A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex
In the skin fragility disorder epidermolysis bullosa simplex (EBS), mutations in keratin 14 (K14, also known as KRT14) or keratin 5 (K5, also known as KRT5) lead to keratinocyte rupture and skin blistering. Severe forms of EBS are associated with cytoplasmic protein aggregates, with elevated kinase...
| Main Authors: | , , , , , , |
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| Format: | Journal Article |
| Language: | English |
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2021
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| Online Access: | https://hdl.handle.net/10356/153607 |
| _version_ | 1826123296806010880 |
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| author | Tan, Tong San Common, John E. A. Lim, John S. Y. Badowski, Cedric Muhammad Jasrie Firdaus Leonardi, Steven S. Lane, E. Birgitte |
| author2 | Lee Kong Chian School of Medicine (LKCMedicine) |
| author_facet | Lee Kong Chian School of Medicine (LKCMedicine) Tan, Tong San Common, John E. A. Lim, John S. Y. Badowski, Cedric Muhammad Jasrie Firdaus Leonardi, Steven S. Lane, E. Birgitte |
| author_sort | Tan, Tong San |
| collection | NTU |
| description | In the skin fragility disorder epidermolysis bullosa simplex (EBS), mutations in keratin 14 (K14, also known as KRT14) or keratin 5 (K5, also known as KRT5) lead to keratinocyte rupture and skin blistering. Severe forms of EBS are associated with cytoplasmic protein aggregates, with elevated kinase activation of ERK1 and ERK2 (ERK1/2; also known as MAPK3 and MAPK1, respectively), suggesting intrinsic stress caused by misfolded keratin protein. Human keratinocyte EBS reporter cells stably expressing GFP-tagged EBS-mimetic mutant K14 were used to optimize a semi-automated system to quantify the effects of test compounds on keratin aggregates. Screening of a protein kinase inhibitor library identified several candidates that reduced aggregates and impacted on epidermal growth factor receptor (EGFR) signalling. EGF ligand exposure induced keratin aggregates in EBS reporter keratinocytes, which was reversible by EGFR inhibition. EBS keratinocytes treated with a known EGFR inhibitor, afatinib, were driven out of activation and towards quiescence with minimal cell death. Aggregate reduction was accompanied by denser keratin filament networks with enhanced intercellular cohesion and resilience, which when extrapolated to a whole tissue context would predict reduced epidermal fragility in EBS patients. This assay system provides a powerful tool for discovery and development of new pathway intervention therapeutic avenues for EBS. |
| first_indexed | 2024-10-01T06:02:21Z |
| format | Journal Article |
| id | ntu-10356/153607 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T06:02:21Z |
| publishDate | 2021 |
| record_format | dspace |
| spelling | ntu-10356/1536072023-03-05T16:46:06Z A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex Tan, Tong San Common, John E. A. Lim, John S. Y. Badowski, Cedric Muhammad Jasrie Firdaus Leonardi, Steven S. Lane, E. Birgitte Lee Kong Chian School of Medicine (LKCMedicine) Skin Research Institute of Singapore, A*STAR Institute of Medical Biology, A*STAR Science::Medicine Epidermolysis Bullosa Simplex Epidermolysis Bullosa Simplex In the skin fragility disorder epidermolysis bullosa simplex (EBS), mutations in keratin 14 (K14, also known as KRT14) or keratin 5 (K5, also known as KRT5) lead to keratinocyte rupture and skin blistering. Severe forms of EBS are associated with cytoplasmic protein aggregates, with elevated kinase activation of ERK1 and ERK2 (ERK1/2; also known as MAPK3 and MAPK1, respectively), suggesting intrinsic stress caused by misfolded keratin protein. Human keratinocyte EBS reporter cells stably expressing GFP-tagged EBS-mimetic mutant K14 were used to optimize a semi-automated system to quantify the effects of test compounds on keratin aggregates. Screening of a protein kinase inhibitor library identified several candidates that reduced aggregates and impacted on epidermal growth factor receptor (EGFR) signalling. EGF ligand exposure induced keratin aggregates in EBS reporter keratinocytes, which was reversible by EGFR inhibition. EBS keratinocytes treated with a known EGFR inhibitor, afatinib, were driven out of activation and towards quiescence with minimal cell death. Aggregate reduction was accompanied by denser keratin filament networks with enhanced intercellular cohesion and resilience, which when extrapolated to a whole tissue context would predict reduced epidermal fragility in EBS patients. This assay system provides a powerful tool for discovery and development of new pathway intervention therapeutic avenues for EBS. Published version This work was supported by DEBRA International grants LANE2/LANE3 to E.B.L., and by grants IAF311011 and SPF2013/004 to E.B.L. and J.E.A.C. from the Biomedical Research Council of Singapore. The funding sources were not involved in the conduct of the research, or writing of the manuscript. No payment was received from any pharmaceutical company or other for-profit agency to write the manuscript. Deposited in PMC for immediate release. 2021-12-10T04:56:56Z 2021-12-10T04:56:56Z 2021 Journal Article Tan, T. S., Common, J. E. A., Lim, J. S. Y., Badowski, C., Muhammad Jasrie Firdaus, Leonardi, S. S. & Lane, E. B. (2021). A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex. Journal of Cell Science, 134(19), jcs258409-. https://dx.doi.org/10.1242/jcs.258409 0021-9533 https://hdl.handle.net/10356/153607 10.1242/jcs.258409 34643242 19 134 jcs258409 en IAF311011 SPF2013/004 Journal of Cell Science © 2021. Published by The Company of Biologists Ltd | Journal of Cell Science (2021). This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed application/pdf |
| spellingShingle | Science::Medicine Epidermolysis Bullosa Simplex Epidermolysis Bullosa Simplex Tan, Tong San Common, John E. A. Lim, John S. Y. Badowski, Cedric Muhammad Jasrie Firdaus Leonardi, Steven S. Lane, E. Birgitte A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| title | A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| title_full | A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| title_fullStr | A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| title_full_unstemmed | A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| title_short | A cell-based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| title_sort | cell based drug discovery assay identifies inhibition of cell stress responses as a new approach to treatment of epidermolysis bullosa simplex |
| topic | Science::Medicine Epidermolysis Bullosa Simplex Epidermolysis Bullosa Simplex |
| url | https://hdl.handle.net/10356/153607 |
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