PROJECT SUMMARY/ABSTRACT MOLECULAR REGULATION OF CARDIOVASCULAR SEVEN TRANSMEMBRANE RECEPTORS. All aspects of cardiovascular function are regulated by members of the seven transmembrane receptor (7TMR or GPCR) family, and they are the commonest target of therapeutic drugs. A universal mechanism regulating these receptors is desensitization of heterotrimeric G protein signaling. Classically, this is mediated by a two-step process in which activated receptors are phosphorylated by G protein-coupled receptor kinases, leading to the binding of a β-arrestin (βarr) molecule which sterically interdicts further activation of the G protein. More recently it has become clear that βarrs can also serve as multifunctional adaptors which act as signal transducers in their own right. Moreover, for many receptors, ligands can be found which disproportionately activate either G protein- or βarr-mediated signaling—i.e., "biased ligands" which may possess greater specificity of action and fewer side effects. Biased drugs for GPCR's represent a major, as yet largely unmined opportunity for drug development. Another such untapped opportunity to develop drugs with greater specificity and safety is presented by allosteric ligands which, rather than binding to the orthosteric site of the receptors, where endogenous ligands bind, interact with other sites. Accordingly, this proposal has three closely linked aims which involve laying the groundwork for targeting both of these largely untapped opportunities by developing the first biased allosteric ligands for a receptor of great cardiovascular significance, the angiotensin II type I receptor (AT1R) as well as for the βarrs. In the process we aim to also develop a molecular and atomic level understanding of how biased allosteric GPCR drugs regulate signaling. These aims are: 1. To discover the first small molecule biased allosteric modulators of the AT1R; 2. To discover novel small molecule allosteric modulators of the signal transducer βarr; 3. To elucidate the mechanisms of allosteric regulation of downstream effectors of βarrs. The insights which we will generate have the potential to guide the design of powerful new cardiovascular drugs and will further our understanding of the conserved signaling mechanisms of the greater GPCR family.