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Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia
Choroideremia (CHM) is an X-linked chorioretinal dystrophy leading to progressive retinal degeneration that results in blindness by late adulthood. It is caused by mutations in the CHM gene encoding the Rab Escort Protein 1 (REP1), which plays a crucial role in the prenylation of Rab proteins ensuri...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10607001/ https://www.ncbi.nlm.nih.gov/pubmed/37894906 http://dx.doi.org/10.3390/ijms242015225 |
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author | Toualbi, Lyes Toms, Maria Almeida, Patrick Vingadas Harbottle, Richard Moosajee, Mariya |
author_facet | Toualbi, Lyes Toms, Maria Almeida, Patrick Vingadas Harbottle, Richard Moosajee, Mariya |
author_sort | Toualbi, Lyes |
collection | PubMed |
description | Choroideremia (CHM) is an X-linked chorioretinal dystrophy leading to progressive retinal degeneration that results in blindness by late adulthood. It is caused by mutations in the CHM gene encoding the Rab Escort Protein 1 (REP1), which plays a crucial role in the prenylation of Rab proteins ensuring correct intracellular trafficking. Gene augmentation is a promising therapeutic strategy, and there are several completed and ongoing clinical trials for treating CHM using adeno-associated virus (AAV) vectors. However, late-phase trials have failed to show significant functional improvements and have raised safety concerns about inflammatory events potentially caused by the use of viruses. Therefore, alternative non-viral therapies are desirable. Episomal scaffold/matrix attachment region (S/MAR)-based plasmid vectors were generated containing the human CHM coding sequence, a GFP reporter gene, and ubiquitous promoters (pS/MAR-CHM). The vectors were assessed in two choroideremia disease model systems: (1) CHM patient-derived fibroblasts and (2) chm(ru848) zebrafish, using Western blotting to detect REP1 protein expression and in vitro prenylation assays to assess the rescue of prenylation function. Retinal immunohistochemistry was used to investigate vector expression and photoreceptor morphology in injected zebrafish retinas. The pS/MAR-CHM vectors generated persistent REP1 expression in CHM patient fibroblasts and showed a significant rescue of prenylation function by 75%, indicating correction of the underlying biochemical defect associated with CHM. In addition, GFP and human REP1 expression were detected in zebrafish microinjected with the pS/MAR-CHM at the one-cell stage. Injected chm(ru848) zebrafish showed increased survival, prenylation function, and improved retinal photoreceptor morphology. Non-viral S/MAR vectors show promise as a potential gene-augmentation strategy without the use of immunogenic viral components, which could be applicable to many inherited retinal disease genes. |
format | Online Article Text |
id | pubmed-10607001 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-106070012023-10-28 Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia Toualbi, Lyes Toms, Maria Almeida, Patrick Vingadas Harbottle, Richard Moosajee, Mariya Int J Mol Sci Article Choroideremia (CHM) is an X-linked chorioretinal dystrophy leading to progressive retinal degeneration that results in blindness by late adulthood. It is caused by mutations in the CHM gene encoding the Rab Escort Protein 1 (REP1), which plays a crucial role in the prenylation of Rab proteins ensuring correct intracellular trafficking. Gene augmentation is a promising therapeutic strategy, and there are several completed and ongoing clinical trials for treating CHM using adeno-associated virus (AAV) vectors. However, late-phase trials have failed to show significant functional improvements and have raised safety concerns about inflammatory events potentially caused by the use of viruses. Therefore, alternative non-viral therapies are desirable. Episomal scaffold/matrix attachment region (S/MAR)-based plasmid vectors were generated containing the human CHM coding sequence, a GFP reporter gene, and ubiquitous promoters (pS/MAR-CHM). The vectors were assessed in two choroideremia disease model systems: (1) CHM patient-derived fibroblasts and (2) chm(ru848) zebrafish, using Western blotting to detect REP1 protein expression and in vitro prenylation assays to assess the rescue of prenylation function. Retinal immunohistochemistry was used to investigate vector expression and photoreceptor morphology in injected zebrafish retinas. The pS/MAR-CHM vectors generated persistent REP1 expression in CHM patient fibroblasts and showed a significant rescue of prenylation function by 75%, indicating correction of the underlying biochemical defect associated with CHM. In addition, GFP and human REP1 expression were detected in zebrafish microinjected with the pS/MAR-CHM at the one-cell stage. Injected chm(ru848) zebrafish showed increased survival, prenylation function, and improved retinal photoreceptor morphology. Non-viral S/MAR vectors show promise as a potential gene-augmentation strategy without the use of immunogenic viral components, which could be applicable to many inherited retinal disease genes. MDPI 2023-10-16 /pmc/articles/PMC10607001/ /pubmed/37894906 http://dx.doi.org/10.3390/ijms242015225 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Toualbi, Lyes Toms, Maria Almeida, Patrick Vingadas Harbottle, Richard Moosajee, Mariya Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia |
title | Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia |
title_full | Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia |
title_fullStr | Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia |
title_full_unstemmed | Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia |
title_short | Gene Augmentation of CHM Using Non-Viral Episomal Vectors in Models of Choroideremia |
title_sort | gene augmentation of chm using non-viral episomal vectors in models of choroideremia |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10607001/ https://www.ncbi.nlm.nih.gov/pubmed/37894906 http://dx.doi.org/10.3390/ijms242015225 |
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