Zebrafish as a Model to Evaluate a CRISPR/Cas9-Based Exon Excision Approach as a Future Treatment Option for EYS-Associated Retinitis Pigmentosa

Schellens, R., de Vrieze, E., Graave, P., Broekman, S., Nagel-Wolfrum, K., Peters, T., Kremer, H., Collin, R.W.J., van Wijk, E.
International Journal of Molecular Sciences   22(17): (Journal)
Registered Authors
Collin, Rob, de Vrieze, Erik, Kremer, Hannie, van Wijk, Erwin
CRISPR/Cas9, EYS, antisense oligonucleotides, exon skipping therapy, photoreceptors, retinitis pigmentosa, zebrafish
MeSH Terms
  • Animals
  • CRISPR-Cas Systems
  • Disease Models, Animal*
  • Exons
  • Eye Proteins/chemistry
  • Eye Proteins/genetics*
  • Eye Proteins/metabolism
  • Genetic Therapy/methods
  • Phenotype
  • Protein Domains
  • Retinitis Pigmentosa/genetics*
  • Retinitis Pigmentosa/pathology
  • Retinitis Pigmentosa/therapy
  • Zebrafish
  • Zebrafish Proteins/chemistry
  • Zebrafish Proteins/genetics*
  • Zebrafish Proteins/metabolism
34502064 Full text @ Int. J. Mol. Sci.
Retinitis pigmentosa (RP) is an inherited retinal disease (IRD) with an overall prevalence of 1 in 4000 individuals. Mutations in EYS (Eyes shut homolog) are among the most frequent causes of non-syndromic autosomal recessively inherited RP and act via a loss-of-function mechanism. In light of the recent successes for other IRDs, we investigated the therapeutic potential of exon skipping for EYS-associated RP. CRISPR/Cas9 was employed to generate zebrafish from which the region encompassing the orthologous exons 37-41 of human EYS (eys exons 40-44) was excised from the genome. The excision of these exons was predicted to maintain the open reading frame and to result in the removal of exactly one Laminin G and two EGF domains. Although the eysΔexon40-44 transcript was found at levels comparable to wild-type eys, and no unwanted off-target modifications were identified within the eys coding sequence after single-molecule sequencing, EysΔexon40-44 protein expression could not be detected. Visual motor response experiments revealed that eysΔexon40-44 larvae were visually impaired and histological analysis revealed a progressive degeneration of the retinal outer nuclear layer in these zebrafish. Altogether, the data obtained in our zebrafish model currently provide no indications for the skipping of EYS exons 37-41 as an effective future treatment strategy for EYS-associated RP.
Genes / Markers
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Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Engineered Foreign Genes