Antibodies against Alpha-Synuclein Reduce Oligomerization in Living Cells

Abstract

Recent research implicates soluble aggregated forms of α-synuclein as neurotoxic species with a central role in the pathogenesis of Parkinson's disease and related disorders. The pathway by which α-synuclein aggregates is believed to follow a step-wise pattern, in which dimers and smaller oligomers are initially formed. Here, we used H4 neuroglioma cells expressing α-synuclein fused to hemi:GFP constructs to study the effects of α-synuclein monoclonal antibodies on the early stages of aggregation, as quantified by Bimolecular Fluorescence Complementation assay. Widefield and confocal microscopy revealed that cells treated for 48 h with monoclonal antibodies internalized antibodies to various degrees. C-terminal and oligomer-selective α-synuclein antibodies reduced the extent of α-synuclein dimerization/oligomerization, as indicated by decreased GFP fluorescence signal. Furthermore, ELISA measurements on lysates and conditioned media from antibody treated cells displayed lower α-synuclein levels compared to untreated cells, suggesting increased protein turnover. Taken together, our results propose that extracellular administration of monoclonal antibodies can modify or inhibit early steps in the aggregation process of α-synuclein, thus providing further support for passive immunization against diseases with α-synuclein pathology.

Immunotherapeutic Approaches Targeting Amyloid-β, α-Synuclein, and Tau for the Treatment of Neurodegenerative Disorders

“…Interestingly, and as mentioned before, antibodies against α-syn may not only reduce α-syn levels, but also reduce its oligomerization and fibrillization in living cells, thus reducing the pathology in mouse models of PD [66,124,125]. Furthermore, antibodies may also prevent cell-to-cell propagation of α-syn and facilitate the clearance of extracellular α-syn [84,121,122] (Fig.…”

Identification of Protein Interfaces between α-Synuclein, the Principal Component of Lewy Bodies in Parkinson Disease, and the Molecular Chaperones Human Hsc70 and the Yeast Ssa1p

“…Our findings together with those where antibodies directed against ␣-Syn assembly are used (39,40) lay the foundations for a therapeutic strategy targeting soluble ␣-Syn and inhibiting its assembly into fibrils or modifying the surface properties of fibrils in such a way that they become less toxic and/or intercellular propagation is perturbed. Indeed, our results are invaluable for the design of future therapeutic tools in PD as we describe within a molecular chaperone client binding domain what is necessary and possibly sufficient to bind ␣-Syn.…”

Targeting α-Synuclein as a Parkinson’s Disease Therapeutic

“…In active immunization paradigms, it was noted that many of the high affinity α-synuclein antibodies were directed against the C-terminus, perhaps because this portion of the protein is exposed to the extracellular space upon α-synuclein oligomer binding to and partial penetration of the plasma membrane [299]. Passive immunization with antibody 9E4, directed against amino acids 118-126 of α-synuclein, results in clearance of α-synuclein in the neocortex and hippocampus of PDGFβ-human α-synuclein transgenic mice.…”