TARGETING Α-SYNUCLEIN AGGREGATION AND INFLAMMATION IN PARKINSON¿S DISEASE: THE ROLE OF POLY-Γ-GLUTAMIC ACID

Parkinson’s disease (PD) is a complex multifactorial neurodegenerative disorder affecting more than 10 million people worldwide. As a synucleinopathy, PD is characterized by the pathological accumulation of misfolded and aggregated α-synuclein in both the central and peripheral nervous system. In this context, many efforts have been made to counteract α-synuclein aggregation, propagation, and associated inflammatory responses, which are key features of the disease. However, despite the tremendous effort and research investment, no disease-modifying therapies are currently available, and symptomatic approaches remain the standard cure for the treatment of PD. In this work we aimed to evaluate the therapeutic potential of commercially available poly-γ-glutamic acid (γ-PGA) polymers, a natural, non-toxic and non-immunogenic biopolymer, in different in vitro models of PD, focusing on its properties as anti-inflammatory agent and modulator of aggregation. Indeed γ-PGA has gained significant interest in biomedicine due to its antioxidant, anti-inflammatory, and neuroprotective effects, providing evidence for its beneficial value in neurodegenerative disorders. To investigate these biological properties, we selected complementary in vitro models that recapitulate different aspects of PD pathogenesis. Primary murine astrocytes were employed for their central role in α-synuclein clearance and neuroinflammation, given their involvement in the interplay between protein aggregation and immune responses in the central nervous system. Macrophages were selected to assess peripheral immune cell activation, which is strongly implicated in PD pathogenesis. Finally, neuronal in vitro model and cell free system assay were employed to assess the specific effect of γ-PGA in interfering with α-synuclein aggregation. Using primary murine astrocytes exposed to α-synuclein preformed fibrils (PFFs) we demonstrated that γ-PGA restored cell viability, reduced astrocyte inflammation, and decreased α-synuclein pathology burden. These effects were associated with the reduced internalization of aggregates and the direct interference with α-synuclein aggregation, as confirmed by cell-free system assays. In addition, we examined the impact of α-synuclein aggregates on macrophage immune response, focusing on their polarization. Using an in vitro model of macrophages, we observed that PFFs induced a mixed M1/M2 phenotype without affecting cytotoxicity and ROS production, and that γ-PGA administration attenuated this polarization, suggesting a modulatory role in α-synuclein-induced immune response. Finally, in a neuronal in vitro model, γ-PGA polymers significantly reduced the burden of early-stage α-synuclein aggregates supporting its direct anti-aggregation capacity. 4 Overall, this work provides in vitro proof-of-concept evidence that γ-PGA can modulate α-synuclein aggregation and aggregate-induced inflammatory response, highlighting its potential as a safe and effective therapeutic strategy for PD. Interestingly, given its additional role as a prebiotic able to modulate the gut microbiome, and the emerging evidence that dysbiosis is linked to PD, future research that explores γ-PGA effects on the gut-brain axis and focuses on inflammation and microbiota modulation in the gut, as well as α-synuclein propagation and neuronal function in PD pathogenesis, holds promise.