Project Summary Extracellular ligands are among the most valuable drug targets, but are much less exploited for disease therapy than plasma membrane receptors. This is mainly because the technical hurdles for ligand discovery are much greater than those for receptors, the latter of which can be conveniently identified based on their transmembrane domains. Current omics technologies, such as proteomics, functional proteomics and transcriptomics, are not designed to map extracellular ligands. Consequently, extracellular ligands are traditionally identified and characterized on a case-by-case basis with inherent technical challenges. It is even more daunting to delineate ligands with therapeutic potential. To address these challenges, we recently developed a unique platform technology of ligandomics for global mapping of cell-wide ligands as well as disease-selective ligands in the absence of receptor information. The validity and utility of this innovative technology has been demonstrated by efficient discovery of highly disease-selective angiogenic factors and rapid development of novel anti-angiogenic therapies with high efficacy and minimal side effects on normal vessels. However, this first-generation ligandomics technology has the limited application only to homogeneous cells. In this project, we will develop next-generation ligandomics technology that is applicable to any type of cells. In Aim 1, we will develop the next- generation ligandomics technology to map ligand-secreting and binding cells connected through different ligand signaling pathways. In Aim 2, we will apply the new technology to diabetic and healthy retina to systematically identify diabetes-selective cellular ligands. The successful implementation of this project will markedly improve the capacity of ligandomics to identify cellular ligands and disease-selective ligands for different cells. Our new ligandomics technology developed in this project is universally applicable to any cells, tissues and diseases, and therefore may have the potential to impact on a broad field of cell biology research and drug target discovery.