Abstract
Currently, there are no effective treatments for patients with neurodegenerative diseases. Acetylcholinesterase (AChE) inhibitors are the main drugs used in the clinic for the treatment of Alzheimer's disease (AD), but the drugs presently in use have several limitations. There remains an immense challenge regarding the development of effective compounds with lower cytotoxicity and side effects for patients, as well as the need to understand the mechanisms responsible for the development of the disease, since the elucidation of these mechanisms may direct the research to therapeutic targets more feasible and effective.The present proposal is based on results of preliminary studies with new molecules that are inhibitors of cholinesterases, acetyl- (AChE) and butyryl cholinesterases (BuChE), synthesized by Chierrito et al. (Eur. J. Med. Chem., 139: 773-791, 2017) and Andrade et al. (Bioorg. Med. Chem. 2019, 15;27(6):931-943), respectively. The hypothesis of the work is based on the effects of these compounds as neuroprotective agents in cells (neural models) submitted in vitro to neurotoxic stimuli and oxidative stress, leading to an increase in cell survival and recovery. From a mechanistic point of view, it is assumed that several signaling pathways (PI3K/AKT, nAChR-PI3K, oxidative stress and DNA repair) are implicated in the responses to the treatments with the compounds, which will be evaluated by the alterations observed in gene expression profiles.Thus, the general objectives of the project are: 1) To characterize the therapeutic potential of new AChE inhibitory molecules, which are hybrids of donepezil-tacrine (TA8Amino and TAHB3), and inhibitors of BuChE (synthetic compounds C9 and C11) in SH-SY5Y neuronal cell line and in cortical neurons and astrocytes derived from NPCs - Neural Progenitor Cells (derived from patients with sporadic AD and healthy individuals), tested under the neurotoxic stimulus and oxidative stress induced by hydrogen peroxide, ²-amyloid peptide (1-42) and L-Glutamate; 2) Elucidate the molecular mechanisms of neuroprotection in the same cells, using LY294002 (PI3K/AKT inhibitor) and Methyl-licaconitine (nAChRs antagonist), or knockdown strategy, to investigate the pathways or molecular processes involved in the cellular responses to treatments.The ability of the synthetic compounds to confer neuroprotection against the damage caused by genotoxic stress will be assessed in human cells by various methods: evaluation of DNA breaks, micronuclei, cell cycle kinetics, viability and detection of cell death, alterations in mitochondria; detection of total intracellular and mitochondrial ROS, evaluation of gene expression for key genes of the above-mentioned signaling pathways. In addition, experiments will be conducted for neuro-differentiation analysis in SH-SY5Y cells and NPCs, with the aforementioned parameters being evaluated in differentiated cells, as well as the application of chemical inhibition or knockdown strategies. This approach will allow a study of some aspects of the neuroprotection mechanisms conferred by the new synthetic compounds (inhibitors of ChE).The present research project to be conducted in in vitro human cellular models aims to expand the knowledge about the action of novel potential drugs developed for the therapy of patients with AD, as well as to study aspects of the molecular mechanisms of neuroprotection and neurogenesis implicated in the cellular responses displayed by compounds that are inhibitors of AchE and BuChE. (AU)
Scientific publications(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)