title: Inhibition of protease-resistant prion protein formation in a transformed deer cell line infected with chronic wasting disease.
abstract: Chronic wasting disease (CWD) is an emerging transmissible spongiform encephalopathy (prion disease) of North American cervids, i.e., mule deer, white-tailed deer, and elk (wapiti). To facilitate in vitro studies of CWD, we have developed a transformed deer cell line that is persistently infected with CWD. Primary cultures derived from uninfected mule deer brain tissue were transformed by transfection with a plasmid containing the simian virus 40 genome. A transformed cell line (MDB) was exposed to microsomes prepared from the brainstem of a CWD-affected mule deer. CWD-associated, protease-resistant prion protein (PrP(CWD)) was used as an indicator of CWD infection. Although no PrP(CWD) was detected in any of these cultures after two passes, dilution cloning of cells yielded one PrP(CWD)-positive clone out of 51. This clone, designated MDB(CWD), has maintained stable PrP(CWD) production through 32 serial passes thus far. A second round of dilution cloning yielded 20 PrP(CWD)-positive subclones out of 30, one of which was designated MDB(CWD2). The MDB(CWD2) cell line was positive for fibronectin and negative for microtubule-associated protein 2 (a neuronal marker) and glial fibrillary acidic protein (an activated astrocyte marker), consistent with derivation from brain fibroblasts (e.g., meningeal fibroblasts). Two inhibitors of rodent scrapie protease-resistant PrP accumulation, pentosan polysulfate and a porphyrin compound, indium (III) meso-tetra(4-sulfonatophenyl)porphine chloride, potently blocked PrP(CWD) accumulation in MDB(CWD) cells. This demonstrates the utility of these cells in a rapid in vitro screening assay for PrP(CWD) inhibitors and suggests that these compounds have potential to be active against CWD in vivo.
authors.affiliation: Laboratory of Persistent Viral Diseases, NIAID, NIH, Rocky Mountain Labs, 903 S. 4th St., Hamilton, MT 59840, USA.
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Raymond
authors.forename: Emily A
authors.initials: EA
authors.lastname: Olsen
authors.forename: Kil Sun
authors.initials: KS
authors.lastname: Lee
authors.forename: Lynne D
authors.initials: LD
authors.lastname: Raymond
authors.forename: P Kruger
authors.initials: PK
authors.lastname: Bryant
authors.forename: Gerald S
authors.initials: GS
authors.lastname: Baron
authors.forename: Winslow S
authors.initials: WS
authors.lastname: Caughey
authors.forename: David A
authors.initials: DA
authors.lastname: Kocisko
authors.forename: Linda E
authors.initials: LE
authors.lastname: McHolland
authors.forename: Cynthia
authors.initials: C
authors.lastname: Favara
authors.forename: Jan P M
authors.initials: JP
authors.lastname: Langeveld
authors.forename: Fred G
authors.initials: FG
authors.lastname: van Zijderveld
authors.forename: Richard T
authors.initials: RT
authors.lastname: Mayer
authors.forename: Michael W
authors.initials: MW
authors.lastname: Miller
authors.forename: Elizabeth S
authors.initials: ES
authors.lastname: Williams
authors.forename: Byron
authors.initials: B
authors.lastname: Caughey
mesh_terms: D000818:Animals; D002461:Cell Line, Transformed / Q000378:metabolism*; D002478:Cells, Cultured; D003670:Deer; D004351:Drug Resistance; D010426:Pentosan Sulfuric Polyester / Q000494:pharmacology*; D010447:Peptide Hydrolases / Q000494:pharmacology*; D011166:Porphyrins / Q000494:pharmacology*; D011328:Prions / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D034081:Wasting Disease, Chronic
chemical_list: D011166:Porphyrins; D011328:Prions; D010426:Pentosan Sulfuric Polyester; D010447:Peptide Hydrolases