Exploiting retinal endothelial heterogeneity for disease-targeted therapy of diabetic retinopathy

Abstract

Purpose : The purpose is to investigate contributions of retinal endothelial heterogeneity to the mechanism of disease-targeted therapy of diabetic retinopathy (DR) by antibodies against secretogranin III (Scg3), a DR-restricted angiogenic and vascular leakage factor.

Methods : Scg3- and VEGF-induced retinal vascular leakage in healthy and streptozotocin-induced 4-month diabetic mice were visualized and quantified using FITC-dextran. CD31 expression in the capillary plexuses of flat-mounted retinas was analyzed by immunohistochemistry (IHC). Scg3 and VEGF binding to diabetic and healthy mouse retinas was quantified by an innovative method of in vivo ligand binding (IVLB) assay. Binding sites of Scg3 and VEGF in the retinal vascular plexuses were visualized by a unique technique of functional IHC (FIHC). The therapeutic efficacy of anti-Scg3 humanized antibody (hAb) to ameliorate retinal vascular leakage in wild-type and Scg3^-/- diabetic mice was analyzed by Evans Blue assay and compared to aflibercept.

Results : FITC-dextran assay showed that Scg3 selectively stimulates retinal vascular leakage in diabetic but not healthy mice, whereas VEGF induces leakage indiscriminately. IHC revealed that the expression of endothelial junction protein CD31 is significantly downregulated in the deep plexus of diabetic retina. In contrast, both diabetic and healthy retinas showed similar levels and patterns of CD31 expression in the superficial and intermediate plexuses. IVLB detected a 22.7-fold increase in Scg3 binding to diabetic vs. healthy retinas. Importantly, FIHC revealed that Scg3 selectively binds to the CD31^- deep plexus but not relatively healthy CD31⁺ superficial and intermediate plexuses of the same diabetic retina. In contrast, minimal VEGF binding was detected by IVLB and FIHC in healthy and diabetic retinas. Evans blue assay revealed similar efficacies for both anti-Scg3 hAb and aflibercept to alleviate DR leakage in wild-type, whereas aflibercept, but not anti-Scg3 hAb, is equally effective in Scg3^-/- mice.

Conclusions : Scg3 selectively binds to CD31^- endothelial cells of the deep retinal plexus in DR mice, supporting a role for endothelial heterogeneity in Scg3-mediated regulation of DR leakage. Such heterogeneity provides an avenue for anti-Scg3 therapy to alleviate DR leakage by preferentially targeting the deep plexus without affecting the relatively healthy superficial and intermediate plexuses.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.