In vitro characterization of urea derivatives to inhibit alpha-synuclein early-stage aggregation

Abstract Amyloidosis, a group of diseases caused by the fibrillation of prone-to-aggregate proteins, remains one of the most difficult ailments to treat human medicine. This study focuses on the amyloid protein, alpha-synuclein (α-syn), the major pathogenetic contributor to well-known diseases such as Parkinson's Disease (PD) and multiple system atrophy (MSA). We examine the effectiveness of our novel family of molecules, diaryl derivatives of urea, on the inhibition of early- and late-stage α-syn aggregation. Screening with Thioflavin-T (ThT) fluorescence assay has identified one compound 12 as a promising inhibitor for fibril formation (late-stage aggregation). Utilizing the Photo-induced Cross-linking of Unmodified Proteins (PICUP) assays, compound 12 impeded α-syn oligomerization (early-stage aggregation). In contrast to resveratrol, a polyphenol used as positive control, compound 12 delayed the lag time and did not generate globular structures of 5 nm per transmission electron microscopy (TEM). EC50 of compound 12 is 27.6 ± 1.3 µM. Compound 12 reduced cell inclusions and cytotoxicity in a dose-dependent manner using the inclusion-forming neuroblastoma cell-based assays. This family of compound has the potential to become an invaluable tool to abrogate, at the early-stage, α-syn aggregation.