Intraocular Penetration of a vNAR: In Vivo and In Vitro VEGF165 Neutralization

Abstract

Variable new antigen receptor domain (vNAR) antibodies are novel, naturally occurring antibodies that can be isolated from naïve, immune or synthetic shark libraries. These molecules are very interesting to the biotechnology and pharmaceutical industries because of their unique characteristics related to size and tissue penetrability. There have been some approved anti-angiogenic therapies for ophthalmic conditions, not related to vNAR. This includes biologics and chimeric proteins that neutralize vascular endothelial growth factor (VEGF)165, which are injected intravitreal, causing discomfort and increasing the possibility of infection. In this paper, we present a vNAR antibody against human recombinant VEGF165 (rhVEGF165) that was isolated from an immunized Heterodontus francisci shark. A vNAR called V13, neutralizes VEGF165 cytokine starting at 75 μg/mL in an in vitro assay based on co-culture of normal human dermal fibroblasts (NHDFs) and green fluorescence protein (GFP)-labeled human umbilical vein endothelial cells (HUVECs) cells. In the oxygen-induced retinopathy model in C57BL/6:Hsd mice, we demonstrate an endothelial cell count decrease. Further, we demonstrate the intraocular penetration after topical administration of 0.1 μg/mL of vNAR V13 by its detection in aqueous humor in New Zealand rabbits with healthy eyes after 3 h of application. These findings demonstrate the potential of topical application of vNAR V13 as a possible new drug candidate for vascular eye diseases.

Keywords

• vNAR
• single chain binding domain
• VEGF165
• intraocular penetration
• Heterodontus fransisci
• horn shark
• age-related macular degeneration
• diabetic retinopathy