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Series GSE128579 Query DataSets for GSE128579
Status Public on Mar 20, 2019
Title Cas9/sgRNA selective targeting of the P23H Rhodopsin mutant allele for treating retinitis pigmentosa by intravitreal AAV9.PHP.B-based delivery
Organism Mus musculus
Experiment type Other
Summary P23H is the most common mutation in the RHODOPSIN (RHO) gene leading to a dominant form of retinitis pigmentosa (RP), a rod photoreceptor degeneration that invariably causes vision loss. Specific disruption of the disease P23H RHO mutant while preserving the wild-type (WT) functional allele would be an invaluable therapy for this disease. However, various technologies tested in the past failed to achieve effective changes and consequently therapeutic benefits. We validated a CRISPR/Cas9 strategy to specifically inactivate the P23H RHO mutant, while preserving the WT allele in vitro. We, then, translated this approach in vivo by delivering the CRISPR/Cas9 components in murine Rho+/P23H mutant retinae. Targeted retinae presented a high rate of cleavage in the P23H but not WT Rho allele. This gene manipulation was sufficient to slow photoreceptor degeneration and improve retinal functions. To improve the translational potential of our approach, we tested intravitreal delivery of this system by means of adeno-associated viruses (AAVs). To this purpose, the employment of the AAV9-PHP.B resulted the most effective in disrupting the P23H Rho mutant. Finally, this approach was translated successfully in human cells engineered with the homozygous P23H RHO gene mutation. Overall, this is a significant proof-of-concept that gene allele specific targeting by CRISPR/Cas9 technology is specific and efficient and represents an unprecedented tool for treating RP and more broadly dominant genetic human disorders affecting the eye, as well as other tissues.
Overall design 3x AAV-transduced, 2x electroporated, and 1x control retinae
Contributor(s) Broccoli V, Giannelli SG, Massimino L
Citation(s) 29281027
Submission date Mar 20, 2019
Last update date Mar 23, 2019
Contact name Luca Massimino
Phone 3389039500
Organization name Ospedale San Raffaele
Department Gastroenterology
Street address Via Olgettina, 58
City Milano
State/province MI
ZIP/Postal code 20100
Country Italy
Platforms (1)
GPL16417 Illumina MiSeq (Mus musculus)
Samples (6)
GSM3680565 AAV1
GSM3680566 AAV2
GSM3680567 AAV3
BioProject PRJNA528206
SRA SRP188941

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Supplementary file Size Download File type/resource
GSE128579_RAW.tar 20.0 Kb (http)(custom) TAR (of BEDGRAPH)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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