| 名稱(chēng) | Belnacasan |
| 描述 | Belnacasan (VX-765) is an IL-converting enzyme (ICE)/caspase-1 inhibitor and an orally bioactive prodrug of VRT-043198. Belnacasan modifies the active site of caspase-1 and leads to caspase-1 blockade, which has been associated with tissue protection against inflammatory diseases in animal models. |
| 細(xì)胞實(shí)驗(yàn) | VX-765 is solubilized in DMSO and stored, and then diluted with RPMI 1640 complete medium (DMSO 0.2%) before use[1]. A total of 2×105 cells/well (100 μL cell suspension) is distributed in triplicate in flat-bottom 96-well plates. Either 50 μL of VX-765 (40 μM in RPMI 1640 complete medium containing 0.2% DMSO) or vehicle control is added to appropriate wells. Following a 30-min incubation at 37°C, 50 μL of LPS diluted in RPMI 1640 complete medium is added at final concentrations varying from 0.001 to 10 ng/mL. Cells are returned to a 37°C incubator. At 4 h after LPS addition, 75 μL of supernatant is removed from wells, cleared by centrifugation for 5 min at 1500 rpm, and stored at 4°C until assayed. Cells are returned to a 37°C incubator until 24 h after LPS addition, at which time 100 μL of supernatant is removed, cleared by centrifugation, and stored at 4°C. Supernatants are tested using ELISA kits for IL-1β, IL-6, IL-18, and IL-1α[1]. |
| 激酶實(shí)驗(yàn) | Enzyme inhibition is assayed by tracking of the rate of hydrolysis of an appropriate substrate labeled with either p-nitroaniline or aminomethyl coumarin (AMC) as follows: ICE/caspase-1, suc-YVAD-p-nitroanilide; caspase-4, Ac-WEHD-AMC; caspase-6, Ac-VEID-AMC; caspase-3, -7, -8, and -9, Ac-DEVD-AMC; and granzyme B, Ac-IEPD-AMC. Enzymes and substrates are incubated in a reaction buffer [10 mM Tris, pH 7.5, 0.1% (w/v) CHAPS, 1 mM dithiothreitol, and 5% (v/v) DMSO] for 10 min at 37°C. Glycerol is added to the buffer at 8% (v/v) for caspase-3, -6, and -9 and granzyme B to improve stability of enzymes. The rate of substrate hydrolysis is monitored using a fluorometer. Assays for cathepsin B and trypsin are performed[2]. |
| 體外活性 | 方法：人 PBMCs 和全血用 LPS 和 Belnacasan 處理 18 h，使用 ELISA 檢測(cè)細(xì)胞因子。
結(jié)果：Belnacasan 抑制 PBMC 和全血的 IL-1β 釋放，IC50 值為 0.67 μM 和 1.9 μM。[1] |
| 體內(nèi)活性 | 方法：為測(cè)試體內(nèi)活性，將 Belnacasan (25-200 mg/kg) 灌胃給藥給 CD-1 小鼠，1 h 后靜脈注射 LPS (2 mg/kg)。
結(jié)果：Belnacasan 在體內(nèi)抑制 LPS 誘導(dǎo)的 IL-1β 產(chǎn)生。[1]
方法：為檢測(cè)抗癲癇活性，將 Belnacasan (12.5-200 mg/kg) 腹腔注射給慢性癲癇 C57BL6 小鼠模型，每天兩次，持續(xù)四天。
結(jié)果：Belnacasan 的反復(fù)全身給藥以劑量依賴(lài)的方式顯著降低了小鼠的慢性癲癇活動(dòng)。[2] |
| 存儲(chǔ)條件 | Powder: -20°C for 3 years | In solvent: -80°C for 1 year
Shipping with blue ice/Shipping at ambient temperature. |
| 溶解度 | 10% DMSO+40% PEG300+5% Tween 80+45% Saline : 9.3 mg/mL (18.27 mM), Solution.
Ethanol : 93 mg/mL (182.71 mM), Sonication is recommended.
DMSO : 240 mg/mL (471.52 mM), Sonication is recommended.
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| 關(guān)鍵字 | VX765 | VX 765 | Inhibitor | inhibit | Caspase-4 | Caspase-1 | Caspase | Belnacasan |
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