| 名稱 | Sotrastaurin |
| 描述 | Sotrastaurin (AEB071) is a potent pan-PKC inhibitor (Kis: 0.95/0.64/2.1/3.2/1.8/0.22 nM for PKCα/βI/δ/ε/η/θ). |
| 細胞實驗 | Jurkat cells (5×10^6 cells) were pretreated for 4 h with 500 nM AEB071 and loaded for 30 min at 37°C in the dark with 5 μM fura-2 acetoxymethyl ester. Dye excess was removed by washing in Hanks' balanced salt solution. Samples were prewarmed to 37°C and baseline Ca2+ levels were determined for 100 s on a Spex Fluorolog 2 spectrofluorometer equipped with two excitation monochrometers and a Cooper system. At this point, anti-CD3 antibody was added to a final concentration of 10 μg/ml, and data were collected over 6.5 min. The maximal and minimal Ca2 levels were determined by adding an excess of ionomycin and EGTA. Experiments were performed at least four times with similar outcomes [1]. |
| 激酶實驗 | Classical and novel PKC isotypes were assayed by scintillation proximity assay technology. In brief, the assay was performed in 20 mM Tris-HCl buffer, pH 7.4, and 0.1% bovine serum albumin by incubating 1.5 μM of the peptide substrate with 10 μM [33P]ATP, 10 mM Mg(NO3)2, 0.2 mM CaCl2, and PKC at a protein concentration varying from 25 to 400 ng/ml, and lipid vesicles containing 30 mol% phosphatidylserine, 5 mol% diacylglycerol (DAG), and 65 mol% phosphatidylcholine at a final lipid concentration of 0.5 μM. Incubation was performed for 60 min at room temperature. The reaction was stopped by adding 50 μl of a mixture containing 100 mM EDTA, 200 μM ATP, 0.1% Triton X-100, and 0.375 μg/well streptavidin-coated scintillation proximity assay beads in PBS without Ca2+ and Mg2+. Incorporated radioactivity was measured in a MicroBetaTrilux counter for 1 min. In situ Thr-219 autophosphorylation status analysis of PKC was done by a phospho-site-specific antibody [1]. |
| 動物實驗 | 6–8 week nu/nu SCID female mice bearing subcutaneously injected 92.1 tumors (7 mice/group) of 100mm3 diameter were treated with vehicle, AEB071 (80mg/kg/d) TID and or BYL719 orally (50mg/kg/d) QD as single agents and in combination, 5 days/week for 2 weeks. After 2 weeks, two animals from each group were sacrificed and tumors were collected to analyze for Western blot. For Omm1 xenografts, 6–8 weeks athymic female mice bearing subcutaneously injected Omm1 tumors (7 mice/group) of 100 mm3 diameter were treated with vehicle, AEB071 (80mg/kg/d) TID and or BYL719 orally (50mg/kg/d) QD as single agents and in combination, 5 days/week for 3 weeks. Tumors were homogenized with grinding resins kits as per manufacturer's instructions. Tumors were collected to analyze for H&E and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Tumors were measured every 2 to 3 days with calipers, and tumor volumes were calculated by the formula 4/3 × r3 [r = (larger diameter + smaller diameter)/4. Toxicity was monitored by weight loss [3]. |
| 體外活性 | 在無細胞激酶測定中，Sotrastaurin (AEB071) 抑制了PKC，其K(i)值在亞納摩爾至低納摩爾范圍內。當T細胞受到刺激時，AEB071顯著抑制了原位PKC的催化活性。在原代人類及小鼠T細胞中，低納摩爾濃度的AEB071處理有效阻斷了早期T細胞激活標志[1]。在GNAQ/GNA11突變細胞中，與野生型細胞相比，AEB071觀察到生長抑制作用。在GNAQ突變細胞中，AEB071降低了myristoylated alanine-rich C-kinase substrate（PKC的一個底物）、ERK1/2和核糖體S6的磷酸化，但AKT激活仍然持續(xù)存在[2]。 |
| 體內活性 | 每日口服Sotrastaurin（80 mg/kg，tid）的處理相較于對照組（vehicle-treated animals）顯示出了統(tǒng)計學意義上的腫瘤生長抑制效果，相應地，與對照組相比，治療組的腫瘤體積變化率為17% [2]。與單獨使用AEB071或BYL719相比，聯(lián)合療法在腫瘤體積減少方面顯著更有效。與對照組（vehicle control）相比，這種效果甚至更加顯著 [3]。 |
| 存儲條件 | Powder: -20°C for 3 years | In solvent: -80°C for 1 year | Shipping with blue ice. |
| 溶解度 | DMSO : 81 mg/mL (184.7 mM)
Ethanol : 2 mg/mL (4.56 mM)
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| 關鍵字 | inhibit | Sotrastaurin | Inhibitor | Protein kinase C | AEB-071 | AEB 071 | PKC |
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