The use of proteomics from in vitro to in vivo
in neurotrophic research
Proteomics is a key technique in drug discovery
The lack of therapeutic options to prevent or treat neurodegenerative diseases is of concern, as the prevalence and disability resulting from these disorders are predicted to increase significantly (Abrahams et al. 2019).Since proteins are responsible for almost all biological processes in an organism, changes in the concentration and/or their structure are likely to reflect the effects of a disease (Davidsson and Sjögren, 2005). Specifically, the presence of protein aggregates in the cells of the central nervous system is a hallmark of neurodegenerative diseases. As a consequence, most drug targets are proteins, enabling proteomics for drug discovery, development and clinical practice (Tyers and Mann, 2003).
Neurotrophins are a family of four similar proteins: NGF, BDNF, NT-3 and NT-4 (Allen et al. 2011); that are important regulators of neuronal survival, development, function, and plasticity (Huang and Reichardt, 2001). Since, in EuroNeurotrophin, we are interested in the study of the therapeutic potential of neurotrophins mimetics, the solution we propose is to create novel small molecules that mimic neurotrophins.
At the Department of Molecular and Cellular Neurobiology, of Vrije Universiteit of Amsterdam, we attempt to use proteomics in order to study the effectiveness of the neurotrophins mimetic molecules that other ESRs are developing in the project. To do it, two different approaches have been established.
The first approach is focused on an in vitro strategy, where primary hippocampal neurons from embryonic wild type mouse have been used in order to create a cell-based model. These cultures have been challenged with APP/Tau/β-Amyloid to obtain an Alzheimer’s disease (AD) model in which we can test the novel molecules that have been created. To analyse the effects of the compounds in our cell model, a high throughput imaging screening is used. This technique gives us information about morphological development, as for example, neurite outgrowth and length; cell proliferation, among many other parameters.
This will help us to understand if there are any differences in the compound treated cells versus the non-treated and provide information concerning if the compound is a good candidate for further studies.
The second approach is an in vivo one which focuses on the examination of the temporal effects of neurotrophin mimetics on the brain tissue and synaptic proteomes in mouse models of neurodegeneration (APPswe/PS1dE9 and 5xFAD transgenic mice for AD; cuprizone mouse model of MS). Large scale proteomics, gives the opportunity to gain deep knowledge with broad information. Data-independent acquisition (DIA) methods such as sequential windowed acquisition of all theoretical fragment ion spectra, SWATH analysis, is ideally suited to high-throughput tissue research.
Specifically, these kind of approaches result in comprehensive peptide data capture. Currently, the chance of analyses by various proteomic software pipelines with high confidence is available with false discovery rates (FDR) usage to control error propagation. The large-scale proteomics data can be hypothesis generating that guide the subsequent functional studies to explain the mechanistic aspects of the disorders and their rescues by the neurotrophin mimetics.
About the Authors
ESR10: Débora Pita
Débora’s research focuses on Cell-Based Models for Neurotrophic Therapeutic Testing. She is hosted at the Vrije Universiteit Amsterdam, Center for Neurogenomics and Cognitive Research.
Evangelia works at the VU University of Amsterdam, Center for Neurogenomics and Cognitive Research in Vrije Universiteit Amsterdam, Netherlands. The main focus of the project is the examination of the temporal effect of the already existed neu-rosteroids and the new neurotrophin mimetics on synapse pro-teome and synapse density in mice models of AD (APPswe/PS1dE9 and 5xFAD transgenic mice).