Anti-Prion Screening for Acridine, Dextran, and Tannic Acid using Real Time–Quaking Induced Conversion: A Comparison with PrPSc-Infected Cell Screening

Prion propagation is mediated by the structural alteration of normal prion protein (PrP C ) to generate pathogenic prion protein (PrP Sc ). To date, compounds for the inhibition of prion propagation have mainly been screened using PrP Sc -infected cells. Real time–quaking-induced conversion (RT-QuIC) is one alternative screening method. In this study, we assessed the propagation inhibition effects of known anti-prion compounds using RT-QuIC and compared the results with those from a PrP Sc -infected cell assay. Compounds were applied to RT-QuIC reactions at 0 h or 22 h after prion propagation to determine whether they inhibited propagation or reduced amplified aggregates. RT-QuIC reactions in presence of acridine, dextran sulfate sodium (DSS), and tannic acid inhibited seeded aggregation with sporadic Creutzfeldt-Jakob disease at 0 h. After treatment at 22 h, amplified fluorescence was decreased in wells treated with either acridine or tannic acid. Compound activities were verified by western blot of RT-QuIC products and in a dye-independent conversion assay, the Multimer Detection System. Protease K-resistant PrP Sc fragments (PrP res ) were reduced by DSS and tannic acid in the PrP Sc -infected cell assay. Importantly, these inhibitory effects were similar despite different treatment times (0 h versus 3 days). Consequentially, RT-QuIC enabled the more specific classification of compounds according to action (i.e., inhibition of prion propagation versus reduction of amplified aggregates). RT-QuIC addresses the limitations of cell-based screening methods and can be used to further aid our understanding of the mechanisms of action of anti-prion compounds.