JNK Inhibitor VIII (TCS JNK 6o) is a selective and ATP-competitive inhibitor of the JNK-1, -2, and -3 c-Jun N-terminal kinases, with Kis of 2 nM, 4 nM, and 52 nM, respectively, and IC50 values of 45 nM and 160 nM for JNK-1 and -2, respectively.

Physicochemical Properties

Molecular Formula	C₁₈H₂₀N₄O₄
Molecular Weight	356.38
Exact Mass	356.148
Elemental Analysis	C, 60.66; H, 5.66; N, 15.72; O, 17.96
CAS #	894804-07-0
Related CAS #	894804-07-0
PubChem CID	11624601
Appearance	White to off-white solid powder
Density	1.3±0.1 g/cm3
Boiling Point	658.6±55.0 °C at 760 mmHg
Flash Point	352.1±31.5 °C
Vapour Pressure	0.0±2.0 mmHg at 25°C
Index of Refraction	1.599
LogP	4.86
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	7
Rotatable Bond Count	7
Heavy Atom Count	26
Complexity	509
Defined Atom Stereocenter Count	0
SMILES	N#CC1C(N)=CC(NC(CC2C(OC)=CC=C(OC)C=2)=O)=NC=1OCC
InChi Key	KQMPRSZTUSSXND-UHFFFAOYSA-N
InChi Code	InChI=1S/C18H20N4O4/c1-4-26-18-13(10-19)14(20)9-16(22-18)21-17(23)8-11-7-12(24-2)5-6-15(11)25-3/h5-7,9H,4,8H2,1-3H3,(H3,20,21,22,23)
Chemical Name	N-(4-amino-5-cyano-6-ethoxypyridin-2-yl)-2-(2,5-dimethoxyphenyl)acetamide
Synonyms	TCS-JNK-6o; TCS-JNK 6o; TCS JNK-6o; JNK Inhibitor VIII; c-Jun N-terminal Kinase Inhibitor VIII
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	JNK1 (Ki = 2 nM); JNK1 (IC50 = 45 nM); JNK2 (Ki = 4 nM); JNK2 (Ki = 160 nM); JNK3 (Ki = 52 nM)
ln Vitro	TCS-JNK-6a binds to the ATP site in a competitive and reversible manner. This substance exhibits an impressive selectivity profile. In hepG2 cells, it also exhibits some inhibition of c-Jun phosphorylation. TCS-JNK-6o is a 2,5-dimethoxy analogue of TCS-JNK-6a in which the phenyl ring has been replaced by a dimethoxy group. TCS-JNK-6o exhibits additional increases in potency. [1]
ln Vivo	JNK Inhibitor VIII (TCS-JNK-6o) pharmacokinetic profiles are established in Sprague-Dawley rats. The JNK Inhibitor VIII (TCS-JNK-6o) exhibits a brief half-life of about 1 h, rapid clearance, and hardly detectable bioavailability. Studies on microsomal incubation have shown that this compound has a very quick oxidative metabolism. [1]
Enzyme Assay	Radioactive FlashPlates are used as the assay platform for Ser/Thr-kinase assays. In this format, biotinylated substrate peptide (2 μM), γ-[33P]-ATP (5 μM, 2 mCi/μmol), inhibitors (3-10000 nM in 2% DMSO), and enzyme are incubated for 1 h in buffer containing 25 mM Hepes, pH 7.5, 1 mM DTT, 10 mM MgCl2, 100 μM Na3VO4, and 0.075 mg/mL Triton X-100, stopped with 80 μL of stop buffer containing 100 mM EDTA and 4 M NaCl, transferred to streptavidin-coated 384-well FlashPlates, which are then washed 3 times and read using a TopCount microplate reader.
Cell Assay	HepG2 human hepatoma cells (ATCC) are grown in low glucose MEM with 1×NEAA, 1×sodium pyruvate, and 10% FBS as dietary supplements. Cells are seeded at 5×104 cells/well in 500 μL of complete media on 24-well collagen-coated plates for P-c-Jun assays, and the plates are incubated overnight. The final inhibitor concentrations are provided by adding 5 μL directly to the media on the cells after serial compound dilutions in DMSO at 100×. Cells are stimulated for 30 minutes with TNFR or vehicle control after 1 hour, harvested in 70 μL of lysis buffer (TBS (54 mM Tris-HCl, pH 7.6, 150 mM NaCl), 1% TritonX-100,0.5% Nonidet P-40, 0.25% sodium deoxycholate, 1 mM EDTA, 1 mM EGTA, 0.5 mM sodium fluoride, 1 mM pervanadate, 1 μM microcystin, 1 mM AEBSF, 1 tablet of complete EDTA Free-Mini inhibitor cocktail) and then frozen at -80 degrees Celsius for use in the P-c-Jun assay.
Animal Protocol	Sprague-Dawley rat 5 mg/kg IV, Oral gavage
References	[1]. Aminopyridine-based c-Jun N-terminal kinase inhibitors with cellular activity and minimal cross-kinase activity. J Med Chem. 2006 Jun 15;49(12):3563-80.

Solubility Data

Solubility (In Vitro)

DMSO: 71~250 mg/mL (199.2~701.5 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.08 mg/mL (5.84 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	2.8060 mL	14.0300 mL	28.0599 mL
	5 mM	0.5612 mL	2.8060 mL	5.6120 mL
	10 mM	0.2806 mL	1.4030 mL	2.8060 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.