Introduction

The intestinal epithelium as dynamic cellular interface integrates signals from the microbial and metabolic environment to sustain tissue homeostasis. We established a clear mechanistic link between mitochondrial dysfunction of the epithelium and the development and progression of inflammatory bowel diseases (IBD). Several clinical trials targeting disease conditions associated with impaired mitochondrial function use mitochondria-targeted pharmaceutical molecules. Thus, we hypothesize that mitochondria-protective targets can be repurposed for adjuvant IBD therapy. We aim to characterize the effect of 10 metabolic drugs targeting various mitochondrial functions on IBD patient-derived organoids, as well as IBD mouse and pig models.

Aims & Methods

Cellular toxicity of the drugs Acipimox, AICAR, ALCAR, Bezafibrate, Epi-743, Idebenone, Metformin, SS-31 and TUDCA was assessed in the intestinal epithelial cell line Mode-K and murine wild-type (WT) small intestinal (SI) organoids. Furthermore, intestinal organoids from TNF^ΔARE mice, a model for Crohn's Disease (CD) like ileitis, were used to mimic IBD-related inflammatory conditions and treated with a predefined concentration of all drugs. Additionally, the corresponding TNF^ΔARE pig model and organoids generated from resected tissue of CD patients were treated with the compounds. Epithelial stemness and differentiation were analysed via imaging. Drug-related changes on mitochondrial respiration were assessed using respirometry.

Results

Cytotoxicity tests were performed in cell and WT organoid cultures to determine suitable concentration ranges for further treatment studies. Exposure of cells to Metformin inhibited cellular respiration, while Idebenone bypassed complex I inhibition and increased ATP production by 57 %. Furthermore, the 10 tested drugs differentially mediated concentration-dependent effects on seeding efficiency in murine WT organoids. To assess the rescuing capacity of the drugs in an inflammatory context, the predefined concentrations of all 10 compounds were applied to SI organoids generated from the inflamed ileum of TNF^ΔARE mice. Three drugs were excluded from further experiments based on reduced organoid seeding efficiency and de novo crypt formation. In comparison, the remaining seven drugs either did not influence organoid growth and differentiation or were able to increase them. Those seven compounds demonstrated similar results using crypts isolated from acutely inflamed ileal tissue sections of the TNF^ΔARE pig model with Epi-743, Idebenone and SS-31 showing a significant increase in de novo crypt formation. Those three drugs did not affect seeding efficiency and de novo crypt formation of human CD organoids from the resection border, while Acipimox, ALCAR, Bezafibrate and Metformin significantly reduced them. Treatment of strongly inflamed parts of resected CD patient tissue specimens with Epi-743, Idebenone and SS-31 restored growth and differentiation capacity in seeded crypts, indicating an increased stemness upon mitochondrial drug treatment. Those drugs will be tested in vivo in the DSS, TNF^ΔARE and the metabolic injury mouse model Hsp60^Δ/ΔIEC; Il10^-/- as well as the TNF^ΔARE pig model.

Conclusion

Selected drugs targeting mitochondrial functions restored seeding efficiency, differentiation, and stemness of intestinal organoids from inflamed tissue sections of TNF^ΔARE mice, TNF^ΔARE pigs, and CD patients. These findings clearly support the potential of metabolic drug repurposing for adjuvant IBD therapy and indicate putative risks for Metformin.