Modulation of cell growth and metabolism by natural and synthetic iminosugars in intestinal tumor cell lines

Metabolic and Structural Impact of Food Iminosugars and Synthetic Derivatives in Colon Cancer Cell Lines T. Montebugloni¹, C. Grootaert², A. Bordoni¹,³, A. Rajkovic², S. De Maeseneire⁴, J. Van Camp², M. L. De Mol⁴ ¹ Department of Agricultural and Food Sciences, University of Bologna, Cesena, Italy ² Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering, Ghent University, Ghent, Belgium ³ Interdepartmental Centre of Agrifood Industry Research, University of Bologna, Cesena, Italy ⁴ Department of Biotechnology, Faculty of Bioscience Engineering, Ghent University, Ghent, Belgium Introduction. Iminosugars are sugar analogs where nitrogen replaces oxygen in the ring, conferring glycosidase inhibition properties that contribute to anti-hyperlipidemic and anti-hyperglycemic effects. Found in mulberries, buckwheat, rice, and fermented vegetables, they have been highlighted as lead candidates for the treatment of a various diseases, including cancer. Despite their therapeutic potential, their role in cancer therapy remains underexplored. Objectives. This study aimed to evaluate the anticancer potential of six iminosugars (1-Deoxynojirimycin, N-nonyl-deoxynojirimycin, Miglitol, 2,3,4,6-Tetra-O-benzyl-(+)-1-deoxynojirimycin, Fagomine, Isofagomine) by applying in vitro screening tools. Methods. Caco-2 and HCT-116 2D cell lines cultured in DMEM with or without glucose were treated with the iminosugars. Cell growth and behavior were monitored for 2 or 13 days in real-time using Incucyte, followed by a cytotoxicity assay. For 3D HCT-116 spheroids, cells were cultured in DMEM with glucose, monitored via Incucyte for 13 days, and analyzed with the Seahorse XF Analyzer for bioenergetic assessment. AdipoRed staining was used to quantify lipid content in 2D cultures, while PAS staining assessed glycogen distribution. In 3D cultures, the Seahorse XF Analyzer was used for metabolic profiling. Results. Iminosugars reduced Caco-2 and HCT-116 cell viability, with effects depending on the iminosugar type, glucose concentration, and culture architecture. HCT-116 spheroids showed unchanged basal respiration but exhibited reduced spare mitochondrial capacity and increased lactate production. Substrate oxidation assays indicated enhanced lipid oxidation following synthetic iminosugar treatment. Microscopy analysis revealed reduced lipid storage in Caco-2, but not HCT-116, after N-nonyl-deoxynojirimycin, 2,3,4,6-Tetra-O-benzyl-(+)-1-deoxynojirimycin, and Fagomine treatment. Both natural and synthetic iminosugars altered intracellular glycogen distribution. Conclusions. Using colorectal cancer models, this study evidenced the different mechanisms of action of natural and synthetic iminosugars, and highlighted the possibility of discriminating them using in vitro screening tools. Acknowledgements. We thank the Fonds Wetenschappelijk Onderzoek (FWO-Flanders) for funding the Incucyte SX5 (FWO-WEAVE G000123N) and Ghent University’s Special Research Fund for the Seahorse equipment (BOF.BAS.2020.0035.01).