Novel Trafficking Assays For Pharmacochaperone Discovery Using Enzyme Fragment Complementation Applied To CFTR-ΔF508
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DiscoveRx® has pioneered a novel enzyme complementation system to monitor cellular events such as protein translocation, protein interactions, and degradation in an HTS friendly, cell-based format. This unique technology provides a basis for the generation of novel cellular assays for intractable targets, and simplify the detection of cellular signaling events using a simple chemiluminescent based protocol. We have applied this technology towards the development of cell-based assays designed to detect pharmacochaperone mediate trafficking of proteins with mutations that otherwise degrade normal trafficking to the cell membrane. Analysis of a mutant form of the ion channel cystic fibrosis transmembrane conductance regulator (CFTR) containing the single amino acid-residue deletion ΔF508 was conducted. The deletion results in a missense mutation that leads to protein misfolding and poor trafficking to the cell membrane. DiscoveRx has developed the PathHunter® CFTR-ΔF508 Pharmacotrafficking assay for discovery of pharmacochaperone compounds that can facilitate proper protein folding and trafficking of the mutant ion channel. The pharmacochaperone activity of two compounds, C4 Corrector and VX-809, is demonstrated. Additional disease relevant trafficking mutants (ion channel and GPCRs) linked to long QT syndrome, retinitis pigmentosa, severe early-onset obesity, basal cell carcinoma and nephrogenic diabetes insipidus will also be highlighted. Overall, these results indicate this novel assay provides a powerful method for screening small molecule libraries to discover pharmacochaperones of disease relevant or orphan GPCRs, ion channels, and transporters.