Cell-Based Quantification of Chronic Wasting Disease Prions

Bian, Jifeng; Napier, Dana; Khaychuck, Vadim; Angers, Rachel; Graham, Catherine; Telling, Glenn C.

doi:10.1128/jvi.00633-10

Cited by 72 publications

(109 citation statements)

References 23 publications

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“…We previously developed a cell-based assay for quantifying CWD prions with sensitivity comparable with the CWD bioassay in susceptible Tg mice (16). We used this cervid prioncell assay (CPCA) to compare levels of CWD prions in Elk21 + with quinacrine-treated cells.…”

Section: Significancementioning

confidence: 99%

“…Our first approach was to assess the efficacy of compounds with known effects on rodent prions. Consistent with its effect on mouse prion infectivity, sustained treatment of Elk21 + cells with dextran sulfate 500 (DS-500) resulted in PrP Sc clearance, which did not reemerge in the resulting Elk21 − cells after more than 40 passages, and inoculation of susceptible transgenic (Tg) mice showed that Elk21 − cells were cured of CWD prion infection (16). The present study was initially designed to assess the effects of quinacrine on CWD.…”

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confidence: 99%

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Quinacrine promotes replication and conformational mutation of chronic wasting disease prions

Bian

Kang

Telling

2014

Proc. Natl. Acad. Sci. U.S.A.

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Quinacrine's ability to reduce levels of pathogenic prion protein (PrP Sc ) in mouse cells infected with experimentally adapted prions led to several unsuccessful clinical studies in patients with prion diseases, a 10-y investment to understand its mechanism of action, and the production of related compounds with expectations of greater efficacy. We show here, in stark contrast to this reported inhibitory effect, that quinacrine enhances deer and elk PrP Sc accumulation and promotes propagation of prions causing chronic wasting disease (CWD), a fatal, transmissible, neurodegenerative disorder of cervids of uncertain zoonotic potential. Surprisingly, despite increased prion titers in quinacrine-treated cells, transmission of the resulting prions produced prolonged incubation times and altered PrP Sc deposition patterns in the brains of diseased transgenic mice. This unexpected outcome is consistent with quinacrine affecting the intrinsic properties of the CWD prion. Accordingly, quinacrine-treated CWD prions were comprised of an altered PrP Sc conformation. Our findings provide convincing evidence for drug-induced conformational mutation of prions without the prerequisite of generating drug-resistant variants of the original strain. More specifically, they show that a drug capable of restraining prions in one species/strain setting, and consequently used to treat human prion diseases, improves replicative ability in another and therefore force reconsideration of current strategies to screen antiprion compounds.prion therapeutics | prion enhancing drugs P rions are protein-based infectious agents that cause an increasing variety of fatal, infectious neurodegenerative disorders, frequently as epidemics. Variant Creutzfeldt-Jakob disease (CJD) resulting from bovine spongiform encephalopathy exemplifies prion zoonosis, and the continued, unpredictable emergence of additional epidemics in other species, particularly chronic wasting disease (CWD) in deer, elk, and moose, raises additional concerns. Its unprecedented contagious spread, widening host range, and conflicting evidence surrounding its zoonotic potential place CWD at the forefront of public health concerns. Although the notion, that prion replication occurs by corruption of host-encoded cellular prion protein (PrP C ) by abnormally conformed PrP Sc , is now widely accepted, the details of this mechanism remain enigmatic.Except under certain experimental conditions, treatments capable of arresting or of effectively modifying the course of disease do not exist for prion disorders. Compounds targeting prevention of PrP misfolding have been aggressively pursued as therapeutics. Cells chronically infected with rodent prions are used as a means either to screen compounds inhibiting PrP Sc accumulation (1-5) or to confirm the proposed antiprion effects of compounds discovered by other means (6, 7). Such strategies rest on the assumption that compounds with efficacy against experimentally adapted rodent prions will be effective against naturally occurring prions, in ...

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Section: Significancementioning

confidence: 99%

mentioning

confidence: 99%

Quinacrine promotes replication and conformational mutation of chronic wasting disease prions

Bian

Kang

Telling

2014

Proc. Natl. Acad. Sci. U.S.A.

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“…12 Several other groups also reported the permissiveness of RK13-cells expressing mouse, bank vole, deer and elk PrP C to propagate prions from the corresponding species. [13][14][15] Scrapie prion propagation was not observed in various established cell lines (non-transfectants) that naturally express endogenous PrP C , including the 2 cell lines derived from fetal tissues of goats. 20 Therefore, we decided to use the RK13 cell line to express caprine PrP C and then assess its ability to propagate brain-derived classical caprine scrapie prion isolates.…”

Section: Cprk13 Cells Support Propagation Of Classical Caprine Scrapimentioning

confidence: 99%

“…The susceptibility of RK13-based transfectants to corresponding rodent-and cervid-derived prion propagation has been reported. [13][14][15] In this study, we demonstrate for the first time that RK13 cells stably expressing caprine PrP C (cpRK13) was permissive to certain classical caprine scrapie prion isolates prepared from the brain tissues of scrapie-infected goats and ovinized transgenic mice.…”

Section: Introductionmentioning

confidence: 99%

A transfectant RK13 cell line permissive to classical caprine scrapie prion propagation

Dassanayake

Zhuang

Truscott

et al. 2016

Prion

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ABSTRACT. To assess scrapie infectivity associated with caprine-origin tissues, bioassay can be performed using kids, lambs or transgenic mice expressing caprine or ovine prion (PRNP) alleles, but the incubation periods are fairly long. Although several classical ovine scrapie prion permissive cell lines with the ability to detect brain-derived scrapie prion have been available, no classical caprine scrapie permissive cell line is currently available. Therefore, the aims of this study were to generate a rabbit kidney epithelial cell line (RK13) stably expressing caprine wild-type PRNP (cpRK13) and then to assess permissiveness of cpRK13 cells to classical caprine scrapie prion propagation. The cpRK13 and plasmid control RK13 (pcRK13) cells were incubated with brain-derived classical caprine scrapie inocula prepared from goats or ovinized transgenic mice (Tg338, express ovine VRQ allele) infected with caprine scrapie. Significant PrP Sc accumulation, which is indicative of scrapie prion propagation, was detected by TSE ELISA and immunohistochemistry in cpRK13 cells inoculated with classical caprine scrapie inocula. Western blot analysis revealed the typical proteinase K-resistant 3 PrP res isoforms in the caprine scrapie prion inoculated cpRK13 cell lysate. Importantly, PrP Sc accumulation was not detected in similarly inoculated pcRK13 cells, whether by TSE ELISA, immunohistochemistry, or western blot. These findings suggest that caprine scrapie prions can be propagated in cpRK13 cells, thus this cell line may be a useful tool for the assessment of classical caprine prions in the brain tissues of goats.

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“…13 -15 Cell-based assays for the titration of prion infectivity Due to substantial methodological advancements it has become possible to titrate the infectivity of certain murine scrapie prions (22L, RML) in quantitative cell-based assays using subcloned neuroblastoma (N2a) cells. 10 -12 Furthermore, RK13 cells transgenically expressing PrP from mouse, sheep or cervids were shown to allow the titration of RML prions, 16 a natural sheep scrapie isolate (PG127), 16 and chronic wasting disease agent, 17 respectively.…”

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confidence: 99%

Towards further reduction and replacement of animal bioassays in prion research by cell and protein misfolding cyclic amplification assays

et al. 2013

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Laboratory animals have long since been used extensively in bioassays for prions in order to quantify, usually in terms of median infective doses [ID 50 ], how infectious these pathogens are in vivo. The identification of aberrant prion protein as the main component and self-replicating principle of prions has given rise to alternative approaches for prion titration. Such approaches often use protein misfolding cyclic amplification (PMCA) for the cell-free biochemical measurement of prionassociated seeding activity, or cell assays for the titration of in vitro infectivity. However, median seeding and cell culture infective doses (SD 50 and CCID 50 , respectively) of prions are neither formally congruent nor definitely representative for ID 50 titres in animals and can be therefore only tentatively translated into the latter. This may potentially impede the acceptance and use of alternative methods to animal bioassays in prion research. Thus, we suggest performing PMCA and cell assays jointly, and to check whether these profoundly different test principles deliver consistent results in order to strengthen the reliability and credibility of prion ID 50 assessments by in vitro methods. With regard to this rationale, we describe three pairs of PMCA and glial cell assays for different hamster-adapted prion agents (the frequently used 263K scrapie strain, and 22A-H scrapie and BSE-H). In addition, we report on the adaptation of quantitative PMCA to human variant Creutzfeldt -Jakob disease (vCJD) prions on steel wires for prion disinfection studies. Our rationale and methodology can be systematically extended to other types of prions and used to further reduce or replace prion bioassays in rodents.

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Cell-Based Quantification of Chronic Wasting Disease Prions

Cited by 72 publications

References 23 publications

Quinacrine promotes replication and conformational mutation of chronic wasting disease prions

Quinacrine promotes replication and conformational mutation of chronic wasting disease prions

A transfectant RK13 cell line permissive to classical caprine scrapie prion propagation

Towards further reduction and replacement of animal bioassays in prion research by cell and protein misfolding cyclic amplification assays

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