Screening of small molecules using the inhibition of oligomer formation in \u03b1-synuclein aggregation as a selection parameter

Roxine Staats,Robert I Horne,Thomas C T Michaels,Christopher M Dobson,Johnny Habchi,Tuomas P J Knowles,Sara Linse,Michele Vendruscolo,Sean Chia,Patrick Flagmeier

doi:10.1038/s42004-020-00412-y

Roxine Staats, Robert I Horne + Show 8 more

Open Access

https://doi.org/10.1038/s42004-020-00412-y

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Abstract

The aggregation of α-synuclein is a central event in Parkinsons’s disease and related synucleinopathies. Since pharmacologically targeting this process, however, has not yet resulted in approved disease-modifying treatments, there is an unmet need of developing novel methods of drug discovery. In this context, the use of chemical kinetics has recently enabled accurate quantifications of the microscopic steps leading to the proliferation of protein misfolded oligomers. As these species are highly neurotoxic, effective therapeutic strategies may be aimed at reducing their numbers. Here, we exploit this quantitative approach to develop a screening strategy that uses the reactive flux toward α-synuclein oligomers as a selection parameter. Using this approach, we evaluate the efficacy of a library of flavone derivatives, identifying apigenin as a compound that simultaneously delays and reduces the formation of α-synuclein oligomers. These results demonstrate a compound selection strategy based on the inhibition of the formation of α-synuclein oligomers, which may be key in identifying small molecules in drug discovery pipelines for diseases associated with α-synuclein aggregation.

Highlights

The aggregation of α-synuclein is a central event in Parkinsons’s disease and related synucleinopathies
In the case of α-synuclein, developing drug discovery strategies based on our understanding of its aggregation process is important given that the specific microscopic steps in the aggregation of this protein are highly sensitive to the cellular environment and solution conditions
We set out to develop a strategy for the identification of small molecules able to interfere with the aggregation process of α-synuclein in a specific manner, whereby the flux towards oligomeric species is reduced

Summary

Introduction

The aggregation of α-synuclein is a central event in Parkinsons’s disease and related synucleinopathies. Since pharmacologically targeting this process, has not yet resulted in approved disease-modifying treatments, there is an unmet need of developing novel methods of drug discovery In this context, the use of chemical kinetics has recently enabled accurate quantifications of the microscopic steps leading to the proliferation of protein misfolded oligomers. Our approach is based on recent advances in the understanding of the conditions that influence α-synuclein aggregation, such as pH and the effects of varying ratios of seed fibrils to free monomers This increased understanding has given rise to a set of different experimental assays through which α-synuclein aggregation may be measured by thioflavin T (ThT) fluorescence, which individually recapitulate fibril elongation and secondary nucleation, whereby new aggregates nucleate on existing fibril surfaces. These experiments occur under quiescent conditions and have been shown to be reproducible under consistent solution conditions[35,36]

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