Design and Development of Potential α-Synuclein PET Tracers: Synthesis and Evaluation of Diarylpyrazole- and 2-Styrylbenzothiazole-based Libraries

Abstract

Parkinson’s disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB) are collectively identified as synucleinopathies, a group of neurodegenerative diseases which are defined by the accumulation of α-synuclein (αSYN) fibrils as the characteristic hallmark of pathogenesis. [1] A technique to detect and quantify these aggregates in vivo would be critical to the advancement of the field, drastically transforming the understanding, diagnosis and treatment of synucleinopathies. Multiple studies across the recent literature have highlighted some promising αSYN ligands, [2] however no candidate has yet displayed a suitable binding profile to overcome all challenges set by this target and to successfully reach clinical application. Overall, this study aimed at enhancing the affinity and selectivity of attractive scaffolds from the literature, with the goal of ultimately establishing an αSYN PET tracer. The rational drug design technique of molecular hybridization was applied to the SIL and MODAG scaffolds to develop a library of diarylpyrazole- (DAP) based hybrid compounds in the pursue of an enhancement of their binding properties. The in vitro assessment of DAPs via [3H]SIL26 and [3H]MODAG-001 competition binding assays disproved the original hypothesis for this attachment strategy and demonstrated a shift of selectivity towards amyloid β (Aβ) instead, indicating a theranostic potential for Alzheimer’s disease (AD). [3] The flexibility increase induced by the ring-opening modification on the phenothiazine moiety of DAPs did not result in an enhanced αSYN affinity but instead a complete loss of competition, along with a considerable decrease in Aβ binding. [3] Simultaneously, this study investigated a 2-styrylbenzothiazole-based library, originating from the αSYN selective fluorescent probe RB1. The lead compound PFSB and its less lipophilic derivative MFSB were selected from a set of 42 novel compounds for their favorable binding affinity. Both [3H]PiB competition assays and in vitro autoradiography on human brain slices demonstrated a remarkable αSYN/Aβ selectivity. Together with the display of blood-brain barrier penetration and brain-uptake in the in vivo PET evaluation of [18F]MFSB, these findings highlight 2-styrylbenzothiazoles as a potential critical step forward in the development of αSYN radioligands, although requiring further structural optimization to improve their pharmacokinetics as well as the evaluation of off-target binding in direct assays. [4] The research presented in this thesis overall provides additional insights on the structural features influencing selectivity among neurodegeneration-associated misfolded proteins and describes a novel candidate for the in vivo detection of αSYN.