| 名稱 | MK-5108 |
| 描述 | MK-5108 (VX-689) is a highly potent and specific Aurora-A kinase inhibitor with an IC50 value of 0.064 nM. |
| 細(xì)胞實(shí)驗(yàn) | HeLa-S3 cells are synchronized at the G1-S phase boundary by double thymidine block with 2 mM thymidine. Cells are washed and seeded to 96-well cell culture plates. After 4 hours, an equal volume of medium containing MK-5108 is added to each well. Nocodazole (300 nM) is used as a 100% control. The cells are fixed overnight with cold methanol 12 hours after seeding. Then, the cells are stained with rabbit anti-phospho-histone H3 Ser28 antibody and then with anti-rabbit IgG-Cy5. Total nuclei are stained with 10 mg/mL 4′,6-diamidino-2-phenylindole. Immunostained images are acquired using the IN Cell Analyzer1000 with ×10 objective lens. After acquisition of images, data are analyzed. The %pHH3-positive index is determined by measuring the %pHH3-positive cell counts per total nuclei counts for each sample, then by normalizing with respect to nocodazole-treated cells. (Only for Reference) |
| 激酶實(shí)驗(yàn) | Biochemical kinase assays: Recombinant His-tagged human Aurora-A protein is expressed in Escherichia coli and is purified with HisTrap HP column. Purified recombinant human Aurora-B and Aurora-C protein are purchased. Experiments are done in quintuplicate in 96-well plates. The Aurora-A assay reaction is conducted in the presence of 20 μM ATP, 25 μM Tetra-Kemptide [RRR(GLRRASLG)4R-NH2], 1.0 μCi per well [γ-33P]-ATP, 0.1 ng per well Aurora-A in 50 mM Tris-HCl (pH 7.4), 15 mM Mg(OAc)2, and 0.2 mM EDTA at 30°C for 40 minutes. To investigate the inhibition mode of MK-5108 for Aurora-A, the IC50 values of MK-5108 are determined in the presence of different concentrations of ATP. Then, the IC50 value is plotted as a function of ATP concentration to analyze the effect of ATP concentration on the IC50 value of MK-5108. The Aurora-B assay reaction is conducted in the presence of 15 μM ATP, 100 μM Kemptide (GLRRASLG-NH2), 1.0 μCi per well [γ-33P]-ATP, 5.0 ng per well Aurora-B in 50 mM Tris-HCl (pH 7.4), 15 mM Mg(OAc)2, and 0.2 mM EDTA at 30 °C for 20 minuts. The Aurora-C assay reaction is conducted in the presence of 40 μM ATP, 100 μM Kemptide, 1.0 μCi per well [γ-33P]-ATP, 15 ng per well Aurora-C in 10 mM MOPS-NaOH (pH 7.4), 5 mM Mg(OAc)2, 1 mM (±) DTT, and 1 mM EGTA at 30 °C for 20 minutes. After kinase reactions are terminated by adding 2.0% phosphoric acid, Tetra-Kemptide or Kemptide is trapped on the MultiScreen-PH plate. Wells are washed five times with 0.64% phosphoric acid and then monitored for radioactivity in a liquid scintillation counter. |
| 體外活性 | MK-5108以ATP競(jìng)爭性方式抑制Aurora-A活性，對(duì)其他家族激酶Aurora-B（220倍）和Aurora-C（190倍）顯示出強(qiáng)大的選擇性。MK-5108對(duì)Aurora-A與其他蛋白激酶的選擇性也很高，僅對(duì)一種激酶（TrkA）顯示出<100倍的選擇性，可能比MLN8054對(duì)Aurora-A的選擇性更高。MK-5108誘導(dǎo)pHH3陽性細(xì)胞的產(chǎn)生，與G2-M期細(xì)胞堆積一致。在生化檢測(cè)中，MK-5108抑制包括HCC1143、AU565、MCF-7、HCC1806和CAL85-1在內(nèi)的腫瘤細(xì)胞增殖，其IC50分別為0.42μM、0.45μM、0.52μM、0.56μM和0.74μM。MK-5108以劑量依賴性方式降低包括LEIO285、LEIO505和SK-LSM1細(xì)胞線在內(nèi)的所有三種細(xì)胞線的細(xì)胞活性，IC50約為100nM。LEIO285細(xì)胞在與MK-5108孵育后，48小時(shí)和72小時(shí)后G2/M期細(xì)胞比例增加。與DMSO處理的對(duì)照組相比，MK-5108顯著提高Caspase 3/7活性。在LEIO505細(xì)胞中，MK-5108導(dǎo)致更多細(xì)胞在24小時(shí)累積在G2/M期，但在48小時(shí)或72小時(shí)沒有此現(xiàn)象。MK-5108在M期阻滯ULMS細(xì)胞線，降低LEIO285細(xì)胞中g(shù)emcitabine的IC50值，但在LEIO505和SK-LMS1細(xì)胞中升高。 |
| 體內(nèi)活性 | MK-5108在16 mg/kg和32 mg/kg劑量下誘導(dǎo)pHH3陽性細(xì)胞。MK-5108在8 mg/kg和16 mg/kg劑量下的血漿濃度分別是1.7 μM和4.4 μM。MK-5108處理在腫瘤和皮膚組織中誘導(dǎo)pHH3的表達(dá)，從2小時(shí)開始，4小時(shí)達(dá)到最大。以15 mg/kg和30 mg/kg劑量進(jìn)行的MK-5108處理顯著抑制腫瘤生長，治療組平均腫瘤體積變化百分比相對(duì)于對(duì)照組（%T/C）在第11天分別為10%和-6%，而在第18天分別為17%和5%。MK-5108在這兩個(gè)劑量下耐受性良好，體重輕微減少。MK-5108還通過間歇給藥在裸鼠負(fù)載SW48腫瘤模型中展現(xiàn)顯著的抗腫瘤活性，15 mg/kg和45 mg/kg劑量的MK-5108引起劑量依賴性的腫瘤生長抑制，第10天的%T/C分別為35%和7%，第27天分別為58%和32%。[1] |
| 存儲(chǔ)條件 | Powder: -20°C for 3 years | In solvent: -80°C for 1 year | Shipping with blue ice. |
| 溶解度 | Ethanol : < 1 mg/mL (insoluble or slightly soluble)
H2O : < 1 mg/mL (insoluble or slightly soluble)
DMSO : 85 mg/mL (184 mM)
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| 關(guān)鍵字 | Aurora Kinase | inhibit | Autophagy | MK-5108 | VX 689 | MK 5108 | VX689 | Inhibitor |
| 相關(guān)產(chǎn)品 | Guanidine hydrochloride | Naringin | Valproic Acid | Taurine | Gefitinib | Aceglutamide | Hydroxychloroquine | Curcumin | Stavudine | Salicylic acid | Paeonol | Sodium 4-phenylbutyrate |
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