Potent α-Synuclein Aggregation Inhibitors, Identified by High-Throughput Screening, Mainly Target the Monomeric State

alpha-Synuclein (alpha SN) aggregation is central to the etiology of Parkinson's disease (PD). Large-scale screening of compounds to identify aggregation inhibitors is challenged by stochastic alpha SN aggregation and difficulties in detecting early-stage oligomers (alpha SOs). We developed a high-throughput screening assay combining SDS-stimulated alpha SN aggregation with FRET to reproducibly detect initial stages in alpha SN aggregation. We screened 746,000 compounds, leading to 58 hits that markedly inhibit alpha SN aggregation and reduce alpha SOs' membrane permeabilization activity. The most effective aggregation inhibitors were derivatives of (4-hydroxynaphthalen-1-yl) sulfonamide. They interacted strongly with the N-terminal part of monomeric alpha SN and reduced alpha SO-membrane interactions, possibly by affecting electrostatic interactions. Several compounds reduced alpha SO toxicity toward neuronal cell lines. The inhibitors introduced chemical modifications of alpha SN that were, however, not a prerequisite for inhibitory activity. We also identified several phenyl-benzoxazol compounds that promoted alpha SN aggregation proaggregators). These compounds may be useful tools to modulate alpha SN aggregation in cellula.