EN4 is a covalent ligand for cysteine 171 (C171) of MYC. EN4 is more selective for c-MYC than N-MYC and L-MYC. EN4 inhibits MYC transcriptional activity, downregulates MYC targets, and Has potential anticancer/anti-tumor effects.

Physicochemical Properties

Molecular Formula	C25H24N2O4
Molecular Weight	416.477
Exact Mass	416.173
Elemental Analysis	C, 72.10; H, 5.81; N, 6.73; O, 15.37
CAS #	1197824-15-9
Related CAS #	1197824-15-9
PubChem CID	45917225
Appearance	White to light yellow solid powder
Density	1.2±0.1 g/cm3
Boiling Point	564.1±50.0 °C at 760 mmHg
Flash Point	295.0±30.1 °C
Vapour Pressure	0.0±1.5 mmHg at 25°C
Index of Refraction	1.614
LogP	3.54
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	9
Heavy Atom Count	31
Complexity	580
Defined Atom Stereocenter Count	0
SMILES	O(C1C=CC(=CC=1)OCC)C1=CC=CC=C1NC(C1C=CC(=CC=1)CNC(C=C)=O)=O
InChi Key	BHCJZNGDYXVVII-UHFFFAOYSA-N
InChi Code	InChI=1S/C25H24N2O4/c28-25(29)18-30-17-23-9-7-20(16-27-12-3-11-26-27)15-24(23)31-13-10-19-6-8-21-4-1-2-5-22(21)14-19/h1-9,11-12,14-15H,10,13,16-18H2,(H,28,29)
Chemical Name	2-[[2-(2-naphthalen-2-ylethoxy)-4-(pyrazol-1-ylmethyl)phenyl]methoxy]acetic acid
Synonyms	EN-4; EN4; EN 4; EN4 MYC inhibitor
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 Note: This product requires protection from light (avoid light exposure) during transportation and storage.
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	MYC
ln Vitro	En4 inhibits MYC transcriptional activity, downregulates numerous MYC transcriptional targets, directly targets MYC in cells, decreases MYC and MAX thermal stability, and hinders tumorigenesis. Treatment with EN4 significantly reduces the proliferation of 231MFP breast cancer cells in a dose- and time-dependent manner. Mammary epithelial MCF10A cells that have undergone MYC transformation are less likely to survive when exposed to EN4, but not parental MCF10A cells that are not MYC-dependent. [1]
ln Vivo	In immune-deficient mice, EN4 treatment that is started after the establishment of a 231MFP breast tumor xenograft significantly slows tumor growth in vivo.[1]
Cell Assay	In 6 cm dishes, 231MFP cells are plated, and they are then given DMSO vehicle or EN4 (50 μM) treatment for 24 hours. Scraping is used to collect the cells, and the Qiagen RNeasy mini kit and Qiagen DNase max kit are used to isolate the RNA and remove any DNA contamination. The KAPA mRNA Capture Kit's Oligo dT beads are used to enrich for mRNA.
References	[1]. Discovery of a Functional Covalent Ligand Targeting an Intrinsically Disordered Cysteine within MYC. Cell Chem Biol. 2021 Jan 21;28(1):4-13.e17.

Solubility Data

Solubility (In Vitro)

DMSO: 83~125 mg/mL (199.3~300.1 mM)

Solubility (In Vivo)

Solubility in Formulation 1: 2.08 mg/mL (4.99 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 20.8 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: 2.08 mg/mL (4.99 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 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. Solubility in Formulation 3: ≥ 2.08 mg/mL (4.99 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.4011 mL	12.0054 mL	24.0108 mL
	5 mM	0.4802 mL	2.4011 mL	4.8022 mL
	10 mM	0.2401 mL	1.2005 mL	2.4011 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.