Library-Derived Peptide Aggregation Modulators of Parkinson?s Disease Early-Onset ?-Synuclein Variants

Parkinson's Disease (PD) is characterized by the accumulation of Lewy bodies in dopaminergic neurons. The main protein component of Lewy bodies, alpha-synuclein (alpha S), is also firmly linked to PD through the identification of a number of single point mutations that are autosomal dominant for early-onset disease. Consequently, the misfolding and subsequent aggregation of alpha S is thought to be a key stage in the development and progression of PD. Therefore, modulating the aggregation pathway of alpha S is an attractive therapeutic target. Owing to the fact that all but one of the familial mutations is located in the preNAC 45-54 region of alpha S, we previously designed a semi-rational library using this sequence as a design scaffold. The 45-54 peptide library was screened using a protein-fragment complementation assay approach, leading to the identification of the 4554W peptide. The peptide was subsequently found to be effective in inhibiting primary nucleation of alpha S, the earliest stage of the aggregation pathway. Here, we build upon this previous work by screening the same 45-54 library against five of the known alpha S single-point mutants that are associated with early-onset PD (A30P, E46K, H50Q, G51D, and A53T). These point mutations lead to a rapid acceleration of PD pathology by altering either the rate or type of aggregates formed. All ultimately lead to earlier disease onset and were therefore used to enforce increased assay stringency during the library screening process. The ultimate aim was to identify a peptide that is effective against not only the familial alpha S variant from which it has been selected but that is also effective against WT alpha S. Screening resulted in five peptides that shared common residues at some positions, while deviating at others. All reduced aggregation of the respective target, with several also identified to be effective at reducing aggregation when incubated with other variants. In addition, our results demonstrate that a previously optimized peptide, 4554W(N6A), is highly effective against not only WT alpha S but also several of the single-point mutant forms and hence is a suitable baseline for further work toward a PD therapeutic.