Lentiviral mediated delivery of CRISPR/Cas9 reduces intraocular pressure in a mouse model of myocilin glaucoma.

Publisher: Nature Publishing Group Country of Publication: England NLM ID: 101563288 Publication Model: Electronic Cited Medium: Internet ISSN: 2045-2322 (Electronic) Linking ISSN: 20452322 NLM ISO Abbreviation: Sci Rep Subsets: MEDLINE

Original Publication: London : Nature Publishing Group, copyright 2011-

Cytoskeletal Proteins* ; Glaucoma, Open-Angle*/genetics ; Glaucoma, Open-Angle*/therapy ; Glaucoma, Open-Angle*/metabolism ; Glycoproteins*; Animals ; Mice ; CRISPR-Cas Systems ; Disease Models, Animal ; Eye Proteins/genetics ; Eye Proteins/metabolism ; Intraocular Pressure/genetics ; Lentivirus/genetics ; Trabecular Meshwork/metabolism

Mutations in myocilin (MYOC) are the leading known genetic cause of primary open-angle glaucoma, responsible for about 4% of all cases. Mutations in MYOC cause a gain-of-function phenotype in which mutant myocilin accumulates in the endoplasmic reticulum (ER) leading to ER stress and trabecular meshwork (TM) cell death. Therefore, knocking out myocilin at the genome level is an ideal strategy to permanently cure the disease. We have previously utilized CRISPR/Cas9 genome editing successfully to target MYOC using adenovirus 5 (Ad5). However, Ad5 is not a suitable vector for clinical use. Here, we sought to determine the efficacy of adeno-associated viruses (AAVs) and lentiviruses (LVs) to target the TM. First, we examined the TM tropism of single-stranded (ss) and self-complimentary (sc) AAV serotypes as well as LV expressing GFP via intravitreal (IVT) and intracameral (IC) injections. We observed that LV_GFP expression was more specific to the TM injected via the IVT route. IC injections of Trp-mutant scAAV2 showed a prominent expression of GFP in the TM. However, robust GFP expression was also observed in the ciliary body and retina. We next constructed lentiviral particles expressing Cas9 and guide RNA (gRNA) targeting MYOC (crMYOC) and transduction of TM cells stably expressing mutant myocilin with LV_crMYOC significantly reduced myocilin accumulation and its associated chronic ER stress. A single IVT injection of LV_crMYOC in Tg-MYOC ^Y437H mice decreased myocilin accumulation in TM and reduced elevated IOP significantly. Together, our data indicates, LV_crMYOC targets MYOC gene editing in TM and rescues a mouse model of myocilin-associated glaucoma.
(© 2024. The Author(s).)

Update of: Res Sq. 2023 Dec 19:rs.3.rs-3740880. doi: 10.21203/rs.3.rs-3740880/v1. (PMID: 38196579)

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EY034070 United States EY NEI NIH HHS; R01 EY034238 United States EY NEI NIH HHS; R01 EY030366 United States EY NEI NIH HHS; R01 EY026177 United States EY NEI NIH HHS; R01 EY024259 United States EY NEI NIH HHS; EY026177 United States EY NEI NIH HHS; EY030366 United States EY NEI NIH HHS; R01 EY030967 United States EY NEI NIH HHS; EY025580 United States EY NEI NIH HHS

Keywords: ER stress; Gene therapy; Genome editing for glaucoma; Intraocular pressure; Lentiviral particles; Myocilin-associated Glaucoma; Trabecular meshwork; Viral vectors

0 (Cytoskeletal Proteins)
0 (Eye Proteins)
0 (Glycoproteins)
0 (trabecular meshwork-induced glucocorticoid response protein)

Date Created: 20240324 Date Completed: 20240325 Latest Revision: 20240810

20240812

PMC10960846

10.1038/s41598-024-57286-6

38521856