PROJECT SUMMARY Myeloid-derived suppressor cells (MDSCs) have emerged as key suppressor cells that inhibit effector immunity both in tumors and in the TB granuloma. In addition, MDSCs have been implicated in HIV progression. We recently demonstrated that high glutamine (Gln) levels in the tumor microenvironment (TME) foster immunotolerance and tumor progression. Furthermore, we showed that inhibitors of glutamine metabolism demonstrate potent anticancer activity in part by reprogramming MDSCs to pro-inflammatory M1-type macrophages. Based on seminal work from Marcus Horwitz at UCLA from 1994-2005 which demonstrated the essentiality of a secreted Mycobacterium tuberculosis (Mtb) glutamine synthase and therapeutic benefit with Gln synthase inhibitors in animal models, we investigated whether high Gln levels in the TB granuloma may similarly suppress host immune function. In a three-way collaboration between a tumor immunobiologist (Powell), a chemist with expertise in Gln metabolism (Slusher), and a TB expert (Bishai), we have evaluated novel Gln metabolism inhibitors that are active anticancer drugs for their effectiveness against TB. In contrast to the earlier work of Horwitz et al., we focused on Gln metabolism inhibitors with improved safety profiles and bioavailability, some of which are currently entering human clinical trials as anticancer agents. We observed potent anti-TB activity with Gln metabolism inhibitors in mice with both reductions in Mtb organ burden and prolongation of survival. This was accompanied by significant reductions in lung MDSCs, a corresponding increase in pro-inflammatory M1-type macrophages, and an increase in activated CD8 T cells in murine TB. Our central scientific premises are that (i) a novel Mtb virulence mechanism is release of excess Gln within granulomas leading to MDSC expansion and an immunotolerant microenvironment that enables pathogen survival and (ii) that Gln metabolism inhibitors may represent a valuable host-directed therapy (HDT) approach for the treatment of TB via MDSC inhibition and enhancement of effector T cell immunity. This application will further define the immunosuppressive roles Gln in the TB granuloma and also investigate a panel of new Gln metabolism inhibitors as TB therapeutics. In Aim 1 we will assess novel Gln metabolism inhibitors for their anti-TB therapeutic efficacy using validated animal models. In Aim 2 we will assess the impact of Gln metabolism inhibitors on myeloid cell populations--including MDSCs--during murine Mtb infection. And in Aim 3 we will evaluate the impact of Gln metabolism inhibitors on lymphoid cell activity during murine Mtb infection. These studies may pave the way for Gln metabolism inhibitors that are currently being developed as anticancer drugs to be repurposed as host-directed therapies for TB and TB-HIV.