A charged tail on anti-α-Synuclein antibodies does not enhance their affinity to α-Synuclein fibrils

TL;DR

Adding a negatively charged tail to antibodies targeting α-Synuclein fibrils did not improve their binding and may have reduced it due to repulsion and interference.

Contribution

Demonstrates that a charged peptide tail does not enhance antibody binding to α-Synuclein aggregates and identifies potential reasons for the failure.

Findings

The negatively charged peptide tail reduced the binding strength of antibodies to α-Synuclein aggregates.

The tail may interfere with the antibody's binding site, preventing it from attaching to α-Synuclein fibrils.

Improving the peptide tail's design to avoid repulsion and mimic heparan sulphates could enhance future antibody therapies.

Abstract

The aggregation of α-Synuclein (αSyn) is strongly linked to neuronal death in Parkinson’s disease and other synucleinopathies. The spreading of aggregated αSyn between neurons is at least partly dependent on electrostatic interactions between positively charged stretches on αSyn fibrils and the negatively charged heparan sulphate proteoglycans on the cell surface. To date there is still no therapeutic option available that could halt the progression of Parkinson’s disease and one of the major limitations is likely the relatively low proportion of αSyn aggregates accessible to drugs in the extracellular space. Here, we investigated whether a negatively charged peptide tail fused to the αSyn aggregate-specific antibodies SynO2 and 9E4 could enhance the antibodies’ avidity to αSyn aggregates in order to improve their potential therapeutic effect through inhibiting cell-to-cell spreading and enhancing the clearance of extracellular aggregates. We performed ELISAs to test the avidity to αSyn aggregates of both monovalent and bivalent antibody formats with and without the peptide tail. Our results show that the addition of the negatively charged peptide tail decreased the binding strength of both antibodies to αSyn aggregates at physiological salt conditions, which can likely be explained by intermolecular repulsions between the tail and the negatively charged C-terminus of αSyn. Additionally, the tail might interact with the paratopes of the SynO2 antibody abolishing its binding to αSyn aggregates. Conclusively, our peptide tail did not fulfil the required characteristics to improve the antibodies’ binding to αSyn aggregates. Fine-tuning the design of the peptide tail to avoid its interaction with the antibodies’ CDR and to better mimic relevant characteristics of heparan sulphates for αSyn aggregate binding may help overcome the limitations observed in this study.

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