AZ20-DataSheet-MedChemExpress

1、Hotline: 400-820-3792Inhibitors Agonists Screening Librarieswww.MedChemEAZ20Cat. No.: HY-15557CAS No.: 1233339-22-4分式: CHNOS分量: 412.51作靶點(diǎn): ATM/ATR作通路: Cell Cycle/DNA Damage; PI3K/Akt/mTOR儲存式: Powder -20C 3 years4C 2 yearsIn solvent -80C 6 months-20C 1 month溶解性數(shù)據(jù)體外實(shí)驗(yàn) DMSO : 100 mg/mL (242.42 mM)* mea

2、ns soluble, but saturation unknown.Mass Solvent1 mg 5 mg 10 mg Concentration制備儲備液1 mM 2.4242 mL 12.1209 mL 24.2418 mL5 mM 0.4848 mL 2.4242 mL 4.8484 mL10 mM 0.2424 mL 1.2121 mL 2.4242 mL請根據(jù)產(chǎn)品在不同溶劑中的溶解度，選擇合適的溶劑配制儲備液，并請注意儲備液的保存式和期限。體內(nèi)實(shí)驗(yàn) 請根據(jù)您的實(shí)驗(yàn)動物和給藥式選擇適當(dāng)?shù)娜芙獍?，配制前請先配制澄清的儲備液，再依次添加助溶?為保證實(shí)驗(yàn)結(jié)果的可靠性，體內(nèi)實(shí)驗(yàn)的作液，

3、建議您現(xiàn)現(xiàn)配，當(dāng)天使；澄清的儲備液可以根據(jù)儲存條件，適當(dāng)保存；以下溶劑前的百分 指該溶劑在您配制終溶液中的體積占)：1. 請依序添加每種溶劑： 10% DMSO 40% PEG300 5% Tween-80 45% salineSolubility: 2.5 mg/mL (6.06 mM); Clear solution2. 請依序添加每種溶劑： 10% DMSO 90% (20% SBE-CD in saline)Solubility: 2.5 mg/mL (6.06 mM); Clear solution1/3 Master of Small Molecules 您邊的抑制劑師www.Me

4、dChemEBIOLOGICAL ACTIVITY物活性 AZ20種有效的，選擇性的 ATR 抑制劑，IC50 值為 5 nM；同時(shí)可抑制 mTOR 活性，IC50 值為 38 nM。IC50 & Target ATR mTOR PI3K5 nM (IC50) 38 nM (IC50) 13000 nM (IC50)體外研究 AZ20 inhibits ATR immunoprecipitated from HeLa nuclear extracts with an IC50 of 5 nM and ATR mediatedphosphorylation of Chk1 in HT29 col

5、orectal adenocarcinoma tumor cells with an IC50 of 50 nM 1.體內(nèi)研究 AZ20 (25, 50 mg/kg, p.o.) has high permeability combined with good stability to rat hepatocytes and, despitethe lack of progress in achieving markedly higher solubility, has respectable bioavailability in a low dose ratPK study. AZ20 (2

6、5, 50 mg/kg, p.o.) leads to significant tumor growth inhibition in female nude mice bearingLoVo tumors 1.PROTOCOLKinase Assay 1 ATR for use in the in vitro enzyme assay is obtained from HeLa nuclear extract by immunoprecipitation withrabbit polyclonal antiserum raised to amino acids 400-480 of ATR c

7、ontained in the following buffer: 25 mMHEPES (pH 7.4), 2 mM MgCl2, 250 mM NaCl, 0.5 mM EDTA, 0.1 mM Na3VO4, 10% v/v glycerol, and 0.01%v/v Tween 20. ATR-antibody complexes are isolated from nuclear extract by incubating with protein A-Sepharose beads for 1 h and then through centrifugation to recove

8、r the beads. In the well of a 96-well plate,10 L ATR-containing Sepharose beads are incubated with 1 g of substrate glutathione S-transferase-p53N66 (NH2-terminal 66 amino acids of p53 fused to glutathione S-transferase are expressed in E. coli) inATR assay buffer (50 mM HEPES (pH 7.4), 150 mM NaCl,

9、 6 mM MgCl2, 4 mM MnCl2, 0.1 mM Na3VO4, 0.1mM DTT, and 10% (v/v) glycerol) at 37C in the presence or absence of inhibitor. After 10 min with gentleshaking, ATP is added to a final concentration of 3 M and the reaction continued at 37C for an additional 1h. The reaction is stopped by addition of 100

10、L of PBS, and the reaction is transferred to a white opaqueglutathione coated 96-well plate and incubated overnight at 4C. This plate is then washed with PBS/0.05%(v/v) Tween 20, blotted dry, and analyzed by a standard ELISA technique with a phosphoserine 15 p53antibody. The detection of phosphoryla

11、ted glutathione S-transferase-p53N66 substrate is performed incombination with a goat anti-mouse horseradish peroxidase-conjugated secondary antibody. Enhancedchemiluminescence solution is used to produce a signal, and chemiluminescent detection is carried out via aTopCount plate reader. The resulti

12、ng calculated % enzyme activity is then used to determine the IC50 valuesfor the compounds (IC50 taken as the concentration at which 50% of the enzyme activity is inhibited).MCE has not independently confirmed the accuracy of these methods. They are for reference only.Cell Assay 1 Compound dose rang

13、es are created by diluting in 100% DMSO and then further into assay medium (EMEM,10% FCS, 1% glutamine) using a Labcyte Echo acoustic dispensing instrument. Cells are plated in 384-wellCostar plates at 9104 cells per mL in 40 L of EMEM, 10% FCS, 1% glutamine and grown for 24 h.Following addition of

14、compound the cells are incubated for 60 min. A final concentration of 3 M 4NQO (prepared in 100% DMSO) is then added using the Labcyte Echo, and the cells are incubated for afurther 60 min. The cells are fixed by adding 40 L of 3.7% v/v formaldehyde solution for 20 min. Afterremoval of fix, cells ar

15、e washed with PBS and permeabilized in 40 L of PBS containing 0.1% Triton X-100.2/3 Master of Small Molecules 您邊的抑制劑師www.MedChemEThe cells are then washed, and 15 L primary antibody solution (pChk1 Ser345) is added. The plates areincubated at 4C overnight. The primary antibody is then washed off, an

16、d 20 L of secondary antibodysolution and 1 M Hoechst 33258 added for 90 min at room temperature. The plates are washed and left in40 L of PBS. Plates are then read on an ArrayScan VTI instrument to determine staining intensities, anddose responses are obtained and used to determine the IC50 values f

17、or the compounds.MCE has not independently confirmed the accuracy of these methods. They are for reference only.Animal Female Swiss nu/nu mice are housed in negative pressure isolators. LoVo tumor xenografts are establishedAdministration 1 in 8- to 12-week-old mice by injecting 1107 tumor cells subc

18、utaneously (100 L in serum free medium) onthe left dorsal flank. Animals are randomized into treatment groups when tumors become palpable. AZ20 isprepared in 10% DMSO/40% propylene glycol/50% water and administered orally. Tumors are measured upto three times per week with calipers. Tumor volumes are calculated and the data plotted using the geometricmean for each group versus time.MCE has not independently confirmed the accuracy of these methods. They are for reference only.戶使本產(chǎn)品發(fā)表的科研獻(xiàn) Nat Commun. 2019 Jul 2;10(1):2910. Cell Syst. 2018 Dec. Cell Deat