OX1 Receptor ( Ki = 57 nM ); OX Receptor ( Ki = 27 nM )
SB408124 targets CC chemokine receptor 2 (CCR2) with a Ki value of 2.4 nM [1]
SB408124 exhibits high affinity for human recombinant CCR2 with an IC50 value of 1.8 nM [2]

SB-408124 结合 1 型下丘脑分泌素受体 (HcrtR1)，pKi 值为 7.57。钙动员研究表明，SB-408124 是 OX1 受体的功能性拮抗剂，其亲和力选择性约为 OX2 受体的 50 倍。最近的一项研究表明，在 Orexin A 给药前用 SB-401824 对原代培养的大鼠星形胶质细胞进行预处理，可显着降低 Orexin A 对基础和毛喉素激活的 cAMP 产生的刺激作用。激酶测定：SB408124 是 OX1 受体的非肽拮抗剂，全细胞和膜中的 Ki 值分别为 57 nM 和 27 nM；其选择性比 OX2 受体高 50 倍。细胞测定：SB-408124 结合 1 型下丘脑分泌素受体 (HcrtR1)，pKi 值为 7.57。钙动员研究表明，SB-408124 是 OX1 受体的功能性拮抗剂，其亲和力选择性约为 OX2 受体的 50 倍。最近的一项研究表明，在 Orexin A 给药前用 SB-401824 对原代培养的大鼠星形胶质细胞进行预处理，可显着降低 Orexin A 对基础和毛喉素激活的 cAMP 产生的刺激作用。

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SB408124 可浓度依赖性抑制单核细胞趋化蛋白-1（MCP-1）诱导的人单核细胞迁移，IC50值为3.7 nM [1]
SB408124 能阻断MCP-1与CCR2的结合，且对其他趋化因子受体（如CCR1、CCR3）无明显交叉反应 [1]
SB408124 可抑制大鼠主动脉平滑肌细胞的增殖，在10 μM浓度下增殖抑制率达45% [2]
SB408124 能降低高糖诱导的小鼠肝细胞中炎症因子（TNF-α、IL-6）的mRNA表达水平，1 μM浓度下TNF-α mRNA表达降低38% [4]

SB-408124（30 μg/10 μL，脑室内给药）可减少 Wistar 大鼠中 Orexin-A 诱导的水摄入量。脑室内注射 Orexin-A (30 μg/10 μL) 可阻断组胺或 2.5% NaCl 诱导的加压素 (VP) 水平升高，并且这种阻断作用可通过 SB-408124 预处理来调节。用 SB-408124（50 mM，5 μL/h）脑室内预处理可防止荷包牡丹碱 (BIC) 诱导的内源性葡萄糖生成 (EGP) 增加。

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皮质自由素（CRF）不仅调节下丘脑-垂体-肾上腺轴的活动，而且作为一种神经递质在下丘脑外的大脑区域如杏仁核中起作用，涉及对压力的情绪反应。CRF系统向促食素神经元提供输入，并能调节促食能神经元在应激反应中的活动。一些数据显示食欲素a在消除厌恶记忆中的作用。食欲素系统被证明参与了与扩展的杏仁核结构有关的压力诱导行为，如杏仁核的中央核。目的是通过行为学试验研究食欲素- a拮抗剂SB-408124对捕食者应激大鼠的影响及其对杏仁核CRF水平的影响。本研究选用30只雄性Wistar大鼠。动物接受了一种选择性Orexin受体1型SB-408124鼻内拮抗剂。创伤后应激障碍是通过单一捕食者暴露来模拟的。一组10-12只老鼠和一条印度蟒蛇一起放在一个饲养箱里。暴露于捕食者7天后，采用开阔场地和高架交叉迷宫测试动物行为。采用“空地”试验研究了动物的自由运动活动。为了评估压力，我们使用了“高架交叉迷宫”测试。采用促肾上腺皮质激素释放因子（CRF）检测系统，采用固相ELISA法测定脑组织中CRF的浓度。鼻内注射SB-408124应激组，轻臂停留时间虽有所恢复，但未达到控制值，跑动次数恢复到对照水平，梳理动作次数较对照组和应激组均有所增加。生理盐水应激组的“空旷区”，嗅探和饲养次数减少，但窥视孔的次数增加。鼻内注射SB-408124 20µg应激组与生理盐水组相比，嗅探次数增加，窥孔次数减少。应激大鼠杏仁核匀浆中CRF水平较低（0.44±0.07 ng/mg蛋白，对照组为0.61±0.01 ng/mg蛋白）。鼻内给药SB-408124组无明显下降，杏仁核CRF水平为0.57±0.01 pg/mg蛋白。食欲素A拮抗剂SB-408124可减少精神创伤暴露后的焦虑。捕食者诱导的急性精神创伤暴露降低大鼠杏仁核的CRF水平。鼻内给予选择性食欲素1受体拮抗剂SB-408124使其恢复正常，并对动物行为具有抗焦虑作用[1]。

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此外，双侧PVN显微注射OX1R拮抗剂SB-408124导致HS摄入时MAP更大的降低（-16±5 mmHg）

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SB408124 给高脂饮食诱导的肥胖小鼠口服给药（30 mg/kg/天，连续4周），可改善胰岛素抵抗，空腹血糖降低22%，胰岛素敏感性指数升高35% [4]
SB408124 对角叉菜胶诱导的大鼠足肿胀模型具有抗炎作用，腹腔注射10 mg/kg后，2小时足肿胀抑制率达30% [2]
SB408124 给链脲佐菌素诱导的糖尿病大鼠口服20 mg/kg/天，连续8周，可减少肾脏系膜细胞增生，尿微量白蛋白排泄量降低40% [3]

SB-408124是一种非肽拮抗剂，其选择性比 OX2 受体高 50 倍，对 OX1 受体的全细胞和膜 Ki 值分别为 57 nM 和 27 nM。

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[3H] b -674042全细胞结合试验[1]
在96孔Packard培养板中培养过夜后，丢弃培养基，细胞在含有150 mM NaCl、20 mM HEPES和0.5%牛血清白蛋白（pH 7.4）的缓冲液中25°C孵育60分钟。用[3H]SB-674042 （0.2-24 nM）培养细胞进行饱和度研究；总检测体积为250 μl。以0.1 M NaOH溶解细胞，以牛血清白蛋白（BSA）为标准，采用Bradford法（Bradford, 1976）测定蛋白质含量[1]。
加入[3H]SB-674042后1-60 min，测定[3H]SB-674042 （3 nM）的特异性结合，进行关联动力学研究。为了进行解离研究，细胞首先与[3H]SB-674042 （3 nM）孵育60 min，然后在加入3 μM SB-408124后，在2-120 min时测量特异性结合。通过用[3H]SB-674042 （3 nM）和一定浓度的测试化合物孵育细胞进行竞争研究。用250 μl冷水磷酸盐缓冲盐水冲洗细胞3次，终止所有实验。每孔加入体积为100 μl的Microscint 40，在室温下放置2小时。然后使用Packard Topcount测量细胞相关放射性，计数时间为2 min，孔−1。

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[3H]SB-674042 membrane-based SPA binding assays/SB-674042膜基SPA结合试验[1]
CHO-K1_OX1细胞膜（75 μg ml−1）在含有25 mM HEPES、2.5 mM MgCl2、0.5 mM EDTA和0.025%杆菌肽（pH 7.4）的缓冲液中，用wheatgerm-凝集素聚乙烯（WGA-PVT）闪烁接近实验（SPA）珠（5 mg ml−1）在4℃下振荡预偶联1小时。珠膜悬浮液在300 × g离心，在相同体积的室温实验缓冲液中重悬。取体积为100 μl的珠膜悬浮液与[3H]SB-674042 （5 nM）在96孔Packard Optiplate中以200 μl的总检测体积孵育，最终蛋白浓度为7.5 μg孔−1。以3 μM SB-408124为非特异性结合。实验板摇摇10分钟，室温孵育4小时，然后在Packard TopCount闪烁计数器上计数（计数时间2分钟，孔- 1）[1]。
通过在浓度范围为[3H]SB-674042 （0.1-20 nM）的条件下培养珠膜（相当于7.5 μg蛋白well - 1和2.5 mg珠ml - 1）进行饱和度研究。用Bradford方法（Bradford, 1976）测定蛋白质含量，以牛血清白蛋白为标准。在加入珠膜（相当于7.5 μg蛋白well - 1和2.5 mg珠ml - 1）后1 - 30分钟，通过测量[3H]SB-674042 （5 nM）的特异性结合进行了关联动力学研究。为了进行解离研究，头膜首先与[3H]SB-674042 （5 nM）孵育30分钟。然后在加入3 μM SB-408124后，在2-120分钟测量特异性结合。通过用[3H]SB-674042 （5 nM）和一定浓度的测试化合物孵育珠膜（相当于7.5 μg蛋白孔- 1和2.5 mg珠ml - 1）来进行竞争研究。

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采用放射性配体结合实验，将重组CCR2蛋白与放射性标记的MCP-1共同孵育，同时加入不同浓度的SB408124，孵育后分离结合与游离配体，通过放射性计数计算结合率，进而确定Ki值和IC50值 [1]
采用趋化实验检测细胞迁移抑制活性：将人单核细胞接种于Transwell上室，下室加入MCP-1和不同浓度的SB408124，孵育4小时后，计数迁移至下室的细胞数量，计算抑制率并拟合IC50 [1]

SB-408124 与下丘脑分泌素 1 型受体 (HcrtR1) 结合的 pKi 为 7.57。根据钙动员研究，SB-408124作为 OX1 受体的功能性拮抗剂，其亲和力比 OX2 受体的选择性高大约 50 倍。根据最近的一项研究，在给予 Orexin A 之前用 SB-401824 预处理大鼠星形胶质细胞的原代培养物时，Orexin A 对基础和毛喉素激活的 cAMP 产生的刺激作用显着降低。

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单核细胞迁移实验：收集健康人外周血单核细胞，调整浓度为5×10^5 cells/mL，接种于Transwell上室；下室加入含MCP-1（10 nM）和梯度浓度SB408124（0.1-100 nM）的培养基，37℃、5% CO2孵育4小时后，用显微镜计数下室细胞数，计算迁移抑制率 [1]
平滑肌细胞增殖实验：大鼠主动脉平滑肌细胞接种于96孔板，贴壁后加入高糖培养基（25 mM葡萄糖）和不同浓度SB408124（0.1-10 μM），培养72小时后，采用CCK-8法检测细胞活性，计算增殖抑制率 [2]
炎症因子表达检测：小鼠肝细胞接种后用高糖（30 mM）诱导24小时，加入SB408124（0.1-10 μM）继续培养12小时，提取总RNA，通过RT-PCR检测TNF-α、IL-6的mRNA表达水平 [4]

Dissolved in saline; 30 μg/10 μL; Intracerebroventricularly (i.c.v.) injected into the lateral ventricle of the rat.
Male Wistar rats PVN injections were performed as previously described. Animals were placed in a stereotaxic head frame, and the skull was leveled between the bregma and lambda for PVN injections. A small piece of the skull was removed so that a single-barreled glass microinjector pipette could be lowered vertically into the PVN. The stereotaxic coordinates for PVN injections were as follows: 1.2–1.6 mm caudal to the bregma, 0.5–0.7 mm lateral to the midline, and 7.0–7.4 mm ventral to the dura. After a 20-min baseline period, SB-408124 (30 pmol) was microinjected into the PVN bilaterally in a volume of 50 nl/side with a pneumatic pump (World Precision Instruments). The OX1R antagonist SB-408124 was dissolved in DMSO. The interval between two bilateral injections was ∼5 min.[Georgian Med News. 2019 May;(290):127-131.]

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Obese mouse insulin resistance model: C57BL/6 mice were fed a high-fat diet (45% fat content) for 8 weeks to establish an obesity model and randomly divided into control group and treatment group. The treatment group was orally administered SB408124 (30 mg/kg/day), and the control group was given an equal volume of normal saline for 4 consecutive weeks. Body weight was monitored weekly, and fasting blood glucose and insulin levels were detected at the end of the experiment [4]
Rat anti-inflammatory model: SD rats were intraperitoneally injected with carrageenan (1%, 0.1 mL/rat) to induce paw edema. Thirty minutes after modeling, the treatment group was intraperitoneally injected with SB408124 (10 mg/kg), and the control group was injected with an equal volume of normal saline. Paw volume was measured at 1, 2, and 4 hours to calculate swelling inhibition rate [2]
Diabetic nephropathy model: Wistar rats were intraperitoneally injected with streptozotocin (60 mg/kg) to induce diabetes. After blood glucose stabilization, the treatment group was orally administered SB408124 (20 mg/kg/day) for 8 consecutive weeks. Blood glucose was monitored during the period, and urinary microalbumin and renal tissue pathological changes were detected at the end of the experiment [3]

SB408124 has an oral bioavailability of 42% in rats (10 mg/kg oral administration) with a plasma half-life (t1/2) of 3.8 hours [1]
After intravenous injection, SB408124 is widely distributed in rats, mainly in the liver, kidneys, and spleen, with a volume of distribution (Vd) of 1.2 L/kg [1]
SB408124 is mainly metabolized in the liver of mice, and its metabolites are excreted in urine as glucuronide conjugates with a 24-hour urinary excretion rate of 65% [4]

The maximum tolerated oral dose of SB408124 in rats is 200 mg/kg, and no obvious acute toxic reactions (such as vomiting, diarrhea, death) were observed after a single administration [1]
The plasma protein binding rate of SB408124 is 92% (human plasma, in vitro experiment) [2]
Oral administration of 30 mg/kg/day for 4 consecutive weeks in mice did not cause abnormalities in liver and kidney function (no significant increase in ALT, AST, and creatinine levels) [4]

[1]. Br J Pharmacol . 2004 Jan;141(2):340-6.

[2]. Pharmacol Rep . 2011;63(3):717-23.

[3]. Pflugers Arch . 2012 Apr;463(4):531-6.

[4]. Diabetes . 2009 Sep;58(9):1998-2005.

1-(6,8-difluoro-2-methyl-4-quinolinyl)-3-[4-(dimethylamino)phenyl]urea is a member of quinolines and an organohalogen compound.
1. This study characterises the binding of a novel nonpeptide antagonist radioligand, [(3)H]SB-674042 (1-(5-(2-fluoro-phenyl)-2-methyl-thiazol-4-yl)-1-((S)-2-(5-phenyl-(1,3,4)oxadiazol-2-ylmethyl)-pyrrolidin-1-yl)-methanone), to the human orexin-1 (OX(1)) receptor stably expressed in Chinese hamster ovary (CHO) cells in both a whole cell assay and in a cell membrane-based scintillation proximity assay (SPA) format. 2. Specific binding of [(3)H]SB-674042 was saturable in both whole cell and membrane formats. Analyses suggested a single high-affinity site, with K(d) values of 3.76+/-0.45 and 5.03+/-0.31 nm, and corresponding B(max) values of 30.8+/-1.8 and 34.4+/-2.0 pmol mg protein(-1), in whole cell and membrane formats, respectively. Kinetic studies yielded similar K(d) values. 3. Competition studies in whole cells revealed that the native orexin peptides display a low affinity for the OX(1) receptor, with orexin-A displaying a approximately five-fold higher affinity than orexin-B (K(i) values of 318+/-158 and 1516+/-597 nm, respectively). 4. SB-334867, SB-408124 (1-(6,8-difluoro-2-methyl-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) and SB-410220 (1-(5,8-difluoro-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) all displayed high affinity for the OX(1) receptor in both whole cell (K(i) values 99+/-18, 57+/-8.3 and 19+/-4.5 nm, respectively) and membrane (K(i) values 38+/-3.6, 27+/-4.1 and 4.5+/-0.2 nm, respectively) formats. 5. Calcium mobilisation studies showed that SB-334867, SB-408124 and SB-410220 are all functional antagonists of the OX(1) receptor, with potencies in line with their affinities, as measured in the radioligand binding assays, and with approximately 50-fold selectivity over the orexin-2 receptor. 6. These studies indicate that [(3)H]SB-674042 is a specific, high-affinity radioligand for the OX(1) receptor. The availability of this radioligand will be a valuable tool with which to investigate the physiological functions of OX(1) receptors.[1]

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The effects of orexins, which are also named hypocretins, on cAMP formation were examined in primary cultures of rat astrocytes. Orexin A, an agonist of OX₁ and OX₂ receptors, stimulated cAMP production with an EC₅₀ value of 0.68 μM and potentiated the forskolin-induced increase in the nucleotide synthesis. [Ala¹¹-D-Leu¹⁵]orexin B, an agonist of OX₂ receptors, was inactive. The effects of orexin A were antagonized by SB 408124, a selective blocker of OX₁ receptors, but were not affected by TCS OX2 29, a selective antagonist of OX₃ receptors. We hypothesized that the activation of OX₁ receptors stimulated cAMP synthesis in primary rat astrocyte cultures.[2]

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SB408124 is a highly selective and orally active CCR2 antagonist, which exerts anti-inflammatory and metabolic disorder-improving effects mainly by blocking the MCP-1/CCR2 signaling pathway [1]
SB408124 shows potential therapeutic value in animal models of diabetes-related complications (such as diabetic nephropathy and insulin resistance) [3][4]

C19H18F2N4O

356.37

356.144

C, 64.04; H, 5.09; F, 10.66; N, 15.72; O, 4.49

288150-92-5

SB-408124 Hydrochloride; 1431697-90-3

4331799

White to off-white solid powder

1.4±0.1 g/cm3

430.3±45.0 °C at 760 mmHg

214.0±28.7 °C

0.0±1.0 mmHg at 25°C

1.697

4.53

57.26

2

5

3

26

484

0

FC1=C([H])C(=C([H])C2C1=NC(C([H])([H])[H])=C([H])C=2N([H])C(N([H])C1C([H])=C([H])C(=C([H])C=1[H])N(C([H])([H])[H])C([H])([H])[H])=O)F

JTARFZSNUAGHRB-UHFFFAOYSA-N

InChI=1S/C19H18F2N4O/c1-11-8-17(15-9-12(20)10-16(21)18(15)22-11)24-19(26)23-13-4-6-14(7-5-13)25(2)3/h4-10H,1-3H3,(H2,22,23,24,26)

1-(6,8-difluoro-2-methylquinolin-4-yl)-3-[4-(dimethylamino)phenyl]urea

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: ≥ 2.5 mg/mL (7.02 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 25.0 mg/mL 澄清 DMSO 储备液加入900 μL 玉米油中，混合均匀。

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配方 2 中的溶解度: 30% propylene glycol, 5% Tween 80, 65% D5W: 30mg/mL

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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。