Belnacasan 是一种具有口服活性的 IL 转换酶/Caspase-1 抑制剂，是 VRT-043198 的口服生物活性前药，作用于外周血单核细胞，可抑制 LPS 诱导的 IL-1β 和 IL-18 释放。

VX-765 is an orally active IL-converting enzyme/caspase-1 inhibitor.

VX-765通过防止前脑星形胶质细胞中IL-1β的增加阻断大鼠体内的癫痫发生,而对后放电持续时间没有明显影响.在患有遗传性失神癫痫的成年大鼠体内注射3天VX-765后,通过选择性阻断IL-1β生物合成,明显降低累积持续时间,减少平均55%的棘慢波放电.50 mg/kg-200 mg/kg VX-765在急性癫痫小鼠模型中,通过延迟首次癫痫开始时间,并减少平均50%的癫痫发作次数以及64%的总持续时间,产生抗癫痫作用.200 mg/kg VX-765在胶原诱导的关节炎小鼠模型中,抑制60%脂多糖诱导的IL-1β产生,并导致炎症评分剂量依赖性明显降低,有效保护关节病变.

VRT-043198会抑制IL-1β从外周血单个核细胞和全血中的释放,IC50分别为0.67 μM和1.9 μM。VX-765是一种口服可吸收的VRT-043198前药,显示出对ICE /半胱天冬酶-1和半胱天冬酶-4的有效抑制作用,Ki分别为0.8 nM和<0.6 nM。

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].

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].

273404-37-8

C24H33ClN4O6

509.0

VX-765

Ethanol:93 mg/mL (182.7 mM)
DMSO:93 mg/mL (182.7 mM)
Powder: -20°C for 3 years
In solvent: -80°C for 2 years