Obesity is a growing epidemic in the United States, leading to increases in cases of nonalcoholic fatty liver disease (NAFLD), cardiovascular disease, and type 2 diabetes. A common characteristic of these diseases is aberrant lipid and glucose metabolism. This proposal centers on the nuclear hormone receptor, Liver Receptor Homolog 1 (LRH-1), which acts as an important regulator of lipid metabolism, reverse cholesterol transport, glucose sensing, and homeostasis. As such, LRH-1 represents a novel therapeutic target for metabolic diseases. LRH-1 binds to phospholipids (PLs), but until recently, the role of PLs in receptor activation was unclear. Recent studies identified dilauroylphosphatidylcholine (DLPC) as a specific LRH-1 agonist with potent anti-diabetic effects. Despite this untapped therapeutic potential, mechanisms through which LRH-1 is regulated by ligands remain poorly understood. The discovery that LRH-1 is regulated by PL ligands reveals an exciting potential to tune LRH-1 activity for the treatment of metabolic diseases. However, PLs (such as DLPC) are labile and not suitable for clinical use, necessitating the development of small molecule agonists. This has proved challenging thus far, since very few small molecules are capable of displacing endogenous lipids from the large, lipophilic binding pocket. Recent preliminary studies in our lab have characterized a potent class of small molecules that are capable of this feat. Our crystal structures of LRH-1 bound to a set of these agonists have uncovered novel mechanisms of receptor activation and have provided insights into strategies to improve agonist activity. The overall goal of this proposal is to develop improved small molecule modulators using a structure-based, rational design approach. X-ray crystallography combined with hydrogen deuterium exchange mass spectrometry, cellular assays, and animal studies will be used to address this goal in a series of three Aims. Aim 1. Design and synthesize potent, effective, and selective LRH-1 modulators Aim 2. Determine the molecular basis of allosteric modulation of LRH-1 by RJW100 analogues Aim 3. Evaluate the efficacy of LRH-1 modulators as anti-diabetic agents in obesity.