ABSTRACT This application seeks to address the unmet need for curative therapies in pulmonary arterial hypertension (PAH), a fatal disease with a dismal prognosis. A key feature of the pathobiology of PAH is the profound pulmo- nary vascular remodeling for which new therapeutic strategies are woefully lacking. We successfully employed genomic?intensive approaches to identify nicotinamide phosphoribosyltransferase (NAMPT) as a novel PAH target that is robustly upregulated in PBMCs from PAH patients. We also demonstrated markedly increased NAMPT expression in remodeled vessels from human PAH subjects and in preclinical PAH models (mice and rats) with NAMPT localized to lung endothelial cells (ECs). We further reported that reducing the availability of secreted or extracellular NAMPT or eNAMPT (via eNAMPT neutralizing antibodies, siRNAs, Nampt+/- mice) dra- matically attenuated PAH severity in our preclinical PAH models. To test the hypothesis that NAMPT promotes vascular remodeling during PH development and serves as a novel PAH therapeutic target, Specific Aim #1 (SA #1) will further characterize the regulation of NAMPT expression in response to PAH stimuli (PDGF, VEGF, PHD2 inhibitor, endothelin-1) focusing on promoter activity and epigenetic regulation (DNA methylation, miR- NAs) and specific transcription factors (HIF-2a, STAT5, SOX17) that we have shown to regulate NAMPT tran- scription. SA #2 will mechanistically examine the contribution of extracellular NAMPT (eNAMPT) to vascular remodeling via influences on EC apoptosis and smooth muscle cell (SMC) activation (Ca2+ signaling and prolif- eration). We will specifically focus on the interaction of eNAMPT with Toll?like receptor 4 (TLR4), that we recently identified as the NAMPT receptor and explore eNAMPT-TLR4 mediated NF?B transcriptional activities as a novel mechanism by which eNAMPT may influence vascular remodeling. SA #3 will leverage our prior published stud- ies and preliminary data demonstrating that NAMPT 5? promoter polymorphisms (SNPs) alter NAMPT promoter activity which also confer significantly increased risk for susceptibility and severity in acute respiratory distress syndrome (ARDS) and will be assessed in PAH. SA #3 studies will determine the influence of NAMPT SNPs on transcriptional regulation, on eNAMPT-TLR4 binding, and NF?B activation in PAH. Finally, SA #4 will define in established PAH, the therapeutic efficacy of reduced NAMPT expression (conditional EC knockout mice), eN- AMPT elimination (neutralizing antibodies), inhibition of NAMPT enzymatic activity (novel FK-866 analogues), and TLR4 antagonism (novel peptide inhibitors). Supported by intimate involvement of outstanding investigators and substantial highly translational published/preliminary data highlighting NAMPT as a novel innate immunity modulator, this application will successfully define NAMPT participation in PAH susceptibility, pathobiology, and severity.