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Purity: ≥98%
DIM-C-pPhOCH3 is an agonist Nur77 (Nerve growth factor-induced Bα, abbreviated as NGFI-Bα or Nur77). Nur77 is an orphan nuclear receptor with no known endogenous ligands and was reported to be overexpressed in colon tumors compared with normal colon tissue. In RKO colon cancer cells, DIM-C-phOCH3 was able to reduce survival and trigger apoptosis; this was accompanied by an increase in the protein known as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). A small Nur77 inhibitory RNA (iNur77) greatly reduced the induction of TRAIL and apoptosis by DIM-C-pPhOCH3, but RNA interference studies clearly demonstrated that DIM-C-pPhOCH3 also induced Nur77-independent apoptosis. Microarray analysis of DIM-C-phOCH3-induced gene expression revealed a number of proapoptotic genes. Reverse transcription-PCR analysis conducted in the presence or absence of iNur77 revealed that induction of gene 1 for programmed cell death was dependent on Nur77, while induction of cystathionase and activating transcription factor 3 was independent of Nur77. Furthermore, in athymic nude mice with RKO cell xenografts, DIM-C-pPhOCH3 (25 mg/kg/d) inhibited tumor growth. These findings demonstrate that a novel class of anti-colon cancer medications, known as DIM-C-phOCH3, functions via both receptor-dependent and receptor-independent mechanisms.
| Targets |
Nur77
Nur77 (Nuclear Receptor Subfamily 4, Group A, Member 1, also known as NGFI-Balpha), an orphan nuclear receptor [1] |
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| ln Vitro |
DIM-C-pPhOCH3 causes tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) protein to be produced in conjunction with a decrease in survival and an increase in apoptosis in RKO colon cancer cells. Apoptosis that is Nur77-independent is also induced by DIM-C-phOCH3. After treatment, DIM-C-pHOCH3 (10 μM) inhibits cell growth for 24, 48, or 72 hours. The maximum inhibitory response is seen after 72 hours, at which point there is a significant amount of dead cells and cell detachment. Following a 72-hour period, DIM-C-phOCH3 was shown to exhibit growth-inhibitory effects along with a number of apoptosis markers, such as PARP cleavage and cleavage of caspase-3, caspase-9, and caspase-8. Following DIM-C-pHOCH3 treatment of RKO cells for 48 hours, PARP cleavage is also seen[1].
1. Decreases survival and induces apoptosis in RKO colon cancer cells: Treatment with DIM-C-pPhOCH3 leads to reduced viability and enhanced apoptotic responses in RKO colon cancer cells. This apoptotic induction is accompanied by the upregulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) protein expression [1] 2. Exhibits both Nur77-dependent and Nur77-independent apoptotic pathways: The induction of apoptosis and TRAIL expression by DIM-C-pPhOCH3 is significantly inhibited by small inhibitory RNA for Nur77 (iNur77), indicating a Nur77-dependent component. However, RNA interference studies confirm that DIM-C-pPhOCH3 also induces apoptosis through Nur77-independent mechanisms [1] 3. Regulates the expression of proapoptotic genes: Microarray analysis identifies multiple proapoptotic genes induced by DIM-C-pPhOCH3. Reverse transcription-PCR (RT-PCR) analysis in the presence or absence of iNur77 shows that the induction of programmed cell death gene 1 is Nur77-dependent, while the induction of cystathionase and activating transcription factor 3 is Nur77-independent [1] 4. Nur77 is overexpressed in colon cancer models: Nur77 is expressed in several colon cancer cell lines, including RKO, SW480, HCT-116, HT-29, and HCT-15. Immunostaining results demonstrate that Nur77 is overexpressed in colon tumors compared with normal colon tissue [1] |
| ln Vivo |
DIM-C-pPhOCH3 (25 mg/kg/d) also prevents tumor growth in RKO cell xenografted athymic nude mice. In athymic nude mice carrying RKO cell xenografts, the effects of DIM-C-pPhOCH3 (25 mg/kg/d) on colon tumor growth are also examined. Tumor volumes and final tumor weights are significantly reduced after treatment with DIM-C-phOCH3 when compared to corn oil controls[1].
Inhibits tumor growth in xenograft models: Athymic nude mice bearing RKO cell xenografts are treated with DIM-C-pPhOCH3 at a dose of 25 mg/kg per day. The treatment results in significant inhibition of tumor growth [1] |
| Cell Assay |
For two and six hours, RKO cells are exposed to either 12.5 μM DIM-C-pHOCH3 or DMSO. For the reverse transcription-PCR (RT-PCR) experiment, RNA is isolated, examined for gene expression, and three replicates are calculated for every time point and the DMSO control. An analysis is done on the microarray data[1].
1. Colon cancer cell viability and apoptosis assay: RKO colon cancer cells are cultured and treated with DIM-C-pPhOCH3. After a specific incubation period, cell survival is evaluated to assess the inhibitory effect of the drug on cell viability. Apoptotic responses are detected through relevant experimental methods, and the expression level of TRAIL protein is analyzed to confirm the apoptotic pathway activation [1] 2. RNA interference (RNAi) experiment for Nur77: RKO colon cancer cells are transfected with small inhibitory RNA for Nur77 (iNur77) or control RNA. Subsequently, the transfected cells are treated with DIM-C-pPhOCH3. The effects of iNur77 on DIM-C-pPhOCH3-induced apoptosis and TRAIL expression are analyzed to distinguish Nur77-dependent and Nur77-independent pathways [1] 3. Gene expression analysis by microarray and RT-PCR: RKO colon cancer cells are treated with DIM-C-pPhOCH3, and total RNA is extracted. Microarray analysis is performed to screen for proapoptotic genes induced by the drug. For validation, RT-PCR is conducted on selected genes (programmed cell death gene 1, cystathionase, activating transcription factor 3) in cells with or without iNur77 transfection to determine the dependence on Nur77 for their induction [1] 4. Nur77 expression detection in colon cancer cells and tissues: Multiple colon cancer cell lines (RKO, SW480, HCT-116, HT-29, HCT-15) are cultured, and Nur77 expression is detected. Additionally, immunostaining is performed on colon tumor tissues and normal colon tissues to compare the expression levels of Nur77 [1] |
| Animal Protocol |
Mice: The mice used are male athymic nude mice (Foxn1nu, aged 7-8 weeks). The mice are kept in laminar flow cabinets with particular pathogen-free conditions. In vitro cultured RKO cells (5×106 per 150 μL) are injected subcutaneously into each mouse's flank to create a xenograft. Four days are given to tumor growth before they become palpable. After that, mice are divided into two groups of six mice each, and they are given oral gavage doses of either corn oil or 25 mg/kg/d DIM-C-phOCH3 for a period of 21 days. Both the tumor size and the mice's weight are determined[1].
Xenograft tumor inhibition experiment in athymic nude mice: Athymic nude mice are implanted with RKO colon cancer cells to establish xenograft tumors. Once the tumors reach an appropriate size, the mice are administered DIM-C-pPhOCH3 at a dose of 25 mg/kg per day. The drug is administered continuously for a specified period, and tumor volume or weight is measured regularly during the treatment to evaluate the tumor growth inhibitory effect of the drug [1] |
| References | |
| Additional Infomation |
1. DIM-C-pPhOCH3 is a methylene-substituted diindolemethane (C-DIM) compound that has been identified as a Nur77 agonist [1]. 2. Nur77 is an orphan nuclear receptor with no known endogenous ligands [1]. 3. DIM-C-pPhOCH3 represents a novel class of anti-colon cancer drugs that can exert therapeutic effects through both Nur77-dependent and non-Nur77-dependent pathways [1].
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| Molecular Formula |
C24H20N2O
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| Molecular Weight |
352.43
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| Exact Mass |
352.158
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| Elemental Analysis |
C, 81.79; H, 5.72; N, 7.95; O, 4.54
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| CAS # |
33985-68-1
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| Related CAS # |
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| PubChem CID |
11371604
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| Appearance |
White to off-white solid powder
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| LogP |
5.838
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
1
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| Rotatable Bond Count |
4
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| Heavy Atom Count |
27
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| Complexity |
457
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| Defined Atom Stereocenter Count |
0
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| SMILES |
COC1=CC=C(C(C2=CNC3=C2C=CC=C3)C4=CNC5=C4C=CC=C5)C=C1
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| InChi Key |
ZCCMKJAXOIFTHH-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C24H20N2O/c1-27-17-12-10-16(11-13-17)24(20-14-25-22-8-4-2-6-18(20)22)21-15-26-23-9-5-3-7-19(21)23/h2-15,24-26H,1H3
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| Chemical Name |
3-[1H-indol-3-yl-(4-methoxyphenyl)methyl]-1H-indole
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| Synonyms |
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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 |
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| 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) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (7.09 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% 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 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 (7.09 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. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.8374 mL | 14.1872 mL | 28.3744 mL | |
| 5 mM | 0.5675 mL | 2.8374 mL | 5.6749 mL | |
| 10 mM | 0.2837 mL | 1.4187 mL | 2.8374 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.
Inhibition of tumor growth by DIM-C-pPhOCH3.Cancer Res.2007 Jan 15;67(2):674-83. |
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Induction of gene expression by DIM-C-pPhOCH3.Cancer Res.2007 Jan 15;67(2):674-83. |
Nur77-dependent induction of TRAIL and apoptosis in RKO cells. Induction of TRAIL by DIM-C-pPhOCH3Cancer Res.2007 Jan 15;67(2):674-83. |
Nur77-active C-DIMs decrease cell proliferation and induce apoptosis in RKO cells and also act through nuclear Nur77DIM-C-pPhOCH3.Cancer Res.2007 Jan 15;67(2):674-83. |
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Nur77 expression and induced transactivation in colon cancer cells and tumors.Cancer Res.2007 Jan 15;67(2):674-83. |
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