Home Epigenetics Histone Methyltransferase inhibitor - EZH1/2 inhibitor - Autophagy activator GSK343 For research use only.

GSK343

GSK343 is a potent and selective EZH2 inhibitor with IC50 of 4 nM in a cell-free assay, showing 60 fold selectivity against EZH1, and >1000 fold selectivity against other histone methyltransferases. GSK343 induces autophagy.

GSK343 Chemical Structure

GSK343 Chemical Structure

CAS No. 1346704-33-3

Purity & Quality Control

GSK343 Related Products

Signaling Pathway

Histone Methyltransferase Signaling Pathways

Histone Methyltransferase Signaling Pathway

Cell Data

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
2D10  Function Assay 0.25-2.0 μM 96 h reduces total cellular trimethylated H3K27 in a time- and dose-dependent manner 26041287
MDA-MB-231 Growth Inhibition Assay 0-20 μM 72 h shows cytotoxicity in a dose-dependent manner 25203626
MDA-MB-231 Function Assay 10 μM 72 h reduces the level of H3K27-me3 25203626
MDA-MB-231 Function Assay 10 μM 72 h induces LC3-II accumulation 25203626
MDA-MB-231 Function Assay 10 μM 24 h induces autophagy 25203626
HepG2 Growth Inhibition Assay 0-20 μM 72 h shows cytotoxicity in a dose-dependent manner 25203626
A549 Growth Inhibition Assay 0-20 μM 72 h shows cytotoxicity in a dose-dependent manner 25203626
HepG2 Function Assay 10 μM 24 h induces autophagy 25203626
A549 Function Assay 10 μM 24 h induces autophagy 25203626
OVCAR10 Growth Inhibition Assay 1 μM 0-12 d DMSO inhibits cell growth time dependently 23759589
UPN289 Growth Inhibition Assay 1 μM 0-12 d DMSO inhibits cell growth time dependently 23759589
SKOV3  Growth Inhibition Assay 1 μM 0-12 d DMSO inhibits cell growth time dependently 23759589
SKOV3  Apoptosis Assay 1 μM 4 d DMSO induces apoptosis cultured in 3D conditions 23759589
HCC1806 Function assay 72 hrs Inhibition of EZH2-mediated nuclear H3K27 methylation in human HCC1806 cells after 72 hrs by immunofluorescence analysis, IC50 = 0.174 μM. 24900432
LNCaP Function assay 6 days Inhibition of EZH2-mediated proliferation of human LNCaP cells after 6 days by chemiluminescence analysis, IC50 = 2.9 μM. 24900432
DU145 Function assay 6 days Inhibition of EZH2-mediated proliferation of human DU145 cells after 6 days by chemiluminescence analysis 24900432
SKBR3 Function assay 6 days Inhibition of EZH2-mediated proliferation of human SKBR3 cells after 6 days by chemiluminescence analysis 24900432
PC3 Function assay 6 days Inhibition of EZH2-mediated proliferation of human PC3 cells after 6 days by chemiluminescence analysis 24900432
ZR-75-1 Function assay 6 days Inhibition of EZH2-mediated proliferation of human ZR-75-1 cells after 6 days by chemiluminescence analysis 24900432
HCC180 Function assay 6 days Inhibition of EZH2-mediated proliferation of human HCC180 cells after 6 days by chemiluminescence analysis 24900432
Click to View More Cell Line Experimental Data

Biological Activity

Description GSK343 is a potent and selective EZH2 inhibitor with IC50 of 4 nM in a cell-free assay, showing 60 fold selectivity against EZH1, and >1000 fold selectivity against other histone methyltransferases. GSK343 induces autophagy.
Features A chemical probe for the SGC epigenetics initiative. Potential use in a variety of solid tumors.
Targets
EZH2 [1]
(Cell-free assay)		 EZH1 [1]
(Cell-free assay)
4 nM 240 nM
In vitro
In vitro

GSK343 inhibits trimethylation of H3K27 (H3K27me3) with IC50 of 174 nM in HCC1806 breast cancer cells. GSK343 potently inhibits cell proliferation in breast cancer cells and prostate cancer cells, and the prostate cancer cell line LNCaP is the most sensitive to GSK343, with IC50 of 2.9 μM. [1]

GSK343 significantly suppresses the growth of EOC cells cultured in 3D matrigel extracellular matrix (ECM), which mimics the tumor microenvironment in vivo. In addition, GSK343 also induces apoptosis of EOC cells in 3D and significantly inhibits the invasion of EOC cells. [2]
Kinase Assay In vitro biochemical assays against histone methyltransferases
Activity against EZH2 is assessed using 5 member PRC2 complex (Flag-EZH2, EED, SUZ12, AEBP2, RbAp48). The assay protocol may be summarized as follows: 10 mM stocks of compounds are prepared from solid in 100% DMSO. An 11 point serial dilution master plate is prepared in 384 well format (1:3 dilution, columns 6 and 18 were equal volume DMSO controls) and dispensed to assay ready plates using acoustic dispensing technology to create a 100 nL stamp of compound and DMSO controls. The assay additions consisted of equal volume additions of 10 nM EZH2 and the substrate solution (5 µg/mL HeLa nucleosomes and 0.25 µM [3H]-SAM) dispensed into assay plates using a multi-drop combi dispense. Reaction plates are incubated for 1 hr and quenched with an equal volume addition of 0.5 mg/mL PS-PEI Imaging Beads (RPNQ0098) containing 0.1 mM unlabeled SAM. The plates are sealed, dark adapted for 30 minutes, and a 5 minute endpoint luminescence image is acquired using a Viewlux imager. Plate statistics such as Z’ and signal to background as well as dose response curves are analyzed using Activity BaseXE. The in vitro biochemical activity of EZH1 is assessed as part of a 5 member PRC2 complex using a 384 well SPA assay identical to EZH2. Buffer components, reagent dispensing, compound plate preparation, quench conditions and data analysis are identical for EZH1 and EZH2 with final assay concentrations of 20 nM EZH1, 5 μM/mL HeLa nucleosomes and 0.25 μM [3H]-SAM. Further data analysis, pIC50 pivots and visualizations are enabled by TIBCO Spotfire. Compounds are profiled at Reaction Biology Corp. (Malvern, PA) to assess inhibition in their panel of histone methyltransferase assays. Methyltransferase activity is assessed using HotSpot technology, a miniaturized radioisotope-based filter binding assay. Inhibitors are dissolved in dimethyl sulfoxide (DMSO) and tested at concentrations up to 100 uM with a final DMSO concentration of 2%. Buffer containing the methyltrasferase at the listed concentration and its preferred substrate as shown in the accompanying table is preincubated with compound for 10 min. Reactions are initiated by the addition of 1 uM S-adenosyl-L-[methyl-3H]methionine (SAM), allowed to incubate for 60 min at 30C followed by transfer to P81 filter-paper and PBS wash before detection.
Cell Research Cell lines Breast cancer cell lines (HCC1806, Sk-Br-3, ZR-75-1), prostate cancer cell lines (DU145, PC3, LNCaP)
Concentrations ~50 μM
Incubation Time 6 days
Method

To account for varying doubling rates among cancer cell lines, the optimal cell seeding is determined empirically for all cell lines by examining their growth in a 384-well plate over 6 days with a wide range of seeding densities. Cells are then plated at the optimal seeding density and allowed to adhere overnight. Cells are treated in duplicate with a 20-point 2-fold dilution series of compound or 0.147% DMSO (vehicle control) and incubated for 6 days at 37C in 5% CO2. Cells are then lysed with 25 μl CellTiter-Glo per well and chemiluminescence is quantified with a TECAN Safire2 microplate reader. In addition, an untreated plate of cells is harvested at the time of compound addition (T0) to quantify the starting number of cells. CTG values after 6 days of treatment were expressed as a percent of the T0 value and plotted against compound concentration. Data are fit with a 4-parameter equation to generate a concentration response curve and the concentration of compound required to inhibit 50% of growth (gIC50) is determined
Experimental Result Images Methods Biomarkers Images PMID
Western blot H3K27me3 EZH2 / EED / SUZ12 N-cadherin / Vimentin / Snail / Slug / MMP2 / MMP9 29228694
Immunofluorescence N-cadherin / H3K27me3 / Vimentin 29228694
Growth inhibition assay Cell viability 26973856
In Vivo
In vivo

GSK343, a selective EZH2 inhibitor, inhibits phagocytosis.

Chemical Information & Solubility

Molecular Weight 541.69 Formula

C31H39N7O2
CAS No. 1346704-33-3 SDF Download GSK343 SDF
Smiles CCCC1=C(C(=O)NC(=C1)C)CNC(=O)C2=C3C=NN(C3=CC(=C2)C4=CC(=NC=C4)N5CCN(CC5)C)C(C)C
Storage (From the date of receipt)

In vitro
Batch:

Ethanol : 4 mg/mL

DMSO : 1 mg/mL ( (1.84 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Water : Insoluble


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Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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