Endolysosomal N-glycan processing is critical to attain the most active form of the enzyme acid alpha-glucosidase.

Acid alpha-glucosidase (GAA) is a lysosomal glycogen-catabolizing enzyme, the deficiency of which leads to Pompe disease. Pompe disease can be treated with systemic recombinant human GAA (rhGAA) enzyme replacement therapy (ERT), but the current standard of care exhibits poor uptake in skeletal muscles, limiting its clinical efficacy. Furthermore, it is unclear how the specific cellular processing steps of GAA after delivery to lysosomes impact its efficacy. GAA undergoes both proteolytic cleavage and glycan trimming within the endolysosomal pathway, yielding an enzyme that is more efficient in hydrolyzing its natural substrate, glycogen. Here, we developed a tool kit of modified rhGAAs that allowed us to dissect the individual contributions of glycan trimming and proteolysis on maturation-associated increases in glycogen hydrolysis using in vitro and in cellulo enzyme processing, glycopeptide analysis by MS, and high-pH anion-exchange chromatography with pulsed amperometric detection for enzyme kinetics. Chemical modifications of terminal sialic acids on N-glycans blocked sialidase activity in vitro and in cellulo, thereby preventing downstream glycan trimming without affecting proteolysis. This sialidase-resistant rhGAA displayed only partial activation after endolysosomal processing, as evidenced by reduced catalytic efficiency. We also generated enzymatically deglycosylated rhGAA that was shown to be partially activated despite not undergoing proteolytic processing. Taken together, these data suggest that an optimal rhGAA ERT would require both N-glycan and proteolytic processing to attain the most efficient enzyme for glycogen hydrolysis and treatment of Pompe disease. Future studies should examine the amenability of next-generation ERTs to both types of cellular processing.

Domestic harm and neglect among lesbian, gay, and bisexual older adults.

This study examined harm, hurt, and neglect by caregivers as well as self-neglect and physical and mental health status among 113 lesbian, gay, and bisexual (LGB) older adults aged 60-88 years, who attended community-based social and recreation programs or groups in the United States. Reporting on their experiences with caregivers, 22.1% of the participants experienced at least one type of harm, including physical, emotional, verbal, sexual, financial, and neglectful; additionally, 25.7% of the participants reported they knew LGB older adults who experienced at least one type of harm from his or her caregiver. With regard to self-neglect, 62.8% reported experiencing it; those indicating positive psychological health reported fewer experiences with self-neglect.