Genetic Correction of the Most Common Mutation Causing Primary Hyperoxaluria Restores Enzyme Localization and Oxalate Metabolism.

Our research aimed to model primary hyperoxaluria type 1 in vitro using a stem cell model and assess the potential of adenine base editors in correcting the most common pathogenic AGXT genetic variant, c.508G>A (Gly170Arg), which leads to oxalate accumulation due to alanine-glyoxylate aminotransferase mislocalization. Patient-derived fibroblasts were induced to pluripotent stem cells, genetically corrected with adenine base editing, and subsequently differentiated into hepatocyte-like cells in parallel with their non-corrected isogenic counterparts. Enzyme localization was assessed through immunocytochemistry and confocal microscopy. The key metabolites associated with the disease were analyzed using liquid chromatography-mass spectrometry to evaluate the metabolic phenotype. Finally, lipid nanoparticle formulations were designed and tested as an in vivo-applicable delivery method for base editors. All induced pluripotent stem cell lines successfully differentiated into hepatocyte-like cells and expressed essential hepatocyte markers, including ALB, HNF1A, and AGXT. Adenine base editor-mediated genetic correction of the pathogenic AGXT mutation restored enzyme localization into peroxisomes and diminished oxalate accumulation without significant off-target effects. Base editor mRNA and AGXT variant targeting single guide RNA encapsulated within lipid nanoparticles mediated gene correction in the hepatocyte-like cell model. Using an in vitro model of primary hyperoxaluria type 1, we showed that base editor-mediated genetic correction of the most common hyperoxaluria-causing variant corrects enzyme mislocalization from mitochondria to peroxisomes and improves metabolic function. These results propose gene correction as a potential therapeutic approach to hyperoxaluria.