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Assay for testing inhibitors of α-synuclein amyloid fibril formation in cells
Galkin, Maksym ; Yushchenko, Dmytro (advisor) ; Hudeček, Jiří (referee) ; Schneider, Bohdan (referee)
Parkinson's disease (PD) is the second most common neurodegenerative disease worldwide and has no cure yet. During PD progressive neuronal loss is accompanied by the formation of Lewy Bodies - abnormal intracellular aggregates, which are mostly formed by fibrils of neuronal protein α-synuclein (αSyn). Aggregation of αSyn was shown to correlate with the pathology progression during PD. Therefore, inhibition of αSyn aggregation and subsequent pathology spreading is a promising approach to the disease treatment. In previous years, a high variety of such inhibitors were developed including small molecules, antibodies, and fibril end-targeting proteins. While these inhibitors were profoundly tested in solutions, their performance in cell- based assays remained unclear. The main reason for this was the absence of a reliable high-throughput cell-based assay for αSyn aggregation evaluation. The application of such an assay is particularly important in the case of molecules that target amyloid fibrils because they can lack specificity and interact with other β-sheet rich proteins present in cells. Moreover, the application of the inhibitors can be severely limited due to poor membrane penetration, intracellular degradation or off-target interactions. The main goal of my work is the development of an assay...
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Development of inhibitors of amyloid fibril formation
Priss, Anastasiia ; Yushchenko, Dmytro (advisor) ; Schneider, Bohdan (referee) ; Hudeček, Jiří (referee)
α-Synuclein (α-Syn) is a small neuronal protein that is present in synapses, and, presumably, regulates the vesicle-mediated neurotransmitter release. Misfolding of α-Syn into amyloid fibrils is linked to the progression of synucleinopathies, including Parkinson's disease, the second most common neurodegenerative disorder. To date, no Parkinson's disease treatment that stops or retards neurodegeneration was established. Several direct and indirect approaches of influence on the α-Syn aggregation are extensively developed in search of potential drug candidates. Recently proposed inhibition of the α-Syn fibrils growth by blocking their ends is one of the most promising known strategies, as it opens an opportunity to design fibrillization inhibitors that act at very low concentrations. However, rational design of inhibitors of this type was not performed yet. The ultimate goal of this work was the development of highly efficient and specific inhibitors of α-Syn fibrillization. Therefore, careful structure-activity relationship analysis of the fibril end blocking inhibitors was performed and the affinity to fibril end was defined as the crucial factor for their inhibition activity. Using the recent data on the fibril structure, we designed inhibitors that are able to bind to the fibril ends with...
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