RAF265 (CHIR-265)

製品コードS2161 バッチS216103

化学情報

	Synonyms	N/A	Storage (From the date of receipt)	3 years -20°C powder 1 years -80°C in solvent
	化学式	C₂₄H₁₆F₆N₆O
	分子量	518.41	CAS No.	927880-90-8
Solubility (25°C)*	体外	DMSO	100 mg/mL (192.89 mM)
		Ethanol	100 mg/mL (192.89 mM)
		Water	Insoluble
* <1 mg/ml means slightly soluble or insoluble. * Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.

溶剤液（一定の濃度）を調合する

生物活性

製品説明	RAF265 (CHIR-265) is a potent selective inhibitor of C-Raf/B-Raf/B-Raf V600E with IC50 of 3-60 nM, and exhibits potent inhibition on VEGFR2 phosphorylation with EC50 of 30 nM in cell-free assays. RAF265 (CHIR-265) induces cell cycle arrest and apoptosis. Phase 2.
in vitro	RAF265 inhibits C-Raf, wild type B-Raf and mutant (V600E) B-Raf. RAF265 effectively block phosphorylation of Raf's downstream substrates MEK and ERK in cells and also kill melanoma and colorectal cancer cell lines harboring B-Raf mutations independent of PTEN mutation status. Raf kinase inhibition by RAF265 in mutant B-Raf melanoma cell lines causes cell cycle arrest and induces apoptosis, mimicking the effect of Raf RNAi in these cells. RAF265 also potently inhibits the phosphorylation of VEGFR2 and proliferation of VEGF-stimulated hMVEC. ^[1] In HT29 and MDAMB231 cells, RAF265 shows inhibitory activity with IC20 of 1 to 3 μM and IC50 of 5 to 10 μM, respectively. While RAF265 leads to a significant decrease in clonogenic survival in all tested cell lines, which means that RAF265 induces a dominant effect on clonogenic survival. Addition of RAF265 to RAD001 in HCT116 cells could lead to moderately decreased AKT, S6 protein, and 4EBP1 phosphorylation. ^[2] Raf265 markedly reduces the protein level of Bcl-2 and great inhibitory in CM- and NCI-H727 cells, while having no effect on the TRAIL susceptibility of BON1 and GOT1 cells. ^[3] Protein kinase D3 (PRKD3) that when knocked down could enhance cell killing by RAF265 in A2058 melanoma cells, which prevent reactivation of MAPK signaling, induce PARP cleavage, increase caspase activity, interrupt cell-cycle progression, and inhibit colony formation. ^[4]
in vivo	RAF265 shows 71% to 72% TVI% (tumor volume inhibition percentage) in HCT116 xenografts at 12 mg/kg. While the combination of RAF265 and RAD001 shows enhanced antitumor activity with increased T10 (time to achieve a relative tumor volume of 10 times the initial tumor volume) and tumor growth delay. The combination of RAD001 and RAF265 also significantly enhances the activation of caspase-3 in HCT116 and MDAMB231 but not in A549 xenografts. ^[2] RAF265 inhibits FDG (2-deoxy-2-[¹⁸F]fluoro-d-glucose) accumulation and decreases the tumor volumes in A375M xenografts by orally dosed of 100 mg/kg. ^[5]

プロトコル(参考用のみ)

キナーゼアッセイ	Assay Protocol
キナーゼアッセイ	Raf and Mek are combined at 2 × final concentrations in assay buffer (50 mM Tris, pH 7.5, 15 mM MgCl₂. 0.1 mM EDTA and 1 mM DTT) and dispensed 15 μL per well in polypropylene assay plates. Background levels are determined in wells containing Mek and DMSO without Raf. To the Raf/Mek containing wells is added 3 μL of 10 × of RAF265 diluted in 100% DMSO. The raf kinase activity reaction is started by the addition of 12 μL per well of 2.5 × ³³P-ATP diluted in assay buffer. After 45-60 minutes, the reactions are stopped with the addition of 70 μL of stop reagent (30 mM EDTA). Filtration plates are pre-wetted for 5 min with 70% ethanol, and then rinsed by filtration with wash buffer. Samples (90 μL) from the reaction wells are then transferred to the filtration plates. The filtration plates are washed 6 × with wash buffer using Millipore filtration apparatus. The plates are dried and 100 μL per well of scintillation fluid is added. The CPM is then determined using a Wallac Microbeta 1450 reader.
細胞アッセイ	細胞株	Human A549 and H460 lung, HT29 and HCT 116 colon, and MDAMB231 breast cancer cell lines
	濃度	0.1 - 10 μM
	反応時間	48 hours
	実験の流れ	The MTT assay and Bliss additivism model are used to assess the effect of RAF265 on cell viability. In each well of a 96-well plate, 1 × 10⁴ cells are grown in 200 μL of medium. After 24 hours, RAF265 is added to achieve a final concentration of 0.1 to 10 μM. After 48 hours of treatment, 20 μL of 5 mg/mL MTT solution in PBS is added to each well. After 4 hours, supernatant is removed and formazan crystals are discarded in 200 μL of DMSO. Absorbance is then measured at 595 nm using an absorbance plate reader. Data are expressed as the percentage of viable cells.
動物実験	動物モデル	A549, H460, HCT116, or MDAMB231 cells are injected s.c. into the flank region of 6-wk-old female athymic mice.
	投薬量	12 mg/kg
	投与方法	Orally administered daily

参考

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カスタマーフィードバック

: Data from [Pigment Cell Melanoma Res, 2014, 27(1), 124-33]

: Data from [Data independently produced by Hepatology, 2012, 56(6), 2363-74]

: Data from [Data independently produced by , , Cell Rep, 2014, 8(5):1475-83]

Selleckの高級品が、幾つかの出版された研究調査結果（以下を含む）で使われた：

Integrative analysis of drug response and clinical outcome in acute myeloid leukemia [ Cancer Cell, 2022, S1535-6108(22)00312-9]	PubMed: 35868306
Comprehensive drug response profiling and pan-omic analysis identified therapeutic candidates and prognostic biomarkers for Asian cholangiocarcinoma [ iScience, 2022, 25(10):105182]	PubMed: 36248745
Identification of New Vulnerabilities in Conjunctival Melanoma Using Image-Based High Content Drug Screening [ Cancers (Basel), 2022, 14(6)1575]	PubMed: 35326726
B-Raf inhibitor vemurafenib counteracts sulfur mustard-induced epidermal impairment through MAPK/ERK signaling [ Drug Chem Toxicol, 2022, 1-10]	PubMed: 34986718
RAF1 amplification drives a subset of bladder tumors and confers sensitivity to MAPK-directed therapeutics [ J Clin Invest, 2021, e147849]	PubMed: 34554931
RAF1 amplification drives a subset of bladder tumors and confers sensitivity to MAPK-directed therapeutics [ J Clin Invest, 2021, e147849]	PubMed: 34554931
SHOC2 phosphatase-dependent RAF dimerization mediates resistance to MEK inhibition in RAS-mutant cancers. [ Nat Commun, 2019, 10(1):2532]	PubMed: 31182717
RAF kinases are stabilized and required for dendritic cell differentiation and function. [ Cell Death Differ, 2019, 10.1038/s41418-019-0416-4]	PubMed: 31541179
Genetic Heterogeneity of BRAF Fusion Kinases in Melanoma Affects Drug Responses. [ Cell Rep, 2019, 29(3):573-588]	PubMed: 31618628
Mitochondrial metabolic reprograming via BRAF inhibition ameliorates senescence. [ Exp Gerontol, 2019, 126:110691]	PubMed: 31421186

長期の保管のために-20°Cの下で製品を保ってください。

人間や獣医の診断であるか治療的な使用のためにでない。

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