| Size | Price | Stock | Qty |
|---|---|---|---|
| 5mg |
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| 10mg |
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| 25mg |
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| 50mg |
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| 100mg | |||
| Other Sizes |
Purity: =100%
| Targets |
WWP2 ubiquitin ligase (E3) [1]
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|---|---|
| ln Vitro |
- NSC2805 (Compound 1) was identified as a hit compound from the NCI Diversity Set V high-throughput screen targeting WWP2 auto-ubiquitination. At a concentration of 10 μM and 0.1% DMSO, it significantly decreased WWP2 activity in single-shot assays and was selected for further testing based on residual activity thresholds. [1]
- In dose-dependent WWP2 inhibition assays, compound 1 demonstrated an IC50 value in the low micromolar range (exact IC50 value not specified in the text for compound 1; see Supporting Information Table S1). [1] - In counter assays against other ubiquitin ligases, compound 1 showed no significant difference in inhibition between WWP2, WWP1, and Nedd4, indicating that this compound is not selective towards a single HECT E3 ligase. All three proteins showed a significant reduction in activity in the presence of compound 1. [1] - Compound 1 was tested in E1 (Uba1) and E2 (UbcH7) counter assays to determine whether it affects upstream steps in the ubiquitination cascade. No significant decrease in E2 activity was observed in the His-E2 ubiquitination assay. In the E1 gel-based assay, no discernible impact on E1 ubiquitin conjugation was found in the presence of compound 1. [1] - In substrate-specific PTEN ubiquitination assays, compound 1 robustly inhibited WWP2-dependent PTEN ubiquitination as analyzed by western blotting. [1] - Ligand-based NMR spectroscopy (STD NMR and CPMG NMR) confirmed binding of compound 1 to the WWP2 HECT catalytic domain, showing positive binding results in both experiments. [1] NSC2805 exhibits inhibitory effect against WWP2, as evidenced by its IC50 of 0.38 μM[1]. The three proteins' (WWP2, WWP1, and Nedd4) dramatically lower activity in NSC2805 [1]. Potent inhibition of WWP2-dependent PTEN ubiquitination is possible with NSC2805 [1]. |
| Enzyme Assay |
- Auto-ubiquitination assay for WWP2 inhibition: Full-length WWP2 expressed as a GST fusion protein was attached onto glutathione-coated plates and incubated with recombinant E1, E2 enzymes, and Flag-tagged ubiquitin. Compounds were tested at 10 μM and 0.1% DMSO. WWP2 auto-ubiquitination was detected using HRP-conjugated anti-Flag antibody and TMB substrate. The specificity was confirmed using a catalytically inactive WWP2-FL C838A mutant. Z' values were over 0.5, indicating robust plate assays. Results were normalized using high (DMSO) and low (E3 only) controls to calculate percentage residual activity compared to the DMSO control. [1]
- Counter assay for E2 (UbcH7) activity: Bacterially expressed His-tagged UbcH7 was bound onto nickel-coated 96-well plates. Compounds were tested at the same initial concentrations as used in the GST-WWP2-FL screening. No significant decrease in activity was observed for all compounds in the His-E2 ubiquitination assay. [1] - Counter assay for E1 (Uba1) activity: Recombinant His-Uba1 and ubiquitin were incubated together to undergo E1-mediated transthiolation, forming higher molecular weight mono-Ub-E1 conjugates. Reactions were separated by SDS-PAGE and stained with Coomassie Blue. In the presence of compound 1, no discernible impact on E1 ubiquitin conjugation was found. [1] - Counter assays against Nedd4 and WWP1: GST-tagged Nedd4 and His-tagged WWP1 were immobilized onto glutathione-coated and nickel-coated plates, respectively. Auto-ubiquitination levels were determined using HRP-conjugated anti-Flag antibody and TMB substrate. Both Nedd4 and WWP1 produced significant increases in OD when in the presence of E1, E2, and ubiquitin. Compound 1 showed no significant difference in inhibition between WWP2, WWP1, and Nedd4, indicating non-selective inhibition across these HECT E3 ligases. [1] - Substrate-specific PTEN ubiquitination assay: Reactions contained E1, E2, WWP2, PTEN, ubiquitin, and ATP, and were analyzed by western blotting using anti-GST and anti-His antibodies. Compound 1 robustly inhibited WWP2-dependent PTEN ubiquitination. [1] |
| References |
[1]. Discovery of Small Molecule WWP2 Ubiquitin Ligase Inhibitors. Chemistry. 2018 Dec 3;24(67):17677-17680.
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| Additional Infomation |
- NSC2805 (Compound 1) was identified from the NCI Diversity Set V small molecule library (1,593 compounds) through a high-throughput screen targeting WWP2 auto-ubiquitination. Initial single-shot assays were run at 10 μM and 0.1% DMSO. A threshold of 70% (>2 SD from the DMSO control mean) was used to determine initial hits. Compound 1 was among 24 initial hits from the Diversity Set V screen. After triplicate testing, 17 of 24 hits demonstrated significant decrease in WWP2 activity, and compound 1 was further selected based on <20% residual activity thresholds and dose-dependent WWP2 inhibition. [1]
- WWP2 is an E3 ubiquitin ligase associated with tumour outgrowth and spread. It causes ubiquitin-dependent degradation of specific tumour suppressor proteins commonly lost in many types of cancer, including Smad transcription factors, PTEN, and Oct4. [1] |
| Molecular Formula |
C14H14O4
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|---|---|
| Molecular Weight |
246.26
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| Exact Mass |
246.089
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| Elemental Analysis |
C, 68.28; H, 5.73; O, 25.99
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| CAS # |
4371-34-0
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| Related CAS # |
4371-34-0
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| PubChem CID |
220284
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| Appearance |
Pale purple to gray solid powder
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| Density |
1.36g/cm3
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| Boiling Point |
501.6ºC at 760 mmHg
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| Flash Point |
248.2ºC
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| Index of Refraction |
1.675
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| LogP |
2.792
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| Hydrogen Bond Donor Count |
4
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
1
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| Heavy Atom Count |
18
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| Complexity |
255
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| Defined Atom Stereocenter Count |
0
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| SMILES |
OC1=CC(C)=C(O)C=C1C2=CC(O)=C(C)C=C2O
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| InChi Key |
DSVRCBOSZSZMRX-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C14H14O4/c1-7-3-13(17)9(5-11(7)15)10-6-12(16)8(2)4-14(10)18/h3-6,15-18H,1-2H3
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| Chemical Name |
2-(2,5-dihydroxy-4-methylphenyl)-5-methylbenzene-1,4-diol
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| Synonyms |
NSC-2805; NSC 2805; 4371-34-0; 4,4'-dimethyl[1,1'-biphenyl]-2,2',5,5'-tetrol; NSC-2,805; 2-(2,5-dihydroxy-4-methylphenyl)-5-methylbenzene-1,4-diol; NSC2805
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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. |
| 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 : ~10 mg/mL (~40.61 mM)
H2O : < 0.1 mg/mL |
|---|---|
| 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 | 4.0607 mL | 20.3037 mL | 40.6075 mL | |
| 5 mM | 0.8121 mL | 4.0607 mL | 8.1215 mL | |
| 10 mM | 0.4061 mL | 2.0304 mL | 4.0607 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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