Targeting α-synuclein oligomers by protein-fragment complementation for drug discovery in synucleinopathies

Abstract

Objective: Reducing the burden of α-synuclein oligomeric species represents a promising approach for disease-modifying therapies against synucleinopathies such as Parkinson’s disease and dementia with Lewy bodies. However, the lack of efficient drug discovery strategies that specifically target α-synuclein oligomers has been a limitation to drug discovery programs.

Research design and methods: Here we describe an innovative strategy that harnesses the power of bimolecular protein-fragment complementation to monitor synuclein–synuclein interactions. We have developed two robust models to monitor α-synuclein oligomerization by generating novel stable cell lines expressing α-synuclein fusion proteins for either fluorescent or bioluminescent protein-fragment complementation under the tetracycline-controlled transcriptional activation system.

Main outcome measures: A pilot screen was performed resulting in the identification of two potential hits, a p38 MAPK inhibitor and a casein kinase 2 inhibitor, thereby demonstrating the suitability of our protein-fragment complementation assay for the measurement of α-synuclein oligomerization in living cells at high throughput.

Conclusions: The application of the strategy described herein to monitor α-synuclein oligomer formation in living cells with high throughput will facilitate drug discovery efforts for disease-modifying therapies against synucleinopathies and other proteinopathies.

Keywords:

• α-synuclein
• high-throughput screening
• oligomers
• Parkinson’s disease
• protein-fragment complementation
• synucleinopathy

Acknowledgments

The authors thank SW Michnick for the kind gift of leucine zipper venus and luciferase constructs, EL Moussaud-Lamodière, F Fiesel and W Springer, for intellectual contributions and editorial help and L Lewis-Tuffin, M DeJesus-Hernandez, V Rossi, M Van Blitterswijk, TA Evans and A-M Baine for excellent technical assistance.

Declaration of interest

This work was supported by a Mayo Clinic/SBMRI collaborative agreement (SM), a grant from the Florida Translational Research Program (FTRP) and NIH NS063963 (PJM). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

Notes