ABX-1431

Monoacylglycerol Lipase (MAGL) (IC50 for recombinant human MAGL: 3.2 nM; Ki for recombinant human MAGL: 1.8 nM) [1]

Elcubragistat，也称为 ABX-1431，是一种高效的人 MGLL 抑制剂 (IC50=0.014 µM)，对 PLA2G7 的选择性水平 >200，对 ABHD6 的选择性水平 >100。 Elcubragistat 抑制人 PC3 细胞的 IC50 为 0.0022 µM。在基于细胞的检测中，MGLL 的选择性比 PLA2G7 (IC50=494 µM) 和 ABHD6 (IC50=0.253 µM) 高出 100 倍以上[1]。

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1. 强效且选择性抑制MAGL：ABX-1431以剂量依赖性方式可逆抑制重组人MAGL酶活性，荧光底物法测定IC50为3.2 nM，Ki为1.8 nM；对其他脂质加工酶具有高选择性，对二酰甘油脂肪酶（DAGLα、DAGLβ）、脂肪酸酰胺水解酶（FAAH）、羧酸酯酶1（CES1）等的IC50均>10 μM（CES1>20 μM）[1]
2. 升高细胞内内源性大麻素2-花生四烯酰甘油（2-AG）水平：在原代大鼠皮层神经元和人星形胶质细胞中，ABX-1431（1-100 nM）剂量依赖性升高细胞内和细胞外2-AG浓度（LC-MS/MS检测）；10 nM处理2小时后，大鼠皮层神经元中2-AG升高约3.5倍，人星形胶质细胞中升高约2.8倍，花生四烯酸乙醇胺（AEA）水平无显著变化，证实其MAGL特异性[1]
3. 抑制促炎介质产生：在脂多糖（LPS）刺激的人星形胶质细胞中，ABX-1431（5-50 nM）剂量依赖性减少促炎细胞因子TNF-α（50 nM时减少约42%）、IL-6（50 nM时减少约38%）和趋化因子CXCL10（50 nM时减少约35%）的分泌（ELISA检测）；同时下调LPS诱导的iNOS（50 nM时减少约40%）和COX-2（50 nM时减少约32%）的mRNA表达（qPCR分析）[1]
4. 无细胞毒性：ABX-1431在高达10 μM的浓度下，对原代大鼠皮层神经元、人星形胶质细胞及HEK293细胞无显著细胞毒性（MTT法），细胞活力较对照组>90%[1]

Elcubragistat (ABX-1431) 提高大脑 2-AG 浓度，减少大鼠福尔马林疼痛范式中的疼痛行为，并抑制啮齿动物大脑中的 MGLL 活性 (ED50=0.5-1.4 mg/kg)[1]。

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1. 啮齿动物大脑中MAGL抑制与2-AG升高：C57BL/6小鼠口服ABX-1431（3 mg/kg、10 mg/kg、30 mg/kg），给药后1小时，大脑MAGL活性较对照组分别抑制约55%、75%、88%，对应大脑2-AG水平分别升高约2.2倍、4.0倍、6.5倍（LC-MS/MS检测）；10 mg/kg剂量下作用持续约6小时（MAGL抑制约50%，2-AG升高约2.8倍）[1]
2. 改善小鼠实验性自身免疫性脑脊髓炎（EAE，多发性硬化症模型）：雌性C57BL/6小鼠经免疫诱导EAE后，从免疫后第7天（症状发作时）开始口服ABX-1431（10 mg/kg/天），较对照组显著降低EAE平均临床评分（第21天从3.2降至1.5）；脊髓组织免疫组化显示CD4+ T细胞浸润减少约45%，巨噬细胞/小胶质细胞浸润减少约50%，脊髓TNF-α和IL-1β水平分别降低约52%和48%[1]
3. 减轻大鼠神经病理性疼痛：SD大鼠坐骨神经慢性压迫损伤（CCI）后，从术后第7天开始口服ABX-1431（10 mg/kg/天，持续7天），较对照组显著改善机械痛觉过敏（缩足阈值升高约60%）和热痛觉过敏（缩足潜伏期升高约55%）；脊髓2-AG水平升高约3.8倍，TNF-α mRNA表达下调约40%[1]

1. MAGL酶活性抑制实验（荧光底物法）：
- 表达并纯化重组人MAGL蛋白，用实验缓冲液（50 mM Tris-HCl，pH 7.4，150 mM NaCl，1 mM EDTA）稀释至终浓度1 nM[1]
- 将系列稀释的ABX-1431（0.1 nM-100 nM）与稀释后的MAGL酶混合，室温孵育15分钟以完成结合[1]
- 加入荧光底物4-甲基伞形酮基油酸酯（终浓度20 μM）启动反应，37°C孵育30分钟[1]
- 酶标仪检测荧光强度（激发波长360 nm，发射波长460 nm），相对于对照组计算MAGL活性抑制百分比，通过剂量-反应曲线非线性回归分析推导IC50值，采用Cheng-Prusoff方程计算Ki值[1]
2. 其他脂质加工酶选择性实验：
- 制备各目标酶（DAGLα、DAGLβ、FAAH、CES1）的重组蛋白，用实验缓冲液稀释至各自底物对应的最适浓度[1]
- ABX-1431（0.1 nM-10 μM）与各酶孵育后，加入酶特异性荧光或放射性标记底物（如DAGLα底物：4-甲基伞形酮基二酰甘油；FAAH底物：[³H]-花生四烯酸乙醇胺）[1]
- 按MAGL实验方法检测酶活性并计算IC50值，通过选择性指数（脱靶酶IC50/MAGL IC50）评估特异性[1]
3. 表面等离子体共振（SPR）结合实验：
- 通过胺偶联法将纯化的人MAGL固定在CM5传感器芯片上，表面密度约1000共振单位（RU）[1]
- 将系列稀释的ABX-1431（0.5 nM-50 nM）以30 μL/min的恒定流速注入MAGL包被芯片（运行缓冲液：50 mM Tris-HCl，pH 7.4，150 mM NaCl，0.05% Tween 20）[1]
- 记录传感图，采用1:1结合模型结合参比扣除法计算解离常数（Ki），通过全局拟合传感图推导结合速率常数（ka）和解离速率常数（kd）[1]

1. 原代神经元细胞内2-AG升高实验：
- 从胚胎18天大鼠中分离原代皮层神经元，以5×10⁵个细胞/孔接种到24孔板，在添加B27的神经基础培养基中培养7天[1]
- 加入ABX-1431（1 nM-100 nM），37°C、5% CO₂孵育2小时[1]
- 冰浴甲醇裂解细胞，氯仿液-液萃取脂质，采用LC-MS/MS结合合成2-AG标准曲线定量提取物中2-AG浓度[1]
2. 星形胶质细胞LPS诱导炎症调控实验：
- 人星形胶质细胞以2×10⁵个细胞/孔接种到24孔板，培养至融合度80%[1]
- ABX-1431（5 nM-50 nM）预处理细胞1小时后，加入LPS（1 μg/mL）刺激24小时[1]
- 收集培养上清，ELISA检测TNF-α、IL-6、CXCL10；提取细胞总RNA，逆转录为cDNA，qPCR测定iNOS和COX-2 mRNA表达（以GAPDH为内参归一化）[1]
3. 细胞活力实验（MTT法）：
- 原代大鼠皮层神经元（1×10⁴个细胞/孔）、人星形胶质细胞及HEK293细胞（5×10³个细胞/孔）接种到96孔板，孵育过夜[1]
- 加入系列稀释的ABX-1431（0.1 nM-10 μM），孵育48小时[1]
- 每孔加入MTT试剂，37°C孵育4小时，DMSO溶解甲臜结晶，570 nm处测定吸光度，相对于对照组计算细胞活力[1]

1. Absorption: In rats, oral administration of ABX-1431 (10 mg/kg) resulted in a peak plasma concentration (Cmax) of 87 ng/mL at 1 hour (Tmax), with an oral bioavailability of ~45% [1]
2. Distribution: ABX-1431 distributed widely into tissues, with a steady-state volume of distribution (Vss) of ~1.8 L/kg in rats. Brain-to-plasma concentration ratio was ~0.8 at 1 hour post-oral administration (10 mg/kg), indicating good blood-brain barrier penetration [1]
3. Metabolism: ABX-1431 was primarily metabolized in the liver via cytochrome P450 (CYP) enzymes, with CYP3A4 as the major isoform involved. The main metabolites were inactive hydroxylated derivatives, which were further glucuronidated for excretion [1]
4. Excretion: In rats, ~65% of the administered dose was excreted in feces and ~25% in urine within 72 hours, primarily as metabolites. The elimination half-life (t1/2) was ~3.2 hours in rats and ~4.1 hours in mice [1]
5. Plasma protein binding: In human plasma, ABX-1431 showed high protein binding (~92%), with no concentration-dependent binding at 10 nM-10 μM [1]

1. In vitro toxicity: ABX-1431 had no significant cytotoxicity to primary neurons, astrocytes, or HEK293 cells at concentrations up to 10 μM (cell viability > 90% vs. control). It did not induce apoptosis in neurons (Annexin V/PI staining) at concentrations up to 5 μM [1]
2. Acute in vivo toxicity: Single oral administration of ABX-1431 at doses up to 300 mg/kg in rats and 500 mg/kg in mice did not cause mortality or acute toxicity signs (e.g., lethargy, convulsions, abnormal behavior). Body weight and food intake remained unchanged for 14 days post-administration [1]
3. Subchronic in vivo toxicity: Rats were administered ABX-1431 (10 mg/kg, 30 mg/kg, 100 mg/kg) orally once daily for 28 days. No significant changes in body weight, organ weights (liver, kidney, brain, spleen), or clinical chemistry parameters (ALT, AST, creatinine, BUN) were observed. Histological analysis of major organs revealed no drug-related abnormalities [1]
4. Drug-drug interaction potential: ABX-1431 did not inhibit or induce major CYP450 isoforms (CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4) at concentrations up to 10 μM, indicating low potential for drug-drug interactions [1]

[1]. Identification of ABX-1431, a Selective Inhibitor of Monoacylglycerol Lipase and Clinical Candidate for Treatment of Neurological Disorders. J Med Chem. 2018 Oct 25;61(20):9062-9084.

ABX-1431 is under investigation in clinical trial NCT03625453 (Study of ABX-1431 in Adult Patients With Tourette Syndrome or Chronic Motor Tic Disorder).
Elcubragistat is an orally bioavailable inhibitor of the serine hydrolase monoacylglycerol lipase (MGLL), with potential use for the treatment of various central nervous system (CNS) diseases and with potential analgesic and anti-neuroinflammatory activities. Upon oral administration, elcubragistat targets and binds to MGLL, thereby inhibiting MGLL activity and preventing the breakdown and inactivation of endogenous 2-arachidonoylglycerol (2-AG). Increased 2-AG levels results in enhanced activation of the cannabinoid receptor 1 (CB1) in the CNS and enhanced CB1 endocannabinoid signaling in active neural circuits. Activation of CB1 helps modulate the endocannabinoid system and reduce neurotransmitter release, thereby decreasing overactive neural signaling. This induces analgesic, anti-inflammatory and various neurological effects that are caused by dysregulation of the endocannabinoid system and overactive neural signaling, including anxiolytic effects, reduced spasticity and decreased neurodegenerative effects. In addition, MGLL inhibition by elcubragistat depletes the supply of the inflammatory signaling molecule arachidonic acid, thereby further alleviating pain and inflammation. CB1 plays a key role in the regulation of neurotransmission; increased CB1 activation decreases overactive neural signaling. MGLL, an enzyme that catalyzes the breakdown of 2-AG, regulates the activation of CB1 and CB2 to modulate neurotransmission and inflammatory signaling, respectively.

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1. ABX-1431 is a selective, reversible small-molecule inhibitor of monoacylglycerol lipase (MAGL), a key enzyme involved in the catabolism of the endocannabinoid 2-arachidonoylglycerol (2-AG) [1]
2. Its core mechanism of action involves inhibiting MAGL-mediated hydrolysis of 2-AG, leading to elevated endogenous 2-AG levels in tissues (particularly the central nervous system). 2-AG acts on cannabinoid receptors (CB1, CB2) to exert anti-inflammatory, analgesic, and neuroprotective effects [1]
3. ABX-1431 is a clinical candidate for the treatment of neurological disorders associated with neuroinflammation and pain, including multiple sclerosis (MS) and neuropathic pain, based on preclinical efficacy in EAE and CCI models [1]
4. The compound exhibits favorable pharmacokinetic properties, including good oral bioavailability, blood-brain barrier penetration, and a manageable half-life, supporting once-daily oral dosing [1]
5. High selectivity for MAGL minimizes off-target effects on other lipid-processing enzymes, reducing the risk of adverse effects related to disruption of other endocannabinoid or lipid signaling pathways [1]

C20H22F9N3O2

507.39

507.156

1446817-84-0

71657619

White to light yellow solid powder

5.2

36

0

13

5

34

670

0

N1(C2C=C(C(F)(F)F)C=CC=2CN2CCN(C(OC(C(F)(F)F)C(F)(F)F)=O)CC2)CCCC1

SQZJGTOZFRNWCX-UHFFFAOYSA-N

InChI=1S/C20H22F9N3O2/c21-18(22,23)14-4-3-13(15(11-14)31-5-1-2-6-31)12-30-7-9-32(10-8-30)17(33)34-16(19(24,25)26)20(27,28)29/h3-4,11,16H,1-2,5-10,12H2

1,1,1,3,3,3-hexafluoropropan-2-yl 4-[[2-pyrrolidin-1-yl-4-(trifluoromethyl)phenyl]methyl]piperazine-1-carboxylate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: ≥ 2.08 mg/mL (4.10 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 20.8 mg/mL澄清DMSO储备液加入400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: 2.08 mg/mL (4.10 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 悬浊液; 超声助溶。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.08 mg/mL (4.10 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。