AAV2 Delivery of Flt23k Intraceptors Inhibits Murine Choroidal Neovascularization

Abstract

Long-term inhibition of extracellular vascular endothelial growth factor (VEGF) in the treatment of age-related macular degeneration (AMD) may induce retinal neuronal toxicity and risk other side effects. We developed a novel strategy which inhibits retinal pigment epithelium (RPE)-derived VEGF, sparing other highly sensitive retinal tissues. Flt23k, an intraceptor inhibitor of VEGF, was able to inhibit VEGF in vitro. Adeno-associated virus type 2 (AAV2)-mediated expression of Flt23k was maintained for up to 6 months postsubretinal injection in mice. Flt23k was able to effectively inhibit laser-induced murine choroidal neovascularization (CNV). VEGF levels in the RPE/choroid complex decreased significantly in AAV2.Flt23k treated eyes. Neither retinal structure detected by Heidelberg Spectralis nor function measured by electroretinography (ERG) was adversely affected by treatment with AAV2.Flt23k. Hence AAV2.Flt23k can effectively maintain long-term expression and inhibit laser-induced CNV in mice through downregulation of VEGF while maintaining a sound retinal safety profile. These findings suggest a promising novel approach for the treatment of CNV.

Subretinal AAV2.COMP-Ang1 suppresses choroidal neovascularization and vascular endothelial growth factor in a murine model of age-related macular degeneration

“…When tracked by OCT, the retina typically reattaches as the injected subretinal fluid dissipates, without any concomitant impairment to visual function on electroretinography (ERG) or optokinetic tracking (OKT) (Zhang et al, 2015), likely helped by the fact that the penetration of the needle is in the peripheral retina distant from the macula.…”

“…Subretinal AAV2 localization was first tested with well-established reporter genes: AcGFP, which has a fluorescent product (Cereso et al, 2014; Cronin et al, 2012; Li et al, 2008; Zhang et al, 2012, 2015), and LacZ, which has a chromogenic product (Hojo et al, 2004), to validate successful inoculation of the targeted tissue.…”

“…ELISA is a common way to examine VEGF levels in the RPE/ choroid (Itaya et al, 2007; Kim et al, 2008; Zhang et al, 2015). …”

“…The sawtooth pattern of drug delivery entailed by the short half-life of recombinant protein (Zhu et al, 2011) creates a peak-trough problem with drug availability (Zhang et al, 2015) which could be circumvented by long-acting gene therapy with viral vectors (Campochiaro, 2011; Colella and Auricchio, 2010; Roy et al, 2010). The efficacy of Ad.COMP-Ang1 therapy is limited by its transient expression (i.e.…”

“…With specificity for the outer retina and the RPE/choroid complex (Cronin et al, 2012; Muhlfriedel et al, 2013; Zhang et al, 2012, 2015), a subretinal route for AAV2 offers stronger and more diffuse transfection of these layers, along with a milder immune reaction, compared to an intravitreal route (Campochiaro, 2011; Li et al, 2008). Targeting this space may also mitigate the danger of medication-induced global retinal toxicity linked with the intravitreal approach (Campochiaro, 2011; Grunwald et al, 2014; Rofagha et al, 2013; Zhang et al, 2015). …”