Selective binding of C-6 OH sulfated hyaluronic acid to the angiogenic isoform of VEGF(165)

Abstract

Vascular endothelial growth factor 165 (VEGF(165)) is an important extracellular protein involved in pathological angiogenesis in diseases such as cancer, wet age-related macular degeneration (wet-AMD) and retinitis pigmentosa. VEGF(165) exists in two different isoforms: the angiogenic VEGF-(165a), and the anti-angiogenic VEGF(165b). In some angiogenic diseases the proportion of VEGF(165b) may be equal to or higher than that of VEGF(165a). Therefore, developing therapeutics that inhibit VEGF(165a) and not VEGF(165b) may result in greater anti-angiogenic activity and therapeutic benefit. To this end, we report the selective binding properties of sulfated hyaluronic acid (s-HA). Selective biopolymers offer several advantages over antibodies or aptamers including cost effective and simple synthesis, and the ability to make nano-particles or hydrogels for drug delivery applications or VEGF(165a) sequestration. Limiting sulfation to the C-6 hydroxyl (C-6 OH) in the N-acetyl-glucosamine repeat unit of hyaluronic acid (HA) resulted in a polymer with strong affinity for VEGF(165a) but not VEGF(165b). Increased sulfation beyond the C-6 OH (i.e. greater than 1 sulfate group per HA repeat unit) resulted in s-HA polymers that bound both VEGF(165a) and VEGF(165b). The C-6 OH sulfated HA (Mw 150 kDa) showed strong binding properties to VEGF(165a) with a fast association rate constant (K-a; 2.8 x 10(6) M-1 s(-1)), slow dissociation rate constant (K-d; 2.8 x 10(-3) s(-1)) and strong equilibrium binding constant (K-D); similar to 1.0 nM)), which is comparable to the non-selective VEGF(165) binding properties of the commercialized therapeutic anti-VEGF antibody (Avastin (R)). The C-6 OH sulfated HA also inhibited human umbilical vein endothelial cell (HUVEC) survival and proliferation and human dermal microvascular endothelial cell (HMVEC) tube formation. These results demonstrate that the semi-synthetic natural polymer, C-6 OH sulfated HA, may be a promising biomaterial for the treatment of angiogenesis-related disease. (C) 2015 Elsevier Ltd. All rights reserved.