AG-1478

Cell lines

Human breast cancer cell lines MDA-MB-231 and MCF-7

Preparation Method

The human breast cancer cell lines MDA-MB-231 and MCF-7 were obtained from American Type Culture Collection. Cells were grow in RPMI-1640 supplemented with 10% FBS, 100U/ml penicillin and 100μg/ml streptomycin, in a humidified atmosphere of 5% CO₂ at 37˚C. AG-1478 was dissolved in DMSO for preparation of 10mM stock. For the experiments, the tyrphostin was diluted with RPMI-1640 containing 10% FBS. MDA-MB-231 and MCF-7 cells were treated with serial concentrations of Tyrphostin AG-1478 (5-40μM) for 72h. DMSO (0.1%) was used as a control. The antiproliferative effect was evaluated by MTT assay. ApoDETECT™ Annexin V-FITC Kit and flow cytometry were used for apoptosis assessment.

Reaction Conditions

5-40μM; 72h

Applications

AG-1478 suppressed cellular growth by inhibiting cell proliferation, inducing apoptosis.

Animal models

Male C57BL/6 mice and Male ApoE^−/− mice

Preparation Method

Male C57BL/6 mice and Male ApoE^−/− mice weighing 18–20g aged 8 weeks were housed at a constant room temperature with a 12:12h light-dark cycle and fed with a standard rodent diet and water. After an acclimatization period of one week, 28 C57BL/6 or ApoE^−/− mice were randomly divided into four weight-matched groups. 7 mice were fed with standard animal low-fat diet containing 10kcal.% fat, 20kcal.% protein and 70kcal.% carbohydrate served as a normal control group (Control or ApoE-LF), while the remaining 21 mice were fed with high-fat diet containing 60kcal.% fat, 20kcal.% protein and 20kcal.% carbohydrate for 16 weeks(HFD). Since 9th week, AG-1478 was given daily by oral gavage at a dose of 10mg/kg/day for the next 8 weeks. Mice in the Control and HFD groups were gavaged with vehicle (1% CMC-Na solution) only. At the day before the sacrifice of ApoE^−/− mice, doppler analysis was performed to determine the pathologic cardiac hypertrophy. Mice was anesthetized with isoflurane, echocardiography was performed by SONOS 5500 ultrasound with a 15-MHz linear array ultrasound transducer. At the end of experimental period, all the animals were sacrificed by cervical decapitation. The body weight was recorded. Blood samples were collected and centrifuged at 4°C at 3000rpm for 10min for collecting the serum. The heart was excised aseptically, blotted dry and the weight was recorded followed by immediate freezing in liquid nitrogen and then stored at −80°C before further analyses.

Dosage form

10mg/kg/day ; p.o.; 8 weeks

Applications

AG-1478 significantly blocked high-fat diet-induced cardiac EGFR phosphorylation, and reduced myocardial inflammation, fibrosis, apoptosis, and dysfunction without affecting plasma LDL and TG levels.

References:
[1] Zhang, Y. G., Du, Q., Fang, W. G., Jin, M. L., & Tian, X. X. (2008). Tyrphostin AG1478 suppresses proliferation and invasion of human breast cancer cells. International journal of oncology, 33(3), 595–602.
[2] Li, W., Fang, Q., Zhong, P., Chen, L., Wang, L., Zhang, Y., Wang, J., Li, X., Wang, Y., Wang, J., & Liang, G. (2016). EGFR Inhibition Blocks Palmitic Acid-induced inflammation in cardiomyocytes and Prevents Hyperlipidemia-induced Cardiac Injury in Mice. Scientific reports, 6, 24580.