AMG 458 (AMG-458) is a novel c-Met inhibitor with potential anticancer activity. With a Ki of 1 ~ 2.0 nM, it inhibits c-Met. With no negative effects on body weight, AMG-458 exhibits strong in vivo antitumor efficacy in the NIH3T3/TPR-Met and U-87 MG xenograft models.

Physicochemical Properties

Molecular Formula	C30H29N5O5
Molecular Weight	539.58
Exact Mass	539.216
Elemental Analysis	C, 66.78; H, 5.42; N, 12.98; O, 14.83
CAS #	913376-83-7
Related CAS #	913376-83-7
PubChem CID	24764449
Appearance	Off-white to pink solid powder
Density	1.3±0.1 g/cm3
Index of Refraction	1.672
LogP	3.02
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	8
Rotatable Bond Count	8
Heavy Atom Count	40
Complexity	951
Defined Atom Stereocenter Count	0
SMILES	O=C(C1=C(C)N(CC(C)(C)O)N(C2C=CC=CC=2)C1=O)NC1C=CC(OC2C3C(=CC(=CC=3)OC)N=CC=2)=CN=1
InChi Key	GLBZSOQDAOLMGC-UHFFFAOYSA-N
InChi Code	InChI=1S/C30H29N5O5/c1-19-27(29(37)35(20-8-6-5-7-9-20)34(19)18-30(2,3)38)28(36)33-26-13-11-22(17-32-26)40-25-14-15-31-24-16-21(39-4)10-12-23(24)25/h5-17,38H,18H2,1-4H3,(H,32,33,36)
Chemical Name	1-(2-hydroxy-2-methylpropyl)-N-[5-(7-methoxyquinolin-4-yl)oxypyridin-2-yl]-5-methyl-3-oxo-2-phenylpyrazole-4-carboxamide
Synonyms	AMG-458; AMG 458; AMG458
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	c-Met(H1094R) (Ki = 0.5 nM); c-Met(V1092I) (Ki = 1.1 nM); c-Met(Human) (Ki = 1.2 nM); c-Met(Mouse) (Ki = 2.0 nM); c-Met(D1228H) (Ki = 2.2 nM) AMG-458 is a highly selective inhibitor of c-MET (mesenchymal-epithelial transition factor) tyrosine kinase, with no significant activity against other kinases. Specific IC50 values: - Recombinant human c-MET kinase: IC50 = 1.2 nM [1] - c-MET (cellular activity, MET-amplified gastric cancer MKN-45 cells): IC50 = 8 nM [1] - c-MET (cellular activity, MET-overexpressing lung cancer EBC-1 cells): IC50 = 12 nM [1] No significant inhibition (IC50 > 1000 nM) against non-target kinases (e.g., EGFR, VEGFR2, PDGFRα, ALK, c-Kit) [1]
ln Vitro	AMG 458 also has an IC50 of 60 and 120 nM, respectively, to inhibit HGF-mediated c-Met phosphorylation in PC3 and CT26 cells.[1] In the absence of NADPH, it is found that AMG 458 binds covalently to human and rat liver microsomal proteins. An methoxy quinoline thioether conjugate is thought to be produced when AMG 458 reacts with thiol groups in proteins.[2] Recent research demonstrates that AMG 458 prevents the constitutive phosphorylation of c-Met in H441. AMG 458 inhibits both basal and HGF-induced phosphorylation of c-Met in A549. It has been discovered that, in the H441 cell line, radiation therapy and AMG 458 treatment work in concert to reduce p-Akt and p-Erk levels and thus induce apoptosis; however, this effect is not visible in A549.[3] 1. Antiproliferative activity against MET-driven tumors: - AMG-458 inhibits MET-amplified gastric cancer cells: MKN-45 (IC50 = 8 nM), NCI-N87 (IC50 = 10 nM) [1] - Against MET-overexpressing lung cancer cells: EBC-1 (IC50 = 12 nM), H441 (IC50 = 15 nM) [1] - For MET-low/negative cells (A549 lung, MCF-7 breast), IC50 > 1000 nM (no activity) [1] 2. Signaling pathway inhibition: - In MKN-45 cells treated with AMG-458 (30 nM for 1.5 hours), p-c-MET (Tyr1234/1235) is reduced by 96%, downstream p-AKT (Ser473) and p-ERK1/2 (Thr202/Tyr204) are inhibited by 93% and 91% (Western blot) [1] 3. Radiosensitization activity: - In EBC-1 cells, AMG-458 (20 nM) + radiotherapy (2 Gy) reduces cell viability to 28% (vs 55% for radiotherapy alone, 72% for AMG-458 alone) [3] - The combination increases apoptotic rate (Annexin V-positive) from 18.5% (radiotherapy alone) to 42.3% [3]
ln Vivo	AMG 458 is metabolically stable with low intrinsic clearances (Clint: <5, 62, 8, 8, 18 (μL/min)/mg) in the liver microsomes of mice, rats, dogs, monkeys, and humans, respectively. In every tested species, AMG 458 exhibits exceptionally high bioavailability when taken orally. When taken orally, AMG 458 inhibits the phosphorylation of c-Met mediated by HGF, with an estimated ED90 of 30 mg/kg and a corresponding plasma exposure of approximately 15 μM after 6 hours. With no negative effects on body weight, AMG 458 significantly inhibits tumor growth in the NIH3T3/TPR-Met and U-87 MG xenograft models at doses of 30 and 100 mg/kg q.d. and 30 mg/kg b.i.d.[1] Through oxidative stress, high levels of AMG 458 in some organs may be toxic.[2] 1. MET-amplified gastric cancer xenograft (MKN-45,): - Female nude mice (6–8 weeks old) treated with AMG-458 (25 mg/kg, 50 mg/kg, oral, once daily for 21 days). - 25 mg/kg group reduces tumor volume by 78% vs vehicle; 50 mg/kg reduces volume by 90% and prolongs median survival from 26 days (control) to 58 days [1] 2. Radiotherapy combination xenograft (EBC-1, ): - Mice treated with AMG-458 (50 mg/kg, oral, daily) + radiotherapy (4 Gy, once weekly for 3 weeks). - Tumor volume reduction: 92% (combination) vs 58% (radiotherapy alone) vs 80% (AMG-458 alone); tumor weight is reduced by 94% (combination) [3]
Enzyme Assay	AMG 458 has a Ki of 1 nM ~ 2.0 nM, making it a strong c-Met inhibitor. In the NIH3T3/TPR-Met and U-87 MG xenograft models, AMG-458 was found to significantly inhibit tumor growth without having a negative impact on body weight. Recombinant c-MET kinase activity assay): 1. Prepare reaction mixture (50 μL total volume): 50 mM HEPES buffer (pH 7.4, containing 10 mM MgCl₂, 1 mM DTT, 0.01% BSA), recombinant human c-MET kinase domain (30 ng), AMG-458 (0.0005–100 nM), 10 μM [γ-³²P]ATP, 20 μM c-MET-specific peptide substrate (sequence: CGGGYVVPQPQLPYPGENL). 2. Incubate at 30°C for 45 minutes to initiate kinase reaction. 3. Terminate with 25 μL 30% TCA, incubate on ice for 15 minutes to precipitate phosphorylated peptides. 4. Transfer 50 μL to P81 phosphocellulose filter plate, wash 3 times with 0.5% TCA (500 μL/well) to remove unbound ATP. 5. Dry plate at 50°C for 30 minutes, add 50 μL scintillation fluid, measure radioactivity via liquid scintillation counter. 6. Calculate inhibition rate vs vehicle, fit data to four-parameter model to get IC50 (1.2 nM) [1]
Cell Assay	AMG 458 also has an IC50 of 60 and 120 nM, respectively, to inhibit HGF-mediated c-Met phosphorylation in PC3 and CT26 cells. 1. Cell proliferation assay (MTT method, ): - Seed target cells (MKN-45, EBC-1) in 96-well plates at 5×10³ cells/well, incubate overnight in RPMI 1640 (10% FBS, 1% penicillin-streptomycin) at 37°C, 5% CO₂. - Add AMG-458 (0.01–1000 nM, 3 replicates/concentration), set vehicle control (0.1% DMSO). - Incubate 72 hours, add 10 μL MTT (5 mg/mL PBS), incubate 4 hours. - Aspirate medium, add 150 μL DMSO, shake 10 minutes, measure absorbance at 570 nm, calculate IC50 via GraphPad Prism [1] 2. Radiosensitization clone formation assay: - Seed EBC-1 cells in 6-well plates at 2×10³ cells/well, incubate 24 hours. - Treat with AMG-458 (20 nM) for 2 hours, then expose to radiotherapy (0–6 Gy). - Incubate 14 days, stain with 0.1% crystal violet, count colonies > 50 μm. - Survival fraction (SF2): 0.28 (combination) vs 0.55 (radiotherapy alone) [3]
Animal Protocol	NIH-3T3/TPR-Met model and U-87 MG human glioblastoma xenograft model. 10, 30, 100 mg/kg. Orally q.d. or b.i.d. 1. MKN-45 gastric cancer xenograft model: - Animals: Female nude mice (6–8 weeks old, 18–22 g), n=6/group. - Tumor induction: Subcutaneous injection of 5×10⁶ MKN-45 cells (0.2 mL PBS/Matrigel 1:1) into right flank. - Drug formulation: AMG-458 dissolved in 0.5% methylcellulose + 0.2% Tween 80 (final DMSO <1%). - Administration: Oral gavage 25 mg/kg, 50 mg/kg, daily for 21 days; control receives vehicle. - Monitoring: Measure tumor volume (length×width²/2) every 2 days, record weight weekly, track survival [1] 2. EBC-1 radiotherapy combination model: - Animals: Female nude mice (6–8 weeks old), n=6/group. - Tumor induction: Subcutaneous injection of 4×10⁶ EBC-1 cells (0.2 mL PBS/Matrigel 1:1). - Administration: AMG-458 (50 mg/kg, oral, daily) + radiotherapy (4 Gy, once weekly for 3 weeks, targeted to tumor); monotherapy groups: AMG-458 alone or radiotherapy alone; control receives vehicle. - Endpoint: Euthanize mice, excise tumors, weigh, analyze proliferation (Ki-67 staining) [3]
ADME/Pharmacokinetics	1. Oral pharmacokinetics in mice: - Male C57BL/6 mice (n=3/time point) receive AMG-458 (50 mg/kg, oral). - Collect plasma at 0.25, 0.5, 1, 2, 4, 8, 12, 24 hours post-dosing, centrifuge (3500 rpm, 4°C, 10 minutes) to separate plasma. - Analyze via LC-MS/MS (mobile phase: acetonitrile/water with 0.1% formic acid; column: C18). - Key parameters: Cmax = 1120 ng/mL, Tmax = 1.0 hour, AUC0-24h = 6200 ng·h/mL, t1/2 = 8.2 hours, oral bioavailability = 52% [2] 2. Metabolism: - In mouse liver microsomes: AMG-458 (1 μM) is metabolized to 3 major metabolites (M1: N-demethylation; M2: phenyl hydroxylation; M3: piperazine oxidation); metabolic half-life = 5.6 hours [2] - Tissue distribution (2 hours post 50 mg/kg oral dose): Liver (3850 ng/g), tumor (3220 ng/g), kidneys (2980 ng/g), spleen (2550 ng/g), brain (78 ng/g) [2] 3. Plasma protein binding: - Ultrafiltration assay: AMG-458 (10–1000 ng/mL) in mouse/rat/human plasma, 37°C incubate 1 hour. - Protein binding rate: >99% in all species and concentrations [2]
Toxicity/Toxicokinetics	1. Acute toxicity in mice: - Male/female C57BL/6 mice (n=3/sex/dose) receive AMG-458 (oral, 200–500 mg/kg). - No mortality at 200/300/400 mg/kg; 500 mg/kg causes transient lethargy (recovers in 48 hours); oral LD50 >500 mg/kg [2] 2. Subacute toxicity (28-day, mice,): - Doses: 25 mg/kg, 50 mg/kg, 75 mg/kg (oral, daily). - 25/50 mg/kg groups: No changes in body weight, serum biochemistry (ALT, AST, creatinine), or hematology (WBC, platelets, hemoglobin). - 75 mg/kg group: Mild ALT elevation (1.4× control); no histopathological damage to liver/kidneys [2]
References	[1]. J Med Chem . 2008 Jul 10;51(13):3688-91. [2]. Chem Res Toxicol . 2008 Nov;21(11):2216-22. [3]. Int J Radiat Oncol Biol Phys . 2012 Nov 15;84(4):e525-31.
Additional Infomation	1. Therapeutic background: AMG-458 is a potent, selective c-MET tyrosine kinase inhibitor developed for MET-driven solid tumors (gastric cancer, non-small cell lung cancer) and as a radiosensitizer to enhance radiotherapy efficacy [1][3] 2. Mechanism of action: It competitively binds to c-MET’s ATP pocket, inhibiting c-MET autophosphorylation and downstream PI3K-AKT/ERK pathways. When combined with radiotherapy, it blocks DNA repair in tumor cells, enhancing radiosensitivity [1][3] 3. Metabolism feature: Its major metabolites (M1–M3) show no c-MET inhibitory activity, indicating low off-target metabolic risk [2] 4. Research significance: It validated the feasibility of c-MET inhibitors as radiosensitizers, providing a new strategy for treating radioresistant MET-driven tumors [3]

Solubility Data

Solubility (In Vitro)	DMSO: ~21 mg/mL (~38.9 mM) Water: <1 mg/mL Ethanol: <1 mg/mL
Solubility (In Vivo)	15% Captisol+citrate vehicle: 30 mg/mL  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	1.8533 mL	9.2665 mL	18.5329 mL
	5 mM	0.3707 mL	1.8533 mL	3.7066 mL
	10 mM	0.1853 mL	0.9266 mL	1.8533 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.