Home Epigenetics Histone Demethylase inhibitor - JMJD inhibitor GSK J4 HCl For research use only.

GSK J4 HCl

GSK J4 HCl is a cell permeable prodrug of GSK J1, which is the first selective inhibitor of the H3K27 histone demethylase JMJD3 and UTX with IC50 of 60 nM in a cell-free assay and inactive against a panel of demethylases of the JMJ family.

GSK J4 HCl Chemical Structure

GSK J4 HCl Chemical Structure

CAS No. 1797983-09-5

Purity & Quality Control

GSK J4 HCl Related Products

Signaling Pathway

Histone Demethylase Signaling Pathways

Histone Demethylase Signaling Pathway

Cell Data

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
CUTLL1 Growth inhibitory assay 2 μM DMSO affects cell growth 25132549
CUTLL1 Apoptosis assay 2 μM DMSO induces apoptosis 25132549
CUTLL1 Function assay 2 μM DMSO induces cell cycle arrest 25132549
CUTLL1 Kinase assay 6 μM DMSO leads to increased H3K27me3 25132549
SF7761 Kinase assay 6 μM DMSO increases K27 methylation 25401693
SF8628 Kinase assay 6 μM DMSO increases K28 methylation 25401693
H3.3 Kinase assay 6 μM DMSO increases K29 methylation 25401693
SF9012 Kinase assay 6 μM DMSO increases K30 methylation 25401693
SF9402 Kinase assay 6 μM DMSO increases K31 methylation 25401693
SF9427 Kinase assay 6 μM DMSO increases K32 methylation 25401693
human astrocytes Kinase assay 6 μM DMSO increases K33 methylation 25401693
SF7761 Growth inhibitory assay 6 μM DMSO inhibits K27M glioma cell growth 25401693
SF8628 Growth inhibitory assay 6 μM DMSO inhibits K28M glioma cell growth 25401693
H3.3 Growth inhibitory assay 6 μM DMSO inhibits K29M glioma cell growth 25401693
SF9012 Growth inhibitory assay 6 μM DMSO inhibits K30M glioma cell growth 25401693
SF9402 Growth inhibitory assay 6 μM DMSO inhibits K31M glioma cell growth 25401693
SF9427 Growth inhibitory assay 6 μM DMSO inhibits K32M glioma cell growth 25401693
human astrocytes Growth inhibitory assay 6 μM DMSO inhibits K33M glioma cell growth 25401693
TG neurons Function assay 50 μM DMSO inhibits HSV-1 reactivation from sensory neurons 25552720
Th17 Function assay 80 nM DMSO inhibits cell differentiation 25840993
β-cells Function assay 20 μM DMSO blunts IFNγ, Il-1β, and TNFα-induced chemokine gene expression 26505193
β-cells Function assay 20 μM DMSO induces β-cell dysfunction 26505193
ESCs Function assay 1.8 µM DMSO induces DNA damage along with activation of the DNA damage response 26759175
Raw 264.7 Function assay 0.8192 µM DMSO inhibits TNF-α production 26776360
MCF7 Function assay 1 to 10 uM 30 hrs Inhibition of KDM5A in human MCF7 cells assessed as effect on H3K4me3 methylation levels at 1 to 10 uM after 30 hrs by Western blot analysis 30392349
MCF7 Function assay 1 to 10 uM 30 hrs Inhibition of KDM5A in human MCF7 cells assessed as effect on H3K27me3 methylation levels at 1 to 10 uM after 30 hrs by Western blot analysis 30392349
RAW264.7 0.82 uM 24 hrs Inhibition of LPS-induced TNFalpha production in mouse RAW264.7 cells 0.82 uM after 24 hrs by ELISA 26776360
RAW264.7 24 hrs Inhibition of LPS-induced TNFalpha production in mouse RAW264.7 cells after 24 hrs by ELISA 26776360
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Biological Activity

Description GSK J4 HCl is a cell permeable prodrug of GSK J1, which is the first selective inhibitor of the H3K27 histone demethylase JMJD3 and UTX with IC50 of 60 nM in a cell-free assay and inactive against a panel of demethylases of the JMJ family.
Targets
JMJD3 [1]
(Cell-free assay)
60 nM
In vitro
In vitro

GSK J4 HCl is an ethyl ester derivative of the JMJD3 selective histone demethylase inhibitor GSK-J1 with an IC50 value greater than 50 μM in vitro. GSK J4 HCl is used to probe the consequences of demethylation of H3K27me3. In human primary macrophages, GSK-J4 inhibits the lipopolysaccharide-induced production of cytokines, including pro-inflammatory tumour necrosis factor (TNF). In addition, GSK-J4 prevents the lipopolysaccharide-induced loss of H3K27me3 associated with the TNF transcription start sites and blocked the recruitment of RNA polymerase II. [1]
Kinase Assay Histone Demethylase AlphaScreen
Inhibition of histone demethylases is assessed using the histone demethylase AlphaScreen assay (Amplified Luminescence Proximity Homogenous Assay). This assay uses a biotinylated peptide substrate and relies on detection of the product methyl mark using a specific antibody coupled to protein-A acceptor beads and a Steptavidin donor bead to capture the peptide. In brief, recombinant demethylase enzymes are incubated in the presence of Fe2+ in the form of Ferrous Ammonium Sulphate (FAS), -ketoglutarate (KG) and biotinylated peptide substrate. L-Ascorbic Acid is included to provide a reducing environment and prevent oxidation of Fe2+. After incubation with peptide substrate the presence of the product is detected using AlphaScreen technology. The demethylase AlphaScreen assays are performed in 384-well plate format using white proxiplates. All steps are carried out in assay buffer (50 mM HEPES pH 7.5, 0.1% (w/v) BSA and 0.01 % (v/v) Tween-20). FAS is dissolved fresh each day in 20 mM HCl to a concentration of 400 mM and diluted to 1.0 mM in deionized water. All other components are dissolved fresh each day in deionized water. For IC50 determinations 5 μL of assay buffer containing demethylase enzyme is transferred to wells of a 384-well proxiplate. Titrations of compound (0.1 μL) are transferred to each well and the enzymes allowed to pre-incubate for 15 minutes with compound (final concentration of DMSO is 1%). The enzyme reaction is initiated by addition of 5 μL of a substrate mix consisting of α-KG, FAS, L-Ascorbic Acid and biotinylated peptide substrate and the reaction incubated for the indicated time at room temperature. The enzyme reaction is stopped after the indicated time by addinton of 5 μL of EDTA (7.5 mM final concentration in assay buffer). Streptavidin Donor beads (0.08 mg/ml) and Protein-A conjugated acceptor beads (0.08 mg/ml) are pre-incubated for 1 hour with an antibody to the product methyl mark and the presence of biotin-H3-product is detected by addition of 5 μL of the preincubated AlphaScreen beads (final concentrations of 0.02 mg/ml with respect to acceptor and donor beads). Detection is allowed to proceed for 1 hour at room temperature and the assay plates read in a BMG Labtech Pherastar FS plate reader. Data are normalized to the no enzyme control and the IC50 determined from the nonlinear regression curve fit using GraphPad Prism 5.
Cell Research Cell lines Mouse podocytes
Concentrations 5 μM
Incubation Time 48 h
Method

Cells were serum starved for 4 hours, followed by treatment with EPZ-6438 (10 μM) or GSK-J4 (5 μM) for 48 hours.
In Vivo
In vivo

GSK-J4 hydrochloride is a potent dual inhibitor of H3K27me3/me2-demethylases JMJD3/KDM6B and UTX/KDM6A. It inhibits LPS-induced TNF-α production in human primary macrophages. GSK-J4 hydrochloride is a cell permeable prodrug of GSK-J1.
Animal Research Animal Models BALB/c mice
Dosages 10 mg/kg
Administration i.p.

Chemical Information & Solubility

Molecular Weight 453.96 Formula

C24H27N5O2.HCl
CAS No. 1797983-09-5 SDF Download GSK J4 HCl SDF
Smiles CCOC(=O)CCNC1=CC(=NC(=N1)C2=CC=CC=N2)N3CCC4=CC=CC=C4CC3.Cl
Storage (From the date of receipt)

In vitro
Batch:

DMSO : 91 mg/mL ( (200.45 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Ethanol : 91 mg/mL

Water : 10 mg/mL


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In vivo
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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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