Inhibition of corneal neovascularization with rAAV-anti-VEGF in mouse corneal injury models

Abstract

Purpose : Corneal neovascularization (CoNV) can lead to visual impairment, affecting over 1.4 million people in the United States. It is caused by a variety of pathologies associated with angiogenic stimuli. Several medical and surgical management regimens are currently available, but they are only partially effective. Conbercept (KH902), an anti-vascular endothelial growth factor (VEGF) drug, can successfully inhibit ocular neovascularization, but it requires repeated dosing. In this study, we investigated the long-acting anti-angiogenesis and safety of rAAV-delivered KH902 in mouse corneal injury models.

Methods : We employed two rAAV serotypes with different transduction efficiencies to deliver KH902 gene via single intrastromal or subconjunctival administration. rAAV-mediated eGFP expression was used as a guide to determine transduction efficiency and cell tropism in the cornea. The levels of KH902 mRNA expression mediated by each rAAV serotype were analyzed by Droplet Digital PCR. The potential toxicity was determined through analyzing the central corneal thickness at various timepoints and immune response at two weeks after high- and low-dose rAAV gene delivery. We tracked and quantified the CoNV progression for 12 weeks post-injection of rAAV-KH902 in CoNV models in vivo. The levels of DLL4/Notch signaling and ERK activation, markers for VEGF stimulation, were tested by Western blot.

Results : We found that the two different serotypes when administered by intrastromal injection conferred KH902 expression in corneal keratocytes with different efficiencies, while rAAV rarely transduced cornea tissue via subconjunctival injection. After reaching peak expression at 1-2weeks, KH902 mRNA expression in the cornea gradually decreased but was still detectable at three months following a single intrastromal injection. High doses of rAAV-KH902 induced a strong corneal inflammatory response; while at low doses, inflammation was at a minimum. Furthermore, rAAV-KH902 reduced DLL4/Notch signaling and ERK activation in alkali burn-induced CoNV mouse model. Our data suggest that rAAV-mediated KH902 gene therapy can dramatically inhibit CoNV for an extended period of time in both alkali burn- and suture-induced CoNV mouse models without adverse events.

Conclusions : Our study demonstrates the potential and relative safety of rAAV-based anti-angiogenesis therapy in the treatment of corneal neovascularization progression.

This is a 2021 ARVO Annual Meeting abstract.