VII-31 is an activator of the NEDDylation pathway and can inhibit tumors in vitro and in vivo. VII-31 also causes apoptosis.

Physicochemical Properties

Molecular Formula	C23H25NO5S
Molecular Weight	427.5133
Exact Mass	427.145
CAS #	2305757-96-2
PubChem CID	146673076
Appearance	White to off-white solid powder
LogP	4
Hydrogen Bond Donor Count	0
Hydrogen Bond Acceptor Count	6
Rotatable Bond Count	9
Heavy Atom Count	30
Complexity	514
Defined Atom Stereocenter Count	0
SMILES	S1C([H])=C([H])C([H])=C1C([H])([H])C(N(C1C([H])=C(C(=C(C=1[H])OC([H])([H])[H])OC([H])([H])[H])OC([H])([H])[H])C([H])([H])C1C([H])=C([H])C(=C([H])C=1[H])OC([H])([H])[H])=O
InChi Key	CGVFRZMQURRQIE-UHFFFAOYSA-N
InChi Code	InChI=1S/C23H25NO5S/c1-26-18-9-7-16(8-10-18)15-24(22(25)14-19-6-5-11-30-19)17-12-20(27-2)23(29-4)21(13-17)28-3/h5-13H,14-15H2,1-4H3
Chemical Name	N-[(4-methoxyphenyl)methyl]-2-thiophen-2-yl-N-(3,4,5-trimethoxyphenyl)acetamide
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	NEDDylation[1]
ln Vitro	VII-31 (100 nM, 200 nM; 48 hours) reduces the cell viability of the gastric cell line MGC803 with an IC50 of 0.09±0.01 μM. Additionally, VII-31 inhibits MCF-7 and PC-3's ability to proliferate, with IC50 values of 0.10±0.006 and 1.15±0.28μM, respectively[1]. In G2/M phase, VII-31 (50-150 nM; 24 hr) stops the cycle of MGC803 cells[1]. Apoptosis is induced by VII-31 (50-150 nM; 48 hours) through both intrinsic and extrinsic pathways[1]. In MGC803 cells, VII-31 (50-150 nM; 24 hours) promotes NEDDylation[1]. Pro-apoptotic proteins FADD, Fasl, PIDD, Bax, and Bad are up-regulated by VII-31 (50-150 nM; 48 hours), whereas anti-apoptotic proteins Bcl-xL, Bcl-2, XIAP, and c-IAP1 are down-regulated[1].
ln Vivo	While exhibiting no overt harm to mice, VII-31 suppresses the growth of tumors in vivo[1].
Cell Assay	Cell Viability Assay[1] Cell Types: Gastric cancer MGC803 cells Tested Concentrations: 100, 200 nM Incubation Duration: 48 hrs (hours) Experimental Results: Inhibited the cell viability in dose-dependent manner. Cell Cycle Analysis[1] Cell Types: MGC803 cells Tested Concentrations: 50, 100, 150 nM Incubation Duration: 24 hrs (hours) Experimental Results: Arrested cells in G2/M phase, and a clear sub-G1 peak was observed in the high dose group. Apoptosis Analysis[1] Cell Types: MGC803 cells Tested Concentrations: 50, 75, 100, and 150 nM Incubation Duration: 48 hrs (hours) Experimental Results: High dose (150 nM) treatment Dramatically elevated the early and late apoptosis rate to 92.8% from 4.8%. Western Blot Analysis[1] Cell Types: MGC803 cells Tested Concentrations: 50, 100, 150 nM Incubation Duration: 24 hrs (hours) Experimental Results: Resulted in NEDDylation activation of MGC803 cells, the NEDDylation of 3 important proteins NAE1, Ubc12 and CUL1 has been activated.
Animal Protocol	Animal/Disease Models: Mice bearing MGC803 xenograft tumors[1] Doses: 50, 100, 150 mg/kg Route of Administration: subcutaneous (sc)injection; daily for 28 days Experimental Results: The mice had a much smaller tumor compared with vehicle control. The tumor volumes of middle/high dose treated mice at certain time points were evidently diminished comparing comparing with untreated group.
References	[1]. Discovery of Novel Tertiary Amide Derivatives as NEDDylation Pathway Activators to Inhibit the Tumor Progression in Vitro and in Vivo. Eur J Med Chem. 2020 Apr 15;192:112153.

Solubility Data

Solubility (In Vitro)

DMSO : 250 mg/mL (584.78 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.87 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 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 (4.87 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 (4.87 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	2.3391 mL	11.6956 mL	23.3913 mL
	5 mM	0.4678 mL	2.3391 mL	4.6783 mL
	10 mM	0.2339 mL	1.1696 mL	2.3391 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.