DI-1548
DI-1548 is a potent, selective, and irreversible covalent inhibitor of DCN1 (IC50 = 4.6 nM). DI-1548 potently and selectively inhibits cullin 3 neddylation at nanomolar concentrations, and exhibits no obvious effect on the neddylation of other cullin members (including cullin 1, 2, 4A, 4B, and 5). DI-1548 exhibits no cytotoxicity. DI-1548 covalently binds to DCN1, disrupting the DCN1-UBC12 interaction, causing the collapse of the neddylation complex, inactivating CRL3, leading to NRF2 accumulation and upregulation of its target genes, and ultimately protecting mice from liver damage. DI-1548 can be used in liver injury research.
For research use only. We do not sell to patients.
- CAS No.: 2247060-97-3
- Formula: C32H47N5O4S
- Molecular Weight:597.81
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Storage:
Please store the product under the recommended conditions in the Certificate of Analysis.
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
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Biological Activity
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Chemical Information
- Protocols
- Purity & Documentation
- References
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Help & FAQs
Biological Activity
Description
In Vitro
DI-1548 (1-1000 nM, 24 h) inhibits cullin 3 neddylation at concentrations as low as 1 nM and are approximately 1000 times more potent than DI-591 (HY-124602) in U2OS cells[1].
DI-1548 (0.3-1000 nM, 24 h) inhibits the neddylation of cullin 3 with over 1000-fold selectivity over other cullin members in various cell lines, including U2OS, MDA-MB-231, KYSE70, and HCT116[1].
DI-1548 (0-1000 nM, 72 h) shows no cytotoxicity in U2OS, MDA-MB-231, HCT116 and KYSE70 cells at concentrations up to 1000 nM[1].
DI-1548 (30 nM, 0-48 h) has fast kinetics in selective inhibition of the neddylation of cullin 3 over cullin 1[1].
DI-1548 (0.3-1000 nM, 1 h) selectively and potently stabilizes cellular DCN1 in a dose-dependent manner at nanomolar concentrations, with no observable effect on DCN3[1].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only. Further protocols information, click here.
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Cell Line:U2OS cells
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Concentration:1, 10, 100 and 1000 nM
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Incubation Time:24 h
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Result:Reduced N-Cul3 levels at 1 nM and was approximately 1000 times more potent than DI-591.
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Cell Line:U2OS, MDA-MB-231, HCT116 and KYSE70 cells
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Concentration:0.3, 1, 3, 10, 30, 100, and 1000 nM
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Incubation Time:24 h
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Result:Significantly reduced the level of N-CUL3 at concentrations as low as 0.3 nM, and achieved profound inhibition of the neddylation of cullin 3 at 1-3 nM.
Demonstrate over 300-fold greater potency than DI-591 across various cell lines.
Had no obvious effect on the neddylation of other cullin members that were examined, including cullin 1, 2, 4A, 4B, and 5 at concentrations up to 1000 nM.
Induced significant accumulation of NRF2 protein at concentrations as low as 0.3-1 nM, and was 100-1000 times more potent than DI-591.
Had no obvious effect on protein levels of KEAP1 and p62 in U2OS cells.
Had no effect on the levels of p21 and BIM proteins, substrates of CRL1, and CDT1, a substrate of CRL4A, at concentrations up to 1000 nM, consistent with its inability to inhibit neddylation of these cullin members.
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Cell Line:U2OS, MDA-MB-231, HCT116 and KYSE70 cells
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Concentration:0-1000 nM
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Incubation Time:72 h
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Result:Showed no cytotoxicity in four cancer cell lines at concentrations up to 1000 nM.
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Cell Line:U2OS cells
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Concentration:30 nM
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Incubation Time:5 and 30 min,6, 24, and 48 h
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Result:Inhibited the neddylation of cullin 3 within 5min but had no effect on the neddylation of cullin 1 with treatment time up to 48 h.
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Cell Line:U2OS cells
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Concentration:0.3, 1, 3, 10, 30, 100, and 1000 nM
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Incubation Time:1 h
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Result:Enhanced the thermal stability of DCN1 protein in a dose-dependent manner at concentrations as low as 0.3 nM.
Exhibited over 1000-fold greater potency than DI-591 in enhancing the thermal stability of cellular DCN1 protein.
Showed no discernible effect on the thermal stability of cellular DCN3 protein at concentrations up to 1000 nM.
In Vivo
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.
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Animal Model:Male C57BL/6 WT mice (~8 weeks) [1]
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Dosage:25 mg/kg
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Administration:i.p., once
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Result:Induced robust upregulation of NRF2 protein in the mouse liver.
Produced a sustained PD effect in the liver despite being cleared from systemic circulation within 3 hours.
Chemical Information
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CAS No. 2247060-97-3
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Molecular Weight 597.81
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Formula C32H47N5O4S
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SMILES
C=C(CN1CCOCC1)C(NC[C@H](C2CCCCC2)NC([C@@H](NC(CC)=O)CC(SC3=C4)=NC3=CC=C4C(C)C)=O)=O
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Shipping
Room temperature in continental US; may vary elsewhere.
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Storage
Please store the product under the recommended conditions in the Certificate of Analysis.
Protocols
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Cell Cytotoxicity Assay
Cytotoxicity assays are usually based on the assessment of cell membrane damage, which can also be indirectly detected by measuring cell viability. Detection methods include MTT assay, CKK-8 assay, LDH assay and ATP assay, etc.
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Mammalian live/dead viability and cytotoxicity staining
Live/dead viability and cytotoxicity staining assays are based on the simultaneous detection of intracellular esterase activity in metabolically active (viable) cells and membrane integrity loss in non-viable cells. In commonly used dual-staining approaches, membrane-permeant fluorogenic substrates are converted by intracellular esterases into fluorescent products in live cells, while impermeant DNA-binding dyes selectively enter cells with compromised plasma membranes and label nucleic acids in dead or dying cells, enabling discrimination between viable and non-viable populations by fluorescence microscopy or flow cytometry.
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Hepatotoxicity Study
This protocol evaluates hepatotoxicity using complementary in vivo mouse APAP acute liver injury and in vitro hepatocyte-based cytotoxicity readouts. In vivo APAP injury is assessed by serum ALT/AST, liver histology, hepatic glutathione, protein adducts, necrosis, inflammation, and regeneration-related endpoints. In vitro hepatotoxicity is assessed by loss of viability, leakage of ALT/AST/LDH, oxidative-stress markers, mitochondrial function, nuclear morphology, intracellular calcium, and high-content imaging endpoints.
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Cell Viability Determination by MTT Colorimetric Assay
The following protocol uses the MTT colorimetric assay as a classic literature-established method for assessing cell viability/metabolic activity in cultured mammalian cells. MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] is reduced by metabolically active cells to a colored formazan product; the amount of formazan is quantified spectrophotometrically and provides an indirect measure of metabolically active viable cells. Importantly, MTT reduction reflects cellular oxidoreductase/metabolic activity rather than an absolute direct count of living cells, so changes in cellular metabolism can alter the signal independently of cell number.
Purity & Documentation
References
Calculators
Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
DI-1548 Related Classifications
HY-156240 Related Classifications
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