Berberine inhibits gastric cancer progression by targeting canonical Wnt/β-catenin signaling in AGS cells.

Gastric cancer (GC) progression is strongly associated with dysregulated proliferation, apoptosis resistance, epithelial-mesenchymal transition, invasion, and aberrant Wnt/β-catenin signaling. Berberine has shown anti-cancer activity in several tumor models; however, its mechanistic effects on Wnt/β-catenin-driven GC phenotypes require further clarification. This study investigated whether berberine suppresses AGS GC cell progression through inhibition of canonical Wnt/β-catenin signaling. AGS cells were treated with berberine at low, medium, and high concentrations, with CHIR99021 used for Wnt/β-catenin pathway activation and XAV939 as a pathway inhibitory control. Cell viability, proliferation, colony formation, cell cycle distribution, apoptosis, migration, invasion, EMT marker expression, β-catenin localization, TOP/FOP luciferase activity, ROS/RNS production, mitochondrial membrane potential, and glucose uptake were assessed using functional, molecular, and fluorescence-based assays. CTNNB1 knockdown was used to validate β-catenin-dependent mechanisms. Berberine reduced AGS cell viability, EdU incorporation, clonogenic survival, migration, invasion, and glucose uptake while inducing G0/G1 arrest, mitochondrial depolarization, oxidative/nitrosative stress, caspase activation, and apoptosis. Berberine increased E-cadherin and decreased N-cadherin, Vimentin, Snail, Slug, ZEB1, and MMP9, indicating suppression of EMT. Mechanistically, berberine reduced β-catenin expression, nuclear localization, TOP/FOP transcriptional activity, and downstream Wnt targets including CCND1, MYC, BIRC5, AXIN2, and MMP9. CHIR99021 partially reversed berberine-induced anti-tumor effects, whereas XAV939 and CTNNB1 knockdown mimicked or enhanced berberine activity. Berberine suppresses AGS GC cell progression by inhibiting canonical Wnt/β-catenin signaling, thereby reducing proliferation, EMT, invasion, metabolic activity, and survival while promoting cell cycle arrest and apoptosis.