ETP-46464

製品コードS8050 バッチS805002

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	C₃₀H₂₂N₄O₂
	分子量	470.52	CAS No.	1345675-02-6
Solubility (25°C)*	体外	DMSO	14 mg/mL (29.75 mM)
		Water	Insoluble
		Ethanol	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	ETP-46464 is a potent and selective inhibitor of ATR with IC50 of 25 nM.
in vitro	ETP-46464 shows almost full activity against ATR at 100 nM. ETP-46464 inhibits the activity of PI3Kα, mTOR and DNA-PKcs with IC50s of 170, 0.6 and 36 nM. ETP-46464 is able to promote the breakage of stalled replication forks. Exposure to ETP-46464 leads the generation of substantial amounts of DNA damage in replicating cells. 1 μM ETP-46464 abrogates the ionizing radiation-induced G2/M checkpoint and leads to the presence of micronuclei or completely fragmented nuclei in cells. ETP-46464 is particularly toxic for p53-deficient cells, which is exacerbated by replicative stress-generating conditions such as the overexpression of cyclin E. ^[1]
特徴	Selective ATR inhibitor and particularly toxic to p53-deficient cancer cells. Often used as a pharmacologic probe due to its ideal pharmacological properties.

プロトコル(参考用のみ)

キナーゼアッセイ	ATR kinase activity assay
キナーゼアッセイ	For the in vitro kinase assay flag-tagged ATR kinase is immunoprecipitated from 293T cells using Anti-flag M2 affinity gel. As a substrate, p53/1-101 peptide fused to GST is purified from IPTG stimulated BL21 bacteria by conventional GST purification methods. Briefly, pBJF-flag-ATR plasmids (wild type and kinase dead versions) is transfected into 293T cells. 48 hours after transfection, cells are lysed in TGN buffer (50 mM Tris (pH 7.5), 50 mM glycerophosphate, 150 mM NaCl, 10% glycerol, 1% tween-20 and protease inhibitors cocktail) for 15 minutes on ice and, after centrifugation, the supernatant is pre-cleared for 1h with unspecific rabbit IgG and protein A on a rotator at 4 degrees. Next, the supernatant is incubated with M2-slurry for 2 h at 4 degrees. For a confluent p100 plate of 293T cells, 400 μL of buffer, 10 μg of IgG, 30 μL of protein A slurry, and 25 μL of M2 beads are used. Anti flag M2 beads are then washed 5 times in ice cold TNG, 3 times in ice cold LiCl buffer (100mM Tris pH 7.5, 0.5 M LiCl), and 5 times in ice cold Kinase buffer (10 mM Hepes (pH 7.5), 50 mM NaCl, 10 mM MgCl₂, 10 mM Mn₂, 1 mM DTT). After the last wash, IPs are splitted in the proper number of tubes for the assay (approximately one confluent 293T p100 is used per condition). For the kinase reaction, 100 μM ‘cold’ ATP is added to the buffer. Only fresh IPs are used. Prior to every experiment a mix is prepared containing 2 μg of GST-p53 and 10 μ Ci of ³²P-γATP per condition. Inhibitor are first added to the beads. Last, 25μL of the mix is added to each tube. Reactions are performed for 20 minutes at 30 degrees and stopped by adding 15 microliters of 4 × NuPAGE SDS buffer and boiling for 5 minutes. Beads are pelleted and supernatants are resolved by SDS-PAGE on a polyacrylamide gel. Radioactive gels are slightly washed and directly exposed to Phospor-Imager screens for 2h to 16h. Radioactivity is then detected using a STORM scanner. Finally, gels are stained with coomasie to check for protein levels.

参考

[1] Toledo LI, et al. Nat Struct Mol Biol, 2011, 18(6), 721-727.

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：

Calpain-2 mediates SARS-CoV-2 entry via regulating ACE2 levels [ mBio, 2024, e0228723.]	PubMed: 38349185
A function for ataxia telangiectasia and Rad3-related (ATR-kinase in cytokinetic abscission. [ iScience, 2023, 25(7)]	PubMed: None
Increased replication origin firing links replication stress to whole chromosomal instability in human cancer [ Cell Rep, 2022, 41(11):111836]	PubMed: 36516748
A function for ataxia telangiectasia and Rad3-related (ATR) kinase in cytokinetic abscission [ iScience, 2022, 25(7):104536]	PubMed: 35754741
ATAD5 promotes replication restart by regulating RAD51 and PCNA in response to replication stress. [ Nat Commun, 2019, 10(1):5718]	PubMed: 31844045
Mild replication stress causes chromosome mis-segregation via premature centriole disengagement. [ Nat Commun, 2019, 10(1):3585]	PubMed: 31395887
Targeting the cell cycle in head and neck cancer by Chk1 inhibition: a novel concept of bimodal cell death. [ Oncogenesis, 2019, 8(7):38]	PubMed: 31209198
DNA Damage Signaling Instructs Polyploid Macrophage Fate in Granulomas [Herrtwich L Cell, 2016, 167(5):1264-1280]	PubMed: 28084216

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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