Neuroprotective Effect of JGK-263 in Transgenic SOD1-G93A Mice Model of Amyotrophic Lateral Sclerosis

Abstract

Introduction: Glycogen synthase kinase-3β (GSK-3β) activity plays a central role and has been identified as one of the important pathogenic mechanisms in the regulation of a wide range of cellular functions, and additionally, GSK-3β inhibitor has been investigated as a modulator of apoptosis and has been shown to confer significant protective effects on cell death in various neurodegenerative diseases. Especially, GSK-3β inhibitors have been shown to prolong motor neuron survival and suppress disease progression of amyotrophic lateral sclerosis (ALS). In this study, we evaluated the neuroprotective effects of a new GSK-3β inhibitor, JGK-263, on ALS in G93A superoxide dismutase 1 (SOD1) transgenic mice as a promising novel therapeutics.

Method: Before in vivo study, the biochemical efficacy of a new GSK-3β inhibitor, JGK-263, was evaluated via treating mouse motor neuron-like hybrid cells (NSC-34) with low to high doses of JGK-263 after 48 h of serum withdrawal. And biochemical changes of the apoptotic pathway components, including cleaved caspase-3, cytochrome c, Fas, Fas-associated protein with death domain (FADD), and cleaved caspase-8 were measured. Then JGK-263 was administered orally to 93 transgenic mice with the human G93A-mutated SOD1 gene. The study was designed into three groups: a group administered 20 mg/kg JGK-263, a group administered 50 mg/kg JGK-263, and a control group not administered with JGK-263. Clinical status, rotarod test, onset of symptom and survival of transgenic mice with ALS were evaluated. In addition, sixteen mice from each group were selected for further biochemical studies of motor neuron count, apoptosis, and cell survival signals.

Results: An in vitro study on NSC-34 motor neuron revealed that treatment with JGK-263 reduced serum-deprivation-induced motor neuronal apoptosis, The cell viability was peaked after treatment of serum-deprived cells with 50 μM JGK-263. In vivo study in the SOD1-G93A transgenic mice, JGK-263 administration remarkably improved motor function and prolonged the time until symptom onset, and rotarod failure, and survival in transgenic SOD1-G93A mice with ALS compared to control mice. In both 20 mg/kg and 50 mg/kg JGK-263 treated groups, choline acetyltransferase (ChAT) staining in the ventral horn of the lower lumbar spinal cord showed a large number of viable motor neurons, suggesting normal morphology. The neuroprotective effects of JGK-263 in ALS mice were also identified by western blot analysis of spinal cord tissues in transgenic ALS mice. The apoptosis signals were reduced by JGK-263 administration in G93A transgenic ALS mice. By contrast, the levels of phosophoinositide 3-kinase (PI3K) p85 and phosphorylated Akt (p-Akt), both of which are cell survival markers, in the spinal cords were elevated in G93A transgenic ALS mices treated with JGK-263 compared to control mice

Conclusions: These studies suggest that JGK-263, a novel oral GSK-3β inhibitor, has a neuroprotective effect through modulation of the apoptosis and cell survival pathway in motor neuron degeneration of the NSC-34 cells and SOD1-G93A transgenic mice. Therefore the JGK-263 administration could be a promising therapeutic strategy for ALS.