GDC-0623 (G-868)

MEK1 (Ki = 0.13 nM)

GDC-0623 (RG 7421) 和 G-573 在体外能够阻止 CRAF 引起的 MEK 磷酸化，并能够阻断 BRAF(V600E) 引起的 MEK 磷酸化[1]。尽管 GDC-0623 (RG 7421) 是一种强大的 ATP 非竞争性 MEK1 抑制剂，但与其他两种抑制剂相比，它在细胞活性方面表现出明显的变化，EC50 仅降低 6 倍[2]。

GDC-0623 (RG 7421)（40 mg/kg，口服）在 MiaPaCa-2 异种移植模型中表现出 %TGI 或肿瘤生长抑制百分比。在所有三种 KRAS 模型中，GDC-0623 (RG 7421) 和 G-573 表现出比 GDC-0623 (RG 7421) 更强的抗肿瘤活性[1]。

在 15 μL 激酶缓冲液（20 mM MOPS pH 7.2、25 mM β 甘油磷酸、5 mM EGTA、1 mM 原钒酸钠、1 mM DTT、100 μM ATP、15 mM MgCl2）中，0.14 μM 纯化的无活性重组 MEK-1 （北部）蛋白质与抑制剂一起预孵育。 30°C 孵育 10 分钟后，将 1 ng BRAF、CRAF 或 BRAF V600E 添加到反应中，总体积为 20 μL。还添加了 0.5 μg 无活性的重组 ERK2。在 30°C 孵育 30 分钟后，添加 Laemmle 样品缓冲液来终止反应。磷酸-MEK 水平的 SDS-PAGE 分析提供了酶活性的指示。 SuperSignal West Pico 化学发光底物可实现免疫反应蛋白的可视化。

QuickChange 定点诱变试剂盒用于产生 Flag-MEK1 的 S212P 和 S212A 突变体。 HCT116 细胞表达来自哺乳动物表达载体的 N 末端带有 Flag 标签的 MEK-1。第二天，将 1.8×106 HCT116 细胞铺板于 10 cm 板中，并使用 lipofectamine 2000 用 17 g 表达构建体转染细胞。 48 小时后，收获细胞并在用抑制剂处理指定时间后在 100 L 细胞提取缓冲液中裂解。 SDS-PAGE 用于检查每个样品的细胞裂解物。将膜与磷酸-MEK S221、磷酸-ERK1/2 和磷酸-MEK1 一抗一起孵育后，使用 SuperSignal West Pico 化学发光底物检查免疫反应蛋白。

Colo205 xenografts are created by injecting 6–8 week old female nude (nu/nu) mice with 5×106 cells resuspended in Hank's Balanced Salt Solution (HBSS) subcutaneously (s.c.) into the rear right flank. In order to create NCI-H2122 xenografts, 6–8 week old female nu/nu mice are injected with 1×107 cells that have been resuspended in Hank's Balanced Salt Solution (HBSS) and matrigel (growth factor reduced). In order to start an A375 or MiaPaca-2 xenograft, 1 mm3 tumor fragments from the corresponding passaged tumors are injected subcutaneously (s.c.) into the flank of athymic nu/nu mice. When tumors have grown to approximately 200 mm3, mice are randomized and treated with either vehicle (methylcellulose 0.1% tween 80 0.1% (MCT)), GDC-0973 (at 10 mg/kg), GDC-0623 (RG 7421) (at 40 mg/kg), or G-573 (at 100 mg/kg). All MEK inhibitor doses corresponded to maximally tolerated doses (MTDs), which did not cause weight loss of more than 15% to 20% of body weight. Using digital calipers and the equation (L×W×W)/2, tumor volumes are calculated. Tumor growth inhibition (%TGI) is calculated as a proportion of the area under the fitted curve (AUC) for the corresponding dose group daily in comparison to the vehicle. Animal weights are measured twice a week, and mice are taken out of the study if they lose ≥20% or more of their body weight. Complete responses (CRs) are defined as any tumor demonstrating a 100% reduction in tumor volume at any point during the study, whereas partial responses (PRs) are defined as any tumor demonstrating a ≥ 50% decrease in tumor volume.

[1]. Mechanism of MEK inhibition determines efficacy in mutant KRAS- versus BRAF-driven cancers. Nature. 2013 Sep 12;501(7466):232-6.

[2]. Elucidating the Mechanisms of Formation for Two Unusual Cytochrome P450-Mediated Fused Ring Metabolites of GDC-0623, a MAPK/ERK Kinase Inhibitor. Drug Metab Dispos. 2015 Dec;43(12):1929-1933.

GDC-0623 is a member of the class of imidazopyridines that is imidazo[1,5-a]pyridine substituted by (2-fluoro-4-iodophenyl)amino and (2-hydroxyethoxy)aminoacyl groups at positions 5 and 6. It is a potent ATP non-competitive inhibitor of MEK1 (Ki = 0.13nM) and also has efficacy against both mutant BRAF and mutant KRAS. It is in clinical development for treatment of patients with locally advanced or metastatic solid tumors. It has a role as an EC 2.7.12.2 (mitogen-activated protein kinase kinase) inhibitor, an antineoplastic agent and an apoptosis inducer. It is a substituted aniline, a member of monofluorobenzenes, an organoiodine compound, an imidazopyridine, a secondary amino compound, a hydroxamic acid ester and a primary alcohol.
GDC-0623 has been used in trials studying the treatment of Solid Cancers.
MEK Inhibitor GDC-0623 is an orally active, selective MEK inhibitor with potential antineoplastic activity. MEK inhibitor GDC-0623 specifically inhibits mitogen-activated protein kinase kinase (MEK or MAP/ERK kinase), resulting in inhibition of growth factor-mediated cell signaling and tumor cell proliferation. MEK 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.

C16H14FIN4O3

456.21

456.009

C, 42.12; H, 3.09; F, 4.16; I, 27.82; N, 12.28; O, 10.52

1168091-68-6

42642654

Light yellow to gray solid powder

1.8±0.1 g/cm3

1.703

4.16

87.89

3

6

6

25

461

0

IC1C=CC(=C(C=1)F)NC1=C(C(NOCCO)=O)C=CC2=CN=CN12

RFWVETIZUQEJEF-UHFFFAOYSA-N

InChI=1S/C16H14FIN4O3/c17-13-7-10(18)1-4-14(13)20-15-12(16(24)21-25-6-5-23)3-2-11-8-19-9-22(11)15/h1-4,7-9,20,23H,5-6H2,(H,21,24)

5-(2-fluoro-4-iodoanilino)-N-(2-hydroxyethoxy)imidazo[1,5-a]pyridine-6-carboxamide

G-868; GDC 0623; G868; GDC 0632; GDC0632; G 868

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 (5.48 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 (5.48 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: Methylcellulose 0.1% tween 80 0.1% (MCT): 5mg/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。