Base editing of the GLB1 gene is therapeutic in GM1 gangliosidosis patient-derived cells.

GM1 gangliosidosis is an autosomal recessive neurodegenerative lysosomal storage disease caused by pathogenic variants in the GLB1 gene, limiting the production of active lysosomal β-galactosidase. Phenotypic heterogeneity is due in part to variant type, location within GLB1, and the amount of residual enzyme activity; in the most severe form, death occurs in infancy. With no FDA approved therapeutics, development of efficacious strategies for the disease is pivotal. CRISPR/Cas based approaches have revolutionized precision medicine and have been indispensable to the development of treatments for several monogenic disorders with bespoke strategies central to current research pipelines. We used CRISPR/Cas-adenine base editing to correct the GLB1 c.380G>A (p.Cys127Tyr) variant in patient-derived dermal fibroblasts compound heterozygous with the GLB1 c.481T>G (p.Trp161Gly) pathogenic variant. Nucleofection of plasmids encoding the target sgRNA and ABEmax restored the canonical guanine (32.2 ± 2.2 % of the target allele) and synthesis of active β-galactosidase. Analysis of cellular markers of pathology revealed normalization of both primary glycoconjugate storage and lysosomal pathology. Furthermore, analysis of off-target sites nominated by the in silico tools Cas-OFFinder and/or CRISTA revealed no significant editing or indels. This study supports the use of CRISPR/Cas-based approaches for the treatment of GM1 gangliosidosis, and provides foundational data for future translational studies.