WYE-354(IC50=5 nM) is an effective, selective and ATP-competitive mTOR inhibitor. It blocks mTORC2/P-AKT(S473) and mTORC1/P-S6K(T389), not P-AKT(T308). The selectivity for mTOR is higher than PI3Kα (>100-fold) and PI3Kγ (>500-fold).

PI3Kα:1.89 μM, PI3Kγ:7.37 μM, mTOR:5 nM

在HEK293细胞中，WYE-354（0.2 μM–5 μM）有效抑制了mTORC1和mTORC2。在U87 mg和MDA361细胞中，WYE-354（0.3 μM–10 μM）显著阻断了mTOR信号传导和Akt激活。在包括MDA-MB-361、MDA-MB-231、MDA-MB-468、LNCap、A498和HCT116在内的肿瘤细胞系中，WYE-354有效抑制了增殖，IC50值范围为0.28 μM至2.3 μM。在内皮HUVEC细胞中，WYE-354也抑制了mTORC1和mTORC2信号传导，IC50值从10 nM到1 μM不等，这一效果通过S6核糖体蛋白和Akt的去磷酸化得到验证。WYE-354引起的凋亡伴随着G1期细胞周期阻滞和caspases激活。WYE-354（10 nM–1 μM）激活了丝裂原激活蛋白激酶（MAPK）信号传导，这可能是由于其对mTORC1的抑制作用。

WYE-354 (50 mg/kg) 在PTEN-null PC3 mM2肿瘤的小鼠异种移植模型中，强效抑制mTOR信号传导和肿瘤生长。

The assays were incubated at room temperature for 2 hours. In 96-well plates (25 μL) contains 6 nM Flag-TOR(3.5) (estimated 5-10% purity), 1 μM His6-S6K and 100 μM ATP.For inhibitor versus ATP matrix competition, mTOR kinase reactions are carried out with varying concentrations of ATP (0, 25, 50 100, 200, 400 and 800 μM) in combination with varying concentrations of inhibitor.The assays are performed and detected by DELFIA employing the Euphospho-p70S6K T389 antibody.

Tumor cell lines including MDA-MB-361, MDA-MB-231, MDA-MB-468, LNCap, DU145, A498, and HCT116, are plated in 96-well plates at 1000 to 3000 cells per well for 24 hours, treated with DMSO or varying concentrations of WYE-354(0–50 μM, dissolved in DMSO) for72 hours.

Nude mice (BALB/c, nu/nu, female) bearing PC3 mM2 xenograft were intraperitoneally injected with WYE-354（50 mg/kg）dissolved in 5% ethanol, 5% polysorbate 80, 5% PEG-400.