more relevant large-animal TSE disease models, these factors make in vivo testing impractical for high-throughput screening.
A variety of scrapie-and CWD-infected cell-culture models, and some yeast models [14], have been developed and serve as in vitro surrogates for animal studies in screening for anti-prion compounds. These models have the advantage of high throughput and low cost compared to animal testing. Extensive compound screening in cell cultures has identified a number of different classes of anti-prion compounds which have then shown efficacy in animal models. However, as expected with any in vitro drug screen, the predictive accuracy of cell-culture screens for efficacy in vivo is far from 100%. Furthermore, although some compounds appear to have broad-based anti-prion efficacy, striking variation has been observed in drug effectiveness between different prion-infected models and prion strains [15][16][17]. This variation emphasizes the need for testing in experimental models that approximate specific TSE disease targets as closely as possible. Unfortunately, in terms of cell culture models, no cell lines have been stably infected with two of the most important TSE agents, CJD and BSE.
Another advantage of cell cultures is the ability to probe mechanisms of prion inhibition. One can often see that anti-prion compounds that bind to PrP c cause its clustering and internalization. The result is the sequestration of PrP c in a state and/or subcellular location that is incompatible with conversion to PrP res [18][19][20][21].
Noncellular in vitro methods have also been employed to assess a wide range of therapeutic candidates. These assays usually look for competitive binding of PrP c and PrP res or the prevention of PrP amyloid fibril formation. Recently developed techniques include surface plasmon resonance [22], fluorescence correlation spectroscopy [23], semiautomated cell-free conversion [24], and a fluorescence-polarization-based competitive binding assay [25]. Some have even turned to computer ''in silico'' modeling to predict binding partners [26,27].
Ultimately, compounds that look promising in vitro require testing in animals and in humans. Of the many compounds studied in rodent models, only a few have made their way into human trials or case reports. The primary outcome measures of therapy in animal studies include incubation and survival times, PrP res accumulation, histopathological changes, and some take into account behavioral and clinical aspects of the disease. Key factors affecting these outcomes include the dose and route of therapeutic agent, dose and route of disease inoculation, and most important, the time at which treatment is started. Generally, the effectiveness of compounds given at the onset of clinical symptoms, or when there is significant neuropathology, is limited. However, many compounds show some effectiveness in prophylaxis or early treatment, and may have a significant role to play in decontamination of sources of infection, such as blood in the case of human variant CJD. Such drugs need not cross the blood-brain barrier, and might have value in livestock or wildlife disease management. Also, for the many prion diseases that arise following oral INTRODUCTION