| Size | Price | Stock | Qty |
|---|---|---|---|
| 10mg |
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| Other Sizes |
Purity: ≥98%
| Targets |
G9a (IC50=4.7 μM), SUV39H1(H320R) (IC50=1.16 μM), PRMT1 , GLP, ESET, SET7/9 (inhibited at 15 μM). [1]
BIX01338 targets G9a (also known as EHMT2), a histone lysine methyltransferase that catalyzes the dimethylation of histone H3 at lysine 9 (H3K9me2), a mark associated with transcriptional repression. By inhibiting G9a, BIX01338 modulates epigenetic gene silencing. |
|---|---|
| ln Vitro |
In enzyme assays, BIX-01338 inhibited G9a with an IC50 of 4.7 μM and SUV39H1(H320R) with an IC50 of 1.16 μM. It started to affect methylation activity of all tested HMTases at around 5 μM and neutralized almost all enzymes at 15 μM, as determined by mass spectrometry. The compound displayed competitive inhibition with SAM in Eadie-Hofstee analysis. [1]
In cellular assays, treatment of mouse embryonic stem (ES) cells with 3.3 μM BIX-01338 for 2 days resulted in no detectable differences in methylation states at H3K9 and H3K27 compared to mock-treated cells, as measured by quantitative mass spectrometry. No alterations were observed for H3K36 and H3K79 methylation either. Although the compound is known to enter mammalian cells, it did not reduce H3K9me2 levels in vivo. [1] BIX01338 inhibits G9a histone methyltransferase activity with an IC50 of 4.7 µM. It was discovered alongside BIX01294 through high-throughput screening efforts to identify small molecule inhibitors of G9a. The compound represents a moderate-potency G9a inhibitor for epigenetic research. |
| ln Vivo |
In vivo activity data for BIX01338 are limited. As a moderate-potency G9a inhibitor (IC50 = 4.7 µM), its utility for in vivo applications may be limited compared to more potent G9a inhibitors. Further in vivo characterization is needed to evaluate its efficacy in animal models of diseases where G9a is implicated, such as cancer, neurological disorders, and metabolic diseases.
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| Enzyme Assay |
The high-throughput screen used a DELFIA (dissociation enhanced lanthanide fluoroimmunoassay) with recombinant GST-G9a (amino acids 685-1000) fusion protein, biotinylated histone H3 peptide, and 20 μM SAM. The H3K9me2 reaction product was bound to neutravidin-coated 384-well microtiter plates and detected with an H3K9me2 antibody followed by europium-labeled secondary antibody. Hits were scored as reduction of europium signal below 65% of control. [1]
For dose-response assays, BIX-01338 was tested on G9a, SUV39H1(H320R), and PRMT1 using DELFIA. IC50 values were calculated. [1] To determine mode of action, reaction kinetics were measured at varying SAM concentrations. Eadie-Hofstee plot analysis showed that BIX-01338 acts as a competitive inhibitor with respect to SAM. [1] For mass spectrometry-based validation, HMTase reactions were performed with nonbiotinylated H3(1-20)-cys peptide (for G9a, SUV39H1, GLP, SET7/9) or H4(1-20)-cys (for PRMT1). Reactions were stopped by dilution with 0.1% trifluoroacetic acid. Peptides were separated by C18 reverse-phase HPLC coupled to mass spectrometers, and peak areas of 3+ and 4+ peptide ions were quantified against synthetic peptide standards. [1] Cell-free histone methyltransferase assays for BIX01338 utilize purified recombinant G9a enzyme and histone H3 peptide or nucleosome substrates. The methylation reaction is performed in the presence of S-adenosylmethionine (SAM) as the methyl donor and increasing concentrations of the compound. Methylation of H3K9 is quantified using radiometric (3H-SAM) or fluorescence-based detection methods. IC50 values are determined from concentration-response curves. |
| Cell Assay |
Mouse ES cells (wild-type and G9a-deficient), mouse embryonic fibroblasts (MEFs), and human HeLa cells were treated with BIX-01338 at 3.3 μM for 2 days. Nuclear extracts were prepared, and histone H3 was isolated. After propionylation of lysines and carbamidomethylation of cysteines, tryptic digestion was performed. Peptides were separated by reverse-phase HPLC coupled to mass spectrometry. Quantification was performed by comparison of peak areas to external synthetic peptide standards containing the same posttranslational modifications. No changes in H3K9me2, H3K9me3, H3K9me1, H3K27me, H3K36me, or H3K79me were detected in BIX-01338-treated cells compared to mock-treated controls. [1]
Cells (e.g., cancer cell lines) are treated with BIX01338 at various concentrations (typically 1–50 µM) for specified durations. Global H3K9me2 levels are assessed by Western blot or immunofluorescence using H3K9me2-specific antibodies. Gene expression changes (particularly of G9a-repressed genes) are analyzed by qRT-PCR. Cell proliferation, differentiation, or apoptosis assays can be performed to evaluate functional consequences of G9a inhibition. |
| Animal Protocol |
In vivo animal studies for BIX01338 have not been extensively reported. For potential cancer or neurological applications, the compound could be administered via intraperitoneal or oral routes in rodent xenograft or disease models. Target engagement can be assessed by measuring H3K9me2 levels in tumor or tissue samples. Efficacy endpoints would depend on the specific disease model being studied.
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| ADME/Pharmacokinetics |
Pharmacokinetic properties of BIX01338 hydrate have not been comprehensively characterized. As a small molecule (MW 621.56, formula C32H26F3N3O7) with moderate molecular weight, its oral bioavailability and brain penetration would need to be evaluated. The compound is likely soluble in DMSO. Detailed PK parameters remain to be determined.
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| Toxicity/Toxicokinetics |
Toxicity data for BIX01338 are limited. As an epigenetic modulator targeting histone methylation, standard toxicological assessments would be required for development. G9a is involved in various cellular processes including development and differentiation, so on-target toxicity could be a concern. Off-target effects on other histone methyltransferases would need to be evaluated.
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| References | |
| Additional Infomation |
BIX-01338 belongs to a class of inhibitors containing a 2-(N-acyl)-aminobenzimidazole backbone, which is also found in a number of biologically active molecules. It is related to BIX-01294 but acts broadly. The compound was identified from the Boehringer Ingelheim chemical library. Unlike BIX-01294, which specifically inhibits G9a, BIX-01338 is non-selective and competes with SAM. [1]
BIX01338 (molecular formula C32H26F3N3O7, MW 621.56) is a histone lysine methyltransferase inhibitor discovered alongside the better-known G9a inhibitor BIX01294. It is available as a hydrate. The compound serves as a research tool for studying the role of G9a and H3K9 methylation in epigenetic regulation. It is intended for research use only. |
| Molecular Formula |
C32H26F3N3O7
|
|---|---|
| Molecular Weight |
621.559958934784
|
| Exact Mass |
621.172
|
| CAS # |
1228184-65-3
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| PubChem CID |
71311908
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| Appearance |
Light blue to blue solid powder
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| Hydrogen Bond Donor Count |
3
|
| Hydrogen Bond Acceptor Count |
11
|
| Rotatable Bond Count |
10
|
| Heavy Atom Count |
45
|
| Complexity |
990
|
| Defined Atom Stereocenter Count |
0
|
| SMILES |
FC(C1C=CC(=CC=1)C(NC1=NC2C=C(C(=O)O)C=CC=2N1CCC1C=CC(=CC=1)OC(C1C=CC(=CC=1)OC)=O)=O)(F)F.O
|
| InChi Key |
LLTPCGBKXOESTJ-UHFFFAOYSA-N
|
| InChi Code |
InChI=1S/C32H24F3N3O6.H2O/c1-43-24-13-6-21(7-14-24)30(42)44-25-11-2-19(3-12-25)16-17-38-27-15-8-22(29(40)41)18-26(27)36-31(38)37-28(39)20-4-9-23(10-5-20)32(33,34)35;/h2-15,18H,16-17H2,1H3,(H,40,41)(H,36,37,39);1H2
|
| Chemical Name |
1-[2-[4-(4-methoxybenzoyl)oxyphenyl]ethyl]-2-[[4-(trifluoromethyl)benzoyl]amino]benzimidazole-5-carboxylic acid;hydrate
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| Synonyms |
BIX-01338; BIX01338; BIX 01338
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ~200 mg/mL (~321.77 mM)
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|---|---|
| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.6089 mL | 8.0443 mL | 16.0886 mL | |
| 5 mM | 0.3218 mL | 1.6089 mL | 3.2177 mL | |
| 10 mM | 0.1609 mL | 0.8044 mL | 1.6089 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution 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 stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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