Dual Inhibition of GSK3β and CDK5 Protects the Cytoskeleton of Neurons from Neuroinflammatory-Mediated Degeneration In Vitro and In Vivo

Abstract

SummaryNeuroinflammation is a hallmark of neurological disorders and is accompanied by the production of neurotoxic agents such as nitric oxide. We used stem cell-based phenotypic screening and identified small molecules that directly protected neurons from neuroinflammation-induced degeneration. We demonstrate that inhibition of CDK5 is involved in, but not sufficient for, neuroprotection. Instead, additional inhibition of GSK3β is required to enhance the neuroprotective effects of CDK5 inhibition, which was confirmed using short hairpin RNA-mediated knockdown of CDK5 and GSK3β. Quantitative phosphoproteomics and high-content imaging demonstrate that neurite degeneration is mediated by aberrant phosphorylation of multiple microtubule-associated proteins. Finally, we show that our hit compound protects neurons in vivo in zebrafish models of motor neuron degeneration and Alzheimer's disease. Thus, we demonstrate an overlap of CDK5 and GSK3β in mediating the regulation of the neuronal cytoskeleton and that our hit compound LDC8 represents a promising starting point for neuroprotective drugs.

Glycogen Synthase Kinase 3β in Cancer Biology and Treatment

“…Clinical trials have evaluated seed compounds for GSK3β inhibitors (e.g., tideglusib) in the treatment of Alzheimer's disease and bipolar disorder (reviewed in [15]). A recent study showed that dual inhibition of GSK3β and CDK5 protects the cytoskeleton of neurons from neuroinflammatory-mediated degeneration, a common biological characteristic of neurodegenerative disorders [246]. Co-administration of pharmacological GSK3β inhibitors prevents apoptosis of neural precursor cells and peripheral neuropathy induced by camptothecin and paclitaxel without impairing their chemotherapeutic efficacy [247,248].…”

Type 1 Interleukin-4 Signaling Obliterates Mouse Astroglia in vivo but Not in vitro

“…However, reductionist models of human diseases in appropriate organisms are quite powerful for addressing particular aspects of pathologies or for designing experimental treatment options that may defy that particular disease. For instance, zebrafish proposed many signaling pathways that could be harnessed for enhanced responses in tissue counterparts in humans (Zon, 1999;Tomasiewicz et al, 2002;Poss et al, 2003;Rubinstein, 2003;Lieschke and Currie, 2007;Newman et al, 2010;Diep et al, 2011;Kizil et al, 2012a,b;Kyritsis et al, 2012;Gemberling et al, 2013;MacRae and Peterson, 2015;Mokalled et al, 2016;Papadimitriou et al, 2018;Celikkaya et al, 2019;Cosacak et al, 2019;Reinhardt et al, 2019). Other disease models in zebrafish yielded in useful information on the pathological mechanisms and led to the development of promising drugs (Cully, 2019).…”

Repurposing cell growth-regulating compounds identifies kenpaullone which ameliorates pathologic pain via normalization of inhibitory neurotransmission

“…Using measures of Kcc2 promoter activity, Kcc2 mRNA abundance, and [Cl-]i, we conducted an unbiased screen of two NCI libraries containing 1057 compounds that inhibit growth of transformed cells. This screen identified kenpaullone (KP), a GSK3/CDK kinase inhibitor with neuroprotective properties (Liu et al, 2016;Reinhardt et al, 2019;Schultz et al, 1999;Skardelly et al, 2011;Yang et al, 2013;Zaharevitz et al, 1999). Our studies of KP revealed the following principal findings: 1) KP enhances Kcc2/KCC2 gene expression in a concentration-dependent manner and lowers [Cl-]i in cultured mouse, rat, and human neurons; 2) Systemic administration of KP to mice attenuates measures of nerve injury pain and chronic itch in preclinical models;…”

“…This concept will permit rational exploration of links between GSK3ßèd-cat and attenuated Kcc2/KCC2 gene expression and the resulting malfunction of inhibitory neurotransmission in several other relevant neurologic and psychiatric conditions such as Alzheimer's Disease and other neurodegenerative diseases, psychoses, traumatic brain/spinal cord injury, Rett Syndrome, Autism Spectrum Disorders, and epilepsy (Boulenguez et al, 2010;Chen et al, 2018;Ferando et al, 2016;Freund and Meskenaite, 1992;Huberfeld et al, 2007;Hyde et al, 2011;Kahle et al, 2014b;Tang et al, 2016;Tao et al, 2012). We selected KP because of its previously reported neuroprotective properties for spinal motoneurons, brainstem auditory relay neurons, and hypoxia-injured hippocampal neurons (Liu et al, 2016;Reinhardt et al, 2019;Skardelly et al, 2011;Teitz et al, 2018;Winkelmann et al, 2015;Yang et al, 2013). It is possible there was a unifying mechanism of Kcc2 expression enhancement by KP in these previous studies.…”

“…1A). Of these we identified kenpaullone (KP), a GSK3/CDK kinase inhibitor, as a promising compound for further study based on its previous record of neuroprotection in translationally relevant preclinical models (Liu et al, 2016;Reinhardt et al, 2019;Skardelly et al, 2011;Yang et al, 2013).…”