PROJECT ABSTRACT This proposed career development award will provide Dr. Danielle Alfano, MD with targeted mentored training to ensure she develops into an independent researcher utilizing both experimental approaches, mouse models, and “omics”, to probe how specific pathogens harness ADAM10 leading to the dysregulated endothelial and host response in sepsis. Sepsis is a complex syndrome defined as a ‘dysregulated host response to infection leading to life-threatening organ dysfunction’. While many studies in the field have focused on an aberrant host immune response as the instigator of severe sepsis, all efforts have failed to translate this into new therapies. The endothelium plays a critical role in the host response to infection and organ injury. We have recently demonstrated that ADAM10, the eukaryotic receptor for S. aureus α-toxin, acts a molecular specifier of sepsis, mediating mortality and endothelial injury to a diverse subset of pathogens. However, the precise molecular mechanisms remain poorly understood. The proposed research plan aims to close key knowledge gaps regarding how specific pathogens harness ADAM10 in disease and the molecular mechanisms of endothelial injury. To do so, the PI will leverage multiple live pathogens and unique mouse lines to fully characterize the nature of ADAM10 functions that contribute to sepsis progression. The PI aims to 1) understand the molecular mechanisms by which diverse pathogens activate endothelial ADAM10, 2) characterize ADAM10 specific substrates released during systemic infection, and 3) examine how an ADAM10 SNP polymorphism may confer increased risk of lethal disease. The proposed 5-year career development and training plan incorporates strategically designed didactic learning, mentored practical training, and career advising to complement the PI’s expertise in ways that are critical to completion of her research and career goals. The specific career development goals outlined in this application include developing mechanistic expertise in 1) vascular biology and intracellular signaling pathways; 2) host-pathogen interactions; and 3) proteomics. She will be training at WUSM, a world- class center for basic and translational research and an excellent environment for physician- scientist training with experts in all aspects of the proposed training. She will be closely mentored by Dr. Juliane Bubeck Wardenburg, an expert in S. aureus, ADAM10, host-pathogen interactions, and immunology. The long- term goal is to provide Dr. Alfano with the skills required to become an independent, R01- funded faculty member working to study pathogen-specific molecular determinants of sepsis and factors that confer increased risk of life-threatening sepsis to certain populations to provide novel insights into future therapeutic developments.

PROJECT NARRATIVE Sepsis is a complex clinical infectious disease that represents one of the leading causes of death across the world. Decades of research have failed to produce new clinically efficacious treatments, mandating the development of novel strategies to prevent and treat infection. The design of such strategies necessitates that we obtain a clear understanding of the mechanisms by which diverse human pathogens injure the endothelium causing lethal disease, which forms the focus of this proposal.