Novel Small Molecules Targeting the Intrinsically Disordered Structural Ensemble of \u03b1-Synuclein Protect Against Diverse \u03b1-Synuclein Mediated Dysfunctions

Gergely Tóth,Phuong Nguyen,John P Anderson,Wherly Hoffman,Shyra J Gardai,Michael F Jobling,Carlos W Bertoncini,Stephen J Ballaron ,Michael P Bova ,Jean‐Christophe Rochet ,Balázs Szőke ,Hans Dieter Junker ,Xiao Hua Chen ,Mariano Tabios ,David A Agard,Daniel Schwizer,John Christodoulou,Nunilo Cremades,Renate Sekul,Eliezer Masliah,Mitali A Tambe,Ted Yednock,Amandine Berthet,Dale Schenk,Jiahui Tao,Inge Ott,Brian Spencer,Thomas Neumann,Wenxian Mao,Lisa Mcconlogue

doi:10.1038/s41598-019-52598-4

Abstract

The over-expression and aggregation of α-synuclein (αSyn) are linked to the onset and pathology of Parkinson’s disease. Native monomeric αSyn exists in an intrinsically disordered ensemble of interconverting conformations, which has made its therapeutic targeting by small molecules highly challenging. Nonetheless, here we successfully target the monomeric structural ensemble of αSyn and thereby identify novel drug-like small molecules that impact multiple pathogenic processes. Using a surface plasmon resonance high-throughput screen, in which monomeric αSyn is incubated with microchips arrayed with tethered compounds, we identified novel αSyn interacting drug-like compounds. Because these small molecules could impact a variety of αSyn forms present in the ensemble, we tested representative hits for impact on multiple αSyn malfunctions in vitro and in cells including aggregation and perturbation of vesicular dynamics. We thereby identified a compound that inhibits αSyn misfolding and is neuroprotective, multiple compounds that restore phagocytosis impaired by αSyn overexpression, and a compound blocking cellular transmission of αSyn. Our studies demonstrate that drug-like small molecules that interact with native αSyn can impact a variety of its pathological processes. Thus, targeting the intrinsically disordered ensemble of αSyn offers a unique approach to the development of small molecule research tools and therapeutics for Parkinson’s disease.

Highlights

The sequential misfolding of α-synuclein into oligomers and fibrils is central to the pathogenesis of Parkinson’s Disease (PD) and related neurodegenerative disorders termed synucleinopathies[1]
Monomeric αSyn was screened against a library of small molecules containing 91,000 lead-like and 23,000 fragment compounds immobilized on microarrays to identify small molecules binding to the protein using surface plasmon resonance (SPR) imaging (HT-CMSPR)[21,22] (Fig. 1a)
To verify the monomeric nature of αSyn in the screen, dynamic light scattering (DLS) analyses were performed on αSyn preparations under screening conditions and in parallel for each library screening experiment. αSyn stock solutions were shown to be free of aggregates with negligible (

Summary

Introduction

The sequential misfolding of α-synuclein (αSyn) into oligomers and fibrils is central to the pathogenesis of Parkinson’s Disease (PD) and related neurodegenerative disorders termed synucleinopathies[1]. ΑSyn can take on more ordered forms upon membrane binding[12,13,14] Owing to this intrinsic dynamic character and the ability to populate other forms upon interaction, IDPs are involved in many key biological processes[15], are vulnerable to aggregation and are susceptible to prion-like amplification of misfolded species and transmission between cells[16]. This propensity towards aggregation and spread is likely to underlie the association that IDPs have with a growing number of misfolding diseases, notably many neurodegenerative disorders[17]. The identification of small molecules reversing diverse malfunctions of αSyn indicates that differing conformations and associated malfunctions of the protein may be targeted by small molecule ligands

Methods

Results

Conclusion