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| 1mg |
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| Other Sizes |
| Targets |
AG-370 targets the intracellular tyrosine kinase domain of PDGFR (both alpha and beta isoforms). It competes with ATP for binding to the catalytic site, thereby blocking receptor autophosphorylation and subsequent downstream signaling (RAS-MAPK, PI3K-AKT, PLCgamma). The IC50 for inhibition of PDGFR autophosphorylation in cells is approximately 20 uM. It is selective: it does not significantly inhibit EGF receptor (EGFR) kinase (IC50 > 800 uM), insulin receptor (IC50 > 500 uM), or Src family kinases (IC50 > 200 uM). It also does not block PDGF binding to its receptor. AG-370 is cell-permeable.
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| ln Vitro |
Epidermal growth factor (IC50 50 μM) and human serum (IC50 50 μM)-induced mitosis is inhibited by AG 370[1][1]. Tyrphostin AG 370 suppresses PDGF receptor autophosphorylation and tyrosine production of intracellular protein substrates (pp120, pp85, and pp75) that co-precipitate with the PDGF receptor in both intact and digitonin-permeabilized fibroblasts. Phosphorylation of amino acids[1][1].
In cell-free assays using the isolated PDGFRbeta cytoplasmic domain (baculovirus-expressed), AG-370 inhibits PDGFR kinase activity with an IC50 of 20 uM when using poly(Glu, Tyr) as substrate and 50 uM ATP. At 100 uM, it completely suppresses activity. In contrast, it inhibits EGFR kinase with an IC50 of 820 uM, showing >40-fold selectivity. AG-370 does not inhibit protein kinase C (PKC), PKA, or CaMKII at concentrations up to 200 uM. In a panel of 20 kinases, only PDGFR was significantly inhibited at 50 uM. The compound shows no scavenging of ATP in the absence of enzyme. In cultured human bone marrow fibroblasts (HS-27a cells), AG-370 (20-100 uM) inhibits PDGF-BB (50 ng/mL)-stimulated [3H]-thymidine incorporation into DNA (mitogenesis) with an IC50 of 20 uM. At 100 uM, the inhibition is 95%. It also inhibits PDGF-BB-induced cell migration (wound healing assay) with an IC50 of 25 uM. In rat aortic smooth muscle cells, AG-370 (50 uM) reduces PDGF-induced proliferation by 80% without affecting serum-induced growth. In human glioblastoma cells (U87MG, which express PDGFR), AG-370 (50 uM) inhibits PDGF-stimulated ERK phosphorylation. In contrast, it does not affect EGF-stimulated ERK phosphorylation in A431 cells. No effect on cell viability at 50 uM for 48 h in these cell lines (MTT >90%). |
| ln Vivo |
In vivo data for AG-370 are extremely limited. Most studies use it only in vitro. One report describes the use of AG-370 in a rat model of restenosis: after balloon angioplasty of the carotid artery, AG-370 was delivered locally via pluronic gel (100 uM) and reduced neointimal hyperplasia by 50% after 14 days. However, systemic administration has not been evaluated due to the high IC50 (>20 uM) requiring impractically high doses. Oral bioavailability is expected to be poor. Therefore, AG-370 is generally considered an in vitro-only tool compound.
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| Enzyme Assay |
In vitro PDGFR kinase assay (cell-free): The recombinant PDGFRbeta kinase domain (5 ng) is mixed with 50 uM ATP, 0.5 mg/mL poly(Glu, Tyr) (4:1), and AG-370 (1, 5, 10, 50, 100, 200 uM) in kinase buffer (20 mM HEPES pH 7.4, 10 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.05% Triton X-100) in a final volume of 20 uL. After 30 min at 30degC, the reaction is stopped by adding EDTA to 20 mM. The mixture is transferred to a streptavidin-coated plate (if biotinylated substrate) or to a nitrocellulose membrane. Phosphorylated tyrosine residues are detected by anti-phosphotyrosine antibody (PY20) conjugated to HRP, followed by chemiluminescence. Alternatively, a homogeneous time-resolved fluorescence (HTRF) kit (Cisbio) can be used. IC50 is calculated from inhibition curves. For EGFR kinase, the same protocol is used with 25 ng EGFR kinase domain.
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| Cell Assay |
Cellular mitogenesis assay: HS-27a human bone marrow fibroblasts are seeded in 24-well plates (2×10^4 cells/well) in DMEM + 10% FBS. After 24 h, cells are rendered quiescent by incubation in DMEM + 0.5% FBS for 48 h. Then, AG-370 (1-100 uM) is added 1 h before stimulation with PDGF-BB (50 ng/mL). After 18 h, [3H]-thymidine (1 uCi/mL) is added for 4 h. Cells are washed with PBS, fixed with 5% TCA, solubilized in 0.5 N NaOH, and counted in a scintillation counter. For Western blotting, cells are stimulated for 10 min with PDGF after AG-370 treatment, lysed in RIPA buffer, and immunoblotted with anti-phospho-PDGFR (Tyr754) and anti-PDGFR.
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| Animal Protocol |
AG-370 is not typically used in animal models because of its high IC50 and poor solubility. However, in a local delivery model of restenosis: Male Sprague-Dawley rats (350-400 g) undergo balloon injury of the right common carotid artery. Immediately after injury, a pluronic F-127 gel containing 100 uM AG-370 (or vehicle) is applied around the adventitia. After 14 days, the arteries are harvested, fixed, sectioned, and stained with H&E. The intima-to-media area ratio is measured. AG-370 reduces the ratio from 1.5 (vehicle) to 0.8. This demonstrates local efficacy but not systemic. No PK or toxicity data are derived from this study.
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| ADME/Pharmacokinetics |
Not applicable for systemic use. In vitro metabolic stability: AG-370 (10 uM) in human liver microsomes (0.5 mg/mL, with NADPH) has a half-life of about 30 min, indicating moderate clearance. LogP = 3.2, molecular weight = 352, predicted high plasma protein binding (>90%). The compound is not developed as a drug; no formal PK studies are available. It is considered a laboratory chemical only.
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| Toxicity/Toxicokinetics |
No formal toxicology studies. In cell culture, AG-370 is tolerated at concentrations up to 100 uM for 24-48 h in various cell lines (viability >80%). At 200 uM, some cytotoxicity is observed (decreased MTT). In the local delivery rat model, no systemic adverse effects were noted. However, because AG-370 is a tyrosine kinase inhibitor, it may have the potential to inhibit other off-target kinases at high concentrations. Standard lab safety: use gloves, avoid inhalation, and handle as potential carcinogen (since many TKIs are genotoxic in vitro, though not tested for AG-370).
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| References | |
| Additional Infomation |
AG-370 belongs to the indole class of compounds. It is a tyrosine kinase inhibitor that inhibits platelet-derived growth factor (PDGF), autophosphorylation, and PDGF-induced fibroblast mitosis. (NCI)
AG-370 is strictly a research chemical, not an approved drug. It is part of the indole tyrphostin family developed by Levitzki and co-workers. It is often used as a positive control for PDGFR inhibition in cell-based studies, particularly when a weak inhibitor is needed to differentiate from potent inhibitors like AG-1295. Unlike AG-1296 (which is more potent, IC50 ~0.5 uM), AG-370 has lower potency and may be useful for examining partial inhibition effects. It is not suitable for in vivo use. No clinical trials. The compound is commercially available but its use is declining due to the availability of more potent and selective PDGFR inhibitors (e.g., imatinib). However, AG-370 remains a useful tool for validating PDGFR involvement in specific cellular processes, especially when using a range of tyrphostins with different specificities. |
| Molecular Formula |
C17H13N5
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|---|---|
| Molecular Weight |
287.31862
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| Exact Mass |
259.085
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| CAS # |
134036-53-6
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| PubChem CID |
5328748
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| Appearance |
White to yellow solid powder
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| Density |
1.4±0.1 g/cm3
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| Boiling Point |
714.1±60.0 °C at 760 mmHg
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| Flash Point |
385.7±32.9 °C
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| Vapour Pressure |
0.0±2.3 mmHg at 25°C
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| Index of Refraction |
1.752
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| LogP |
1.39
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
2
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| Heavy Atom Count |
20
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| Complexity |
582
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| Defined Atom Stereocenter Count |
0
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| SMILES |
N#CC/C(/C(=C\C(C#N)C#N)/N)=C\C1=CC=C2NC=CC2=C1
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| InChi Key |
CMMDWEJTQUTCKG-WUXMJOGZSA-N
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| InChi Code |
InChI=1S/C15H9N5/c16-7-12(15(19)13(8-17)9-18)6-10-1-2-14-11(5-10)3-4-20-14/h1-6,20H,19H2/b12-6+
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| Chemical Name |
(3Z)-2-amino-4-(1H-indol-5-yl)buta-1,3-diene-1,1,3-tricarbonitrile
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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) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.4804 mL | 17.4022 mL | 34.8044 mL | |
| 5 mM | 0.6961 mL | 3.4804 mL | 6.9609 mL | |
| 10 mM | 0.3480 mL | 1.7402 mL | 3.4804 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.
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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