Searching for anti-prion compounds: cell-based high-throughput in vitro assays and animal testing strategies.
The transmissible spongiform encephalopathies (TSEs) or prion diseases are infectious neurodegenerative diseases of mammals. Protease-resistant prion protein (PrP-res) is only associated with TSEs and thus has been a target for therapeutic intervention. The most effective compounds known against scrapie in vivo are inhibitors of PrP-res in infected cells. Mouse neuroblastoma (N2a) cells have been chronically infected with several strains of mouse scrapie including RML and 22L. Also, rabbit epithelial cells that produce sheep prion protein in the presence of doxycycline (Rov9) have been infected with sheep scrapie. Here a high-throughput 96-well plate PrP-res inhibition assay is described for each of these scrapie-infected cell lines. With this dot-blot assay, thousands of compounds can easily be screened for inhibition of PrP-res formation. This assay is designed to find new PrP-res inhibitors, which may make good candidates for in vivo anti-scrapie testing. However, an in vitro assay can only suggest that a given compound might have in vivo anti-scrapie activity, which is typically measured as increased survival times. Methods for in vivo testing of compounds for anti-scrapie activity in transgenic mice, a much more lengthy and expensive process, are also discussed.
Kocisko
David A
DA
Laboratory of Persisten Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana 59840, USA.
Caughey
Byron
B
D000818:Animals; D001681:Biological Assay; D002460:Cell Line; D045744:Cell Line, Tumor; D015195:Drug Design; D004847:Epithelial Cells / Q000166:cytology; D066298:In Vitro Techniques; D051379:Mice; D008822:Mice, Transgenic; D009447:Neuroblastoma; D018620:PrPSc Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011817:Rabbits; D012608:Scrapie / Q000188:drug therapy* / Q000378:metabolism*; D012756:Sheep; D013045:Species Specificity
D018620:PrPSc Proteins