CXCL5-mediated accumulation of SiglecFhigh neutrophils in lung tumor tissues impairs CD8 T cell responses and limits the efficacy of PD-L1 checkpoint blockade

Neutrophils are recruited in different type of cancer, including lung cancer, where they emerged as the most significant negative prognostic factor. Most experimental reports converge toward a rotumorigenic role of neutrophils via direct induction of cancer cell proliferation, promotion of metastasis, stimulation of angiogenesis and modulation of T cell responses. Nevertheless, in some cancer type and tumor stages, neutrophils exert anti-tumoral properties through direct killing of cancer cells and activation of T cell-dependent anti-tumor immunity. Thus, neutrophils in cancer display an enormous plasticity and heterogeneity which may be strongly influenced by the tissue and the constellation of immune modulating factors. Evaluate the complexity of neutrophils in tissue specific tumor microenvironment to unequivocal distinguish immunosuppressive neutrophils from other neutrophil subsets represent an important challenge in the field. In addition, understanding how these cells accumulate in tissues and how polarize to a pro- or anti-tumorigenic phenotype is crucial to develop successful cancer therapies. A critical subset of neutrophils expressing high levels of the sialic-acid-binding protein SiglecF (neuSiglecFhigh) was identified in KrasG12D/+; Trp53-/-(KP) mouse lung adenocarcinoma that correspond to a transcriptionally related human counterpart associated to negative outcomes. Neu-SiglecFhigh are mature, long-lived, cells that display tumor promoting functions associated to angiogenesis, matrix remodeling and production of ROS. However, the mechanism of recruitment of neu-SiglecFhigh in lung cancer and their impact on endogenous anti-tumor T cell responses are still unknown.Here, we used a transplantable KP line to investigate the role of the C-X-C motif chemokine 5 (CXCL5) in recruitment and accumulation of neu-SiglecFhigh in the microenvironment of lung tumors. By genome editing we abrogated the expression of CXCL5 in immunogenic KP cells (KP OVA KOCXCL5) and we characterized neutrophils frequencies and T cell activation within lung tumor tissues. We observed a drastic decrease of Cxcl5 transcripts followed by a strong reduction of neu-SiglecFhigh in KOCXCL5 tumors proving that the chemokine is a key player for their accumulation. Moreover, phenotypic and functional analysis of endogenous anti-cancer responses revealed a significant expansion of highly activated and cytotoxic tumor specific CD8+ T cells in tumor lacking neuSiglecFhigh. Immunofluorescence analysis of lung tissues shown tight CD8 T cell-neutrophils interactions, suggesting a contact-mediated mechanism of inhibition. Moreover, administration of antibodies to PD-L1 during challenge proved that neu-SiglecFhigh, due to a high expression of PD-L1, hamper the full activity of checkpoint blockade. Thus, we infer that targeting the CXCL5-axis could be a viable improvement to existing immunotherapy.