RO4987655

MEK1 (IC50 = 5.2 nM); MEK2 (IC50 = 5.2 nM)

RO4987655 有效抑制丝裂原激活蛋白激酶信号通路激活和肿瘤细胞生长，抑制 MEK1/2 的体外 IC50 为 5.2 nM[1]。 RO4987655 以剂量依赖性方式抑制 NCI-H2122 细胞的增殖，IC50 值为 0.0065 μM。在 0.1 至 1.0 μM 的剂量范围内，RO4987655 最早在治疗后 2 小时即可抑制 pERK1/2[2]。

在异种移植模型中，RO4987655 (CH4987655) 作为单一药物口服给药可完全根除肿瘤。 RO4987655 的 tmax 不到一小时，会被迅速吸收。 0.5 至 4 mg 的暴露量与剂量成正比。终末 t1/2 小于 25 小时，该处置是双相的。观察到较低的受试者间变异性； Cmax 和曲线下面积 (AUC) 的范围分别为 9%–23% 和 14%–25%。在较高剂量下，pERK 抑制率超过 80%，并且具有暴露依赖性。抑制性 Emax 模型（Emax ~100%；IC50 40.6 ng/mL）用于非线性混合效应模型来描述药代动力学-药效关系[1]。随机分配的研究组由雌性无胸腺裸鼠组成。使用数字卡尺，在第 0、1 和 3 天，使用剂量为 1.0、2.5 和 5.0 mg/kg RO4987655 来估计肿瘤大小。在此期间，载体治疗不会阻止 NCI-H2122 肿瘤异种移植物的生长。另一方面，RO4987655 治疗第 3 天，1.0 mg/kg 时的肿瘤生长抑制 (TGI) 为 119%，2.5 mg/kg 时为 145%，5.0 mg/kg 时为 150%。PET 成像表明[18F ] RO4987655 给药后 24 小时（第 1 天），异种移植物中的 FDG 摄取减少[2]。

RO4987655（也称为 CH-4987655）是一种新型、口服生物可利用的、特异性的 MEK 激酶小分子抑制剂，对 MEK1/MEK2 的 IC50 为 5.2 nM。丝裂原激活蛋白激酶激酶 1 (MAP2K1/MEK1) 可能具有抗肿瘤活性，是该药物的靶标。 MEK1/2抑制的体外IC50为5.2 nmol/L，可有效阻止丝裂原激活蛋白激酶信号通路的激活和肿瘤细胞的生长。

将热灭活的胎牛血清和 L-谷氨酰胺在人肺腺癌细胞系 NCI-H2122 的指定培养基中维持在指定浓度。在 37 摄氏度和 5% 二氧化碳的条件下，细胞会发育。将细胞在 96 孔板中暴露于不同浓度（0.00001、0.001、0.1 和 10 μM）的 RO4987655 72 小时后，使用 Cell Counting Kit-8 对活细胞进行计数 [2]。

Mice: Mice that are athymic and naked in females It uses balb nu/nu that are 5 to 6 weeks old (18 to 22 g). Balb-nu/nu mice receive a subcutaneous injection of NCI-H2122 cells ((4×106/mouse). Mice are randomized into groups with comparable mean tumor volumes at the beginning of the study once tumors are established (100 to 200 mm3). On days 0, 1, and 3 with doses of 1.0, 2.5, and 5.0 mg/kg RO4987655, PET scans are used to estimate the tumor size. Days 0 (baseline), 1, 2, 3, and 9 of [18F] FDG-PET imaging are used to measure tumor volume and body weight. Calculations are made to determine tumor growth inhibition[2].

Purpose: CH4987655 (RO4987655) is an orally active and highly selective small-molecule MEK inhibitor. It potently inhibits mitogen-activated protein kinase signaling pathway activation and tumor cell growth, with an in vitro IC(50) of 5.2 nmol/L for inhibition of MEK1/2. Single-agent oral administration of CH4987655 resulted in complete tumor regressions in xenograft models. [1]
Experimental design: All 40 subjects received a single oral dose followed by 72 hrs of pharmacokinetic, pharmacodynamic, and safety/tolerability assessments. The pharmacodynamics were measured by changes in phosphorylated extracellular signal-regulated kinase (pERK) levels in a surrogate tissue peripheral blood mononuclear cells ex vivo stimulated by PMA. [1]
Results: Doses of 0.5, 1, 2, 3, and 4 mg were safe and well tolerated. No clinically significant safety event was observed. A total of 26 adverse events (n = 15) were reported: 21 mild, 5 moderate, and none severe. Moderate adverse events were experienced by one subject at 1 mg (autonomic nervous system imbalance) and three subjects at 4 mg (diarrhea, abdominal pain, autonomic nervous system and acne). CH4987655 was rapidly absorbed with a t(max) of approximately 1 h. Exposures were dose proportional from 0.5 to 4 mg. The disposition was biphasic with a terminal t(1/2) of approximately 25 hr. Intersubject variability was low, 9% to 23% for C(max) and 14% to 25% for area-under-the-curve (AUC). pERK inhibition was exposure dependent and was greater than 80% inhibition at higher doses. The pharmacokinetic-pharmacodynamic relationship was characterized by an inhibitory E(max) model (E(max) approximately 100%; IC(50) 40.6 ng/mL) using nonlinear mixed-effect modeling. [1]
Conclusions: A significant extent of pERK inhibition was achieved for a single dose that was considered to be safe and well tolerated in healthy volunteers.

[1]. The safety, tolerability, pharmacokinetics, and pharmacodynamics of single oral doses of CH4987655 in healthy volunteers: target suppression using a biomarker. Clin Cancer Res. 2009 Dec 1;15(23):7368-74.

[2]. Tegnebratt T, et al. Evaluation of efficacy of a new MEK inhibitor, RO4987655, in human tumor xenografts by [(18)F] FDG-PET imaging combined with proteomic approaches. EJNMMI Res. 2014 Dec;4(1):34.

RO4987655 has been used in trials studying the treatment of Neoplasms.
MEK Inhibitor RO4987655 is an orally active small molecule, targeting mitogen-activated protein kinase kinase 1 (MAP2K1 or MEK1), with potential antineoplastic activity. MEK inhibitor RO4987655 binds to and inhibits MEK, which may result in the inhibition of MEK-dependent cell signaling and the inhibition of tumor cell proliferation. MEK, a dual specificity threonine/tyrosine kinase, is a key component of the RAS/RAF/MEK/ERK signaling pathway that regulates cell growth; constitutive activation of this pathway has been implicated in many cancers.

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Inhibition of mitogen-activated protein kinase (MEK, also known as MAPK2, MAPKK), a key molecule of the Ras/MAPK (mitogen-activated protein kinase) pathway, has shown promising effects on B-raf-mutated and some RAS (rat sarcoma)-activated tumors in clinical trials. The objective of this study is to examine the efficacy of a novel allosteric MEK inhibitor RO4987655 in K-ras-mutated human tumor xenograft models using [(18)F] FDG-PET imaging and proteomics technology. Methods: [(18)F] FDG uptake was studied in human lung carcinoma xenografts from day 0 to day 9 of RO4987655 therapy using microPET Focus 120 (CTI Concorde Microsystems, Knoxville, TN, USA). The expression levels of GLUT1 and hexokinase 1 were examined using semi-quantitative fluorescent immunohistochemistry (fIHC). The in vivo effects of RO4987655 on MAPK/PI3K pathway components were assessed by reverse phase protein arrays (RPPA). Results: We have observed modest metabolic decreases in tumor [(18)F] FDG uptake after MEK inhibition by RO4987655 as early as 2 h post-treatment. The greatest [(18)F] FDG decreases were found on day 1, followed by a rebound in [(18)F] FDG uptake on day 3 in parallel with decreasing tumor volumes. Molecular analysis of the tumors by fIHC did not reveal statistically significant correlations of GLUT1 and hexokinase 1 expressions with the [(18)F] FDG changes. RPPA signaling response profiling revealed not only down-regulation of pERK1/2, pMKK4, and pmTOR on day 1 after RO4987655 treatment but also significant up-regulation of pMEK1/2, pMEK2, pC-RAF, and pAKT on day 3. The up-regulation of these markers is interpreted to be indicative of a reactivation of the MAPK and activation of the compensatory PI3K pathway, which can also explain the rebound in [(18)F] FDG uptake following MEK inhibition with RO4987655 in the K-ras-mutated human tumor xenografts. Conclusions: We have performed the first preclinical evaluation of a new MEK inhibitor, RO4987655, using a combination of [(18)F] FDG-PET imaging and molecular proteomics. These results provide support for using preclinical [(18)F] FDG-PET imaging in early, non-invasive monitoring of the effects of MEK and perhaps other Ras/MAPK signaling pathway inhibitors, which should facilitate a wider implementation of clinical [(18)F] FDG-PET to optimize their clinical use.[2]

C20H19F3IN3O5

565.28

565.032

C, 42.49; H, 3.39; F, 10.08; I, 22.45; N, 7.43; O, 14.15

874101-00-5

11548630

White to off-white solid powder

1.7±0.1 g/cm3

1.638

5.49

103.62

3

9

8

32

652

0

O=C(C1C(NC2C(F)=CC(I)=CC=2)=C(F)C(F)=C(CN2C(=O)CCCO2)C=1)NOCCO

FIMYFEGKMOCQKT-UHFFFAOYSA-N

InChI=1S/C20H19F3IN3O5/c21-14-9-12(24)3-4-15(14)25-19-13(20(30)26-31-7-5-28)8-11(17(22)18(19)23)10-27-16(29)2-1-6-32-27/h3-4,8-9,25,28H,1-2,5-7,10H2,(H,26,30)

3,4-difluoro-2-(2-fluoro-4-iodoanilino)-N-(2-hydroxyethoxy)-5-[(3-oxooxazinan-2-yl)methyl]benzamide

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 (4.42 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 (4.42 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 (4.42 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 储备液加入到 900 μL 玉米油中并混合均匀。

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配方 4 中的溶解度: 10% DMSO+ 40% PEG300+ 5% Tween-80+ 45% saline: ≥ 2.5 mg/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；
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7、 以上所有助溶剂都可在 Invivochem.cn网站购买。