title: Utility of RNAi-mediated prnp gene silencing in neuroblastoma cells permanently infected by prions: potentials and limitations.
abstract: Prion diseases are incurable, transmissible neurodegenerative disorders in humans and animals. Because the disease-associated isoform of prion protein, PrP(Sc), is conformationally converted from cellular prion protein, PrP(C), knockdown of PrP(C) expression by RNA interference (RNAi) implicates therapy for prion diseases. In this study, introduction of small interfering (si) and small hairpin (sh) RNAs targeting the prion protein gene (prnp) transcripts triggered specific gene silencing and reduced the PrP(C) level in both prion-free and -infected neuroblastoma cell lines. Furthermore, this approach suppressed PrP(Sc) formation and ultimately eliminated PrP(Sc) from prion-infected cell lines. However, prolonged culture of cured cells resulted in reappearance of PrP(Sc) in the cell population, presumably by de novo PrP(Sc) formation from residual PrP(C) uncontrolled by RNAi and PrP(Sc) remained under the detection limit. Protein misfolding cyclic amplification assays further confirmed that lysate of cured cells was sufficient to support PrP(Sc) propagation. Our data not only suggest a potential treatment option but also implicate a caveat for using an RNAi approach for prion diseases. These findings provide critical information required to advance RNAi-based prevention and therapy for prion diseases of humans and animals.
authors.affiliation: Sanders-Brown Center on Aging, Department of Microbiology, Immunology and Molecular Genetics, College of Medicine, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Younghwan
authors.initials: Y
authors.lastname: Kim
authors.forename: Boram
authors.initials: B
authors.lastname: Han
authors.forename: William
authors.initials: W
authors.lastname: Titlow
authors.forename: Charles E
authors.initials: CE
authors.lastname: Mays
authors.forename: Moosik
authors.initials: M
authors.lastname: Kwon
authors.forename: Chongsuk
authors.initials: C
authors.lastname: Ryou
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D015536:Down-Regulation; D005786:Gene Expression Regulation; D020868:Gene Silencing; D006801:Humans; D009447:Neuroblastoma / Q000378:metabolism*; D018621:PrPC Proteins / Q000235:genetics / Q000378:metabolism*; D018620:PrPSc Proteins / Q000235:genetics / Q000378:metabolism*; D017096:Prion Diseases / Q000628:therapy; D011328:Prions / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D017510:Protein Folding; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D018621:PrPC Proteins; D018620:PrPSc Proteins; D011328:Prions; D034741:RNA, Small Interfering