Disruption of NANOG-driven epithelial-mesenchymal transition (EMT) and self-renewal restores drug sensitivity in colorectal cancer
Kiarash Saleki, Sameerah Shaheen, Miao Xue, Amirreza Mazloomi, Sepideh Youssefi, Hossein Kashfi, Mehreen Ahmed, Roya Babaei‐Jadidi, Bradley Spencer‐Dene, Dominique Bonnet, Chris Denning, Abdolrahman S. Nateri
Abstract
AIMS: To investigate the regulatory role of NANOG in genes associated with stemness, symmetric division, and therapeutic resistance in colorectal cancer stem-like cells (CRC-SCs), with a focus on ERK/GSK-3β/β-catenin signalling and epithelial-mesenchymal transition (EMT), in order to evaluate the translational potential of targeting NANOG-associated signalling pathways. METHODS: Stemness, signalling activity, and cell division modes were analysed using 3D colonospheres enriched for CRC-SCs. Drug responses to the MEK inhibitor U0126 and the GSK-3β inhibitor TDZD-8 were assessed in CRC patient-derived organoids (PDOs), alongside molecular assays, immunohistochemistry with H-score quantification in xenograft models, and molecular dynamics simulations. RESULTS: NANOG overexpression enhanced the expression of stemness-associated genes, promoted symmetric cell division, and activated ERK/GSK-3β signalling, contributing to increased sphere formation. Inhibition of MEK and GSK-3β reduced EMT, cell proliferation, and symmetric division in CRC-SCs. NANOG-mediated dysregulation of ERK/GSK-3β altered β-catenin signalling and disrupted E-cadherin-dependent cell-cell adhesion. Molecular simulations and drug assays demonstrated that TDZD-8 and U0126 interfere with NANOG-DNA binding and β-catenin/E-cadherin interactions. CONCLUSIONS: NANOG drives CRC-SC maintenance via ERK/GSK-3β/β-catenin signalling and EMT modulation. This study offers significant insights into the translational impact of targeting NANOG and its downstream pathways with small-molecule inhibitors U0126 and TDZD-8 and presents a promising strategy to reduce CRC-SCs stemness, functionality, and tumourigenicity.