Home Epigenetics Histone Methyltransferase inhibitor - G9a/GLP inhibitor - Antiviral chemical UNC0638 For research use only.

UNC0638

UNC0638 is a potent, selective and cell-penetrant chemical probe for G9a and GLP histone methyltransferase with IC50 of <15 nM and 19 nM, respectively, shows selectivity over a wide range of epigenetic and non-epigenetic targets. UNC0638 has anti-viral activities.

UNC0638 Chemical Structure

UNC0638 Chemical Structure

CAS No. 1255580-76-7

Purity & Quality Control

UNC0638 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
22Rv1 cells Function assay Inhibition of G9a in human 22Rv1 cells assessed as reduction of H3K9me2 after 48 hrs by In-Cell Western assay, IC50=0.048 μM 21780790
PC3 cells Function assay Inhibition of G9a in human PC3 cells assessed as reduction of H3K9me2 after 48 hrs by In-Cell Western assay, IC50=0.059 μM 21780790
MCF7 cells Function assay Inhibition of G9a in human MCF7 cells assessed as reduction of H3K9me2 after 48 hrs by In-Cell Western assay, IC50=0.07 μM 21780790
MDA-MB-231 cells Function assay 48 h Inhibition of G9a in human MDA-MB-231 cells after 48 hrs by clonogenic assay, IC50=0.081 μM 22975593
IMR90 cells Function assay Inhibition of G9a in human IMR90 cells assessed as reduction of H3K9me2 after 48 hrs by In-Cell Western assay, IC50=0.12 μM 21780790
HCT116 cells Function assay Inhibition of G9a in human HCT116 cells assessed as reduction of H3K9me2 after 48 hrs by In-Cell Western assay, IC50=0.21 μM 21780790
H1299 cells Function assay 0.25 uM 24 h Inhibition of G9a expressed in H1299 cells assessed as inhibition of dimethylation of p53 at lys 373 at 0.25 uM after 24 hrs by Western blot analysis 21780790
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Biological Activity

Description UNC0638 is a potent, selective and cell-penetrant chemical probe for G9a and GLP histone methyltransferase with IC50 of <15 nM and 19 nM, respectively, shows selectivity over a wide range of epigenetic and non-epigenetic targets. UNC0638 has anti-viral activities.
Targets
G9a [1]
(Cell-free assay)		 GLP [1]
(Cell-free assay)
<15 nM 19 nM
In vitro
In vitro UNC0638 is a potent, selective and cell-penetrant chemical probe for G9a and GLP, with a toxicity/function ratio of >100, compared to <6 for BIX01294. UNC0638 is a selectivite inhibitor of G9a and GLP over a wide range of epigenetic and non-epigenetic targets. UNC0638 is more than 10,000-fold selective against SET7/9 (a H3K4 HMTase), SET8 (a H4K20 HMTase), PRMT3, and SUV39H2. In MDA-MB-231 cells, UNC0638 (48 h exposure) reduces H3K9me2 levels in a concentration-dependent manner with an IC50 of 81 nM. UNC0638 treatment of a variety of cell lines results in lower global H3K9me2 levels, equivalent to levels observed for small hairpin RNA knockdown of G9a and GLP with the functional potency of UNC0638 being well separated from its toxicity. UNC0638 markedly reduces the clonogenicity of MCF7 cells, reduces the abundance of H3K9me2 marks at promoters of known G9a-regulated endogenous genes and disproportionately affected several genomic loci encoding microRNAs. In mouse embryonic stem cells, UNC0638 reactivates G9a-silenced genes and a retroviral reporter gene in a concentration-dependent manner without promoting differentiation. [1]
Kinase Assay SAHH-coupled assays
This assay utilizes SAHH to hydrolyze the methyltransfer product S-adenosylhomocysteine to homocysteine and adenosine in the presence of adenosine deaminase which converts adenosine to inosine. The homocysteine concentration is then determined through conjugation of its free sulfhydryl moiety to a thiol-sensitive fluorophore, ThioGlo. For IC50 determinations, assay mixtures are prepared in 25 mM Potassium Phosphate buffer pH 7.5, 1 mM EDTA, 2 mM MgCl2, 0.01% Triton X 100 with 5 μM SAHH , 0.3 U/mL of adenosine deaminase, 25 μM SAM, and 15 μM ThioGlo. G9a, EHMT1, SETD7, SETD8, PRMT3 and SUV39H2 are assayed at 25 nM, 100 nM, 200 nM, 250 nM, 1 μM and 100 nM, respectively. UNC0638 is added at concentrations ranging from 4 nM to 16 μM. After 2 min incubation, reactions are initiated by addition of the histone peptides: 10 μM H3(1-25) for G9a, 20 μM H3(1-25) for EHMT1, 100 μM H3(1-25) for SETD7, 500 μM H4(1-24) for SETD8, 10 μM H4(1-24) for PRMT3 and 200 μM H3K9Me1 (1-15) for SUV39H2. The methylation reaction is followed by monitoring the increase in fluorescence using Biotek Synergy2 plate reader with 360/40 nm excitation filter and 528/20 nm emission filter for 20 min in 384 well-plate format. Activity values are corrected by subtracting background caused by the peptide or the protein. IC50 values are calculated using Sigmaplot. Standard deviations are calculated from two independent experiments.
Cell Research Cell lines MCF7, U2OS, H1299 cell lines
Concentrations 3 μM
Incubation Time 65 h
Method

Approximately 5×105 cells were plated onto 75 cm2 flasks. MCF7 and U2OS cells were grown in Minimal Essential Media (MEM) without phenol red supplemented with Earl's salts, 10% FBS, 1 mM Pyruvate, 2 mM Glutamine, and 1×non-essential amino acids. H1299 cells were grown in Dulbecco's Modified Eagle's Medium (DMEM) with phenol red supplemented with 10% FBS and 1×Anti-Anti. At the time of cell plating, media was supplemented with 3 μM UNC638A or the equivalent DMSO vehicle. The media with 3 μM UNC638A but without any cells was also used as a control. All flasks were incubated at 37℃/5% CO2. After 65 hours, media was collected from the cells and centrifuged to remove any cell debris. The media (10 to 15 mL) from each of study groups was mixed with HPLC grade chloroform (7 to 12 mL). After the mixture was gently shaken, it was allowed to sit until the organic layer and the aqueous layer separated. The organic layer was collected, dried over anhydrous Na2SO4, and concentrated in vacuo. The resulting residue was dissolved in 100 μL of methanol and the solution was analyzed using an Agilent 6110 LC/MS Series system with UV detector set to at 254 nm. Samples were injected (10 μL) onto a Phenomenex Kinetex 2.1 x 100 nm, 2.6 μM, C18 column at room temperature and eluted with 72% B (MeCN) with A being H2O + 0.1% formic acid. The flow rate was 0.3 mL/min. No degradation products of UNC0638 were observed for any of treatment groups.
In Vivo
In vivo In mouse drug metabolism and pharmacokinetic studies, UNC0638 had high clearance, short half-life, high volume distribution and low exposure after intravenous, oral or intraperitoneal administration. Thus, although UNC0638 is probably not suitable for in vivo animal studies owing to low exposure levels, its high stability under cellular assay conditions, in combination with high potency and selectivity, makes UNC0638 an ideal chemical tool for cell-based studies[1].
Animal Research Animal Models Swiss albino mice
Dosages IV, 1 mg/kg; PO, 3 mg/kg; IP, 2.5 mg/kg
Administration i.v./i.p./oral

Chemical Information & Solubility

Molecular Weight 509.73 Formula

C30H47N5O2
CAS No. 1255580-76-7 SDF Download UNC0638 SDF
Smiles CC(C)N1CCC(CC1)NC2=NC(=NC3=CC(=C(C=C32)OC)OCCCN4CCCC4)C5CCCCC5
Storage (From the date of receipt)

In vitro
Batch:

DMSO : 100 mg/mL ( (196.18 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Water : Insoluble

Ethanol : Insoluble


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In vivo
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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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