Antibodies against the C-terminus of α-synuclein modulate its fibrillation

•Antibodies targeting the C-terminus of alpha synuclein inhibit its fibrillation.•Antibody-bound oligomers show reduced permeabilization of lipid membranes.•Increased avidity for oligomers and fibrils is ascribed to multiple epitopes.

The 140-residue natively disordered protein α-synuclein (aSN) is a central component in the development of a family of neurodegenerative diseases termed synucleinopathies. This is attributed to its ability to form cytotoxic aggregates such as oligomers and amyloid fibrils. Consequently there have been intense efforts to avoid aggregation or reroute the aggregation pathway using pharmaceutical agents such as small molecules, chaperones and antibodies. aSN's lack of persistent structure in the monomeric state, as well as the multitude of different oligomeric and even different fibrillar states, makes it difficult to raise antibodies that would be efficacious in neutralizing all conformations of aSN. However, the C-terminal 20-40 residues of aSN are a promising epitope for antibody development. It is primarily disordered in both monomeric and aggregated forms, and an anti-C-terminal antibody will therefore be able to bind all forms. Furthermore, it might not interfere with the folding of aSN into membranes, which could be important for its physiological role. Here we report a screen of a series of monoclonal antibodies, which all target the C-terminal of aSN. According to dot blot analyses, different antibodies bound different forms of aSN with different preferences and showed reduced binding to monomeric compared to aggregated (oligomeric and fibrillary) aSN. Consequently they have different effects on aSN's ability to fibrillate and permeabilize membranes. Generally, the antibodies with strongest binding to aggregated aSN in dot blot, also inhibited fibrillation and membrane permeabilization the most, and promoted formation of amorphous aggregates surrounded by small and thin fibers. This suggests that the development of antibodies that targets the C-terminus, exposed in the aggregated forms of aSN, may be beneficial for improved immunotherapy against PD.

Graphical abstractBinding of a bivalent antibody to an alpha-synuclein oligomer leads to a decrease in membrane permeabilization, as measured by a reduction in calcein release. The antibody is a space-filled cartoon model of an IgG1 molecule (PDB codes 1BJ1 and 1IGY) with the heavy chains colored purple and light chains colored red.Download full-size image