Successes and challenges in phenotype-based lead discovery for prion diseases

Abstract

Creutzfeldt-Jakob disease (CJD) is a rare but invariably fatal neurodegenerative disease caused by misfolding of an endogenous protein into an alternative pathogenic conformation. The details of protein misfolding and aggregation are not well understood nor are the mechanism(s) by which the aggregated protein confers cellular toxicity. While there is as yet no clear consensus about how best to intervene therapeutically in CJD, prion infections can be propagated in cell culture and in experimental animals, affording both in vitro and in vivo models of disease. Here we review recent lead discovery efforts for CJD, with a focus on our own efforts to optimize 2-aminothiazole analogues for anti-prion potency in cells and for brain exposure in mice. The compounds that emerged from this effort were found to be efficacious in multiple animal models of prion disease even as they revealed new challenges for the field, including the emergence of resistant prion strains.

Figure 1

Mechanism of prion propagation and potential points of therapeutic intervention. (A) The NMR structure of the folded domain of human PrP^C is shown. (B) PrP^Sc forms fibrillar aggregates as shown in the electron micrograph (bar indicates 1000 Å). The β solenoid structure of the fungal Het-S prion provides one possible model of the pathogenic fold. (C) A simplified model for the propagation of PrP^Sc suggests several potential sites for therapeutic intervention.

Figure 2

Approved drugs studied in human clinical trials for CJD.

Figure 3

Cell-based assay for identification of anti-prion compounds. (A) Neuroblastoma cell lines are exposed to PrP^Sc to initiate the infection process. Once infected, cells continuously transmit PrP^Sc to daughter cells upon passage. (B) Infected cells are exposed to test compounds at various concentrations and time durations. The effect of a compound on PrP^Sc levels is quantified by treating cell lysates with proteinase K and quantifying the level of remaining PrP^Sc by ELISA.

Chart 1. Selected Small Molecules with Anti-Prion Activitya

Chart 2. Pharmacologic Agents Bearing the 2-Aminothiazole Ring System

Figure 4

Summary of anti-prion SAR for aminothiazole analogues.

Chart 3. Select Early AMT Analogues Evaluated for Brain Exposure in Micea

Chart 4. Anti-Prion Activities (ScN2a-cl3 Assay) for Structural Analogues of 7a

Figure 5

Kaplan–Meier survival curves for mice infected with ME7 (top) and RML prions (bottom) . The vehicle-treated cohort is shown as a solid black line. Treated cohorts received compound 7 from day 1 postinoculation (solid gray line) or from day 60 postinoculation (dashed gray line).