Project Summary Pulmonary diseases are a major health concern, with pneumonia the leading infectious cause of mortality for young children. Respiratory infections in early life are associated with increased susceptibility to asthma; an inflammatory disorder that afflicts more than 250 million people each year and has increased in prevalence during the past decade. However, the lack of vaccines for many of the pathogens that cause pneumonia and the emergence of antibiotic resistant bacteria necessitate alternative approaches. Mucosal-associated invariant T (MAIT) cells comprise a substantial effector population within human lungs, reaching up to 9% of pulmonary T cells in healthy individuals. MAIT cells recognize microbial derivatives of riboflavin (vitamin B2) synthesis presented by the MHC class-I related (MR1) molecule. Due to the broad conservation of this biosynthetic pathway among bacteria and fungi, MAIT cells promote immunity to a wide array of respiratory pathogens. MAIT cells have also been implicated in preventing asthma and reducing airway inflammation. However, in addition to presenting derivatives of vitamin synthesis, recent work has demonstrated that MR1 can also bind drugs and drug metabolites that can either be agonists or non-agonists of MAIT cells, suggesting that drugs may hinder MR1-mediated presentation of endogenous microbial metabolites, which could have implications for the susceptibility to pneumonia or asthma. While derivatives of riboflavin synthesis increase MAIT cell abundance and cytokine production in vivo, the instability of these molecules results in rapid degradation under physiological conditions, limiting their therapeutic potential. We will perform the first evaluation of how MR1 binding drugs impact the development of MAIT cells and the first in vivo assessment of how non-vitamin derived MR1 ligands affect MAIT cell responses to physiologically relevant pulmonary conditions. We will also perform the most extensive virtual screen to date for novel MAIT cell agonists using Reactive Docking to simulate the formation of a Schiff base with K43 of MR1 – a key characteristic of the strongest known MAIT cell agonists the promotes MR1 folding for surface expression. Following validation of MR1 binding and MAIT cell agonistic activity in vitro, the immunomodulatory effects of these novel agonists will be established in vivo. We hypothesize that we will identify novel MAIT cell agonists that enhance pulmonary immunity, while administration of non-agonistic MR1 binding drugs can inhibit the development and function of MAIT cells in vivo.