ab1793) (Abcam). mechanisms whereby immune checkpoint therapy for cancer paradoxically activates TB infection. In patients, PD-1 is expressed in (Mtb)-infected lung tissue but is absent in areas of immunopathology. In the microsphere model, PD-1 ligands are up-regulated by infection, and the PD-1/PD-L1 axis is further induced by hypoxia. Inhibition of PD-1 signalling increases Mtb growth, and augments cytokine secretion. TNF- Bromperidol is responsible for accelerated Rabbit Polyclonal to FSHR Mtb growth, and TNF- neutralisation reverses augmented Mtb growth caused by anti-PD-1 treatment. In human TB, pulmonary TNF- immunoreactivity is increased and circulating PD-1 expression negatively correlates with sputum TNF- concentrations. Together, our findings demonstrate that PD-1 regulates the immune response in TB, and inhibition of PD-1 accelerates Mtb growth via excessive TNF- secretion. (Mtb) having undergone prolonged co-evolution (Menardo et al., 2019). TB has often been thought to primarily result from loss of immune control, because approximately 90% individuals infected with TB never progress to active disease, and this progression is increased in the context of immune deficiency; such as in cases of HIV infection, in infants, people with genetic deficiency of the IL-12/IFN- signalling pathway or after anti-TNF- antibody treatment (O’Garra et al., 2013). However, an emerging concept is that an excessive immune response to Mtb may be equally harmful. Standard disease paradigms predict that immune activation resulting from the administration of checkpoint inhibitors should lead to better control of Mtb infection (Zumla et al., 2016). However, counter-intuitively these agents seem to be activating TB, as evidenced by recent reports of TB developing in patients treated for malignancy with immune checkpoint inhibition, often rapidly after commencing therapy (Fujita et al., 2016; Lee et al., 2016; Chu et al., 2017; Picchi et al., 2018; Jensen et al., 2018; Elkington et al., 2018; He et al., 2018; Takata et al., 2019; Barber et al., 2019; Tsai et al., 2019; van Eeden et al., 2019). Consistent with this emerging clinical phenomenon, programmed death (PD-1) deficient mice are highly susceptible to TB, dying Bromperidol more rapidly than T-cell deficient mice (Lzr-Molnr et al., 2010; Barber et al., 2011). PD-1 and its ligand PD-L1 are expressed in human granulomas (Elkington et al., 2018), suggesting a regulatory role at the site of disease. TB granulomas are hypoxic (Belton et al., 2016), and PD-L1 is up-regulated by hypoxia (Noman et al., 2014), further suggesting a mechanistic link between hypoxia and the PD-1/PD-L1 axis within TB lesions. In this study, we investigate the expression patterns of PD-1 and PDL-1 within TB infected human lung tissue and the relationship between PD-1 and anti-TB immunity. Next, using a human 3D cell culture model of TB (Tezera et al., 2017a), we show that hypoxia increases Bromperidol expression of PD-1 and its ligands, that PD-1 inhibition increases Mtb growth. Surprisingly, TNF- is primarily responsible for this effect, and TNF- neutralisation reverses the anti-PD-1 induced phenotype. Results PD-1 is expressed in human TB granulomas but not in areas of immunopathology First, we investigated the presence and localisation of PD-1-expressing T cells in human pulmonary TB. We hypothesised that PD-1 would be expressed by T cells in the lung of patients with TB, and at a higher frequency than in the blood. In thirty-five patients undergoing medically indicated lung resection to treat TB or TB sequalae, PD-1 expression was measured on T cells isolated from the lung and matched blood Bromperidol samples, available for 23 patients, by flow cytometry. Overall, PD-1 expression in homogenized lung tissue was highly variable, with a trend towards increased PD-1 in both CD4 and CD8 T-cells from the lung compared to matched blood, which reached statistical significance for CD8 T-cells (Figure 1A). Lung tissue from healthy individuals was not available for study, however, the median frequency of 11% and 14% Bromperidol of PD-1+ CD4 and CD8 T-cells observed are generally lower than recently reported for healthy human lung tissue from organ.