NVP-AEW541

In vitro kinase assays

NVP-AEW541 was dissolved in DMSO (10 mM) and stored at -20°C. Dilutions were freshly made in DMSO/water 1:1. The final concentration of DMSO in the enzyme assays was < 0.5 %. The protein kinase assays were carried out in 96-well plates at RT and terminated by the addition of 20 μL of 125 mM EDTA. Subsequently, 30 μL (c-Abl, c-Src, IGF-1R) of the reaction mixture were transferred onto Immobilon-PVDF presoaked for 5 mins with methanol, rinsed with water, then soaked for 5 mins with 0.5 % H3PO4 and mounted on vacuum manifold. After spotting all samples, vacuum was connected and each well rinsed with 200 μL 0.5 % H3PO4. Membranes were removed and washed 4 times on a shaker with 1.0 % H3PO4, once with ethanol. After drying, mounting in Packard TopCount 96-well frame, and adding of 10 μL/well of Microscint, membranes were counted. The IC50 values were calculated by linear regression analysis of the percentage inhibition of NVP-AEW541 in duplicate, at 4 concentrations (usually 0.01, 0.1, 1, and 10 μM). One unit of protein kinase activity was defined as 1 nmol of 33P transferred from [γ33P]ATP to the substrate protein per minute per mg of protein at 37°C.

Cell lines

MCF-7 cells

Preparation method

The solubility of this compound in DMSO is >10 mM. General tips for obtaining a higher concentration: Please warm the tube at 37 ℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months.

Reaction Conditions

~ 10 μM; 72 hrs

Applications

In MCF-7 cells, NVP-AEW541 suppressed the IGF-I-mediated survival, soft agar and cell proliferation with IC50 values of 0.162 μM, 0.105 μM and 1.64 μM, respectively.

Animal models

Female Harlan athymic nude mice with NWT-21 cells

Dosage form

20, 30, or 50 mg/kg; p.o.; twice daily, 7 days/week

Applications

NVP-AEW541 dose-dependently inhibited tumor growth with T/C values of 32%, 28% and 14% at the doses of 20 mg, 30 mg, or 50 mg, respectively. NVP-AEW541 was well tolerated at the indicated doses, and the recorded variations in body weight were not statistically significant.

Other notes

Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal.

References:

[1]. García-Echeverría C, Pearson MA, Marti A, et al. In vivo antitumor activity of NVP-AEW541-A novel, potent, and selective inhibitor of the IGF-IR kinase. Cancer Cell, 2004, 5(3): 231-239.