Nutlin-3

Alias: Nutlin-3
Cat No.:V0040 Purity: ≥98%
Nutlin-3, a tetra-substituted imidazoline, is a novel potent and selective small-molecule antagonist/inhibitor of murine double minute 2 (MDM2) with potential antitumor activities.
Nutlin-3 Chemical Structure CAS No.: 890090-75-2
Product category: Mdm2
This product is for research use only, not for human use. We do not sell to patients.
Size Price Stock Qty
5mg
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25mg
50mg
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Other Forms of Nutlin-3:

  • Nutlin-3a
  • Nutlin (3)
Official Supplier of:
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Purity & Quality Control Documentation

Purity: ≥98%

Product Description

Nutlin-3, a tetra-substituted imidazoline, is a novel potent and selective small-molecule antagonist/inhibitor of murine double minute 2 (MDM2) with potential antitumor activities. In essence, Nutlin-3 is a protein-protein interaction (PPI) inhibitor. It works by occupying the p53 binding site in MDM2 and preventing MDM2 from binding to p53, which breaks down the autoregulator feedback loop and strengthens the p53 tumor suppressor network. Additionally, it interacts with murine double minute 4 (MDM4), a different element of the p35 tumor surveillance pathway. By attaching to MDM2 in the TP53-binding pocket, Nutlin-3 prevents MDM2-directed TP53 degradation. disrupts its interaction with p53, which causes p53 to become stabilized and active.

Biological Activity I Assay Protocols (From Reference)
Targets
MDM2 (IC50 = 180 nM)
ln Vitro
Nutlin-3 potently inhibits the MDM2-p53 interaction, leading to the activation of the p53 pathway. Only in cells with wild-type p53, such as HCT116, RKO, and SJSA-1, but not in the mutant p53 cell lines SW480 and MDA-MB-435, does nutlin-3 treatment induce the expression of MDM2 and p21 and exhibit strong antiproliferative activity with an IC50 of 1.5 μM. A 48-hour treatment with 10 μM of Nutlin-3 significantly increases the amount of caspase-dependent cell apoptosis in SJSA-1 cells by about ~45%. Although Nutlin-3 also inhibits the growth and viability of human skin (1043SK) and mouse embryo (NIH/3T3) with IC50 values of 2.2 M and 1.3 M, respectively, cells remain viable one week after treatment even at 10 M of Nutlin-3, in contrast to the SJSA-1 cells, whose viability was lost at a concentration of 3 M of Nutlin-3 treatment. [1] When compared to phosphorylated p53 produced by the genotoxic drugs doxorubicin and etoposide, nutlin-3 does not cause the phosphorylation of p53 on critical serine residues. It also shows no difference in their capacity to transactivate p53 target genes and sequence-specific DNA binding. These findings show that phosphorylation of p53 on critical serines is not necessary for transcriptional activation and apoptosis.[2] Nutlin-3 has a lower affinity for MDMX than it does for MDM2, but it still has the ability to block the MDMX-p53 interaction and activate the p53 pathway in retinoblastoma cells (Weri1) with an IC50 of 0.7 μM. [3] Apoptosis is induced and cell growth is inhibited in a dose-dependent manner in cells lacking wild-type p53 due to nutlin-3 at a concentration of 30 M, which also disrupts the interaction between endogenous p73 and HDM2 and increases the stability and proapoptotic activities of p73. [4]
ln Vivo
Nutlin-3 200 mg/kg orally twice daily for three weeks significantly slows the growth of SJAS-1 xenograft tumors by 90%, which is comparable to the effect of doxorubicin treatment, which inhibits tumor growth by 81%. [1]
Enzyme Assay
Competition assay is performed on a Biacore S51. A PentaHis antibody is immobilized on a Series S Sensor chip CM5 in order to capture p53 that has been His-tagged. The level of capture is 200 response units (one response unit equals one pg of protein per mm2). MDM2 protein is maintained at a constant concentration of 300 nM. In order to create a concentration series of Nutlin-3 in each MDM2 test sample, Nutlin-3 is first dissolved in DMSO at a concentration of 10 mM and then further diluted. The assay is carried out at 25 °C in running buffer (10 mM Hepes, 0.15 M NaCl, 2% DMSO). MDM2-p53 binding in the presence of Nutlin-3 is calculated as a percentage of binding in the absence of Nutlin-3, and IC50 is calculated.
Cell Assay
Nutlin-3 is present in cells for 8, 24, and 48 hours at various concentrations. Real-time PCR and western blotting are used to assess the protein levels of the p21 and MDM2 genes' transcriptional activity. Using the MTT assay, cell viability is evaluated. Deoxyuridine triphosphate nick end labeling (TUNEL) staining, which is carried out by terminal deoxynucleotidyl transferase and used in flow cytometry and fluorescence microscopy, is used to identify cell apoptosis.
Animal Protocol
Athymic female nude mice (Nu/Nu-nuBR) injected subcutaneously with SJSA-1 cells
200 mg/kg
Orally, twice a day
References

[1]. Science . 2004 Feb 6;303(5659):844-8.

[2]. J Biol Chem . 2004 Dec 17;279(51):53015-22.

[3]. Nature . 2006 Nov 2;444(7115):61-6.

[4]. Oncogene . 2008 Feb 7;27(7):997-1003.

[5]. Study on p53-MDM2 Interaction Inhibitors as a Novel Anticancer Agent. 2013, Hokkaido University.

These protocols are for reference only. InvivoChem does not independently validate these methods.
Physicochemical Properties
Molecular Formula
C30H30CL2N4O4
Molecular Weight
581.5
CAS #
890090-75-2
SMILES
CC(C)OC1=C(C=CC(=C1)OC)C2=NC(C(N2C(=O)N3CCNC(=O)C3)C4=CC=C(C=C4)Cl)C5=CC=C(C=C5)Cl
InChi Key
BDUHCSBCVGXTJM-UHFFFAOYSA-N
InChi Code
InChI=1S/C30H30Cl2N4O4/c1-18(2)40-25-16-23(39-3)12-13-24(25)29-34-27(19-4-8-21(31)9-5-19)28(20-6-10-22(32)11-7-20)36(29)30(38)35-15-14-33-26(37)17-35/h4-13,16,18,27-28H,14-15,17H2,1-3H3,(H,33,37)
Chemical Name
4-[4,5-bis(4-chlorophenyl)-2-(4-methoxy-2-propan-2-yloxyphenyl)-4,5-dihydroimidazole-1-carbonyl]piperazin-2-one
Synonyms
Nutlin-3
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 Data
Solubility (In Vitro)
DMSO: ~100 mg/mL (~171.9 mM)
Water: <1 mg/mL (slightly soluble or insoluble)
Ethanol: ~30 mg/mL (~51.6 mM)
Solubility (In Vivo)
O=C(N1C(C2=C(C=C(C=C2)OC)OC(C)C)=N[C@H](C3=CC=C(C=C3)Cl)[C@@H]1C4=CC=C(C=C4)Cl)N5CC(NCC5)=O.
 (Please use freshly prepared in vivo formulations for optimal results.)
Preparing Stock Solutions 1 mg 5 mg 10 mg
1 mM 1.7197 mL 8.5985 mL 17.1969 mL
5 mM 0.3439 mL 1.7197 mL 3.4394 mL
10 mM 0.1720 mL 0.8598 mL 1.7197 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.

Calculator

Molarity Calculator allows you to calculate the mass, volume, and/or concentration required for a solution, as detailed below:

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An example of molarity calculation using the molarity calculator is shown below:
What is the mass of compound required to make a 10 mM stock solution in 5 ml of DMSO given that the molecular weight of the compound is 350.26 g/mol?
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  • The answer of 17.513 mg appears in the Mass box. In a similar way, you may calculate the volume and concentration.

Dilution Calculator allows you to calculate how to dilute a stock solution of known concentrations. For example, you may Enter C1, C2 & V2 to calculate V1, as detailed below:

What volume of a given 10 mM stock solution is required to make 25 ml of a 25 μM solution?
Using the equation C1V1 = C2V2, where C1=10 mM, C2=25 μM, V2=25 ml and V1 is the unknown:
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  • The answer of 62.5 μL (0.1 ml) appears in the Volume (Start) box
g/mol

Molecular Weight Calculator allows you to calculate the molar mass and elemental composition of a compound, as detailed below:

Note: Chemical formula is case sensitive: C12H18N3O4  c12h18n3o4
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Definitions of molecular mass, molecular weight, molar mass and molar weight:
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In vivo Formulation Calculator (Clear solution)
Step 1: Enter information below (Recommended: An additional animal to make allowance for loss during the experiment)
Step 2: Enter in vivo formulation (This is only a calculator, not the exact formulation for a specific product. Please contact us first if there is no in vivo formulation in the solubility section.)
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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.

Biological Data
  • Nutlin-3

  • Nutlin-3

  • Nutlin-3

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