Anle138b and related compounds are aggregation specific fluorescence markers and reveal high affinity binding to α-synuclein aggregates

Abstract

Background: Special diphenyl-pyrazole compounds and in particular anle138b were found to reduce the progression of prion and Parkinson's disease in animal models. The therapeutic impact of these compounds was attributed to the modulation of α-synuclein and prion-protein aggregation related to these diseases. Methods: Photophysical and photochemical properties of the diphenyl-pyrazole compounds anle138b, anle186b and sery313b and their interaction with monomeric and aggregated α-synuclein were studied by fluorescence techniques. Results: The fluorescence emission of diphenyl-pyrazole is strongly increased upon incubation with α-synuclein fibrils, while no change in fluorescence emission is found when brought in contact with monomeric α-synuclein. This points to a distinct interaction between diphenyl-pyrazole and the fibrillar structure with a high binding affinity (K d = 190 ± 120 nM) for anle138b. Several α-synuclein proteins form a hydrophobic binding pocket for the diphenyl-pyrazole compound. A UV-induced dehalogenation reaction was observed for anle138b which is modulated by the hydrophobic environment of the fibrils. Conclusion: Fluorescence of the investigated diphenyl-pyrazole compounds strongly increases upon binding to fibrillar α-synuclein structures. Binding at high affinity occurs to hydrophobic pockets in the fibrils. General significance: The observed particular fluorescence properties of the diphenyl-pyrazole molecules open new possibilities for the investigation of the mode of action of these compounds in neurodegenerative diseases. The high binding affinity to aggregates and the strong increase in fluorescence upon binding make the compounds promising fluorescence markers for the analysis of aggregation-dependent epitopes.

High-Throughput Screening Methodology to Identify Alpha-Synuclein Aggregation Inhibitors

“…There is strong interest in the discovery of small compounds that can act as chemical chaperones modulating the aggregation of α-syn [15,16,17,18,19,20]. In the absence of a defined 3D-structure to target, screening of large collections of chemically diverse compounds is a useful approach toward the discovery of novel bioactive molecules exhibiting an α-syn anti-aggregational effect.…”

11C Radiolabeling of anle253b: a Putative PET Tracer for Parkinson's Disease That Binds to α‐Synuclein Fibrils in vitro and Crosses the Blood‐Brain Barrier

“…[9] Diphenylpyrazoles such as anle138b (1) were identified from a large-scale screening to inhibit pathological aggregation of prion protein (PrP sc ) and αSyn in vivo [10] and their direct binding to αSyn fibrils was demonstrated in vitro by fluorescence spectroscopy. [11] This led us to the hypothesis that the anle138b family might represent good candidates for the development of PET tracers for αSyn aggregation. As anle253b (2) from the compound list originally reported [12] bears a methyl group suitable for 11 C-methylation (t 1/2 = 20.3 min) we selected this compound as a starting point for tracer development.…”

Progress of Pharmacological Approaches in Parkinson's Disease

“…3‐(1,3‐benzodioxol‐5‐yl)‐5‐(3‐bromophenyl)‐1H‐pyrazole (ANLE138b) has been the first oligomer modulator providing an approach to modify neurodegenerative diseases, such as Prion Disease and PD . ANLE138b reduced the aggregation of aSYN and inhibited disease progression in a PD mouse model, even when treatment had been started after disease onset.…”