<b>Neutrophil extracellular trap inhibition revitalizes PDAC immunotherapy responsiveness via reduced fibrosis and TCF1</b> ^<b>+</b> <b>CD8</b> ^<b>+</b> <b>progenitor T-cell expansion</b>

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease with poor survival. The immunosuppressive tumor microenvironment (TME) drives resistance to therapy, including immunotherapy. This may be, in part, mediated by the formation of neutrophil extracellular traps (NETs), formed when neutrophils release their intracellular contents. NETs are elevated in PDAC and are associated with nearly every stage of tumor progression. We investigated the influence of NETs on the PDAC TME and immunotherapy response. An orthotopic PDAC model was utilized in C57BJ6 or PAD4^-/- mice receiving one of the following: control, DNase, anti-PD-1 therapy, or DNase and anti-PD-1. NET markers, fibrosis, and the TME immune profile were evaluated. Human PDAC patients were also evaluated for levels of NETs and tumor-infiltrating T cells. Circulating NET markers correlated with intra-tumoral CD8⁺ cells in PDAC patients. Patients with high NET levels also experienced more post-operative complications. NET inhibition in mice reduced tumor growth and enhanced survival. Decreased expression of collagen and matrix metalloproteinase (MMP) genes, as well as reduced intra-tumoral collagen deposition were found in NET deficient mice. Additionally, an increase in TCF1⁺PD-1⁺CD44⁺CD8⁺ progenitor T cells, a subpopulation of T cells responsive to immunotherapy, were identified. These changes resulted in further tumor burden reduction and prolonged survival when anti-PD-1 therapy was given in conjunction with NET inhibition. NETs influence extracellular matrix remodeling and the T cell response to PDAC, allowing for a significant response to anti-PD-1 therapy. These findings support the combination therapy of immunotherapy and NET inhibition in patients with PDAC.