Aliertib 是一种口服活性和选择性的AuRORa A激酶抑制剂，IC50值为 1.2 nM。它通过靶向白血病细胞中的Akt/mTOR/AMPK/p38途径诱导其凋亡和自噬，具有抗肿瘤活性。

Alisertib (MLN8237) is a specific Aurora A inhibitor (IC50: 1.2 nM). The selectivity of Alisertib is >200-fold higher for Aurora A than Aurora B.

Treatment of cultured MM cells with Alisertib (MLN8237) results in mitotic spindle abnormalities, mitotic accumulation, as well as inhibition of cell proliferation through apoptosis and senescence. In addition, MLN8237 up-regulates p53 and tumor suppressor genes p21 and p27. Combining MLN8237 with dexamethasone, doxorubicin, or bortezomib induces synergistic/additive anti-MM activity in vitro [1]. Alisertib inhibited AAK over ABK with a selectivity of more than 200-fold in cells. Alisertib inhibited proliferation of human tumor cell lines in vitro [2]. A T217D/W277E double mutant exhibits superior levels of resistance to MLN8237, with the I50 value increasing approximately 20-fold from 30 to 650 nM in the presence of TPX2 [3].

Tumor burden was significantly reduced and overall survival was significantly increased in animals treated with 30 mg/kg MLN8237 for 21 days. Induction of apoptosis and cell death by MLN8237 were confirmed in tumor cells excised from treated animals by TdT-mediated dUTP nick end labeling assay [1]. Alisertib produced a dose-dependent decrease in bipolar and aligned chromosomes in the HCT-116 xenograft model. Alisertib produced tumor growth inhibition in solid tumor xenograft models and regressions in in vivo lymphoma models. In addition, a dose of alisertib that caused tumor stasis, as measured by volume, resulted in a decrease in FLT uptake [2]. Nude mice bearing human tumor xenografts treated with a single oral dose of alisertib (20 mg/kg) displayed a phenotype consistent with inhibition of Aurora A [4].

Recombinant murine Aurora A and Aurora B protein were expressed in Sf9 cells and purified with GST affinity chromatography. The peptide substrate for Aurora A was conjugated with biotin (Biotin-GLRRASLG). Aurora A kinase (5 nM) was assayed in 50 mM Hepes (pH 7.5)/10 mM MgCl2/5 mM DTT/0.05% Tween 20/2 μM peptide substrate/3.3 μCi/ml [γ-33P]ATP at 2 μM by using Image FlashPlates. Aurora B kinase (2 nM) was assayed with 10 μM biotinylated peptide Biotin-TKQTARKSTGGKAPR in 50 mM Tricine (pH 8.0)/2.5 mM MgCl2/5 mM DTT/10% glycerol/2% BSA/40 μCi/ml [γ-33P]ATP at 250 μM. The conditions for all other in vitro kinase assays are available upon request. MLN8054 was run in a 226 kinase screen at a 1 μM compound concentration with an ATP concentration of 10 μM for all assays [2].

HCT-116 colorectal carcinoma cells were plated on 6-well dishes (2 × 10^5 per well) and propagated in McCoy's 5A media supplemented with 10% FBS. After 18 hours, alisertib at a final concentration of 0.050, 0.250, or 1.000 μmol/L was added, and the cells were grown for an additional 24 hours. Cells treated with dimethyl sulfoxide (DMSO; 0.2%) served as the untreated vehicle control. The cells were harvested with trypsin EDTA 1×, washed once with PBS, fixed in 70% ethanol, and stored at 4°C for 1 hour. The cells were resuspended in propidium iodide (1:40) and RNAse A (1:5,000) in PBS for 30 minutes at 4°C. Cell-cycle distributions were determined by measuring DNA content using flow cytometry, and samples were analyzed using Winlist 5.0 software [2].

Mice were irradiated (200 cGy), and then 5 × 106 MM1.S cells were inoculated subcutaneously in the right flank. When tumor growth was measurable (～ 2 weeks after the injection), mice were assigned into 4 groups (10 mice each) receiving vehicle orally (100 μL of 10% 2-hydroxypropyl-β-cyclodextrin/1% sodium bicarbonate) or MLN8237 (7.5 mg/kg, 15 mg/kg, and 30 mg/kg in a final formulation in 10% 2-hydroxypropyl-β-cyclodextrin/1% sodium bicarbonate) for 21 consecutive days. The maximal tolerated dose of MLN8237 in most mouse strains (continuous dosing for 21 days) is approximately 20 mg/kg twice a day (40 mg/kg per day). Tumor volumes were measured by a Vernier caliper every alternate day and calculated using the following formula: length × width2 × 0.5. Tumor growth inhibition (TGI) was calculated using the formula (Δcontrol average volume ? Δtreated average volume) × 100/(Δcontrol average volume). Mice were killed at the end of the treatment, 2 hours after the last treatment, or when tumor reached 2 cm^3; tumors were immediately collected from mice and evaluated for induction of apoptosis and cell death by TdT-mediated dUTP nick end labeling (TUNEL) assay [1].

1028486-01-2

C27H20ClFN4O4

518.92

MLN 8237;4-[[9-氯-7-(2-氟-6-甲氧基苯基)-5H-嘧啶并[5,4-D][2]苯并氮杂卓-2-基]氨基]-2-甲氧基苯甲酸

DMSO:26 mg/mL (50.1 mM)
H2O:<1 mgml
Ethanol:<1 mgml
Powder: -20°C for 3 years
In solvent: -80°C for 2 years