CP-91149

399, 87

Obese, diabetic male C57BL/6J-Lep(ob/ob) mice and their lean, nondiabetic C57BL/6J- /+ littermates

Dissolved in DMSO, final concentrations ca.50 uM

Formulated in vehicle consisting of either 0.25% (wt/vol) methyl cellulose in water or 0.1% Pluronic P105 Block Copolymer Surfactant in 0.1% saline

CP-91149 displays 200-fold higher inhibitory activity against human liver glycogen phosphorylase a (HLGPa) than caffeine (IC50 = 26 uM). CP-91149 (10-100 uM) inhibits glucagon-stimulated glycogenolysis in isolated rat hepatocytes in a dose-dependent manner, and in primary human hepatocytes with IC50 of ca.2.1 uM. [1] CP-91149 also potently inhibits the activities of human muscle phosphorylase a and b with IC50 of 0.2 uM and ca.0.3 uM, respectively. CP-91149 treatment at 2.5 uM induces inactivation of phosphorylase and sequential activation of glycogen synthase in hepatocytes, and increases glycogen synthesis by 7-fold at 5 mM glucose and by 2-fold at 20 mM glucose. CP-91149 can partially counteract the effects of phosphorylase overexpression. [2] CP-91149 also potently inhibits brain GP with IC50 of 0.5 uM in A549 cells. CP-91149 treatment at 10-30 uM causes significant glycogen accumulation in A549 and HSF55 cells. CP-91149 treatment increases G1-phase cells with a significant reduction of the S-phase population in HSF55 cells, correlated with increased expression of p21 and p27. [3] CP-91149 also promotes the dephosphorylation and activation of GS (glycogen synthase) in non-engineered or GP-overexpressing cultured human muscle cells, but exclusively in glucose-deprived cells. [4]

72 hours

Cells are exposed to various concentrations of CP-91149 for 72hours. Viability is determined with manual cell counts following staining with trypan blue exclusion assay. Cells are fixed with 70% ethanol. DNA is stained with propidium iodide and the intensity of fluorescence is measured using a Becton-Dickinson flow cytometer at 488nm for excitation and at 650nm for emission. The cell cycle profile is analyzed using Modifit's Sync Wizard.

CP-91149 is a selective glycogen phosphorylase (GP) inhibitor with IC50 of 0.13 μM in the presence of glucose, 5- to 10-fold less potent in the absence of glucose.