| Size | Price | |
|---|---|---|
| 500mg | ||
| 1g | ||
| Other Sizes |
| Targets |
WEE1
|
|---|---|
| ln Vitro |
In this study, we discovered a covalent small-molecule recruiter EN523 for the K48-ubiquitin chain-specific DUB OTUB1. We demonstrated that this recruiter can be used incorporated into fully synthetic heterobifunctional DUBTACs by linking a DUB recruiter to protein targeting ligands to enable TPS of actively degraded target proteins in cells. We showed two successful examples of TPS with ΔF508-CFTR and WEE1. For ΔF508-CFTR, we also demonstrated that we not only heightened the levels of the mutant protein, but also improved cell surface chloride channel conductance of CFTR with our DUBTAC in combination with the potentiator ivacaftor, compared to lumacaftor and ivacaftor treatments. While we showed early validation of the DUBTAC platform here, there are many avenues for future exploration. These include further optimization of DUB recruiters against OTUB1 to improve their potency and proteome-wide selectivity, as well as the discovery of new recruiters against other candidate DUBs. For exploring optimization of CFTR DUBTACs, further improvement of the linker between lumacaftor and the DUB recruiter could improve potency and degree of CFTR stabilization. In addition, elucidating the mechanism, structural underpinnings, and kinetics in the formation of the ternary complex formed between CFTR and OTUB1 and understanding how CFTR is deubiquitinated by the DUBTAC will be important. Furthermore, better understanding of whether we are disrupting endogenous OTUB1 function would be important to understanding the mechanism and safety of DUBTACs[1].
WEE1 stabilization: Treatment of HEP3B hepatoma cancer cell lines with LEB-03-146, a DUBTAC linking AZD1775 to the OTUB1 recruiter EN523 through a PEG linker, showed significant stabilization of WEE1 protein levels comparable to those observed with bortezomib treatment. This stabilization was not observed with treatment of EN523 or AZD1775 alone. [1] |
| Cell Assay |
Western blotting: Used to assess WEE1 protein levels in HEP3B cells treated with LEB-03-146, EN523, AZD1775, or bortezomib. [1]
|
| References | |
| Additional Infomation |
Many diseases are driven by aberrant ubiquitination and degradation of proteins. Targeted protein stabilization (TPS) holds promise for the treatment of these diseases. This article introduces deubiquitinase-targeting chimeras (DUBTACs), heterobifunctional small molecules composed of a deubiquitinase recruiter linked to a protein-targeting ligand, designed to stabilize the levels of specific proteins that degrade in a ubiquitin-dependent manner. Using chemical proteomics, we discovered a covalent ligand, EN523, that targets the non-catalytic allosteric cysteine C23 in the K48-ubiquitin-specific deubiquitinase OTUB1. We demonstrate that DUBTACs, composed of the EN523 OTUB1 recruiter linked to lumacaftor (a drug for treating cystic fibrosis that binds to ΔF508-cystic fibrosis transmembrane conductance regulator (CFTR)), significantly stabilize ΔF508-CFTR protein levels, thereby improving chloride channel conductance in human cystic fibrosis bronchial epithelial cells. We also demonstrate that DUBTACs stabilize the tumor suppressor kinase WEE1 in hepatocellular carcinoma cells. Our study demonstrates the application of covalent chemical proteomics methods in developing novel therapeutic modalities based on induced proximity effects and introduces the DUBTAC platform for TPS. [1]
LEB-03-146 is a deubiquitinase-targeting chimera (DUBTAC) designed to stabilize intracellular WEE1 kinase levels by linking the WEE1 inhibitor AZD1775 to the OTUB1 recruitment protein EN523 via a PEG linker. This method utilizes the induced proximity effect of deubiquitinase (DUB) with the target protein to prevent its ubiquitination and subsequent proteasome degradation. [1] |
| Molecular Formula |
C46H57N11O8
|
|---|---|
| Molecular Weight |
892
|
| Exact Mass |
891.4391578
|
| CAS # |
2858812-91-4
|
| PubChem CID |
162624810
|
| Appearance |
Light yellow to yellow solid powder
|
| LogP |
2.7
|
| SMILES |
CC(C)(C1=NC(=CC=C1)N2C3=NC(=NC=C3C(=O)N2CC=C)NC4=CC=C(C=C4)N5CCN(CC5)CCOCCOCCNC(=O)CCC6=CC=C(O6)N7CCN(CC7=O)C(=O)C=C)O
|
| InChi Key |
ZYUOWVKGQBMNGW-UHFFFAOYSA-N
|
| InChi Code |
InChI=1S/C46H57N11O8/c1-5-19-56-44(61)36-31-48-45(51-43(36)57(56)38-9-7-8-37(50-38)46(3,4)62)49-33-10-12-34(13-11-33)53-22-20-52(21-23-53)26-28-64-30-29-63-27-18-47-39(58)16-14-35-15-17-42(65-35)55-25-24-54(32-41(55)60)40(59)6-2/h5-13,15,17,31,62H,1-2,14,16,18-30,32H2,3-4H3,(H,47,58)(H,48,49,51)
|
| Chemical Name |
N-[2-[2-[2-[4-[4-[[1-[6-(2-hydroxypropan-2-yl)pyridin-2-yl]-3-oxo-2-prop-2-enylpyrazolo[3,4-d]pyrimidin-6-yl]amino]phenyl]piperazin-1-yl]ethoxy]ethoxy]ethyl]-3-[5-(2-oxo-4-prop-2-enoylpiperazin-1-yl)furan-2-yl]propanamide
|
| Synonyms |
LEB-03-146; SCHEMBL27163559; LEB03-146; SCHEMBL-27163559;
|
| HS Tariff Code |
2934.99.9001
|
| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
|
| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
|
|---|---|
| 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 | 1.1211 mL | 5.6054 mL | 11.2108 mL | |
| 5 mM | 0.2242 mL | 1.1211 mL | 2.2422 mL | |
| 10 mM | 0.1121 mL | 0.5605 mL | 1.1211 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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