Increasing the Endoplasmic Reticulum Pool of the F508del Allele of the Cystic Fibrosis Transmembrane Conductance Regulator Leads to Greater Folding Correction by Small Molecule Therapeutics
| العنوان: | Increasing the Endoplasmic Reticulum Pool of the F508del Allele of the Cystic Fibrosis Transmembrane Conductance Regulator Leads to Greater Folding Correction by Small Molecule Therapeutics |
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| المؤلفون: | W. Joon Chung, Adam B. Keeton, Allan M. Weissman, Zackery E. Plyler, E. Lucile White, Jeffrey L. Brodsky, Marina Mazur, Viktória Havasi, Eric J. Sorscher, Lynn Rasmussen, Gary A. Piazza, Jennifer L. Goeckeler-Fried, Jeong S. Hong, Annette N. Chiang, R. Aldrin Denny, Steven M. Rowe |
| المصدر: | PLoS ONE, Vol 11, Iss 10, p e0163615 (2016) PLoS ONE |
| بيانات النشر: | Public Library of Science (PLoS), 2016. |
| سنة النشر: | 2016 |
| مصطلحات موضوعية: | 0301 basic medicine, Protein Folding, Cystic Fibrosis, Pulmonology, Cell Membranes, Cystic Fibrosis Transmembrane Conductance Regulator, lcsh:Medicine, Bioinformatics, Endoplasmic Reticulum, Hydroxamic Acids, Cystic fibrosis, Benzoates, Biochemistry, Epithelium, Mutant protein, Animal Cells, Medicine and Health Sciences, lcsh:Science, Cells, Cultured, Vorinostat, Multidisciplinary, biology, Organic Compounds, Messenger RNA, Small molecule, Cystic fibrosis transmembrane conductance regulator, 3. Good health, Cell biology, Nucleic acids, Chemistry, Genetic Diseases, High-content screening, Physical Sciences, Cell lines, Cellular Types, Anatomy, Cellular Structures and Organelles, Biological cultures, Research Article, Biotechnology, Library Screening, Research and Analysis Methods, Small Molecule Libraries, 03 medical and health sciences, Autosomal Recessive Diseases, medicine, Humans, RNA, Messenger, Furans, Molecular Biology Techniques, Molecular Biology, Alleles, Clinical Genetics, Molecular Biology Assays and Analysis Techniques, Endoplasmic reticulum, Organic Chemistry, HEK 293 cells, lcsh:R, Wild type, Ubiquitination, Chemical Compounds, Biology and Life Sciences, Epithelial Cells, Cell Biology, Cell Cultures, medicine.disease, Fibrosis, High-Throughput Screening Assays, Protein Structure, Tertiary, 030104 developmental biology, HEK293 Cells, Biological Tissue, Microscopy, Fluorescence, Small Molecules, biology.protein, Pyrazoles, RNA, lcsh:Q, Gene Deletion, HeLa Cells, Developmental Biology |
| الوصف: | Small molecules that correct the folding defects and enhance surface localization of the F508del mutation in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) comprise an important therapeutic strategy for cystic fibrosis lung disease. However, compounds that rescue the F508del mutant protein to wild type (WT) levels have not been identified. In this report, we consider obstacles to obtaining robust and therapeutically relevant levels of F508del CFTR. For example, markedly diminished steady state amounts of F508del CFTR compared to WT CFTR are present in recombinant bronchial epithelial cell lines, even when much higher levels of mutant transcript are present. In human primary airway cells, the paucity of Band B F508del is even more pronounced, although F508del and WT mRNA concentrations are comparable. Therefore, to augment levels of "repairable" F508del CFTR and identify small molecules that then correct this pool, we developed compound library screening protocols based on automated protein detection. First, cell-based imaging measurements were used to semi-quantitatively estimate distribution of F508del CFTR by high content analysis of two-dimensional images. We evaluated ~2,000 known bioactive compounds from the NIH Roadmap Molecular Libraries Small Molecule Repository in a pilot screen and identified agents that increase the F508del protein pool. Second, we analyzed ~10,000 compounds representing diverse chemical scaffolds for effects on total CFTR expression using a multi-plate fluorescence protocol and describe compounds that promote F508del maturation. Together, our findings demonstrate proof of principle that agents identified in this fashion can augment the level of endoplasmic reticulum (ER) resident "Band B" F508del CFTR suitable for pharmacologic correction. As further evidence in support of this strategy, PYR-41-a compound that inhibits the E1 ubiquitin activating enzyme-was shown to synergistically enhance F508del rescue by C18, a small molecule corrector. Our combined results indicate that increasing the levels of ER-localized CFTR available for repair provides a novel route to correct F508del CFTR. |
| اللغة: | English |
| تدمد: | 1932-6203 |
| الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::548cb6f2b3cb88d56f75f466b19a5a57Test http://europepmc.org/articles/PMC5061379?pdf=renderTest |
| حقوق: | OPEN |
| رقم الانضمام: | edsair.doi.dedup.....548cb6f2b3cb88d56f75f466b19a5a57 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 19326203 |
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