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

Abstract 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.