Pompe disease (PD) is a rare genetic disorder stemming from mutations inactivating the gene encoding acid maltase (GAA), the enzyme responsible for glycogen breakdown within lysosomes. GAA deficiency leads to pathological glycogen accumulation in several tissues; in the infantile form, this triggers a severe cardiomyopathy that can be successfully reverted by enzyme replacing therapy. Moreover, both infantile and late onset (LOPD) forms exhibit a progressive and ERT resistant skeletal muscle wasting that forces patients to mechanical ventilation and leads to death.
Studies clarifying the pathogenic cascade guided the development and optimization of next-generation ERT; however, no systematic assessment of lysosomal signaling abnormalities during PD progression has been made due to major technical challenges in the isolation and characterization of functional organelles. Moreover, if and how glycogen accumulation can be considered the unique priming event remains unclear.
For this reason, we optimized a robust protocol for rapid lysosomes immunopurification (LysoIP) starting from skeletal muscles, diaphragm and heart. Through LysoIP proteomics, we captured the enrichment of all ten glycolytic enzymes in muscular lysosomal fraction of healthy mice, leading us to hypothesize the existance of an intimate relationship between GAA function, lysosomal glycogen and glycolytic enzyme enrichment. To link this phenomenon to PD pathogenesis and rule out if lysosomal failure may be not driven by glycogen accumulation alone but could be a consequence of aberrant GAA signaling, we generated HEK GAA KO cells and identified Cathepsin B activity failure as a primary hallmark of GAA deficiency independently from glycogen accumulation. Moreover, this CTSB failure was also reported in GAA deficient skeletal muscles, while heart and liver are unaffected. We confirmed this observation also in Pompe patient derived myoblasts, suggesting that the lack of lysosomal glucosidase activity could impair fundamental cellular processes, such as lysosomal CSTB activity and lysosomal degradative capacity, through a defect in glycolytic enzymes signaling. Thus, despite further investigations are needed to mechanistically clarify this phenomenon, our findings identified a glycogen accumulation independent hallmark of lysosomal failure that could link a signaling phenomenon originating on lysosomal surface to the comprehension on novel events driving GAA deficiency pathogenesis.