Wnt/β-catenin; c-Met (IC50 = 0.13 nM)

卡马替尼 (INCB28060) 在约 4 nM 的剂量和约 1 nM 的 IC50 值下抑制 c-MET 磷酸化。超过90%的c-MET被它抑制。这是可逆的，去除该物质数小时后，效果会大大减弱。 48 小时后，完全消失[1]。 Capmatinib (INCB28060)（0–1000 nM；72 小时）抑制 SNU-5、S114、H441 和 U-87MG 增殖 [1]。卡马替尼 (INCB28060) 有效抑制 c-MET 和 c-MET 通路下游效应器（包括 ERK1/2、AKT、FAK、GAB1 和 STAT3/5）的磷酸化（0.06-62.25 nM；2 小时）[1 ]。抑制剂卡马替尼 (INCB28060)（0.24-63 nM；过夜）可阻止 HGF 诱导的 H441 细胞迁移 [1]。 Capmatinib (INCB28060) 快速抑制 EGFR 和 HER-3 磷酸化（0.5–50 nM；20 分钟）[1]。在 SNU-5 细胞中，卡马替尼 (INCB28060)（0-333 nM；24 小时）会导致细胞凋亡 [1]。

卡马替尼 (INCB28060)（1-30 mg/kg；口服，每天两次，持续两周）显示出剂量依赖性的肿瘤发展减少，并且在治疗期间的所有剂量下均具有良好的耐受性，没有证据表明存在明显的毒性或体重减轻U-87MG肿瘤小鼠模型[1]。 Capmatinib (INCB28060)（0.03-10 mg/kg；口服，单剂量）可抑制 S114 肿瘤小鼠模型中的 c-MET 磷酸化 [1]。

测定缓冲液的成分如下：pH 7.8、50 mM Tris-HCl、10 mM MgCl2、100 mM NaCl、0.1 mg/ml BSA 和 5 mM DTT。在 HTS 的 384 孔板上点样的是溶解在 DMSO 中的 0.8 μL 5 mM INCB28060。根据DMSO滴定，4%的溶剂浓度是可以耐受的最高浓度。 INCB28060 板通过三个和十一个点的连续稀释来制备，以测量 IC50。从 INCB28060 板中转移 0.8 μL INCB28060 的 DMSO 溶液。 DMSO 的终浓度为 2%。在测定缓冲液中，制备 0.5 nM 磷酸化 c-Met 或 8 nM 非磷酸化 c-Met 溶液。在含有 400 μM ATP（未磷酸化 c-Met）或 160 uM ATP（磷酸化 c-Met）的测定缓冲液中，将溶解在 DMSO 中的肽底物生物素-EQEDEPEGDYFEWLE-酰胺的 1 mM 储备液稀释至 1 μM。要开始反应，向每个板的相应孔中添加 20 μL 体积的酶溶液（或酶空白的测定缓冲液）后，每孔添加 20 μL 底物溶液。将板在 25°C 避光条件下孵育 90 分钟。为了终止反应，引入 20 μL 包含 45 mM EDTA、50 mM Tris-HCl、50 mM NaCl、0.4 mg/ml BSA、200 nM SA-APC 和 3 nM EUPy20 的混合物。将板在室温下孵育 15-30 分钟后，Perkin Elmer Fusion α-FP 仪器测量均质时间分辨荧光 (HTRF)。使用以下 HTRF 程序设置：330/30 主激励滤波器，主窗口 200 uSec，主延迟 50 uSec，总共 15 次闪烁。读板时间：2000

细胞活力测定[1]
细胞类型： SNU-5、S114、H441 和 U-87MG
测试浓度： 0-10000 nM
孵育持续时间：72小时
实验结果：抑制SNU-5和S114 H441和U-87MG的细胞活力和集落形成IC50 值分别为 1.2 nM、12.4 nM、~0.5 nM 和 2 nM。

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细胞迁移测定[1]
细胞类型： H441（用 50 ng/mL 重组人 HGF 刺激 24 小时）
测试浓度：0.24、1、4、16 和 63 nM
孵育时间：过夜
实验结果：防止 HGF 刺激H441细胞迁移，IC50约2nM，细胞迁移为16nM。

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蛋白质印迹分析[1]
细胞类型： SNU-5
测试浓度： 0.06、0.24、0.98、3.91、15.63和 62.25 nM
孵育时间： 2 小时
实验结果： 有效抑制 c-MET 和下游效应子的磷酸化c-MET 通路，例如 ERK1/2、AKT、FAK、GAB1 和 STAT3/5。

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蛋白质印迹分析[1]
细胞类型： H1993 细胞
测试浓度： 0.5、5 和 50 nM
孵育时间：20分钟
实验结果：快速抑制EGFR和HER-的磷酸化

Animal/Disease Models: Female Balb/c nu/nu (nude) mice (subcutaneously (sc) (sc) inoculated with 5×106 U-87MG glioblastoma cells) [1]
Doses: 1, 3, 10 and 30 mg/kg
Route of Administration: Orally, daily Two times for 2 weeks.
Experimental Results: 1 mg/kg and 3 mg/kg one time/day had a dose-dependent inhibitory effect on tumor growth, which was 35% and 76% respectively; among 10 U-87MG tumor-bearing mice, Six animals experienced partial regression after taking the 10 mg/kg daily dose; and all doses were well tolerated during treatment, with no evidence of significant toxicity or weight loss.

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Animal/Disease Models: Female Balb/c nu/nu (nude) mice (subcutaneously (sc) (sc) inoculated with 4×106 S114 tumor cells) [1]
Doses: 0.03, 0.1, 0.3, 1, 3 and 10 mg/kg
Doses: po (po (oral gavage)) single dose
Experimental Results: Causes approximately 50% and 90% inhibition of c-MET phosphorylation 30 minutes after administration of 0.03 and 0.3 mg/kg, and more than 90% inhibition of phosphorylation of c-MET after 7 hrs (hrs (hours)).

Absorption
The oral bioavailability of capmatinib is estimated to be >70%. Following oral administration, maximum plasma concentrations are achieved within 1 to 2 hours (Tmax). Co-administration with a high-fat meal increased capmatinib AUC by 46% with no change in Cmax (as compared to fasted conditions), and co-administration with a low-fat meal had no clinically meaningful effects on exposure.

Route of Elimination
Following oral administration of radiolabeled capmatinib, approximately 78% of the radioactivity is recovered in feces, of which ~42% is unchanged parent drug, and 22% is recovered in the urine, of which a negligible amount remains unchanged parent drug.

Volume of Distribution
The apparent volume of distribution at steady-state is 164 L.

Clearance
The mean apparent clearance of capmatinib at steady-state is 24 L/h.

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Metabolism / Metabolites
Capmatinib undergoes metabolism primarily via CYP3A4 and aldehyde oxidase. Specific biotransformation pathways and metabolic products have yet to be elucidated.

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Biological Half-Life
The elimination half-life is 6.5 hours.

Hepatotoxicity
In the prelicensure clinical trials of capmatinib in patients with solid tumors harboring MET mutations, liver test abnormalities were frequent although usually self-limited and mild. Some degree of ALT elevations arose in 39% of capmatinib treated patients and were above 5 times the upper limit of normal (ULN) in 7%. In these trials that enrolled 373 patients, capmatinib was discontinued early due to increased AST or ALT in only 1% of patients. The liver test abnormalities had a median onset of 2 months after initiation of therapy. While serum aminotransferase elevations were occasionally quite high (5 to 20 times upper limit of normal), there were no accompanying elevations in serum bilirubin and no patient developed clinically apparent liver injury with jaundice. The product label for capmatinib recommends monitoring for routine liver tests before, at 2 week intervals during the first 3 months of therapy, and monthly thereafter as clinically indicated.
Likelihood score: E* (unproven but suspected rare cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the clinical use of capmatinib during breastfeeding. Because capmatinib is 96% bound to plasma proteins, the amount in milk is likely to be low. The manufacturer recommends that breastfeeding be discontinued during capmatinib therapy and for 1 week after the last dose.

◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.

◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date. Plasma protein binding is approximately 96% and is independent of drug serum concentration.

[1]. A novel kinase inhibitor, INCB28060, blocks c-MET-dependent signaling, neoplastic activities, and cross-talk with EGFR and HER-3. Clin Cancer Res. 2011 Nov 15;17(22):7127-38.

[2]. Capmatinib (INC280) Is Active Against Models of Non-Small Cell Lung Cancer and Other Cancer Types with Defined Mechanisms of MET Activation. Clin Cancer Res. 2019 May 15;25(10):3164-3175.

[3]. Dhillon S. Capmatinib: First Approval. Drugs. 2020 Jul;80(11):1125-1131.

Purpose: The c-MET receptor tyrosine kinase plays important roles in the formation, progression, and dissemination of human cancer and presents an attractive therapeutic target. This study describes the preclinical characterization of INCB28060, a novel inhibitor of c-MET kinase.
Experimental design: Studies were conducted using a series of in vitro and in vivo biochemical and biological experiments.
Results: INCB28060 exhibits picomolar enzymatic potency and is highly specific for c-MET with more than 10,000-fold selectivity over a large panel of human kinases. This inhibitor potently blocks c-MET phosphorylation and activation of its key downstream effectors in c-MET-dependent tumor cell lines. As a result, INCB28060 potently inhibits c-MET-dependent tumor cell proliferation and migration and effectively induces apoptosis in vitro. Oral dosing of INCB28060 results in time- and dose-dependent inhibition of c-MET phosphorylation and tumor growth in c-MET-driven mouse tumor models, and the inhibitor is well tolerated at doses that achieve complete tumor inhibition. In a further exploration of potential interactions between c-MET and other signaling pathways, we found that activated c-MET positively regulates the activity of epidermal growth factor receptors (EGFR) and HER-3, as well as expression of their ligands. These effects are reversed with INCB28060 treatment. Finally, we confirmed that circulating hepatocyte growth factor levels are significantly elevated in patients with various cancers.
Conclusions: Activated c-MET has pleiotropic effects on multiple cancer-promoting signaling pathways and may play a critical role in driving tumor cell growth and survival. INCB28060 is a potent and selective c-MET kinase inhibitor that may have therapeutic potential in cancer treatment.[1]

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Purpose: The selective MET inhibitor capmatinib is being investigated in multiple clinical trials, both as a single agent and in combination. Here, we describe the preclinical data of capmatinib, which supported the clinical biomarker strategy for rational patient selection.
Experimental design: The selectivity and cellular activity of capmatinib were assessed in large cellular screening panels. Antitumor efficacy was quantified in a large set of cell line- or patient-derived xenograft models, testing single-agent or combination treatment depending on the genomic profile of the respective models.
Results: Capmatinib was found to be highly selective for MET over other kinases. It was active against cancer models that are characterized by MET amplification, marked MET overexpression, MET exon 14 skipping mutations, or MET activation via expression of the ligand hepatocyte growth factor (HGF). In cancer models where MET is the dominant oncogenic driver, anticancer activity could be further enhanced by combination treatments, for example, by the addition of apoptosis-inducing BH3 mimetics. The combinations of capmatinib and other kinase inhibitors resulted in enhanced anticancer activity against models where MET activation co-occurred with other oncogenic drivers, for example EGFR activating mutations.
Conclusions: Activity of capmatinib in preclinical models is associated with a small number of plausible genomic features. The low fraction of cancer models that respond to capmatinib as a single agent suggests that the implementation of patient selection strategies based on these biomarkers is critical for clinical development. Capmatinib is also a rational combination partner for other kinase inhibitors to combat MET-driven resistance.[2]

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Capmatinib (Tabrecta™) is an oral, small molecule mesenchymal-epithelial transition (MET) inhibitor being developed by Novartis Oncology, under a license from Incyte Corporation, for the treatment of lung cancer. Capmatinib targets and selectively binds to MET, including the mutant variant produced by exon 14 skipping, and inhibits cancer cell growth driven by the mutant MET variant. In May 2020, oral capmatinib received its first global approval in the USA for the treatment of adults with metastatic non-small cell lung cancer (NSCLC) whose tumours have a mutation that leads to MET exon 14 skipping, as detected by an FDA-approved test. Clinical development for the treatment of glioblastoma, liver cancer, malignant melanoma, breast cancer, colorectal cancer, head and neck cancer and solid tumours is ongoing in several countries. This article summarizes the milestones in the development of capmatinib leading to its first approval.[3]

C23H18CLFN6O

448.880026340485

448.121

1029714-89-3

Capmatinib;1029712-80-8;Capmatinib dihydrochloride hydrate;1865733-40-9;Capmatinib dihydrochloride;1197376-85-4

137347172

Solid

85.1Ų

2

6

4

32

637

0

JJBXRCLAAKZSNF-UHFFFAOYSA-N

InChI=1S/C23H17FN6O.ClH/c1-25-22(31)18-6-5-16(11-19(18)24)21-13-28-23-27-12-17(30(23)29-21)10-14-4-7-20-15(9-14)3-2-8-26-20;/h2-9,11-13H,10H2,1H3,(H,25,31);1H

2-fluoro-N-methyl-4-[7-(quinolin-6-ylmethyl)imidazo[1,2-b][1,2,4]triazin-2-yl]benzamide;hydrochloride

NVP-INC 280AAA; INCB 028060; Capmatinib hydrochloride; INCB28060; INC280; INC-280; Capmatinib HCl; Capmatinib xHCl; 1029714-89-3; 2-Fluoro-N-methyl-4-(7-(quinolin-6-ylmethyl)imidazo[1,2-b][1,2,4]triazin-2-yl)benzamide hydrochloride; 2126164-56-3; INC 280; INCB028060; INCB-028060; INCB-28060; INCB 28060.

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)

DMSO: > 10 mM

注意: 如下所列的是一些常用的体内动物实验溶解配方，主要用于溶解难溶或不溶于水的产品（水溶度<1 mg/mL）。 建议您先取少量样品进行尝试，如该配方可行，再根据实验需求增加样品量。

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注射用配方1: DMSO : Tween 80： Saline = 10 : 5 : 85 (如： 100 μL DMSO → 50 μL Tween 80 → 850 μL Saline)
*生理盐水/Saline的制备：将0.9g氯化钠/NaCl溶解在100 mL ddH ₂ O中，得到澄清溶液。
注射用配方 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL DMSO → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)
注射用配方 3: DMSO : Corn oil = 10 : 90 (如： 100 μL DMSO → 900 μL Corn oil)
示例: 以注射用配方 3 (DMSO : Corn oil = 10 : 90) 为例说明, 如果要配制 1 mL 2.5 mg/mL的工作液, 您可以取 100 μL 25 mg/mL 澄清的 DMSO 储备液，加到 900 μL Corn oil/玉米油中, 混合均匀。

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注射用配方 4: DMSO : 20% SBE-β-CD in Saline = 10 : 90 [如：100 μL DMSO → 900 μL (20% SBE-β-CD in Saline)]
*20% SBE-β-CD in Saline的制备（4°C，储存1周）：将2g SBE-β-CD (磺丁基-β-环糊精) 溶解于10mL生理盐水中，得到澄清溶液。
注射用配方 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (如： 500 μL 2-Hydroxypropyl-β-cyclodextrin (羟丙基环胡精)→ 500 μL Saline)
注射用配方 6: DMSO : PEG300 : Castor oil : Saline = 5 : 10 : 20 : 65 (如： 50 μL DMSO → 100 μL PEG300 → 200 μL Castor oil → 650 μL Saline)
注射用配方 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (如： 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
注射用配方 8: 溶解于Cremophor/Ethanol (50 : 50), 然后用生理盐水稀释。
注射用配方 9: EtOH : Corn oil = 10 : 90 (如： 100 μL EtOH → 900 μL Corn oil)
注射用配方 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL EtOH → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)

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口服配方 1: 悬浮于0.5% CMC Na (羧甲基纤维素钠)
口服配方 2: 悬浮于0.5% Carboxymethyl cellulose (羧甲基纤维素)
示例: 以口服配方 1 (悬浮于 0.5% CMC Na)为例说明, 如果要配制 100 mL 2.5 mg/mL 的工作液, 您可以先取0.5g CMC Na并将其溶解于100mL ddH2O中，得到0.5%CMC-Na澄清溶液；然后将250 mg待测化合物加到100 mL前述 0.5%CMC Na溶液中，得到悬浮液。

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口服配方 3: 溶解于 PEG400 (聚乙二醇400)
口服配方 4: 悬浮于0.2% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 5: 溶解于0.25% Tween 80 and 0.5% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 6: 做成粉末与食物混合

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注意: 以上为较为常见方法，仅供参考， InvivoChem并未独立验证这些配方的准确性。具体溶剂的选择首先应参照文献已报道溶解方法、配方或剂型，对于某些尚未有文献报道溶解方法的化合物，需通过前期实验来确定（建议先取少量样品进行尝试），包括产品的溶解情况、梯度设置、动物的耐受性等。

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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。