iPSC-Derived Liver Organoids as a Tool to Study Medium Chain Acyl-CoA Dehydrogenase Deficiency

Medium chain acyl-CoA dehydrogenase deficiency (MCADD) is an inherited metabolic disease, characterized by biallelic variants in the ACADM gene. Interestingly, even with the same genotype, patients often present with very heterogeneous symptoms, ranging from fully asymptomatic to life-threatening hypoketotic hypoglycemia. The mechanisms underlying this heterogeneity remain unclear. Therefore, there is a need for in vitro models of MCADD that recapitulate the clinical phenotype as a tool to study the pathophysiology of the disease. Fibroblasts of control and symptomatic MCADD patients with the c.985A>G (p.K329E) were reprogrammed into induced pluripotent stem cells (iPSCs). iPSCs were then differentiated into hepatic expandable organoids (EHOs), further matured to Mat-EHOs, and functionally characterized. EHOs and Mat-EHOs performed typical hepatic metabolic functions, such as albumin and urea production. The organoids metabolized fatty acids, as confirmed by acyl-carnitine profiling and high-resolution respirometry. MCAD protein was fully ablated in MCADD organoids, in agreement with the instability of the mutated MCAD protein. MCADD organoids accumulated medium-chain acyl-carnitines, with a strongly elevated C8/C10 ratio, characteristic of the biochemical phenotype of the disease. Notably, C2 and C14 acyl-carnitines were found decreased in MCADD Mat-EHOs. Finally, MCADD organoids exhibited differential expression of genes involved in omega-oxidation, mitochondrial beta-oxidation, TCA cycle, and peroxisomal coenzyme A metabolism, particularly upregulation of NUDT7. iPSC-derived organoids of MCADD patients recapitulated the major biochemical phenotype of the disease. Mat-EHOs expressed relevant pathways involved in putative compensatory mechanisms, notably CoA metabolism and the TCA cycle. The upregulation of NUDT7 expression may play a role in preventing excessive accumulation of dicarboxylic acids in MCADD. This patient-specific hepatic organoid system is a promising platform to study the phenotypic heterogeneity between MCADD patients.