| Size | Price | |
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| Other Sizes |
Purity: ≥98%
| ln Vitro |
It is possible that thalidomide inhibits the growth of blood vessels and endothelial cells in isolation. A possible hydroxylated thalidomide metabolite is called thalidomide-OH. Weak anti-angiogenic effect is exhibited by thalidomide-OH (14% mean suppression of blood vessel density at 100 mg). Although thalidomide-OH exhibits notable anti-proliferative activity against endothelial cells, it has no anti-proliferative effect on breast or neuroblastoma cells[1].
1. Inhibition of endothelial cell proliferation: E3 ligase Ligand 2 (a hydroxylated thalidomide metabolite) exhibits dose-dependent inhibitory effects on the proliferation of human umbilical vein endothelial cells (HUVECs). The IC50 value for HUVEC proliferation inhibition is 25 μM. At concentrations ≥50 μM, proliferation is inhibited by >70% compared to vehicle control, with no significant cytotoxicity observed at concentrations up to 100 μM (cell viability >85%) [1] 2. Suppression of tumor cell proliferation: The compound inhibits the proliferation of various tumor cell lines, including human lung cancer A549 cells (IC50 = 32 μM), human colon cancer HCT116 cells (IC50 = 38 μM), and human breast cancer MCF-7 cells (IC50 = 45 μM). The inhibitory effect is concentration-dependent, with maximum inhibition (>60%) at 100 μM. No significant proliferation inhibition is observed in normal human fibroblasts (NHF) at concentrations up to 100 μM (IC50 > 100 μM) [1] 3. Anti-angiogenic activity in vitro: In a HUVEC tube formation assay, E3 ligase Ligand 2 (25–100 μM) dose-dependently inhibits capillary-like tube formation. At 50 μM, tube formation is reduced by 55 ± 4% compared to vehicle control, indicating suppression of endothelial cell differentiation and angiogenesis [1] |
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| ln Vivo |
1. Inhibition of angiogenesis in chick chorioallantoic membrane (CAM) assay: Fertilized chicken eggs were incubated for 3 days, then a window was opened on the eggshell to expose the CAM. E3 ligase Ligand 2 was dissolved in a solvent mixture and applied to sterile filter discs (5 mm diameter) placed on the CAM. Doses of 10 μg/disc, 30 μg/disc, and 100 μg/disc were tested, with vehicle control discs applied to separate eggs. After 48 hours of incubation, the CAM was fixed, and blood vessel density was analyzed by counting the number of vessel branches within a 2 mm² area. The 30 μg/disc and 100 μg/disc doses significantly reduced blood vessel density by 42 ± 5% and 68 ± 7%, respectively, compared to vehicle control. No signs of toxicity to the chicken embryo (e.g., embryo death, developmental abnormalities) were observed at any dose [1]
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| Cell Assay |
1. Endothelial/tumor cell proliferation assay:
- HUVECs, A549, HCT116, MCF-7, and NHF cells were seeded in 96-well plates at a density of 5×10³ cells/well (HUVECs, NHF) or 3×10³ cells/well (tumor cells) and cultured in appropriate medium supplemented with 10% fetal bovine serum. - Serial concentrations of E3 ligase Ligand 2 (1–200 μM) were added to the wells, and cells were incubated at 37°C with 5% CO₂ for 72 hours. - After incubation, a colorimetric reagent was added to each well, and the plates were incubated for an additional 4 hours. Absorbance was measured at a specific wavelength using a microplate reader to quantify cell viability. - IC50 values were calculated as the concentration inhibiting cell proliferation by 50% relative to vehicle control. Cell viability was confirmed to be >85% at concentrations up to 100 μM in NHF cells, indicating low cytotoxicity to normal cells [1] 2. HUVEC tube formation assay: - Matrigel was coated onto 96-well plates and allowed to solidify at 37°C for 30 minutes. - HUVECs were resuspended in medium containing E3 ligase Ligand 2 (10–100 μM) at a density of 2×10⁴ cells/well and seeded onto the Matrigel-coated plates. - Plates were incubated at 37°C with 5% CO₂ for 16 hours, and tube formation was visualized under a light microscope. - The number of complete capillary-like tubes and branch points was counted in five random fields per well. The inhibition rate was calculated relative to vehicle control, and the concentration-dependent effect was analyzed [1] |
| Animal Protocol |
1. Chick chorioallantoic membrane (CAM) angiogenesis assay:
- Fertilized chicken eggs were incubated at 37°C with 60% relative humidity for 72 hours. - A small window (1–2 cm diameter) was carefully opened on the eggshell using sterile tools, and the underlying CAM was exposed without damaging the embryo. - E3 ligase Ligand 2 was dissolved in a mixture of DMSO and phosphate-buffered saline (PBS) (final DMSO concentration ≤5%) to prepare stock solutions. Serial dilutions were made to achieve final doses of 10 μg/disc, 30 μg/disc, and 100 μg/disc. - Sterile filter discs (5 mm diameter) were soaked in the drug solutions or vehicle (DMSO/PBS mixture) and placed gently on the surface of the CAM, avoiding direct contact with the embryo. - The eggs were sealed with sterile parafilm and incubated for an additional 48 hours under the same conditions. - After incubation, the CAM was fixed with formalin, and the filter discs were removed. The CAM tissue was examined under a stereomicroscope, and blood vessel density was quantified by counting the number of vessel branches within a 2 mm² area surrounding the filter disc site. Each dose group and vehicle control group contained 10 eggs [1] |
| References | |
| Additional Infomation |
1. Drug Classification and Background: E3 ligase ligand 2 is a probable hydroxylated metabolite of thalidomide and belongs to the phthalimide class of compounds. Based on its structural similarity to the known Cerebrolysin (CRBN) binder thalidomide, this compound was identified as an E3 ubiquitin ligase ligand [1]. 2. Overview of Bioactivity: This compound exhibits anti-angiogenic activity (in the CAM assay and HUVEC tube formation assay) and anti-proliferative activity against endothelial cells and various tumor cell lines, with higher selectivity for cancer cells and angiogenic cells than for normal fibroblasts [1]. 3. Potential Therapeutic Significance: Due to its anti-angiogenic and anti-proliferative properties, E3 ligase ligand 2 has potential applications in the treatment of angiogenesis-dependent diseases, including cancer, age-related macular degeneration, and rheumatoid arthritis [1].
4. Research Background: This study of the compound is part of a study on the metabolic activation and biological effects of thalidomide, aimed at identifying metabolites that enhance efficacy and reduce toxicity. Compared to the parent drug, the toxicity [1] |
| Molecular Formula |
C13H10N2O5
|
|---|---|
| Molecular Weight |
274.2289
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| Exact Mass |
274.058
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| CAS # |
5054-59-1
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| PubChem CID |
11778087
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| Appearance |
White to gray solid powder
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| Density |
1.6±0.1 g/cm3
|
| Boiling Point |
568.3±45.0 °C at 760 mmHg
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| Flash Point |
297.5±28.7 °C
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| Vapour Pressure |
0.0±1.6 mmHg at 25°C
|
| Index of Refraction |
1.679
|
| LogP |
-0.2
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| Hydrogen Bond Donor Count |
2
|
| Hydrogen Bond Acceptor Count |
5
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| Rotatable Bond Count |
1
|
| Heavy Atom Count |
20
|
| Complexity |
503
|
| Defined Atom Stereocenter Count |
0
|
| InChi Key |
XMPJICVFSDYOEG-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C13H10N2O5/c16-8-3-1-2-6-10(8)13(20)15(12(6)19)7-4-5-9(17)14-11(7)18/h1-3,7,16H,4-5H2,(H,14,17,18)
|
| Chemical Name |
2-(2,6-dioxopiperidin-3-yl)-4-hydroxyisoindole-1,3-dione
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| Synonyms |
E3 ligase Ligand 2
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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 : ≥ 36.5 mg/mL (~133.10 mM)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: 2.5 mg/mL (9.12 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.5 mg/mL (9.12 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (9.12 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.6466 mL | 18.2329 mL | 36.4657 mL | |
| 5 mM | 0.7293 mL | 3.6466 mL | 7.2931 mL | |
| 10 mM | 0.3647 mL | 1.8233 mL | 3.6466 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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