In vivo targeting of a variant causing vanishing white matter using CRISPR/Cas9

Vanishing white matter (VWM) is a leukodystrophy caused by recessive variants in subunits of eIF2B. At present, no curative treatment is available and patients often die at young age. Due to its monogenic nature, VWM is a promising candidate for the development of CRISPR/Cas9-mediated gene therapy....

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Bibliographic Details
Main Authors: Anne E.J. Hillen, Martina Hruzova, Tanja Rothgangl, Marjolein Breur, Marianna Bugiani, Marjo S. van der Knaap, Gerald Schwank, Vivi M. Heine
Format: Article
Language:English
Published: Elsevier 2022-06-01
Series:Molecular Therapy: Methods & Clinical Development
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Online Access:http://www.sciencedirect.com/science/article/pii/S2329050122000225
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Summary:Vanishing white matter (VWM) is a leukodystrophy caused by recessive variants in subunits of eIF2B. At present, no curative treatment is available and patients often die at young age. Due to its monogenic nature, VWM is a promising candidate for the development of CRISPR/Cas9-mediated gene therapy. Here we tested a dual-AAV approach in VWM mice encoding CRISPR/Cas9 and a DNA donor template to correct a pathogenic variant in Eif2b5. We performed sequencing analysis to assess gene correction rates and examined effects on the VWM phenotype, including motor behavior. Sequence analysis demonstrated that over 90% of CRISPR/Cas9-induced edits at the targeted locus are insertion or deletion (indel) mutations, rather than precise corrections from the DNA donor template by homology-directed repair. Around half of the CRISPR/Cas9-treated animals died prematurely. VWM mice showed no improvement in motor skills, weight, or neurological scores at 7 months of age, and CRISPR/Cas9-treated controls displayed an induced VWM phenotype. In conclusion, CRISPR/Cas9-induced DNA double-strand breaks (DSBs) at the Eif2b5 locus did not lead to sufficient correction of the VWM variant. Moreover, indel formation in Eif2b5 induced an exacerbated VWM phenotype. Therefore, DSB-independent strategies like base- or prime editing might better suited for VWM correction.
ISSN:2329-0501