A-078 Impaired Fluxomics in Congenital Disease of Glycosylation” to “Impaired Fluxomics in Congenital Disorders of Glycosylation

Background Congenital disorders of glycosylation (CDG) represent a complex and heterogeneous group of inherited metabolic disorders often characterized by impaired protein glycosylation. With over 170 identified types, CDG encompasses a broad spectrum of genetic mutations that affect glycosylation processes, thereby making it one of the most extensive groups of inborn errors of metabolism. Among these, phosphomannomutase 2-associated CDG (PMM2-CDG) is the most prevalent, accounting for over 50% of CDG cases in numerous populations. Recent advancements in therapeutic approaches have yielded promising outcomes for the management of CDG, particularly PMM2-CDG. Innovative strategies, including aldolase reductase inhibitors and mannose-1-phosphate-based therapies, have demonstrated potential in enhancing monosaccharide flux into the biosynthesis of N-linked glycoproteins. These emerging treatments provide hope for improvement in the quality of lives among affected children while addressing the intricate metabolic disruptions characteristic of the disorder. The application of Fluxomics, an advanced methodology for the systematic study of metabolic pathway dynamics and metabolite flow, provides a comprehensive framework for understanding the imechanisms that underlie glycosylation deficits. This approach empowers researchers to detect subtle metabolic alterations, thereby facilitating the identification of crucial biomarkers and therapeutic targets for CDGs. Methods Stable isotope labeling experiments were performed in human fibroblast cell lines to trace the flow of monosaccharide precursors and nucleotide sugar donors through glycosylation pathways into N-glycans. High-resolution mass spectrometry detected isotopic enrichments, while the isotopic ratios quantified fluxes across various pathways and into N-glycans. Additionally, comparative metabolomics revealed compensatory shifts in alternative routes, such as the hexosamine biosynthetic pathway, glycolysis, and salvage pathway. Results The study revealed significant metabolic differences between various CDG cell lines. PMM2 cells showed a marked decrease in flux through essential glycosylation steps and a partial diversion of metabolic intermediates into alternative pathways. This metabolic redirection reduced N-glycan species* flux, underscoring the metabolic challenges inherent in PMM2-CDG. In contrast, ALG13 cells exhibited a fascinating metabolic profile characterized by increased flux into N-glycan synthesis. This finding suggests an overcompensatory mechanism in cell lines with deficiencies in key glycan assembly steps, offering valuable insights into the adaptive metabolic responses of these CDG cells. Conclusion Our research highlights the transformative potential of fluxomics in clarifying the pathophysiological mechanisms that underlie CDG. By offering a comprehensive view of metabolic dynamics, this approach opens new pathways for understanding disease mechanisms, identifying possible therapeutic targets, and a novel strategy to monitor the restoration of normal glycosylation patterns.