sb-415286 and Neurodegenerative-Diseases

Glycogen synthase kinase-3beta (GSK-3beta) is a crucial component in the cascade of events that culminate in a range of neurodegenerative diseases. It is controlled by several pathways, including calpain-mediated cleavage. Calpain mediates in cell death induced by 3-nitropropionic acid (3-NP), but GSK-3beta regulation has not been demonstrated. Here we studied changes in total GSK-3beta protein levels and GSK-3beta phosphorylation at Ser-9 in this model. The 3-NP treatment induced GSK-3beta truncation. This regulation was dependent on calpain activation, since addition of calpeptin to the medium prevented this cleavage. While calpain inhibition prevented 3-NP-induced neuronal loss, inhibition of GSK-3beta by SB-415286 did not. Furthermore, inhibition of cdk5, a known target of calpain involved in 3-NP-induced cell death, also failed to rescue neurons in our model. Our results point to a new target of calpain and indicate possible cross-talk between calpain and GSK-3beta in the 3-NP toxicity pathway. On the basis of our findings, we propose that calpain may modulate 3-NP-induced neuronal loss.

Topics: Amino Acid Chloromethyl Ketones; Aminophenols; Animals; Calpain; Caspases; Cell Survival; Cells, Cultured; Convulsants; Disease Models, Animal; Embryo, Mammalian; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hippocampus; Male; Maleimides; Mice; Neurodegenerative Diseases; Neurons; Neuroprotective Agents; Nitro Compounds; Propionates; Purines; Rats; Roscovitine; Signal Transduction; Time Factors

Glycogen synthase kinase-3 (GSK-3) is a serine/threonine protein kinase, the activity of which is inhibited by a variety of extracellular stimuli including insulin, growth factors, cell specification factors and cell adhesion. Consequently, inhibition of GSK-3 activity has been proposed to play a role in the regulation of numerous signalling pathways that elicit pleiotropic cellular responses. This report describes the identification and characterisation of potent and selective small molecule inhibitors of GSK-3.. SB-216763 and SB-415286 are structurally distinct maleimides that inhibit GSK-3alpha in vitro, with K(i)s of 9 nM and 31 nM respectively, in an ATP competitive manner. These compounds inhibited GSK-3beta with similar potency. However, neither compound significantly inhibited any member of a panel of 24 other protein kinases. Furthermore, treatment of cells with either compound stimulated responses characteristic of extracellular stimuli that are known to inhibit GSK-3 activity. Thus, SB-216763 and SB-415286 stimulated glycogen synthesis in human liver cells and induced expression of a beta-catenin-LEF/TCF regulated reporter gene in HEK293 cells. In both cases, compound treatment was demonstrated to inhibit cellular GSK-3 activity as assessed by activation of glycogen synthase, which is a direct target of this kinase.. SB-216763 and SB-415286 are novel, potent and selective cell permeable inhibitors of GSK-3. Therefore, these compounds represent valuable pharmacological tools with which the role of GSK-3 in cellular signalling can be further elucidated. Furthermore, development of similar compounds may be of use therapeutically in disease states associated with elevated GSK-3 activity such as non-insulin dependent diabetes mellitus and neurodegenerative disease.

Topics: Adenosine Triphosphate; Aminophenols; beta Catenin; Binding, Competitive; Calcium-Calmodulin-Dependent Protein Kinases; Cell Line; Cytoskeletal Proteins; Diabetes Mellitus, Type 2; Enzyme Activation; Gene Expression Regulation; Genes, Reporter; Glycogen; Glycogen Synthase; Glycogen Synthase Kinase 3; Glycogen Synthase Kinases; Humans; Indoles; Kinetics; Liver; Maleimides; Molecular Structure; Neurodegenerative Diseases; Protein Kinases; Recombinant Proteins; Signal Transduction; Trans-Activators; Transcription, Genetic