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生物活性
产品描述
A-674563是一种Akt抑制剂，作用于Akt1，Ki为11 nM，对PKA适度有效，作用于Akt1比 作用于PKC选择性高20倍以上。
靶点
Akt1
PKA
CDK2
 
 
 
IC50
11 nM (Ki)
16 nM (Ki)
46 nM (Ki) [1]
 
 
 
体外研究
A-674563 is achieved from A-443654 by replacing the indole with a phenyl moiety and getting oral activity. A-674563 slows proliferation of tumor cells with EC50 of 0.4 μM. [1] A-674563 does not inhibit Akt phosphorylation per se, but blocks the phosphorylation of Akt downstream targets in a dose-dependent manner. A-674563 induced Akt blockade results in decreased STS cell downstream target phosphorylation and tumor cell growth inhibiton. A-674563 induces G2 cell cycle arrest and apoptosis in STS cells. [2]
体内研究
20 mg/kg A-674563 increases plasma insulin in an oral glucose tolerance test. A-674563 shows no significant monotherapy tumor inhibitory activity; the efficacy of the combination therapy is significantly improved compared to paclitaxel monotherapy. [1] A674563-treated (20 mg/kg/bid, p.o.) mice exhibits slower tumor growth and more than 50% decrease in the tumor volume at the termination of the study compared with that in control group. [2] A-674563 is identified to have drastically improved PK profile with oral bioavailability of 67% in mouse, but is 70-fold less active than A-443654. [3]
特征
 
推荐的实验操作(此推荐来自于公开的文献所以Selleck并不保证其有效性)
激酶实验: [1]
Akt Kinase Assay
The kinase assay uses His-Akt1 and a biotinylated mouse Bad peptide as substrate. The kinase assay is carried out at room temperature for 30 minutes in 50 μL of reaction buffer [20 mM HEPES, pH 7.5, 10 mM MgCl2, 0.1% (w/v) Triton X-100, 5 μM ATP (Km = 40 μM), 5 μM peptide (Km = 15 μM), 1 mM DTT, 60 ng of Akt1, and 0.5 μCi of [γ-33P]ATP] in the presence of different concentrations of A-674563. Each reaction is stopped by adding 50 μL of termination buffer (0.1 M EDTA, pH 8.0, and 4 M NaCl). The biotinylated Bad peptides are immobilized on streptavidin-coated FLASH plates. After being washed with PBS-Tween 20 (0.05%), the 33P phosphopeptide captured on the FLASH plates is measured with a TopCount Packard Instruments γ counter.
细胞试验: [1]
细胞系
MiaPaCa-2 cells
浓度
0-30 μM
处理时间
48 hours
方法
The cells on 96-well plates are gently washed with 200 μL of PBS. Alamar Blue reagent is diluted 1:10 in normal growth media. The diluted Alamar Blue reagent (100 M) is added to each well on the 96-well plates and incubated until the reaction is complete as per manufacturers instructions. Analysis is done using an fmax Fluorescence Microplate Reader, set at the excitation wavelength of 544 nm and emission wavelength of 595 nm. Data are analyzed using SOFTmax PRO software provided by the manufacturer.
动物实验: [1]
动物模型
Fasted mice
配制
Dissolved in DMSO.
剂量
20 or 100 mg/kg
给药处理
A-674563 is administered 30 minutes before the 1 g/kg glucose challenge.
溶解度
Saline, 30 mg/mL
参考文献
[1] Luo Y, et al, Mol Cancer Ther, 2005, 4(6), 977-986.
[2] Zhu QS, et al, Cancer Res, 2008, 68(8), 2895-2903.
[3] Tatsuya Okuzumi, et al, Mol Biosyst, 2010, 6(8), 1389-1402.