Therapeutic mRNA delivery of CRISPR-Cas9 to the trabecular meshwork reverses ocular hypertension in myocilin glaucoma.

Mutations in the myocilin gene (MYOC) are the leading genetic cause of primary open angle glaucoma (POAG), the most common glaucoma type. These mutations trigger a toxic gain-of-function phenotype, causing the misfolded MYOC protein to accumulate in the endoplasmic reticulum (ER) of trabecular meshwork (TM) cells, leading to ER stress, TM cell death, and elevation of intraocular pressure (IOP). Here, we demonstrate that the delivery of Cas9 mRNA via a cationic lipid polymer (lipoplex) targets the TM selectively and edits the MYOC gene, thereby rescuing a mouse model of glaucoma. A single intracameral injection of Cre-mRNA lipoplex resulted in mutant MYOC expression in the TM, leading to glaucoma in a recently developed Cre-inducible mouse model of glaucoma. Lipoplex encapsulating Cas9 and guide RNA targeting MYOC knocked out MYOC, reduced intracellular accumulation of mutant MYOC, and relieved ER stress, thereby rescuing a mouse model of MYOC-associated glaucoma. Our studies further establish the ocular safety and efficacy of non-viral Cas9-sgRNA delivery to the TM, offering a novel, one-time therapeutic strategy for inherited glaucoma caused by MYOC mutations.