Project Summary Diabetic retinopathy (DR) is a leading cause of vision loss in working adults, affecting more than 100 million people worldwide. A breakthrough in DR therapy is the advent and approval of vascular endothelial growth factor (VEGF) inhibitors as the first pharmacological treatment but with limited efficacy. Additionally, anti-VEGF therapy indiscriminately inhibits both diseased and healthy vessels as well as neuronal function and may indirectly reduce the efficacy to improve visual acuity. An unmet clinical need is to develop drug therapies against VEGF- independent angiogenic factors for alternative or combination therapy to improve efficacy with optimal safety. We recently discovered a novel disease-restricted pro-angiogenic factor that selectively binds to and stimulates angiogenesis and vascular leakage of diabetic but not healthy vessels, whereas VEGF indiscriminately induces angiogenesis and leakage. We further revealed that the new pro-angiogenic factor selectively binds to the DR- stressed deep retinal vascular plexus with compromised endothelial junctions, but not the relatively healthy superficial and intermediate retinal plexuses in diabetic mice with intact endothelial junctions. Therefore, inhibition of this disease-restricted angiogenic factor may selectively target DR-stressed deep retinal plexus, while sparing the relatively healthy superficial and intermediate vascular plexuses within the same diabetic retina. To exploit this unique disease-targeted DR therapy, we generated neutralizing monoclonal antibodies and a related humanized antibody (hAb) against the new angiogenic factor and demonstrated their high efficacy to alleviate DR leakage in diabetic mice. We confirmed that the hAb stringently targets diseased but not healthy vessels. More importantly, combination therapy simultaneously targeting VEGF and the new angiogenic factor with distinct receptor signaling pathways synergistically improved the efficacy to ameliorate DR leakage. The objective of this project is to complete all required preclinical studies for Investigational New Drug (IND) application. In Aim 1, we will analyze IND-enabling pharmacokinetic studies and develop formulation and bioanalytic assays. In Aim 2, we will submit a Pre-IND application to the FDA and seek the agency’s guidance on relevant requirements for a formal IND application. In Aim 3, we will complete IND-enabling toxicology studies. Successful implementation of the proposed studies will advance this first-in-class disease-targeted anti- angiogenic therapy to IND application and clinical trials.