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Jeffrey O'Callaghan, Darragh Crosbie, Paul Cassidy, Marian M Humphries, Anna-Sophia Kiang, Joseph M. Sherwood, Darryl R. Overby, W Daniel Stamer, Lawrence Tam, Peter Humphries; AAV-mediated MMP3 expression from corneal endothelium as a targeted gene therapy approach for glaucoma. Invest. Ophthalmol. Vis. Sci. 2016;57(12):793.
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© ARVO (1962-2015); The Authors (2016-present)
Matrix metalloproteinases (MMP) play an important role in remodeling of the extracellular matrix (ECM) within the drainage channels of the eye to control outflow resistance and thus IOP. This study assesses the therapeutic potential of increasing outflow facility via inoculation of an AAV virus expressing MMP3 in the corneal endothelium.
Transendothelial electrical resistance (TEER) and FITC-dextran assays were carried out to evaluate the effect of recombinant MMP-3 on the paracellular permeability of Schlemm’s canal endothelial (SCE) monolayers. Wildtype murine eyes (C57/BL6J) were intracamerally injected with an AAV2/9 vector (1x1011 AAV particles per eye) expressing a murine MMP3 cDNA driven by a CMV promoter, with contralateral AAV-2/9 expressing the CMV promoter with no transgene as controls. After 4 weeks, mouse eyes were enucleated and outflow facility was measured using the i-Perfusion system.
Treatment of SCE cells with recombinant MMP-3 significantly increased paracellular permeability to 70kDa dextran 114-fold (p=0.0005) and reduced TEER values by 23% (p=0.015). Histological analysis of anterior segment flat mounts of C57/BL6J mice showed that intracameral inoculation of AAV2/9 specifically transduced the corneal endothelium, indicating the use of this serotype as a means to deliver MMP-3 to the conventional outflow pathway by following the natural flow of aqueous humour in the anterior chamber. We observed that intracameral delivery of an AAV2/9 vector expressing MMP3 from the corneal endothelium significantly increased outflow facility, on average, by 39% (p=0.008, N=7), as compared to contralateral controls.
The use of AAV vectors in directing expression of potentially therapeutic genes to the anterior chamber of the eye has not been explored for gene therapy as extensively as other ocular regions such as the retina. The secretory nature of MMPs renders the corneal endothelium a favourable site for protein expression, since the flow of aqueous humor will direct the secreted protein into the trabecular meshwork. In the current study, outflow facility was significantly increased, suggesting a realistic rationale for gene-based therapy.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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