PROJECT SUMMARY Glaucoma is a major cause of blindness, affecting over 80 million people worldwide. Glaucoma is a neurodegenerative optic neuropathy caused by the loss of retinal ganglion cells (RGC), leading to loss of vision. Current therapies are all directed at lowering intraocular pressure (IOP), and yet RGC loss still continues in many patients despite IOP lowering. The identification of an agent that complements IOP lowering by promoting RGC survival would be a significant advance toward improving the visual outcomes of patients with glaucoma. Using cultures of primary RGC, we screened more than 10,000 compounds and identified candidates with potent neuroprotective properties, including a drug that is FDA-approved for an unrelated indication. We further characterized the novel pathway through which these compounds act to protect RGC, thus identifying a novel drug/drug target combination for neuroprotection. We have also developed a novel thermosensitive gel-forming eye drop drug delivery system that provides efficacious drug delivery to the posterior segment, even in large animals (rabbits, pigs). Importantly, we observed that the combination of more effective intraocular drug penetration provided by the gel-forming eye drop with a drug that binds to melanin in the eye, led to RGC protection in vivo with once weekly topical dosing. In this application, we are screening additional neuroprotective drugs for melanin binding, cell uptake, and intraocular penetration to compare head- to-head for pharmacokinetics and efficacy in an optic nerve crush rat model and a bead injection mouse model of IOP elevation. The goal is to develop an efficacious eye drop for neuroprotection that requires once weekly, or ideally once monthly maintenance dosing. In Aim 1, we will make further formulation changes in the eye drop to increase intraocular drug absorption, and formulate additional melanin-binding neuroprotective drugs. In Aim 2, we will characterize the pharmacokinetics and efficacy of a variety of dosing regimens and formulations to identify the most effective dosing approach with the lowest dosing frequency. In Aim 3, we will perform dose-ranging studies in rabbits to achieve similar drug concentrations in target tissues that were shown to be effective in Aim 2. We will also perform safety evaluations with longitudinal dosing, including fundus exams, IOP, and retinal morphology analyses. The demonstration of efficacy in rodent models of neurodegeneration along with similar pharmacokinetics and no overt toxicity in the rabbit eye, would provide evidence of the therapeutic potential of our neuroprotective drug delivery strategy for the treatment of glaucoma.