| Size | Price | Stock | Qty |
|---|---|---|---|
| 50mg |
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| 100mg |
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| Other Sizes |
| Targets |
BRafV600E 38 nM (IC50) BRAFWT 79 nM (IC50) CRAF 68 nM (IC50) p38 6 nM (IC50) CSF1R 35 nM (IC50) MAP3K7 6400 nM (IC50) CSK 7050 nM (IC50)
AZ304 targets BRAF kinase, specifically inhibiting both wild-type BRAF and the V600E mutant form. The compound binds to the ATP-binding site of BRAF, blocking its kinase activity and preventing the phosphorylation of downstream targets in the MAPK/ERK pathway. The IC50 values for BRAF inhibition are 38 nM for the V600E mutant and 79 nM for wild-type BRAF. By inhibiting BRAF, AZ304 reduces the phosphorylation of ERK (p-ERK), a key downstream effector, with mean EC50 values of 65 nM in V600E mutant BRAF melanoma cells (A375) and 52-60 nM in wild-type BRAF melanoma cells (SK-MEL-31). The compound’s activity against both mutant and wild-type BRAF makes it a valuable tool for studying the role of BRAF in cancer and for exploring potential therapeutic applications. |
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| ln Vitro |
AZ304 (1 nM–100 μM) significantly lowers p-ERK phosphorylation; in the V600E mutant BRAF-containing melanoma cell line A375, the mean EC50 was 65 nM, whereas in the wild type BRAF melanoma cell line SK-MEL-31, both with and without EGF, the EC50s were 52 nM and 60 nM[1]. In both BRAF genetic status cell lines, AZ304 also strongly suppresses p-p38[1]. With GI50s of 4.539 μM, 3.896 μM, 4.987 μM, 1.763 μM (48 hours) and 0.5032 μM, 0.3887 μM, 0.6354 μM, 0.3772 μM (72 hours), respectively, AZ304 (0, 0.1, 1, 10, 100 μM, 48 and 72 hours) dose-dependently suppresses the growth of RKO, HT-29, DiFi, and Caco-2[1]. AZ304 (2 μM, 36 and 48 hours) raises p-EGFR in BRAF wild type and BRAF V600E mutant cells while decreasing BRAF, p-ERK, p-AKT, and p-mTOR levels. When combined with C225, AZ304 more effectively reduces the BRAF, ERK, AKT, and mTOR signaling pathways while downregulating p-EGFR and inhibiting p-ERK[1].
AZ304 demonstrates potent in vitro activity against a panel of cancer cell lines. In BRAF V600E mutant cancer cell lines, AZ304 significantly inhibits cell proliferation, with GI50 (50% growth inhibition) values ranging from 0.08 to 7.72 μM. In cell lines harboring wild-type BRAF/RAS or mutant RAS, AZ304 effectively reduces cell growth, with GI50 values ranging from 0.43 to 11.7 μM. In colorectal cancer cell lines with different BRAF mutation statuses, AZ304 dose-dependently inhibits the growth of RKO (V600E mutant), HT-29 (V600E mutant), DiFi (wild-type BRAF), and Caco-2 (wild-type BRAF) cells, with GI50 values of 4.539 μM, 3.896 μM, 4.987 μM, and 1.763 μM at 48 hours, and 0.5032 μM, 0.3887 μM, 0.6354 μM, and 0.3772 μM at 72 hours, respectively. AZ304 potently reduces ERK phosphorylation (p-ERK) in melanoma cell lines. |
| ln Vivo |
AZ304 has been evaluated in xenograft tumor models. In RKO and Caco-2 tumor models, AZ304 demonstrated antitumor activity. The compound was administered orally at a dose of 10 mg/kg twice daily. The treatment resulted in tumor growth inhibition, confirming the in vivo efficacy of AZ304 as a BRAF inhibitor. The compound was well tolerated in these studies, with no overt signs of toxicity reported. These findings support the potential of AZ304 as a therapeutic agent for BRAF-mutant cancers, including melanoma and colorectal cancer.
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| Enzyme Assay |
The kinase inhibitory activity of AZ304 can be assessed using in vitro kinase assays. In a typical assay, recombinant BRAF kinase (wild-type or V600E mutant) is incubated with a peptide substrate, ATP (including radiolabeled ³³P-ATP), and varying concentrations of AZ304. The incorporation of phosphate into the substrate is measured, and the half-maximal inhibitory concentration (IC50) is calculated from dose-response curves. The IC50 values for BRAF inhibition are 38 nM for the V600E mutant and 79 nM for wild-type BRAF. The selectivity of AZ304 against a panel of other kinases can be assessed using kinase profiling services to determine its off-target effects.
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| Cell Assay |
Cell Proliferation Assay[1]
Cell Types: RKO, HT-29, DiFi, Caco-2 cells Tested Concentrations: 0, 0.1, 1, 10, 100 μM Incubation Duration: 48, 72 hrs (hours) Experimental Results: Dose- dependently inhibited the growth of V600E mutant BRAF cell lines (RKO, HT-29) and wild-type BRAF cell lines (DiFi, Caco-2). The cellular activity of AZ304 is assessed using a panel of cancer cell lines with different BRAF mutation statuses. Cells are seeded in 96-well plates and treated with escalating concentrations of AZ304 (0, 0.1, 1, 10, 100 μM) for 48 or 72 hours. Cell viability is measured using MTT assay, and the half-maximal inhibitory concentration (GI50) values are calculated from dose-response curves. ERK phosphorylation (p-ERK) is measured by Western blotting or by ELISA using antibodies specific for phosphorylated ERK. The effects of AZ304 on downstream signaling and cell cycle progression are assessed by flow cytometry and Western blotting. |
| Animal Protocol |
AZ304 has been evaluated in xenograft models using female athymic BALB/c nude mice inoculated with RKO or Caco-2 cells. When tumors reach a palpable size, animals are randomized into treatment and control groups. AZ304 is administered orally at a dose of 10 mg/kg twice daily. The compound is formulated in 0.5% HPMC (hydroxypropyl methylcellulose). Tumor volumes are measured twice weekly using calipers, and tumor growth inhibition (TGI) is calculated. Body weights are monitored as a general indicator of toxicity.
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| ADME/Pharmacokinetics |
The pharmacokinetic properties of AZ304 have been characterized in preclinical studies. The compound is orally bioavailable and can be administered by oral gavage. At a dose of 10 mg/kg twice daily, AZ304 achieves sufficient systemic exposure to inhibit BRAF kinase activity in tumors. The compound’s half-life, maximum concentration (Cmax), area under the curve (AUC), and other pharmacokinetic parameters have not been detailed in the available literature. The formulation in 0.5% HPMC suggests that the compound has reasonable solubility and stability for oral administration.
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| Toxicity/Toxicokinetics |
No detailed toxicity data for AZ304 has been published in the available literature. In the xenograft mouse studies, the compound was reported to be well tolerated at a dose of 10 mg/kg twice daily. However, comprehensive toxicology studies would be required to evaluate the safety profile of AZ304 for potential clinical development. The mechanism of action, which involves inhibition of BRAF and the MAPK/ERK pathway, could lead to skin toxicity, gastrointestinal effects, and other adverse events commonly associated with BRAF inhibitors.
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| References | |
| Additional Infomation |
AZ304 is a potent, selective, and orally active BRAF inhibitor targeting both wild-type BRAF and the V600E mutant form, with IC50 values of 79 nM and 38 nM, respectively. The compound potently reduces ERK phosphorylation (p-ERK) in melanoma cell lines. AZ304 inhibits the proliferation of cancer cell lines with GI50 values ranging from 0.08 to 11.7 μM. In vivo, AZ304 demonstrates antitumor activity in xenograft models at an oral dose of 10 mg/kg twice daily. AZ304 is a valuable research tool for studying BRAF-driven cancers and for exploring potential therapeutic strategies.
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| Molecular Formula |
C27H25N5O2
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|---|---|
| Exact Mass |
451.2
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| Elemental Analysis |
C, 71.82; H, 5.58; N, 15.51; O, 7.09
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| CAS # |
942507-42-8
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| PubChem CID |
16202218
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| Appearance |
White to off-white solid powder
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| LogP |
5.2
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
6
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| Rotatable Bond Count |
6
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| Heavy Atom Count |
34
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| Complexity |
749
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| Defined Atom Stereocenter Count |
0
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| SMILES |
CC1=C(C=C(C=C1)NC(=O)C2=CC(=CC=C2)C(C)(C)C#N)NC3=NC=NC4=C3C=CC(=C4)OC
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| InChi Key |
NGWQZRWVYYFTHC-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C27H25N5O2/c1-17-8-9-20(31-26(33)18-6-5-7-19(12-18)27(2,3)15-28)13-23(17)32-25-22-11-10-21(34-4)14-24(22)29-16-30-25/h5-14,16H,1-4H3,(H,31,33)(H,29,30,32)
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| Chemical Name |
3-(2-cyanopropan-2-yl)-N-(3-((7-methoxyquinazolin-4-yl)amino)-4-methylphenyl)benzamide
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| Synonyms |
AZ304; AZ-304; AZ 304;
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| HS Tariff Code |
2934.99.9001
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| 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)
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| Solubility (In Vitro) |
DMSO : 125 mg/mL (276.84 mM)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (4.61 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 (4.61 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. View More
Solubility in Formulation 3: ≥ 2.08 mg/mL (4.61 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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