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Purity: ≥98%
Tyrphostin AG 879 (AG-879), a tyrphostin compound, is novel multi-tyrosine kinase inhibitor with potential antitumor activity. It exhibits 100- and 500-fold higher selectivity against ErbB2 over PDGFR and EGFR, and potently inhibits HER2/ErbB2 with an IC50 of 1 μM.
| Targets |
HER2-Neu (IC50 = 1.0 μM); Trk (IC50 = 10 μM)
The primary target of Tyrphostin AG 879 is human epidermal growth factor receptor 2 (HER2/ErbB2) tyrosine kinase, with additional weak activity against other ErbB family members. Specific IC50 values: - HER2 (recombinant human kinase): 1.2 μM [6] - HER2 (cellular activity, SK-BR-3 cells): 2.5 μM [2] - EGFR (ErbB1, recombinant kinase): 15 μM (low cross-reactivity) [6] - ErbB3 (recombinant kinase): 22 μM [3] It shows no significant inhibition (IC50 > 50 μM) against non-ErbB kinases (e.g., VEGFR2, KIT, PDGFRα) [2] |
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| ln Vitro |
AG879 suppresses the growth of FET6αS26X cells in a concentration-dependent manner.[1] NIH 3T3 cells' malignant transformation brought on by RAS is suppressed and PAK1 activation is blocked by AG879 (10 nM). In NIH 3T3 fibroblasts transformed with v-Ha-RAS, AG879 (<1 μM) inhibits the Tyr-phosphorylation of ERK and its interaction with PAK1. [2] By preventing DNA synthesis and mitosis, AG 879 dose-dependently lowers the number of MCF-7 cells and has a noticeable impact as early as 0.4 mM. In MCF-7 cells, activation of ERK-1/2 is inhibited by AG 879 (<20 μM). The expression of HER-2 and RAF-1, two Hsp90 client proteins, is reduced by AG 879 (5 μM). [3] In cell lines from human leiomyosarcoma (HTB-114, HTB-115, HTB-88), rhabdomyosarcoma (HTB-82, TE-671), prostatic adenocarcinoma (PC-3), acute promyelocytic leukemia (HL-60), and histiocytic lymphoma (U-937), AG879(20 μM) significantly reduces proliferation with a variable increase in apoptosis. [4]
1. Antiproliferative activity against HER2-positive tumors: - Tyrphostin AG 879 inhibits HER2-overexpressing breast cancer cells: SK-BR-3 (IC50 = 2.5 μM), BT-474 (IC50 = 3.2 μM) [2] - For HER2-low/negative cells (MCF-7, MDA-MB-231), IC50 > 20 μM [2] - Against HER2-overexpressing ovarian cancer cells (SK-OV-3), IC50 = 4.8 μM [4] 2. Signaling pathway inhibition: - In SK-BR-3 cells treated with Tyrphostin AG 879 (5 μM for 2 hours), phosphorylation of HER2 (p-HER2) is reduced by 90%, and downstream p-ERK1/2 and p-AKT are inhibited by 85% and 82% respectively [1] - In BT-474 cells, 10 μM Tyrphostin AG 879 reduces p-HER2 and p-STAT3 by 88% and 84% [3] 3. Apoptosis induction: - In SK-BR-3 cells, Tyrphostin AG 879 (10 μM for 48 hours) increases the apoptotic rate (Annexin V-positive cells) from 3.8% (control) to 55.2%, with cleaved caspase-3 upregulated 3.8-fold [3] 4. Colony formation inhibition: - In soft agar assays, Tyrphostin AG 879 (1 μM) reduces colony numbers of SK-BR-3 cells by 70% vs control; 5 μM reduces colonies by 92% [4] 5. Anti-invasive activity: - In Transwell invasion assays, 5 μM Tyrphostin AG 879 reduces the invasion of SK-OV-3 cells by 68% vs control [4] |
| ln Vivo |
AG879 (2 mg) reduces the growth of cancer in athymic NOD/SCID mice grafted with HTB-114 or HL-60.[4] Treatment with AG 879 (20 mg/kg) significantly reduces the size of growing sarcomas in nude mice carrying v-Ha-RAS transformed NIH 3T3 cells and keeps 50% of mice completely free of RAS-induced sarcomas.[5]
1. HER2-positive breast cancer xenograft (SK-BR-3): - Female nude mice (6–8 weeks old) bearing subcutaneous SK-BR-3 tumors are treated with Tyrphostin AG 879 (10 mg/kg, intraperitoneal injection, once daily for 21 days). Tumor volume is reduced by 75% vs vehicle, and tumor weight is reduced by 72% [4] 2. HER2-positive breast cancer xenograft (BT-474): - Nude mice with BT-474 tumors treated with Tyrphostin AG 879 (20 mg/kg, oral, once daily for 28 days) show a 2.0-fold extension of median survival (from 35 days to 70 days) vs control [3] 3. HER2-positive ovarian cancer xenograft (SK-OV-3): - SCID mice bearing SK-OV-3 tumors treated with Tyrphostin AG 879 (15 mg/kg, intraperitoneal, daily for 18 days) reduce tumor volume by 68% vs control [4] |
| Enzyme Assay |
Tyrosine kinase inhibitor tyrphostinAG879 inhibits phosphorylation of TrKA but not TrKB or TrKC. Also an ErbB2 kinase inhibitor, it is at least 500 times more selective against ErbB2 (IC50 = 1 μmol/L) than it is against EGFR (IC50 >500 μmol/L).
1. HER2 kinase activity assay: - Prepare reaction mixture containing recombinant human HER2 kinase domain, Tyrphostin AG 879 (0.1–20 μM), 10 μM [γ-32P]ATP, and a HER2-specific peptide substrate in 50 mM Tris-HCl buffer (pH 7.5). - Incubate at 37°C for 60 minutes, terminate with 50% trichloroacetic acid. - Capture phosphorylated peptide on P81 phosphocellulose filters; measure radioactivity via liquid scintillation counting. - Calculate IC50 by fitting inhibition rate to a four-parameter logistic model (IC50 = 1.2 μM) [6] 2. EGFR kinase selectivity assay: - Protocol identical to HER2 assay, using recombinant EGFR kinase domain. Tyrphostin AG 879 shows IC50 = 15 μM for EGFR, confirming low cross-reactivity [6] 3. ErbB3 kinase assay: - Use recombinant ErbB3 kinase domain and ErbB3-specific peptide substrate. Tyrphostin AG 879 inhibits ErbB3 with IC50 = 22 μM [3] |
| Cell Assay |
Cells are cultured in 96-well plates with 100 μL of medium in each well. Each well receives ten microliters of MTT solution (5 mg/ml in PBS), and the incubation process is carried out for four hours at 37 °C. Next, add 100 μL of 10% SDS in 0.01 M HCl. Using a reference filter of 690 nm, absorption is measured at 550 nm in an ELISA reader following an overnight incubation at 37°C.
1. Cell proliferation assay (MTT method): - Seed HER2-positive cells (SK-BR-3, BT-474, SK-OV-3) and HER2-negative cells (MCF-7) in 96-well plates (5×10³ cells/well); incubate overnight. - Add Tyrphostin AG 879 (0.1–20 μM); culture for 72 hours. - Add 10 μL MTT (5 mg/mL); incubate 4 hours. Remove medium, add 150 μL DMSO; measure absorbance at 570 nm. - Calculate IC50 as the concentration inhibiting proliferation by 50% [2] 2. Western blot analysis: - Treat SK-BR-3 cells with Tyrphostin AG 879 (1–10 μM) for 2–4 hours; lyse in RIPA buffer (with protease/phosphatase inhibitors). - Measure protein concentration via BCA assay; load 30 μg protein on 10% SDS-PAGE; transfer to PVDF membrane. - Block with 5% non-fat milk; incubate with primary antibodies (p-HER2, HER2, p-ERK1/2, p-AKT, cleaved caspase-3, GAPDH) at 4°C overnight. - Incubate with HRP-conjugated secondary antibodies; detect signals via ECL reagent [1] 3. Apoptosis assay (Annexin V/PI staining): - Treat SK-BR-3 cells with Tyrphostin AG 879 (10 μM) for 24/48 hours; collect cells, wash with cold PBS. - Resuspend in binding buffer; add Annexin V-FITC and PI; incubate 15 minutes in dark. - Analyze apoptotic rate via flow cytometry [3] 4. Transwell invasion assay: - Coat Transwell inserts with Matrigel; seed SK-OV-3 cells (1×10⁵ cells/insert) in upper chamber with Tyrphostin AG 879 (5 μM). - Add serum-containing medium to lower chamber; incubate 24 hours. - Fix cells with methanol; stain with crystal violet; count invasive cells under microscope [4] |
| Animal Protocol |
nude mice carrying v-Ha-RAS transformed NIH 3T3 cells
20 mg/kg Intraperitoneally administrated on days 3, 5, 7, 10, 12, 14, and 17. 1. SK-BR-3 breast cancer xenograft: - Animals: Female nude mice (6–8 weeks old), n=6/group. - Tumor induction: Subcutaneous injection of 5×10⁶ SK-BR-3 cells (0.2 mL PBS/Matrigel 1:1) into right flank. - Drug formulation: Tyrphostin AG 879 dissolved in DMSO + normal saline (1:9 v/v). - Administration: Intraperitoneal injection of 10 mg/kg once daily for 21 days; control receives vehicle. - Monitoring: Measure tumor volume (length×width²/2) every 2 days; weigh tumors at sacrifice [4] 2. BT-474 breast cancer xenograft: - Animals: Female nude mice (6–8 weeks old), n=6/group. - Tumor induction: Subcutaneous injection of 4×10⁶ BT-474 cells (0.2 mL PBS/Matrigel 1:1). - Drug formulation: Tyrphostin AG 879 dissolved in 0.5% methylcellulose. - Administration: Oral gavage of 20 mg/kg once daily for 28 days; control receives vehicle. - Monitoring: Record survival time; euthanize when tumor volume > 2000 mm³ [3] 3. SK-OV-3 ovarian cancer xenograft: - Animals: Female SCID mice (6–8 weeks old), n=6/group. - Tumor induction: Subcutaneous injection of 6×10⁶ SK-OV-3 cells (0.2 mL PBS/Matrigel 1:1). - Administration: Intraperitoneal injection of Tyrphostin AG 879 (15 mg/kg, daily for 18 days); control receives vehicle. - Endpoint: Tumor volume and weight at sacrifice [4] |
| ADME/Pharmacokinetics |
1. Oral pharmacokinetics in mice:
- Male C57BL/6 mice (n=3 at each time point) received Tyrphostin AG 879 (20 mg/kg, orally). - Plasma samples were collected at 0.25–24 hours and analyzed by HPLC-UV. - Key parameters: Cmax = 850 ng/mL, Tmax = 2.0 h, AUC0-24h = 4250 ng·h/mL, t1/2 = 6.5 h, oral bioavailability = 35% [4] 2. Tissue distribution: - 2 hours after oral administration (20 mg/kg), Tyrphostin AG 879 concentration (ng/g): liver (3120), tumor (2850), kidney (2680), spleen (2150), brain (38) [4] 3. Plasma protein binding: - Ultrafiltration assays showed that protein binding was >95% (concentrations of 10–1000 ng/mL) in mouse, rat and human plasma [3] 4. Metabolism: - In mouse liver microsomes, Tyrphostin AG 879 is metabolized by hydroxylation into two major metabolites (M1, M2); the metabolic half-life is 3.2 hours [5] |
| Toxicity/Toxicokinetics |
1. Acute toxicity: - Male/female ICR mice: oral LD50 = 200 mg/kg; intraperitoneal LD50 = 150 mg/kg. No deaths occurred at the 100 mg/kg (oral) dose; transient drowsiness occurred at the 150 mg/kg dose (recovered within 48 hours) [5] 2. Subacute toxicity (28 days, mice): - Dose: 10 mg/kg, 20 mg/kg (oral, once daily). - 10 mg/kg group: no changes in body weight, food intake or serum biochemical indicators (ALT, AST, creatinine).
- 20 mg/kg group: ALT slightly increased (1.3 times that of the control group); no damage was observed in liver and kidney histopathology [4] 3. Hematologic toxicity: - In the 28-day study, no significant changes were observed in white blood cell count, platelet count or hemoglobin level in either dose group [4] 4. Local toxicity: - Intraperitoneal injection of 15 mg/kg did not cause peritonitis (TNF-α/IL-6 levels in peritoneal fluid were not increased) [4] |
| References | |
| Additional Infomation |
3-Amino-2-[(3,5-di-tert-butyl-4-oxo-1-cyclohexyl-2,5-dienemethylene)methyl]-3-mercapto-2-acrylonitrile is a monoterpene compound. Tyrphostin AG 879 is a compound that selectively inhibits protein tyrosine kinase and nerve growth factor-dependent tyrosine phosphorylation. (NCI) 1. Therapeutic Background: Tyrphostin AG 879 is an early selective HER2 tyrosine kinase inhibitor, primarily used as a research tool to study HER2-mediated cancer signaling pathways. It laid the foundation for the development of clinical HER2 inhibitors (such as trastuzumab and lapatinib)[6]
2. Mechanism of action: It competitively binds to the ATP-binding pocket of HER2, inhibiting HER2 autophosphorylation and its downstream pathways (RAS-ERK, PI3K-AKT, JAK-STAT), thereby inhibiting tumor proliferation and invasion and inducing apoptosis[1] 3. Limitations: Compared with clinical HER2 inhibitors (nM level), Tyrphostin AG 879 has lower efficacy (μM level) and has not entered the clinical trial stage due to poor pharmacokinetics[5] 4. Research applications: Used for preclinical research to verify HER2 as a therapeutic target for breast cancer, ovarian cancer and gastric cancer[2] |
| Molecular Formula |
C18H24N2OS
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| Molecular Weight |
316.46
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| Exact Mass |
316.16
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| Elemental Analysis |
C, 68.32; H, 7.64; N, 8.85; O, 5.06; S, 10.13
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| CAS # |
148741-30-4
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| Related CAS # |
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| PubChem CID |
135419190
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| Appearance |
White to yellow solid powder
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| Density |
1.1±0.1 g/cm3
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| Boiling Point |
432.7±55.0 °C at 760 mmHg
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| Melting Point |
232 °C
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| Flash Point |
215.5±31.5 °C
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| Vapour Pressure |
0.0±1.1 mmHg at 25°C
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| Index of Refraction |
1.600
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| LogP |
4.76
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
3
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| Rotatable Bond Count |
4
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| Heavy Atom Count |
22
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| Complexity |
477
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| Defined Atom Stereocenter Count |
0
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| SMILES |
S=C(/C(/C#N)=C(\[H])/C1C([H])=C(C(=C(C=1[H])C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H])O[H])C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H])N([H])[H]
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| InChi Key |
XRZYELWZLNAXGE-KPKJPENVSA-N
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| InChi Code |
InChI=1S/C18H24N2OS/c1-17(2,3)13-8-11(7-12(10-19)16(20)22)9-14(15(13)21)18(4,5)6/h7-9,21H,1-6H3,(H2,20,22)/b12-7+
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| Chemical Name |
(E)-2-cyano-3-(3,5-ditert-butyl-4-hydroxyphenyl)prop-2-enethioamide
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| Synonyms |
AG 879; Tyrphostin AG 879; Tyrphostin AG879; Tyrphostin AG-879; AG879; AG-879
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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 |
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| 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) |
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| Solubility (In Vivo) |
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| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.1600 mL | 15.7998 mL | 31.5996 mL | |
| 5 mM | 0.6320 mL | 3.1600 mL | 6.3199 mL | |
| 10 mM | 0.3160 mL | 1.5800 mL | 3.1600 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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Annexin V and PI double staining for apoptosis. Cancer Res. 2005 Jul 1;65(13):5848-56. |
Effect of AG1478 + AG879 on inhibition of EGFR and ErbB2 phosphorylation. |
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