When more than two value sets were compared, we used one-way ANOVA followed by the Dunnett test when the data involved three or more groups. mass cytometry and multiplex immunofluorescence techniques. We uncovered that IL17 recruits neutrophils, triggers neutrophil extracellular traps (NETs), and excludes cytotoxic CD8 T cells from tumors. Additionally, IL17 blockade increases immune checkpoint blockade (PD-1, CTLA4) sensitivity. Inhibition of neutrophils or Padi4-dependent NETosis phenocopies IL17 neutralization. NMR spectroscopy revealed changes in tumor lactate as a potential early biomarker for IL17/PD-1 combination efficacy. Higher expression of IL17 and PADI4 in human PDAC corresponds with poorer prognosis, and the serum of patients with PDAC has higher potential for NETosis. Clinical studies with IL17 and checkpoint blockade represent a novel combinatorial therapy with potential efficacy for this lethal disease. Graphical Abstract Open in a separate window Introduction Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest malignancies, with limited treatment options. Several immunotherapy approaches have emerged in the past decade, and immune checkpoint blockade has been approved for use in several cancer types (Le et al., 2015; Brahmer et al., 2015; Diaz and Le, 2015; Wolchok et al., 2017; RGS9 DAngelo et al., 2017; Long et al., 2017). Unfortunately, immune checkpoint inhibitors have not proved efficacious in treating TD-106 pancreatic cancer (Royal et al., 2010; Brahmer et al., 2012; Herbst et al., 2014; Patnaik et al., 2015; Balachandran et al., 2019). Recently, considerable interest has been focused on combining checkpoint inhibitors with other immunotherapies, antibodies, or vaccines to target or prime the tumor microenvironment, one of the main drivers of the immunosuppression that prevails in pancreatic cancer (Lutz et al., 2014; Highfill et al., 2014; Zhu et al., 2014; Winograd et al., 2015). Several cell types have been implicated in contributing to the immunosuppressive microenvironment that supports PDAC growth: macrophages, myeloid-derived suppressor cells (MDSC), fibroblasts, and T regulatory cells (Clark et al., 2007; Bayne et al., 2012). Strategies that target these cell types in combination with immune checkpoint inhibitors have been proved to have synergistic antitumoral effects in preclinical models of pancreatic cancer as well as other cancer types (Zhang et al., 2017; Zhu et al., 2014; Highfill et al., 2014; Feig et al., 2013; Provenzano et al., 2012), and some of them are being tested in ongoing clinical trials (“type”:”clinical-trial”,”attrs”:”text”:”NCT02452424″,”term_id”:”NCT02452424″NCT02452424, “type”:”clinical-trial”,”attrs”:”text”:”NCT02777710″,”term_id”:”NCT02777710″NCT02777710). We have previously described that IL17, a cytokine secreted mostly by CD4+ and T cells during pancreatic tumorigenesis, is involved in the initiation and development of pancreatic precursor lesions of PDAC (McAllister et al., 2014). The interaction between IL17 and IL17RA, which is overexpressed in the epithelium upon Kras activation, promotes a stemness signature in premalignant lesions (McAllister and Leach, 2014; TD-106 Zhang et al., 2018). In the present study, we detected high levels of IL17 in tumors from both autochthonous and transplanted orthotopic PDAC mouse models. We found that IL17 sustains immunosuppression by decreasing CD8+ T cell recruitment and activation while promoting neutrophil recruitment in the tumor microenvironment. IL17 blockade sensitizes tumors to checkpoint inhibitors in a CD8+ T cellCdependent manner. We explored mechanisms implicated in this process as well as biomarkers. Results IL17-secreting cells are increased in murine and human pancreatic adenocarcinoma Given the key role that IL17 exerts in the initiation and progression of pancreatic premalignant lesions (McAllister et al., 2014), we first aimed to determine the dynamic systemic levels of IL17 that change with progression in pancreatic adenocarcinoma mouse models. To this end, we measured the serum concentration of a panel of cytokines in the autochthonous K-rasLSL.G12D/+;p53R172H/+;PdxCre (KPC) pancreatic adenocarcinoma mouse model at two time points (1 and 6 mo). We found that serum levels of several cytokines were increased in KPC mice in an age-dependent manner (Fig. 1 A). When we compared KPC mice with age-matched control PdxCre mice, we found that serum levels of IL17 drastically increased with PDAC progression (Fig. 1, A and B). We then measured IL17 in an orthotopic PDAC mouse model in which KPC cells were allografted into the pancreas of syngeneic animals and found that IL17 mRNA expression in orthotopic pancreatic tumors was significantly upregulated compared with normal pancreas (Fig. 1 C). We also detected T helper type 17 (Th17) cells in human PDAC tissue, which are absent or in very low numbers in normal tissue (Fig. 1, D and E). We also found that patients with higher IL17A expression in their pancreatic tumors, based on The Cancer Genome Atlas (TCGA), had significantly worse prognosis than those with lower IL17 expression (hazard ratio [HR], 2.2; P = 0.0021; Fig. 1 F). Open in a separate window Figure 1. IL17-secreting cells are increased in murine and human pancreatic adenocarcinoma TD-106 carcinoma. (A) TD-106 Heat map representing serum levels of cytokines from a spontaneous pancreatic.