Neutrophils in advanced non-small cell lung cancer

Abstract

Lung cancer is the third most common cancer in the UK; 87% are histologically nonsmall cell lung cancer (NSCLC). At the time of diagnosis, patients frequently have advanced disease, with poor one-year survival due to metastatic burden. The immune system should act as a key line of defence; recognising, killing and clearing malignant cells. However, ineffective immune responses contribute to cancer progression. Neutrophils are active players in the metastatic environment, but there is limited data regarding human neutrophil populations in advanced lung cancer. Greater understanding of the interplay between extrinsic cell signals and intrinsic cell features in determining neutrophil roles, is also needed. In this study, to investigate how the extrinsic metastatic NSCLC pleural environment influences the function of neutrophils, it was modelled in vitro by culturing healthy donor neutrophils in conditions emulating the pleural space. Tumour necrosis factor alpha (TNFa) and hypoxia acted as pro-survival signals leading to neutrophil persistence and NSCLC pleural fluid conditioned neutrophils to suppress CD8+ T cells through mechanisms including programmed death-ligand 1 (PD-L1) expression. To establish the phenotype of neutrophil populations in advanced NSCLC, neutrophils were extracted from the pleural fluid (metastatic site) and blood of patients. Cellular morphology, surface marker expression and functional assays were utilised. There was an expanded population of low-density blood neutrophils in advanced NSCLC, that were not present in health, and a proportion of which were immature with banded nuclei. Similar immature neutrophils were seen in NSCLC pleural fluid. NSCLC pleural fluid neutrophils expressed PD-L1 and were long-lived. To understand how cell-intrinsic features of neutrophil populations in advanced NSCLC may determine their role in cancer, the populations were defined by their transcriptomic and proteomic signatures. This provided evidence that implied NSCLC low-density blood neutrophils represent immature cells that have been recruited from the bone marrow; they may retain proliferative capacity, have reduced neutrophil granules, and are pro-survival. In summary, the data favours a phenotype of sustained neutrophilic inflammation that is immunosuppressive at the NSCLC metastatic site, with a subpopulation of immature cells that may be ineffective/ detrimental. This is likely to permit tumour progression, leading to adverse patient outcomes.