Exploring Novel\nGSK-3β Inhibitors for Anti-Neuroinflammatory\nand Neuroprotective Effects: Synthesis, Crystallography, Computational\nAnalysis, and Biological Evaluation

In the pathogenesis of Alzheimer’s disease, the\noverexpression\nof glycogen synthase kinase-3β (GSK-3β) stands out due\nto its multifaced nature, as it contributes to the promotion of amyloid\nβ and tau protein accumulation, as well as neuroinflammatory\nprocesses. Therefore, in the present study, we have designed, synthesized,\nand evaluated a new series of GSK-3β inhibitors based on the <i>N</i>-(pyridin-2-yl)cyclopropanecarboxamide scaffold. We identified\ncompound <b>36</b>, demonstrating an IC<sub>50</sub> of 70 nM\nagainst GSK-3β. Subsequently, through crystallography studies\nand quantum mechanical analysis, we elucidated its binding mode and\nidentified the structural features crucial for interactions with the\nactive site of GSK-3β, thereby understanding its inhibitory\npotency. Compound <b>36</b> was effective in the cellular model\nof hyperphosphorylated tau-induced neurodegeneration, where it restored\ncell viability after okadaic acid treatment and showed anti-inflammatory\nactivity in the LPS model, significantly reducing NO, IL-6, and TNF-α\nrelease. In ADME-tox in vitro studies, we confirmed the beneficial\nprofile of <b>36,</b> including high permeability in PAMPA (<i>Pe</i> equals 9.4) and high metabolic stability in HLMs as well\nas lack of significant interactions with isoforms of the CYP enzymes\nand lack of considerable cytotoxicity on selected cell lines (IC<sub>50</sub> > 100 μM on HT-22 cells and 89.3 μM on BV-2\ncells). Based on promising pharmacological activities and favorable\nADME-tox properties, compound <b>36</b> may be considered a\npromising candidate for in vivo research as well as constitute a reliable\nstarting point for further studies.