(E)-AG 99 ((E)-Tyrphostin 46) is a potent EGFR inhibitor.

Physicochemical Properties

Molecular Formula	C10H8N2O3
Molecular Weight	204.18212
Exact Mass	204.053
CAS #	122520-85-8
Related CAS #	122520-85-8
PubChem CID	5328768
Appearance	Light yellow to yellow solid
Density	1.5±0.1 g/cm3
Boiling Point	544.4±50.0 °C at 760 mmHg
Melting Point	231 °C
Flash Point	283.0±30.1 °C
Vapour Pressure	0.0±1.5 mmHg at 25°C
Index of Refraction	1.711
LogP	0.02
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	2
Heavy Atom Count	15
Complexity	329
Defined Atom Stereocenter Count	0
SMILES	O=C(N)/C(C#N)=C/C1=CC=C(O)C(O)=C1
InChi Key	USOXQZNJFMKTKJ-XVNBXDOJSA-N
InChi Code	InChI=1S/C10H8N2O3/c11-5-7(10(12)15)3-6-1-2-8(13)9(14)4-6/h1-4,13-14H,(H2,12,15)/b7-3+
Chemical Name	(E)-2-cyano-3-(3,4-dihydroxyphenyl)prop-2-enamide
Synonyms	(E)-AG 99; (E)-Tyrphostin 46; (E)-Tyrphostin AG 99
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	EGFR Epidermal growth factor receptor (EGFR) inhibitor.[1]
ln Vitro	(E)-AG 99 stimulates cell death via the activation of caspase-like proteases and efficiently inhibits the tyrosine phosphorylation of p145met[1]. AG99 (5 μM) was shown to inhibit tyrosine phosphorylation of p145met (a 145-kDa protein later identified as the β-subunit of c-Met/HGF receptor) in human bladder carcinoma 5637 cells that had been serum-starved for 24 hours. This inhibition was partial compared to complete inhibition by Src family kinase (SFK) inhibitors (PP2, SU6656).[1] AG99 demonstrated a growth-inhibitory effect on 5637 cells under both normal culture conditions (with serum) and serum-starved conditions.[1] The inactive analog AG9 did not show any inhibitory effect on tyrosine phosphorylation of p145met under the same conditions.[1]
Cell Assay	Human bladder carcinoma 5637 cells were cultured in normal medium (containing 10% fetal calf serum, FCS) or serum-free medium for 24 hours. For the final 1 hour of serum starvation treatment, cells were incubated in the absence or presence of AG99 (5 μM). Whole cell extracts were then prepared and analyzed by immunoblotting with anti-phosphotyrosine antibody to assess the phosphorylation status of proteins, specifically the 145-kDa protein (p145met).[1] The growth-inhibitory effect of AG99 was evaluated by comparing cell numbers under different culture conditions (with or without serum) in the presence of the inhibitor.[1]
References	[1]. Tyrosine phosphorylation of p145met mediated by EGFR and Src is required for serum-independent survival of human bladder carcinoma cells. J Cell Sci. 2006 Nov 15;119(Pt 22):4623-33.
Additional Infomation	Tyrphostin A46 is a protein tyrosine kinase inhibitor that selectively inhibits EGFR kinase, involved in signaling pathways. (NCI) AG99 is used as a pharmacological tool to inhibit EGFR activity in this study investigating the mechanism of serum-independent survival in bladder cancer cells.[1] Its inhibitory effect on p145met phosphorylation, along with data using other inhibitors and antibodies, supports the conclusion that a signaling cascade involving EGFR ligands, EGFR, Src family kinases (SFKs), and p145met is crucial for suppressing caspase activation and enabling survival of 5637 cells under serum-free conditions.[1] The study notes that since EGFR activation was not observed in normal culture conditions, the growth-inhibitory effect of AG99 under normal conditions suggests it may have alternative target(s) besides EGFR, or that EGFR's importance is not limited to serum-independent growth.[1]

Solubility Data

Solubility (In Vitro)

DMSO: 41~165 mg/mL (200.8~808.1 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.75 mg/mL (13.47 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 27.5 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: ≥ 2.75 mg/mL (13.47 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 27.5 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	4.8976 mL	24.4882 mL	48.9764 mL
	5 mM	0.9795 mL	4.8976 mL	9.7953 mL
	10 mM	0.4898 mL	2.4488 mL	4.8976 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.