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

Sam Yacoub; Balasankara Reddy Kaipa; Linya Li; Sarahi Rios; Ramesh Kasetti; Prabhavathi Maddineni; Abbot F Clark; Gulab S Zode

doi:10.1016/j.omtm.2025.101614

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

Mol Ther Methods Clin Dev. 2025 Oct 11;33(4):101614. doi: 10.1016/j.omtm.2025.101614. eCollection 2025 Dec 11.

Authors

Sam Yacoub¹, Balasankara Reddy Kaipa², Linya Li², Sarahi Rios², Ramesh Kasetti¹, Prabhavathi Maddineni³, Abbot F Clark¹, Gulab S Zode²

Affiliations

¹ Department of Pharmacology and Neuroscience and North Texas Eye Research Institute, University of North Texas Health Science Center, Fort Worth, TX 76107, USA.
² Gavin Herbert Eye Institute-Center for Translational Vision Research, Department of Ophthalmology, Department of Physiology and Biophysics, University of California, Irvine, Irvine School of Medicine, Irvine, CA 92697, USA.
³ Department of Ophthalmology, School of Medicine, University of Missouri, Columbia, MO 65212, USA.

Abstract

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.

Keywords: Cas9; gene editing; glaucoma; juvenile-onset glaucoma; liposomes; mRNA delivery; mouse model of glaucoma; myocilin; ocular hypertension; protein misfolding; trabecular meshwork.