Suppression of Choroidal Neovascularization and Fibrosis by a Novel RNAi Therapeutic Agent against (Pro)renin Receptor

Abstract

The receptor-associated prorenin system refers to the pathogenic mechanism whereby prorenin binding to (pro)renin receptor [(P)RR] dually activates the tissue renin-angiotensin system (RAS) and RAS-independent signaling, and its activation contributes to the molecular pathogenesis of various ocular diseases. We recently developed a new single-stranded RNAi agent targeting both human and mouse (P)RR ((P)RR-proline-modified short hairpin RNA [(P)RR-PshRNA]), and confirmed its therapeutic effect on murine models of ocular inflammation. Here, we investigated the efficacy of (P)RR-PshRNA against laser-induced choroidal neovascularization (CNV) and subretinal fibrosis, both of which are involved in the pathogenesis of age-related macular degeneration (AMD). Administration of (P)RR-PshRNA in mice significantly reduced CNV formation, together with the expression of inflammatory molecules, macrophage infiltration, and extracellular signal-regulated kinase (ERK) 1/2 activation. In addition, (P)RR-PshRNA attenuated subretinal fibrosis, together with epithelial-mesenchymal transition (EMT)-related markers including phosphorylated SMAD2. The suppressive effect of (P)RR-PshRNA is comparable with aflibercept, an anti-vascular endothelial growth factor drug widely used for AMD therapy. AMD patient specimens demonstrated (P)RR co-localization with phosphorylated ERK1/2 in neovascular endothelial cells and retinal pigment epithelial cells. These results indicate that (P)RR contributes to the ocular pathogenesis of both inflammation-related angiogenesis and EMT-driven fibrosis, and that (P)RR-PshRNA is a promising therapeutic agent for AMD.

Receptor-Associated Prorenin System in the Trabecular Meshwork of Patients with Primary Open-Angle Glaucoma and Neovascular Glaucoma

“…Our clinical sample data also revealed (P)RR involvement in retinal angiogenesis and systemic inflammation in patients with PDR, showing a close association of RAPS with the molecular mechanism of PDR [37][38][39]. Moreover, increasing evidence has suggested RAPS activation in various human eye disorders including age-related macular degeneration [40], idiopathic epiretinal membrane [41], noninfectious uveitis [42], and conjunctival lymphoma [43].…”

“…In the mouse model of diabetic retinopathy, leukocyte adhesion to the retinal vasculature was revealed to be governed independently by both AT1R and (P)RR [17,32]. Molecules induced via (P)RR downstream pathway and their cellular sources were reported to include monocyte chemotactic protein 1 in brain capillary endothelial cells [34], vascular endothelial growth factor A in macrophages [34] and retinal microvascular endothelial cells [37], transforming growth factor-β1 in retinal pigment epithelial cells [40], and fibroblast growth factor 2 in Müller glial cells [41]. (P)RR signaling would thus lead to various pathologic conditions such as inflammation, angiogenesis, and fibrosis, depending on different cellular reactions in diabetic retinopathy [37], age-related macular degeneration [40], and idiopathic epiretinal membrane [41].…”

The Role of Inflammation in Age-Related Macular Degeneration

“…Subretinal fibrosis is a clinical manifestation of later period of nAMD [91], which is a wound healing response after CNV, together with the damage of photoreceptors, RPE and choroidal blood vessels, causing irreparable visual impairment [19]. Cellular and ECM constituents, and the growth factor mediated EMT act as important roles in the RPE and the complex signaling networks of fibrosis in AMD [28].…”

Blockade of Platelet-Derived Growth Factor Signaling Inhibits Choroidal Neovascularization and Subretinal Fibrosis in Mice

“…The laser-induced CNV model was generated as described previously [ 20 ]. In brief, laser injury (532 nm, 180 mW, 75 μm, 100 ms, Novus Spectra; Lumenis, Tokyo, Japan) was conducted around the optic disc using a slit-lamp delivery system.…”

“…The size of CNVs and subretinal fibrosis was measured at 7 and 21 days after laser injury, as described previously [ 20 ]. In brief, the anterior segment and retina were removed from the eye after fixation in 4% paraformaldehyde.…”