AZD8330 (AZD-8330; ARRY-704; ARRY-424704) is an orally bioactive and non-ATP competitive (Allosteric) MEK 1/2 inhibitor with potential anticancer activity. It has an IC50 of 7 nM for MEK1/2 inhibition.

Physicochemical Properties

Molecular Formula	C16H17FIN3O4
Molecular Weight	461.23
Exact Mass	461.024
Elemental Analysis	C, 41.67; H, 3.72; F, 4.12; I, 27.51; N, 9.11; O, 13.88.
CAS #	869357-68-6
Related CAS #	869357-68-6
PubChem CID	16666708
Appearance	Off-white to pink solid powder
Density	1.7±0.1 g/cm3
Index of Refraction	1.659
LogP	2.05
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	6
Rotatable Bond Count	6
Heavy Atom Count	25
Complexity	596
Defined Atom Stereocenter Count	0
SMILES	O=C(C1=C(NC2C(F)=CC(I)=CC=2)N(C)C(=O)C(C)=C1)NOCCO
InChi Key	RWEVIPRMPFNTLO-UHFFFAOYSA-N
InChi Code	InChI=1S/C16H17FIN3O4/c1-9-7-11(15(23)20-25-6-5-22)14(21(2)16(9)24)19-13-4-3-10(18)8-12(13)17/h3-4,7-8,19,22H,5-6H2,1-2H3,(H,20,23)
Chemical Name	2-(2-fluoro-4-iodoanilino)-N-(2-hydroxyethoxy)-1,5-dimethyl-6-oxopyridine-3-carboxamide
Synonyms	ARRY704; AZD 8330; AZD8330; ARRY424704; ARRY-704; ARRY 424704; ARRY 704; ARRY-424704; AZD-8330
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	MEK1 (IC50 = 7 nM); MEK2 (IC50 = 7 nM)
ln Vitro	AZD8330 is a selective allosteric MEK1/ MEK2 inhibitor. Loss of ERK phosphorylation indicates effective MEK inhibition when the human osteosarcoma cell lines MOS, U2OS, and 143B are exposed to 0.5 μM of trametinib, AZD8330, or TAK-733 for 6 hours. Six osteosarcoma cell lines—MOS, U2OS, KPD, ZK58, 143b, and Saos-2—are used to test the activity of these three inhibitors at different concentrations. All three inhibitors have a significant negative impact on 143b and reduce the viability of MOS and U2OS. However, none of the three inhibitors affect the viability of KPD, ZK58, or Saos-2[2].
ln Vivo	At tolerated doses (1.0 mg/kg once daily [OD]), AZD8330 exhibits a dose-dependent tumour growth inhibition of 90% in tumor xenograft models[1].
Enzyme Assay	MEK1 (S218D, S222D ΔR4F) is expressed in baculovirus-infected Hi5 insect cells and purified using immobilized metal affinity chromatography, ion exchange, and gel filtration. MEK1 is also NH2-terminally hexahistidine-tagged and constitutively active. When [γ- 33P]phosphate from [γ- 33P]ATP is incorporated onto ERK2, it is possible to measure the activity of MEK1 in the body. An incubation solution (100 μL) consisting of 25 mM HEPES (pH 7.4), 10 mM MgCl2, 5 mM β-glycerolphosphate, 100 μM sodium orthovanadate, 5 mM DTT, 5 nM MEK1, 1 μM ERK2, and 0 to 80 nM AZD8330 (final concentration of 1% DMSO) is used in the assay. The addition of 10 μM ATP (with 0.5 μC k[γ-33P]ATP/well) starts the reactions, which are then allowed to proceed for 45 min at room temperature. For the purpose of stopping the reaction and precipitating the proteins, an equal volume of 25% trichloracetic acid is added. The excess labeled ATP is removed with 0.5% phosphoric acid, the precipitated proteins are trapped on glass fiber B filter plates, and radioactivity is measured using a liquid scintillation counter. By changing the concentration of ATP in the reaction mixture, ATP dependence can be identified. The data have been fitted globally.
Cell Assay	Malme-3M melanoma cells are plated in 96-wells and subjected to AZD8330 treatments in a range of concentrations for an hour at 37 °C. Anti-phospho-ERK and anti-ERK 1/2 antibodies are incubated with the cells after they have been fixed and permeabilized. Secondary antibodies that have been fluorescently labeled are added after washing the plates. Utilizing a LICOR fluorescence imager, plates are examined. The total ERK signal is used to normalize the pERK signal.
Animal Protocol	Female nude rats (NIH rnu/rnu) with Calu-6 cells, nude rats with SW620 cells 0.3 mg/kg, 1 mg/kg Oral administration
References	[1]. A phase I dose-finding, safety and tolerability study of AZD8330 in patients with advanced malignancies. Eur J Cancer. 2013 May;49(7):1521-9./a > [2]. MEK inhibition induces apoptosis in osteosarcoma cells with constitutive ERK1/2 phosphorylation. Genes Cancer. 2015 Nov;6(11-12):503-12.
Additional Infomation	2-(2-fluoro-4-iodoanilino)-N-(2-hydroxyethoxy)-1,5-dimethyl-6-oxo-3-pyridinecarboxamide is a pyridinecarboxamide. It is functionally related to a nicotinamide. AZD-8330 is a potent, selective, orally active MEK inhibitor that blocks signal transduction pathways implicated in cancer cell proliferation and survival. AZD-8330 has shown tumor suppressive activity in multiple preclinical models of human cancer including melanoma, pancreatic, colon, lung, and breast cancers. MEK Inhibitor AZD8330 is an orally active, selective MEK inhibitor with potential antineoplastic activity. MEK inhibitor AZD8330 specifically inhibits mitogen-activated protein kinase kinase 1 (MEK or MAP/ERK kinase1), resulting in inhibition of growth factor-mediated cell signaling and tumor cell proliferation. MEK is a key component of the RAS/RAF/MEK/ERK signaling pathway that regulates cell growth; constitutive activation of this pathway has been implicated in many cancers. Drug Indication Investigated for use/treatment in cancer/tumors (unspecified). Mechanism of Action AZD8330 specifically inhibits mitogen-activated protein kinase kinase 1 (MEK or MAP/ERK kinase1), resulting in inhibition of growth factor-mediated cell signaling and tumor cell proliferation. MEK is a key component of the RAS/RAF/MEK/ERK signaling pathway that regulates cell growth; constitutive activation of this pathway has been implicated in many cancers.

Solubility Data

Solubility (In Vitro)

DMSO: ~92 mg/mL (~199.5 mM) Water: <1 mg/mL Ethanol: ~92 mg/mL (~199.5 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 3.25 mg/mL (7.05 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 32.5 mg/mL clear DMSO stock solution to 400 μL of PEG300 and mix evenly; then add 50 μL of Tween-80 to the above solution and mix evenly; then add 450 μL of 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: ≥ 3.25 mg/mL (7.05 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 32.5 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. Solubility in Formulation 3: ≥ 3.25 mg/mL (7.05 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 32.5 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Solubility in Formulation 4: 0.5% hydroxyethyl cellulose+0.1% Tween 80: 30mg/mL  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.1681 mL	10.8406 mL	21.6812 mL
	5 mM	0.4336 mL	2.1681 mL	4.3362 mL
	10 mM	0.2168 mL	1.0841 mL	2.1681 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.