Exploring non-coding variants and evaluation of antisense oligonucleotides for splicing redirection in Usher syndrome.
Por:
García-Bohórquez B, Barberán-Martínez P, Aller E, Jaijo T, Mínguez P, Rodilla C, Fernández-Caballero L, Blanco-Kelly F, Ayuso C, Sanchis-Juan A, Broekman S, de Vrieze E, van Wijk E, García-García G and Millán JM
Publicada:
10 dic 2024
Resumen:
Exploring non-coding regions is increasingly gaining importance in the diagnosis of inherited retinal dystrophies. Deep-intronic variants causing aberrant splicing have been identified, prompting the development of antisense oligonucleotides (ASOs) to modulate splicing. We performed a screening of five previously described USH2A deep-intronic variants among USH2A monoallelic patients with Usher syndrome (USH) or isolated retinitis pigmentosa. Sequencing of entire USH2A or USH genes was then conducted in unresolved or newly monoallelic cases. The splicing impact of identified variants was assessed using minigene assays, and ASOs were designed to correct splicing. The screening allowed to diagnose 30.95% of the studied patients. The sequencing of USH genes revealed 16 new variants predicted to affect splicing, with four confirmed to affect splicing through minigene assays. Two of them were unreported deep-intronic variants and predicted to include a pseudoexon in the pre-mRNA, and the other two could alter a regulatory cis -element. ASOs designed for three USH2A deep-intronic variants successfully redirected splicing in vitro . Our study demonstrates the improvement in genetic characterization of IRDs when analyzing non-coding regions, highlighting that deep-intronic variants significantly contribute to USH2A pathogenicity. Furthermore, successful splicing modulation through ASOs highlights their therapeutic potential for patients carrying deep-intronic variants.
Filiaciones:
:
Molecular, Cellular and Genomics Biomedicine, Health Research Institute La Fe, 46026 Valencia, Spain
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Joint Unit CIPF-IIS La Fe Molecular, Cellular and Genomic Biomedicine, 46026 Valencia, Spain
Barberán-Martínez P:
Molecular, Cellular and Genomics Biomedicine, Health Research Institute La Fe, 46026 Valencia, Spain
Joint Unit CIPF-IIS La Fe Molecular, Cellular and Genomic Biomedicine, 46026 Valencia, Spain
:
Molecular, Cellular and Genomics Biomedicine, Health Research Institute La Fe, 46026 Valencia, Spain
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Joint Unit CIPF-IIS La Fe Molecular, Cellular and Genomic Biomedicine, 46026 Valencia, Spain
University and Polytechnic La Fe Hospital of Valencia, 46026 Valencia, Spain
:
Molecular, Cellular and Genomics Biomedicine, Health Research Institute La Fe, 46026 Valencia, Spain
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Joint Unit CIPF-IIS La Fe Molecular, Cellular and Genomic Biomedicine, 46026 Valencia, Spain
University and Polytechnic La Fe Hospital of Valencia, 46026 Valencia, Spain
Mínguez P:
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain
Bioinformatics Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain
Rodilla C:
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain
Fernández-Caballero L:
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain
Blanco-Kelly F:
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain
Ayuso C:
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain
Sanchis-Juan A:
Center for Genomic Medicine, Massachusetts General Hospital, Boston, MA, USA
Program in Medical and Population Genetics and Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA, USA
Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA
Broekman S:
Department of Otorhinolaryngology, Radboud University Medical Center, 6525 GA Nijmegen, the Netherlands
de Vrieze E:
Department of Otorhinolaryngology, Radboud University Medical Center, 6525 GA Nijmegen, the Netherlands
van Wijk E:
Department of Otorhinolaryngology, Radboud University Medical Center, 6525 GA Nijmegen, the Netherlands
:
Molecular, Cellular and Genomics Biomedicine, Health Research Institute La Fe, 46026 Valencia, Spain
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Joint Unit CIPF-IIS La Fe Molecular, Cellular and Genomic Biomedicine, 46026 Valencia, Spain
:
Molecular, Cellular and Genomics Biomedicine, Health Research Institute La Fe, 46026 Valencia, Spain
Center for Biomedical Network Research on Rare Diseases (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain
Joint Unit CIPF-IIS La Fe Molecular, Cellular and Genomic Biomedicine, 46026 Valencia, Spain
University and Polytechnic La Fe Hospital of Valencia, 46026 Valencia, Spain
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