Aloxistatin-E64d-DataSheet-生命科学试剂-MedChemExpress

1、Hotline: 400-820-3792Inhibitors Agonists Screening Librarieswww.MedChemEAloxistatinCat. No.: HY-100229CAS No.: 88321-09-9Synonyms: E64d; E64c ethyl ester分式: CHNO分量: 342.43作靶点: Cathepsin作通路: Metabolic Enzyme/Protease储存式: Powder -20C 3 years4C 2 yearsIn solvent -80C 6 months-20C 1 month溶解性数据体外实验 DMSO

2、: 23 mg/mL (67.17 mM)* means soluble, but saturation unknown.Mass Solvent1 mg 5 mg 10 mg Concentration制备储备液1 mM 2.9203 mL 14.6015 mL 29.2030 mL5 mM 0.5841 mL 2.9203 mL 5.8406 mL10 mM 0.2920 mL 1.4602 mL 2.9203 mL请根据产品在不同溶剂中的溶解度，选择合适的溶剂配制储备液，并请注意储备液的保存式和期限。体内实验 请根据您的实验动物和给药式选择适当的溶解案，配制前请先配制澄清的储备液，再依次

3、添加助溶剂(为保证实验结果的可靠性，体内实验的作液，建议您现现配，当天使；澄清的储备液可以根据储存条件，适当保存；以下溶剂前的百分 指该溶剂在您配制终溶液中的体积占)：1. 请依序添加每种溶剂： 10% DMSO 40% PEG300 5% Tween-80 45% salineSolubility: 2.5 mg/mL (7.30 mM); Suspended solution2. 请依序添加每种溶剂： 10% DMSO 90% corn oilSolubility: 2.5 mg/mL (7.30 mM); Clear solution1/4 Master of Small Molecul

4、es 您边的抑制剂师www.MedChemEBIOLOGICAL ACTIVITY物活性 Aloxistatin (E64d)可渗透细胞,不可逆的谱半胱氨酸蛋酶 (cysteine protease) 抑制剂。IC50 & Target Cysteine protease 1体外研究 Inhibition of protease-resistant prion protein (PrP-res) accumulation in ScNB cells by cysteine proteaseinhibitor Aloxistatin (E64d) with IC50 of 0.50.11 M.

5、For the cell surface PrP-sen detection, PrP-sen isimmunoprecipitated from media treated with phosphatidylinositol-specific phospholipase C (PIPLC) torelease pulse-35S-labeled PrP-sen from the cell surface. Aloxistatin is maintained at 15 M, respectively, inthe labeling media of all but the control c

6、ells 1. Aloxistatin (E64d) (which specifically blocks cysteineproteases, but not serine proteases such as granzymes) is able to completely block turnover of the CatLsubstrate Z-Phe-Arg-aminomethylcoumarin, when pre-incubated with NK-92 or YT 5 cells 2. Aloxistatin(E64d) is a broad-spectrum cell-perm

7、eable inhibitor of cysteine proteases 3.体内研究 Oral administration of Aloxistatin (E64d) to guinea pigs results in a dose-dependent reduction in brain, CSFand plasma A(40) and A(42). Aloxistatin also causes a biphasic dose-dependent reduction in brain CTF.Aloxistatin causes a dose-dependent increase i

8、n brain sAPP. The mean sAPP levels are significantlyhigher than the no dose group for Aloxistatin doses of 5 mg/kg/day or greater with the highest Aloxistatindose resulting in the maximum increase in sAPP of about 54% more than the control group. Similar to theA effect, oral Aloxistatin administrati

9、on produces a biphasic dose-dependent reduction in brain cathepsin Bactivity. The minimum effective dose is about 1 mg/kg/day with the highest Aloxistatin dose causing themaximum reduction in brain cathepsin B activity of about 91% lower than that of the control group. Thus,Aloxistatin reduces guine

10、a pig brain cathepsin B activity in a manner which is consistent with the compoundinhibiting cathepsin B -secretase activity 4. Aloxistatin (E64d) inhibits the increases in the expression ofAT1AR and ACE genes in rats. Administration of Olmesartan or Aloxistatin reduces the increase in thesuperoxide

11、 production of the intramyocardial coronary arteries in HF rats 5.PROTOCOLKinase Assay 2 The CTLs and NK cells (0.8106/mL) are treated with the inhibitors L1 (10-20 M) or Aloxistatin (20-30 M)for 24 hr at 37C in 24-well plates. Cells are then used in 51Cr-release assays or are lysed to examineperfor

12、in in Western blots. The inhibitor is also added at the same concentration during the 4 hr reactions insome 51Cr-release assays, as indicated. Cell lysates are prepared using NP-40 lysis buffer (25 mM HEPES,250 mM NaCl, 2.5 mM ethylenediaminetetraacetic acid, 0.1% volume/volume Nonidet P-40) and tot

13、al proteinconcentration is determined using the Bradford assay. Equal amounts of protein are loaded and resolved on8% SDSgels. Human or mouse perforin is detected using the appropriate antibodies as indicated.Anti-actin antibody is used as a loading control 2.MCE has not independently confirmed the

14、accuracy of these methods. They are for reference only.Cell Assay 3 Cell proliferation and apoptosis are assessed by staining for a proliferation marker, Ki67, or an apoptoticmarker, cleaved caspase 3, following the protocol described above for the polarity markers. MCF10 variants2/4 Master of Small

15、 Molecules 您边的抑制剂师www.MedChemEare grown in 3D rBM overlay cultures for 4 days and are treated with 0.1 % DMSO, 5 M CA074Me or 5 MAloxistatin. The percentage of structures that are positive for Ki67 or cleaved caspase 3 is determined bycounting a total of 100 structures on two separate coverslips wit

16、h a Zeiss Axiophot epifluorescentmicroscope. Structures are considered Ki67 positive if they contained at least one cell staining for Ki67.Structures are considered to be caspase 3 positive if they contained at least one cell that is positive forcleaved caspase 3 and the positive cell(s) is not loca

17、lized in the center of a developing lumen 3.MCE has not independently confirmed the accuracy of these methods. They are for reference only.Animal Mice and Pigs 4Administration 45 Guinea Pigs (male, Hartley strain, average weight 400 g corresponding to animals about 6 weeks old) areused. Male transge

18、nic mice expressing human APP containing the wt -secretase site and the Londonmutant -secretase site sequences are used. Delivering a drug by gavage offers the advantage of accuratedosing but is traumatic and thus only suitable for relatively short dosing periods (up to about a week).Delivery by gav

19、age is used for the guinea pig studies. Aloxistatin is suspended in Me2SO at the indicatedconcentrations (0.1, 1.0, 5, and 10 mg/kg) and administered by gavage daily using a feeding tube. Vehiclecontrol animals are treated by gavage of Me2SO alone.Rats 5Male inbred DS rats are used. Weaned rats are

20、fed laboratory chow containing 0.3% NaCl until 7 weeks ofage. DS rats fed an 8% NaCl diet after 7 weeks manifest compensated concentric left ventricular (LV)hypertrophy secondary to hypertension at 12 weeks and a distinct stage of fatal LV failure with lungcongestion at 19 weeks. DS rats are therefo

21、re fed an 8% NaCl diet from 7 weeks of age and are randomizedto an HF group, an Aloxistatin group (10 mg per kg of body mass per day, administered intraperitoneallyevery other day), or an olmesartan group (3 mg/kg per day in chow) from 12 to 19 weeks of age (n=10 foreach group). The doses of olmesar

22、tan (an ARB) and Aloxistatin are determined in preliminary experimentsand previous studies. DS rats maintained on the 0.3% NaCl diet served as age-matched controls (controlgroup, n=10). At 19 weeks of age, all of the rats are euthanized by an intraperitoneal overdose of sodiumpentobarbital (50 mg/kg

23、), and the hearts are removed for biological and histological analyses. Arterial bloodis collected from the abdominal aorta for the measurement of renin activity. Systolic blood pressure and heartrate are measured in conscious rats from 7 weeks of age, every week, using a noninvasive tail-cuff metho

24、d.In separate experiments, 12-week-old DS rats, fed a low-salt diet from 7 weeks of age, are given vehicle,olmesartan, or Aloxistatin in the same manner as in the above experiments (n=5 for each group), and the LVtissues for measuring targeting mRNAs and protein levels are immediately placed in liqu

25、id nitrogen andstored at -80C.MCE has not independently confirmed the accuracy of these methods. They are for reference only.户使本产品发表的科研献 Environ Sci Technol. 2017 Dec 5;51(23):13938-13948. Toxicol Appl Pharmacol. 2018 Oct 1;356:159-171. Biochem Biophys Res Commun. 2018 Sep 3;503(1):297-303. Mol Med

26、Rep. 2019 Jan;19(1):41-50.See more customer validations on HYPERLINK / www.MedChemE3/4 Master of Small Molecules 您边的抑制剂师www.MedChemEREFERENCES1. Doh-Ura K, et al. Lysosomotropic agents and cysteine protease inhibitors inhibit scrapie-associated prion protein accumulation. J Virol.2000 May;74(10):4894-7.2. Konjar S, et al. Human and mouse perforin are processed in part through cleavage by the lysosomal cysteine proteinase cathepsin L.Immunology. 2010 O