PROJECT SUMMARY: Myeloid derived suppressor cells (MDSCs) are immature myeloid cells with potent inhibitory properties for macrophages and T cells. The link between MDSCs and tuberculosis (TB), a disease caused by Mycobacterium tuberculosis (Mtb), is supported by work in mice where MDSC recruitment to the lungs correlates with disease severity, and in humans, where peripheral blood MDSC abundance positively correlates with TB progression. Work in mice also demonstrates that MDSCs are recruited to sites of BCG vaccination where they phagocytose bacteria and suppress local T cell activation, suggesting that MDSCs may reduce the effectiveness of anti-TB vaccination and contribute to TB’s status as a global public health concern. The relationship between MDSCs and TB has generated interest in targeting them with host-directed therapies to improve TB treatment. Recent work using tasquinimod (TSQ), an FDA-approved small molecule MDSC inhibitor, in guinea pigs and mice finds that targeting MDSCs lowers reduces granuloma formation, lowers bacteria loads, and improves the treatment efficacy of anti-TB drugs. These results are encouraging but critical questions need to be answered before MDSCs can be safely targeted in human TB. Knowledge gaps include an incomplete understanding of how MDSCs interact with macrophages in diseased tissue, if MDSCs restrain or promote pathologic inflammation in TB, how MDSC activity relates to bacterial burden in granulomas, and if MDSCs permit or restrict Mtb replication. These gaps are particularly acute in human TB where progress is inhibited by the inability to access granulomas to study MDSCs in situ and fundamental differences between mouse and human TB pathophysiology. Our proposal addresses these gaps with innovative studies using MDSCs from human cord blood and a nonhuman primates (NHPs) model that accurately reflects human TB pathophysiology. We hypothesize that MDSCs suppress macrophage-mediated anti-Mtb activity and are permissive hosts for Mtb and inhibiting their functions will improve immunity, restrict bacterial persistence, and improve outcomes in TB. In Aim 1, we propose mechanistic studies investigating relationships between MDSCs, macrophages, and Mtb that influence immunity. We will determine if cell-free DNA release is a novel MDSC effector that suppresses macrophage function. We also determine if MDSCs support Mtb as a permissive host cell and define molecular features at the phagosomal level that contribute to competency for hosting Mtb. In Aim 2, we infect NHPs with mCherry-expressing Mtb and use TSQ-mediated MDSC inhibition as an intervention to assess how MDSCs regulate lung inflammation, immunity, and if they host Mtb. Here, we use PET/CT and fluorescence imaging, quantitative microbiology, and flow cytometry to identify MDSC-regulated correlates of immunity in granulomas. We also define MDSC transcriptional profiles in scRNAseq and use CosMx spatial molecular imaging to identify how MDSCs regulate...