In vivo base editing rescues liver pathophysiology and peroxisome dysfunction in a mouse model of Zellweger spectrum...

Zellweger spectrum disorder (ZSD) is caused by biallelic loss-of-function variants in PEX genes required for peroxisome biogenesis, which is critical for normal cellular metabolism and signalling. The PEX1-p.G843D (c.2528G>A) allele, present in approximately 30% of individuals with ZSD, frequently results in chronic liver disease that can progress to cirrhosis, hepatocellular carcinoma and degraded neurological health. Here we report the development and application of an adenine base editing strategy to correct an established homozygous Pex1-p.G844D ZSD mouse model that manifests liver pathologies and metabolic dysfunction found in patients. Through intravenous delivery of AAV9 encoding ABE8e-V106W into both neonatal and 4-week-old mice, we achieved up to 60% pathogenic allele correction in the bulk liver. By restoring peroxisome function, base editing eliminated bulk accumulation of very long-chain and branched-chain fatty acids, and toxic C27-bile acid intermediates. Increased levels of phytanic acid, a branched-chain fatty acid that becomes harmful when accumulated, were normalized in blood, liver and brain tissue. Treatment of homozygous Pex1-p.G844D mice resulted in the progressive, dose-dependent normalization of liver transcriptomes and histopathology, accompanied by gains in body weight. Non-viral lipid nanoparticle delivery of ABE8e-V106W mRNA to 4-week-old mice also yielded correction of the Pex1-p.G844D allele in 27% of bulk liver cells. In patient-derived fibroblasts, base editing corrected >80% of PEX1-p.G843D alleles and restored peroxisome homeostasis. Genome-wide experimental and computational off-target analyses found minimal off-target editing in the mouse or human genome. Collectively, these findings suggest that liver base editing over a range of ages may benefit individuals with ZSD and provides a foundation for developing precision gene correction treatments that address the root cause of a wide range of peroxisomal disorders.