mTOR Inhibition Drives Mutation-Specific Remodeling of Lysosomal and Autophagic Pathways and GCase Activity in PBMC-Derived Macrophages from Patients with GBA1-Associated Parkinson's Disease.

To date, we and others have demonstrated that GBA1-associated Parkinson's disease (GBA1-PD) exhibits hyperactivation of mTOR and impairment of mTOR-regulated autophagy. Our previous study showed that the degree of autophagy impairment depends on the type of GBA1 mutation in peripheral blood mononuclear cell (PBMC)-derived macrophages. Moreover, the type of GBA1 mutation ("mild"-e.g., p.N370S or "severe"-e.g., p.L444P) correlates with PD severity and may influence therapeutic response. Here, we investigated the dose-dependent effects of GCase inhibition by conduritol β-epoxide (CBE) in SH-SY5Y cells on mTOR signaling, as well as the effects of mTOR inhibition by Torin 1 on mTOR-dependent autophagy-related proteins, lysosomal morphology, and lysosomal hydrolase activities in PBMC-derived macrophages from PD patients carrying GBA1-L444P or GBA1-N370S mutations. CBE induced dose-dependent activation of mTOR signaling in SH-SY5Y, as evidenced by dose-dependent accumulation of p-RPS6 (Ser235/236). mTOR inhibition decreased Beclin-1 protein levels while increasing the LC3B-II/LC3B-I ratio, LC3B-lysosome colocalization, and lysosome number regardless of mutation type in PBMC-derived macrophages. However, Torin1 reduced p62 levels in GBA1-N370S-PD, whereas lysosomal size decreased in GBA1-L444P-PD. Interestingly, Torin 1 increased GCase activity in both patient groups. These findings suggest that mTOR inhibition restores GCase function and autophagy and may represent a potential therapeutic strategy for GBA1-PD.