anti-cancer compound 42 (compound 10d) is an orally bioactive anti-cancer compound with strong anti-tumor effects on MDA-MB-231 cells, with IC50 of 0.07 μM. anti-cancer compound 42 exerts anti-cancer activities by activating the apoptotic pathway and p53 expression. anti-cancer compound 42 may be utilized in the research/study of metastatic breast cancer.

Physicochemical Properties

Molecular Formula	C19H16CL2N2O3
Molecular Weight	391.2479429245
Exact Mass	390.053
CAS #	2687265-18-3
PubChem CID	163322347
Appearance	Typically exists as solid at room temperature
LogP	3.8
Hydrogen Bond Donor Count	1
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	2
Heavy Atom Count	26
Complexity	697
Defined Atom Stereocenter Count	0
InChi Key	QGROEQOAPSQCOH-UHFFFAOYSA-N
InChi Code	InChI=1S/C19H16Cl2N2O3/c1-10-13(11-4-2-3-5-12(11)22-10)14-15(20)19(25)16(21)17(18(14)24)23-6-8-26-9-7-23/h2-5,22H,6-9H2,1H3
Chemical Name	2,6-dichloro-3-(2-methyl-1H-indol-3-yl)-5-morpholin-4-ylcyclohexa-2,5-diene-1,4-dione
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

ln Vitro	On MDA-MB-231 cells, anticancer agent 42 (compound 10d) (0–20 μM, 4 hours) demonstrates strong antitumor activity [1]. In MDA-MB-231 cells, anticancer drug 42 (10 μM, 24 hours) causes G2 and S phase arrest [1]. In MDA-MB-231 cells, anticancer drug 42 (10 μM, 24 h) causes apoptosis via controlling the expression of proteins linked to apoptosis [1]. Apoptosis results from the depolarization of the mitochondrial membrane and the reduction of mitochondrial membrane potential caused by anticancer drug 42 (0-1 μM) [1]. Anticancer agent 42 (0–1 μM, 24 hours) causes a significant quantity of ROS production in cells [1].
ln Vivo	Anticancer agent 42 (compound 10d) has a very low oral toxicity (Kunming mice, 5000 mg/kg, gavage, once) [1]. The LD50 of anticancer agent 42 (Kunming mice, 238-600 mg/kg, IP, once) is 374 mg/kg, and there is no discernible liver or kidney damage in mice[1]. Mild liver and kidney damage was brought on by anticancer agent 42 (Kunming mice, 25 mg/kg, IP, once every two days) [1]. Breast cancer 4T1 tumor growth is inhibited by anticancer agent 42 (BALB/c mice). When paired with cyanoacrylate (CA), it has a stronger anti-tumor effect and can penetrate the skin to do so. [1].
Cell Assay	Cell viability assay Cell Types: A549, MDA-MB-231, HeLa[1] Tested Concentrations: 0-20 μM Incubation Duration: 4 hrs (hours) Experimental Results: demonstrated effective activity against MDA-MB-231 with IC50 of 0.07 μM. Cell cycle analysis Cell Types: MDA-MB-231 cells [1] Tested Concentrations: 10 μM Incubation Duration: 24 hrs (hours) Experimental Results: Induced G2 and S phase arrest of MDA-MB-231 cells; resulted in a significant proportion of MDA-MB-231 cells in the G1 phase The proportion of cells in G1 phase increased Dramatically (from 74.44% to 16.48%), the proportion of cells in G1 phase increased (from 16.61% to 28.47%), and the proportion of cells in G2 phase increased Dramatically (from 8.95%). % to 55.05%). Apoptosis analysis Cell Types: MDA-MB-231 cells [1] Tested Concentrations: 10 μM Incubation Duration: 24 h Experimental Results: Cell apoptosis was induced, and the apoptosis rate was 31.69%. Western Blot Analysis Cell Types: MDA-MB-231 cells [1] Tested Concentrations: 100 nM Incubation Duration: 48 hrs (hours) Experimental Results: Human apoptosis-related proteins (pro-caspase 3, catalase
Animal Protocol	Animal/Disease Models: Kunming mice (n=10, 5 males and 5 females) [1] Doses: 5000mg/kg Route of Administration: intragastric (po) (po)administration, once. Experimental Results: Oral toxicity is extremely low, and 5000mg/kg will not cause morbidity. Rat died. Model: Kunming mice [1] Doses: 600, 476, 378, 300, 238 mg/kg Route of Administration: IP, one time Experimental Results:no obvious liver and kidney damage to mice, LD50 is 374 mg/kg. Kunming mice (n=3) [1] Doses: 25 mg/kg Route of Administration: IP, once every two days Experimental Results: Mild liver and kidney damage was caused after administration, and ALT, AST and BUN in mice were slightly increased. Animal/Disease Models: balb/c (Bagg ALBino) mouse (4T1 tumor-bearing, female, 8 groups, 6 mice in each group) [1] Doses: 10d (50 mg/kg) + CA; 10d (50 mg/kg) + saline; 10d (200 mg/kg) + CA Route of Administration: intratumoral injection (50 mg/kg) every four days; smear (200 mg/kg) every two days for 14 days. Experimental Results: There was obvious anti-tumor effect from the 8th day; it had a protective effect on the spleen of tumor-
References	[1]. Design, synthesis, and evaluation of a novel series of mono-indolylbenzoquinones derivatives for the potential treatment of breast cancer. Eur J Med Chem. 2022 Apr 16;237:114375.

Solubility Data

Solubility (In Vitro)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

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	2.5559 mL	12.7796 mL	25.5591 mL
	5 mM	0.5112 mL	2.5559 mL	5.1118 mL
	10 mM	0.2556 mL	1.2780 mL	2.5559 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.