Anti-Prion Activity of Poly-L-Amino Acids and Related Peptides

Abstract

Prion diseases or transmissible spongiform encephalopathies (TSE) are lethal neurodegenerative conditions in humans and animals that initiate spontaneously, genetically or by infection. Normal cellular prion protein (PrPC) is conformationally changed into the pathological, disease-linked isoform of prion protein (PrPSc), which is the sole constituent of the infectious agents referred as prion. Previously, some cationic poly-amino acids demonstrated the ability to eliminate PrPSc. In this study, additional cationic, anionic, and uncharged poly-amino acids were examined for anti-prion activity, as well as self-assembled cationic peptide nanostructures (SPNs). Poly-L-arginine (PLR) was investigated to establish the anti-prion activity. PLR decreased the level of PrPSc in cultured cells permanently infected with prions in a concentration- and PLR mass-reliant manner. The PrPSc inhibition by PLR exceeded that of previously identified prion-suppressant poly-L-lysine (PLK). The effective concentration of PLR to inhibit prions was achieved securely underneath the cytotoxic concentrations, and overall cytotoxicity of PLR was alike to that of PLK. PLR did not modify the PrPC level and was not capable to alter the states of preformed recombinant PrP aggregates and PrPSc from prion-infected cells. These results suggest that the action mechanism of PLR is independent of depletion of PrPC or pre-existing PrPSc. The plasminogen–PLR complex established the superior positive surface charge values than the comparable complex with PLK, raising the possibility that PLR interferes with the function of plasminogen, the cofactor for PrPSc production. Furthermore, additional cationic poly-amino acids, poly-L-ornithine (PLO) and poly-L-histadine (PLH) also exhibited strong PrPSc inhibitory effects like PLK and PLR. On the other hand, poly-L-glutamic acid (PLE), an anionic poly-amino acid, and poly-L-threonine (PLT), a polar but electrically uncharged poly-amino acid showed no dramatic changes in the PrPSc level of the prion-infected cells. Cytotoxicity results supported that the concentrations of PLR, PLH, PLO, and PLK, at which PrPSc was entirely eradicated, were not lethal for cells. PrP aggregate formation assay data showed that PLR, PLO, and PLH delayed the generation of PrP aggregates. Next, self-assembled nanostructures based on oligoarginine were designed as an inhibitor of prions, which may lower cytotoxicity while increasing anti-prion activity. By using the copolypeptide block of an oligoarginine and a self-assembling peptide as building blocks, correlation among the nanostructural state, anti-prion activity, and cytotoxicity was investigated. The optimal bioactivity (i.e., the highest anti-prion activity and lowest cytotoxicity) was obtained when the building blocks existed in a unimolecular/unordered state was transformed into high-charge-density vesicles in the low pH endosome/lysosomes during the cell internalization progression. This resulted in the significant enhancement of anti-prion activity. In particular, the in-cell self-assembly concept and cationic polymer and peptides present a practical methodology to developing therapeutics against protein misfolding diseases.