BRD4 (pIC50 = 6.1); BRD2 (pIC50 = 6.3); BRD3 (pIC50 = 6.6)[1]

I-BET151（1 μM；72 小时）处理表明，大多数活细胞处于 G0 期，这与细胞增殖的剂量和时间依赖性下降以及溴脱氧尿苷积累的消除一致[2]。 I-BET151（100 nM；72 小时）会导致 S/G2 期骨髓瘤细胞比例出现剂量和时间依赖性下降 [2]。

I-BET151 在大鼠中表现出良好的口服全身暴露和低血液清除率（大约 20% 肝血流量），这转化为良好的口服生物利用度。狗表现出高清除率，约占肝血流量的 95%。在狗中，由于全身暴露量低，口服生物利用度可低至 16%。虽然大鼠和小鼠的低内在清除率（小鼠 IVC 1.6 mL/min/g；CLb 8 mL/min/kg）与这些物种的体内血液清除率较低相关，但狗的高血液清除率与在狗微粒体和肝细胞中观察到的高内在清除率。由于狗的全身暴露量较低，小型猪被作为可能的第二个毒理学评估物种进行研究。在这些动物中，I-BET151 表现出良好的生物利用度 (65%) 和较低的清除率（~32% 肝血流量）[1]。

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临床前工具化合物I-BET151先前已被证明具有抗白血病活性，我们的数据显示，它在体内对骨髓瘤也有活性[2]。

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建立HLA-B27/β2m转基因AS Lewis大鼠模型，用30mg/kg I-BET151治疗5周。使用ELISA在体内测量核因子-κB配体受体激活物（RANKL）、骨保护素（OPG）、基质金属蛋白酶（MMP）3和MMP9的水平，并在体外用蛋白质印迹和聚合酶链式反应进行检测。AS血清、AS血清处理的MG63细胞和HLA-B27/β2m转基因AS大鼠的RANKL、OPG、MMP3和MMP9水平上调。相反，在用I-BET151处理的细胞或动物中，RANKL、OPG、MMP3和MMP9的水平受到显著抑制。总体而言，本研究的结果表明，BET抑制剂I-BET151在体内和体外抑制AS中RANKL、OPG、MMP3和MMP9的水平。I-BET151可能显示出用作治疗as患者的治疗剂的潜力。

根据文献报道方法[J.Med.Chem.，54（2011），p.3827]，在BRD2、BRD3和BRD4荧光各向异性（FP）测定中评估了化合物的靶点结合活性。异恶唑喹啉类似物与FP配体竞争，以亚微摩尔IC50与溴结构域结合，如表1所示。通过浸泡在BRD2 N-末端溴结构域的晶体中，获得了化合物1的1.8Å分辨率X射线晶体结构，6揭示了其结合模式（图1A）[1]。

细胞活力测定[2]
细胞类型： H929 细胞
测试浓度： 1 μM
孵育时间： 72 hrs（小时）
实验结果：显示大多数活细胞处于 G0 期，并与细胞增殖的剂量和时间依赖性减少以及溴脱氧尿苷掺入的废除相称。

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细胞增殖测定[2]
细胞类型： H929 细胞
测试浓度： 100 nM
孵育时间：72小时
实验结果：引起S/G2期骨髓瘤细胞比例显着的剂量和时间依赖性下降。

Animal/Disease Models: Mice (model of subcutaneous (sc) myeloma)[2]
Doses: 50 mg/kg
Route of Administration: Ip; daily for 21 days
Experimental Results: decreased rate of tumor size doubling than vehicle-treated mice.

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NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice were bred and maintained in-house at Imperial College in accordance with the 1986 Animal Scientific Procedures Act and under a United Kingdom Government Home Office–approved project license. In total, 5 × 106 KMS11 myeloma cells were injected subcutaneously into 9- to 12-week-old NSG mice. When tumors were ≥5 mm in maximum diameter, mice were randomized to receive once daily intraperitoneal injection of either I-BET151 30 mg/kg in 0.9% NaCl plus Kleptose hydroxypropyl betadex 10% (w/v) and DMSO 5% (v/v) pH 5.0 or vehicle solution for a maximum of 21 days[2].

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AS animal model HLA-B27/β2 m transgenic AS Lewis rat model was constructed as previously described. A total of 20 AS rats were constructed and all animals (including normal Lewis rats, n=10) were housed in standard conditions under a 12-h light/dark cycle with free access to food and water. For I-BET151 treatment, 20 transgenic rats were intraperitoneally administrated with 30 mg/kg of I-BET151 (n=10; GlaxoSmithKline) and equal volume normal saline (n=10) once per day for 5 weeks. At the end of 5 weeks, all animals were anesthetized and 0.5 ml of blood samples were collected before sacrifice. [3]

[1]. Identification of a novel series of BET family bromodomain inhibitors: Binding mode and profile of I-BET151 (GSK1210151A). Bioorg Med Chem Lett. 2012 Apr 15;22(8):2968-72.
[2]. Chaidos A, et al. Potent antimyeloma activity of the novel bromodomain inhibitors I-BET151 and I-BET762. Blood. 2014 Jan 30;123(5):697-705.
[3]. I-BET151 inhibits expression of RANKL, OPG, MMP3 and MMP9 in ankylosing spondylitis in vivo and in vitro. Exp Ther Med . 2017 Nov;14(5):4602-4606.

A novel series of quinoline isoxazole BET family bromodomain inhibitors are discussed. Crystallography is used to illustrate binding modes and rationalize their SAR. One member, I-BET151 (GSK1210151A), shows good oral bioavailability in both the rat and minipig as well as demonstrating efficient suppression of bacterial induced inflammation and sepsis in a murine in vivo endotoxaemia model.[1]

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The bromodomain and extraterminal (BET) protein BRD2-4 inhibitors hold therapeutic promise in preclinical models of hematologic malignancies. However, translation of these data to molecules suitable for clinical development has yet to be accomplished. Herein we expand the mechanistic understanding of BET inhibitors in multiple myeloma by using the chemical probe molecule I-BET151. I-BET151 induces apoptosis and exerts strong antiproliferative effect in vitro and in vivo. This is associated with contrasting effects on oncogenic MYC and HEXIM1, an inhibitor of the transcriptional activator P-TEFb. I-BET151 causes transcriptional repression of MYC and MYC-dependent programs by abrogating recruitment to the chromatin of the P-TEFb component CDK9 in a BRD2-4-dependent manner. In contrast, transcriptional upregulation of HEXIM1 is BRD2-4 independent. Finally, preclinical studies show that I-BET762 has a favorable pharmacologic profile as an oral agent and that it inhibits myeloma cell proliferation, resulting in survival advantage in a systemic myeloma xenograft model. These data provide a strong rationale for extending the clinical testing of the novel antimyeloma agent I-BET762 and reveal insights into biologic pathways required for myeloma cell proliferation.[2]

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Ankylosing spondylitis (AS) is characterized by osteoclastogenesis and inflammatory bone resorption. The present study aimed to investigate the effect of bromodomain and extra-terminal domain (BET) protein inhibitor I-BET151 on AS process. A total of 38 AS Chinese patients were recruited and a further 38 sex- and age-matched healthy participants were selected as control. The Bath AS Function Index and Bath AS Disease Activity Index were assessed in AS patients and levels of erythrocyte sedimentation rate and C-reactive protein were measured in AS and healthy groups. Serum from AS patients was used to induce MG63 osteoblasts and BET inhibitor I-BET151 at concentrations of 50, 100 and 200 ng/ml used for treatment of the cells. A HLA-B27/β2m transgenic AS Lewis rat model was established and treated with 30 mg/kg I-BET151 for 5 weeks. Levels of receptor activator of nuclear factor-κB ligand (RANKL), osteoprotegerin (OPG), matrix metalloproteinase (MMP)3, and MMP9 were measured using ELISA in vivo and additionally detected with western blotting and polymerase chain reaction in vitro. The levels of RANKL, OPG, MMP3 and MMP9 were upregulated in AS serum, AS serum treated MG63 cells and HLA-B27/β2m transgenic AS rats. Conversely, levels of RANKL, OPG, MMP3 and MMP9 were significantly inhibited in cells or animals treated with I-BET151. Overall, the results of the present study demonstrated that BET inhibitor I-BET151 suppresses levels of RANKL, OPG, MMP3 and MMP9 in AS in vivo and in vitro. I-BET151 may exhibit the potential to be used as a therapeutic in the treatment of AS patients.[3]

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7-(3,5-dimethyl-4-isoxazolyl)-8-methoxy-1-[(1R)-1-(2-pyridinyl)ethyl]-3H-imidazo[4,5-c]quinolin-2-one is an imidazoquinoline.

C23H21N5O3

415.44

415.164

C, 66.49; H, 5.09; N, 16.86; O, 11.55

1300031-49-5

I-BET151 dihydrochloride;1883545-47-8

52912189

Typically exists as White to khaki solids at room temperature

1.3±0.1 g/cm3

1.651

2.28

98.83

C[C@@H](N1C(C(C=C(OC)C(C2=C(C)ON=C2C)=C3)=C3N=C4)=C4NC1=O)C5=CC=CC=N5

VUVUVNZRUGEAHB-CYBMUJFWSA-N

InChI=1S/C23H21N5O3/c1-12-21(14(3)31-27-12)16-9-18-15(10-20(16)30-4)22-19(11-25-18)26-23(29)28(22)13(2)17-7-5-6-8-24-17/h5-11,13H,1-4H3,(H,26,29)/t13-/m1/s1

7-(3,5-dimethyl-1,2-oxazol-4-yl)-8-methoxy-1-[(1R)-1-pyridin-2-ylethyl]-3H-imidazo[4,5-c]quinolin-2-one

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 (6.02 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 25.0 mg/mL澄清DMSO储备液加入到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: ≥ 2.5 mg/mL (6.02 mM) (饱和度未知) 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), 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 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: ≥ 2.5 mg/mL (6.02 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 储备液添加到 900 μL 玉米油中并混合均匀。

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配方 4 中的溶解度: ≥ 2.5 mg/mL (6.02 mM) (饱和度未知) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
*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 5 中的溶解度: ≥ 2.5 mg/mL (6.02 mM) (饱和度未知) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
*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 6 中的溶解度: 0.5 mg/mL (1.20 mM) in 1% DMSO 99% 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。
*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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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。