Name: BIX-01294 Trihydrochloride
Brand: Selleck Chemicals
SKU: S8006
Availability: InStock
Rating: 5 (49 reviews)

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Quality Control

Batch: Purity: 100.00%

100.00

Related Targets	HDAC JAK BET PKC PARP HIF PRMT EZH2 AMPK Histone Acetyltransferase
Other Histone Methyltransferase Inhibitors	Pinometostat (EPZ5676) 3-Deazaneplanocin A (DZNep) Hydrochloride UNC1999 EPZ015666 (GSK3235025) EPZ004777 MM-102 (HMTase Inhibitor IX) Chaetocin SGC 0946 EPZ005687 UNC0638

Cell Culture, Treatment & Working Concentration

Cell Lines	Assay Type	Concentration	Incubation Time	Activity Description	PMID
HeLa cells	Function assay			Inhibition of ERK5 phosphorylation in sorbitol-stimulated human HeLa cells, IC50=0.059 μM	23656407
HeLa	Function assay		4.5 hrs	Inhibition of Ebolavirus glycoprotein/matrix protein VP40 entry in human HeLa cells after 4.5 hrs beta-lactamase reporter assay, IC50 = 0.966 μM.	29624387
BL21 (DE3)-Gold	Function assay		20 mins	Inhibition of N-terminal hexahistidine-tagged human G9a (913 to 1193 residues) expressed in Escherichia coli BL21 (DE3)-Gold cells using histone H3 (1 to 15 residues) as substrate preincubated for 20 mins followed by substrate addition by mass spectrometr, IC50 = 1.9 μM.	27720557
HL60	Antiproliferative assay		48 hrs	Antiproliferative activity against human HL60 cells after 48 hrs by CCK8 assay, IC50 = 2.2 μM.	27393948
A549	Antiproliferative assay		48 hrs	Antiproliferative activity against human A549 cells after 48 hrs by CCK8 assay, IC50 = 2.8 μM.	27393948
HepG2	Antiproliferative assay		48 hrs	Antiproliferative activity against human HepG2 cells after 48 hrs by CCK8 assay, IC50 = 3.7 μM.	27393948
K562	Antiproliferative assay		48 hrs	Antiproliferative activity against human K562 cells after 48 hrs by CCK8 assay, IC50 = 3.9 μM.	27393948
MDA-MB-231	Function assay		2 days	Inhibition of G9a in human MDA-MB-231 cells assessed as reduction in H3K9me2 level after 2 days by immunofluorescence in-cell western analysis, IC50 = 4.563 μM.	29541353
K562	Function assay	0.5 to 2 uM	72 hrs	Inhibition of histone methyltransferase G9a in human K562 cells assessed as reduction in histone H3K9me2 levels at 0.5 to 2 uM after 72 hrs by Western blot analysis	27393948
TC32	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for TC32 cells	29435139
DAOY	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for DAOY cells	29435139
SJ-GBM2	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SJ-GBM2 cells	29435139
A673	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for A673 cells	29435139
SK-N-MC	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SK-N-MC cells	29435139
BT-37	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for BT-37 cells	29435139
NB-EBc1	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for NB-EBc1 cells	29435139
Saos-2	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Saos-2 cells	29435139
SK-N-SH	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SK-N-SH cells	29435139
NB1643	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for NB1643 cells	29435139
LAN-5	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for LAN-5 cells	29435139
BT-12	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for BT-12 cells	29435139
Rh18	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Rh18 cells	29435139
OHS-50	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for OHS-50 cells	29435139
RD	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for RD cells	29435139
MG 63 (6-TG R)	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for MG 63 (6-TG R) cells	29435139
wild type fibroblast cells	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for control Hh wild type fibroblast cells	29435139
Rh41	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Rh41 cells	29435139
NB1643	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for NB1643 cells	29435139
A673	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for A673 cells)	29435139
SK-N-MC	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for SK-N-MC cells	29435139
SJ-GBM2	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for SJ-GBM2 cells	29435139
BT-37	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for BT-37 cells	29435139
TC32	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for TC32 cells	29435139
Rh41	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for Rh41 cells	29435139
RD	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for RD cells	29435139
Rh30	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for Rh30 cells	29435139
OHS-50	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for OHS-50 cells	29435139
SK-N-SH	qHTS assay			qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Confirmatory screen for SK-N-SH cells	29435139
Click to View More Cell Line Experimental Data

Solubility

In vitro
Batch:

DMSO : 98 mg/mL (163.32 mM)
(Moisture-contaminated DMSO may reduce solubility. Use fresh, anhydrous DMSO.)

Water : 98 mg/mL

Ethanol : 8 mg/mL

Molarity Calculator

Mass	Concentration	Volume	Molecular Weight
Mass value Mass unit	Concentration value Concentration unit	Volume value Volume unit	Molecular weight (g/mol)

Dilution Calculator Molecular Weight Calculator

In vivo batch selection In vivo Batch:
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)

Dosage: mg/kg

Average weight of animals: g

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Working concentration: mg/ml;

Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such
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Chemical Information, Storage & Stability

Molecular Weight	600.02	Formula	C₂₈H₃₈N₆O₂.3HCl	Storage (From the date of receipt)	3 years-20°Cpowder
CAS No.	1392399-03-9	Download SDF		Storage of Stock Solutions	1 year-80°Cin solvent 1 month-20°Cin solvent
Synonyms	N/A			Smiles	CN1CCCN(CC1)C2=NC3=CC(=C(C=C3C(=N2)NC4CCN(CC4)CC5=CC=CC=C5)OC)OC.Cl.Cl.Cl

Mechanism of Action

Targets/IC₅₀/Ki	G9a (Cell-free assay) 2.7 μM
In vitro	BIX01294 is a selective inhibitor of G9a histone methyltransferase and does not affect SUV39H1(H320R) and PRMT1 within the tested concentration range. BIX-01294 specifically inhibited G9a (H3K9me2) and, to a lesser extent, the closely related GLP enzyme (primarily H3K9me3), with an IC50 of 1.7 μM for G9a and 38 μM for GLP. BIX-01294 inhibits G9a in an uncompetitive manner with SAM. BIX-01294 (4.1μM) reduces H3K9me2 Levels in Bulk Histone Preparations from wt ES cells, mouse embryonic fibroblasts and HeLa cells, but not in G9a deficient stem cells. BIX-01294 is a valuable inhibitor for the transient modulation of chromosomal H3K9me2. BIX-01294 Reduces H3K9me2 at Several G9a Target Genes including Bim1 and Serac1. BIX01294 could reactivate expression of HIV-1 from latently infected cells such as ACH-2 and OM10.1.
Kinase Assay	HMTase Assays
Kinase Assay	Dissociation Enhanced Lanthanide Fluoro-ImmunoAssays (DELFIA) are performed in white, opaque 384-well plates coated with Neutravidin. Test compounds are diluted to 12 μg/ml in 50mM Tris-HCl pH 8.5containing 4% DMSO and 10 μl isdispensed into the wells. Blank and control wells received only compound buffer. GST-G9a at 10 μg/ml and SAM at 40 μM are diluted in 50mM Tris HCl pH 8.5/10 mM DTT and added in a volume of 20 μL. Blank wells received Tris/DTT buffer only. The reactions are initiated by the addition of 800 nM H3(1-20)-cysbiotin substrate in 50 mM Tris pH 8.5 in a volume of 10 μl, and incubated at room temperature for 60 minutes. The plates are washed 3 times with 100 μl of Wash Buffer. Next, 50 μl of Fluoroimmunoassay (FI) Buffer containing 5ng α-2X-di-meth H3-K9 and 5ng goat anti-rabbit Eu chelate isadded to all wells of the plate, and the plate isincubated for an additional hour at room temperature. The plates are washed 3 times with 100 μL of Wash Buffer, and 50 μl of Enhancement Solution isadded to each well. Time resolved fluorescence ismeasured after 45 minutes on a Viewlux Microplate Imager imaging for 15 second.
References	[1] http://www.ncbi.nlm.nih.gov/pubmed/?term=17289593 [2] http://www.ncbi.nlm.nih.gov/pubmed/?term=20335163 [3] https://pubmed.ncbi.nlm.nih.gov/33147463/

Applications

Methods	Biomarkers	PMID
Western blot	Cyclin D1 / CDK2 / CDK4 / CDK 6 / Cyclin E G9a / H3K9me2 Atg7 / Atg5 / Atg5-Atg12 / Atg3 / Atg12 / LC3B	25198515
Immunofluorescence	Ran / H3K9me3	30565836
Growth inhibition assay	Cell viability	26013382