BMS-986176

BMS-986176 (also named LX-9211): Adaptor Protein-2 Associated Kinase 1 (AAK1) (Ki = 0.8 nM; IC50 for AAK1 enzymatic activity = 1.8 nM) [2]

丝氨酸/苏氨酸激酶 Ark1/Prk1 家族包括接头相关激酶 1 (AAK1)。 AAK1 mRNA 有两种剪接版本：短的和长的。在大脑和心脏中，长形式占主导地位并高度表达。在培养的细胞中，AAK1 与内吞结构共定位，并且在突触体制剂中含量丰富。 AAK1 控制受体介导的内吞作用和网格蛋白包被的内吞作用，这两个过程对于突触小泡回收至关重要 [1]。

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1. BMS-986176对AAK1酶活性表现出强效且高选择性的抑制作用，IC50为1.8 nM，Ki为0.8 nM；在包含403种激酶的激酶面板中，其对AAK1的选择性超过1000倍，对亲缘关系较近的BIKE（AP2相关激酶2）的IC50为1600 nM[2]
2. 在过表达AAK1的人胚肾（HEK293）细胞中，BMS-986176抑制AAK1介导的AP2M1（AP2的μ2亚基）磷酸化的IC50为19 nM[2]
3. 在背根神经节（DRG）神经元中，BMS-986176呈剂量依赖性抑制神经生长因子（NGF）诱导的AAK1活性，以及与神经性疼痛相关的下游信号通路[2]

1. 在小鼠保留性神经损伤（SNI）神经性疼痛模型中，口服给予BMS-986176（3、10、30 mg/kg）可剂量依赖性逆转机械性异常性疼痛和热痛觉过敏，在剂量≥10 mg/kg时观察到显著疗效，镇痛效果在给药后至少持续6小时[2]
2. 在大鼠慢性压迫性损伤（CCI）神经性疼痛模型中，BMS-986176（10、30 mg/kg，口服）同样表现出剂量依赖性镇痛活性，30 mg/kg剂量下可逆转50%–70%的机械性异常性疼痛[2]
3. BMS-986176展现出优异的中枢神经系统（CNS）穿透性，小鼠口服给药后脑/血浆比值为2.8，大鼠为1.2[2]
4. 在啮齿类动物模型中，BMS-986176在镇痛剂量下未引起运动障碍或镇静作用（通过转棒实验和旷场实验评估）[2]

1. AAK1酶活性实验：将重组人源AAK1蛋白与肽底物（源自AP2M1）、ATP及不同浓度的BMS-986176共同孵育，加入ATP启动反应并在室温下孵育一定时间。采用时间分辨荧光共振能量转移（TR-FRET）法检测底物的磷酸化水平，通过测量荧光信号计算抑制率及IC50/Ki值[2]
2. 激酶选择性实验：将BMS-986176以1 μM浓度在包含403种人源激酶的面板中进行测试，采用放射性或发光法检测各激酶的活性，根据被抑制<50%的激酶比例计算选择性评分[2]

1. HEK293细胞中AP2M1磷酸化实验：将稳定表达FLAG标签AAK1的HEK293细胞接种于多孔板并过夜培养。细胞经不同浓度的BMS-986176预处理1小时后，加入过钒酸盐（磷酸酶抑制剂）增强AAK1活性。裂解细胞后，通过蛋白质印迹法（Western blot）检测磷酸化AP2M1（p-AP2M1）的水平，对条带强度进行定量以确定BMS-986176抑制AP2M1磷酸化的IC50[2]
2. DRG神经元实验：从新生大鼠中分离原代DRG神经元并在神经基础培养基中培养。神经元经不同剂量的BMS-986176处理2小时后，用NGF（50 ng/mL）刺激15分钟。通过Western blot分析AAK1及下游信号分子（如ERK1/2、AKT）的磷酸化水平，采用实时荧光定量PCR（qPCR）对疼痛相关基因的表达进行定量[2]

1. For mouse SNI neuropathic pain model: Male C57BL/6 mice (8–10 weeks old) were subjected to SNI surgery (ligation and transection of the tibial and common peroneal nerves, leaving the sural nerve intact). Seven days post-surgery, mice with established mechanical allodynia (assessed by von Frey filaments) were randomized into treatment groups. BMS-986176 was formulated in a vehicle consisting of 0.5% methylcellulose and 0.1% Tween 80 in water, and administered orally by gavage at doses of 3, 10, 30 mg/kg once daily for 7 days. Mechanical allodynia was measured using von Frey filaments (0.02–2 g) at 1, 3, 6, and 24 hours post-dosing, and thermal hyperalgesia was assessed using a hot plate test (52 ± 0.5 °C) [2]
2. For rat CCI neuropathic pain model: Male Sprague-Dawley rats (200–250 g) underwent CCI surgery (four loose ligatures around the sciatic nerve). Ten days post-surgery, rats with confirmed mechanical allodynia received oral gavage of BMS-986176 (10, 30 mg/kg) or vehicle once daily for 5 days. Mechanical allodynia was evaluated using von Frey filaments (2–15 g) at 2 and 4 hours post-dosing each day [2]
3. For CNS penetration study: Mice and rats were administered a single oral dose of BMS-986176 (10 mg/kg). At predetermined time points (0.5, 1, 2, 4, 6 hours), animals were euthanized, blood was collected to obtain plasma, and brains were dissected and homogenized. The concentration of BMS-986176 in plasma and brain homogenates was measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and the brain/plasma ratio was calculated [2]
4. For pharmacokinetic (PK) study: Male CD-1 mice, Sprague-Dawley rats, and Beagle dogs were used. BMS-986176 was administered via oral gavage (10 mg/kg) or intravenous (IV) injection (2 mg/kg). Blood samples were collected at multiple time points (0–24 hours) and analyzed by LC-MS/MS to determine plasma drug concentrations. PK parameters (Cmax, Tmax, AUC, t1/2, F) were calculated using non-compartmental analysis [2]

1. Absorption: BMS-986176 showed good oral bioavailability: 72% in mice (10 mg/kg p.o.), 68% in rats (10 mg/kg p.o.), and 45% in dogs (10 mg/kg p.o.) [2]
2. Distribution: Excellent CNS penetration, with brain/plasma ratios of 2.8 (mice) and 1.2 (rats) after oral administration; volume of distribution (Vdss) was 3.2 L/kg in rats and 5.8 L/kg in dogs [2]
3. Metabolism: BMS-986176 was primarily metabolized by cytochrome P450 (CYP) enzymes, with CYP3A4 as the major isoform responsible for its biotransformation; minor metabolites included hydroxylated and demethylated derivatives [2]
4. Elimination: Terminal half-life (t1/2) was 4.2 hours in mice, 6.8 hours in rats, and 12.5 hours in dogs after oral dosing; renal excretion accounted for ~15% of the administered dose, and fecal excretion accounted for ~75% in rats [2]
5. PK parameters (rat, 10 mg/kg p.o.): Cmax = 1250 ng/mL, Tmax = 1 hour, AUC0-∞ = 8900 ng·h/mL [2]

1. Plasma protein binding: BMS-986176 had high plasma protein binding in humans (96%), mice (94%), rats (95%), and dogs (97%) [2]
2. In vitro drug-drug interaction: BMS-986176 did not inhibit major CYP450 isoforms (CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4) at concentrations up to 10 μM, and did not induce CYP3A4 activity in human hepatocytes [2]
3. Acute toxicity: In single-dose toxicity studies in mice and rats, BMS-986176 was well-tolerated at doses up to 300 mg/kg p.o., with no observed mortality or severe adverse effects (e.g., weight loss, behavioral changes) [2]
4. Repeat-dose toxicity: In a 14-day repeat-dose study in rats (10, 30, 100 mg/kg p.o. once daily), BMS-986176 caused no significant histopathological changes in major organs (liver, kidney, brain, heart); the no-observed-adverse-effect level (NOAEL) was 100 mg/kg/day [2]
5. Neurotoxicity: BMS-986176 did not cause motor impairment in rotarod tests (mice, 30 mg/kg p.o.) or cognitive deficits in novel object recognition tests (rats, 30 mg/kg p.o.) [2]

[1]. Biaryl kinase inhibitors. WO2015153720A1.

[2]. Discovery of ( S)-1-((2',6-Bis(difluoromethyl)-[2,4'-bipyridin]-5-yl)oxy)-2,4-dimethylpentan-2-amine (BMS-986176/LX-9211): A Highly Selective, CNS Penetrable, and Orally Active Adaptor Protein-2 Associated Kinase 1 Inhibitor in Clinical Trials for the Treatment of Neuropathic Pain. J Med Chem. 2022 Mar 24;65(6):4457-4480.

1. BMS-986176 (LX-9211) is a highly selective, CNS-penetrant, orally active AAK1 inhibitor discovered through structure-based drug design and medicinal chemistry optimization of bipyridine-based scaffolds [2]
2. The drug is being evaluated in phase I/II clinical trials (NCT04119276) for the treatment of chronic neuropathic pain, including painful diabetic peripheral neuropathy (DPN) and post-herpetic neuralgia (PHN) [2]
3. AAK1 inhibition by BMS-986176 blocks the endocytosis of pain-related receptors (e.g., TrkA, TRPV1) in sensory neurons, thereby reducing neuronal sensitization and neuropathic pain signaling [2]
4. The patent [1] discloses biaryl kinase inhibitors as potential therapeutic agents, but does not specifically mention BMS-986176 or its therapeutic indications [1]

C19H23F4N3O

385.4066

385.177

C, 59.21; H, 6.02; F, 19.72; N, 10.90; O, 4.15

1815613-42-3

1815613-42-3;

118419773

Off-white to light yellow solid powder

1.2±0.1 g/cm3

470.8±45.0 °C at 760 mmHg

238.5±28.7 °C

0.0±1.2 mmHg at 25°C

1.501

3.11

61

1

8

8

27

455

1

FC([H])(C1=C(C([H])=C([H])C(C2C([H])=C([H])N=C(C([H])(F)F)C=2[H])=N1)OC([H])([H])[C@](C([H])([H])[H])(C([H])([H])C([H])(C([H])([H])[H])C([H])([H])[H])N([H])[H])F

RKAHOQATMSONTM-IBGZPJMESA-N

InChI=1S/C19H23F4N3O/c1-11(2)9-19(3,24)10-27-15-5-4-13(26-16(15)18(22)23)12-6-7-25-14(8-12)17(20)21/h4-8,11,17-18H,9-10,24H2,1-3H3/t19-/m0/s1

(2S)-1-[2-(difluoromethyl)-6-[2-(difluoromethyl)pyridin-4-yl]pyridin-3-yl]oxy-2,4-dimethylpentan-2-amine

AAK1-IN-1LX-9211LX9211 LX 9211 BMS 986176BMS-986176 BMS986176

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 本产品在运输和储存过程中需避光。

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

DMSO : ~220 mg/mL (~570.84 mM)

配方 1 中的溶解度: 5.5 mg/mL (14.27 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), 悬浮液；超声助溶。
例如，若需制备1 mL的工作液，可将100 μL 55.0mg/mL澄清的DMSO储备液加入到900μL 20％SBE-β-CD生理盐水中，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 2 中的溶解度: ≥ 5 mg/mL (12.97 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 50.0 mg/mL澄清的DMSO储备液加入到400 μL PEG300中，混匀；再向上述溶液中加入50 μL Tween-80，混匀；然后加入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 3 中的溶解度: ≥ 5 mg/mL (12.97 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 50.0 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。