Neutrophil extracellular traps in cancer: From mechanisms to treatments

Abstract

Neutrophil extracellular traps (NETs) are reticular ultrastructures released by activated neutrophils. As the reaction products of neutrophils, NETs have been identified as crucial effectors in pathogen defence and autoimmune diseases. Recently, increasing evidence suggest that this process also occurs in cancer. The formation and clearance of NETs are dynamically influenced by the tumour microenvironment, while NETs reciprocally play a dual role in either promoting or inhibiting tumour progression through their DNA scaffold, proteases and other granule-derived proteins. Given the interplay between NETs and tumours, active exploration is currently underway to harness their potential as tumour biomarkers and therapeutic targets. Here, we delve into the biochemical and immunological mechanisms underlying NETs formation within the tumour microenvironment, along with recent advances elucidating their multifaceted roles in tumourigenesis, metastasis and tumour-associated co-morbidities. Furthermore, we present emerging strategies for NETs-based tumour diagnostic approaches and therapeutics, with a special focus on the challenging questions that need to be answered within this field. KEY POINTS: The formation and clearance of NETs are dynamically influenced by the tumor microenvironment. NETs are engaged in tumorigenesis, formation, metastatic spread, and cancer-associated co-morbidities. NETs-based tumor biomarkers and therapeutic strategies warrant significant attention.

Keywords: biomarkers; cancer; neutrophil extracellular traps (NETs); therapeutic target.

FIGURE 1

Schematic representation of neutrophil extracellular traps (NETs) formation. Three mechanisms of NETs formation (NETosis) have been described: suicidal NETosis, vital NETosis and mitochondrial NETosis, which differ in their stimuli, morphodynamics and biological mechanisms. Among them, suicidal NETosis, as the first mechanism to be described, activates many signalling pathways after being stimulated by bacteria, chemicals and immune complexes, leading to a variety of morphological changes in neutrophils, including nuclear structure changes, disaggregated chromatin, increased permeability of nuclear and granular membranes and the release of DNA and particles into the extracellular space. These can be detected several hours after neutrophils are activated. In contrast to suicidal NETosis, vital NETosis is Nox‐independent and does not require ROS formation. Moreover, neutrophils of this mechanistic type can survive after activation, while the nucleus, membrane and plasma membrane remain intact and maintain chemotactic and phagocytotic activities. Particulate matter released through vesicles can be detected minutes after cell activation. Mitochondrial NETosis forms NETs made of mitochondrial DNA rather than nuclear DNA, similar to suicidal NETosis, which also relies on ROS but does not lead to membrane rupture and cell death and is formed in a short time. C5a, complement component 5a; CitH3, citrullinated histone H3; ERK, extracellular regulated protein kinases; FcR, Fc receptor; GM‐CSF, granulocyte/macrophage colony‐stimulating factor; GPCRs, G protein‐coupled receptors; GSDMD, gasdermin D; LPS, lipopolysaccharide; MAPK, mitogen‐activated protein kinase; MEK, mitogen‐activated extracellular signal‐regulated kinase; MPO, myeloperoxidase; NE, neutrophil elastase; NOX, nicotinamide adenine dinucleotide phosphate oxidase; PAD4, protein arginine deiminase type IV; PMA, phorbol 12‐myristate 13‐acetate; Rac, Ras‐related C3 botulinum toxin substrate; RAF, rapid acceleration fibrosarcoma; ROS, reactive oxygen species. Image created with BioRender.com with permission.

FIGURE 2

Mechanisms of neutrophil extracellular traps (NETs) formation in cancer. NETs formation is significantly influenced in the tumour microenvironment (TME), particularly through multiple cell interactions and signals from the extracellular matrix. Specifically, tumour cells can secrete cytokines, such as C5a, TNF‐α, IL‐8, IL‐17A and Chi3l1, which activate signalling pathways mediated by TLR4, TLR9, Src, p38, PI3K, AKT and ERK to induce NETosis. Endothelial cells and TH17 cells contribute to this process by secreting IL‐8 and IL‐17, respectively. Mesenchymal stromal cells regulate NETosis via the up‐regulation of complement C3 through the C3–C3aR axis or via vesicle‐mediated Fas pathways. Amyloid beta secreted by cancer‐associated fibroblasts also drives NETosis. The effects of complex protein components within the three‐dimensional non‐cellular framework of the ECM, such as SPARC and collagen, on NETosis are currently under investigation. Additionally, small molecules recruited to the TME during tumour development, such as platelets and albumin, influence NETosis by promoting ROS accumulation through distinct pathways. AKT, Ras‐related C3 botulinum toxin substrate, Rac; APT, abnormal prothrombin; CAFs, cancer‐associated fibroblasts; Chi3l1, chitinase‐3‐like protein 1; CTSC, cathepsin C; CXCR2, C–X–C motif chemokine receptor 2; C5a, complement component 5a; C3, complement receptor 3; ECs, endothelial cells; ECM, extracellular matrix; ERK, extracellular regulated protein kinases; EVs, extracellular vesicles; Fas, apo‐1/CD95/TNFRSF6; HMGB1, high mobility group box‐1 protein; IL‐1β, interleukin‐1 β; IL‐17, interleukin‐17; Mac‐1, macrophages‐1 antigen; MSCs, mesenchymal stromal cells; PI3K, phosphatidylinositol‐3‐kinase; PR3, proteinase 3; PSGL‐1, P‐selectin glycoprotein ligand‐1; ROS, reactive oxygen species; SPARC, secreted protein, acidic and rich in cysteine; SRC, proto‐oncogene tyrosine‐protein kinase Src; TAM, tumour‐associated macrophage; Th17 cells, helper T cell 17; TNF‐α, tumour necrosis factor‐α; vWF, von Willebrand factor. Image created with BioRender.com with permission.

FIGURE 3

Roles of neutrophil extracellular trap (NETs) in cancer. NETs have been identified in a diverse range of human cancer types and are intricately associated with tumourigenesis, metastasis and tumour‐associated co‐morbidities. In addition to augmenting tumour cell proliferation, NETs can facilitate metastasis through distinct mechanisms, including promoting the epithelial–mesenchymal transition, awakening dormant tumour cells, trapping circulating tumour cells and establishing pre‐metastatic niches. Importantly, NETs likely contribute to the increased risk of thrombosis and even organ dysfunction observed in cancer patients. It is noteworthy that NETs may exhibit either pro‐ or anti‐tumour functions. AKT, protein kinase B; CCDC25, coiled‐coil domain containing 25; CDC42, cell division cycle 42; cGAS, cyclic GMP‐AMP synthase; cox78, cyclooxygenase 78; EGFR, epithelial growth factor receptor; ERK, extracellular regulated protein kinases; IL‐1β, interleukin‐1 beta; IL‐8, interleukin‐8; Irs‐1, insulin receptor substrate‐1; MEPK, mitogen‐Activated protein kinase; MIR503HG, MIR503 host gene; MMP9, matrix metallopeptidase 9; MPO, myeloperoxidase; mTOR, mammalian target of rapamycin; NE, neutrophil elastase; NF‐κB, nuclear factor kappa‐light‐chain‐enhancer of activated B cells; NLRP3, nucleotide‐binding oligomerisation domain, leucine‐rich repeat and pyrin domain‐containing 3; PAD4, protein arginine deiminase type IV; PI3K, phosphoinositide 3‐kinase; RAC, Ras‐related C3 botulinum toxin substrate; RAGE, receptor of advanced glycation endproducts; ROS, reactive oxygen species; STING, stimulator of interferon genes; TGF‐β, transforming growth factor beta; TLR‐4, Toll‐like receptor 4; TLR‐9, Toll‐like receptor 9; vWF, von Willebrand factor. Image created with BioRender.com with permission.

FIGURE 4

The clinical application of neutrophil extracellular traps (NETs) in cancer. The application of NETs in cancer clinical practice benefits from the progressive advancement of detection techniques, such as multi‐immunofluorescence, multi‐spectral imaging flow cytometry, flow cell‐omics platform and microfluidic devices. Aberrant levels of NETs markers have been widely observed in the blood and tissues of cancer patients, potentially aiding in cancer diagnosis, prognosis assessment and treatment efficacy evaluation. Furthermore, the therapeutic strategies for NETs primarily revolve around inhibition of NETs formation (NETosis) and disruption of already formed NETs. CEACAM1, carcinoembryonic antigen‐related cellular adhesion molecule1; cfDNA, circulating free DNA; cfmtDNA, circulating free mitochondrial DNA; CitH3, citrullinated histone H3; G‐CSF, granulocyte colony‐stimulating factor; GSDMD, gasdermin D; MPO, myeloperoxidase; NE, neutrophil elastase; PAD4, protein arginine deiminase Type IV. Image created with BioRender.com with permission.