Cyclic AMP (cAMP) is an important second messenger signaling molecule that is involved in wide variety of physiological processes. The canonical cAMP pathway is initiated when hormones or neurotransmitters bind to G- protein coupled receptors (GPCRs) that are coupled to G[alpha]s. This leads to activation of adenylyl cyclases (ACs) that convert ATP to cAMP. There are two main intracellular effectors of cAMP-mediated signaling: Protein Kinase A (PKA) and Exchange Protein Activated by cAMP (Epac). Dysregulation of the cAMP pathway is implicated in a variety of diseases including diabetes mellitus, heart failure, and cancer. While much work has been done to develop pharmacological tools to study the cAMP signaling system, most of these efforts have focused on PKA, GPCRs, and ACs. Studies on Epac are more limited. The work to be described in this dissertation analyzes the dysregulation of the cAMP effector Epac in chronic lymphocytic leukemia (CLL, Chapter 2). I show that Epac is up-regulated at the mRNA and protein level and this contributes to the anti-apoptotic phenotype of the disease. I further show that Lef-1, a transcription factor up-regulated in leukemia, mediates the increased expression of Epac1. Understanding the dysregulation of Epac signaling, such as occurs in CLL, would be enhanced with new pharmacological tools to study Epac. To assist in the development of new tools I developed a BRET-based assay that takes advantage of the conformational changes that Epac undergoes when activated to analyze modulators of Epac activity in vitro (Chapter 3). I demonstrated that this assay can identify competitive inhibitors, such as the cAMP analogs CPT-N6-cAMP and (Rp)-CPT-cAMPS, and can identify allosteric inhibitors such as CE3F4. I further verified these findings in cells by measuring Rap1 activation. I used this assay to experimentally test compounds that were predicted to inhibit Epac activity allosterically (Chapter 4). I identified a novel thiobarbituric acid derivative that binds to the hinge region of the cyclic nucleotide binding domain of Epac1 and Epac2 and inhibits their activity. Together these data implicate Epac as an important signaling protein that is increased in cancer and identify a new approach to identify inhibitors of Epac