Prion diseases are fatal, untreatable neurodegenerative diseases caused by the accumulation of the misfolded, infectious isoform of the prion protein (PrP), termed PrP(Sc). In an effort to identify novel inhibitors of prion formation, we utilized a high-throughput enzyme-linked immunosorbent assay (ELISA) to evaluate PrP(Sc) reduction in prion-infected neuroblastoma cell lines (ScN2a). We screened a library of approximately 10,000 diverse small molecules in 96-well format and identified 121 compounds that reduced PrP(Sc) levels at a concentration of 5 microM. Four chemical scaffolds were identified as potential candidates for chemical optimization based on the presence of preliminary structure-activity relationships (SAR) derived from the primary screening data. A follow-up analysis of a group of commercially available 2-aminothiazoles showed this class as generally active in ScN2a cells. Our results establish 2-aminothiazoles as promising candidates for efficacy studies of animals and validate our drug discovery platform as a viable strategy for the identification of novel lead compounds with antiprion properties.