NOVEL SMALL-MOLECULE PERK INHIBITOR RESCUES NEURODEGENERATION IN PARKINSON'S DISEASE IN VITRO MODEL
Natalia Siwecka1, Wioletta Rozpędek1, Adrianna Cieślak1, Adam Wawrzynkiewicz1, Dariusz Pytel2, John Alan Diehl2, Ireneusz Majsterek1
1 Department of Clinical Chemistry and Biochemistry, Medical University of Lodz, Poland; 2Department of Biochemistry and Molecular Biology, Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425, USA
Parkinson’s disease (PD), a neurodegenerative disease characterized by selective loss of midbrain dopamine neurons, is caused by accumulation of abnormal aggregates of α-synuclein called Lewy bodies. Mentioned event evokes Endoplasmic Reticulum (ER) stress and triggers activation of the Unfolded Protein Response (UPR) signaling pathway. Among three major branches of the UPR, the protein kinase RNA-like endoplasmic reticulum kinase (PERK)-dependent one plays a key role in neurodegeneration, since it may orchestrate neuronal cell apoptosis under chronic ER stress conditions via induction of pro-apoptotic CCAAT-enhancer-binding protein homologous protein (CHOP). Thus, the aim of the study was to evaluate the effectiveness of the selected PERK inhibitor. Experiments were performed on SH-SY5Y human neuroblastoma cell line, which represents a common cellular model for PD. To assess the effectiveness of the selected compound, cells were incubated with the inhibitor over the concentration range of 3µM-50µM for 1h, and next with an ER stress inducer, thapsigargin (500nM), for 2h. The level of phosphorylation of the main substrate of PERK, eukaryotic initiation factor 2α (eIF2α), was evaluated by Western blotting. Evaluation of the inhibitor cytotoxicity was carried out using the colorimetric XTT assay, over a wide concentration range of 0,75µM-50µM and additionally 0,5mM. The results showed that the tested compound significantly inhibited eIF2α phosphorylation at 25µM concentration (52%). In addition, the PERK inhibitor did not induce a cytotoxic effect at any concentration and incubation time used. Therefore, we can conclude that investigated PERK inhibitor may constitute a promising, innovative therapeutic strategy for PD treatment in the future.