Supramolecular Self‐Assembled Multivalent Anti‐VEGF Protein Nanospheres for Prolonged Ocular Retention

To meet the therapeutic demands for long-acting anti-vascular endothelial growth factor (VEGF) agents in retinal vascular diseases-specifically prolonged half-life, high bioactivity, and high concentration with low viscosity-a supramolecular anti-VEGF protein nanosphere (Fer-VD) featuring enlarged hydrodynamic size and multivalent binding capacity is developed and evaluated. Fer-VD is engineered by genetically fusing the VEGF-binding domain of VEGF receptor (VEGFR) to a ferritin monomer via a flexible linker, enabling self-assembly into a well-defined 24-mer nanostructure (≈29 nm diameter, ≈1100 kDa). The resulting construct retains high-affinity VEGF binding and demonstrates potent inhibition of VEGF-induced signaling in vitro and in cellular assays. Biophysical characterization reveals that Fer-VD possesses excellent thermal and colloidal stability, maintains solution homogeneity at high concentrations, and exhibits favorable viscosity profiles compatible with intravitreal administration. In rabbit intravitreal pharmacokinetic studies, Fer-VD achieves a two-fold extension in vitreous half-life compare to aflibercept, indicating enhanced ocular retention. Immunohistochemical analysis further demonstrates that Fer-VD localized primarily to the inner retinal surface post-injection, consistent with diffusion-limited transport due to its supramolecular size. Together, these findings support Fer-VD as a promising long-acting VEGF antagonist with improved pharmacokinetic properties.