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Title: Integrated organotypic slice cultures and RT-QuIC (OSCAR) assay: implications for translational discovery in protein misfolding diseases

item KONDRU, NAVEEN - Iowa State University
item MANNE, SIREESHA - Iowa State University
item Greenlee, Justin
item WEST GREENLEE, M - Iowa State University
item ANANTHARAM, VELLAREDDY - Iowa State University
item HALBUR, PATRICK - Iowa State University
item KANTHASAMY, ARTHI - Iowa State University
item KANTHASAMY, ANUMANTHA - Iowa State University

Submitted to: Scientific Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/19/2017
Publication Date: 2/24/2017
Citation: Kondru, N., Manne, S., Greenlee, J., West Greenlee, H., Anantharam, V., Halbur, P., Kanthasamy, A., Kanthasamy, A. 2017. Integrated organotypic slice cultures and RT-QuIC (OSCAR) assay: implications for translational discovery in protein misfolding diseases. Scientific Reports. 7:43155. doi:10.1038/srep43155.

Interpretive Summary: Prion diseases are invariably fatal neurologic diseases for which there is no known prevention or cure. Because of long incubation times and knowledge gaps in how the disease progresses, there is not a well-defined model for testing potential cures or preventative measures. This manuscript describes an ultrasensitive two-step screening model for potential antiprion compounds. In the first step, prion agents are cultured on a brain slice (Organotypic Slice Culture Assay) derived from a prion-susceptible mouse to which various potential treatment compounds can be applied. In the second step, an in vitro prion amplification technique (Real-Time Quaking Induced Conversion or RT-QuIC) is used to test for reductions in prion amplification due to application of the test compound. Collectively, the method is referred to as OSCAR. This paper demonstrates the sensitivity of the OSCAR method and describes the outcome of 3 compounds with known antiprion capabilities. Results suggest that the OSCAR method would be useful in pre-screening potential treatment compounds prior to conducting animal tests. Because mechanisms of neurodegeneration in prion disease are similar to other protein misfolding diseases such as Alzheimer's disease and Parkinson’s disease, use of this model could have a major impact on improving treatments for other neurodegenerative diseases.

Technical Abstract: Protein misfolding is a key pathological event in neurodegenerative diseases like prion diseases, synucleinopathies, and tauopathies that are collectively termed protein misfolding disorders (PMD). Prions are a prototypic model to study protein aggregation biology and therapeutic development. Attempts to develop anti-prion therapeutics have been impeded by the lack of screening models that faithfully replicate prion diseases and the lack of rapid, sensitive biological screening systems. Therefore, a sensitive model encompassing prion replication and neurotoxicity would be indispensable to the pursuit of intervention strategies. We present an ultrasensitive screening system coupled to ex vivo prion organotypic slice culture model to rapidly advance rationale-based high-throughput therapeutic strategies. This hybrid Organotypic Slice Culture Assay coupled with RT-QuIC (OSCAR) permits rapid, sensitive, specific and quantitative detection of prions from an infectious slice culture model, making it amenable to high-throughput. We demonstrate that the anti-prion activity of test compounds can be readily resolved based on the power and kinetics of seeding activity in the OSCAR screening platform and that the prions generated in slice cultures are biologically active. Collectively, our results imply that OSCAR is a robust model of prion diseases that offers a promising platform for understanding prion proteinopathies and advancing anti-prion therapeutics.