCF/Wnt reporter activity and CellTiterGlo signal were determined and normalised to DMSO controls. d Time-dependent activation of Wnt signalling by trametinib. SW480 cells were treated for different time periods with 1 µM of trametinib and expression of Wnt target gene AXIN2 was measured by qPCR. e Trametinib activates Wnt signalling in different colorectal cancer cell lines. HT29, HCT116 and SW403 cells were treated with indicated concentrations of trametinib for 24 h and AXIN2 transcript levels were determined by qPCR. The tankyrase inhibitor XAV939 serves as negative and the GSK3 inhibitor CHIR99021 as positive control. a – e Data from three independent experiments are presented as mean ± s.e.m. * p < 0.05, ** p < 0.01, two-sided Student’s t -test

Article Snippet: Trametinib, ICG-001, PRI-724, dabrafenib, GDC-0994, PD318088, selumetinib were purchased from Selleckchem.

Techniques: Activation Assay, Activity Assay, Targeted Gene Expression, Luciferase, Expressing, Concentration Assay, Positive Control