Protein delivery to the eye: assessing therapeutic potential across inner and outer retina.

Protein-based therapeutics represent a promising approach for treating diseases of the retina, yet effective delivery systems and translational models remain limited. We developed a robust ex vivo framework utilizing murine and porcine retinal explants to investigate protein delivery mechanisms for both peripheral and central retinal regions. Using guanylate cyclase-activating protein-1 (GCAP1) as a retina-specific model protein and mCardinal2 as a non-retina-specific control, we demonstrated spontaneous protein internalization and accumulation in photoreceptors and ganglion cells following single-dose administration. In murine explants, myristoylated GCAP1 exhibited sustained retention and appropriate subcellular targeting in rod photoreceptors over 96 h without inducing tissue damage. Porcine explants, which better recapitulate human macular architecture, revealed differential protein trafficking dependent on post-translational modifications. Retinal tissue showed preferential uptake and retention of retina-specific proteins over foreign proteins, suggesting sophisticated protein-specific recognition mechanisms. Liposomal encapsulation enhanced initial uptake of non-retina-specific proteins but did not improve long-term retention. This experimental platform provides a valuable tool for screening protein therapeutics, optimizing delivery formulations, and investigating protein-selective cellular uptake mechanisms, with direct implications for developing treatments for inherited retinal degenerations, age-related macular degeneration, and optic nerve pathologies.