SP6.06 Intestinal stem cell dysfunction is driven by loss of HMGCS2-mediated ketogenic response in Ulcerative colitis and represents a novel therapeutic target

Abstract Background Intestinal stem cells (ISCs) continuously divide and differentiate into multiple enterocyte lineages, a process requiring high energy. Recent evidence implicates bioenergetic failure of ISCs with non-resolving inflammation in Ulcerative Colitis (UC). Methods In treatment-naïve UC patients, we characterized mitochondrial oxidative phosphorylation (OXPHOS) in epithelial and ISCs, performed single-cell RNA sequencing (scRNA-seq), and functional metabolism with SCENITH*. Results Single-cell quantitative immunofluorescence showed loss of OXPHOS Complex I/IV in UC (n=10/group; p<0.0001) and UC-derived organoids (n=20/group; p<0.0001), suggesting bioenergetic failure transmitted from ISCs to daughter epithelial cells. scRNA-seq of ISCs from active UC revealed significant ketogenesis pathway downregulation, with HMGCS2 as the top downregulated gene (FC-2.76, p<0.0001). HMGCS2, the rate-limiting mitochondrial matrix ketogenesis enzyme, produces the ketone body βHB which, provides metabolites for the TCA cycle and has important roles in stem cell fate-signalling. Moreover, we find colonic organoids produce βHB, but this is reduced in active UC (n=5/group, p <0.001). βHB supplementation of UC organoids restores ISC function, measured via growth assay (n=5/group, p<0.05. Functionally, Functionally, βHB restored OXPHOS capacity in UC organoids using SCENITH (n=3/group, p<0.05). In UC organoids, fenofibrate (PPARα agonist), induced HMGCS2 expression (p<0.001) and βHB production (p<0.05). Finally, HMGCS2 protein expression significantly increased in responders to therapy with mucosal healing (n=4, p<0.05). Conclusions We provide the first combined evidence of mitochondrial dysfunction and ketogenesis failure (‘primary’ and ‘backup’ energy failure) in UC. Crucially, restoring ketogenesis reverses ISC pathogenic defects, opening a novel ‘immuno-metabolic’ therapeutic approach in UC. * Single Cell Energetic Metabolism by Profiling Translation Inhibition