Physicochemical Properties
| Molecular Formula | C33H42N6O3 |
| Molecular Weight | 570.72 |
| Exact Mass | 570.331 |
| CAS # | 2632259-92-6 |
| Related CAS # | ZW4864;2632259-93-7 |
| PubChem CID | 156180089 |
| Appearance | Light yellow to yellow solid powder |
| LogP | 3.4 |
| Hydrogen Bond Donor Count | 2 |
| Hydrogen Bond Acceptor Count | 6 |
| Rotatable Bond Count | 9 |
| Heavy Atom Count | 42 |
| Complexity | 914 |
| Defined Atom Stereocenter Count | 1 |
| SMILES | C1(N2C[C@H](C(=O)N(C3CC3)CC3C=CC(C4C=NNC=4)=CC=3)CCC2)=CC(OC(C)(C)C(=O)N2CCNCC2)=CC=C1 |
| InChi Key | DEGJSKLSOCKWSF-AREMUKBSSA-N |
| InChi Code | InChI=1S/C33H42N6O3/c1-33(2,32(41)37-17-14-34-15-18-37)42-30-7-3-6-29(19-30)38-16-4-5-26(23-38)31(40)39(28-12-13-28)22-24-8-10-25(11-9-24)27-20-35-36-21-27/h3,6-11,19-21,26,28,34H,4-5,12-18,22-23H2,1-2H3,(H,35,36)/t26-/m1/s1 |
| Chemical Name | (3R)-N-cyclopropyl-1-[3-(2-methyl-1-oxo-1-piperazin-1-ylpropan-2-yl)oxyphenyl]-N-[[4-(1H-pyrazol-4-yl)phenyl]methyl]piperidine-3-carboxamide |
| 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) |
Biological Activity
| Targets | IC50: 0.87 μM (β catenin/BCL9 PPI)[1]. Ki: 0.76 μM (β catenin/BCL9 PPI)[1] |
| ln Vitro | The free base ZW4864 (10~40 μM; 24 hours; SW480 and MBA-MD-231 cells) lowers the expression levels of the proteins Axin2 and cyclin D1 [1]. ZW4864 (10~40 μM; 72 hours; MDA-MB231, MCF10A, and MDA-MB-468 cells) (free base) preferentially induces hyperactive beta-catenin signaling-driven fast death in triple-negative breast cancer cells while sparing normal MCF10A breast epithelial cells [1]. ZW4864 (10~40 μM; 24 hours; SW480 and MBA-MD-231 cells) (free base) inhibited the transcription of β-catenin target genes in a concentration-dependent manner in both SW480 and Wnt 3a-activated MDA-MB-231 cells without changing the expression of the housekeeping gene HPRT [1]. Binding to β-catenin, ZW4864 (free base) specifically breaks down the protein-protein interaction (PPI) between β-catenin and B-cell lymphoma 9 (BCL9), leaving the β-catenin/E-cadherin PPI intact. ZW4864 is a free base that dose-dependently inhibits the activation of β-catenin signaling, downregulates the expression of oncogenic β-catenin target genes, and removes the invasiveness of cancer cells that rely on β-catenin. ZW4864 (free base) exhibits a dose-dependent inhibition of TOPFlash luciferase activity in HEK293 cells expressing β-catenin, with an IC50 of 11 μM. With an IC50 of 7.0 and 6.3 μM, respectively, ZW4864 (free base) also reduces TOPFlash luciferase activity in SW480- and Wnt 3a-activated MDA-MB-468 cells in a dose-dependent manner. ZW4864, a free base, inhibits β-catenin signaling transactivation specifically [1]. |
| ln Vivo | Oral bioavailability (F) of ZW4864 (20 mg/kg; po) (free base) is 83%, indicating favorable pharmacokinetic features[1]. In mice, ZW4864 (90 mg/kg; po) (free base) exhibits alterations in tumor growth[1]. In patient-derived xenograft mouse models, ZW4864 free base efficiently suppresses β-catenin target gene expression and exhibits good pharmacokinetic characteristics[1]. |
| Cell Assay |
Western Blot Analysis[1] Cell Types: SW480 and MBA-MD-231 cells Tested Concentrations: 10~40 μM Incubation Duration: 24 hrs (hours) Experimental Results: diminished the expression levels of Axin2 and cyclin D1 proteins. Apoptosis Analysis[1] Cell Types: MDA -MB231, MCF10A and MDA-MB-468 cells Tested Concentrations: 10~40 μM Incubation Duration: 72 hrs (hours) Experimental Results: Selectively triggered rapid apoptosis of triple-negative breast cancer cells with hyperactive β-catenin signaling while sparing normal mammary epithelial MCF10A cells. RT-PCR[1] Cell Types: SW480 and MBA-MD-231 cells Tested Concentrations: 10~40 μM Incubation Duration: 24 hrs (hours) Experimental Results: Suppressed the transcription of β-catenin target genes in a concentration-dependent manner without affecting the expression of HPRT , a house-keeper gene, in both SW480 and Wnt 3a-activated MDA-MB-231 cells. |
| Animal Protocol |
Animal/Disease Models: C57BL/6 mice[1] Doses: 20 mg/kg (pharmacokinetic/PK Analysis) Route of Administration: Po Experimental Results: demonstrated good pharmacokinetic/PK properties with an oral bioavailability (F) of 83%. Animal/Disease Models: Mice[1] Doses: 90 mg/kg Route of Administration: Po Experimental Results: demonstrated a variation in tumor growth in mice. |
| References |
[1]. Discovery of an Orally Bioavailable Small-Molecule Inhibitor for the β-Catenin/B-Cell Lymphoma 9 Protein-Protein Interaction. J Med Chem. 2021;64(16):12109-12131. |
Solubility Data
| Solubility (In Vitro) | DMSO: 100 mg/mL (175.22 mM) |
| 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 (e.g. IP/IV/IM/SC) Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline) *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). 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)] *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. Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline) Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline) Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline) Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil) Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL 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). Oral Formulation 3: Dissolved in PEG400 Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose Oral Formulation 6: Mixing with food powders Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.7522 mL | 8.7609 mL | 17.5217 mL | |
| 5 mM | 0.3504 mL | 1.7522 mL | 3.5043 mL | |
| 10 mM | 0.1752 mL | 0.8761 mL | 1.7522 mL |