A Genome Engineered Human Embryonic Stem Cell line to Investigate Pompe Disease

Background Pompe disease is an autosomal recessive lysosomal storage disorder caused by mutations in the GAA gene, leading to acid alpha-glucosidase deficiency and pathological glycogen accumulation, primarily in cardiac and skeletal muscle. While enzyme replacement therapy (ERT) has improved clinical outcomes, its limited efficacy especially in skeletal muscle underscores the need for improved disease models and novel therapeutic strategies. Induced pluripotent stem cells (iPSCs) from Pompe patients have facilitated mechanistic studies; however, their utility is restricted by limited patient sample availability. Methods To address this limitation, we employed CRISPR-Cas9 genome editing to disrupt GAA in a well-characterized human embryonic stem cell (hESC) line, BJNhem20, thereby generating a Pompe disease model independent of patient material. Results The edited hESC line exhibited markedly reduced GAA enzymatic activity while maintaining pluripotency and trilineage differentiation potential. Upon directed differentiation, cardiomyocytes displayed pronounced lysosomal accumulation and increased glycogen storage, whereas skeletal myotubes exhibited elevated cell death and a marginal increase in glycogen content. Conclusions These findings demonstrate that genome-edited hESC for Pompe disease can recapitulate key pathological features, providing a robust and scalable platform for disease modelling and therapeutic screening. This approach offers a valuable alternative to patient-derived iPSCs for studying rare genetic disorders and for the development of targeted interventions.