Mechanisms underlying prion protein toxicity and therapeutic strategies

Abstract

Prion diseases such as Creutzfeldt-Jakob disease (CJD) and Kuru in humans, scrapie in sheep and bovine spongiform encephalopathy (BSE) in cattle are a group of neurodegenerative diseases that invariably lead to death. The current hypothesis states that the cellular prion protein (PrPC) gets converted into a misfolded form PrPSc, characterized by a high β-sheet content (Aguzzi and Calella, 2009). The misfolded PrPSc oligomerizes and grows into fibrils. Broken fibrils can then serve as a seed and lead to further conversion and oligomerization, and therefore be used as a surrogate for infectivity (Knowles et al., 2009). The in vivo conformation of prion fibrils is not defined; hence, developing specific inhibitors remains challenging. Other therapies targeting both prion replication and the intracellular signalling pathways that mediate neurotoxicity have not been successful. Consequently, to date no effective prion therapy exists. Prion disease represents one if not the best-studied protein aggregation disease. An in vitro model for prion-induced pathology has been established in our laboratory. When cerebellar organotypic cultured slices (COCS) are infected with prions, they exhibit all the characteristic features as prion replication, astro- and microgliosis, vacuolation and neurotoxicity (Falsig et al., 2008; Falsig et al., 2012). Luminescent conjugated polythiophenes (LCP) are polymeric fluorescent molecules that preferentially bind to protein aggregates with regular cross-β-sheet structures, including those formed by PrPSc, and can be used to stain many different amyloids in tissues (Klingstedt and Nilsson, 2012). Recently, our laboratory found that treatment of prion-infected brain homogenates and prion-infected COCS with LCPs reduced infectivity. Interestingly, the prionostatic effect seems to rely on hyperstabilization, rather than dissociation, of PrP aggregates (Margalith et al., 2012). More recent findings from our lab have shown that full length, monovalent antibodies or single chain antibodies that target the globular domain (termed globular domain ligands; GDL) of prion protein (PrPC) lead to dramatic neuronal cell loss when applied in cultured organotypic cerebellar slices or stereotactically injected in the cerebellum of mice (Sonati et al., 2013). It was also found that neurotoxicity involves the production of reactive oxygen species and activation of calpains. This thesis focuses on the evaluation of LCPs as a therapy in a mouse model of prion diseases, the comparison of the pathogenetic mechanisms underlying neurotoxicity elicited by GDL or prions and signaling mechanisms involved in prion induced neuronal cell death.