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Selisistat (EX 527)

Selisistat (EX 527)

Manufacturer	Selleckchem
Category	Oncology Neuroscience Metabolism Diabetes Glucose Homeostasis Regulation of Appetite Molecular Targets for Neuroscience Targets for Rodent Neurosciences Inhibitors & Compound Libraries Small Molecules By Organ Lung Cancer Bladder Cancer Colorectal Cancer Breast Cancer Testicular & Prostate Cancer
Type	Inhibitors
Specific against	other
Amount	10 mM/1 ml
Item no.	S1541-10mM
Targets	SIRT1
CASRN	49843-98-3
eClass 6.1	30220300
eClass 9.0	32160605
Available
Administration	Intracerebroventricular injection
Animal Models	Male Sprague-Dawley rats
Cell lines	NCI-H460, MCF-7, U-2 OS and HMEC
Chemical Name	(S)-6-chloro-2, 3, 4, 9-tetrahydro-1H-carbazole-1-carboxamide
Concentrations	Dissolved in DMSO, final concentration 1 uM
Description	EX 527 is a potent and selective SIRT1 inhibitor with IC50 of 38 nM.
Dosages	ca.5 ug/rat
Features	EX 527 has more potency, specificity, stability, and lower toxicity than other inhibitors of SIRT1 catalytic activity identified to date.
Formulation	Dissolved in DMSO in a total volume of 5 uL
IC50	38 nM [1], 38 nM [1], 38 nM [1], 38 nM [1], 38 nM [1], 38 nM [1]
In vitro	EX 527 exhibits potently inhibitory effect against SIRT1 deacetylase activity in a concentration-dependent manner with an IC50 of 38 nM, displays much lower activity against SIRT2 and SIRT3 with IC50 values of 19.6 uM and 48.7 uM, respectively. EX 527 does not inhibit SIRT4-7 and class I/II HDAC activity at concentrations up to 100 uM. EX-527 alone (1 uM) has no detectable effect on the acetylation of p53 lysine 382 in NCI-H460 cells. EX-527 significantly increases the amount of acetylated p53 in NCI-H460 cells, human mammary epithelial cells, U-2 OS and MCF-7 cells subjected to genotoxic agents such as Etoposide, Doxorubicin, Hydroxyurea, and Hydrogen peroxide, which is more effective than that caused by Nicotinamide (5 mM). But surprisingly EX 527 does not result in detectable effects on p53-controled gene expression, cell survival, or cell proliferation. [1] EX 527 causes a 90% increase in cell number of HCT116 cells after 7 days in the condition of 0.1% serum but not 10% serum, suggesting that SIRT1 is a significant regulator of cell proliferation during growth factor deprivation conditions. [2] EX 527 abrogates resveratrol effects on glucose responses, and prevents resveratrol-induced up-regulation of Glut2, glucokinase, Pdx-1, and Tfam in INS-1E Cells, due to the opposite effect of EX 527 and resveratrol on SIRT1 deacetylase activity. [3]
In vivo	Administration of EX 527 (ca.10 ug) to rats increases hypothalamic acetyl-p53 levels by inhibiting hypothalamic SIRT1 activity. Co-administration of EX 527 with ghrelin markedly blunts the orexigenic action of ghrelin by decreasing the pAMPK levels, increasing the ACC levels, and abolishing the higher expression of the transcription factors FoxO1, pCREB, and Bsx and the neuropeptides NPY and AgRP in the hypothalamic arcuate nucleus. [4]
Incubation Time	48 or 72 hours
Kinase Assay	Inhibition of GST-SIRT1 deacetylase activity, 293T cells are transiently transfected with GST-tagged human SIRT1 in the pDEST27 Gateway vector using FuGENE-6. After 48 hours, the cells are lysed with 50 mM Tris, pH 8.0, 120 mM NaCl, 1 mM EDTA, and 0.5% Nonidet P-40, supplemented with Complete Mini protease inhibitor cocktail tablets. GST-SIRT1 is purified from lysates using glutathione-Sepharose beads and washed extensively in the above buffer. The deacetylation assay is performed with approximately 30 ng of GST-SIRT1 in the presence of EX 527 (48 pM to 100 uM). Deacetylation is measured using the Fluor de Lys kit using a fluorogenic peptide encompassing residues 379 to 382 of p53, acetylated on lysine 382. The acetylated lysine residue is coupled to an aminomethylcoumarin moiety. The peptide is deacetylated by SIRT1, followed by the addition of a proteolytic developer that releases the fluorescent aminomethylcoumarin. Briefly, enzyme preparations are incubated with 170 uM NAD+ and 100 uM p53 fluorogenic peptide for 45 minutes at 37 C followed by incubation in developer for 15 minutes at 37 C. Fluorescence is measured by excitation at 360 nm and emission at 460 nm and enzymatic activity is expressed in relative fluorescence units.
Method	For viability assays, cells are treated with EX 527 for 48 hours. Cell viability is then determined using the Cell Titer-Glo luminescent assay, which measures total ATP level as an index of cell number. Luminescence is measured on a Luminoskan Ascent. For the proliferation assay, 0.5 uCi/mL of [14C]thymidine is added to the medium immediately after EX 527. Plates are counted at 48 hours (HMEC) or 72 hours (NCI-H460, MCF-7, and U-2 OS cells) in a Microbeta liquid scintillation counter. Thymidine incorporated by the cells is detected by proximity to the scintillant in the base of the Cytostar-T tissue culture plate.
Molecular Weight (MW)	248, 71
Picture ChemicalStructure Description	EX 527 Chemical Structure
Solubility (25C)	DMSO 50 mg/mL, Water <1 mg/mL, Ethanol 18 mg/mL
Storage	2 years -20CPowder, 2 weeks4Cin DMSO, 2 months-80Cin DMSO

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Amount: 10 mM/1 ml

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Delivery expected until 5/16/2024