In silico and in vitro evaluation of risedronate as a DKK1 antagonist: Implications for neuroprotection through...

Dickkopf-1 (DKK1) mediated dysregulation of the Wnt/β-catenin signaling pathway contributes to synaptic loss in Alzheimer's disease (AD). Risedronate, a third-generation bisphosphonate, shows promise in pathologies related to the Wnt pathway, although its direct interaction with the DKK1-LRP6 complex has yet to be investigated. This study aimed to characterize risedronate as a novel small-molecule DKK1 antagonist and evaluate its capacity to restore canonical Wnt signaling and provide neuroprotection. In silico studies were performed using the Schrödinger Maestro (v2018-3) platform, utilizing molecular docking, molecular simualtion, SiteMap and WaterMap analysis to define the thermodynamic hydration energetics of the DKK1-LRP6 interface. In vitro validation was conducted using LRP6-overexpressing HEK-293T cells, with Wnt modulation quantified via immunofluorescence of membrane-associated DKK1 and β-catenin stabilization assays and neuroprotective assay in presence of glutamate in SHSY5Y cells. In silico analysis identified a high-affinity binding site on the DKK1 C-terminal domain (Dscore: 0.998). WaterMap analysis and simulation study revealed that risedronate binding is driven by the displacement of high-energy "unfavorable" water molecules (ΔG > 0) at the Arg203/Lys222 residue cluster. In HEK-293LRP6 cells, risedronate (IC₅₀: 65.94 μM) significantly attenuated DKK1-mediated LRP6 sequestration (p < 0.0001), acting as a molecular "plug" to restore β-catenin signaling. This restoration successfully suggests a potential modulation of downstream of Wnt pathway restoration. Risedronate demonstrates potential inhibitory effects on DKK1 in in vitro and in silico models. By bridging bone pharmacology and neurobiology, this study provides a mechanistically grounded repurposing strategy to counteract Wnt signaling failure in neurodegenerative disorders.