Small Molecule Inhibitors Of Glycogen Synthase Kinase-3 Research Articles

Advances in the Therapeutic Potential of Inhibitors Targeting Glycogen Synthase Kinase 3 in Inflammatory Diseases.

Glycogen synthase kinase-3 (GSK3) is one of the important serine/threonine protein kinases and has two isoforms, namely, GSK3α and GSK3β. GSK3 inhibits glycogen synthase activity through phosphorylation. It plays a key role in various pathophysiological processes, such as differentiation, immunity, metabolism, cell death, and cell survival. Therefore, GSK3 has evolved as an important therapeutic target for treating neurological diseases, inflammatory diseases, and cancer. In addition, GSK3 regulates inflammatory processes through NF-κB-induced expression of various cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6. Moreover, GSK3 is reported to participate in many signaling pathways related to disease pathology, including PI3K/Akt, Wnt, Hedgehog, cyclic adenosine monophosphate, mitogen-activated protein kinase, and transforming growth factor-β (TGF-β). GSK3 has become a therapeutic target against some inflammatory diseases, including the inclusion body myositis, sepsis, and inflammatory bowel disease. Hence, several GSK3 inhibitors have been under evaluation as new therapeutic strategies in recent years. Two drugs targeting GSK3 have already entered clinical studies, including tideglusib and lithium carbonate. In this study, we analyzed nearly 30 different small-molecule GSK3 inhibitors reported in the past 4 years and classified them into four categories (thiazole, pyridine, F-substituted benzene, and others) according to their structure to conduct further literature research. Moreover, we summarized the optimal compounds and described the process of transformation from the lead compound to the optimal compound. In addition, we aimed to summarize the role of GSK3 in the pathogenesis of inflammatory diseases, with insights into the recent progress in the discovery of GSK3 inhibitors.

Elucidation of the GSK3α Structure Informs the Design of Novel, Paralog-Selective Inhibitors.

Glycogen synthase kinase 3 (GSK3) remains a therapeutic target of interest for diverse clinical indications. However, one hurdle in the development of small molecule GSK3 inhibitors has been safety concerns related to pan-inhibition of both GSK3 paralogs, leading to activation of the Wnt/β-catenin pathway and potential for aberrant cell proliferation. Development of GSK3α or GSK3β paralog-selective inhibitors that could offer an improved safety profile has been reported but further advancement has been hampered by the lack of structural information for GSK3α. Here we report for the first time the crystal structure for GSK3α, both in apo form and bound to a paralog-selective inhibitor. Taking advantage of this new structural information, we describe the design and in vitro testing of novel compounds with up to ∼37-fold selectivity for GSK3α over GSK3β with favorable drug-like properties. Furthermore, using chemoproteomics, we confirm that acute inhibition of GSK3α can lower tau phosphorylation at disease-relevant sites in vivo, with a high degree of selectivity over GSK3β and other kinases. Altogether, our studies advance prior efforts to develop GSK3 inhibitors by describing GSK3α structure and novel GSK3α inhibitors with improved selectivity, potency, and activity in disease-relevant systems.

Regulation of Phosphorylation of Glycogen Synthase Kinase 3α and the Correlation with Sperm Motility in Human.

To unravel the mechanism regulating the phosphorylation of glycogen synthase kinase 3 (GSK3) and the correlation between the inhibitory phosphorylation of GSK3α and sperm motility in human. The phosphorylation and priming phosphorylated substrate-specific kinase activity of GSK3 were examined in human spermatozoa with various motility conditions. In human spermatozoa, GSK3α/β was localized in the head, midpiece, and principal piece of tail and p-GSK3α(Ser21) was enriched in the midpiece. The ratio of p-GSK3α(Ser21)/GSK3α was positively coupled with normal sperm motility criteria of World Health Organization. In high-motility spermatozoa, p-GSK3α(Ser21) phosphotyrosine (p-Tyr) proteins but p-GSK3α(Tyr279) markedly increased together with decreased kinase activity of GSK3 after incubation in Ca2+ containing medium. In high-motility spermatozoa, p-GSK3α(Ser21) levels were negatively coupled with kinase activity of GSK3, and which was deregulated in low-motility spermatozoa. In high-motility spermatozoa, 6-bromo-indirubin-3'-oxime, an inhibitor of kinase activity of GSK3 increased p-GSK3α(Ser21) and p-Tyr proteins. p-GSK3α(Ser21) and p-Tyr protein levels were decreased by inhibition of PKA and Akt. Calyculin A, a protein phosphatase-1/2A inhibitor, markedly increased the p-GSK3α(Ser21) and p-Tyr proteins, and significantly increased the motility of low-motility human spermatozoa. Down regulation of kinase activity of GSK3α by inhibitory phosphorylation was positively coupled with human sperm motility, and which was regulated by Ca2+, PKA, Akt, and PP1. Small-molecule inhibitors of GSK3 and PP1 can be considered to potentiate human sperm motility.

WIN Symposium 2022 - Abstract Poster Presentations.

WIN Symposium 2022 - Abstract Poster Presentations.

Glycogen kinase 3 inhibitor nanoformulation as an alternative strategy to inhibit PD-1 immune checkpoint

Immune checkpoint inhibition with antibodies targeting the programmed cell death-1 (PD-1) pathway is a frontline cancer immunotherapy. Driven by the limited response rates and high off-target toxicity associated to monoclonal antibodies, small molecule inhibitors of PD-1 are under active investigation. Glycogen synthase kinase 3 (GSK3) is an up-stream regulator of PD-1 and small molecule GSK3 inhibitors have been shown to effectively reduce T-cell expression of PD-1 receptors. Towards harnessing the potent anticancer effects of GSK3 inhibition, we report here on the development of a nanoformulation within PEG-PLGA nanoparticles of the small molecule GSK3 inhibitor SB415286. The formulation physicochemical properties were optimised using a novel 3D printed microfluidic nanoprecipitation device and a hydrophobic ion pairing approach was used to increase the loading of the drug. The SB415286 nanoformulation efficiently inhibited PD-1 expression in chimeric antigen receptor (CAR)-T cells co-cultured with tumour cells expressing the CAR target, and improved their survival and proliferation. Treatment of the CAR-T cells with nanoformulation also increased the population of memory T-cells. The nanoformulation of small molecule inhibitor of the GSK3 pathway is a promising alternative to antibody-based checkpoint inhibition that warrants further studies.

CD28 and glycogen synthase kinase-3 (GSK-3) inhibitor combination therapy generates tumor infiltrating T-cells (TILs) with a reversal of markers for T-cell exhaustion

Abstract Immune checkpoint blockade (ICB) of inhibitory receptors such as PD-1 has revolutionized the treatment of cancer. ICB is dependent on CD28 co-receptor expression introducing the question of whether the positive co-receptor could be targeted for the reversal of T-cell exhaustion in immunotherapy. We previously showed that the serine/threonine kinase glycogen synthase kinase-3 (GSK-3) is a central regulator of programmed cell death-1 (PD-1) transcription (Taylor et al., 2016, Immunity) and that small molecule inhibitors of GSK-3 (GSK-3 SMI) are as effective as anti-PD-1 blocking antibody in the control of tumor growth (Taylor et al., 2018, Cancer Research). In this study, we now show that anti-CD28 can synergise with GSK-3α/β inhibitors to regress tumors that are resistant to anti-PD-1 immunotherapy. Combination therapy showed responses in >55% mice with an overall reduction in tumor volume of 70% in responders compared to control mice (182 mm3 vs. 611 mm3). Flow cytometry analysis of tumor infiltrating lymphocytes (TILs) shows that, while non-treated mice were enriched with TOX+TIM3+PD1+CD8+, combination therapy increased tumor infiltration of TOX-negative CD8 T cells with TIM3-PD1int CD8 phenotype. Remarkably, this reversal involved only a single injection of anti-CD28 (PV-1) and GSK-3 SMI (SB415286). These results showed that CD28/GSK-3 combination therapy greatly reduced TOX, TIM-3 and PD1markers indicative of exhaustion. Our findings offer a small molecule approach for synergy with anti-CD28 that leads to an increase tumor infiltration with TOX-negative CD8 T cells that have decrease expression of inhibitory receptors.

Proteomic Analysis Implicates Vimentin in Glioblastoma Cell Migration.

We previously showed lithium chloride (LiCl) and other inhibitors of glycogen synthase kinase-3 (GSK-3) including 6-bromo-indirubin-3-oxime (BIO), can block glioblastoma (GBM) cell migration. To investigate the mechanisms involved we used two-dimensional difference in-gel electrophoresis (2D-DIGE) and mass spectrometry to identify proteins altered after treatment of U251 GBM cells with 20 mM LiCl. Downregulation of the intermediate filament protein vimentin was the most significant change identified. Analysis of patient tumor samples revealed that vimentin is expressed abundantly in GBM, and is prognostic especially in lower grade tumors. Additionally, siRNA-mediated vimentin knockdown impaired GBM migration. Western blotting showed that treatment with LiCl or small molecule GSK-3 inhibitors led to the rapid downregulation of detergent soluble vimentin levels across a panel of GBM-derived cells. Fluorescence reactivation after photobleaching (FRAP) microscopy studies showed a significant reduction in the ability of the vimentin cytoskeleton to recover from photo-bleaching in the presence of LiCl or BIO. Biochemical studies revealed that GSK-3 and vimentin directly interact, and analysis of vimentin revealed a GSK-3 consensus phosphorylation site. We conclude that anti-migratory compounds with the ability to inhibit GSK-3 have effects on vimentin cytoskeletal dynamics, which may play a role in their anti-invasive activity.

9-ING-41, a small-molecule glycogen synthase kinase-3 inhibitor, is active in neuroblastoma.

Advanced stage neuroblastoma is a very aggressive pediatric cancer with limited treatment options and a high mortality rate. Glycogen synthase kinase-3β (GSK-3β) is a potential therapeutic target in neuroblastoma. Using immunohistochemical staining, we observed positive GSK-3β expression in 67% of human neuroblastomas (34 of 51 cases). Chemically distinct GSK-3 inhibitors (AR-A014418, TDZD-8, and 9-ING-41) suppressed the growth of neuroblastoma cells, whereas 9-ING-41, a clinically relevant small-molecule GSK-3β inhibitor with broad-spectrum preclinical antitumor activity, being the most potent. Inhibition of GSK-3 resulted in a decreased expression of the antiapoptotic molecule XIAP and an increase in neuroblastoma cell apoptosis. Mouse xenograft studies showed that the combination of clinically relevant doses of CPT-11 and 9-ING-41 led to greater antitumor effect than was observed with either agent alone. These data support the inclusion of patients with advanced neuroblastoma in clinical studies of 9-ING-41, especially in combination with CPT-11.

Radiosynthesis and in Vivo Evaluation of [11C]A1070722, a High Affinity GSK-3 PET Tracer in Primate Brain.

Dysfunction of glycogen synthase kinase 3 (GSK-3) is implicated in the etiology of Alzheimer's disease, Parkinson's disease, diabetes, pain, and cancer. A radiotracer for functional positron emission tomography (PET) imaging could be used to study the kinase in brain disorders and to facilitate the development of small molecule inhibitors of GSK-3 for treatment. At present, there is no target-specific or validated PET tracer available for the in vivo monitoring of GSK-3. We radiolabeled the small molecule inhibitor [11C]1-(7-methoxy- quinolin-4-yl)-3-(6-(trifluoromethyl)pyridin-2-yl)urea ([11C]A1070722) with high affinity to GSK-3 (Ki = 0.6 nM) in excellent radiochemical yield. PET imaging experiments in anesthetized vervet/African green monkey exhibited that [11C]A1070722 penetrated the blood-brain barrier (BBB) and accumulated in brain regions, with highest radioactivity binding in frontal cortex followed by parietal cortex and anterior cingulate, and with the lowest bindings found in caudate, putamen, and thalamus, similarly to the known distribution of GSK-3 in human brain. Our studies suggest that [11C]A1070722 can be a potential PET radiotracer for the in vivo quantification of GSK-3 in brain.

Small-molecule GSK-3 inhibitor rescued apoptosis and neurodegeneration in anesthetics-injured dorsal root ganglion neurons.

Application of general anesthetics may induce neurotoxicity in dorsal root ganglia (DRG) neurons. In this study, we examined the possible protective mechanism and associated signaling pathways of small-molecule glycogen synthase kinase-3 (GSK-3) inhibitor, SB216763, in bupivacaine-injured mouse DRG neurons in vitro. In vitro DRG explant of 6-week old mice was treated with 5mM bupivacaine to induce neurotoxicity. The explants were also pre-treated with SB216763 for 72h. Neural protection of SB216763 on bupivacaine-injured DRG neurons was investigated by TUNEL assay, neurite outgrowth assay and western blot assay, respectively. Possible downstream gene of GSK-3 signaling pathway, protein kinase C (PKC) was knocked down by siRNA in DRG explant. Its function in regulating GSK-3 inhibition induced DRG neural protection was also examined by TUNEL, neurite outgrowth and western blot assays. Pre-treatment of SB216763 significantly ameliorated bupivacaine induced apoptosis and neurite loss in DRG neurons. Western blot showed that, in addition to the decrease of phosphorylated-GSK-3 α/β protein, SB216763 increased PKC and decreased caspase-3 (Casp-3) in bupivacaine-injured DRG neurons. SiRNA-mediated PKC knockdown was able to reverse the neural protection of SB216763 in bupivacaine-injured DRG neurons. Western blot showed that PKC knockdown increased phosphorylated-GSK-3 α/β and Casp-3 protein in DRG neurons, confirming that PKC was directly involved in GSK-3-inhibition induced neural protection in DRG. GSK-3 inhibitor SB216763, through PKC, is effective in protecting anesthetics-induced neurotoxicity in DRG.

Glycogen Synthase Kinase 3 Protein Kinase Activity Is Frequently Elevated in Human Non-Small Cell Lung Carcinoma and Supports Tumour Cell Proliferation

BackgroundGlycogen synthase kinase 3 (GSK3) is a central regulator of cellular metabolism, development and growth. GSK3 activity was thought to oppose tumourigenesis, yet recent studies indicate that it may support tumour growth in some cancer types including in non-small cell lung carcinoma (NSCLC). We examined the undefined role of GSK3 protein kinase activity in tissue from human NSCLC.MethodsThe expression and protein kinase activity of GSK3 was determined in 29 fresh frozen samples of human NSCLC and patient-matched normal lung tissue by quantitative immunoassay and western blotting for the phosphorylation of three distinct GSK3 substrates in situ (glycogen synthase, RelA and CRMP-2). The proliferation and sensitivity to the small-molecule GSK3 inhibitor; CHIR99021, of NSCLC cell lines (Hcc193, H1975, PC9 and A549) and non-neoplastic type II pneumocytes was further assessed in adherent culture.ResultsExpression and protein kinase activity of GSK3 was elevated in 41% of human NSCLC samples when compared to patient-matched control tissue. Phosphorylation of GSK3α/β at the inhibitory S21/9 residue was a poor biomarker for activity in tumour samples. The GSK3 inhibitor, CHIR99021 dose-dependently reduced the proliferation of three NSCLC cell lines yet was ineffective against type II pneumocytes.ConclusionNSCLC tumours with elevated GSK3 protein kinase activity may have evolved dependence on the kinase for sustained growth. Our results provide further important rationale for exploring the use of GSK3 inhibitors in treating NSCLC.

The negative cell cycle regulators, p27Kip1, p18Ink4c, and GSK-3, play critical role in maintaining quiescence of adult human pancreatic β-cells and restrict their ability to proliferate

Adult human pancreatic β-cells are primarily quiescent (G0) yet the mechanisms controlling their quiescence are poorly understood. Here, we demonstrate, by immunofluorescence and confocal microscopy, abundant levels of the critical negative cell cycle regulators, p27(Kip1) and p18(Ink4c), 2 key members of cyclin-dependent kinase (CDK) inhibitor family, and glycogen synthase kinase-3 (GSK-3), a serine-threonine protein kinase, in islet β-cells of adult human pancreatic tissue. Our data show that p27(Kip1) localizes primarily in β-cell nuclei, whereas, p18(Ink4c) is mostly present in β-cell cytosol. Additionally, p-p27(S10), a phosphorylated form of p27(Kip1), which was shown to interact with and to sequester cyclinD-CDK4/6 in the cytoplasm, is present in substantial amounts in β-cell cytosol. Our immunofluorescence analysis displays similar distribution pattern of p27(Kip1), p-p27(S10), p18(Ink4c) and GSK-3 in islet β-cells of adult mouse pancreatic tissue. We demonstrate marked interaction of p27(Kip1) with cyclin D3, an abundant D-type cyclin in adult human islets, and vice versa as well as with its cognate kinase partners, CDK4 and CDK6. Likewise, we show marked interaction of p18(Ink4c) with CDK4. The data collectively suggest that inhibition of CDK function by p27(Kip1) and p18(Ink4c) contributes to human β-cell quiescence. Consistent with this, we have found by BrdU incorporation assay that combined treatments of small molecule GSK-3 inhibitor and mitogen/s lead to elevated proliferation of human β-cells, which is caused partly due to p27(Kip1) downregulation. The results altogether suggest that ex vivo expansion of human β-cells is achievable via increased proliferation for β-cell replacement therapy in diabetes.

Efficient Derivation of Lateral Plate and Paraxial Mesoderm Subtypes from Human Embryonic Stem Cells Through GSKi-Mediated Differentiation

The vertebrae mesoderm is a source of cells that forms a variety of tissues, including the heart, vasculature, and blood. Consequently, the derivation of various mesoderm-specific cell types from human embryonic stem cells (hESCs) has attracted the interest of many investigators owing to their therapeutic potential in clinical applications. However, the need for efficient and reliable methods of differentiation into mesoderm lineage cell types remains a significant challenge. Here, we demonstrated that inhibition of glycogen synthase kinase-3 (GSK-3) is an essential first step toward efficient generation of the mesoderm. Under chemically defined conditions without additional growth factors/cytokines, short-term GSK inhibitor (GSKi) treatment effectively drives differentiation of hESCs into the primitive streak (PS), which can potentially commit toward the mesoderm when further supplemented with bone morphogenetic protein 4. Further analysis confirmed that the PS-like cells derived from GSKi treatment are bipotential, being able to specify toward the endoderm as well. Our findings suggest that the bipotential, PS/mesendoderm-like cell population exists only at the initial stages of GSK-3 inhibition, whereas long-term inhibition results in an endodermal fate. Lastly, we demonstrated that our differentiation approach could efficiently generate lateral plate (CD34(+)KDR(+)) and paraxial (CD34(-)PDGFRα(+)) mesoderm subsets that can be further differentiated along the endothelial and smooth muscle lineages, respectively. In conclusion, our study presents a unique approach for generating early mesoderm progenitors in a chemically directed fashion through the use of small-molecule GSK-3 inhibitor, which may be useful for future applications in regenerative medicine.

Anti-obesity effects of 3-hydroxychromone derivative, a novel small-molecule inhibitor of glycogen synthase kinase-3

Glycogen synthase kinase 3 (GSK-3) plays a central role in cellular energy metabolism, and dysregulation of GSK-3 activity is implicated in a variety of metabolic disorders, including obesity, type 2 diabetes, and cancer. Hence, GSK-3 has emerged as an attractive target molecule for the treatment of metabolic disorders. Therefore, this research focused on identification and characterization of a novel small-molecule GSK-3 inhibitor. Compound 1a, a structure based on 3-hydroxychromone bearing isothiazolidine-1,1-dione, was identified from chemical library as a highly potent GSK-3 inhibitor. An in vitro kinase assay utilizing a panel of kinases demonstrated that compound 1a strongly inhibits GSK-3β. The potential effects of compound 1a on the inactivation of GSK-3 were confirmed in human liver HepG2 and human embryonic kidney HEK293 cells. Stabilization of glycogen synthase and β-catenin, which are direct targets of GSK-3, by compound 1a was assessed in comparison with two other GSK-3 inhibitors: LiCl and SB-415286. In mouse 3T3-L1 preadipocytes, compound 1a markedly blocked adipocyte differentiation. Consistently, intraperitoneal administration of compound 1a to diet-induced obese mice significantly ameliorated their key symptoms such as body weight gain, increased adiposity, dyslipidemia, and hepatic steatosis due to the marked reduction of whole-body lipid level. In vitro and in vivo effects were accompanied by upregulation of β-catenin stability and downregulation of the expression of several critical genes related to lipid metabolism. From these results, it can be concluded that compound 1a, a novel small-molecule inhibitor of GSK-3, has potential as a new class of therapeutic agent for obesity treatment.

Small-Molecule Inhibitors of GSK-3: Structural Insights and Their Application to Alzheimer's Disease Models

The world health organization (WHO) estimated that 18 million people are struck by Alzheimer's disease (AD). The USA, France, Germany, and other countries launched major programmes targeting the identification of risk factors, the improvement of caretaking, and fundamental research aiming to postpone the onset of AD. The glycogen synthase kinase 3 (GSK-3) is implicated in multiple cellular processes and has been linked to the pathogenesis of several diseases including diabetes mellitus, cancer, and AD. Inhibition of GSK-3 leads to neuroprotective effects, decreased β-amyloid production, and a reduction in tau hyperphosphorylation, which are all associated with AD. Various classes of small molecule GSK-3 inhibitors have been published in patents and original publications. Herein, we present a comprehensive summary of small molecules reported to interact with GSK-3. We illustrate the interactions of the inhibitors with the active site. Furthermore, we refer to the biological characterisation in terms of activity and selectivity for GSK-3, elucidate in vivo studies and pre-/clinical trials.

Evidence for Irreversible Inhibition of Glycogen Synthase Kinase-3β by Tideglusib

Tideglusib is a GSK-3 inhibitor currently in phase II clinical trials for the treatment of Alzheimer disease and progressive supranuclear palsy. Sustained oral administration of the compound to a variety of animal models decreases Tau hyperphosphorylation, lowers brain amyloid plaque load, improves learning and memory, and prevents neuronal loss. We report here that tideglusib inhibits GSK-3β irreversibly, as demonstrated by the lack of recovery in enzyme function after the unbound drug has been removed from the reaction medium and the fact that its dissociation rate constant is non-significantly different from zero. Such irreversibility may explain the non-competitive inhibition pattern with respect to ATP shown by tideglusib and perhaps other structurally related compounds. The replacement of Cys-199 by an Ala residue in the enzyme seems to increase the dissociation rate, although the drug retains its inhibitory activity with decreased potency and long residence time. In addition, tideglusib failed to inhibit a series of kinases that contain a Cys homologous to Cys-199 in their active site, suggesting that its inhibition of GSK-3β obeys to a specific mechanism and is not a consequence of nonspecific reactivity. Results obtained with [(35)S]tideglusib do not support unequivocally the existence of a covalent bond between the drug and GSK-3β. The irreversibility of the inhibition and the very low protein turnover rate observed for the enzyme are particularly relevant from a pharmacological perspective and could have significant implications on its therapeutic potential.

Identification of a Maleimide-Based Glycogen Synthase Kinase-3 (GSK-3) Inhibitor, BIP-135, That Prolongs the Median Survival Time of Δ7 SMA KO Mouse Model of Spinal Muscular Atrophy

The discovery of upregulated glycogen synthase kinase-3 (GSK-3) in various pathological conditions has led to the development of a host of chemically diverse small molecule GSK-3 inhibitors, such as BIP-135. GSK-3 inhibition emerged as an alternative therapeutic target for treating spinal muscular atrophy (SMA) when a number of GSK-3 inhibitors were shown to elevate survival motor neuron (SMN) levels in vitro and to rescue motor neurons when their intrinsic SMN level was diminished by SMN-specific short hairpin RNA (shRNA). Despite their cellular potency, the in vivo efficacy of GSK-3 inhibitors has yet to be evaluated in an animal model of SMA. Herein, we disclose that a potent and reasonably selective GSK-3 inhibitor, namely BIP-135, was tested in a transgenic Δ7 SMA KO mouse model of SMA, and found to prolong the median survival of these animals. In addition, this compound was shown to elevate the SMN protein level in SMA patient-derived fibroblast cells as determined by western blot, and was neuroprotective in a cell-based, SMA-related model of oxidative stress-induced neurodegeneration.

β-Catenin Enhances Oct-4 Activity and Reinforces Pluripotency through a TCF-Independent Mechanism

Understanding the mechanisms regulating pluripotency in embryonic and induced pluripotent stem cells is required to ensure their safe use in clinical applications. Glycogen synthase kinase-3 (GSK-3) has emerged as an important regulator of pluripotency, based primarily on studies with small-molecule GSK-3 inhibitors. Here, we use mouse embryonic stem cells (ESCs) lacking GSK-3 to demonstrate that a single GSK-3 substrate, β-catenin, controls the ability of ESCs to exit the pluripotent state and to differentiate into neurectoderm. Unexpectedly, the effects of β-catenin on pluripotency do not appear to be dependent on TCF-mediated signaling, based on experiments utilizing a β-catenin C-terminal truncation mutant or highly efficient dominant-negative TCF strategies. Alternatively, we find that stabilized β-catenin forms a complex with and enhances the activity of Oct-4, a core component of the transcriptional network regulating pluripotency. Collectively, our data suggest previously underappreciated, divergent TCF-dependent and TCF-independent roles for β-catenin in ESCs.

Small molecule GSK-3 inhibitors increase neurogenesis of human neural progenitor cells

Human neural progenitor cells provide a source for cell replacement therapy to treat neurodegenerative diseases. Therefore, there is great interest in mechanisms and tools to direct the fate of multipotent progenitor cells during their differentiation to increase the yield of a desired cell type. We tested small molecule inhibitors of glycogen synthase kinase-3 (GSK-3) for their functionality and their influence on neurogenesis using the human neural progenitor cell line ReNcell VM. Here we report the enhancement of neurogenesis of human neural progenitor cells by treatment with GSK-3 inhibitors. We tested different small molecule inhibitors of GSK-3 i.e. LiCl, sodium–valproate, kenpaullone, indirubin-3-monoxime and SB-216763 for their ability to inhibit GSK-3 in human neural progenitor cells. The highest in situ GSK-3 inhibitory effect of the drugs was found for kenpaullone and SB-216763. Accordingly, kenpaullone and SB-216763 were the only drugs tested in this study to stimulate the Wnt/β-catenin pathway that is antagonized by GSK-3. Analysis of human neural progenitor differentiation revealed an augmentation of neurogenesis by SB-216763 and kenpaullone, without changing cell cycle exit or cell survival. Small molecule inhibitors of GSK-3 enhance neurogenesis of human neural progenitor cells and may be used to direct the differentiation of neural stem and progenitor cells in therapeutic applications.

Glycogen Synthase Kinase-3 (GSK-3) Inhibitors for the Treatment of Alzheimers Disease

Originally discovered because of its role in the regulation of glucose metabolism, Glycogen Synthase Kinase-3 (GSK-3) it is now recognised as a crucial player in a diverse series of cellular processes involved in Alzheimer's disease (AD) pathology. Besides having been identified as the major tau protein kinase, GSK-3 mediates Aβ neurotoxicity, plays an essential role in synaptic plasticity and memory, might be involved in Aβ formation, and it has an important role in inflammation and neuronal survival, all key features of AD neuropathology. Moreover, AD was one of the earliest disorders linked to GSK-3 dysfunction. Thus, the discovery of small molecule GSK-3 inhibitors has attracted significant attention to the protein both as therapeutic target for the therapeutic intervention in neurodegenerative diseases as well as a means to understand the molecular basis of these disorders.